Significance of biological resource collection and tumor tissue bank creation.

PubMed

Yu, Ying-Yan; Zhu, Zheng-Gang

2010-01-15

Progress in the molecular oncology of gastrointestinal carcinomas depends on high quality cancer tissues for research. Recent acceleration on new technological platforms as well as the "omics" revolution increases the demands on tissues and peripheral blood for research at the DNA, mRNA and protein levels. Tissue bank creation emerges as a priority. Tumor tissue banks are facilities that are organized to collect, store and distribute samples of tumor and normal tissue for further use in basic and translational cancer research. The samples are generally obtained immediately after excision, prior to fixation, to ensure optimal preservation of proteins and nucleic acids. It is possible for surgeons or pathologists to collect fresh tissue prospectively during their routine dissection procedures. Most tissue banks are "project-driven" tumor banks, which are specialized collections of tumor samples on which their research is based. Systematic collection of all available tumor tissue is much rarer. High quality tissue banks need the collaboration of clinicians and basic scientists, but also the informed consent of patients and ethical approval. Through the standard operation procedure, snap frozen fresh tissue collection, storage and quality control for cryopreserved tissues are the pivotal factors on tissue bank construction and maintaining. The purpose of the tissue bank creation is enhancing the quality and speed on both the basic and translational research on gastrointestinal cancer. The quality assurance and quality control are handled based on reviewing HE staining slides or touch imprint cytology by pathologists.

Investigation of the effect of hydration on dermal collagen in ex vivo human skin tissue using second harmonic generation microscopy

NASA Astrophysics Data System (ADS)

Samatham, Ravikant; Wang, Nicholas K.; Jacques, Steven L.

2016-02-01

Effect of hydration on the dermal collagen structure in human skin was investigated using second harmonic generation microscopy. Dog ears from the Mohs micrographic surgery department were procured for the study. Skin samples with subject aged between 58-90 years old were used in the study. Three dimensional Multiphoton (Two-photon and backward SHG) control data was acquired from the skin samples. After the control measurement, the skin tissue was either soaked in deionized water for 2 hours (Hydration) or kept at room temperature for 2 hours (Desiccation), and SHG data was acquired. The data was normalized for changes in laser power and detector gain. The collagen signal per unit volume from the dermis was calculated. The desiccated skin tissue gave higher backward SHG compared to respective control tissue, while hydration sample gave a lower backward SHG. The collagen signal decreased with increase in hydration of the dermal collagen. Hydration affected the packing of the collagen fibrils causing a change in the backward SHG signal. In this study, the use of multiphoton microscopy to study the effect of hydration on dermal structure was demonstrated in ex vivo tissue.

Quantitative Evaluation of Heavy Metals and Trace Elements in the Urinary Bladder: Comparison Between Cancerous, Adjacent Non-cancerous and Normal Cadaveric Tissue.

PubMed

Abdel-Gawad, Mahmoud; Elsobky, Emad; Shalaby, Mahmoud M; Abd-Elhameed, Mohamed; Abdel-Rahim, Mona; Ali-El-Dein, Bedeir

2016-12-01

The role of heavy metals and trace elements (HMTE) in the development of some cancers has been previously reported. Bladder carcinoma is a frequent malignancy of the urinary tract. The most common risk factors for bladder cancer are exposure to industrial carcinogens, cigarette smoking, gender, and possibly diet. The aim of this study was to evaluate HTME concentrations in the cancerous and adjacent non-cancerous tissues and compare them with those of normal cadaveric bladder. This prospective study included 102 paired samples of full-thickness cancer and adjacent non-cancerous bladder tissues of radical cystectomy (RC) specimens that were histologically proven as invasive bladder cancer (MIBC). We used 17 matched controls of non-malignant bladder tissue samples from cadavers. All samples were processed and evaluated for the concentration of 22 HMTE by using Inductively Coupled Plasma Optical Emission Spectrometry (ICP-OES). Outcome analysis was made by the Mann-Whitney U, chi-square, Kruskal-Wallis, and Wilcoxon signed ranks tests. When compared with cadaveric control or cancerous, the adjacent non-cancerous tissue had higher levels of six elements (arsenic, lead, selenium, strontium, zinc, and aluminum), and when compared with the control alone, it had a higher concentration of calcium, cadmium, chromium, potassium, magnesium, and nickel. The cancerous tissue had a higher concentration of cadmium, lead, chromium, calcium, potassium, phosphorous, magnesium, nickel, selenium, strontium, and zinc than cadaveric control. Boron level was higher in cadaveric control than cancerous and adjacent non-cancerous tissue. Cadmium level was higher in cancerous tissue with node-positive than node-negative cases. The high concentrations of cadmium, lead, chromium, nickel, and zinc, in the cancerous together with arsenic in the adjacent non-cancerous tissues of RC specimens suggest a pathogenic role of these elements in BC. However, further work-up is needed to support this conclusion by the application of these HMTE on BC cell lines.

A new standard of visual data representation for imaging mass spectrometry.

PubMed

O'Rourke, Matthew B; Padula, Matthew P

2017-03-01

MALDI imaging MS (IMS) is principally used for cancer diagnostics. In our own experience with publishing IMS data, we have been requested to modify our protocols with respect to the areas of the tissue that are imaged in order to comply with the wider literature. In light of this, we have determined that current methodologies lack effective controls and can potentially introduce bias by only imaging specific areas of the targeted tissue EXPERIMENTAL DESIGN: A previously imaged sample was selected and then cropped in different ways to show the potential effect of only imaging targeted areas. By using a model sample, we were able to effectively show how selective imaging of samples can misinterpret tissue features and by changing the areas that are acquired, according to our new standard, an effective internal control can be introduced. Current IMS sampling convention relies on the assumption that sample preparation has been performed correctly. This prevents users from checking whether molecules have moved beyond borders of the tissue due to delocalization and consequentially products of improper sample preparation could be interpreted as biological features that are of critical importance when encountered in a visual diagnostic. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Impact of Sodium Bicarbonate-Buffered Lidocaine on Patient Pain During Image-Guided Breast Biopsy.

PubMed

Vasan, Alison; Baker, Jay A; Shelby, Rebecca A; Soo, Mary Scott C

2017-09-01

This randomized, double-blind controlled study evaluated the effectiveness of sodium bicarbonate-buffered lidocaine on reducing pain during imaging-guided breast biopsies. This prospective, HIPAA-compliant study randomly assigned 85 women undergoing ultrasound- or stereotactic-guided core-needle breast biopsies to receive intradermally and intraparenchymally either 1% lidocaine buffered with sodium bicarbonate (9:1 ratio) (bicarbonate study group) or 1% lidocaine alone (control group). Pain was evaluated using a 0-to-10 Likert pain scale during both intradermal and intraparenchymal anesthesia injections and during tissue sampling. Prebiopsy breast pain, anxiety, medical history, demographics, biopsy type, radiologist level of training, breast density, and lesion histology were recorded. Data were analyzed using analysis of variance and analysis of covariance. Unadjusted mean pain scores were 1.47 and 2.07 (study and control groups, respectively; P = .15) during intradermal injections, and 1.84 and 2.98 (study and control groups, respectively; P = .03) during intraparenchymal injections. Tissue sampling mean pain scores were .81 and 1.71 (study and control groups, respectively; P = .07). Moderator analyses found (1) among patients with preprocedural pain, those in the bicarbonate group experienced less intradermal injection pain (0.85 ± 1.23) than patients in the control group (2.50 ± 2.09); (2) among patients with fatty or scattered fibroglandular tissue, those in the bicarbonate group (1.35 ± 1.95) experienced less intraparenchymal injection pain than the control group (3.52 ± 3.13); and (3) during ultrasound-guided biopsies, patients in the bicarbonate group experienced less tissue-sampling pain (0.23 ± 0.63) than the control group (1.79 ± 3.05). Overall, buffering lidocaine with sodium bicarbonate significantly reduced pain during intraparenchymal injections, and additional pain reduction was found in certain patient subgroups during intradermal injections, intraparenchymal injections, and tissue sampling. Copyright © 2017 American College of Radiology. Published by Elsevier Inc. All rights reserved.

Determination of naturally occurring formaldehyde levels in sap and wood tissue of maple trees using gas chromatgraphy/mass spectrometry.

PubMed

Lagacé, Luc; Gaudy, Réjean; Perez-Locas, Carolina; Sadiki, Mustapha

2012-01-01

The occurrence of formaldehyde in sap and wood tissue of treated and untreated maple sugar trees was investigated using GC/MS. Samples were collected at different periods of the 2009 season and at different locations in Quebec, Canada. The natural concentration of formaldehyde found in untreated samples varied according to periods and locations and ranged from below the LOQ to 1.82 mg/kg for sap samples and from 2.39 to 8.92 mg/kg of fresh tissue for wood samples. Late season samples tended to have higher concentrations of formaldehyde. Samples of sap and wood tissue from tapholes treated with solutions of formaldehyde showed increased concentrations of formaldehyde for many days after treatment and were clearly distinct from untreated samples. These results will be useful to elaborate new inspection procedures for sugarbushes to control the illegal use of formaldehyde.

Application of the laser capture microdissection technique for molecular definition of skeletal cell differentiation in vivo.

PubMed

Benayahu, Dafna; Socher, Rina; Shur, Irena

2008-01-01

Laser capture microdissection (LCM) method allows selection of individual or clustered cells from intact tissues. This technology enables one to pick cells from tissues that are difficult to study individually, sort the anatomical complexity of these tissues, and make the cells available for molecular analyses. Following the cells' extraction, the nucleic acids and proteins can be isolated and used for multiple applications that provide an opportunity to uncover the molecular control of cellular fate in the natural microenvironment. Utilization of LCM for the molecular analysis of cells from skeletal tissues will enable one to study differential patterns of gene expression in the native intact skeletal tissue with reliable interpretation of function for known genes as well as to discover novel genes. Variability between samples may be caused either by differences in the tissue samples (different areas isolated from the same section) or some variances in sample handling. LCM is a multi-task technology that combines histology, microscopy work, and dedicated molecular biology. The LCM application will provide results that will pave the way toward high throughput profiling of tissue-specific gene expression using Gene Chip arrays. Detailed description of in vivo molecular pathways will make it possible to elaborate on control systems to apply for the repair of genetic or metabolic diseases of skeletal tissues.

Effects of Apollo 12 lunar material on lipid levels of tobacco tissue and slash pine cultures

NASA Technical Reports Server (NTRS)

Weete, J. D.

1972-01-01

Investigations of the lipid components of pine tissues (Pinus elloitii) are discussed, emphasizing fatty acids and steroids. The response by slash pine tissue cultures to growth in contact with Apollo lunar soil, earth basalt, and Iowa soil is studied. Tissue cultures of tobacco grown for 12 weeks in contact with lunar material from Apollo 12 flight contained 21 to 35 percent more total pigment than control tissues. No differences were noted in the fresh or dry weight of the experimental and control samples.

BMTC: --A Tool for Standardized Tissue Engineering on Ground and in Space ---

NASA Astrophysics Data System (ADS)

Kern, Peter; Kemmerle, Kurt; Jones, David

ESA is developing the BMTC (Biotechnology Mammalian Tissue Culture Facility) as ground demonstrator in order to: • establish a well characterised terrestrial platform for tissue engineer-ing under defined, reproducible conditions • prepare for future tissue engineering experiments in space using proven, well characterised, modular equipment. In the beginning the facility will be dedicated to support research of bone and cartilage growth under controlled mechanical and/or biochemical stimulation. Meanwhile, the industrial BMTC team has finalised the first model. The BMTC is highly automated system which provides standardized experiment hardware for tissue cultivation and stimulation under controlled conditions and the reproducible execution of the experiment according pre-programmed protocols. The BMTC consists of an incubator for the control of the experiment environment. Internally it offers all experiment relevant subsystems: • two Cultivation Units, each with eight Experiment Chamber Modules optical in-situ sensors for pO2 and pH • the Liquid Handling Device for medium exchange and sample taking • the handling devices for the internal transport of the experiment chamber modules to different experiment services • workstations for uni-axial loading of tissue samples; ZETOS (for bone tissue) / CHONDROS (for cartilage tissue) provision of reproducible displacement profiles measurement of the resulting forces computation of the visco-eleastic properties of the samples provision of flow induced shear stress fluorescence microscope • two different reactor types are included in the baseline flat reactor for 2D-and flat 3D-cultures with flow induced shear stress stimulation compatible with microscope cylindrical 3D-reactor for cultivation of vital bone and cartilage samples compatible with un-directional stimulation / analysis by ZETOS / CHONDROS. The modular, flexible design of the system allows the servicing and accommodation of a wide range of other experiment specific reactors. The functional principles and the essential features for controlled experiments will be reported. This facility complements the research done on ground on osteoporosis and the bone and muscle loss during bed rest studies during space flights. It is considered to become a new in-orbit research tool for tissue engineering and the verification of mechanical or pharmaceutical countermeasures.

Swab or biopsy samples for bioburden testing of allograft musculoskeletal tissue?

PubMed

Varettas, Kerry

2014-12-01

Swab and biopsy samples of allograft musculoskeletal tissue are most commonly collected by tissue banks for bacterial and fungal bioburden testing. An in vitro study was performed using the National Committee for Clinical Laboratory Standards standard 'Quality control of microbiological transport systems' (2003) to validate and evaluate the recovery of six challenge organisms from swab and biopsy samples of allograft musculoskeletal tissue. On average, 8.4 to >100 and 7.2 to >100 % of the inoculum was recovered from swab and biopsy samples respectively. A retrospective review of donor episodes was also performed, consisting of paired swab and biopsy samples received in this laboratory during the period 2001-2012. Samples of allograft femoral heads were collected from living donors during hip operations. From the 3,859 donor episodes received, 21 paired swab and biopsy samples each recovered an isolate, 247 swab samples only and 79 biopsy samples only were culture positive. Low numbers of challenge organisms were recovered from inoculated swab and biopsy samples in the in vitro study and validated their use for bioburden testing of allograft musculoskeletal tissue. Skin commensals were the most common group of organisms isolated during a 12-year retrospective review of paired swab and biopsy samples from living donor allograft femoral heads. Paired swab and biopsy samples are a suitable representative sample of allograft musculoskeletal tissue for bioburden testing.

The mitochondrial C16069T polymorphism, not mitochondrial D310 (D-loop) mononucleotide sequence variations, is associated with bladder cancer.

PubMed

Shakhssalim, Nasser; Houshmand, Massoud; Kamalidehghan, Behnam; Faraji, Abolfazl; Sarhangnejad, Reza; Dadgar, Sepideh; Mobaraki, Maryam; Rosli, Rozita; Sanati, Mohammad Hossein

2013-12-05

Bladder cancer is a relatively common and potentially life-threatening neoplasm that ranks ninth in terms of worldwide cancer incidence. The aim of this study was to determine deletions and sequence variations in the mitochondrial displacement loop (D-loop) region from the blood specimens and tumoral tissues of patients with bladder cancer, compared to adjacent non-tumoral tissues. The DNA from blood, tumoral tissues and adjacent non-tumoral tissues of twenty-six patients with bladder cancer and DNA from blood of 504 healthy controls from different ethnicities were investigated to determine sequence variation in the mitochondrial D-loop region using multiplex polymerase chain reaction (PCR), DNA sequencing and southern blotting analysis. From a total of 110 variations, 48 were reported as new mutations. No deletions were detected in tumoral tissues, adjacent non-tumoral tissues and blood samples from patients. Although the polymorphisms at loci 16189, 16261 and 16311 were not significantly correlated with bladder cancer, the C16069T variation was significantly present in patient samples compared to control samples (p < 0.05). Interestingly, there was no significant difference (p > 0.05) of C variations, including C7TC6, C8TC6, C9TC6 and C10TC6, in D310 mitochondrial DNA between patients and control samples. Our study suggests that 16069 mitochondrial DNA D-Loop mutations may play a significant role in the etiology of bladder cancer and facilitate the definition of carcinogenesis-related mutations in human cancer.

Light propagation in tissues with controlled optical properties

NASA Astrophysics Data System (ADS)

Tuchin, Valery V.; Maksimova, Irina L.; Zimnyakov, Dmitry A.; Kon, Irina L.; Mavlyutov, Albert H.; Mishin, Alexey A.

1997-10-01

Theoretical and computer modeling approaches, such as Mie theory, radiative transfer theory, diffusion wave correlation spectroscopy, and Monte Carlo simulation were used to analyze tissue optics during a process of optical clearing due to refractive index matching. Continuous wave transmittance and forward scattering measurement as well as intensity correlation experiments were used to monitor tissue structural and optical properties. As a control, tissue samples of the human sclera were taken. Osmotically active solutions, such as Trazograph, glucose, and polyethylene glycol, were used as chemicals. A characteristic time response of human scleral optical clearing the range 3 to 10 min was determined. The diffusion coefficients describing the permeability of the scleral samples to Trazograph were experimentally estimated; the average value was DT approximately equals (0.9 +/- 0.5) X 10-5 cm2/s. The results are general and can be used to describe many other fibrous tissues.

Total protein analysis as a reliable loading control for quantitative fluorescent Western blotting.

PubMed

Eaton, Samantha L; Roche, Sarah L; Llavero Hurtado, Maica; Oldknow, Karla J; Farquharson, Colin; Gillingwater, Thomas H; Wishart, Thomas M

2013-01-01

Western blotting has been a key technique for determining the relative expression of proteins within complex biological samples since the first publications in 1979. Recent developments in sensitive fluorescent labels, with truly quantifiable linear ranges and greater limits of detection, have allowed biologists to probe tissue specific pathways and processes with higher resolution than ever before. However, the application of quantitative Western blotting (QWB) to a range of healthy tissues and those from degenerative models has highlighted a problem with significant consequences for quantitative protein analysis: how can researchers conduct comparative expression analyses when many of the commonly used reference proteins (e.g. loading controls) are differentially expressed? Here we demonstrate that common controls, including actin and tubulin, are differentially expressed in tissues from a wide range of animal models of neurodegeneration. We highlight the prevalence of such alterations through examination of published "-omics" data, and demonstrate similar responses in sensitive QWB experiments. For example, QWB analysis of spinal cord from a murine model of Spinal Muscular Atrophy using an Odyssey scanner revealed that beta-actin expression was decreased by 19.3±2% compared to healthy littermate controls. Thus, normalising QWB data to β-actin in these circumstances could result in 'skewing' of all data by ∼20%. We further demonstrate that differential expression of commonly used loading controls was not restricted to the nervous system, but was also detectable across multiple tissues, including bone, fat and internal organs. Moreover, expression of these "control" proteins was not consistent between different portions of the same tissue, highlighting the importance of careful and consistent tissue sampling for QWB experiments. Finally, having illustrated the problem of selecting appropriate single protein loading controls, we demonstrate that normalisation using total protein analysis on samples run in parallel with stains such as Coomassie blue provides a more robust approach.

Inflammatory Cell Infiltrates in Acute and Chronic Thoracic Aortic Dissection.

PubMed

Wu, Darrell; Choi, Justin C; Sameri, Aryan; Minard, Charles G; Coselli, Joseph S; Shen, Ying H; LeMaire, Scott A

2013-12-01

Thoracic aortic dissection (TAD) is a highly lethal cardiovascular disease. Injury to the intima and media allows pulsatile blood to enter the media, leading to dissection formation. Inflammatory cells then infiltrate the site of aortic injury to clear dead cells and damaged tissue. This excessive inflammation may play a role in aneurysm formation after dissection. Using immunohistochemistry, we compared aortic tissues from patients with acute TAD (n = 11), patients with chronic TAD (n = 35), and donor controls (n = 20) for the presence of CD68+ macrophages, neutrophils, mast cells, and CD3+ T lymphocytes. Tissue samples from patients with acute or chronic TAD generally had significantly more inflammatory cells in both the medial and adventitial layers than did the control samples. In tissues from patients with acute TAD, the adventitia had more of the inflammatory cells studied than did the media. The pattern of increase in inflammatory cells was similar in chronic and acute TAD tissues, except for macrophages, which were seen more frequently in the adventitial layer of acute TAD tissue than in the adventitia of chronic TAD tissue. The inflammatory cell content of both acute and chronic TAD tissue was significantly different from that of control tissue. However, the inflammatory cell profile of aneurysmal chronic TAD was similar to that of acute TAD. This may reflect a sustained injury response that contributes to medial degeneration and aneurysm formation.

Liquid Microjunction Surface Sampling Probe Electrospray Mass Spectrometry for Detection of Drugs and Metabolites in Thin Tissue Sections

DOE Office of Scientific and Technical Information (OSTI.GOV)

Van Berkel, Gary J; Kertesz, Vilmos; Koeplinger, Kenneth A.

2008-01-01

A self-aspirating, liquid micro-junction surface sampling probe/electrospray emitter mass spectrometry system was demonstrated for use in the direct analysis of spotted and dosed drugs and their metabolites in thin tissue sections. Proof-of-principle sampling and analysis directly from tissue without the need for sample preparation was demonstrated first by raster scanning a region on a section of rat liver onto which reserpine was spotted. The mass spectral signal from selected reaction monitoring was used to develop a chemical image of the spotted drug on the tissue. The probe was also used to selectively spot sample areas of sagittal whole mouse bodymore » tissue sections that had been dosed orally (90 mg/kg) with R,S-sulforaphane 3 hrs prior to sacrifice. Sulforaphane and its glutathione and N-acetyl cysteine conjugates were monitored with selected reaction monitoring and detected in the stomach and various other tissues from the dosed mouse. No signal for these species was observed in the tissue from a control mouse. The same dosed tissue section was used to illustrate the possibility of obtaining a line scan across the whole body section. In total these results illustrate the potential for rapid screening of the distribution of drugs and metabolites in tissue sections with the micro-liquid junction surface sampling probe/electrospray mass spectrometry approach.« less

76 FR 66683 - Submission for OMB Review; Comment Request

Federal Register 2010, 2011, 2012, 2013, 2014

2011-10-27

...: National Oceanic and Atmospheric Administration (NOAA). Title: Protocol for Access to Tissue Specimen Samples from the National Marine Mammal Tissue Bank. OMB Control Number: 0648-0468. Form Number(s): NA..., the National Marine Mammal Tissue Bank (NMMTB) was established by the National Marine Fisheries...

Colon Cancer Associated Transcript-1 (CCAT1) Expression in Adenocarcinoma of the Stomach.

PubMed

Mizrahi, Ido; Mazeh, Haggi; Grinbaum, Ronit; Beglaibter, Nahum; Wilschanski, Michael; Pavlov, Vera; Adileh, Muchamad; Stojadinovic, Alexander; Avital, Itzhak; Gure, Ali Osmay; Halle, David; Nissan, Aviram

2015-01-01

Long non-coding RNAs (lncRNAs) have been shown to have functional roles in cancer biology and are dys-regulated in many tumors. Colon Cancer Associated Transcript -1 (CCAT1) is a lncRNA, previously shown to be significantly up-regulated in colon cancer. The aim of this study is to determine expression levels of CCAT1 in gastric carcinoma (GC). Tissue samples were obtained from patients undergoing resection for gastric carcinoma (n=19). For each patient, tumor tissue and normal appearing gastric mucosa were taken. Normal gastric tissues obtained from morbidly obese patients, undergoing laparoscopic sleeve gastrectomy served as normal controls (n=19). A human gastric carcinoma cell line (AGS) served as positive control. RNA was extracted from all tissue samples and CCAT1 expression was analyzed using quantitative real time-PCR (qRT-PCR). Low expression of CCAT1 was identified in normal gastric mucosa samples obtained from morbidly obese patients [mean Relative Quantity (RQ) = 1.95±0.4]. AGS human gastric carcinoma cell line showed an elevated level of CCAT1 expression (RQ=8.02). Expression levels of CCAT1 were approximately 10.8 fold higher in GC samples than in samples taken from the negative control group (RQ=21.1±5 vs. RQ=1.95±0.4, respectively, p<0.001). Interestingly, CCAT1 expression was significantly overexpressed in adjacent normal tissues when compared to the negative control group (RQ = 15.25±2 vs. RQ=1.95±0.4, respectively, p<0.001). Tissues obtained from recurrent GC cases showed the highest expression levels (RQ = 88.8±31; p<0.001). Expression levels increased with tumor stage (T4- 36.4±15, T3- 16.1±6, T2- 4.7±1), however this did not reach statistical significance (p=0.2). There was no difference in CCAT1 expression between intestinal and diffuse type GC (RQ=22.4±7 vs. 22.4±16, respectively, p=0.9). Within the normal gastric tissue samples, no significant difference in CCAT1 expression was observed in helicobacter pylori negative and positive patients (RQ= 2.4±0.9 vs. 0.93±0.2, respectively, p=0.13). CCAT1 is up-regulated in gastric cancer, and may serve as a potential bio-marker for early detection and surveillance.

Association between P16INK4a Promoter Methylation and Non-Small Cell Lung Cancer: A Meta-Analysis

PubMed Central

Zhu, Siwei; Hua, Feng; Zhao, Hui; Xu, Hongrui; You, Jiacong; Sun, Linlin; Wang, Weiqiang; Chen, Jun; Zhou, Qinghua

2013-01-01

Background Aberrant methylation of CpG islands acquired in tumor cells in promoter regions plays an important role in carcinogenesis. Accumulated evidence demonstrates P16INK4a gene promoter hypermethylation is involved in non-small cell lung carcinoma (NSCLC), indicating it may be a potential biomarker for this disease. The aim of this study is to evaluate the frequency of P16INK4a gene promoter methylation between cancer tissue and autologous controls by summarizing published studies. Methods By searching Medline, EMBSE and CNKI databases, the open published studies about P16INK4a gene promoter methylation and NSCLC were identified using a systematic search strategy. The pooled odds of P16INK4A promoter methylation in lung cancer tissue versus autologous controls were calculated by meta-analysis method. Results Thirty-four studies, including 2 652 NSCLC patients with 5 175 samples were included in this meta-analysis. Generally, the frequency of P16INK4A promoter methylation ranged from 17% to 80% (median 44%) in the lung cancer tissue and 0 to 80% (median 15%) in the autologous controls, which indicated the methylation frequency in cancer tissue was much higher than that in autologous samples. We also find a strong and significant correlation between tumor tissue and autologous controls of P16INK4A promoter methylation frequency across studies (Correlation coefficient 0.71, 95% CI:0.51–0.83, P<0.0001). And the pooled odds ratio of P16INK4A promoter methylation in cancer tissue was 3.45 (95% CI: 2.63–4.54) compared to controls under random-effect model. Conclusion Frequency of P16INK4a promoter methylation in cancer tissue was much higher than that in autologous controls, indicating promoter methylation plays an important role in carcinogenesis of the NSCLC. Strong and significant correlation between tumor tissue and autologous samples of P16INK4A promoter methylation demonstrated a promising biomarker for NSCLC. PMID:23577085

Role of inflammatory mediators in patients with recurrent pregnancy loss.

PubMed

Comba, Cihan; Bastu, Ercan; Dural, Ozlem; Yasa, Cenk; Keskin, Gulsah; Ozsurmeli, Mehmet; Buyru, Faruk; Serdaroglu, Hasan

2015-12-01

To examine interleukin-12 (IL-12), IL-18, IFN-γ, intracellular adhesion molecule-1 (ICAM-1), leukemia inhibitory factor (LIF), and migration inhibitory factor (MIF) levels in precisely-timed blood and endometrial tissue samples from women with idiopathic recurrent pregnancy loss (RPL). Case-control study. University hospital. Twenty-one women with RPL and 20 women with proven fertility (controls). Primary endometrial cells and blood samples during the midsecretory phase (days 19-23). Detection of IL-12, IL-18, IFN-γ, ICAM-1, LIF, and MIF via enzyme-linked immunosorbent assay in both blood and endometrial tissue samples. The blood and tissue levels of IL-12, IL-18, and IFN-γ were statistically significantly higher, and the blood and tissue levels of LIF and MIF were statistically significantly lower in patients with RPL. Only the level of tissue ICAM-1 was higher in patients with RPL. There was a strong correlation between blood and tissue level measurements of IL-12, IL-18, LIF, and MIF. Our findings support the hypothesis that inflammatory processes may contribute to pregnancy loss, possibly through their role in implantation. We found that blood and tissue levels of IL-18, LIF, and MIF, and tissue levels of IL-12, IFN-γ, and ICAM-1 have statistically significant prognostic relevance. Copyright © 2015 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Pre-operative intra-articular deep tissue sampling with novel retrograde forceps improves the diagnostics in periprosthetic joint infection.

PubMed

Wimmer, Matthias D; Ploeger, Milena M; Friedrich, Max J; Hügle, Thomas; Gravius, Sascha; Randau, Thomas M

2017-07-01

Histopathological tissue analysis is a key parameter within the diagnostic algorithm for suspected periprosthetic joint infections (PJIs), conventionally acquired in open surgery. In 2014, Hügle and co-workers introduced novel retrograde forceps for retrograde synovial biopsy with simultaneous fluid aspiration of the knee joint. We hypothesised that tissue samples acquired by retrograde synovial biopsy are equal to intra-operatively acquired deep representative tissue samples regarding bacterial detection and differentiation of periprosthetic infectious membranes. Thirty patients (male n = 15, 50%; female n = 15, 50%) with 30 suspected PJIs in painful total hip arthroplasties (THAs) were included in this prospective, controlled, non-blinded trial. The results were compared with intra-operatively obtained representative deep tissue samples. In summary, 27 out of 30 patients were diagnosed correctly as infected (17/17) or non-infected (10/13). The sensitivity to predict a PJI using the Retroforce® sampling forceps in addition to standard diagnostics was 85%, the specificity 100%. Retrograde synovial biopsy is a new and rapid diagnostic procedure under local anaesthesia in patients with painful THAs with similar histological results compared to deep tissue sampling.

Silver halide fiber optic radiometry for temperature monitoring and control of tissues heated by microwave

NASA Astrophysics Data System (ADS)

Shenfeld, Ofer; Belotserkovsky, Edward; Goldwasser, Benad; Zur, Albert; Katzir, Abraham

1993-02-01

The heating of tissue by microwave radiation has attained a place of importance in various medical fields, such as the treatment of malignancies, urinary retention, and hypothermia. Accurate temperature measurements in these treated tissues is important for treatment planning and for the control of the heating process. It is also important to be able to measure spacial temperature distribution in the tissues because they are heated in a nonuniform way by the microwave radiation. Conventional temperature sensors used today are inaccurate in the presence of microwave radiation and require contact with the heated tissue. Fiber optic radiometry makes it possible to measure temperatures accurately in the presence of microwave radiation and does not require contact with the tissue. Accurate temperature measurements of tissues heated by microwave was obtained using a silver halide optic radiometer, enabling control of the heating process in other regions of the tissue samples. Temperature mappings of the heated tissues were performed and the nonuniform temperature distributions in these tissues was demonstrated.

The Genome Austria Tissue Bank (GATiB).

PubMed

Asslaber, M; Abuja, P M; Stark, K; Eder, J; Gottweis, H; Trauner, M; Samonigg, H; Mischinger, H J; Schippinger, W; Berghold, A; Denk, H; Zatloukal, K

2007-01-01

In the context of the Austrian Genome Program, a tissue bank is being established (Genome Austria Tissue Bank, GATiB) which is based on a collection of diseased and corresponding normal tissues representing a great variety of diseases at their natural frequency of occurrence from a non-selected Central European population of more than 700,000 patients. Major emphasis is put on annotation of archival tissue with comprehensive clinical data, including follow-up data. A specific IT infrastructure supports sample annotation, tracking of sample usage as well as sample and data storage. Innovative data protection tools were developed which prevent sample donor re-identification, particularly if detailed medical and genetic data are combined. For quality control of old archival tissues, new techniques were established to check RNA quality and antigen stability. Since 2003, GATiB has changed from a population-based tissue bank to a disease-focused biobank comprising major cancers such as colon, breast, liver, as well as metabolic liver diseases and organs affected by the metabolic syndrome. Prospectively collected tissues are associated with blood samples and detailed data on the sample donor's disease, lifestyle and environmental exposure, following standard operating procedures. Major emphasis is also placed on ethical, legal and social issues (ELSI) related to biobanks. A specific research project and an international advisory board ensure the proper embedding of GATiB in society and facilitate international networking. (c) 2007 S. Karger AG, Basel.

Differential Expression of c-fos Proto-Oncogene in Normal Oral Mucosa versus Squamous Cell Carcinoma

PubMed Central

Krishna, Akhilesh; Bhatt, Madan Lal Brahma; Singh, Vineeta; Singh, Shraddha; Gangwar, Pravin Kumar; Singh, Uma Shankar; Kumar, Vijay; Mehrotra, Divya

2018-01-01

Background: The c-Fos nuclear protein dimerizes with Jun family proteins to form the transcription factor AP-1 complex which participates in signal transduction and regulation of normal cellular processes. In tumorigenesis, c-Fos promotes invasive growth through down-regulation of tumor suppressor genes but its role in oral carcinogenesis is not clear. Objectives: This study concerned c-fos gene expression in normal and malignant tissues of the oral cavity, with attention to associations between expression status and clinico-pathological profiles of OSCC patients. Method: A total of 65 histopathologically confirmed OSCC tissue samples were included in case group along with an equal number of age and sex-matched normal tissue samples of oral cavity for the control group. c-Fos protein and m-RNA expressions were analyzed using immunohistochemistry and qRT-PCR, respectively. Results: A significant low expression of c-Fos protein was observed in OSCC cases than normal control subjects (p= <0.001). The mean percent positivity of c-Fos protein in cases vs. controls was 24.91± 2.7 vs. 49.68± 2.2 (p= <0.001). Most OSCC tissue samples showed weak or moderate c-Fos expression whereas 53.8% of normal tissue sections presented with strong immunostaining. Moreover, the relative m-RNA expression for the c-fos gene was significantly decreased in case group (0.93± 0.48) as compared to the control group (1.22± 0.87). Majority of c-Fos positive cases were diagnosed with well developed tumor. The mean percent positivity of c-Fos protein was significantly lower in higher grade tumor as compared with normal oral mucosa (p= < 0.001). Conclusion: The present study suggested that the c-fos gene is downregulated in oral carcinomas. The disparity of c-Fos protein levels in different pathological grades of tumor and normal oral tissue samples may indicate that loss of c-Fos expression is related with the progression of OSCC. PMID:29582647

Yellow band disease compromises the reproductive output of the Caribbean reef-building coral Montastraea faveolata (Anthozoa, Scleractinia).

PubMed

Weil, Ernesto; Cróquer, Aldo; Urreiztieta, Isabel

2009-11-16

Sexual reproduction is critical to coral population dynamics and the long-term regeneration of coral reefs. Bleaching, disease, and/or anthropogenic-induced tissue/colony loss reduce reproductive output. This is the first attempt to explore the effect of a biotic disease on the reproduction of scleractinian corals. The study aimed to assess the effect of yellow band disease (YBD) on the reproduction of the important Caribbean reef-builder Montastraea faveolata. Tissue samples were collected from diseased, transition, and healthy-looking areas in each of 5 infected colonies and from 5 healthy controls in southwest Puerto Rico. The effect of disease-induced mortality was assessed by collecting samples from the edge and center of surviving small and large, healthy-looking tissue patches from large, previously infected tagged colonies. Fecundity was significantly lower in disease lesions compared to transition and healthy-looking tissues and the controls (99% fewer eggs). Fecundity in transition areas was significantly lower (50%) than in healthy-looking tissues in diseased colonies, which had 23% lower fecundity than control tissues. Although this fecundity drop was not statistically significant, it could indicate a systemic effect of YBD across the colony. Large and small patches had 64 and 84% fewer eggs than controls, respectively, and edge polyps had 97% fewer eggs than those in central control areas. Field observations of the spawning behavior of each tissue area corroborated the histological results. Our results indicate that YBD significantly compromises the reproductive output of M. faveolata, potentially reducing the fitness and consequently, the recovery of this important reef-building species on Caribbean coral reefs.

Development and validation of a UPLC-MS/MS method for the simultaneous determination of paritaprevir and ritonavir in rat liver.

PubMed

Ocque, Andrew J; Hagler, Colleen E; Difrancesco, Robin; Woolwine-Cunningham, Yvonne; Bednasz, Cindy J; Morse, Gene D; Talal, Andrew H

2016-07-01

Determination of paritaprevir and ritonavir in rat liver tissue samples. We successfully validated a UPLC-MS/MS method to measure paritaprevir and ritonavir in rat liver using deuterated internal standards (d8-paritapervir and d6-ritonavir). The method is linear from 20 to 20,000 and 5 to 10,000 pg on column for paritaprevir and ritonavir, respectively, and is normalized per milligram tissue. Interday and intraday variability ranged from 0.591 to 5.33% and accuracy ranged from -6.68 to 10.1% for quality control samples. The method was then applied to the measurement of paritaprevir and ritonavir in rat liver tissue samples from a pilot study. The validated method is suitable for measurement of paritaprevir and ritonavir within rat liver tissue samples for PK studies.

Increased FOXP3 expression in tumour-associated tissues of horses affected with equine sarcoid disease.

PubMed

Mählmann, K; Hamza, E; Marti, E; Dolf, G; Klukowska, J; Gerber, V; Koch, C

2014-12-01

Recent studies suggest that regulatory T cells (Tregs) are associated with disease severity and progression in papilloma virus induced neoplasia. Bovine papilloma virus (BPV) is recognised as the most important aetiological factor in equine sarcoid (ES) disease. The aim of this study was to compare expression levels of Treg markers and associated cytokines in tissue samples of ES-affected equids with skin samples of healthy control horses. Eleven ES-affected, and 12 healthy horses were included in the study. Expression levels of forkhead box protein 3 (FOXP3), interleukin 10 (IL10), interleukin 4 (IL4) and interferon gamma (IFNG) mRNA in lesional and tumour-distant samples from ES-affected horses, as well as in dermal samples of healthy control horses were measured using quantitative reverse transcription polymerase chain reaction (PCR). Expression levels were compared between lesional and tumour-distant as well as between tumour-distant and control samples. Furthermore, BPV-1 E5 DNA in samples of ES-affected horses was quantified using quantitative PCR, and possible associations of viral load, disease severity and gene expression levels were evaluated. Expression levels of FOXP3, IL10 and IFNG mRNA and BPV-1 E5 copy numbers were significantly increased in lesional compared to tumour-distant samples. There was no difference in FOXP3 and cytokine expression in tumour-distant samples from ES- compared with control horses. In tumour-distant samples viral load was positively correlated with IL10 expression and severity score. The increased expression of Treg markers in tumour-associated tissues of ES-affected equids indicates a local, Treg-induced immune suppression. Copyright © 2014 Elsevier Ltd. All rights reserved.

Controlling for anthropogenically induced atmospheric variation in stable carbon isotope studies

USGS Publications Warehouse

Long, E.S.; Sweitzer, R.A.; Diefenbach, D.R.; Ben-David, M.

2005-01-01

Increased use of stable isotope analysis to examine food-web dynamics, migration, transfer of nutrients, and behavior will likely result in expansion of stable isotope studies investigating human-induced global changes. Recent elevation of atmospheric CO2 concentration, related primarily to fossil fuel combustion, has reduced atmospheric CO2 ??13C (13C/12C), and this change in isotopic baseline has, in turn, reduced plant and animal tissue ??13C of terrestrial and aquatic organisms. Such depletion in CO2 ??13C and its effects on tissue ??13C may introduce bias into ??13C investigations, and if this variation is not controlled, may confound interpretation of results obtained from tissue samples collected over a temporal span. To control for this source of variation, we used a high-precision record of atmospheric CO2 ??13C from ice cores and direct atmospheric measurements to model modern change in CO2 ??13C. From this model, we estimated a correction factor that controls for atmospheric change; this correction reduces bias associated with changes in atmospheric isotopic baseline and facilitates comparison of tissue ??13C collected over multiple years. To exemplify the importance of accounting for atmospheric CO2 ??13C depletion, we applied the correction to a dataset of collagen ??13C obtained from mountain lion (Puma concolor) bone samples collected in California between 1893 and 1995. Before correction, in three of four ecoregions collagen ??13C decreased significantly concurrent with depletion of atmospheric CO2 ??13C (n ??? 32, P ??? 0.01). Application of the correction to collagen ??13C data removed trends from regions demonstrating significant declines, and measurement error associated with the correction did not add substantial variation to adjusted estimates. Controlling for long-term atmospheric variation and correcting tissue samples for changes in isotopic baseline facilitate analysis of samples that span a large temporal range. ?? Springer-Verlag 2005.

Evaluation of endogenous control genes for gene expression studies across multiple tissues and in the specific sets of fat- and muscle-type samples of the pig.

PubMed

Gu, Y R; Li, M Z; Zhang, K; Chen, L; Jiang, A A; Wang, J Y; Li, X W

2011-08-01

To normalize a set of quantitative real-time PCR (q-PCR) data, it is essential to determine an optimal number/set of housekeeping genes, as the abundance of housekeeping genes can vary across tissues or cells during different developmental stages, or even under certain environmental conditions. In this study, of the 20 commonly used endogenous control genes, 13, 18 and 17 genes exhibited credible stability in 56 different tissues, 10 types of adipose tissue and five types of muscle tissue, respectively. Our analysis clearly showed that three optimal housekeeping genes are adequate for an accurate normalization, which correlated well with the theoretical optimal number (r ≥ 0.94). In terms of economical and experimental feasibility, we recommend the use of the three most stable housekeeping genes for calculating the normalization factor. Based on our results, the three most stable housekeeping genes in all analysed samples (TOP2B, HSPCB and YWHAZ) are recommended for accurate normalization of q-PCR data. We also suggest that two different sets of housekeeping genes are appropriate for 10 types of adipose tissue (the HSPCB, ALDOA and GAPDH genes) and five types of muscle tissue (the TOP2B, HSPCB and YWHAZ genes), respectively. Our report will serve as a valuable reference for other studies aimed at measuring tissue-specific mRNA abundance in porcine samples. © 2011 Blackwell Verlag GmbH.

Conjunctival Expansion Using a Subtenon's Silicone Implant in New Zealand White Rabbits

PubMed Central

Yoon, Ie-na; Lee, Dong-hoon

2007-01-01

Purpose In the field of ophthalmology, the conjunctival autograft is a useful therapeutic material in many cases, but the small size of the autograft is a disadvantage. Therefore, we evaluated the feasibility of taking an expanded sample of conjunctival tissue using a subtenon's silicone implant. Materials and Methods We included a total of nine rabbits; eight rabbits were operative cases, and one was a control. A portion of conjunctival tissue from the control rabbit, which did not undergo surgery, was dissected and examined to determine whether it was histologically different from the experimental group. The surgical procedure was performed on eight rabbits via a subtenon's insertion of a silicone sponge in the left superior-temporal portion; after surgery, we dropped antibiotics into the eyes. We sacrificed a pair of rabbits every three days (on days 3, 6, 9, and 12) after surgery, removed the expanded conjunctival tissues with the silicone sponge implants, and measured their sizes. Results The mean size of the expanded conjunctival tissues was 194.4 mm2. On the third day, we were able to harvest a 223.56 mm2 section of conjunctival tissue, which was the most expanded sample of tissue in the study. On the twelfth day, we removed a 160.38 mm2 section of conjunctival tissue, which was the least expanded sample of tissue. Statistically, there were no significant differences in the mean dimensions of the expanded conjunctival tissues for each time period. Microscopic examinations showed no histological differences between the expanded conjunctival tissues and the normal conjunctival tissues. Conclusion The results reveal that this procedure is a useful method to expand the conjunctiva for grafting and transplantation. PMID:18159586

Localised drug release using MRI-controlled focused ultrasound hyperthermia.

PubMed

Staruch, Robert; Chopra, Rajiv; Hynynen, Kullervo

2011-01-01

Thermosensitive liposomes provide a mechanism for triggering the local release of anticancer drugs, but this technology requires precise temperature control in targeted regions with minimal heating of surrounding tissue. The objective of this study was to evaluate the feasibility of using MRI-controlled focused ultrasound (FUS) and thermosensitive liposomes to achieve thermally mediated localised drug delivery in vivo. Results are reported from ten rabbits, where a FUS beam was scanned in a circular trajectory to heat 10-15 mm diameter regions in normal thigh to 43°C for 20-30 min. MRI thermometry was used for closed-loop feedback control to achieve temporally and spatially uniform heating. Lyso-thermosensitive liposomal doxorubicin was infused intravenously during hyperthermia. Unabsorbed liposomes were flushed from the vasculature by saline perfusion 2 h later, and tissue samples were harvested from heated and unheated thigh regions. The fluorescence intensity of the homogenised samples was used to calculate the concentration of doxorubicin in tissue. Closed-loop control of FUS heating using MRI thermometry achieved temperature distributions with mean, T90 and T10 of 42.9°C, 41.0°C and 44.8°C, respectively, over a period of 20 min. Doxorubicin concentrations were significantly higher in tissues sampled from heated than unheated regions of normal thigh muscle (8.3 versus 0.5 ng/mg, mean per-animal difference = 7.8 ng/mg, P < 0.05, Wilcoxon matched pairs signed rank test). The results show the potential of MRI-controlled focused ultrasound hyperthermia for enhanced local drug delivery with temperature-sensitive drug carriers.

Multilaboratory trial for determination of ceftiofur residues in bovine and swine kidney and muscle, and bovine milk.

PubMed

Hornish, Rex E; Hamlow, Philip J; Brown, Scott A

2003-01-01

A multilaboratory trial for determining ceftiofur-related residues in bovine and swine kidney and muscle, and bovine milk was conducted following regulatory guidelines of the U.S. Food and Drug Administration, Center for Veterinary Medicine. The methods convert all desfuroylceftiofur-related residues containing the intact beta-lactam ring to desfuroylceftiofur acetamide to establish ceftiofur residues in tissues. Four laboratories analyzed 5 sets of samples for each tissue. Each sample set consisted of a control/blank sample and 3 control samples fortified with ceftiofur at 0.5 Rm, Rm, and 2 Rm, respectively, where Rm is the U.S. tolerance assigned for ceftiofur residue in each tissue/matrix: 0.100 microg/mL for milk, 8.0 microg/g for kidney (both species), 1.0 microg/g for bovine muscle, and 2.0 microg/g for swine muscle. Each sample set also contained 2 samples of incurred-residue tissues (one > Rm and one < Rm) from animals treated with ceftiofur hydrochloride. All laboratories completed the method trial after a familiarization phase and test of system suitability in which they demonstrated > 80% recovery in pretrial fortified test samples. Results showed that the methods met all acceptable performance criteria for recovery, accuracy, and precision. Although sample preparation was easy, solid-phase extraction cartridge performance must be carefully evaluated before samples are processed. The liquid chromatography detection system was easily set up; however, the elution profile may require slight modifications. The procedures could clearly differentiate between violative (> Rm) and nonviolative (< Rm) ceftiofur residues. Participating laboratories found the procedures suitable for ceftiofur residue determination.

A novel semi-quantitative method for measuring tissue bleeding.

PubMed

Vukcevic, G; Volarevic, V; Raicevic, S; Tanaskovic, I; Milicic, B; Vulovic, T; Arsenijevic, S

2014-03-01

In this study, we describe a new semi-quantitative method for measuring the extent of bleeding in pathohistological tissue samples. To test our novel method, we recruited 120 female patients in their first trimester of pregnancy and divided them into three groups of 40. Group I was the control group, in which no dilation was applied. Group II was an experimental group, in which dilation was performed using classical mechanical dilators. Group III was also an experimental group, in which dilation was performed using a hydraulic dilator. Tissue samples were taken from the patients' cervical canals using a Novak's probe via energetic single-step curettage prior to any dilation in Group I and after dilation in Groups II and III. After the tissue samples were prepared, light microscopy was used to obtain microphotographs at 100x magnification. The surfaces affected by bleeding were measured in the microphotographs using the Autodesk AutoCAD 2009 program and its "polylines" function. The lines were used to mark the area around the entire sample (marked A) and to create "polyline" areas around each bleeding area on the sample (marked B). The percentage of the total area affected by bleeding was calculated using the formula: N = Bt x 100 / At where N is the percentage (%) of the tissue sample surface affected by bleeding, At (A total) is the sum of the surfaces of all of the tissue samples and Bt (B total) is the sum of all the surfaces affected by bleeding in all of the tissue samples. This novel semi-quantitative method utilizes the Autodesk AutoCAD 2009 program, which is simple to use and widely available, thereby offering a new, objective and precise approach to estimate the extent of bleeding in tissue samples.

Impact of Tumour Epithelial Subtype on Circulating microRNAs in Breast Cancer Patients

PubMed Central

Brougham, Cathy; Glynn, Claire L.; Wall, Deirdre; Hyland, Peter; Duignan, Maria; McLoughlin, Mark; Newell, John; Kerin, Michael J.

2014-01-01

While a range of miRNAs have been shown to be dysregulated in the circulation of patients with breast cancer, little is known about the relationship between circulating levels and tumour characteristics. The aim of this study was to analyse alterations in circulating miRNA expression during tumour progression in a murine model of breast cancer, and to detemine the clinical relevance of identified miRNAs at both tissue and circulating level in patient samples. Athymic nude mice received a subcutaneous or mammary fat pad injection of MDA-MB-231 cells. Blood sampling was performed at weeks 1, 3 and 6 following tumour induction, and microRNA extracted. MicroRNA microArray analysis was performed comparing samples harvested at week 1 to those collected at week 6 from the same animals. Significantly altered miRNAs were validated across all murine samples by RQ-PCR (n = 45). Three miRNAs of interest were then quantified in the circulation(n = 166) and tissue (n = 100) of breast cancer patients and healthy control individuals. MicroArray-based analysis of murine blood samples revealed levels of 77 circulating microRNAs to be changed during disease progression, with 44 demonstrating changes >2-fold. Validation across all samples revealed miR-138 to be significantly elevated in the circulation of animals during disease development, with miR-191 and miR-106a levels significantly decreased. Analysis of patient tissue and blood samples revealed miR-138 to be significantly up-regulated in the circulation of patients with breast cancer, with no change observed in the tissue setting. While not significantly changed overall in breast cancer patients compared to controls, circulating miR-106a and miR-191 were significantly decreased in patients with basal breast cancer. In tissue, both miRNAs were significantly elevated in breast cancer compared to normal breast tissue. The data demonstrates an impact of tumour epithelial subtype on circulating levels of miRNAs, and highlights divergent miRNA profiles between tissue and blood samples from breast cancer patients. PMID:24626163

Equilibrium Passive Sampling of POP in Lipid-Rich and Lean Fish Tissue: Quality Control Using Performance Reference Compounds.

PubMed

Rusina, Tatsiana P; Carlsson, Pernilla; Vrana, Branislav; Smedes, Foppe

2017-10-03

Passive sampling is widely used to measure levels of contaminants in various environmental matrices, including fish tissue. Equilibrium passive sampling (EPS) of persistent organic pollutants (POP) in fish tissue has been hitherto limited to application in lipid-rich tissue. We tested several exposure methods to extend EPS applicability to lean tissue. Thin-film polydimethylsiloxane (PDMS) passive samplers were exposed statically to intact fillet and fish homogenate and dynamically by rolling with cut fillet cubes. The release of performance reference compounds (PRC) dosed to passive samplers prior to exposure was used to monitor the exchange process. The sampler-tissue exchange was isotropic, and PRC were shown to be good indicators of sampler-tissue equilibration status. The dynamic exposures demonstrated equilibrium attainment in less than 2 days for all three tested fish species, including lean fish containing 1% lipid. Lipid-based concentrations derived from EPS were in good agreement with lipid-normalized concentrations obtained using conventional solvent extraction. The developed in-tissue EPS method is robust and has potential for application in chemical monitoring of biota and bioaccumulation studies.

Colon Cancer Associated Transcript-1 (CCAT1) Expression in Adenocarcinoma of the Stomach

PubMed Central

Mizrahi, Ido; Mazeh, Haggi; Grinbaum, Ronit; Beglaibter, Nahum; Wilschanski, Michael; Pavlov, Vera; Adileh, Muchamad; Stojadinovic, Alexander; Avital, Itzhak; Gure, Ali Osmay; Halle, David; Nissan, Aviram

2015-01-01

Background: Long non-coding RNAs (lncRNAs) have been shown to have functional roles in cancer biology and are dys-regulated in many tumors. Colon Cancer Associated Transcript -1 (CCAT1) is a lncRNA, previously shown to be significantly up-regulated in colon cancer. The aim of this study is to determine expression levels of CCAT1 in gastric carcinoma (GC). Methods: Tissue samples were obtained from patients undergoing resection for gastric carcinoma (n=19). For each patient, tumor tissue and normal appearing gastric mucosa were taken. Normal gastric tissues obtained from morbidly obese patients, undergoing laparoscopic sleeve gastrectomy served as normal controls (n=19). A human gastric carcinoma cell line (AGS) served as positive control. RNA was extracted from all tissue samples and CCAT1 expression was analyzed using quantitative real time-PCR (qRT-PCR). Results: Low expression of CCAT1 was identified in normal gastric mucosa samples obtained from morbidly obese patients [mean Relative Quantity (RQ) = 1.95±0.4]. AGS human gastric carcinoma cell line showed an elevated level of CCAT1 expression (RQ=8.02). Expression levels of CCAT1 were approximately 10.8 fold higher in GC samples than in samples taken from the negative control group (RQ=21.1±5 vs. RQ=1.95±0.4, respectively, p<0.001). Interestingly, CCAT1 expression was significantly overexpressed in adjacent normal tissues when compared to the negative control group (RQ = 15.25±2 vs. RQ=1.95±0.4, respectively, p<0.001). Tissues obtained from recurrent GC cases showed the highest expression levels (RQ = 88.8±31; p<0.001). Expression levels increased with tumor stage (T4- 36.4±15, T3- 16.1±6, T2- 4.7±1), however this did not reach statistical significance (p=0.2). There was no difference in CCAT1 expression between intestinal and diffuse type GC (RQ=22.4±7 vs. 22.4±16, respectively, p=0.9). Within the normal gastric tissue samples, no significant difference in CCAT1 expression was observed in helicobacter pylori negative and positive patients (RQ= 2.4±0.9 vs. 0.93±0.2, respectively, p=0.13). Conclusion: CCAT1 is up-regulated in gastric cancer, and may serve as a potential bio-marker for early detection and surveillance. PMID:25561974

Pathological changes in Alzheimer"s brain evaluated with fluorescence emission analysis (FEA)

NASA Astrophysics Data System (ADS)

Christov, Alexander; Ottman, Todd; Grammas, Paula

2004-07-01

Development of AD is associated with cerebrovascular deposition of amyloid beta (Aβ) as well as a progressive increase in vasular collagen content. Both AΒ and collagen are naturally fluorescent compounds when exposed to UV light. We analyzed autofluorescence emitted from brain tissue samples and isolated brain resistance vessels harvested postmortem from patients with Alzheimer's disease (AD) and age-matched controls. Fluorescence emission, excited at 355 nm with an Nd:YAG laser, was measured using a fiber-optic based fluorescence spectroscopic system for tissue analysis. Significantly higher values of fluorescence emission intensity (P<0.001) in the spectral region from 465 to 490 nm were detected in brain resistance vessel samples from AD patients compared to the normal individuals. Results from western blot analysis showed elevated levels of type I and type III collagen, and reduced levels of type IV collagen in resistance vessels from AD patients, compared to control samples. In addition, using direct scanning of the cortical suface for fluoresxcence emission by the laser-induced fluorescence spectroscopy system we detected a significantly (P<0.05) higher level of apoptosis in AD brain tissue compared to age-matched controls. Fluorescence emission analysis (FEA) appears to be a sensitive technique for detecting structural changes in AD brain tissue.

Ultrasonically Assisted Cutting of Bio-tissues in Microtomy

NASA Astrophysics Data System (ADS)

Wang, Dong; Roy, Anish; Silberschmidt, Vadim V.

Modern-day histology of bio-tissues for supporting stratified medicine diagnoses requires high-precision cutting to ensure high quality extremely thin specimens used in analysis. Additionally, the cutting quality is significantly affected by a wide variety of soft and hard tissues in the samples. This paper deals with development of a next generation of microtome employing introduction of controlled ultrasonic vibration to realise a hybrid cutting process of bio-tissues. The study is based on a combination of advanced experimental and numerical (finite-element) studies of multi-body dynamics of a cutting system. The quality of cut samples produced with the prototype is compared with the state-of-the-art.

Digital immunohistochemistry platform for the staining variation monitoring based on integration of image and statistical analyses with laboratory information system.

PubMed

Laurinaviciene, Aida; Plancoulaine, Benoit; Baltrusaityte, Indra; Meskauskas, Raimundas; Besusparis, Justinas; Lesciute-Krilaviciene, Daiva; Raudeliunas, Darius; Iqbal, Yasir; Herlin, Paulette; Laurinavicius, Arvydas

2014-01-01

Digital immunohistochemistry (IHC) is one of the most promising applications brought by new generation image analysis (IA). While conventional IHC staining quality is monitored by semi-quantitative visual evaluation of tissue controls, IA may require more sensitive measurement. We designed an automated system to digitally monitor IHC multi-tissue controls, based on SQL-level integration of laboratory information system with image and statistical analysis tools. Consecutive sections of TMA containing 10 cores of breast cancer tissue were used as tissue controls in routine Ki67 IHC testing. Ventana slide label barcode ID was sent to the LIS to register the serial section sequence. The slides were stained and scanned (Aperio ScanScope XT), IA was performed by the Aperio/Leica Colocalization and Genie Classifier/Nuclear algorithms. SQL-based integration ensured automated statistical analysis of the IA data by the SAS Enterprise Guide project. Factor analysis and plot visualizations were performed to explore slide-to-slide variation of the Ki67 IHC staining results in the control tissue. Slide-to-slide intra-core IHC staining analysis revealed rather significant variation of the variables reflecting the sample size, while Brown and Blue Intensity were relatively stable. To further investigate this variation, the IA results from the 10 cores were aggregated to minimize tissue-related variance. Factor analysis revealed association between the variables reflecting the sample size detected by IA and Blue Intensity. Since the main feature to be extracted from the tissue controls was staining intensity, we further explored the variation of the intensity variables in the individual cores. MeanBrownBlue Intensity ((Brown+Blue)/2) and DiffBrownBlue Intensity (Brown-Blue) were introduced to better contrast the absolute intensity and the colour balance variation in each core; relevant factor scores were extracted. Finally, tissue-related factors of IHC staining variance were explored in the individual tissue cores. Our solution enabled to monitor staining of IHC multi-tissue controls by the means of IA, followed by automated statistical analysis, integrated into the laboratory workflow. We found that, even in consecutive serial tissue sections, tissue-related factors affected the IHC IA results; meanwhile, less intense blue counterstain was associated with less amount of tissue, detected by the IA tools.

Digital immunohistochemistry platform for the staining variation monitoring based on integration of image and statistical analyses with laboratory information system

PubMed Central

2014-01-01

Background Digital immunohistochemistry (IHC) is one of the most promising applications brought by new generation image analysis (IA). While conventional IHC staining quality is monitored by semi-quantitative visual evaluation of tissue controls, IA may require more sensitive measurement. We designed an automated system to digitally monitor IHC multi-tissue controls, based on SQL-level integration of laboratory information system with image and statistical analysis tools. Methods Consecutive sections of TMA containing 10 cores of breast cancer tissue were used as tissue controls in routine Ki67 IHC testing. Ventana slide label barcode ID was sent to the LIS to register the serial section sequence. The slides were stained and scanned (Aperio ScanScope XT), IA was performed by the Aperio/Leica Colocalization and Genie Classifier/Nuclear algorithms. SQL-based integration ensured automated statistical analysis of the IA data by the SAS Enterprise Guide project. Factor analysis and plot visualizations were performed to explore slide-to-slide variation of the Ki67 IHC staining results in the control tissue. Results Slide-to-slide intra-core IHC staining analysis revealed rather significant variation of the variables reflecting the sample size, while Brown and Blue Intensity were relatively stable. To further investigate this variation, the IA results from the 10 cores were aggregated to minimize tissue-related variance. Factor analysis revealed association between the variables reflecting the sample size detected by IA and Blue Intensity. Since the main feature to be extracted from the tissue controls was staining intensity, we further explored the variation of the intensity variables in the individual cores. MeanBrownBlue Intensity ((Brown+Blue)/2) and DiffBrownBlue Intensity (Brown-Blue) were introduced to better contrast the absolute intensity and the colour balance variation in each core; relevant factor scores were extracted. Finally, tissue-related factors of IHC staining variance were explored in the individual tissue cores. Conclusions Our solution enabled to monitor staining of IHC multi-tissue controls by the means of IA, followed by automated statistical analysis, integrated into the laboratory workflow. We found that, even in consecutive serial tissue sections, tissue-related factors affected the IHC IA results; meanwhile, less intense blue counterstain was associated with less amount of tissue, detected by the IA tools. PMID:25565007

In vitro antimicrobial effect of the tissue conditioner containing silver nanoparticles

PubMed Central

2011-01-01

PURPOSE The aim of this study was to identify in vitro antimicrobial activity of the tissue conditioner containing silver nanoparticles on microbial strains, Staphylococcus aureus, Streptococcus mutans and Candida albicans. MATERIALS AND METHODS Experimental disc samples (20.0×3.0 mm) of tissue conditioner (GC Soft-Liner, GC cooperation, Tokyo, Japan) containing 0.1 - 3.0% silver nanoparticles (0%: control) were fabricated. Samples were placed on separate culture plate dish and microbial suspensions (100 µL) of tested strains were inoculated then incubated at 37℃. Microbial growth was verified at 24 hrs and 72 hrs and the antimicrobial effects of samples were evaluated as a percentage of viable cells in withdrawn suspension (100 µL). Data were recorded as the mean of three colony forming unit (CFU) numerations and the borderline of the antimicrobial effect was determined at 0.1% viable cells. RESULTS A 0.1% silver nanoparticles combined to tissue conditioner displayed minimal bactericidal effect against Staphylococcus aureus and Streptococcus mutans strains, a 0.5% for fungal strain. Control group did not show any microbial inhibitory effect and there were no statistical difference between 24 hrs and extended 72 hrs incubation time (P > .05). CONCLUSION Within the limitation of this in vitro study, the results suggest that the tissue conditioner containing silver nanoparticles could be an antimicrobial dental material in denture plaque control. Further mechanical stability and toxicity studies are still required. PMID:21503189

The Study of Pentoxifylline Drug Effects on Renal Apoptosis and BCL-2 Gene Expression Changes Following Ischemic Reperfusion Injury in Rat

PubMed Central

Hashemi, Mehrdad

2014-01-01

Ischemia Reperfusion injury is the tissue damage caused when blood supply returns to the tissue after a period of ischemia or lack of oxygen. In this study, the effect of pentoxyfylline on BCL-2 gene expression changes and cell injury in kidney of rat following Ischemia Reperfusion were evaluated. In this experimental study, 20 male wistar rats with average weight of 250-300 g were selected and then were accidently divided them on two tenth group of control and treatment groups. In the control group, celiotomy was performed by ventral midline incision. The left kidney was isolated, and then both the renal artery and vein were obstructed. After 60 minutes of warm ischemia, vessel obstruction resolved and the right kidney was removed. 72 hours after reperfusion, tissue samples were taken from left kidney for Tunel assay. We used quantitative real time PCR for detection of BCL-2 gene expression in treated groups and then compared them to control samples. In the treatment group, the cell death changes, showed lower level than the control group. The results also showed the BCL-2 gene expression was declined in ischemia group as campared to PNT drug group. The pentoxyfylline might have a role in control of apoptosis result from Ischemia- reperfusion and quantitative real-time PCR can be used as a direct method for detection BCL-2 gene expression in tested samples and normal samples. PMID:24734070

Quantitative assessment of protein activity in orphan tissues and single cells using the metaVIPER algorithm. | Office of Cancer Genomics

Cancer.gov

We and others have shown that transition and maintenance of biological states is controlled by master regulator proteins, which can be inferred by interrogating tissue-specific regulatory models (interactomes) with transcriptional signatures, using the VIPER algorithm. Yet, some tissues may lack molecular profiles necessary for interactome inference (orphan tissues), or, as for single cells isolated from heterogeneous samples, their tissue context may be undetermined.

In vitro biological evaluation of beta-TCP/HDPE--A novel orthopedic composite: a survey using human osteoblast and fibroblast bone cells.

PubMed

Homaeigohar, S Sh; Shokrgozar, M A; Khavandi, A; Sadi, A Yari

2008-02-01

Beta-tricalcium phosphate reinforced high density polyethylene (beta-TCP/HDPE) was prepared to simulate bone composition and to study its capacity to act as bone tissue. This material was produced by replacing the mineral component and collagen soft tissue of the bone with beta-TCP and HDPE, respectively. The biocompatibility of the composite samples with different volume fractions of TCP (20, 30 and 40 vol %) was examined in vitro using two osteoblast cell lines G-292 and Saos-2, and also a type of fibroblast cell isolated from bone tissue, namely human bone fibroblast (HBF) by proliferation, and cell adhesion assays. Cell-material interaction with the surface of the composite samples was examined by scanning electron microscopy (SEM). The effect of beta-TCP/HDPE on the behavior of osteoblast and fibroblast cells was compared with those of composite and negative control samples; polyethylene (PE) and tissue culture polystyrene (TPS), respectively. In general, the results showed that the composite samples containing beta-TCP as reinforcement supported a higher rate of proliferation by various bone cells after 3, 7, and 14 days of incubation compared to the composite control sample. Furthermore, more osteoblast cells were attached to the surface of the composite samples when compared to the composite control samples after the above incubation periods (p < 0.05), while in the case of HBF an equal or even higher number of cells adhered to PE was observed. The number of adhered osteoblast cells was almost equal and in some days even higher than the number of adhered cells on negative control sample, while in the case of fibroblast this difference was significantly higher than TPS (p < 0.05). Adhered cells presented a normal morphology by SEM and many of the cells were observed to be undergoing cell division. These findings indicate that beta-TCP/HDPE composites are biocompatible, nontoxic, and act to stimulate proliferation and adhesion of the cells, whether osteoblast or fibroblast. (c) 2007 Wiley Periodicals, Inc. J Biomed Mater Res, 2008.

Induction of Macrophage Chemotaxis by Aortic Extracts from Patients with Marfan Syndrome Is Related to Elastin Binding Protein

PubMed Central

Guo, Gao; Gehle, Petra; Doelken, Sandra; Martin-Ventura, José Luis; von Kodolitsch, Yskert; Hetzer, Roland; Robinson, Peter N.

2011-01-01

Marfan syndrome is an autosomal dominantly inherited disorder of connective tissue with prominent skeletal, ocular, and cardiovascular manifestations. Aortic aneurysm and dissection are the major determinants of premature death in untreated patients. In previous work, we showed that extracts of aortic tissues from the mgR mouse model of Marfan syndrome showed increased chemotactic stimulatory activity related to the elastin-binding protein. Aortic samples were collected from 6 patients with Marfan syndrome and 8 with isolated aneurysms of the ascending aorta. Control samples were obtained from 11 organ donors without known vascular or connective tissue diseases. Soluble proteins extracted from the aortic samples of the two patient groups were compared against buffer controls and against the aortic samples from controls with respect to the ability to induce macrophage chemotaxis as measured using a modified Boyden chamber, as well as the reactivity to a monoclonal antibody BA4 against bioactive elastin peptides using ELISA. Samples from Marfan patients displayed a statistically significant increase in chemotactic inductive activity compared to control samples. Additionally, reactivity to BA4 was significantly increased. Similar statistically significant increases were identified for the samples from patients with idiopathic thoracic aortic aneurysm. There was a significant correlation between the chemotactic index and BA4 reactivity, and the increases in chemotactic activity of extracts from Marfan patients could be inhibited by pretreatment with lactose, VGVAPG peptides, or BA4, which indicates the involvement of EBP in mediating the effects. Our results demonstrate that aortic extracts of patients with Marfan syndrome can elicit macrophage chemotaxis, similar to our previous study on aortic extracts of the mgR mouse model of Marfan syndrome (Guo et al., Circulation 2006; 114:1855-62). PMID:21647416

Shaping field for deep tissue microscopy

NASA Astrophysics Data System (ADS)

Colon, J.; Lim, H.

2015-05-01

Information capacity of a lossless image-forming system is a conserved property determined by two imaging parameters - the resolution and the field of view (FOV). Adaptive optics improves the former by manipulating the phase, or wavefront, in the pupil plane. Here we describe a homologous approach, namely adaptive field microscopy, which aims to enhance the FOV by controlling the phase, or defocus, in the focal plane. In deep tissue imaging, the useful FOV can be severely limited if the region of interest is buried in a thick sample and not perpendicular to the optic axis. One must acquire many z-scans and reconstruct by post-processing, which exposes tissue to excessive radiation and is also time consuming. We demonstrate the effective FOV can be substantially enhanced by dynamic control of the image plane. Specifically, the tilt of the image plane is continuously adjusted in situ to match the oblique orientation of the sample plane within tissue. The utility of adaptive field microscopy is tested for imaging tissue with non-planar morphology. Ocular tissue of small animals was imaged by two-photon excited fluorescence. Our results show that adaptive field microscopy can utilize the full FOV. The freedom to adjust the image plane to account for the geometrical variations of sample could be extremely useful for 3D biological imaging. Furthermore, it could facilitate rapid surveillance of cellular features within deep tissue while avoiding photo damages, making it suitable for in vivo imaging.

Effect of Irradiation on Tissue Penetration Depth of Doxorubicin after Pressurized Intra-Peritoneal Aerosol Chemotherapy (PIPAC) in a Novel Ex-Vivo Model.

PubMed

Khosrawipour, Veria; Giger-Pabst, Urs; Khosrawipour, Tanja; Pour, Yousef Hedayat; Diaz-Carballo, David; Förster, Eckart; Böse-Ribeiro, Hugo; Adamietz, Irenäus Anton; Zieren, Jürgen; Fakhrian, Khashayar

2016-01-01

This study was performed to assess the impact of irradiation on the tissue penetration depth of doxorubicin delivered during Pressurized Intra-Peritoneal Aerosol Chemotherapy (PIPAC). Fresh post mortem swine peritoneum was cut into 10 proportional sections. Except for 2 control samples, all received irradiation with 1, 2, 7 and 14 Gy, respectively. Four samples received PIPAC 15 minutes after irradiation and 4 other after 24 hours. Doxorubicin was aerosolized in an ex-vivo PIPAC model at 12 mmHg/36°C. In-tissue doxorubicin penetration was measured using fluorescence microscopy on frozen thin sections. Doxorubicin penetration after PIPAC (15 minutes after irradiation) was 476 ± 74 µm for the control sample, 450 ± 45µm after 1 Gy (p > 0.05), 438 ± 29 µm after 2 Gy (p > 0.05), 396 ± 32 µm after 7 Gy (p = 0.005) and 284 ± 57 after 14 Gy irradiation (p < 0.001). The doxorubicin penetration after PIPAC (24 hours after irradiation) was 428 ± 77 µm for the control sample, 393 ± 41 µm after 1 Gy (p > 0.05), 379 ± 56 µm after 2 Gy (p > 0.05), 352 ± 53 µm after 7 Gy (p = 0.008) and 345 ± 53 after 14 Gy irradiation (p = 0.001). Higher (fractional) radiation dose might reduce the tissue penetration depth of doxorubicin in our ex-vivo model. However, irradiation with lower (fractional) radiation dose does not affect the tissue penetration negatively. Further studies are warranted to investigate if irradiation can be used safely as chemopotenting agent for patients with peritoneal metastases treated with PIPAC.

LASER BIOLOGY: Optomechanical tests of hydrated biological tissues subjected to laser shaping

NASA Astrophysics Data System (ADS)

Omel'chenko, A. I.; Sobol', E. N.

2008-03-01

The mechanical properties of a matrix are studied upon changing the size and shape of biological tissues during dehydration caused by weak laser-induced heating. The cartilage deformation, dehydration dynamics, and hydraulic conductivity are measured upon laser heating. The hydrated state and the shape of samples of separated fascias and cartilaginous tissues were controlled by using computer-aided processing of tissue images in polarised light.

Identifying Reproducible Molecular Biomarkers for Gastric Cancer Metastasis with the Aid of Recurrence Information

PubMed Central

Li, Mengyao; Hong, Guini; Cheng, Jun; Li, Jing; Cai, Hao; Li, Xiangyu; Guan, Qingzhou; Tong, Mengsha; Li, Hongdong; Guo, Zheng

2016-01-01

To precisely diagnose metastasis state is important for tailoring treatments for gastric cancer patients. However, the routinely employed radiological and pathologic tests for tumour metastasis have considerable high false negative rates, which may retard the identification of reproducible metastasis-related molecular biomarkers for gastric cancer. In this research, using three datasets, we firstly shwed that differentially expressed genes (DEGs) between metastatic tissue samples and non-metastatic tissue samples could hardly be reproducibly detected with a proper statistical control when the metastatic and non-metastatic samples were defined by TNM stage alone. Then, assuming that undetectable micrometastases are the prime cause for recurrence of early stage patients with curative resection, we reclassified all the “non-metastatic” samples as metastatic samples whenever the patients experienced tumour recurrence during follow-up after tumour resection. In this way, we were able to find distinct and reproducible DEGs between the reclassified metastatic and non-metastatic tissue samples and concordantly significant DNA methylation alterations distinguishing metastatic tissues and non-metastatic tissues of gastric cancer. Our analyses suggested that the follow-up recurrence information for patients should be employed in the research of tumour metastasis in order to decrease the confounding effects of false non-metastatic samples with undetected micrometastases. PMID:27109211

Bioassay of procoagulant albumin in human plasma.

PubMed

Grosset, A; Liu, L; Parker, C J; Rodgers, G M

1994-09-01

Procoagulant albumin (P-Al) is present in normal human plasma and increases monocyte and endothelial cell expression of tissue factor activity. To develop a bioassay for P-Al, we partially purified plasma from healthy volunteers and several patient groups using BaCl2 and (NH4)2SO4 precipitation. The samples were assayed for tissue factor (TF) inducing activity, expressed as a percentage increase compared to a serum-free media control. Over six months, the assay was reproducible in stored samples and in serial samples from normal volunteers. The plasma P-Al activities of 35 volunteers averaged 141 +/- 8.2% (SEM). There was no diurnal variation. There was no difference in the P-Al activity after a 12 hour fast and 2 hours after a large meal in 4 healthy volunteers. There was no increase in activity (r = 0.16) with the subject's age. The average activity from 16 poorly-controlled diabetics was 131 +/- 11% (SEM). No alteration in activity was seen with samples from patients with uremia, liver dysfunction, hemophilia, thrombotic events, or adenocarcinoma. These results indicate that P-Al activity can be bioassayed in individual patient samples; however, pathologic states associated with abnormal P-Al-induced tissue factor activity presently remain unidentified.

Tissue Distribution of Enrofloxacin in African Clawed Frogs (Xenopus laevis) after Intramuscular and Subcutaneous Administration

PubMed Central

Felt, Stephen; Papich, Mark G; Howard, Antwain; Long, Tyler; McKeon, Gabriel; Torreilles, Stéphanie; Green, Sherril

2013-01-01

As part of an enrofloxacin pharmacokinetic study, concentrations of enrofloxacin and ciprofloxacin (metabolite) were measured in various tissues (brain, heart, kidney, liver, lung, and spleen) collected from treated (subcutaneous delivery, n = 3; intramuscular delivery, n = 3; untreated controls, n = 2) adult female Xenopus laevis by using HPLC. Enrofloxacin was rapidly absorbed after administration by either route and readily diffused into all sampled tissues. Enrofloxacin and ciprofloxacin were present in the tissue samples collected at 8 h. The highest average tissue concentrations for enrofloxacin were found in kidney, with the lowest concentrations in liver. Ciprofloxacin tissue concentrations paralleled but were always lower than those of enrofloxacin for all time points and tissues except brain and kidney. These results, together with previously published pharmacokinetic data and known minimal inhibitory concentrations of common pathogenic bacteria, provide a strong evidence-based rationale for choosing enrofloxacin to treat infectious diseases in X. laevis. PMID:23562103

Tissue distribution of enrofloxacin in African clawed frogs (Xenopus laevis) after intramuscular and subcutaneous administration.

PubMed

Felt, Stephen; Papich, Mark G; Howard, Antwain; Long, Tyler; McKeon, Gabriel; Torreilles, Stéphanie; Green, Sherril

2013-03-01

As part of an enrofloxacin pharmacokinetic study, concentrations of enrofloxacin and ciprofloxacin (metabolite) were measured in various tissues (brain, heart, kidney, liver, lung, and spleen) collected from treated (subcutaneous delivery, n = 3; intramuscular delivery, n = 3; untreated controls, n = 2) adult female Xenopus laevis by using HPLC. Enrofloxacin was rapidly absorbed after administration by either route and readily diffused into all sampled tissues. Enrofloxacin and ciprofloxacin were present in the tissue samples collected at 8 h. The highest average tissue concentrations for enrofloxacin were found in kidney, with the lowest concentrations in liver. Ciprofloxacin tissue concentrations paralleled but were always lower than those of enrofloxacin for all time points and tissues except brain and kidney. These results, together with previously published pharmacokinetic data and known minimal inhibitory concentrations of common pathogenic bacteria, provide a strong evidence-based rationale for choosing enrofloxacin to treat infectious diseases in X. laevis.

Extraction efficiency and implications for absolute quantitation of propranolol in mouse brain, liver and kidney thin tissue sections using droplet-based liquid microjunction surface sampling-HPLC ESI-MS/MS

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kertesz, Vilmos; Weiskittel, Taylor M.; Vavek, Marissa

Currently, absolute quantitation aspects of droplet-based surface sampling for thin tissue analysis using a fully automated autosampler/HPLC-ESI-MS/MS system are not fully evaluated. Knowledge of extraction efficiency and its reproducibility is required to judge the potential of the method for absolute quantitation of analytes from thin tissue sections. Methods: Adjacent thin tissue sections of propranolol dosed mouse brain (10- μm-thick), kidney (10- μm-thick) and liver (8-, 10-, 16- and 24- μm-thick) were obtained. Absolute concentration of propranolol was determined in tissue punches from serial sections using standard bulk tissue extraction protocols and subsequent HPLC separations and tandem mass spectrometric analysis. Thesemore » values were used to determine propranolol extraction efficiency from the tissues with the droplet-based surface sampling approach. Results: Extraction efficiency of propranolol using 10- μm-thick brain, kidney and liver thin tissues using droplet-based surface sampling varied between ~45-63%. Extraction efficiency decreased from ~65% to ~36% with liver thickness increasing from 8 μm to 24 μm. Randomly selecting half of the samples as standards, precision and accuracy of propranolol concentrations obtained for the other half of samples as quality control metrics were determined. Resulting precision ( ±15%) and accuracy ( ±3%) values, respectively, were within acceptable limits. In conclusion, comparative quantitation of adjacent mouse thin tissue sections of different organs and of various thicknesses by droplet-based surface sampling and by bulk extraction of tissue punches showed that extraction efficiency was incomplete using the former method, and that it depended on the organ and tissue thickness. However, once extraction efficiency was determined and applied, the droplet-based approach provided the required quantitation accuracy and precision for assay validations. Furthermore, this means that once the extraction efficiency was calibrated for a given tissue type and drug, the droplet-based approach provides a non-labor intensive and high-throughput means to acquire spatially resolved quantitative analysis of multiple samples of the same type.« less

Extraction efficiency and implications for absolute quantitation of propranolol in mouse brain, liver and kidney thin tissue sections using droplet-based liquid microjunction surface sampling-HPLC ESI-MS/MS

DOE PAGES

Kertesz, Vilmos; Weiskittel, Taylor M.; Vavek, Marissa; ...

2016-06-22

Currently, absolute quantitation aspects of droplet-based surface sampling for thin tissue analysis using a fully automated autosampler/HPLC-ESI-MS/MS system are not fully evaluated. Knowledge of extraction efficiency and its reproducibility is required to judge the potential of the method for absolute quantitation of analytes from thin tissue sections. Methods: Adjacent thin tissue sections of propranolol dosed mouse brain (10- μm-thick), kidney (10- μm-thick) and liver (8-, 10-, 16- and 24- μm-thick) were obtained. Absolute concentration of propranolol was determined in tissue punches from serial sections using standard bulk tissue extraction protocols and subsequent HPLC separations and tandem mass spectrometric analysis. Thesemore » values were used to determine propranolol extraction efficiency from the tissues with the droplet-based surface sampling approach. Results: Extraction efficiency of propranolol using 10- μm-thick brain, kidney and liver thin tissues using droplet-based surface sampling varied between ~45-63%. Extraction efficiency decreased from ~65% to ~36% with liver thickness increasing from 8 μm to 24 μm. Randomly selecting half of the samples as standards, precision and accuracy of propranolol concentrations obtained for the other half of samples as quality control metrics were determined. Resulting precision ( ±15%) and accuracy ( ±3%) values, respectively, were within acceptable limits. In conclusion, comparative quantitation of adjacent mouse thin tissue sections of different organs and of various thicknesses by droplet-based surface sampling and by bulk extraction of tissue punches showed that extraction efficiency was incomplete using the former method, and that it depended on the organ and tissue thickness. However, once extraction efficiency was determined and applied, the droplet-based approach provided the required quantitation accuracy and precision for assay validations. Furthermore, this means that once the extraction efficiency was calibrated for a given tissue type and drug, the droplet-based approach provides a non-labor intensive and high-throughput means to acquire spatially resolved quantitative analysis of multiple samples of the same type.« less

Apparatus and methods for manipulation and optimization of biological systems

NASA Technical Reports Server (NTRS)

Sun, Ren (Inventor); Ho, Chih-Ming (Inventor); Wong, Pak Kin (Inventor); Yu, Fuqu (Inventor)

2012-01-01

The invention provides systems and methods for manipulating, e.g., optimizing and controlling, biological systems, e.g., for eliciting a more desired biological response of biological sample, such as a tissue, organ, and/or a cell. In one aspect, systems and methods of the invention operate by efficiently searching through a large parametric space of stimuli and system parameters to manipulate, control, and optimize the response of biological samples sustained in the system, e.g., a bioreactor. In alternative aspects, systems include a device for sustaining cells or tissue samples, one or more actuators for stimulating the samples via biochemical, electromagnetic, thermal, mechanical, and/or optical stimulation, one or more sensors for measuring a biological response signal of the samples resulting from the stimulation of the sample. In one aspect, the systems and methods of the invention use at least one optimization algorithm to modify the actuator's control inputs for stimulation, responsive to the sensor's output of response signals. The compositions and methods of the invention can be used, e.g., to for systems optimization of any biological manufacturing or experimental system, e.g., bioreactors for proteins, e.g., therapeutic proteins, polypeptides or peptides for vaccines, and the like, small molecules (e.g., antibiotics), polysaccharides, lipids, and the like. Another use of the apparatus and methods includes combination drug therapy, e.g. optimal drug cocktail, directed cell proliferations and differentiations, e.g. in tissue engineering, e.g. neural progenitor cells differentiation, and discovery of key parameters in complex biological systems.

Circular RNA hsa_circ_0000745 may serve as a diagnostic marker for gastric cancer.

PubMed

Huang, Mei; He, Yi-Ren; Liang, Li-Chuan; Huang, Qiang; Zhu, Zhi-Qiang

2017-09-14

To determine whether circular RNAs (circRNAs) are involved in pathological processes of gastric cancer (GC). Three circRNAs with differential expression in GC and colorectal cancer were randomly selected for validation by quantitative reverse transcription-polymerase chain reaction (qRT-PCR), using 20 pairs of gastric tissues and normal tissues. Based on the predicted circRNA-miRNA network, we then focused on hsa_circ_0000745, which was found to be down-regulated in 20 GC tissues compared with normal tissues. The hsa_circ_0000745 levels were further analyzed by qRT-PCR in 60 GC tissues and paired adjacent non-tumor tissues, as well as 60 plasma samples from GC patients and 60 plasma samples from healthy controls. The associations between the levels of hsa_circ_0000745 and the clinicopathological features of GC patients were statistically assessed. A receiver operating characteristic (ROC) curve was used to evaluate the diagnostic value of hsa_circ_0000745 in GC. Hsa_circ_0000745 was down-regulated in GC tissues vs non-tumorous tissues ( P < 0.001) and in plasma samples from patients with GC vs healthy controls ( P < 0.001). The expression level of hsa_circ_0000745 in GC tissues correlated with tumor differentiation, while the expression level in plasma correlated with tumor-node-metastasis stage. The area under the ROC curve (AUC) of hsa_circ_0000745 in plasma was 0.683, suggesting good diagnostic value. Plasma hsa_circ_0000745 level combined with carcinoembryogenic antigen (CEA) level increased the AUC to 0.775. Hsa_circ_0000745 plays an important role in GC and its expression level in plasma in combination with CEA level is a promising diagnostic marker for this malignancy.

A validated UPLC-MS/MS method for the analysis of linezolid and a novel oxazolidinone derivative (PH027) in plasma and its application to tissue distribution study in rabbits.

PubMed

Hedaya, Mohsen A; Thomas, Vidhya; Abdel-Hamid, Mohamed E; Kehinde, Elijah O; Phillips, Oludotun A

2017-01-01

Linezolid is the first approved oxazolidinone antibacterial agent, whereas PH027 is a novel compound of the same class that exhibits good in vitro antibacterial activity. The objective of this study was to develop an UPLC-MS/MS assay for the analysis of linezolid and PH027 in plasma and to apply the method for comparative pharmacokinetic and tissue distribution studies of both compounds. Plasma samples and calibrators were extracted with diethyl ether after addition of the internal standard solution. After evaporation of the ether layer, the residue was reconstituted in mobile phase and injected into UPLC-MS/MS. The mobile phase consisted of 2mM ammonium acetate buffer solution and acetonitrile (70:30) at a flow rate of 0.2ml/min. Separation was achieved using UPLC BEH C 18 column, and quantitative determination of the analytes was performed using multiple-reaction monitoring (MRM) scanning mode. The method was validated by analyzing quality control tissue homogenate samples, and was applied to analyze tissue homogenate samples obtained following IV injections of linezolid and PH027 in rabbits. The developed UPLC-MS/MS method was linear in the concentration range of 50-5000ng/ml. Validation of the method proved that the method's precision, selectivity and stability were all within the acceptable limits. Linezolid and PH027 concentrations were accurately determined in the quality control tissue homogenate samples, and analysis of samples obtained following IV administration of the two compounds showed that the tissue to plasma concentration ratio of PH027 was higher than that of linezolid probably due to its higher lipophilicity. The developed UPLC-MS/MS method for the analysis of linezolid and PH027 in rabbit's plasma can accurately determine the concentrations of these compounds in different tissues. Copyright © 2016 Elsevier B.V. All rights reserved.

Quality control in molecular immunohistochemistry

PubMed Central

2008-01-01

Immunoperoxidase histochemistry is a widespread method of assessing expression of biomolecules in tissue samples. Accurate assessment of the expression levels of genes is critical for the management of disease, particularly as therapy targeted to specific molecules becomes more widespread. Determining the quality of preservation of macromolecules in tissue is important to avoid false negative and false positive results. In this review we discuss (1) issues of sensitivity (false negativity) and specificity (false positivity) of immunohistochemical stains, (2) approaches to better understanding differences in immunostains done by different laboratories (including the recently proposed MISFISHIE specification for tissue localization studies), and (3) approaches to assessing the quality of preservation of macromolecules in tissue, particularly in small biopsy samples. PMID:18648842

A Comparison of RNA-Seq Results from Paired Formalin-Fixed Paraffin-Embedded and Fresh-Frozen Glioblastoma Tissue Samples

PubMed Central

Esteve-Codina, Anna; Arpi, Oriol; Martinez-García, Maria; Pineda, Estela; Mallo, Mar; Gut, Marta; Carrato, Cristina; Rovira, Anna; Lopez, Raquel; Tortosa, Avelina; Dabad, Marc; Del Barco, Sonia; Heath, Simon; Bagué, Silvia; Ribalta, Teresa; Alameda, Francesc; de la Iglesia, Nuria

2017-01-01

The molecular classification of glioblastoma (GBM) based on gene expression might better explain outcome and response to treatment than clinical factors. Whole transcriptome sequencing using next-generation sequencing platforms is rapidly becoming accepted as a tool for measuring gene expression for both research and clinical use. Fresh frozen (FF) tissue specimens of GBM are difficult to obtain since tumor tissue obtained at surgery is often scarce and necrotic and diagnosis is prioritized over freezing. After diagnosis, leftover tissue is usually stored as formalin-fixed paraffin-embedded (FFPE) tissue. However, RNA from FFPE tissues is usually degraded, which could hamper gene expression analysis. We compared RNA-Seq data obtained from matched pairs of FF and FFPE GBM specimens. Only three FFPE out of eleven FFPE-FF matched samples yielded informative results. Several quality-control measurements showed that RNA from FFPE samples was highly degraded but maintained transcriptomic similarities to RNA from FF samples. Certain issues regarding mutation analysis and subtype prediction were detected. Nevertheless, our results suggest that RNA-Seq of FFPE GBM specimens provides reliable gene expression data that can be used in molecular studies of GBM if the RNA is sufficiently preserved. PMID:28122052

SU-F-J-193: Efficient Dose Extinction Method for Water Equivalent Path Length (WEPL) of Real Tissue Samples for Validation of CT HU to Stopping Power Conversion

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, R; Baer, E; Jee, K

Purpose: For proton therapy, an accurate model of CT HU to relative stopping power (RSP) conversion is essential. In current practice, validation of these models relies solely on measurements of tissue substitutes with standard compositions. Validation based on real tissue samples would be much more direct and can address variations between patients. This study intends to develop an efficient and accurate system based on the concept of dose extinction to measure WEPL and retrieve RSP in biological tissue in large number of types. Methods: A broad AP proton beam delivering a spread out Bragg peak (SOBP) is used to irradiatemore » the samples with a Matrixx detector positioned immediately below. A water tank was placed on top of the samples, with the water level controllable in sub-millimeter by a remotely controlled dosing pump. While gradually lowering the water level with beam on, the transmission dose was recorded at 1 frame/sec. The WEPL were determined as the difference between the known beam range of the delivered SOBP (80%) and the water level corresponding to 80% of measured dose profiles in time. A Gammex 467 phantom was used to test the system and various types of biological tissue was measured. Results: RSP for all Gammex inserts, expect the one made with lung-450 material (<2% error), were determined within ±0.5% error. Depends on the WEPL of investigated phantom, a measurement takes around 10 min, which can be accelerated by a faster pump. Conclusion: Based on the concept of dose extinction, a system was explored to measure WEPL efficiently and accurately for a large number of samples. This allows the validation of CT HU to stopping power conversions based on large number of samples and real tissues. It also allows the assessment of beam uncertainties due to variations over patients, which issue has never been sufficiently studied before.« less

An examination of the genetic control of Douglas-fir vascular tissue phytochemicals: implications for black bear foraging.

Treesearch

Bruce A. Kimball; G.R. Johnson; Dale L. Nolte; Doreen L. Griffin

1999-01-01

Silvicultural practices can influence black bear (Ursus americanus) foraging preferences for Douglas-fir (Pseudotsuga menziesii) cambial-zone vascular tissues, but little is known about the role of genetics. To study the impact of genetic selection, vascular tissue samples were collected from Douglas-fir trees in six half-sib families from five...

Controlled dexamethasone delivery via double-walled microspheres to enhance long-term adipose tissue retention

PubMed Central

Kelmendi-Doko, Arta; Rubin, J Peter; Klett, Katarina; Mahoney, Christopher; Wang, Sheri; Marra, Kacey G

2017-01-01

Current materials used for adipose tissue reconstruction have critical shortcomings such as suboptimal volume retention, donor-site morbidity, and poor biocompatibility. The aim of this study was to examine a controlled delivery system of dexamethasone to generate stable adipose tissue when mixed with disaggregated human fat in an athymic mouse model for 6 months. The hypothesis that the continued release of dexamethasone from polymeric microspheres would enhance both adipogenesis and angiogenesis more significantly when compared to the single-walled microsphere model, resulting in long-term adipose volume retention, was tested. Dexamethasone was encapsulated within single-walled poly(lactic-co-glycolic acid) microspheres (Dex SW MS) and compared to dexamethasone encapsulated in a poly(lactic-co-glycolic acid) core surrounded by a shell of poly-l-lactide. The double-walled polymer microsphere system in the second model was developed to create a more sustainable drug delivery process. Dexamethasone-loaded poly(lactic-co-glycolic acid) microspheres (Dex SW MS) and dexamethasone-loaded poly(lactic-co-glycolic acid)/poly-l-lactide double-walled microspheres (Dex DW MS) were prepared using single and double emulsion/solvent techniques. In vitro release kinetics were determined. Two doses of each type of microsphere were examined; 50 and 27 mg of Dex MS and Dex DW MS were mixed with 0.3 mL of human lipoaspirate. Additionally, 50 mg of empty MS and lipoaspirate-only controls were examined. Samples were analyzed grossly and histologically after 6 months in vivo. Mass and volume were measured; dexamethasone microsphere-containing samples demonstrated greater adipose tissue retention compared to the control group. Histological analysis, including hematoxylin and eosin and CD31 staining, indicated increased vascularization (p < 0.05) within the Dex MS-containing samples. Controlled delivery of adipogenic factors, such as dexamethasone via polymer microspheres, significantly affects adipose tissue retention by maintaining healthy tissue formation and vascularization. Dex DW MS provide an improved model to former Dex SW MS, resulting in notably longer release time and, consequently, larger volumes of adipose retained in vivo. The use of microspheres, specifically double-walled, as vehicles for controlled drug delivery of adipogenic factors therefore present a clinically relevant model of adipose retention that has the potential to greatly improve soft tissue repair. PMID:29051810

Insight into mechanism of oxidative DNA damage in angiomyolipomas from TSC patients

PubMed Central

Habib, Samy L

2009-01-01

Background The tuberous sclerosis complex (TSC) is caused by defects in one of two tumor suppressor genes, TSC-1 or TSC-2. TSC-2 gene encodes tuberin, a protein involved in the pathogenesis of kidney tumors, both angiomyolipomas and renal cell carcinomas. Loss of heterozygosity at the 8-oxoG-DNA glycosylase (OGG1) allele is found in human kidney clear cell carcinoma identifying loss of OGG1 function as a possible contributor to tumorigenesis in the kidney. Tuberin regulates OGG1 through the transcription factor NF-YA in cultured cells. The purpose of this study is to determine the effect of tuberin-deficiency on OGG1 protein and mRNA levels as well as on 8-oxodG levels in kidney tumors from patients with TSC. In addition we evaluated the phophorylation level of downstream targets of mTOR, phospho-S70K, in kidney tumor tissue from TSC patients. Results Kidney angiomyolipoma tissue from TSC patients expresses significant levels of phopho-tuberin and low levels of tuberin compared to control kidney tissue. The increase in tuberin phosphorylation and the decrease tuberin expression are associated with decrease in OGG1 protein and mRNA levels in tumor samples compared to normal kidney samples. The decrease OGG1 expression is also associated with significant decrease in the transcription factor, NF-YA, expression in tumor samples compared to normal tissues. In addition, the levels of 8-oxodG are 4-fold higher in tumors compared to control samples. The significant increase of phospho-tuberin expression is associated with increase phosphorylation of S6K in tumor samples compared to controls. Cyclin D1 expression is also 3-fold higher in increase in the tumor tissues compared to normal kidney tissues. Conclusion These data indicate that tuberin deficiency in angiomyolipoma enhances mTOR activation by phosphorylation of S6K and downregulation of protein and mRNA expression of OGG1 resulted in accumulation of oxidized DNA in patients with TSC. These data suggest that tuberin and OGG1 are important proteins in the pathogenesis of angiomyolipoma in TSC patients. PMID:19265534

Transforming growth factor beta-1 expression in macrophages of human chronic periapical diseases.

PubMed

Liang, Z-Z; Li, J; Huang, S-G

2017-03-30

The objective of this study was to observe the distribution of macrophages (MPs) expressing transforming growth factor beta-1 (TGF-β1) in tissue samples from patients with different human chronic periapical diseases. In this study, samples were collected from 75 volunteers, who were divided into three groups according to classified standards, namely, healthy control (N = 25), periapical granuloma (N = 25), and periapical cyst (N = 25). The samples were fixed in 10% buffered formalin for more than 48 h, dehydrated, embedded, and stained with hematoxylin and eosin for histopathology. Double immunofluorescence was conducted to analyze the expression of TGF-β-CD14 double-positive MPs in periapical tissues. The number of double-positive cells (cells/mm 2 ) were significantly higher in the chronic periapical disease tissues (P < 0.01) compared to that in the control tissue; in addition, the density of TGF-β1-CD14 double positive cells was significantly higher in the periapical cyst group than in the periapical granuloma group (P < 0.01). The number of TGF-β1 expressing macrophages varied with human chronic periapical diseases. The TGF-β1-CD14 double-positive cells might play an important role in the pathology of human chronic periapical diseases.

Electroosmotic Sampling. Application to Determination of Ectopeptidase Activity in Organotypic Hippocampal Slice Cultures

PubMed Central

Xu, Hongjuan; Guy, Yifat; Hamsher, Amy; Shi, Guoyue; Sandberg, Mats; Weber, Stephen G.

2010-01-01

We hypothesize that peptide-containing solutions pulled through tissue should reveal the presence and activity of peptidases in the tissue. Using the natural ζ-potential in the organotypic hippocampal slice culture (OHSC), physiological fluids can be pulled through the tissue with an electric field. The hydrolysis of the peptides present in the fluid drawn through the tissue can be determined using capillary HPLC with electrochemical detection of the biuret complexes of the peptides following a postcolumn reaction. We have characterized this new sampling method by measuring the flow rate, examining the use of internal standards, and examining cell death caused by sampling. The sampling flow rate ranges from 60 to 150 nL/min with a 150 μm (ID) sampling capillary with an electric field (at the tip of the capillary) from 30 to 60 V/cm. Cell death can be negligible with controlled sampling conditions. Using this sampling approach, we have electroosmotically pulled Leu-enkephalin through OHSCs to identify ectopeptidase activity in the CA3 region. These studies show that a bestatin-sensitive aminopeptidase may be critical for the hydrolysis of exogenous Leu-enkephalin, a neuropeptide present in the CA3 region of OHSCs. PMID:20669992

The Perspectives of Haematological Cancer Patients on Tissue Banking.

PubMed

Turon, Heidi; Waller, Amy; Clinton-McHarg, Tara; Boyes, Allison; Fleming, Jennifer; Marlton, Paula; Harrison, Simon J; Sanson-Fisher, Rob

2016-01-01

A high level of support for tissue banking has been identified amongst both the general public and patients. However, much debate remains about the regulatory framework of tissue banks. This study explored the views of haematological cancer patients regarding tissue banking and how tissue banks should operate. Haematological cancer patients from three outpatient clinics in Australia completed a questionnaire examining their preferences for tissue banking as well as items about their sociodemographic characteristics, disease and treatment history. The majority of participants (95%) reported being willing to allow their leftover tissue to be used for medical research. Three quarters (76%) supported the idea of their medical record being linked to their tissue sample, and 77% preferred a blanket (one-off) consent model for future research use of their tissue sample. Only 57 (27%) participants had been asked to give a tissue sample for research, 98% of whom gave permission. The majority of haematological cancer patients are willing to donate their leftover tissue to a tissue bank and have their medical records linked to tissue samples and prefer a one-off consent process. These novel data from potential donors inform the debate about how tissue banks might operate. Strategic Research Partnership Grant from the Cancer Council NSW to the Newcastle Cancer Control Collaborative (New-3C) and infrastructure funding from the Hunter Medical Research Institute (HMRI). A.W. is supported by an Australian Research Council DECRA fellowship (DE150101262). T.C.M. was supported by a Leukaemia Foundation of Queensland Post-Doctoral Fellowship. A.B. is supported by National Health and Medical Research Council (APP1073317) and Cancer Institute NSW (13/ECF/1-37) Early Career Fellowships.

UV-laser microdissection and mRNA expression analysis of individual neurons from postmortem Parkinson's disease brains.

PubMed

Gründemann, Jan; Schlaudraff, Falk; Liss, Birgit

2011-01-01

Cell specificity of gene expression analysis is essential to avoid tissue sample related artifacts, in particular when the relative number of target cells present in the compared tissues varies dramatically, e.g., when comparing dopamine neurons in midbrain tissues from control subjects with those from Parkinson's disease (PD) cases. Here, we describe a detailed protocol that combines contact-free UV-laser microdissection and quantitative PCR of reverse-transcribed RNA of individual neurons from postmortem human midbrain tissue from PD patients and unaffected controls. Among expression changes in a variety of dopamine neuron marker, maintenance, and cell-metabolism genes, we found that α-synuclein mRNA levels were significantly elevated in individual neuromelanin-positive dopamine midbrain neurons from PD brains when compared to those from matched controls.

Ex-vivo holographic microscopy and spectroscopic analysis of head and neck cancer

NASA Astrophysics Data System (ADS)

Holler, Stephen; Wurtz, Robert; Auyeung, Kelsey; Auyeung, Kris; Paspaley-Grbavac, Milan; Mulroe, Brigid; Sobrero, Maximiliano; Miles, Brett

2015-03-01

Optical probes to identify tumor margins in vivo would greatly reduce the time, effort and complexity in the surgical removal of malignant tissue in head and neck cancers. Current approaches involve visual microscopy of stained tissue samples to determine cancer margins, which results in the excision of excess of tissue to assure complete removal of the cancer. Such surgical procedures and follow-on chemotherapy can adversely affect the patient's recovery and subsequent quality of life. In order to reduce the complexity of the process and minimize adverse effects on the patient, we investigate ex vivo tissue samples (stained and unstained) using digital holographic microscopy in conjunction with spectroscopic analyses (reflectance and transmission spectroscopy) in order to determine label-free, optically identifiable characteristic features that may ultimately be used for in vivo processing of cancerous tissues. The tissue samples studied were squamous cell carcinomas and associated controls from patients of varying age, gender and race. Holographic microscopic imaging scans across both cancerous and non-cancerous tissue samples yielded amplitude and phase reconstructions that were correlated with spectral signatures. Though the holographic reconstructions and measured spectra indicate variations even among the same class of tissue, preliminary results indicate the existence of some discriminating features. Further analyses are presently underway to further this work and extract additional information from the imaging and spectral data that may prove useful for in vivo surgical identification.

Culture and Sampling of Primary Adipose Tissue in Practical Microfluidic Systems.

PubMed

Brooks, Jessica C; Judd, Robert L; Easley, Christopher J

2017-01-01

Microfluidic culture of primary adipose tissue allows for reduced sample and reagent volumes as well as constant media perfusion of the cells. By continuously flowing media over the tissue, microfluidic sampling systems can more accurately mimic vascular flow in vivo. Quantitative measurements can be performed on or off chip to provide time-resolved secretion data, furthering insight into the dynamics of the function of adipose tissue. Buoyancy resulting from the large lipid storage capacity in this tissue presents a unique challenge for culture, and it is important to account for this buoyancy during microdevice design. Herein, we describe approaches for microfluidic device fabrication that utilize 3D-printed interface templating to help counteract cell buoyancy. We apply such methods to the culture of both isolated, dispersed primary adipocytes and epididymal adipose explants. To facilitate more widespread adoption of the methodology, the devices presented here are designed for user-friendly operation. Only handheld syringes are needed to control flow, and devices are inexpensive and disposable.

Systematic bias in genomic classification due to contaminating non-neoplastic tissue in breast tumor samples.

PubMed

Elloumi, Fathi; Hu, Zhiyuan; Li, Yan; Parker, Joel S; Gulley, Margaret L; Amos, Keith D; Troester, Melissa A

2011-06-30

Genomic tests are available to predict breast cancer recurrence and to guide clinical decision making. These predictors provide recurrence risk scores along with a measure of uncertainty, usually a confidence interval. The confidence interval conveys random error and not systematic bias. Standard tumor sampling methods make this problematic, as it is common to have a substantial proportion (typically 30-50%) of a tumor sample comprised of histologically benign tissue. This "normal" tissue could represent a source of non-random error or systematic bias in genomic classification. To assess the performance characteristics of genomic classification to systematic error from normal contamination, we collected 55 tumor samples and paired tumor-adjacent normal tissue. Using genomic signatures from the tumor and paired normal, we evaluated how increasing normal contamination altered recurrence risk scores for various genomic predictors. Simulations of normal tissue contamination caused misclassification of tumors in all predictors evaluated, but different breast cancer predictors showed different types of vulnerability to normal tissue bias. While two predictors had unpredictable direction of bias (either higher or lower risk of relapse resulted from normal contamination), one signature showed predictable direction of normal tissue effects. Due to this predictable direction of effect, this signature (the PAM50) was adjusted for normal tissue contamination and these corrections improved sensitivity and negative predictive value. For all three assays quality control standards and/or appropriate bias adjustment strategies can be used to improve assay reliability. Normal tissue sampled concurrently with tumor is an important source of bias in breast genomic predictors. All genomic predictors show some sensitivity to normal tissue contamination and ideal strategies for mitigating this bias vary depending upon the particular genes and computational methods used in the predictor.

Effects of spaceflight on the spermatogonial population of rat seminiferous epithelium

NASA Technical Reports Server (NTRS)

Sapp, Walter J.; Philpott, Delbert E.; Williams, Carol S.; Kato, Katharine; Stevenson, Joann; Vasquez, M.; Serova, L. V.

1990-01-01

Testes from rats flown on Cosmos 1887 were compared with vivarium control and synchronous control samples. The mean weights of flight testes, normalized for weight per 100 g, were 6.4 percent less when compared with the vivarium controls. Counts of spermatogonia from tissue sections (seminiferous tubules in maturation stage 6) from five animals in each group revealed 4 percent fewer spermatogonia in flight testes compared with synchronous controls and 11 percent fewer spermatogonia in flight samples compared with vivarium controls.

Improving tissue preparation for matrix-assisted laser desorption ionization mass spectrometry imaging. Part 1: using microspotting.

PubMed

Franck, Julien; Arafah, Karim; Barnes, Alan; Wisztorski, Maxence; Salzet, Michel; Fournier, Isabelle

2009-10-01

Nowadays, matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI MSI) is a powerful technique to obtain the distribution of endogenous and exogenous molecules within tissue sections. It can, thus, be used to study the evolution of molecules across different physiological stages in order to find out markers or get knowledge on signaling pathways. In order to provide valuable information, we must carefully control the sample preparation to avoid any delocalization of molecules of interest inside the tissue during this step. Currently, two strategies can be used to deposit chemicals, such as the MALDI matrix, onto the tissue both involving generation of microdroplets that will be dropped off onto the surface. First strategy involves microspraying of solutions. Here, we have been interested in the development of a microspotting strategy, where nanodroplets of solvent are ejected by a piezoelectric device to generate microspots at the tissue level. Such systems allow one to precisely control sample preparation by creating an array of spots. In terms of matrix crystallization, a microspotting MALDI matrix is hardly compatible with the results by classical (pipetting) methods. We have thus synthesized and studied new solid ionic matrixes in order to obtain high analytical performance using such a deposition system. These developments have enabled optimization of the preparation time because of the high stability of the printing that is generated in these conditions. We have also studied microspotting for performing on-tissue digestion in order to go for identification of proteins or to work from formalin fixed and paraffin embedded (FFPE) tissue samples. We have shown that microspotting is an interesting approach for on tissue digestion. Peptides, proteins, and lipids were studied under this specific preparation strategy to improve imaging performances for this class of molecules.

The Role of Intrinsic Pathway in Apoptosis Activation and Progression in Peyronie's Disease

PubMed Central

Loreto, Carla; Caltabiano, Rosario; Vespasiani, Giuseppe; Castorina, Sergio; Ralph, David J.; Musumeci, Giuseppe; Djinovic, Rados; Sansalone, Salvatore

2014-01-01

Peyronie's disease (PD) is characterized with formation of fibrous plaques which result in penile deformity, pain, and erectile dysfunction. The aim of this study was to investigate the activation of the intrinsic apoptotic pathway in plaques from PD patients. Tunica albuginea from either PD or control patients was assessed for the expression of bax, bcl-2 and caspases 9 and 3 using immunohistochemistry and by measurement of apoptotic cells using TUNEL assay. Bax overexpression was observed in metaplastic bone tissue, in fibroblasts, and in myofibroblast of plaques from PD patients. Little or no bcl-2 immunostaining was detected in samples from either patients or controls. Caspase 3 immunostaining was very strong in fibrous tissue, in metaplasic bone osteocytes, and in primary ossification center osteoblasts. Moderate caspase 9 immunostaining was seen in fibrous cells plaques and in osteocytes and osteoblasts of primary ossification centers from PD patients. Control samples were negative for caspase 9 immunostaining. In PD patients the TUNEL immunoassay showed intense immunostaining of fibroblasts and myofibroblasts, the absence of apoptotic cells in metaplasic bone tissue and on the border between fibrous and metaplastic bone tissue. Apoptosis occurs in stabilized PD plaques and is partly induced by the intrinsic pathway. PMID:25197653

Transcript levels of ten-eleven translocation type 1–3 in cervical cancer and non-cancerous cervical tissues

PubMed Central

Bronowicka-Kłys, Dorota Ewa; Roszak, Andrzej; Pawlik, Piotr; Sajdak, Stefan; Sowińska, Anna; Jagodziński, Paweł Piotr

2017-01-01

Decreased expression of ten-eleven translocation (TET1, TET2 and TET3) proteins has been reported in various types of cancer. However, the expression levels of TET proteins in cervical cancer (CC) remain to be elucidated. The present study determined the levels of TET1, TET2 and TET3 transcripts in cancerous (n=80) and non-cancerous cervical tissues (n=41). The results revealed a significant reduction in TET1 transcripts (P=0.0000001) in cervical tissue samples from patients with primary CC compared with samples from control patients. Significantly decreased TET1 transcript levels, as compared to non-cancerous cervical tissues, were also observed in tissue samples with the following characteristics: Stage I (P=0.016), II (P<0.0001), III (P=0.00007) and grade of differentiation G1 (P=0.026), G2 (P=0.00006), G3 (P=0.0007) and Gx (P=0.0004) and squamous histological type (P<0.00001). TET1 transcript levels were significantly lower in patients aged 45–60 years (P=0.0002) and patients age >60 years (P=0.003), as compared with non-cancerous cervical tissues. TET2 transcript levels were lower in cervical cancer tissues classified as stage II (P=0.043) and TET3 transcript levels were lower in stage III samples (P=0.010), tissue samples with a grade of differentiation of G3 (P=0.025) and tissue with squamous type histology (P=0.047), all compared with non-cancerous cervical tissues. The present study demonstrated a significantly reduced level of TET1 transcripts in cancerous cervical tissues, as compared with non-cancerous tissues. Furthermore, decreased TET1-3 transcript levels were identified when patients with CC were stratified by clinicopathological variables, as compared with non-cancerous cervical tissues. PMID:28521490

Detection and Clinicopathologic Correlation of Human Immunodeficiency Virus (HIV-1) Nucleic Acids and Antigens in Reticuloendothelial and Central Nervous System Tissues, by Immunohistochemistry, in situ Hybridization, and Polymerase Chain Reaction

DTIC Science & Technology

1992-09-30

mantle zones characteristic of HIV-I infection was absent. 5 Giant cells were absent in lynphoid tissues of all seronegative individuals. Warthin ...control samples. Warthin Finckeldey syncytial cells, which are present in hyperplastic or atrophic nodes, were absent in non-infected tissues

Elevated expression of matrix metalloproteinase-9 is associated with bladder cancer pathogenesis.

PubMed

Wu, Gong-Jin; Bao, Jun-Sheng; Yue, Zhong-Jin; Zeng, Fan-Chang; Cen, Song; Tang, Zheng-Yan; Kang, Xin-Li

2018-01-01

This study investigated the association between abnormal matrix metalloproteinase-9 (MMP-9) expression and bladder cancer (BC) development. In a retrospective analysis, this study used tissue samples derived from 92 patients pathologically diagnosed with BC (experimental group), who were hospitalized between September 2012 and June 2014 at the Urinary Surgery of Department of Urology, Lanzhou University Second Hospital. As controls (control group), 63 normal pericancerous bladder mucosal tissues (3 cm distant form edge of BC foci) with confirmed pathology were selected from the same time period. Immunohistochemistry was employed to detect MMP-9 protein expression in the tissues and enzyme-linked immunosorbent assay was performed to measure MMP-9 protein levels in tissue samples of patients and control subjects. Finally, a meta-analysis was conducted to understand the overall impact of MMP-9 on BC pathogenesis. STATA 12.0 software (Stata Corp, College Station, TX, USA) was used for all statistical analyses. The MMP-9 positive expression rate in tissue samples and MMP-9 levels were significantly greater in the experimental group compared to the control group (both P < 0.001). The frequency of MMP-9 positive status showed statistically significant differences between G1 (low-grade) and G3 (high-grade) (P < 0.001), between G2 and G3 (P < 0.05), and between G1/G2 and G3 (P = 0.001). Our meta-analysis findings provided further evidence that MMP-9 positive expression status and MMP-9 levels in the experimental group were significantly higher than the control group (positive expressions: Odds ratio [OR] = 18.59, 95% confidence interval [95% CI] = 11.63-29.71, P < 0.001; expression levels: Standard mean difference = 1.51, 95%CI = 0.63-2.39, P = 0.001). The positive expression status of MMP-9 was notably lower in G1/G2 compared to G3 (OR = 0.24, 95%CI = 0.15-0.36, P < 0.001). Our study demonstrated that both positive expression status in tumor tissue and expression levels of MMP-9 are significantly elevated in BC patients and correlate with disease progression. Thus, MMP-9 can serve as a biomarker to determine the degree of BC malignancy.

Blended shared control utilizing online identification : Regulating grasping forces of a surrogate surgical grasper.

PubMed

Stephens, Trevor K; Kong, Nathan J; Dockter, Rodney L; O'Neill, John J; Sweet, Robert M; Kowalewski, Timothy M

2018-06-01

Surgical robots are increasingly common, yet routine tasks such as tissue grasping remain potentially harmful with high occurrences of tissue crush injury due to the lack of force feedback from the grasper. This work aims to investigate whether a blended shared control framework which utilizes real-time identification of the object being grasped as part of the feedback may help address the prevalence of tissue crush injury in robotic surgeries. This work tests the proposed shared control framework and tissue identification algorithm on a custom surrogate surgical robotic grasping setup. This scheme utilizes identification of the object being grasped as part of the feedback to regulate to a desired force. The blended shared control is arbitrated between human and an implicit force controller based on a computed confidence in the identification of the grasped object. The online identification is performed using least squares based on a nonlinear tissue model. Testing was performed on five silicone tissue surrogates. Twenty grasps were conducted, with half of the grasps performed under manual control and half of the grasps performed with the proposed blended shared control, to test the efficacy of the control scheme. The identification method resulted in an average of 95% accuracy across all time samples of all tissue grasps using a full leave-grasp-out cross-validation. There was an average convergence time of [Formula: see text] ms across all training grasps for all tissue surrogates. Additionally, there was a reduction in peak forces induced during grasping for all tissue surrogates when applying blended shared control online. The blended shared control using online identification more successfully regulated grasping forces to the desired target force when compared with manual control. The preliminary work on this surrogate setup for surgical grasping merits further investigation on real surgical tools and with real human tissues.

An allogenic cell-based implant for meniscal lesions.

PubMed

Weinand, Christian; Peretti, Giuseppe M; Adams, Samuel B; Bonassar, Lawrence J; Randolph, Mark A; Gill, Thomas J

2006-11-01

Meniscal tears in the avascular zones do not heal. Although tissue-engineering approaches using cells seeded onto scaffolds could expand the indication for meniscal repair, harvesting autologous cells could cause additional trauma to the patient. Allogenic cells, however, could provide an unlimited amount of cells. Allogenic cells from 2 anatomical sources can repair lesions in the avascular region of the meniscus. Controlled laboratory study. Both autologous and allogenic chondrocytes were seeded onto a Vicryl mesh scaffold and sutured into a bucket-handle lesion created in the medial menisci of 17 swine. Controls consisted of 3 swine knees treated with unseeded implants and controls from a previous experiment in which 4 swine were treated with suture only and 4 with no treatment. Menisci were harvested after 12 weeks and evaluated histologically for new tissue and percentage of interface healing surface; they were also evaluated statistically. The lesions were closed in 15 of 17 menisci. None of the control samples demonstrated healing. Histologic analysis of sequential cuts through the lesion showed formation of new scar-like tissue in all experimental samples. One of 8 menisci was completely healed in the allogenic group and 2 of 9 in the autologous group; the remaining samples were partially healed in both groups. No statistically significant differences in the percentage of healing were observed between the autologous and allogenic cell-based implants. Use of autologous and allogenic chondrocytes delivered via a biodegradable mesh enhanced healing of avascular meniscal lesions. This study demonstrates the potential of a tissue-engineered cellular repair of the meniscus using autologous and allogenic chondrocytes.

p16 promoter hypermethylation: A useful serum marker for early detection of gastric cancer

PubMed Central

Abbaszadegan, Mohammad Reza; Moaven, Omeed; Sima, Hamid Reza; Ghafarzadegan, Kamran; A'rabi, Azadeh; Forghani, Mohammad Naser; Raziee, Hamid Reza; Mashhadinejad, Ali; Jafarzadeh, Mostafa; Esmaili-Shandiz, Ehsan; Dadkhah, Ezzat

2008-01-01

AIM: To determine p16 promoter hypermethylation in gastric tumoral tissue and serum samples, its impact on p16-protein expression, and correlation with clinical and histological features. METHODS: Samples were obtained from 52 histologically confirmed cases of gastric adenocarcinoma. Gastric tissue and serum of 50 age- and sex-matched individuals with normal gastroscopy and biopsy were obtained as control samples. Methylation-specific polymerase chain reaction (MSP) was used to evaluate methylation status of p16 promoter. p16-protein expression was analyzed by immunohistochemical staining on paraffin-embedded sections. RESULTS: Methylation was detected in 44.2% (23/52) of tumoral tissues. 60.9% of them were also methylated in serum, i.e., 26.9% of all patients (14/52). Methylation was not detected in tissue and sera of control samples. p16-protein expression was decreased in 61.5% of cases (32/52), and was significantly associated with promoter hypermethylation (P < 0.001). Methylation was significantly more frequent in higher pathological grades (P < 0.05). Methylation was not associated with other clinicopathological features and environmental factors including H pylori infection and smoking. CONCLUSION: p16 promoter hypermethylation is an important event in gastric carcinogenesis. It is the principle mechanism of p16 gene silencing. It is related to malignant tumor behavior. Detection of DNA methylation in serum may be a biomarker for early detection of gastric cancer. PMID:18395906

Elevated interleukin-1β in peripheral blood mononuclear cells contributes to the pathogenesis of autoimmune thyroid diseases, especially of Hashimoto thyroiditis.

PubMed

Sun, Li; Zhang, Xiaoxu; Dai, Fang; Shen, Jijia; Ren, Cuiping; Zuo, Chunlin; Zhang, Qiu

2016-08-01

To explore the relationship between IL-1β expression and two common autoimmune thyroid diseases: Hashimoto thyroiditis (HT) and Graves' disease (GD). qRT-PCR, Quantiglo ELISA, and flow cytometry were used to evaluate the expression levels of IL-1β in serum, peripheral blood mononuclear cells (PBMCs), and thyroid tissue samples from patients with HT or GD. Local infiltration of monocytes was assessed by immunohistochemical study of patients' thyroid tissue samples. Although no significant differences in IL-1β levels were found between samples of serum from patients with HT or GD and normal controls, we found that IL-1β mRNA and protein levels in PBMCs of HT patients were significantly higher than those of patients with GD, which were in turn higher than the level in normal controls. In addition, IL-1β mRNA was also increased in thyroid gland tissue from patients with HT compared to those with GD, and this was accompanied by increased local infiltration of monocytes into thyroid tissues. Correlation analysis of the clinical samples validated the association of high IL-1β levels with the pathogenesis of HT. Our study suggests that IL-1β may be an active etiologic factor in the pathogenesis of HT and thus present a new target for novel diagnostics and treatment.

Stability of Bovine viral diarrhea virus 1 nucleic acid in fetal bovine samples stored under different conditions.

PubMed

Ridpath, Julia F; Neill, John D; Chiang, Yu-Wei; Waldbillig, Jill

2014-01-01

Infection of pregnant cattle with both species of Bovine viral diarrhea virus (BVDV) can result in reproductive disease that includes fetal reabsorption, mummification, abortion, stillbirths, congenital defects affecting structural, neural, reproductive, and immune systems, and the birth of calves persistently infected with BVDV. Accurate diagnosis of BVDV-associated reproductive disease is important to control BVDV at the production unit level and assessment of the cost of BVDV infections in support of BVDV control programs. The purpose of the current study was to examine the stability of viral nucleic acid in fetal tissues exposed to different conditions, as measured by detection by polymerase chain reaction. Five different types of fetal tissue, including brain, skin and muscle, ear, and 2 different pooled organ samples, were subjected to conditions that mimicked those that might exist for samples collected after abortions in production settings or possible storage conditions after collection and prior to testing. In addition, tissues were archived for 36 months at -20°C and then retested, to mimic conditions that might occur in the case of retrospective surveillance studies. Brain tissue showed the highest stability under the conditions tested. The impact of fecal contamination was increased following archiving in all tissue types suggesting that, for long-term storage, effort should be made to reduce environmental contaminants before archiving.

Comparative biomarker expression and RNA integrity in biospecimens derived from radical retropubic and robot-assisted laparoscopic prostatectomies.

PubMed

Ricciardelli, Carmela; Bianco-Miotto, Tina; Jindal, Shalini; Dodd, Thomas J; Cohen, Penelope A; Marshall, Villis R; Sutherland, Peter D; Samaratunga, Hemamali; Kench, James G; Dong, Ying; Wang, Hong; Clements, Judith A; Risbridger, Gail P; Sutherland, Robert L; Tilley, Wayne D; Horsfall, David J

2010-07-01

Knowledge of preanalytic conditions that biospecimens are subjected to is critically important because novel surgical procedures, tissue sampling, handling, and storage might affect biomarker expression or invalidate tissue samples as analytes for some technologies. We investigated differences in RNA quality, gene expression by quantitative real-time PCR, and immunoreactive protein expression of selected prostate cancer biomarkers between tissues from retropubic radical prostatectomy (RRP) and robot-assisted laparoscopic prostatectomy (RALP). Sections of tissue microarray of 23 RALP and 22 RRP samples were stained with antibodies to androgen receptor (AR) and prostate-specific antigen (PSA) as intersite controls, and 14 other candidate biomarkers of research interest to three laboratories within the Australian Prostate Cancer BioResource tissue banking network. Quantitative real-time PCR was done for AR, PSA (KLK3), KLK2, KLK4, and HIF1A on RNA extracted from five RALP and five RRP frozen tissue cores. No histologic differences were observed between RALP and RRP tissue. Biomarker staining grouped these samples into those with increased (PSA, CK8/18, CKHMW, KLK4), decreased (KLK2, KLK14), or no change in expression (AR, ghrelin, Ki67, PCNA, VEGF-C, PAR2, YB1, p63, versican, and chondroitin 0-sulfate) in RALP compared with RRP tissue. No difference in RNA quality or gene expression was detected between RALP and RRP tissue. Changes in biomarker expression between RALP and RRP tissue exist at the immunoreactive protein level, but the etiology is unclear. Future studies should account for changes in biomarker expression when using RALP tissues, and mixed cohorts of RALP and RRP tissue should be avoided.

Bacterial contamination of tissue allografts - experiences of the donor tissue bank of Victoria.

PubMed

Ireland, Lyn; Spelman, Denis

2005-01-01

The aim of this study is to report the experience of the Donor Tissue Bank of Victoria with bacteria isolated from musculoskeletal, skin and cardiac allografts retrieved from cadaveric donors. The results of all quality control samples for bacterial culture, taken during retrieval and processing of allografts at the DTBV for a 12 month period, were extracted and analysed. It was found that 15.7% of skin, 15.1% of heart valves and 5.8% of musculoskeletal samples had positive culture results. The number and types of organisms isolated varied with tissue type. The most commonly isolated organisms were Staphylococcus species (including S. aureus). The identity of the isolate and the number of positive specimens from the same donor were considerations in the decision concerning the suitability of tissue for subsequent implantation.

Biological response in vitro of skeletal muscle cells treated with different intensity continuous and pulsed ultrasound fields

NASA Astrophysics Data System (ADS)

Abrunhosa, Viviane M.; Mermelstein, Claudia S.; Costa, Manoel L.; Costa-Felix, Rodrigo P. B.

2011-02-01

Therapeutic ultrasound has been used in physiotherapy to accelerate tissue healing. Although the ultrasonic wave is widely used in clinical practice, not much is known about the biological effects of ultrasound on cells and tissues. This study aims to evaluate the biological response of ultrasound in primary cultures of chick myogenic cells. To ensure the metrological reliability of whole measurement process, the ultrasound equipment was calibrated in accordance with IEC 61689:2007. The skeletal muscle cells were divided in four samples. One sample was used as a control group and the others were submitted to different time and intensity and operation mode of ultrasound: 1) 0.5 W/cm2 continuous for 5 minutes, 2) 0.5 W/cm2 pulsed for 5 minutes, 3) 1.0 W/cm2 pulsed for 10 minutes. The samples were analyzed with phase contrast optical microscopy before and after the treatment. The results showed alignment of myogenic cells in the sample treated with 0.5 W/cm2 continuous during 5 minutes when compared with the control group and the other samples. This study is a first step towards a metrological and scientific based protocol to cells and tissues treatment under different ultrasound field exposures.

Evaluation of the Xpert MTB/RIF Performance on Tissues: Potential Impact on Airborne Infection Isolation at a Tertiary Cancer Care Center.

PubMed

McMillen, Tracy; Usiak, Shauna C; Chen, Liang Hua; Gomez, Luz; Ntiamoah, Peter; Hameed, Meera R; Budvytiene, Indre; Banaei, Niaz; Kamboj, Mini; Babady, N Esther

2018-04-01

OBJECTIVES In this study, we sought to evaluate the performance of the Xpert MTB/RIF (Cepheid) assay for the detection of Mycobacterium tuberculosis (MTB) complex DNA on fresh and formalin-fixed, paraffin-embedded (FFPE) tissue specimens from oncology patients in an area with a low prevalence of tuberculosis. We also aimed to retrospectively assess the potential impact of Xpert MTB/RIF on the duration of airborne infection isolation (AII). SETTING A 473-bed, tertiary-care cancer center in New York City. DESIGN A total of 203 tissue samples (101 FFPE and 102 fresh) were tested using Xpert MTB/RIF, including 133 pulmonary tissue samples (65.5%) and 70 extrapulmonary tissue samples (34.5%). Acid-fast bacilli (AFB) culture was used as the diagnostic gold standard. The limit of detection (LOD) and reproducibility were also evaluated for both samples types using contrived specimens. The potential impact of the Xpert MTB PCR assay on tissue samples from AII patients on AII duration was retrospectively assessed. RESULTS Using the Xpert MTB/RIF for fresh tissue specimens, the sensitivity was 50% (95% CI, 1.3%-98.7%) and the specificity was 99% (95% CI, 94.5%-99.9%). For FFPE tissue specimens, the sensitivity was 100% (95% CI, 63.1%-100%) and the specificity was 98.3% (95% CI, 95.5%-100%. The LOD was 103 colony-forming units (CFU)/mL for both fresh and FFPE tissue specimens, and the Xpert MTB/RIF was 100% reproducible at concentrations 10 times that of the LOD. With an expected turnaround time of 24 hours, the Xpert MTB PCR could decrease the duration of AII from a median of 8 days to a median of 1 day. CONCLUSIONS The Xpert MTB/RIF assay offers a valid option for ruling out Mycobacterium tuberculosis complex (MTBC) on tissue samples from oncology patients and for minimizing AII resource utilization. Infect Control Hosp Epidemiol 2018;39:462-466.

International Cartilage Repair Society (ICRS) Recommended Guidelines for Histological Endpoints for Cartilage Repair Studies in Animal Models and Clinical Trials

PubMed Central

Hoemann, Caroline; Kandel, Rita; Roberts, Sally; Saris, Daniel B.F.; Creemers, Laura; Mainil-Varlet, Pierre; Méthot, Stephane; Hollander, Anthony P.; Buschmann, Michael D.

2011-01-01

Cartilage repair strategies aim to resurface a lesion with osteochondral tissue resembling native cartilage, but a variety of repair tissues are usually observed. Histology is an important structural outcome that could serve as an interim measure of efficacy in randomized controlled clinical studies. The purpose of this article is to propose guidelines for standardized histoprocessing and unbiased evaluation of animal tissues and human biopsies. Methods were compiled from a literature review, and illustrative data were added. In animal models, treatments are usually administered to acute defects created in healthy tissues, and the entire joint can be analyzed at multiple postoperative time points. In human clinical therapy, treatments are applied to developed lesions, and biopsies are obtained, usually from a subset of patients, at a specific time point. In striving to standardize evaluation of structural endpoints in cartilage repair studies, 5 variables should be controlled: 1) location of biopsy/sample section, 2) timing of biopsy/sample recovery, 3) histoprocessing, 4) staining, and 5) blinded evaluation with a proper control group. Histological scores, quantitative histomorphometry of repair tissue thickness, percentage of tissue staining for collagens and glycosaminoglycan, polarized light microscopy for collagen fibril organization, and subchondral bone integration/structure are all relevant outcome measures that can be collected and used to assess the efficacy of novel therapeutics. Standardized histology methods could improve statistical analyses, help interpret and validate noninvasive imaging outcomes, and permit cross-comparison between studies. Currently, there are no suitable substitutes for histology in evaluating repair tissue quality and cartilaginous character. PMID:26069577

Multimodal optical analysis discriminates freshly extracted human sample of gliomas, metastases and meningiomas from their appropriate controls

NASA Astrophysics Data System (ADS)

Zanello, Marc; Poulon, Fanny; Pallud, Johan; Varlet, Pascale; Hamzeh, H.; Abi Lahoud, Georges; Andreiuolo, Felipe; Ibrahim, Ali; Pages, Mélanie; Chretien, Fabrice; di Rocco, Federico; Dezamis, Edouard; Nataf, François; Turak, Baris; Devaux, Bertrand; Abi Haidar, Darine

2017-02-01

Delineating tumor margins as accurately as possible is of primordial importance in surgical oncology: extent of resection is associated with survival but respect of healthy surrounding tissue is necessary for preserved quality of life. The real-time analysis of the endogeneous fluorescence signal of brain tissues is a promising tool for defining margins of brain tumors. The present study aims to demonstrate the feasibility of multimodal optical analysis to discriminate fresh samples of gliomas, metastases and meningiomas from their appropriate controls. Tumor samples were studied on an optical fibered endoscope using spectral and fluorescence lifetime analysis and then on a multimodal set-up for acquiring spectral, one and two-photon fluorescence images, second harmonic generation signals and two-photon fluorescence lifetime datasets. The obtained data allowed us to differentiate healthy samples from tumor samples. These results confirmed the possible clinical relevance of this real-time multimodal optical analysis. This technique can be easily applied to neurosurgical procedures for a better delineation of surgical margins.

Further factors influencing N-nitrosamine formation in bacon.

PubMed

Gray, J I; Skrypec, D J; Mandagere, A K; Booren, A M; Pearson, A M

1984-01-01

The possible relationship of unsaturated fatty acids in adipose tissue to the formation of N-nitrosamines in bacon was evaluated by trials in which pigs were fed regular (control), tallow-, coconut fat- and corn oil-supplemented diets. Bacon prepared from pigs fed corn oil-supplemented diets contained significantly higher levels of N-nitrosopyrrolidine and N-nitrosodimethylamine than did control bacon samples; however, bacon produced from pigs fed a coconut fat-supplemented diet contained significantly lower levels of N-nitrosopyrrolidine. Fatty acid analyses of the adipose tissue of the bacon samples indicated that N-nitrosopyrrolidine levels in bacon correlated well with the degree of unsaturation of the adipose tissue. N-nitrosothiazolidine was detected in both brine-cured and dry-cured bacon at levels generally below 4 micrograms/kg. However, its formation was greatly reduced by the inclusion of alpha-tocopherol in the cure. The role of woodsmoke in N-nitrosothiazolidine formation in bacon is discussed.

Towards High-Resolution Tissue Imaging Using Nanospray Desorption Electrospray Ionization Mass Spectrometry Coupled to Shear Force Microscopy

NASA Astrophysics Data System (ADS)

Nguyen, Son N.; Sontag, Ryan L.; Carson, James P.; Corley, Richard A.; Ansong, Charles; Laskin, Julia

2018-02-01

Constant mode ambient mass spectrometry imaging (MSI) of tissue sections with high lateral resolution of better than 10 μm was performed by combining shear force microscopy with nanospray desorption electrospray ionization (nano-DESI). Shear force microscopy enabled precise control of the distance between the sample and nano-DESI probe during MSI experiments and provided information on sample topography. Proof-of-concept experiments were performed using lung and brain tissue sections representing spongy and dense tissues, respectively. Topography images obtained using shear force microscopy were comparable to the results obtained using contact profilometry over the same region of the tissue section. Variations in tissue height were found to be dependent on the tissue type and were in the range of 0-5 μm for lung tissue and 0-3 μm for brain tissue sections. Ion images of phospholipids obtained in this study are in good agreement with literature data. Normalization of nano-DESI MSI images to the signal of the internal standard added to the extraction solvent allowed us to construct high-resolution ion images free of matrix effects.

Validation of freezing tissues and cells for analysis of DNA strand break levels by comet assay

PubMed Central

Jackson, Petra

2013-01-01

The comet analysis of DNA strand break levels in tissues and cells has become a common method of screening for genotoxicity. The large majority of published studies have used fresh tissues and cells processed immediately after collection. However, we have used frozen tissues and cells for more than 10 years, and we believe that freezing samples improve efficiency of the method. We compared DNA strand break levels measured in fresh and frozen bronchoalveolar cells, and lung and liver tissues from mice exposed to the known mutagen methyl methanesulphonate (0, 25, 75, 112.5mg/kg). We used a high-throughput comet protocol with fully automated scoring of DNA strand break levels. The overall results from fresh and frozen samples were in agreement [R 2 = 0.93 for %DNA in tail (%TDNA) and R 2 = 0.78 for tail length (TL)]. A slightly increased %TDNA was observed in lung and liver tissue from vehicle controls; and TL was slightly reduced in bronchoalveolar lavage cells from the high-dose group. In our comet protocol, a small block of tissue designated for comet analysis is frozen immediately at tissue collection and kept deep frozen until rapidly homogenised and embedded in agarose. To demonstrate the feasibility of long-term freezing of samples, we analysed the day-to-day variation of our internal historical negative and positive comet assay controls collected over a 10-year period (1128 observations, 11 batches of frozen untreated and H2O2-treated A549 lung epithelial cells). The H2O2 treatment explained most of the variation 57–77% and the day-to-day variation was only 2–12%. The presented protocol allows analysis of samples collected over longer time span, at different locations, with reduced variation by reducing number of electrophoreses and is suitable for both toxicological and epidemiological studies. The use of frozen tissues; however, requires great care during preparation before analysis, with handling as a major risk factor. PMID:24136994

Use of a rapid brain-sampling technique in a physiologic preparation: effects of morphine, ketamine, and halothane on tissue energy intermediates.

PubMed

Dedrick, D F; Sherer, Y D; Biebuyck, J F

1975-06-01

A new method of rapid sampling of brain tissue, "freeze-blowing," has been used to compare the neurochemistry of the brain during anesthesia with that in the awake state. The method avoids anoxia associated with the sampling process. Physiologic variables, including body temperature, blood-gas tensions and blood pressure, were carefully monitored and controlled in the experimental animals. None of the agents tested (halothane, morphine, and ketamine) reduced the brain tissue high-energy phosphate reserved. All three drugs doubled glucose levels. Morphine lowered both lactate and the lactate/pyruvate ratio. Uniformly, the three anesthetic agents led to twofold increases of brain cyclic 3'-5' adenosine monophosphate concentrations. These changes suggest a possible role for cyclic nucleotides in central neurotransmission.

Maximizing Science Return from Future Rodent Experiments on the International Space Station (ISS): Tissue Preservation

NASA Technical Reports Server (NTRS)

Choi, S. Y.; Lai, S.; Klotz, R.; Popova, Y.; Chakravarty, K.; Beegle, J. E.; Wigley, C. L.; Globus, R. K.

2014-01-01

To better understand how mammals adapt to long duration habitation in space, a system for performing rodent experiments on the ISS is under development. Rodent Research-1 is the first flight and will include validation of both on-orbit animal support and tissue preservation. To evaluate plans for on-orbit sample dissection and preservation, we simulated conditions for euthanasia, tissue dissection, and prolonged sample storage on the ISS, and we also developed methods for post-flight dissection and recovery of high quality RNA from multiple tissues following prolonged storage in situ for future science return. Livers and spleens from mice were harvested under conditions that simulated nominal, on-orbit euthanasia and dissection procedures including storage at minus 80 degrees Centigrade for 4 months. The RNA recovered was of high quality (RNA Integrity Number, RNA Integrity Number (RIN) greater than 8) and quantity, and the liver enzyme contents and activities (catalase, glutathione reductase, GAPDH) were similar to positive controls, which were collected under standard laboratory conditions. We also assessed the impact of possible delayed on-orbit dissection scenarios (off-nominal) by dissecting and preserving the spleen (RNA, later) and liver (fast-freezing) at various time points post-euthanasia (from 5 minutes up to 105 minutes). The RNA recovered was of high quality (spleen, RIN greater than 8; liver, RIN greater than 6) and liver enzyme activities were similar to positive controls at all time points, although an apparent decline in select enzyme activities was evident at 105 minutes. Additionally, various tissues were harvested from either intact or partially dissected, frozen carcasses after storage for approximately 2 months; most of the tissues (brain, heart, kidney, eye, adrenal glands and muscle) were of acceptable RNA quality for science return, whereas some tissues (small intestine, bone marrow and bones) were not. These data demonstrate: 1) The protocols developed for future flight experiments will support science return despite delayed preservation post-euthanasia or prolonged storage, and 2) High-quality RNA samples from many different tissues can be recovered by dissection following prolonged storage of the tissue in situ at minus 80 degrees Centigrade. These findings have relevance both to high-value, ground-based experiments when sample collection capability is severely constrained, and to future spaceflight experiments that entail on-orbit sample recovery by the ISS crew.

Health implications of radionuclide levels in cattle raised near U mining and milling facilities in Ambrosia Lake, New Mexico

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lapham, S.C.; Millard, J.B.; Samet, J.M.

1989-03-01

This study was conducted to determine radionuclide tissue levels in cattle raised near U mining and milling facilities. Ambrosia Lake, New Mexico, has been the site of extensive U mining for 30 y and contains several underground U mines, a processing mill, and two large U tailings piles. Ten cows were purchased from two grazing areas in Ambrosia Lake and ten control animals were purchased from Crownpoint, New Mexico. Muscle, liver, kidney, and bone tissue taken from these animals, and environmental samples, including water, grasses and soil collected from the animals' grazing areas, were analyzed for /sup 238/U, /sup 234/U,more » /sup 230/Th, /sup 226/Ra, /sup 210/Pb, and /sup 210/Po. Mean radionuclide levels in cattle tissue and environmental samples from Ambrosia Lake were higher in almost every comparison than those found in respective controls. Liver and kidney tissues were particularly elevated in /sup 226/Ra and /sup 210/Po. Radiation dose commitments from eating cattle tissue with these radionuclide concentrations were calculated. We concluded that the health risk to the public from eating exposed cattle is minimal, unless large amounts of this tissue, especially liver and kidney, are ingested.« less

[Value of specific 16S rDNA fragment of algae in diagnosis of drowning: an experiment with rabbits].

PubMed

Li, Peng; Xu, Qu-Yi; Chen, Ling; Liu, Chao; Zhao, Jian; Wang, Yu-Zhong; Yu, Zheng-Liang; Hu, Sun-Lin; Wang, Hui-Jun

2015-08-01

To establish a method for amplifying specific 16S rDNA fragment of algae related with drowning and test its value in drowning diagnosis. Thirty-five rabbits were randomly divided into 3 the drowning group (n=15), postmortem water immersion group (n=15, subjected to air embolism before seawater immersion), and control group(n=5, with air embolism only). Twenty samples of the liver tissues from human corpses found in water were also used, including 14 diatom-positive and 6 diatom-negative samples identified by microwave digestion-vacuum filtration-automated scanning electron microscopy (MD-VF-Auto SEM). Seven known species of algae served as the control algae (Melosira sp, Nitzschia sp, Synedra sp, Navicula sp, Microcystis sp, Cyclotella meneghiniana, and Chlorella sp). The total DNA was extracted from the tissues and algae to amplify the specific fragment of algae followed by 8% polyacrylamide gelelectrophoresis and sliver-staining. In the drowning group, algae was detected in the lungs (100%), liver (86%), and kidney (86%); algae was detected in the lungs in 2 rabbits in the postmortem group (13%) and none in the control group. The positivity rates of algae were significantly higher in the drowning group than in the postmortem group (P<0.05). Of the 20 tissue samples from human corps found in water, 15 were found positive for algae, including sample that had been identified as diatom-negative by MD-VF-Auto SEM. All the 7 control algae samples yielded positive results in PCR. The PCR-based method has a high sensitivity in algae detection for drowning diagnosis and allows simultaneous detection of multiple algae species related with drowning.

Vitamin D receptor levels in colorectal cancer. Possible role of BsmI polymorphism.

PubMed

Parisi, Eva; Reñé, Josep Maria; Cardús, Anna; Valcheva, Petya; Piñol-Felis, Carme; Valdivielso, José Manuel; Fernández, Elvira

2008-07-01

A high expression of vitamin D receptor (VDR) in colorectal cancer (CRC) tumoral tissue has been related to a good prognosis and it has been proposed that it could be a good biological marker of CRC progression. Nevertheless, there are no previous studies that compare the VDR expression in tumoral towards normal tissue of the same CRC patient in relation to VDR BsmI genotype. We collected normal and tumoral tissue samples, as well as blood samples, from CRC patients (n=170) and controls (n=122). VDR genotyping was performed and BsmI homozygous patients were selected (CRC=50, Cont=32). VDR mRNA and protein levels were analyzed. We also measured 25-Hydroxyvitamin D serum levels. We found no differences in the polymorphism distribution in tumoral versus normal tissue (control: BB=15.7%, bb=41.3%, Bb=43%; CRC: BB=14.2%, bb=41.9%, Bb=43.9%). Furthermore, VDR levels decreased in colonic cancer tissue (mean: 3.03) versus normal mucosa (11.62) from the same patient (p<0.001), but this decrease was similar in both genotypes. There were differences in 25-Hydroxyvitamin D(3) levels between the CRC and the control group (CRC=8.65 ng/ml, Cont=18.15 ng/ml). In conclusion, we found a decrease in VDR levels in tumoral compared with normal mucosa from the same patient. This difference is independent of the BsmI polymorphism.

Detection of Leptospira interrogans DNA and antigen in fixed equine eyes affected with end-stage equine recurrent uveitis.

PubMed

Pearce, Jacqueline W; Galle, Laurence E; Kleiboeker, Steve B; Turk, James R; Schommer, Susan K; Dubielizig, Richard R; Mitchell, William J; Moore, Cecil P; Giuliano, Elizabeth A

2007-11-01

Equine recurrent uveitis (ERU) is the most frequent cause of blindness in horses worldwide. Leptospira has been implicated as an etiologic agent in some cases of ERU and has been detected in fresh ocular tissues of affected horses. The objective of this study was to determine the presence of Leptospira antigen and DNA in fixed equine ocular tissues affected with end-stage ERU. Sections of eyes from 30 horses were obtained. Controls included 1) 10 normal equine eyes and 2) 10 equine eyes with a nonrecurrent form of uveitis. The experimental group consisted of 10 eyes diagnosed with ERU based on clinical signs and histologic lesions. Sections were subjected to immunohistochemical staining with an array of rabbit anti-Leptospira polyclonal antibodies. DNA extractions were performed by using a commercial kit designed for fixed tissue. Real-time PCR analysis was completed on extracted DNA. The target sequence for PCR was designed from alignments of available Leptospira 16S rDNA partial sequences obtained from GenBank. Two of 10 test samples were positive for Leptospira antigen by immunohistochemical assay. Zero of 20 controls were positive for Leptospira antigen. All test samples and controls were negative for Leptospira DNA by real-time PCR analysis. Leptospira was detected at a lower frequency than that previously reported for fresh ERU-affected aqueous humor and vitreous samples. Leptospira is not frequently detectable in fixed ocular tissues of horses affected with ERU when using traditional immunohistochemical and real-time PCR techniques.

Investigation of Epstein-Barr virus DNA in formalin-fixed and paraffin- embedded breast cancer tissues.

PubMed

Kalkan, Ahmet; Ozdarendeli, Aykut; Bulut, Yasemin; Yekeler, Hayrettin; Cobanoglu, Bengu; Doymaz, Mehmet Z

2005-01-01

To investigate etiological role of Epstein-Barr virus (EBV) DNA in breast cancer. The presence of EBV DNA in 57 breast cancer tissues was investigated with a sensitive PCR assay. The breast cancer tissues were from invasive ductular (n=28), lobular (n=20) and other miscellaneous carcinomas (n=9). Tissues from normal breasts and patients with various benign breast diseases (n=55): fibrocystic disease (n=34), fibroadenoma (n=16), hyperplasia, and granulomatous mastitis (n=5), were used as control samples. EBV DNA was detected in 13 (23%) cancerous tissues (7 ductular, 4 lobular, 2 other carcinoma) and 19 (35%) in the control tissues. The difference between EBV presence in malignant and benign tissues was not statistically significant (p>0.05). The presence of EBV DNA was detected almost equally in both breast cancer and normal tissues, which indicates no etiological role for EBV in breast cancer. We suggest further etiological studies. Copyright (c) 2005 S. Karger AG, Basel.

Electromechanical Coupling Factor of Breast Tissue as a Biomarker for Breast Cancer.

PubMed

Park, Kihan; Chen, Wenjin; Chekmareva, Marina A; Foran, David J; Desai, Jaydev P

2018-01-01

This research aims to validate a new biomarker of breast cancer by introducing electromechanical coupling factor of breast tissue samples as a possible additional indicator of breast cancer. Since collagen fibril exhibits a structural organization that gives rise to a piezoelectric effect, the difference in collagen density between normal and cancerous tissue can be captured by identifying the corresponding electromechanical coupling factor. The design of a portable diagnostic tool and a microelectromechanical systems (MEMS)-based biochip, which is integrated with a piezoresistive sensing layer for measuring the reaction force as well as a microheater for temperature control, is introduced. To verify that electromechanical coupling factor can be used as a biomarker for breast cancer, the piezoelectric model for breast tissue is described with preliminary experimental results on five sets of normal and invasive ductal carcinoma (IDC) samples in the 25-45 temperature range. While the stiffness of breast tissues can be captured as a representative mechanical signature which allows one to discriminate among tissue types especially in the higher strain region, the electromechanical coupling factor shows more distinct differences between the normal and IDC groups over the entire strain region than the mechanical signature. From the two-sample -test, the electromechanical coupling factor under compression shows statistically significant differences ( 0.0039) between the two groups. The increase in collagen density in breast tissue is an objective and reproducible characteristic of breast cancer. Although characterization of mechanical tissue property has been shown to be useful for differentiating cancerous tissue from normal tissue, using a single parameter may not be sufficient for practical usage due to inherent variation among biological samples. The portable breast cancer diagnostic tool reported in this manuscript shows the feasibility of measuring multiple parameters of breast tissue allowing for practical application.

Histopathological Analysis of PEEK Wear Particle Effects on the Synovial Tissue of Patients

PubMed Central

Jansson, V.; Giurea, A.

2016-01-01

Introduction. Increasing interest developed in the use of carbon-fiber-reinforced-poly-ether-ether-ketones (CFR-PEEK) as an alternative bearing material in knee arthroplasty. The effects of CFR-PEEK wear in in vitro and animal studies are controversially discussed, as there are no data available concerning human tissue. The aim of this study was to analyze human tissue containing CFR-PEEK as well as UHMWPE wear debris. The authors hypothesized no difference between the used biomaterials. Methods and Materials. In 10 patients during knee revision surgery of a rotating-hinge-knee-implant-design, synovial tissue samples were achieved (tibial inserts: UHMWPE; bushings and flanges: CFR-PEEK). One additional patient received revision surgery without any PEEK components as a control. The tissue was paraffin-embedded, sliced into 2 μm thick sections, and stained with hematoxylin and eosin in a standard process. A modified panoptical staining was also done. Results. A “wear-type” reaction was seen in the testing and the control group. In all samples, the UHMWPE particles were scattered in the tissue or incorporated in giant cells. CFR-PEEK particles were seen as conglomerates and only could be found next to vessels. CFR-PEEK particles showed no giant-cell reactions. In conclusion, the hypothesis has to be rejected. UHMWPE and PEEK showed a different scatter-behavior in human synovial tissue. PMID:27766256

Chronic alcohol abuse in men alters bone mechanical properties by affecting both tissue mechanical properties and microarchitectural parameters.

PubMed

Cruel, M; Granke, M; Bosser, C; Audran, M; Hoc, T

2017-06-01

Alcohol-induced secondary osteoporosis in men has been characterized by higher fracture prevalence and a modification of bone microarchitecture. Chronic alcohol consumption impairs bone cell activity and results in an increased fragility. A few studies highlighted effects of heavy alcohol consumption on some microarchitectural parameters of trabecular bone. But to date and to our knowledge, micro- and macro-mechanical properties of bone of alcoholic subjects have not been investigated. In the present study, mechanical properties and microarchitecture of trabecular bone samples from the iliac crest of alcoholic male patients (n=15) were analyzed and compared to a control group (n=8). Nanoindentation tests were performed to determine the tissue's micromechanical properties, micro-computed tomography was used to measure microarchitectural parameters, and numerical simulations provided the apparent mechanical properties of the samples. Compared to controls, bone tissue from alcoholic patients exhibited an increase of micromechanical properties at tissue scale, a significant decrease of apparent mechanical properties at sample scale, and significant changes in several microarchitectural parameters. In particular, a crucial role of structure model index (SMI) on mechanical properties was identified. 3D microarchitectural parameters are at least as important as bone volume fraction to predict bone fracture risk in the case of alcoholic patients. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Human dental pulp stem cells derived from cryopreserved dental pulp tissues of vital extracted teeth with disease demonstrate hepatic-like differentiation.

PubMed

Chen, Y K; Huang, Anderson H C; Chan, Anthony W S; Lin, L M

2016-06-01

Reviewing the literature, hepatic differentiation of human dental pulp stem cells (hDPSCs) from cryopreserved dental pulp tissues of vital extracted teeth with disease has not been studied. This study is aimed to evaluate the hypothesis that hDPSCs from cryopreserved dental pulp tissues of vital extracted teeth with disease could possess potential hepatic differentiation. Forty vital extracted teeth with disease recruited for hDPSCs isolation, stem cell characterization and hepatic differentiation were randomly and equally divided into group A (liquid nitrogen-stored dental pulp tissues) and group B (freshly derived dental pulp tissues). Samples of hDPSCs isolated from groups A and B but without hepatic growth factors formed negative controls. A well-differentiated hepatocellular carcinoma cell line was employed as a positive control. All the isolated hDPSCs from groups A and B showed hepatic-like differentiation with morphological change from a spindle-shaped to a polygonal shape and normal karyotype. Differentiated hDPSCs and the positive control expressed hepatic metabolic function genes and liver-specific genes. Glycogen storage of differentiated hDPSCs was noted from day 7 of differentiation-medium culture. Positive immunofluorescence staining of low-density lipoprotein and albumin was observed from day 14 of differentiation-medium culture; urea production in the medium was noted from week 6. No hepatic differentiation was observed for any of the samples of the negative controls. We not only demonstrated the feasibility of hepatic-like differentiation of hDPSCs from cryopreserved dental pulp tissues of vital extracted teeth with disease but also indicated that the differentiated cells possessed normal karyotype and were functionally close to normal hepatic-like cells. Copyright © 2013 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Microwave fixation versus formalin fixation of surgical and autopsy tissue.

PubMed

Login, G R

1978-05-01

Microwave irradiation of surgical and autopsy tissue penetrates, fixes, and hardens the tissue almost immediately (the fluid media used in the microwave consisted of saline, ten percent phosphate buffered formalin, and distilled water). Tissue sections from a representative sample of organs were tested. Comparable sections were simultaneously fixed in a phosphate buffered ten percent formalin bath in a vaccum oven as a control. Hematoxylin and eosin were used to stain the sections. Results equal to and superior to the control method were obtained. Saline microwave fixation was superior to formalin microwave fixation. Tissues placed in Zenker's solution and fixed in standard microwave oven (for approximately one minute) yielded results at least equal to conventional Zenker fixation (approximately two hours). No tissue hardening resulted from Zenker microwave fixation. A unique time versus temperature graph (microwave heating curve) reduces individual variation with this technique.

3D-templated, fully automated microfluidic input/output multiplexer for endocrine tissue culture and secretion sampling.

PubMed

Li, Xiangpeng; Brooks, Jessica C; Hu, Juan; Ford, Katarena I; Easley, Christopher J

2017-01-17

A fully automated, 16-channel microfluidic input/output multiplexer (μMUX) has been developed for interfacing to primary cells and to improve understanding of the dynamics of endocrine tissue function. The device utilizes pressure driven push-up valves for precise manipulation of nutrient input and hormone output dynamics, allowing time resolved interrogation of the cells. The ability to alternate any of the 16 channels from input to output, and vice versa, provides for high experimental flexibility without the need to alter microchannel designs. 3D-printed interface templates were custom designed to sculpt the above-channel polydimethylsiloxane (PDMS) in microdevices, creating millimeter scale reservoirs and confinement chambers to interface primary murine islets and adipose tissue explants to the μMUX sampling channels. This μMUX device and control system was first programmed for dynamic studies of pancreatic islet function to collect ∼90 minute insulin secretion profiles from groups of ∼10 islets. The automated system was also operated in temporal stimulation and cell imaging mode. Adipose tissue explants were exposed to a temporal mimic of post-prandial insulin and glucose levels, while simultaneous switching between labeled and unlabeled free fatty acid permitted fluorescent imaging of fatty acid uptake dynamics in real time over a ∼2.5 hour period. Application with varying stimulation and sampling modes on multiple murine tissue types highlights the inherent flexibility of this novel, 3D-templated μMUX device. The tissue culture reservoirs and μMUX control components presented herein should be adaptable as individual modules in other microfluidic systems, such as organ-on-a-chip devices, and should be translatable to different tissues such as liver, heart, skeletal muscle, and others.

Analysis of energy dispersive x-ray diffraction profiles for material identification, imaging and system control

NASA Astrophysics Data System (ADS)

Cook, Emily Jane

2008-12-01

This thesis presents the analysis of low angle X-ray scatter measurements taken with an energy dispersive system for substance identification, imaging and system control. Diffraction measurements were made on illicit drugs, which have pseudo- crystalline structures and thus produce diffraction patterns comprising a se ries of sharp peaks. Though the diffraction profiles of each drug are visually characteristic, automated detection systems require a substance identification algorithm, and multivariate analysis was selected as suitable. The software was trained with measured diffraction data from 60 samples covering 7 illicit drugs and 5 common cutting agents, collected with a range of statistical qual ities and used to predict the content of 7 unknown samples. In all cases the constituents were identified correctly and the contents predicted to within 15%. Soft tissues exhibit broad peaks in their diffraction patterns. Diffraction data were collected from formalin fixed breast tissue samples and used to gen erate images. Maximum contrast between healthy and suspicious regions was achieved using momentum transfer windows 1.04-1.10 and 1.84-1.90 nm_1. The resulting images had an average contrast of 24.6% and 38.9% compared to the corresponding transmission X-ray images (18.3%). The data was used to simulate the feedback for an adaptive imaging system and the ratio of the aforementioned momentum transfer regions found to be an excellent pa rameter. Investigation into the effects of formalin fixation on human breast tissue and animal tissue equivalents indicated that fixation in standard 10% buffered formalin does not alter the diffraction profiles of tissue in the mo mentum transfer regions examined, though 100% unbuffered formalin affects the profile of porcine muscle tissue (a substitute for glandular and tumourous tissue), though fat is unaffected.

Colorimeter and scanning electron microscopy analysis of teeth submitted to internal bleaching.

PubMed

Martin-Biedma, Benjamin; Gonzalez-Gonzalez, Teresa; Lopes, Manuela; Lopes, Luis; Vilar, Rui; Bahillo, José; Varela-Patiño, Purificación

2010-02-01

This in vitro study compared the tooth color and the ultrastructure of internal dental tissues before and after internal bleaching. Sodium perborate was placed in the pulp chamber of endodontically treated molars and sealed with intermediate restorative material. The test samples were stored in a physiologic solution, and the bleaching agent was replaced every 7 days. A control group was used. After 1 month, the colors of the test and control samples were measured with a colorimeter, and the internal surfaces were observed under field emission scanning electron microscopy (FESEM). Statistically significant differences were found between the test and control sample colors. The FESEM ultrastructure analysis of the internal enamel and dentin surfaces did not show any changes after the internal bleaching. The results of the present study show that sodium perborate is effective in bleaching nonvital teeth and does not produce ultrastructural changes in the dental tissues. Copyright 2010 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

In vitro color evaluation of esthetic coatings for metallic dental implants and implant prosthetic appliances.

PubMed

Pecnik, Christina M; Roos, Malgorzata; Muff, Daniel; Spolenak, Ralph; Sailer, Irena

2015-05-01

The aim of this study was to characterize the optical properties of newly developed esthetic coatings for metallic implants and components for an improved peri-implant soft tissue appearance. Pig maxillae (n = 6) were used for the in vitro color evaluation of coated and uncoated samples. Three different coating systems (Ti-ZrO(2), Ti-Al-ZrO(2), and Ti-Ag-ZrO(2)) were deposited on titanium substrates, which exhibited different roughness (polished, machined, and sand-blasted) and interference colors (pink, yellow, and white). Spectrophotometric measurements were made of samples below three different mucosa thicknesses (1 mm, 2 mm, and 3 mm) and titanium served as negative control. Color difference ΔE was calculated using ΔL, Δa, and Δb values for each sample (in total 30 samples). ΔE values were significantly above the threshold value of 3.70 for sand-blasted Ti and Ti-ZrO(2) samples when tested below 1 mm thick soft tissue, hence resulted in a dark appearance of the soft tissues. In contrast, Ti-Al-ZrO(2) and Ti-Ag-ZrO(2) samples showed significant ΔL values below 1 mm, which indicates a brightening of the covering tissue. In general, ΔE values decreased with increasing thickness of the tissue. At 3 mm thick tissue, ΔE values were significantly below 3.70 for Ti-Al-ZrO(2) and Ti-Ag-ZrO(2) samples. The preferable substrate surface should be machined due increased color brightness, good soft tissue integration and improved adhesion between coating and substrates. Improvement of the optical appearance of the metal was achieved with the coating systems Ti-Al-ZrO(2) and Ti-Ag-ZrO(2). Darkening effects could not be observed for these systems, and partially light brightening of the tissue was observed. Advantageous colors were suggested to be pink and yellow. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Gamma Radiation Sterilization Reduces the High-cycle Fatigue Life of Allograft Bone.

PubMed

Islam, Anowarul; Chapin, Katherine; Moore, Emily; Ford, Joel; Rimnac, Clare; Akkus, Ozan

2016-03-01

Sterilization by gamma radiation impairs the mechanical properties of bone allografts. Previous work related to radiation-induced embrittlement of bone tissue has been limited mostly to monotonic testing which does not necessarily predict the high-cycle fatigue life of allografts in vivo. We designed a custom rotating-bending fatigue device to answer the following questions: (1) Does gamma radiation sterilization affect the high-cycle fatigue behavior of cortical bone; and (2) how does the fatigue life change with cyclic stress level? The high-cycle fatigue behavior of human cortical bone specimens was examined at stress levels related to physiologic levels using a custom-designed rotating-bending fatigue device. Test specimens were distributed among two treatment groups (n = 6/group); control and irradiated. Samples were tested until failure at stress levels of 25, 35, and 45 MPa. At 25 MPa, 83% of control samples survived 30 million cycles (run-out) whereas 83% of irradiated samples survived only 0.5 million cycles. At 35 MPa, irradiated samples showed an approximately 19-fold reduction in fatigue life compared with control samples (12.2 × 10(6) ± 12.3 × 10(6) versus 6.38 × 10(5) ± 6.81 × 10(5); p = 0.046), and in the case of 45 MPa, this reduction was approximately 17.5-fold (7.31 × 10(5) ± 6.39 × 10(5) versus 4.17 × 10(4) ± 1.91 × 10(4); p = 0.025). Equations to estimate high-cycle fatigue life of irradiated and control cortical bone allograft at a certain stress level were derived. Gamma radiation sterilization severely impairs the high cycle fatigue life of structural allograft bone tissues, more so than the decline that has been reported for monotonic mechanical properties. Therefore, clinicians need to be conservative in the expectation of the fatigue life of structural allograft bone tissues. Methods to preserve the fatigue strength of nonirradiated allograft bone tissue are needed. As opposed to what monotonic tests might suggest, the cyclic fatigue life of radiation-sterilized structural allografts is likely severely compromised relative to the nonirradiated condition and therefore should be taken into consideration. Methods to reduce the effect of irradiation or to recover structural allograft bone tissue fatigue strength are important to pursue.

Psychiatric Brain Banking: Three Perspectives on Current Trends and Future Directions

PubMed Central

Deep-Soboslay, Amy; Benes, Francine M.; Haroutunian, Vahram; Ellis, Justin K.; Kleinman, Joel E.; Hyde, Thomas M.

2011-01-01

Introduction The study of postmortem human brain tissue is central to the advancement of the neurobiological studies of psychiatric illness, particularly for the study of brain-specific isoforms and molecules. Methods The state-of-the-art methods and recommendations for maintaining a successful brain bank for psychiatric disorders are discussed, using the convergence of viewpoints from three brain collections, the National Institute of Mental Health Brain Collection (NIMH), the Harvard Brain Tissue Resource Center (HBTRC), and the Mt. Sinai School of Medicine Brain Bank (MSSM-BB), with diverse research interests and divergent approaches to tissue acquisition. Results While the NIMH obtains donations from medical examiners for its collection, and places particular emphasis on clinical diagnosis, toxicology, and building lifespan control cohorts, the HBTRC is uniquely designed as a repository whose sole purpose is to collect large-volume, high quality brain tissue from community-based donors based on relationships across an expansive nationwide network, and places emphasis on the accessibility of its bank in disseminating tissue and related data to research groups worldwide. The MSSM-BB collection has shown that, with dedication, prospective recruitment is a successful approach to tissue donation, and places particular emphasis on rigorous clinical diagnosis through antemortem contact with donors. The MSSM-BB places great importance on stereological tissue sampling methods for neuroanatomical studies, and frozen tissue sampling approaches that enable multiple assessments (RNA, DNA, protein, enzyme activity, binding, etc.) of the same tissue block. Promising scientific approaches for elucidating the molecular and cellular pathways in brain that may contribute to schizophrenia and/or bipolar disorder, such as cell culture techniques and microarray-based gene expression and genotyping studies are briefly discussed. Conclusions Despite unique perspectives from three established brain collections, there is a consensus that (1) diverse strategies for tissue acquisition, (2) rigor in tissue and diagnostic characterization, (3) the importance of sample accessibility, and (4) continual application of innovative scientific approaches to the study of brain tissue are all integral to the success and future of psychiatric brain banking. The future of neuropsychiatric research depends upon in the availability of high quality brain specimens from large numbers of subjects, including non-psychiatric controls. PMID:20673875

Electric field computation and measurements in the electroporation of inhomogeneous samples

NASA Astrophysics Data System (ADS)

Bernardis, Alessia; Bullo, Marco; Campana, Luca Giovanni; Di Barba, Paolo; Dughiero, Fabrizio; Forzan, Michele; Mognaschi, Maria Evelina; Sgarbossa, Paolo; Sieni, Elisabetta

2017-12-01

In clinical treatments of a class of tumors, e.g. skin tumors, the drug uptake of tumor tissue is helped by means of a pulsed electric field, which permeabilizes the cell membranes. This technique, which is called electroporation, exploits the conductivity of the tissues: however, the tumor tissue could be characterized by inhomogeneous areas, eventually causing a non-uniform distribution of current. In this paper, the authors propose a field model to predict the effect of tissue inhomogeneity, which can affect the current density distribution. In particular, finite-element simulations, considering non-linear conductivity against field relationship, are developed. Measurements on a set of samples subject to controlled inhomogeneity make it possible to assess the numerical model in view of identifying the equivalent resistance between pairs of electrodes.

Soils element activities for the period October 1973--September 1974

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fowler, E.B.; Essington, E.H.; White, M.G.

Soils Element activities were conducted on behalf of the U. S. Atomic Energy Commission's Nevada Applied Ecology Group (NAEG) program to provide source term information for the other program elements and maintain continuous cognizance of program requirements for sampling, sample preparation, and analysis. Activities included presentation of papers; participation in workshops; analysis of soil, vegetation, and animal tissue samples for $sup 238$Pu, $sup 239-240$Pu, $sup 241$Am, $sup 137$Cs, $sup 60$Co, and gamma scan for routine and laboratory quality control purposes; preparation and analysis of animal tissue samples for NAEG laboratory certification; studies on a number of analytical, sample preparation, andmore » sample collection procedures; and contributions to the evaluation of procedures for calculation of specialized counting statistics. (auth)« less

Identification of viral infections in the prostate and evaluation of their association with cancer

PubMed Central

2010-01-01

Background Several viruses with known oncogenic potential infect prostate tissue, among these are the polyomaviruses BKV, JCV, and SV40; human papillomaviruses (HPVs), and human cytomegalovirus (HCMV) infections. Recently, the Xenotropic Murine Leukemia Virus-related gammaretrovirus (XMRV) was identified in prostate tissue with a high prevalence observed in prostate cancer (PC) patients homozygous for the glutamine variant of the RNASEL protein (462Q/Q). Association studies with the R462Q allele and non-XMRV viruses have not been reported. We assessed associations between prostate cancer, prostate viral infections, and the RNASEL 462Q allele in Mexican cancer patients and controls. Methods 130 subjects (55 prostate cancer cases and 75 controls) were enrolled in the study. DNA and RNA isolated from prostate tissues were screened for the presence of viral genomes. Genotyping of the RNASEL R462Q variant was performed by Taqman method. Results R/R, R/Q, and Q/Q frequencies for R462Q were 0.62, 0.38, and 0.0 for PC cases and 0.69, 0.24, and 0.07 for controls, respectively. HPV sequences were detected in 11 (20.0%) cases and 4 (5.3%) controls. XMRV and HCMV infections were detected in one and six control samples, respectively. The risk of PC was significantly increased (Odds Ratio = 3.98; 95% CI: 1.17-13.56, p = 0.027) by infection of the prostatic tissue with HPV. BKV, JCV, and SV40 sequences were not detected in any of the tissue samples examined. Conclusions We report a positive association between PC and HPV infection. The 462Q/Q RNASEL genotype was not represented in our PC cases; thus, its interaction with prostate viral infections and cancer could not be evaluated. PMID:20576103

Platelet-rich plasma enhances the integration of bioengineered cartilage with native tissue in an in vitro model.

PubMed

Sermer, Corey; Kandel, Rita; Anderson, Jesse; Hurtig, Mark; Theodoropoulos, John

2018-02-01

Current therapies for cartilage repair can be limited by an inability of the repair tissue to integrate with host tissue. Thus, there is interest in developing approaches to enhance integration. We have previously shown that platelet-rich plasma (PRP) improves cartilage tissue formation. This raised the question as to whether PRP could promote cartilage integration. Chondrocytes were isolated from cartilage harvested from bovine joints, seeded on a porous bone substitute and grown in vitro to form an osteochondral-like implant. After 7 days, the biphasic construct was soaked in PRP for 30 min before implantation into the core of a donut-shaped biphasic explant of native cartilage and bone. Controls were not soaked in PRP. The implant-explant construct was cultured for 2-4 weeks. PRP-soaked bioengineered implants integrated with host tissue in 73% of samples, whereas controls only integrated in 19% of samples. The integration strength, as determined by a push-out test, was significantly increased in the PRP-soaked implant group (219 ± 35.4 kPa) compared with controls (72.0 ± 28.5 kPa). This correlated with an increase in glycosaminoglycan and collagen accumulation in the region of integration in the PRP-treated implant group, compared with untreated controls. Immunohistochemical studies revealed that the integration zone contained collagen type II and aggrecan. The cells at the zone of integration in the PRP-soaked group had a 3.5-fold increase in matrix metalloproteinase-13 gene expression compared with controls. These results suggest that PRP-soaked bioengineered cartilage implants may be a better approach for cartilage repair due to enhanced integration. Copyright © 2017 John Wiley & Sons, Ltd.

A High-Throughput Method for Direct Detection of Therapeutic Oligonucleotide-Induced Gene Silencing In Vivo

PubMed Central

Coles, Andrew H.; Osborn, Maire F.; Alterman, Julia F.; Turanov, Anton A.; Godinho, Bruno M.D.C.; Kennington, Lori; Chase, Kathryn; Aronin, Neil

2016-01-01

Preclinical development of RNA interference (RNAi)-based therapeutics requires a rapid, accurate, and robust method of simultaneously quantifying mRNA knockdown in hundreds of samples. The most well-established method to achieve this is quantitative real-time polymerase chain reaction (qRT-PCR), a labor-intensive methodology that requires sample purification, which increases the potential to introduce additional bias. Here, we describe that the QuantiGene® branched DNA (bDNA) assay linked to a 96-well Qiagen TissueLyser II is a quick and reproducible alternative to qRT-PCR for quantitative analysis of mRNA expression in vivo directly from tissue biopsies. The bDNA assay is a high-throughput, plate-based, luminescence technique, capable of directly measuring mRNA levels from tissue lysates derived from various biological samples. We have performed a systematic evaluation of this technique for in vivo detection of RNAi-based silencing. We show that similar quality data is obtained from purified RNA and tissue lysates. In general, we observe low intra- and inter-animal variability (around 10% for control samples), and high intermediate precision. This allows minimization of sample size for evaluation of oligonucleotide efficacy in vivo. PMID:26595721

Controlled ice nucleation in cryopreservation--a review.

PubMed

Morris, G John; Acton, Elizabeth

2013-04-01

We review here for the first time, the literature on control of ice nucleation in cryopreservation. Water and aqueous solutions have a tendency to undercool before ice nucleation occurs. Control of ice nucleation has been recognised as a critical step in the cryopreservation of embryos and oocytes but is largely ignored for other cell types. We review the processes of ice nucleation and crystal growth in the solution around cells and tissues during cryopreservation with an emphasis on non IVF applications. The extent of undercooling that is encountered during the cooling of various cryocontainers is defined and the methods that have been employed to control the nucleation of ice are examined. The effects of controlled ice nucleation on the structure of the sample and the outcome of cryopreservation of a range of cell types and tissues are presented and the physical events which define the cellular response are discussed. Nucleation of ice is the most significant uncontrolled variable in conventional cryopreservation leading to sample to sample variation in cell recovery, viability and function and should be controlled to allow standardisation of cryopreservation protocols for cells for biobanking, cell based assays or clinical application. This intervention allows a way of increasing viability of cells and reducing variability between samples and should be included as standard operating procedures are developed. Copyright © 2012 Elsevier Inc. All rights reserved.

Glutamine Provides Effective Protection against Deltamethrin-Induced Acute Hepatotoxicity in Rats But Not Against Nephrotoxicity

PubMed Central

Gündüz, Ercan; Ülger, Burak Veli; İbiloğlu, İbrahim; Ekinci, Aysun; Dursun, Recep; Zengin, Yılmaz; İçer, Mustafa; Uslukaya, Ömer; Ekinci, Cenap; Güloğlu, Cahfer

2015-01-01

Background The aim of this study was to investigate the protective effects of L-glutamine (GLN) against liver and kidney injury caused by acute toxicity of deltamethrin (DLM). Material/Methods Thirty-two rats were indiscriminately separated into 4 groups with 8 rats each: control group (distilled water; 10 ml/kg, perorally [p.o.]), DLM group (35 mg/kg p.o. one dose.), GLN group (1.5 gr/kg, p.o. single dose.) and DLM (35 mg/kg p.o. one dose.) + GLN group (1.5 gr/kg, p.o. one dose after 4 hours.). Testing for total antioxidant status (TAS), total oxidant status (TOS), interleukin-1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6) analyses were performed on tissue samples, and alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH), urea, and creatinine were analyzed on serum samples. Liver and kidney samples were histopathologically analyzed. Results The TOS level in liver was significantly higher in the DLM group than in the control group, and the level in DLM+GLN group was considerably lower than in the DLM group. The TAS level in the DLM+GLN group was considerably higher than in the control and DLM groups. The TAS level in kidney tissues was considerably lower in the DLM group than in controls, but was similar to other groups. Histopathological analyses of liver tissues established a significant difference between DLM and DLM+GLN groups in terms of grade 2 hepatic injury. However, no significant difference was found between DLM and DLM+GLN groups in terms of kidney injury. Conclusions Glutamine leads to significant improvement in deltamethrin-induced acute hepatotoxicity in terms of histopathologic results, tissue oxidative stress parameters, and serum liver function marker enzymes. PMID:25890620

Telomere length in normal and neoplastic canine tissues.

PubMed

Cadile, Casey D; Kitchell, Barbara E; Newman, Rebecca G; Biller, Barbara J; Hetler, Elizabeth R

2007-12-01

To determine the mean telomere restriction fragment (TRF) length in normal and neoplastic canine tissues. 57 solid-tissue tumor specimens collected from client-owned dogs, 40 samples of normal tissue collected from 12 clinically normal dogs, and blood samples collected from 4 healthy blood donor dogs. Tumor specimens were collected from client-owned dogs during diagnostic or therapeutic procedures at the University of Illinois Veterinary Medical Teaching Hospital, whereas 40 normal tissue samples were collected from 12 control dogs. Telomere restriction fragment length was determined by use of an assay kit. A histologic diagnosis was provided for each tumor by personnel at the Veterinary Diagnostic Laboratory at the University of Illinois. Mean of the mean TRF length for 44 normal samples was 19.0 kilobases (kb; range, 15.4 to 21.4 kb), and the mean of the mean TRF length for 57 malignant tumors was 19.0 kb (range, 12.9 to 23.5 kb). Although the mean of the mean TRF length for tumors and normal tissues was identical, tumor samples had more variability in TRF length. Telomerase, which represents the main mechanism by which cancer cells achieve immortality, is an attractive therapeutic target. The ability to measure telomere length is crucial to monitoring the efficacy of telomerase inhibition. In contrast to many other mammalian species, the length of canine telomeres and the rate of telomeric DNA loss are similar to those reported in humans, making dogs a compelling choice for use in the study of human anti-telomerase strategies.

Oxidative stress in juvenile chinook salmon, Oncorhynchus tshawytscha (Walbaum)

USGS Publications Warehouse

Welker, T.L.; Congleton, J.L.

2004-01-01

Juvenile chinook salmon, Oncorhynchus tshawytscha (Walbaum), were held in 8-11??C freshwater, starved for 3 days and subjected to a low-water stressor to determine the relationship between the general stress response and oxidative stress. Lipid peroxidation (LPO) levels (lipid hydroperoxides) were measured in kidney, liver and brain samples taken at the beginning of the experiment (0-h unstressed controls) and at 6, 24 and 48 h after application of a continuous low-water stressor. Tissue samples were also taken at 48 h from fish that had not been exposed to the stressor (48-h unstressed controls). Exposure to the low-water stressor affected LPO in kidney and brain tissues. In kidney, LPO decreased 6 h after imposition of the stressor; similar but less pronounced decreases also occurred in the liver and brain. At 48 h, LPO increased (in comparison with 6-h stressed tissues) in the kidney and brain. In comparison with 48-h unstressed controls, LPO levels were higher in the kidney and brain of stressed fish. Although preliminary, results suggest that stress can cause oxidative tissue damage in juvenile chinook salmon. Measures of oxidative stress have shown similar responses to stress in mammals; however, further research is needed to determine the extent of the stress-oxidative stress relationship and the underlying physiological mechanisms in fish.

Propionibacterium acnes in shoulder surgery: is loss of hair protective for infection?

PubMed

Hudek, Robert; Sommer, Frank; Abdelkawi, Ayman F; Kerwat, Martina; Müller, Hans-Helge; Gohlke, Frank

2016-06-01

Propionibacterium acnes (P acnes) has been linked to chronic infections in shoulder surgery. It was recently observed during first-time shoulder surgery in healthy patients at a rate between 36% and 56%. Male gender and the anterolateral approach were reported risk factors. Because the skin biology greatly differs, we aimed to correlate skin complaints with P acnes-positive intraoperative cultures from different tissue layer samples in patients undergoing shoulder surgery for the first time. Intraoperative samples (1 skin, 1 superficial, 1 deep tissue, and 1 control sample) from 112 patients (70 men, 42 women; aged 59.2 years) were cultured. The association between the presence of P acnes in the deep or superficial tissue, or both, and 10 items of a validated preoperative questionnaire for skin pathology was explored. The cultures were positive for P acnes in 38.4% (n = 43) of the cases. Skin samples were positive for P acnes in 8% (n = 9), superficial samples were positive in 23% (n = 26), and deep samples were positive in 30% (n = 34). Self-reported "loss of hair" was significantly negatively associated with the presence of P acnes in the superficial or deep tissue sample (P = .00028). Patients who report having "loss of hair" show fewer P acnes-positive cultures in intraoperative tissue samples taken during open shoulder surgery. Whether this subgroup is at a lesser risk for P acnes infections remains to be substantiated. Basic Science Study; Microbiology. Copyright © 2016 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Elsevier Inc. All rights reserved.

Raman spectroscopic evidence of tissue restructuring in heat-induced tissue fusion.

PubMed

Su, Lei; Cloyd, Kristy L; Arya, Shobhit; Hedegaard, Martin A B; Steele, Joseph A M; Elson, Daniel S; Stevens, Molly M; Hanna, George B

2014-09-01

Heat-induced tissue fusion via radio-frequency (RF) energy has gained wide acceptance clinically and here we present the first optical-Raman-spectroscopy study on tissue fusion samples in vitro. This study provides direct insights into tissue constituent and structural changes on the molecular level, exposing spectroscopic evidence for the loss of distinct collagen fibre rich tissue layers as well as the denaturing and restructuring of collagen crosslinks post RF fusion. These findings open the door for more advanced optical feedback-control methods and characterization during heat-induced tissue fusion, which will lead to new clinical applications of this promising technology. Copyright © 2014 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim.

Imaging of oxygen and hypoxia in cell and tissue samples.

PubMed

Papkovsky, Dmitri B; Dmitriev, Ruslan I

2018-05-14

Molecular oxygen (O 2 ) is a key player in cell mitochondrial function, redox balance and oxidative stress, normal tissue function and many common disease states. Various chemical, physical and biological methods have been proposed for measurement, real-time monitoring and imaging of O 2 concentration, state of decreased O 2 (hypoxia) and related parameters in cells and tissue. Here, we review the established and emerging optical microscopy techniques allowing to visualize O 2 levels in cells and tissue samples, mostly under in vitro and ex vivo, but also under in vivo settings. Particular examples include fluorescent hypoxia stains, fluorescent protein reporter systems, phosphorescent probes and nanosensors of different types. These techniques allow high-resolution mapping of O 2 gradients in live or post-mortem tissue, in 2D or 3D, qualitatively or quantitatively. They enable control and monitoring of oxygenation conditions and their correlation with other biomarkers of cell and tissue function. Comparison of these techniques and corresponding imaging setups, their analytical capabilities and typical applications are given.

Modified use of methylene blue in the tissue compression technique to detect sarcocysts in meat-producing animals.

PubMed

Ng, Yit Han; Subramaniam, Vellayan; Lau, Yee Ling

2015-11-30

Sarcocystosis in meat-producing animals is a major cause of reduced productivity in many countries, especially those that rely on agriculture. Although several diagnostic methods are available to detect sarcocystosis, many are too time-consuming for routine use in abattoirs and meat inspection centers, where large numbers of samples need to be tested. This study aimed to compare the sensitivity of the methylene blue tissue preparation, unstained tissue preparation and nested PCR in the detection of sarcocysts in tissue samples. Approximately three-fold more sarcocysts were detected in methylene blue-stained tissue compared to unstained controls (McNemar's test: P<0.01). Test sensitivity was comparable to that of the gold standard for sarcocyst detection, nested polymerase chain reaction. These results suggest that methylene blue can be used in tissue compression as a rapid, safe, and inexpensive technique for the detection of ruminant sarcocystosis in abattoirs. Copyright © 2015 Elsevier B.V. All rights reserved.

Waterjet cutting of periprosthetic interface tissue in loosened hip prostheses: an in vitro feasibility study.

PubMed

Kraaij, Gert; Tuijthof, Gabrielle J M; Dankelman, Jenny; Nelissen, Rob G H H; Valstar, Edward R

2015-02-01

Waterjet cutting technology is considered a promising technology to be used for minimally invasive removal of interface tissue surrounding aseptically loose hip prostheses. The goal of this study was to investigate the feasibility of waterjet cutting of interface tissue membrane. Waterjets with 0.2 mm and 0.6 mm diameter, a stand-off distance of 5 mm, and a traverse speed of 0.5 mm/s were used to cut interface tissue samples in half. The water flow through the nozzle was controlled by means of a valve. By changing the flow, the resulting waterjet pressure was regulated. Tissue sample thickness and the required waterjet pressures were measured. Mean thickness of the samples tested within the 0.2 mm nozzle group was 2.3 mm (SD 0.7 mm) and within the 0.6 mm nozzle group 2.6 mm (SD 0.9 mm). The required waterjet pressure to cut samples was between 10 and 12 MPa for the 0.2 mm nozzle and between 5 and 10 MPa for the 0.6 mm nozzle. Cutting bone or bone cement requires about 3 times higher waterjet pressure (30-50 MPa, depending on used nozzle diameter) and therefore we consider waterjet cutting as a safe technique to be used for minimally invasive interface tissue removal. Copyright © 2015 IPEM. Published by Elsevier Ltd. All rights reserved.

Sun exposure in pigs increases the vitamin D nutritional quality of pork

PubMed Central

Wechsler, Perry J.; Hollis, Bruce W.; Makowski, Andrew J.

2017-01-01

There is a high prevalence of vitamin D insufficiency and deficiency worldwide likely because of both limited sun-exposure and inadequate dietary intake. Meat, including pork, is not typically considered a dietary source of vitamin D, possibly because of management practices that raise pigs in confinement. This experiment determined the vitamin D content of loin and subcutaneous adipose tissue in sun-exposed finisher pigs. Two separate groups of pigs were used. The first group (28 white Landrace-Duroc) was assigned at random to either sunlight exposure (SUN) in spring and summer or confinement per standard practice (Control). The second (24 Yorkshire-Duroc-Landrace) underwent the same exposure protocol but was exposed in summer and fall or assigned to control (Control). A subsample of five SUN and four Control pigs, matched for weight and body condition score, was selected for slaughter from each group. Pigs (n = 10 SUN, n = 8 Control) had blood drawn for analysis of 25(OH)D3 concentration before/after sun exposure or control, and tissue samples were taken at slaughter for analysis of tissue vitamin D3 and 25(OH)D3 concentration. Three random samples from a single loin chop and surrounding adipose were collected and analyzed. Serum concentrations of 25(OH)D3 did not differ (P≥0.376) between treatments prior to sun exposure in either group, but was increased (time*treatment interaction, P<0.001) with SUN exposure. Total vitamin D content (D3 plus 25(OH)D3) of loin tissue was increased (P < 0.001) with sun exposure and averaged 0.997±0.094 μg/100g and 0.348±0.027 μg/100g for sun and control pigs, respectively. While exposure to sunlight increased (P = 0.003) tissue content of 25(OH) D in subcutaneous adipose tissue, vitamin D3 content was similar between treatments (P = 0.56). Sunlight exposure in pigs increased the vitamin D content of loin, and may provide an additional source of dietary vitamin D. PMID:29136033

Sun exposure in pigs increases the vitamin D nutritional quality of pork.

PubMed

Larson-Meyer, D Enette; Ingold, Bennett C; Fensterseifer, Samanta R; Austin, Kathleen J; Wechsler, Perry J; Hollis, Bruce W; Makowski, Andrew J; Alexander, Brenda M

2017-01-01

There is a high prevalence of vitamin D insufficiency and deficiency worldwide likely because of both limited sun-exposure and inadequate dietary intake. Meat, including pork, is not typically considered a dietary source of vitamin D, possibly because of management practices that raise pigs in confinement. This experiment determined the vitamin D content of loin and subcutaneous adipose tissue in sun-exposed finisher pigs. Two separate groups of pigs were used. The first group (28 white Landrace-Duroc) was assigned at random to either sunlight exposure (SUN) in spring and summer or confinement per standard practice (Control). The second (24 Yorkshire-Duroc-Landrace) underwent the same exposure protocol but was exposed in summer and fall or assigned to control (Control). A subsample of five SUN and four Control pigs, matched for weight and body condition score, was selected for slaughter from each group. Pigs (n = 10 SUN, n = 8 Control) had blood drawn for analysis of 25(OH)D3 concentration before/after sun exposure or control, and tissue samples were taken at slaughter for analysis of tissue vitamin D3 and 25(OH)D3 concentration. Three random samples from a single loin chop and surrounding adipose were collected and analyzed. Serum concentrations of 25(OH)D3 did not differ (P≥0.376) between treatments prior to sun exposure in either group, but was increased (time*treatment interaction, P<0.001) with SUN exposure. Total vitamin D content (D3 plus 25(OH)D3) of loin tissue was increased (P < 0.001) with sun exposure and averaged 0.997±0.094 μg/100g and 0.348±0.027 μg/100g for sun and control pigs, respectively. While exposure to sunlight increased (P = 0.003) tissue content of 25(OH) D in subcutaneous adipose tissue, vitamin D3 content was similar between treatments (P = 0.56). Sunlight exposure in pigs increased the vitamin D content of loin, and may provide an additional source of dietary vitamin D.

Methylenetetrahydrofolate reductase gene polymorphisms and the risk of colorectal carcinoma in a sample of Egyptian individuals.

PubMed

El Awady, Mostafa K; Karim, Amr M; Hanna, Laila S; El Husseiny, Lamia A; El Sahar, Medhat; Menem, Hanan A Abdel; Meguid, Nagwa A

2009-01-01

The study was planned as a pilot study to investigate two common polymorphisms in the MTHFR gene c.677C > T and c.1298A > C and their association with enhanced risk of colorectal cancer (CRC) in a sample of Egyptian individuals. Venous blood samples were withdrawn from 35 cases of CRC and 68 healthy controls. Specimens from colonic and rectal carcinoma tissues in addition to cancer free tissues were obtained from all cases. Frequencies of MTHFR677T and 1298C alleles were significantly higher among cases of CRC tumor tissues (50% and 56%, respectively) than germ line alleles in CRC patients (33% and 41%, respectively) and healthy controls (21% and 35%, respectively). Frequencies of heterozygous and homoyzgous polymorphisms of MTHFR at positions 677 and 1298 in carcinoma tissues were always the highest. At position 677, TT and CT genotype frequencies were 17% and 66% with an odds ratio {OR} of 11 [95% confidence interval {CI} 2.39-50.59] and OR 8.34 [95%CI 2.97-23.92], respectively, in carcinoma tissues. While in the germ line of patients the genotype frequencies of 677TT and CT were 6% and 54% with OR 1.57 [95%CI 0.26-9.51] and 2.99 [95%CI 1.25-7.12], respectively, compared to controls (6% and 29%, respectively). The combined genotype MTHFR 1298CC + AC frequencies were 86% with OR 3.71 [95%CI 1.28-10.78] in carcinoma tissues, 69% with OR 1.35 [95%CI 0.57-3.21] in germ line of patients and 62% in controls. The combined genotype 677CT plus any of the following genotypes 1298AA, AC or CC enhanced risk of CRC, when comparing germ line DNA polymorphism of patients versus peripheral blood DNA of control subjects with OR 4.5 [95%CI 0.94-21.56], OR 3.12 [95%CI 0.79-12.36] and OR 18 [95%CI 1.56-207.5], respectively, suggesting strong genetic predisposition of certain Egyptian population to CRC. These results suggested that at least one C to T polymorphism at 677MTHFR gene is required to significantly increase the risk for CRC development. Further large scale studies are required to confirm the present findings.

Towards High-Resolution Tissue Imaging Using Nanospray Desorption Electrospray Ionization Mass Spectrometry Coupled to Shear Force Microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nguyen, Son N.; Sontag, Ryan L.; Carson, James P.

Constant mode ambient mass spectrometry imaging (MSI) of tissue sections with high lateral resolution of better than 10 µm was performed by combining shear force microscopy with nanospray desorption electrospray ionization (nano-DESI). Shear force microscopy enabled precise control of the distance between the sample and nano-DESI probe during MSI experiments and provided information on sample topography. Proof-of-concept experiments were performed using lung and brain tissue sections representing spongy and dense tissues, respectively. Topography images obtained using shear force microscopy were comparable to the results obtained using contact profilometry over the same region of the tissue section. Variations in tissue heightmore » were found to be dependent on the tissue type and were in the range of 0-5 µm for lung tissue and 0-3 µm for brain tissue sections. Ion images of phospholipids obtained in this study are in good agreement with literature data. Normalization of nano-DESI MSI images to the signal of the internal standard added to the extraction solvent allowed us to construct high-resolution ion images free of matrix effects.« less

Tissue dissolution ability of sodium hypochlorite activated by photon-initiated photoacoustic streaming technique.

PubMed

Guneser, Mehmet Burak; Arslan, Dilara; Usumez, Aslihan

2015-05-01

The aim of this study was to evaluate the effect of the photon-initiated photoacoustic streaming (PIPS) technique on the pulp tissue-dissolving capacity of sodium hypochlorite (NaOCl) and compare it with the EndoActivator System (Dentsply Tulsa Dental Specialties, Tulsa, OK) and the Er:YAG laser with an endodontic fiber tip. Bovine pulp tissue samples (45 ± 15 mg) and dentin powder (10 mg) were placed in 1.5-mL Eppendorf tubes with 1 mL 5.25% NaOCl (Wizard; Rehber Kimya, Istanbul, Turkey) or distilled water (control) for 5 minutes with activation by the EndoActivator System, the Er:YAG laser with an endodontic fiber tip, and the PIPS technique. Nonactivated NaOCl served as the positive control. All testing procedures were performed at room temperature. The tissue samples were weighed before and after treatment, and the percentage of weight loss was calculated. The differences were statistically analyzed. The highest rate of tissue dissolution was observed in the NaOCl + Er:YAG group (P < .05). The NaOCl + PIPS group dissolved more bovine pulp tissue than the nonactivated NaOCl group (P < .05). There was no statistically significant difference between the rates of tissue dissolution of the NaOCl + EA and the nonactivated NaOCl groups (P > .05). NaOCl activation with the Er:YAG laser with an endodontic fiber tip was the most effective in bovine pulp tissue dissolution. The PIPS technique also promoted superior tissue-dissolving effects when compared with no activation. However, the EndoActivator System had no direct effect on tissue dissolution. Copyright © 2015 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Irradiation Does Not Increase the Penetration Depth of Doxorubicin in Normal Tissue After Pressurized Intra-peritoneal Aerosol Chemotherapy (PIPAC) in an Ex Vivo Model.

PubMed

Khosrawipour, Veria; Bellendorf, Alexander; Khosrawipour, Carolina; Hedayat-Pour, Yousef; Diaz-Carballo, David; Förster, Eckart; Mücke, Ralph; Kabakci, Burak; Adamietz, Irenäus Anton; Fakhrian, Khashayar

To compare the impact of single fractional with bi-fractional irradiation on the depth of doxorubicin penetration into the normal tissue after pressurized intra-peritoneal aerosol chemotherapy (PIPAC) in our ex vivo model. Fresh post mortem swine peritoneum was cut into 12 proportional sections. Two control samples were treated with PIPAC only (no irradiation), one sample on day 1, the other on day 2. Five samples were irradiated with 1, 2, 4, 7 or 14 Gy followed by PIPAC. Four samples were treated on day one with 0.5, 1, 2, 3.5 or 7 Gy and with the same radiation dose 24 h later followed by PIPAC. Doxorubicin was aerosolized in an ex vivo PIPAC model at 12 mmHg/36°C. In-tissue doxorubicin penetration was measured using fluorescence microscopy on frozen thin sections. Doxorubicin penetration (DP) after PIPAC for the control samples was 407 μm and 373 μm, respectively. DP for samples with single fraction irradiation was 396 μm after 1 Gy, 384 μm after 2 Gy, 327 μm after 4 Gy, 280 μm after 7 Gy and 243 μm after 14 Gy. DP for samples with 2 fractions of irradiation was 376 μm after 0.5+0.5 Gy, 363 μm after 1+1 Gy, 372 μm after 2+2 Gy, 341 μm after 3.5+3.5 and 301 μm after 7+7 Gy irradiation. Fractionating of the irradiation did not significantly change DP into normal tissue. Irradiation does not increase the penetration depth of doxorubicin into the normal tissue but might have a limiting impact on penetration and distribution of doxorubicin. Further studies are warranted to investigate the impact of addition of irradiation to PIPAC of tumor cells and to find out if irradiation can be used safely as chemopotenting agent for patients with peritoneal metastases treated with PIPAC. Copyright © 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Use of a new jumbo forceps improves tissue acquisition of Barrett's esophagus surveillance biopsies.

PubMed

Komanduri, Sri; Swanson, Garth; Keefer, Laurie; Jakate, Shriram

2009-12-01

The major risk factors for the development of esophageal adenocarcinoma remain long-standing GERD and resultant Barrett's esophagus (BE). Finding the exact method of adequate tissue sampling for surveillance of dysplasia in BE remains a dilemma. We prospectively compared standard large-capacity biopsy forceps with a new jumbo biopsy forceps for dysplasia detection in BE. Prospective, single-center investigation. We prospectively enrolled 32 patients undergoing surveillance endoscopy for BE. Biopsy samples were obtained in paired fashion alternating between the experimental (jumbo) and control (large-capacity) forceps. Each sample was assessed for histopathology, specimen size, and adequacy. A total of 712 specimens were available for analysis for this investigation. Six patients were found to have dysplasia, and in 5 of those patients, the dysplasia was only detected with the jumbo forceps. The mean width was significantly greater in the Radial Jaw 4 jumbo group (3.3 mm vs 1.9 mm [P < .005]) as was the mean depth (2.0 mm vs 1.1 mm [P < .005]). Sixteen percent of samples obtained with the standard forceps provided an adequate sample, whereas the jumbo forceps provided an adequate sample 79% of the time (P < .05). A lack of a validated index for assessment of tissue adequacy in BE. The Radial Jaw 4 jumbo biopsy forceps significantly improves dysplasia detection and adequate tissue sampling in patients undergoing endoscopy for BE.

A prospective, multicentre study of moxifloxacin concentrations in the sinus mucosa tissue of patients undergoing elective surgery of the sinus.

PubMed

Gehanno, P; Darantière, S; Dubreuil, C; Chobaut, J C; Bobin, S; Pages, J C; Renou, G; Bobin, F; Arvis, P; Stass, H

2002-05-01

A pharmacokinetic study was carried out to determine moxifloxacin concentrations in sinus tissue, after oral moxifloxacin 400 mg once daily for 5 days to patients with chronic sinusitis, undergoing elective sinus surgery. Patients were randomly allocated to one of seven treatment groups, in which tissues were sampled 2, 3, 4, 6, 12, 24 or 36 h post-dose. A control group with non-infected nasal polyps was also included. Forty-eight patients (13 female, 35 male, mean age 47.1 years) were allocated to one of each active treatment group (n = 42) or to the control group (n = 6). Tissue and plasma samples were taken simultaneously and stored frozen until assayed by HPLC. Thirty-nine patients were fully valid for pharmacokinetic analysis. The geometric mean moxifloxacin plasma concentration increased from 2.32 mg/L at 2 h to a maximum of 3.37 mg/L at 4 h post-dose, decreasing to 0.37 mg/L at 36 h post-dose. The moxifloxacin concentration in sinus mucosa was consistently greater than that in plasma being 4.56-5.73 mg/kg from 2 to 6 h and 2.81-1.25 mg/kg from 12 to 36 h post-dose. The elimination rates in plasma and sinus tissues were similar. The tissue/plasma ratio was c. 200% between 2 and 6 h, and up to 328.9% at 36 h. Results were similar whatever the site of tissue sampling (maxillary sinus, anterior ethmoid sinus or nasal polyps). Tissue levels exceeded the MIC(90) of all pathogens commonly causing acute sinusitis (e.g. 5-30 x MIC for Streptococcus pneumoniae: 0.25 mg/L). These results sup-port the use of moxifloxacin 400 mg once daily as a regimen for the treatment of sinus infections.

Inflammatory bowel disease exacerbation associated with Epstein-Barr virus infection.

PubMed

Dimitroulia, Evangelia; Pitiriga, Vassiliki C; Piperaki, Evangelia-Theophano; Spanakis, Nicholas E; Tsakris, Athanassios

2013-03-01

Epstein-Barr virus infection is associated with inflammatory bowel disease, but its role as a pathogenetic or exacerbating factor remains unclear. The aim of this study was to evaluate the association between Epstein-Barr virus infection and inflammatory bowel disease, particularly in regard to exacerbation of disease activity. This was a nonrandomized crosssectional study in subgroups of patients with inflammatory bowel disease compared with a control group with noninflammatory disease. Participants were patients treated for ulcerative colitis or Crohn's disease and individuals undergoing evaluation for noninflammatory disease recruited from 2 urban adult gastrointestinal referral centers in Greece. Diagnosis of inflammatory bowel disease was based on standard clinical and endoscopic criteria. Demographic and clinical characteristics of all participants were recorded. Whole blood samples and fresh tissue samples from biopsy of intestinal sites were obtained from each participant. The presence of Epstein-Barr virus was determined by amplifying the LMP1 gene of the virus in blood and intestinal tissue samples. The study comprised 94 patients with inflammatory bowel disease (63 with ulcerative colitis and 31 with Crohn's disease) and 45 controls with noninflammatory disease. Of the 94 patients, 67 (71.3%) had disease exacerbation and 27 (28.7%) were in remission. The prevalence of Epstein-Barr virus genome was significantly higher in patients than in controls for intestinal tissue (44 patients, 46.8% vs 6 controls, 13.3%; p = 0.001), but not for whole blood (24 patients, 25.5% vs 9 controls, 20%; p = 0.3). The viral genome was found significantly more frequently in intestinal samples from patients with disease exacerbation compared with patients in remission (38 patients with exacerbation, 56.7% vs 6 patients in remission, 22.2%; p = 0.001), but no significant difference was found for whole blood (18 patients with exacerbation, 26.8% vs 6 patients in remission, 22.2%; p = 0.79). Neither disease exacerbation nor the presence of virus genome was related to demographic or clinical characteristics. The exact location of Epstein-Barr virus in the intestinal tissues could not be specified because morphological data by immunohistochemistry or in situ hybridization were not available. Although causality could not be determined, the significantly higher prevalence of Epstein-Barr virus in intestinal tissue from patients with inflammatory bowel disease compared with controls and in patients with exacerbation compared with patients in remission suggests a potential viral involvement in the severity of inflammatory bowel disease. These findings merit further investigation in view of a potential for usefulness of antiviral therapy against Epstein-Barr virus infection in patients with exacerbation of inflammatory bowel disease.

Cell and tissue microarray technologies for protein and nucleic acid expression profiling.

PubMed

Cardano, Marina; Diaferia, Giuseppe R; Falavigna, Maurizio; Spinelli, Chiara C; Sessa, Fausto; DeBlasio, Pasquale; Biunno, Ida

2013-02-01

Tissue microarray (TMA) and cell microarray (CMA) are two powerful techniques that allow for the immunophenotypical characterization of hundreds of samples simultaneously. In particular, the CMA approach is particularly useful for immunophenotyping new stem cell lines (e.g., cardiac, neural, mesenchymal) using conventional markers, as well as for testing the specificity and the efficacy of newly developed antibodies. We propose the use of a tissue arrayer not only to perform protein expression profiling by immunohistochemistry but also to carry out molecular genetics studies. In fact, starting with several tissues or cell lines, it is possible to obtain the complete signature of each sample, describing the protein, mRNA and microRNA expression, and DNA mutations, or eventually to analyze the epigenetic processes that control protein regulation. Here we show the results obtained using the Galileo CK4500 TMA platform.

Effects of oriental sweet gum storax on porcine wound healing.

PubMed

Ocsel, Hakan; Teke, Zafer; Sacar, Mustafa; Kabay, Burhan; Duzcan, S Ender; Kara, Inci Gokalan

2012-08-01

The objective of the present study was to assess the effects of oriental sweet gum (Liquidambar orientalis Mill.) storax on partial-thickness and full-thickness wounds compared to conventional wound dressings in a porcine model. Six young Yorkshire pigs were used. Sixteen square excisional wounds measuring 3 × 3 cm were performed per animal. The wounds were allocated to one of the four treatment modalities: storax, hydrocolloid dressing, silver sulfadiazine, and control groups. Partial-thickness wounds were created in two pigs, and tissue samples were harvested on days 4 and 8, respectively. Full-thickness wounds were created in four pigs, and tissue samples were taken on days 4, 8, 14, and 21, respectively. Histologically, all wounds were examined for re-epithelialization and granulation tissue formation. Tissue hydroxyproline content and wound contraction areas were measured. In storax-applied group, there was a greater depth of granulation tissue at 4 and 8 days compared to all other groups (p < .0125), and there was a faster re-epithelialization at 21 days compared to both hydrocolloid dressing and control groups in full-thickness wounds (p < .0125). Tissue hydroxyproline content and wound contraction did not differ significantly between the groups. The results of this study indicate that topical application of storax enhanced both re-epithelialization and granulation tissue formation in full-thickness wounds. Further studies are indicated in this important area of wound healing research to evaluate the clinical efficacy of this storax and search for the mechanisms that explain its effects.

Chromium in urothelial carcinoma of the bladder.

PubMed

Golabek, Tomasz; Socha, Katarzyna; Kudelski, Jacek; Darewicz, Barbara; Markiewicz-Zukowska, Renata; Chlosta, Piotr; Borawska, Maria

2017-12-23

Many epidemiological and experimental studies report a strong role of chemical carcinogens in the etiology of bladder cancer. However, the involvement of heavy metals in tumourigenesis of urothelial carcinoma of the bladder has been poorly investigated. Therefore, the aim of this study was to examine the relationship between chromium (Cr) and bladder cancer. Chromium concentration in two 36-sample series of bladder cancer tissue and sera from patients with this neoplasm were matched with those of a control group. The amount of trace elements in every tissue sample was determined using atomic absorption spectrometry. This was correlated with tumour stage. While the median chromium concentration levels reached statistically higher values in the bladder cancer tissue, compared with the non-cancer tissue (99.632ng/g and 33.144ng/g, respectively; p<0.001), the median Cr levels in the sera of the patients with this carcinoma showed no statistical difference when compared to those of the control group (0.511μg/l and 0.710μg/l, respectively; p=0.408). The median levels of Cr in the bladder tissue, depending on the stage of the tumour, compared with the tissue without the neoplasm, observed the same relationship for both non-muscle invasive and muscle-invasive tumours (p<0.001 and p<0.01, respectively). This study shows that patients with urothelial carcinoma of the bladder had higher tissue Cr levels than people without tumour, while no difference was found in the Cr serum levels between the two groups of patients under investigation.

Expression of GRIM-19 in adenomyosis and its possible role in pathogenesis.

PubMed

Wang, Jing; Deng, Xiaohui; Yang, Yang; Yang, Xingsheng; Kong, Beihua; Chao, Lan

2016-04-01

To study the expression of the gene associated with retinoid-interferon (IFN)-induced mortality 19 (GRIM-19) in the endometrial tissue of patients with adenomyosis and to describe the possible pathogenic mechanisms of this phenomenon. Experimental study using human samples and cell lines. University-affiliated hospital. Ectopic and eutopic endometrial tissues were obtained from 30 patients with adenomyosis, whereas normal endometrial specimens were obtained from 10 control patients without adenomyosis. Patients with rapid pathology report-confirmed adenomyosis were recruited, and eutopic and ectopic endometrial tissue samples were collected from patients who had undergone hysterectomies by either the transabdominal or laparoscopic method at Qilu Hospital. Normal endometrial tissue was collected from a group of control patients without adenomyosis. Immunohistochemistry (IHC) was performed to evaluate the expression of GRIM-19, phospho-signal transducer and activator of transcription 3 (Y705) (Y705) (pSTAT3(Y705)), and vascular endothelial growth factor (VEGF) in endometrial tissue samples. The protein levels of GRIM-19, pSTAT3(Y705), STAT3, and VEGF were detected by Western blot. Apoptosis in endometrial specimens was assayed by TUNEL. Immunohistochemistry with an antibody directed against CD34 was performed to detect new blood vessels in the endometrial tissue. GRIM-19 small interfering RNA and a recombinant plasmid carrying GRIM-19 were constructed to evaluate the effects of GRIM-19 on the downstream factors pSTAT3(Y705), STAT3, and VEGF in Ishikawa cells. The expression of GRIM-19 was down-regulated in the eutopic endometria of patients with adenomyosis compared with the endometria of patients in the control group, and it was further reduced in the endometrial glandular epithelial cells of adenomyotic lesions. Apoptosis was reduced in the eutopic endometrium compared with the control group, and it was significantly reduced in ectopic endometrial tissues. In addition, the ectopic and eutopic endometria of patients with adenomyosis displayed a much higher microvessel density. In the eutopic and ectopic endometria of patients with adenomyosis, the expression levels of pSTAT3(Y705) and VEGF were significantly higher than in the controls. Furthermore, down-regulation of GRIM-19 in Ishikawa cells significantly promoted the activation of both pSTAT3(Y705) and its dependent gene VEGF. Aberrant expression of GRIM-19 may be associated with adenomyosis through the regulation of apoptosis and angiogenesis. Copyright © 2016 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Exploring Raman spectroscopy for the evaluation of glaucomatous retinal changes

NASA Astrophysics Data System (ADS)

Wang, Qi; Grozdanic, Sinisa D.; Harper, Matthew M.; Hamouche, Nicolas; Kecova, Helga; Lazic, Tatjana; Yu, Chenxu

2011-10-01

Glaucoma is a chronic neurodegenerative disease characterized by apoptosis of retinal ganglion cells and subsequent loss of visual function. Early detection of glaucoma is critical for the prevention of permanent structural damage and irreversible vision loss. Raman spectroscopy is a technique that provides rapid biochemical characterization of tissues in a nondestructive and noninvasive fashion. In this study, we explored the potential of using Raman spectroscopy for detection of glaucomatous changes in vitro. Raman spectroscopic imaging was conducted on retinal tissues of dogs with hereditary glaucoma and healthy control dogs. The Raman spectra were subjected to multivariate discriminant analysis with a support vector machine algorithm, and a classification model was developed to differentiate disease tissues versus healthy tissues. Spectroscopic analysis of 105 retinal ganglion cells (RGCs) from glaucomatous dogs and 267 RGCs from healthy dogs revealed spectroscopic markers that differentiated glaucomatous specimens from healthy controls. Furthermore, the multivariate discriminant model differentiated healthy samples and glaucomatous samples with good accuracy [healthy 89.5% and glaucomatous 97.6% for the same breed (Basset Hounds); and healthy 85.0% and glaucomatous 85.5% for different breeds (Beagles versus Basset Hounds)]. Raman spectroscopic screening can be used for in vitro detection of glaucomatous changes in retinal tissue with a high specificity.

Liquid chromatography tandem mass spectrometry method using solid-phase extraction and bead-beating-assisted matrix solid-phase dispersion to quantify the fungicide tebuconazole in controlled frog exposure study: analysis of water and animal tissue.

PubMed

Hansen, Martin; Poulsen, Rikke; Luong, Xuan; Sedlak, David L; Hayes, Tyrone

2014-11-01

This paper presents the development, optimization, and validation of a LC-MS/MS methodology to determine the concentration of the antifungal drug and fungicide tebuconazole in a controlled exposure study of African clawed frogs (Xenopus laevis). The method is validated on animal tank water and on tissue from exposed and non-exposed adult X. laevis. Using solid-phase extraction (SPE), the analytical method allows for quantification of tebuconazole at concentrations as low as 3.89 pg mL(-1) in 10 mL water samples. Using bead-beating-assisted matrix solid-phase dispersion (MSPD), it was possible to quantify tebuconazole down to 0.63 pg mg(-1) wet weight liver using 150 mg tissue. The deuterated analogue of tebuconazole was used as internal standard, and ensured method accuracy in the range 80.6-99.7% for water and 68.1-109% for tissue samples. The developed method was successfully applied in a 4-week X. laevis repeated-exposure study, revealing high levels of tebuconazole residues in adipose and liver tissue, and with experimental bioconcentration factors up to 18,244 L kg(-1).

Exploring Raman spectroscopy for the evaluation of glaucomatous retinal changes.

PubMed

Wang, Qi; Grozdanic, Sinisa D; Harper, Matthew M; Hamouche, Nicolas; Kecova, Helga; Lazic, Tatjana; Yu, Chenxu

2011-10-01

Glaucoma is a chronic neurodegenerative disease characterized by apoptosis of retinal ganglion cells and subsequent loss of visual function. Early detection of glaucoma is critical for the prevention of permanent structural damage and irreversible vision loss. Raman spectroscopy is a technique that provides rapid biochemical characterization of tissues in a nondestructive and noninvasive fashion. In this study, we explored the potential of using Raman spectroscopy for detection of glaucomatous changes in vitro. Raman spectroscopic imaging was conducted on retinal tissues of dogs with hereditary glaucoma and healthy control dogs. The Raman spectra were subjected to multivariate discriminant analysis with a support vector machine algorithm, and a classification model was developed to differentiate disease tissues versus healthy tissues. Spectroscopic analysis of 105 retinal ganglion cells (RGCs) from glaucomatous dogs and 267 RGCs from healthy dogs revealed spectroscopic markers that differentiated glaucomatous specimens from healthy controls. Furthermore, the multivariate discriminant model differentiated healthy samples and glaucomatous samples with good accuracy [healthy 89.5% and glaucomatous 97.6% for the same breed (Basset Hounds); and healthy 85.0% and glaucomatous 85.5% for different breeds (Beagles versus Basset Hounds)]. Raman spectroscopic screening can be used for in vitro detection of glaucomatous changes in retinal tissue with a high specificity.

Suppression of Botrytis cinerea on necrotic grapevine tissues by early-season applications of natural products and biological control agents.

PubMed

Calvo-Garrido, Carlos; Viñas, Inmaculada; Elmer, Philip A G; Usall, Josep; Teixidó, Neus

2014-04-01

Necrotic tissues within grape (Vitis vinifera) bunches represent an important source of Botrytis cinerea inoculum for Botrytis bunch rot (BBR) at harvest in vineyards. This research quantified the incidence of B. cinerea on necrotic floral and fruit tissues and the efficacy of biologically based treatments for suppression of B. cinerea secondary inoculum within developing bunches. At veraison (2009 and 2010), samples of aborted flowers, aborted fruits and calyptras were collected, and the incidence and sporulation of B. cinerea were determined. Aborted fruits presented significantly higher incidence in untreated samples. Early-season applications of Candida sake plus Fungicover®, Fungicover alone or Ulocladium oudemansii significantly reduced B. cinerea incidence on aborted flowers and calyptras by 46-85%. Chitosan treatment significantly reduced B. cinerea incidence on calyptras. None of the treatments reduced B. cinerea incidence on aborted fruits. Treatments significantly reduced sporulation severity by 48% or more. Treatments were effective at reducing B. cinerea secondary inoculum on necrotic tissues, in spite of the variable control on aborted fruits. This is the first report to quantify B. cinerea on several tissues of bunch trash and to describe the effective suppression of saprophytic B. cinerea inoculum by biologically based treatments. © 2013 Society of Chemical Industry.

Tc17 cells in patients with uterine cervical cancer.

PubMed

Zhang, Yan; Hou, Fei; Liu, Xin; Ma, Daoxin; Zhang, Youzhong; Kong, Beihua; Cui, Baoxia

2014-01-01

The existence of Tc17 cells was recently shown in several types of infectious and autoimmune diseases, but their distribution and functions in uterine cervical cancer (UCC) have not been fully elucidated. The frequency of Tc17 cells in peripheral blood samples obtained from UCC patients, cervical intraepithelial neoplasia (CIN) patients and healthy controls was determined by flow cytometry. Besides, the prevalence of Tc17 cells and their relationships to Th17 cells and Foxp3-expressing T cells as well as microvessels in tissue samples of the patients were assessed by immunohistochemistry staining. Compared to controls, patients with UCC or CIN had a higher proportion of Tc17 cells in both peripheral blood and cervical tissues, but the level of Tc17 cells in UCC tissues was significantly higher than that in CIN tissues. Besides, the increased level of Tc17 in UCC patients was associated with the status of pelvic lymph node metastases and increased microvessel density. Finally, significant correlations of infiltration between Tc17 cells and Th17 cells or Foxp3-expressing T cells were observed in UCC and CIN tissues. This study indicates that Tc17 cell infiltration in cervical cancers is associated with cancer progression accompanied by increased infiltrations of Th17 cells and regulatory T cells as well as promoted tumor vasculogenesis.

Expression of pleiotrophin in small cell lung cancer.

PubMed

Wang, H Q; Wang, J

2015-01-01

Pleiotrophin (PTN) is a kind of heparin binding growth factor closely related to tumor progression. This study aimed to discuss the significance of the expression of PTN in benign and malignant lung cancer tissues, especially small cell lung cancer. Lung cancer samples were collected for study and lung tissue samples with benign lesions were taken as controls. The expression of PTN was detected using tissue chip combined with the immunohistochemical method, and the differences of small cell lung cancer with non-small cell lung cancer and benign lesion tissue were compared. It was found that PTN expression was mainly located in the cytoplasm and membrane of cells; PTN expression in the lung cancer group was higher than that in the control group (p < 0.01), and PTN expression in the small cell cancer group was higher than that in the squamous carcinoma group and glandular cancer group (p < 0.05). In addition, PTN expression quantity in patients with lung cancer were in close correlation with TNM staging, pathological type and tumor differentiation degree (p < 0.05). PTN was found to express abnormally high in lung cancer, especially small cell lung cancer tissue. PTN is most likely to be a new tumor marker for diagnosis and prognosis of lung cancer.

Acaricide Residues in Laying Hens Naturally Infested by Red Mite Dermanyssus gallinae

PubMed Central

Marangi, Marianna; Morelli, Vincenzo; Pati, Sandra; Camarda, Antonio; Cafiero, Maria Assunta; Giangaspero, Annunziata

2012-01-01

In the poultry industry, control of the red mite D. gallinae primarily relies worldwide on acaricides registered for use in agriculture or for livestock, and those most widely used are carbamates, followed by amidines, pyrethroids and organophosphates. Due to the repeated use of acaricides - sometimes in high concentrations - to control infestation, red mites may become resistant, and acaricides may accumulate in chicken organs and tissues, and also in eggs. To highlight some situations of misuse/abuse of chemicals and of risk to human health, we investigated laying hens, destined to the slaughterhouse, for the presence of acaricide residues in their organs and tissues. We used 45 hens from which we collected a total of 225 samples from the following tissues and organs: skin, fat, liver, muscle, hearth, and kidney. In these samples we analyzed the residual contents of carbaryl and permethrin by LC-MS/MS. Ninety-one (40.4%) samples were positive to carbaryl and four samples (1.7%) were positive to permethrin. Concentrations of carbaryl exceeding the detection limit (0.005 ppm) were registered in the skin and fat of birds from two farms (p<0.01), although these concentrations remained below the maximum residue limit (MRLs) (0.05 ppm) (p<0.01). All organs/tissues of hens from a third farm were significantly more contaminated, with skin and muscle samples exceeding the MRL (0.05 ppm) (p<0.01) of carbaryl in force before its use was banned. Out of 45 chickens tested, 37 (82.2%) were found to be contaminated by carbaryl, and 4 (8.8%) by permethrin. The present study is the first report on the presence of pesticides banned by the EU (carbaryl) or not licensed for use (permethrin) in the organs and tissues of laying hens, which have been treated against red mites, and then slaughtered for human consumption at the end of their life cycle. PMID:22363736

Acaricide residues in laying hens naturally infested by red mite Dermanyssus gallinae.

PubMed

Marangi, Marianna; Morelli, Vincenzo; Pati, Sandra; Camarda, Antonio; Cafiero, Maria Assunta; Giangaspero, Annunziata

2012-01-01

In the poultry industry, control of the red mite D. gallinae primarily relies worldwide on acaricides registered for use in agriculture or for livestock, and those most widely used are carbamates, followed by amidines, pyrethroids and organophosphates. Due to the repeated use of acaricides--sometimes in high concentrations--to control infestation, red mites may become resistant, and acaricides may accumulate in chicken organs and tissues, and also in eggs. To highlight some situations of misuse/abuse of chemicals and of risk to human health, we investigated laying hens, destined to the slaughterhouse, for the presence of acaricide residues in their organs and tissues. We used 45 hens from which we collected a total of 225 samples from the following tissues and organs: skin, fat, liver, muscle, hearth, and kidney. In these samples we analyzed the residual contents of carbaryl and permethrin by LC-MS/MS.Ninety-one (40.4%) samples were positive to carbaryl and four samples (1.7%) were positive to permethrin. Concentrations of carbaryl exceeding the detection limit (0.005 ppm) were registered in the skin and fat of birds from two farms (p<0.01), although these concentrations remained below the maximum residue limit (MRLs) (0.05 ppm) (p<0.01). All organs/tissues of hens from a third farm were significantly more contaminated, with skin and muscle samples exceeding the MRL (0.05 ppm) (p<0.01) of carbaryl in force before its use was banned. Out of 45 chickens tested, 37 (82.2%) were found to be contaminated by carbaryl, and 4 (8.8%) by permethrin. The present study is the first report on the presence of pesticides banned by the EU (carbaryl) or not licensed for use (permethrin) in the organs and tissues of laying hens, which have been treated against red mites, and then slaughtered for human consumption at the end of their life cycle.

Incidence of Propionibacterium acnes in initially culture-negative thioglycollate broths-a prospective cohort study at a Danish University Hospital.

PubMed

Kvich, L; Jensen, P Ø; Justesen, U S; Bjarnsholt, T

2016-11-01

The aim of this study was to prospectively investigate the incidence of Propionibacterium acnes in thioglycollate broths reported as culture-negative at the Department of Clinical Microbiology, Rigshospitalet, to evaluate whether 5 days of incubation was enough to find all relevant cases. Five hundred thioglycollate broths reported as culture-negative after 5 days were consecutively collected and incubated for at least a further 9 days (at least 14 days of incubation in total). Only tissue samples from sterile sites of the body (n = 298), bone tissue (n = 197) and foreign material (n = 5) were included in this study. Samples were divided into two groups: infected group and control group. This made it possible to compare findings between groups, thereby making it possible to estimate the level of true-positive findings and contamination. Samples from 296 participants were included in this study. After exclusion criteria were met, P. acnes was cultured from ten out of 151 patients (6.6%) in the infected group and from one out of 138 participants (0.7%) in the control group. This resulted in more findings of P. acnes in the infected group on day 14 than on day 5 (p 0.002). Furthermore, P. acnes was cultured more often from bone tissue and tissue surrounding foreign materials on day 14 than on day 5 (p 0.04). Clinical microbiology laboratories should consider incubating thioglycollate broths for at least 14 days to find all relevant cases of P. acnes, especially when it comes to bone tissue and tissue surrounding foreign materials. Copyright © 2016 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Cobalt deposition in mineralized bone tissue after metal-on-metal hip resurfacing: Quantitative μ-X-ray-fluorescence analysis of implant material incorporation in periprosthetic tissue.

PubMed

Hahn, Michael; Busse, Björn; Procop, Mathias; Zustin, Jozef; Amling, Michael; Katzer, Alexander

2017-10-01

Most resurfacing systems are manufactured from cobalt-chromium alloys with metal-on-metal (MoM) bearing couples. Because the quantity of particulate metal and corrosion products which can be released into the periprosthetic milieu is greater in MoM bearings than in metal-on-polyethylene (MoP) bearings, it is hypothesized that the quantity and distribution of debris released by the MoM components induce a compositional change in the periprosthetic bone. To determine the validity of this claim, nondestructive µ-X-ray fluorescence analysis was carried out on undecalcified histological samples from 13 femoral heads which had undergone surface replacement. These samples were extracted from the patients after gradient time points due to required revision surgery. Samples from nonintervened femoral heads as well as from a MoP resurfaced implant served as controls. Light microscopy and µ-X-ray fluorescence analyses revealed that cobalt debris was found not only in the soft tissue around the prosthesis and the bone marrow, but also in the mineralized bone tissue. Mineralized bone exposed to surface replacements showed significant increases in cobalt concentrations in comparison with control specimens without an implant. A maximum cobalt concentration in mineralized hard tissue of up to 380 ppm was detected as early as 2 years after implantation. Values of this magnitude are not found in implants with a MoP surface bearing until a lifetime of more than 20 years. This study demonstrates that hip resurfacing implants with MoM bearings present a potential long-term health risk due to rapid cobalt ion accumulation in periprosthetic hard tissue. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 1855-1862, 2017. © 2016 Wiley Periodicals, Inc.

Microscopic examination of the intestinal wall and selected organs of minipigs orally supplemented with humic acids.

PubMed

Büsing, Kirsten; Elhensheri, Mohamed; Entzian, Kristin; Meyer, Udo; Zeyner, Annette

2014-04-01

Humic acids are used to prophylactically treat intestinal diseases in a wide number of species, yet the mechanism of action remains unknown. The general assumption has been that humic acids act locally; however studies using young piglets show orally supplemented humic acids can penetrate the intestinal wall, and thus potentially act systemically. The objective of this study was to determine if humic acids could also cross the intestinal barrier in adult pigs and be detected in other organs. Adult minipigs (>18 months old) orally received either 1g humic acids/kg body weight (verum, n=3) or placebo (control, n=3), for 2 weeks. At the end of the feeding period tissue samples were harvested from the intestine, various glands and organs. Unstained tissue samples were examined by light microscopy for the presence of humic acid particles. No humic acid particles were detected in any of the unstained tissues from verum or control pigs. Copyright © 2014 Elsevier Ltd. All rights reserved.

[The expression and significance of RORγT in periapical granulomas and radicular cysts].

PubMed

Lang, Xiao-ying; Li, Song

2014-08-01

To identify retinoic acid-related orphan nuclear receptor-γT (RORγT), the specific markers of T helper 17 (Th17) cells by immunohistochemical analysis to confirm the presence of Th17 cells in periapical lesions. Eighteen radicular cysts (RCs) and 22 periapical granulomas (PGs) were collected in the Department of Oral Pathology after periapical surgery as the experimental samples. Five alveolar bone samples were obtained from a group of impacted third molars recommended for extraction as the control samples. The protein expression of RORγT was measured by immunohistochemical analysis for all samples. In addition, the protein expression of IL-17 was measured at the same time. Statistical analysis was performed using SPSS 17.0 software package to evaluate the differences of expression of RORγT and IL-17 according to type of lesion (PG vs. RC vs. control group) and intensity of the inflammatory infiltration (mild vs. moderate vs. severe vs. control group). RORγT+ cells were detected in all periapical lesions tissues, and the expression of RORγT was significantly higher in periapical lesions than in normal tissues which had no expression of RORγT (P<0.05). Significant differences in the expression of RORγT were observed among healthy tissues, lesions with mild inflammation, moderate inflammation and severe inflammation (P<0.05), respectively. Positive correlations between RORγT and IL-17 protein levels were observed in PGs (r=0.935,P<0.05) and RCs (r=0.803,P<0.05), respectively. The results demonstrates a significant increase in the expression of RORγT in patients suffering from periapical lesions in comparison with normal control subjects, indicating that Th17 cells are more likely to exist in periapical lesions.

The presence of enterovirus, adenovirus, and parvovirus B19 in myocardial tissue samples from autopsies: an evaluation of their frequencies in deceased individuals with myocarditis and in non-inflamed control hearts.

PubMed

Nielsen, Trine Skov; Hansen, Jakob; Nielsen, Lars Peter; Baandrup, Ulrik Thorngren; Banner, Jytte

2014-09-01

Multiple viruses have been detected in cardiac tissue, but their role in causing myocarditis remains controversial. Viral diagnostics are increasingly used in forensic medicine, but the interpretation of the results can sometimes be challenging. In this study, we examined the prevalence of adenovirus, enterovirus, and parvovirus B19 (PVB) in myocardial autopsy samples from myocarditis related deaths and in non-inflamed control hearts in an effort to clarify their significance as the causes of myocarditis in a forensic material. We collected all autopsy cases diagnosed with myocarditis from 1992 to 2010. Eighty-four suicidal deaths with morphologically normal hearts served as controls. Polymerase chain reaction was used for the detection of the viral genomes (adenovirus, enterovirus, and PVB) in myocardial tissue specimens. The distinction between acute and persistent PVB infection was made by the serological determination of PVB-specific immunoglobulins M and G. PVB was detected in 33 of 112 (29 %) myocarditis cases and 37 of 84 (44 %) control cases. All of the samples were negative for the presence of adenovirus and enterovirus. Serological evidence of an acute PVB infection, determined by the presence of immunoglobulin M, was only present in one case. In the remaining cases, PVB was considered to be a bystander with no or limited association to myocardial inflammation. In this study, adenovirus, enterovirus, and PVB were found to be rare causes of myocarditis. The detection of PVB in myocardial autopsy samples most likely represents a persistent infection with no or limited association with myocardial inflammation. The forensic investigation of myocardial inflammation demands a thorough examination, including special attention to non-viral causes and requires a multidisciplinary approach.

PAH bioconcentration in Mytilus sp from Sinclair Inlet, WA

DOE Office of Scientific and Technical Information (OSTI.GOV)

Frazier, J.; Young, D.; Ozretich, R.

1995-12-31

Approximately 20 polynuclear aromatic hydrocarbons (PAH) were measured by GC/MS in seawater and whole soft tissues of the intertidal mussel Mytilus sp. collected in July 1991 within and around Puget Sound`s Sinclair Inlet. Low variability was observed in the water concentrations collected over three days at control sites, yielding reliable values for the exposure levels experienced by this bioindicator mollusk. Mean water concentrations of acenaphthene, phenanthrene, and fluoranthene in the control region were 2.7 {+-} 0.8, 2.8 {+-} 0.8, and 3.1 {+-} 0.7 ng/liter, respectively. Levels measured near sites of vessel activity were higher but much more variable; this reducedmore » the reliability of the tissue/water bioconcentration factors (BCF) obtained from these samples. An empirical model relating values of Log BCF and Log Kow for the control zone samples supports the utility of this estuarine bioindicator for monitoring general levels of PAH in nearshore surface waters.« less

The protective effect of diosmin on hepatic ischemia reperfusion injury: an experimental study

PubMed Central

Tanrikulu, Yusuf; Şahin, Mefaret; Kismet, Kemal; Kilicoglu, Sibel Serin; Devrim, Erdinc; Tanrikulu, Ceren Sen; Erdemli, Esra; Erel, Serap; Bayraktar, Kenan; Akkus, Mehmet Ali

2013-01-01

Liver ischemia reperfusion injury (IRI) is an important pathologic process leading to bodily systemic effects and liver injury. Our study aimed to investigate the protective effects of diosmin, a phlebotrophic drug with antioxidant and anti-inflammatory effects, in a liver IRI model. Forty rats were divided into 4 groups. Sham group, control group (ischemia-reperfusion), intraoperative treatment group, and preoperative treatment group. Ischemia reperfusion model was formed by clamping hepatic pedicle for a 60 minute of ischemia followed by liver reperfusion for another 90 minutes. Superoxide dismutase (SOD) and catalase (CAT) were measured as antioaxidant enzymes in the liver tissues, and malondialdehyde (MDA) as oxidative stress marker, xanthine oxidase (XO) as an oxidant enzyme and glutathione peroxidase (GSH-Px) as antioaxidant enzyme were measured in the liver tissues and the plasma samples. Hepatic function tests were lower in treatment groups than control group (p<0.001 for ALT and AST). Plasma XO and MDA levels were lower in treatment groups than control group, but plasma GSH-Px levels were higher (p<0.05 for all). Tissue MDA levels were lower in treatment groups than control group, but tissue GSH-Px, SOD, CAT and XO levels were higher (p<0.05 for MDA and p<0.001 for others). Samples in control group histopathologically showed morphologic abnormalities specific to ischemia reperfusion. It has been found that both preoperative and intraoperative diosmin treatment decreases cellular damage and protects cells from toxic effects in liver IRI. As a conclusion, diosmin may be used as a protective agent against IRI in elective and emergent liver surgical operations. PMID:24289756

Skin-infiltrating, interleukin-22-producing T cells differentiate pediatric psoriasis from adult psoriasis.

PubMed

Cordoro, Kelly M; Hitraya-Low, Maria; Taravati, Keyon; Sandoval, Priscila Munoz; Kim, Esther; Sugarman, Jeffrey; Pauli, Mariela L; Liao, Wilson; Rosenblum, Michael D

2017-09-01

Evidence from adult psoriasis studies implicates an imbalance between regulatory and effector T cells, particularly T H -17-producing T cells, in the pathogenesis of psoriasis. Little is known about the immunopathology of psoriasis in children. We sought to functionally characterize the inflammatory cell profiles of psoriatic plaques from pediatric patients and compare them with healthy, age-matched controls and adult psoriasis patients. Skin samples from pediatric psoriasis patients and healthy controls were analyzed by multiparameter flow cytometry to determine the dominant immune cell subsets present and cytokines produced. Lesional tissue from pediatric psoriasis patients had significantly increased interleukin (IL) 22 derived from CD4 + and CD8 + cells compared with the tissues from healthy pediatric controls and adult psoriasis patients. Tissue from pediatric psoriasis patients had significantly less elevation of IL-17 derived from CD4 + and CD8 + cells compared with the tissue from adult psoriasis patients. In contrast with the lesions from adult patients, lesional skin in pediatric patients with psoriasis did not have increases in regulatory T cells. This is a pilot study, thus the sample size is small. Significant differences in IL-17 and IL-22 expression were observed in the pediatric psoriasis patients compared with pediatric healthy controls and adult psoriasis patients. IL-22 might be relevant in the pathogenesis of pediatric psoriasis and represents a potential treatment target unique to pediatric psoriasis. Copyright © 2017 American Academy of Dermatology, Inc. Published by Elsevier Inc. All rights reserved.

Maximizing Science Return from Future Rodent Experiments on the International Space Station (ISS): Tissue Preservation

NASA Technical Reports Server (NTRS)

Choi, S. Y.; Lai, S.; Klotz, R.; Popova, Y.; Chakravarty, K.; Beegle, J. E.; Wigley, C. L.; Globus, R. K.

2014-01-01

To better understand how mammals adapt to long duration habitation in space, a system for performing rodent experiments on the ISS is under development; Rodent Research-1 is the first flight and will include validation of both on-orbit animal support and tissue preservation. To evaluate plans for on-orbit sample dissection and preservation, we simulated conditions for euthanasia, tissue dissection, and prolonged sample storage on the ISS, and we also developed methods for post-flight dissection and recovery of high quality RNA from multiple tissues following prolonged storage in situ for future science. Mouse livers and spleens were harvested under conditions that simulated nominal, on-orbit euthanasia and dissection operations including storage at -80 C for 4 months. The RNA recovered was of high quality (RNA Integrity Number, RIN(is) greater than 8) and quantity, and the liver enzyme contents and activities (catalase, glutathione reductase, GAPDH) were similar to positive controls, which were collected under standard laboratory conditions. We also assessed the impact of possible delayed on-orbit dissection scenarios (off-nominal) by dissecting and preserving the spleen (RNAlater) and liver (fast-freezing) at various time points post-euthanasia (from 5 min up to 105 min). The RNA recovered was of high quality (spleen, RIN (is) greater than 8; liver, RIN (is) greater than 6) and liver enzyme activities were similar to positive controls at all time points, although an apparent decline in select enzyme activities was evident at the latest time (105 min). Additionally, various tissues were harvested from either intact or partially dissected, frozen carcasses after storage for approximately 2 months; most of the tissues (brain, heart, kidney, eye, adrenal glands and muscle) were of acceptable RNA quality for science return, whereas some tissues (small intestine, bone marrow and bones) were not. These data demonstrate: 1) The protocols developed for future flight experiments will support science return despite delayed preservation post-euthanasia or prolonged storage, and 2) Many additional tissues for gene expression analysis can be obtained by dissection following prolonged storage of the tissue in situ at -80 C. These findings have relevance both to high value, ground-based experiments when sample collection capability is severely constrained, and to all future spaceflight experiments that entail on-orbit sample recovery by the ISS crew.

Investigation of viability of plant tissue in the environmental scanning electron microscopy.

PubMed

Zheng, Tao; Waldron, K W; Donald, Athene M

2009-11-01

The advantages of environmental scanning electron microscopy (ESEM) make it a suitable technique for studying plant tissue in its native state. There have been few studies on the effects of ESEM environment and beam damage on the viability of plant tissue. A simple plant tissue, Allium cepa (onion) upper epidermal tissue was taken as the model for study. The change of moisture content of samples was studied at different relative humidities. Working with the electron beam on, viability tests were conducted for samples after exposure in the ESEM under different operating conditions to investigate the effect of electron beam dose on the viability of samples. The results suggested that without the electron beam, the ESEM chamber itself can prevent the loss of initial moisture if its relative humidity is maintained above 90%. With the electron beam on, the viability of Allium cepa (onion) cells depends both on the beam accelerating voltage and the electron dose/unit area hitting the sample. The dose can be controlled by several of the ESEM instrumental parameters. The detailed process of beam damage on cuticle-down and cuticle-up samples was investigated and compared. The results indicate that cuticular adhesion to the cell wall is relatively weak, but highly resistant to electron beam damage. Systematic study on the effect of ESEM operation parameters has been done. Results qualitatively support the intuitive expectations, but demonstrate quantitatively that Allium cepa epidermal cells are able to be kept in a hydrated and viable state under relevant operation condition inside ESEM, providing a basis for further in situ experiments on plant tissues.

Soft Tissue Response to Titanium Abutments with Different Surface Treatment: Preliminary Histologic Report of a Randomized Controlled Trial.

PubMed

Canullo, Luigi; Dehner, Jan Friedrich; Penarrocha, David; Checchi, Vittorio; Mazzoni, Annalisa; Breschi, Lorenzo

2016-01-01

The aim of this preliminary prospective RCT was to histologically evaluate peri-implant soft tissues around titanium abutments treated using different cleaning methods. Sixteen patients were randomized into three groups: laboratory customized abutments underwent Plasma of Argon treatment (Plasma Group), laboratory customized abutments underwent cleaning by steam (Steam Group), and abutments were used as they came from industry (Control Group). Seven days after the second surgery, soft tissues around abutments were harvested. Samples were histologically analyzed. Soft tissues surrounding Plasma Group abutments predominantly showed diffuse chronic infiltrate, almost no acute infiltrate, with presence of few polymorphonuclear neutrophil granulocytes, and a diffuse presence of collagenization bands. Similarly, in Steam Group, the histological analysis showed a high variability of inflammatory expression factors. Tissues harvested from Control Group showed presence of few neutrophil granulocytes, moderate presence of lymphocytes, and diffuse collagenization bands in some sections, while they showed absence of acute infiltrate in 40% of sections. However, no statistical difference was found among the tested groups for each parameter (p > 0.05). Within the limit of the present study, results showed no statistically significant difference concerning inflammation and healing tendency between test and control groups.

Fetal Tissue Procurement for Karyotype Analysis: Clinician or Pathologist - Which is Better?

PubMed

Conant, Joanna L; Tang, Mary E; Waters, Brenda L

2016-01-01

Chromosomal abnormalities are detected in up to 13% of stillbirths and over 20% of those with developmental anomalies. These estimates may be low since up to 50% of samples fail to achieve a result due to microbial overgrowth or nonviability. Tissue for cytogenetics can be procured at bedside by the clinician or by the pathologist in the laboratory. With clinical collection, tissue is placed into culture media immediately, increasing chances of growth. However, collection competes for attention with other activities, which may result in microbial overgrowth or selection of maternal rather than fetal tissue. Laboratory procurement occurs in a controlled environment using sterile technique, but delay in collection may decrease viability. Our goal was to determine which collection method yields better results. We reviewed cases from 2007-2013 that had two samples submitted for cytogenetics, one from the clinician and one from the pathologist. Specimen source, delivery, collection, and culture setup times, harvest date, cell growth, microbial overgrowth, maternal contamination and final result were obtained from medical records and cytogenetic culture sheets. There was no difference in growth rate, maternal cell contamination, or reporting time between clinician- and pathologist-procured samples despite delay in collection time for laboratory samples. Clinical samples had more microbial overgrowth. Compared to samples collected at bedside, samples collected in the laboratory had a lower rate of microbial contamination with similar growth and maternal cell contamination rates, despite prolonged time to collection. Collecting samples both at bedside and in the laboratory is unnecessary.

Liquid microjunction surface sampling coupled with high-pressure liquid chromatography-electrospray ionization-mass spectrometry for analysis of drugs and metabolites in whole-body thin tissue sections.

PubMed

Kertesz, Vilmos; Van Berkel, Gary J

2010-07-15

In this work, a commercially available autosampler was adapted to perform direct liquid microjunction (LMJ) surface sampling followed by a high-pressure liquid chromatography (HPLC) separation of the extract components and detection with electrospray ionization mass spectrometry (ESI-MS). To illustrate the utility of coupling a separation with this direct liquid extraction based surface sampling approach, four different organs (brain, lung, kidney, and liver) from whole-body thin tissue sections of propranolol dosed and control mice were examined. The parent drug was observed in the chromatograms of the surface sampling extracts from all the organs of the dosed mouse examined. In addition, two isomeric phase II metabolites of propranolol (an aliphatic and an aromatic hydroxypropranolol glucuronide) were observed in the chromatograms of the extracts from lung, kidney, and liver. Confirming the presence of one or the other or both of these glucuronides in the extract from the various organs was not possible without the separation. These drug and metabolite data obtained using the LMJ surface sampling/HPLC-MS method and the results achieved by analyzing similar samples by conventional extraction of the tissues and subsequent HPLC-MS analysis were consistent. The ability to directly and efficiently sample from thin tissue sections via a liquid extraction and then perform a subsequent liquid phase separation increases the utility of this liquid extraction surface sampling approach.

Quality Control of RNA Preservation and Extraction from Paraffin-Embedded Tissue: Implications for RT-PCR and Microarray Analysis

PubMed Central

Pichler, Martin; Zatloukal, Kurt

2013-01-01

Analysis of RNA isolated from fixed and paraffin-embedded tissues is widely used in biomedical research and molecular pathological diagnostics. We have performed a comprehensive and systematic investigation of the impact of factors in the pre-analytical workflow, such as different fixatives, fixation time, RNA extraction method and storage of tissues in paraffin blocks, on several downstream reactions including complementary DNA (cDNA) synthesis, quantitative reverse transcription polymerase chain reaction (qRT-PCR) and microarray hybridization. We compared the effects of routine formalin fixation with the non-crosslinking, alcohol-based Tissue Tek Xpress Molecular Fixative (TTXMF, Sakura Finetek), and cryopreservation as gold standard for molecular analyses. Formalin fixation introduced major changes into microarray gene expression data and led to marked gene-to-gene variations in delta-ct values of qRT-PCR. We found that qRT-PCR efficiency and gene-to-gene variations were mainly attributed to differences in the efficiency of cDNA synthesis as the most sensitive step. These differences could not be reliably detected by quality assessment of total RNA isolated from formalin-fixed tissues by electrophoresis or spectrophotometry. Although RNA from TTXMF fixed samples was as fragmented as RNA from formalin fixed samples, much higher cDNA yield and lower ct-values were obtained in qRT-PCR underlining the negative impact of crosslinking by formalin. In order to better estimate the impact of pre-analytical procedures such as fixation on the reliability of downstream analysis, we applied a qRT-PCR-based assay using amplicons of different length and an assay measuring the efficiency of cDNA generation. Together these two assays allowed better quality assessment of RNA extracted from fixed and paraffin-embedded tissues and should be used to supplement quality scores derived from automated electrophoresis. A better standardization of the pre-analytical workflow, application of additional quality controls and detailed sample information would markedly improve the comparability and reliability of molecular studies based on formalin-fixed and paraffin-embedded tissue samples. PMID:23936242

Tissue-aware RNA-Seq processing and normalization for heterogeneous and sparse data.

PubMed

Paulson, Joseph N; Chen, Cho-Yi; Lopes-Ramos, Camila M; Kuijjer, Marieke L; Platig, John; Sonawane, Abhijeet R; Fagny, Maud; Glass, Kimberly; Quackenbush, John

2017-10-03

Although ultrahigh-throughput RNA-Sequencing has become the dominant technology for genome-wide transcriptional profiling, the vast majority of RNA-Seq studies typically profile only tens of samples, and most analytical pipelines are optimized for these smaller studies. However, projects are generating ever-larger data sets comprising RNA-Seq data from hundreds or thousands of samples, often collected at multiple centers and from diverse tissues. These complex data sets present significant analytical challenges due to batch and tissue effects, but provide the opportunity to revisit the assumptions and methods that we use to preprocess, normalize, and filter RNA-Seq data - critical first steps for any subsequent analysis. We find that analysis of large RNA-Seq data sets requires both careful quality control and the need to account for sparsity due to the heterogeneity intrinsic in multi-group studies. We developed Yet Another RNA Normalization software pipeline (YARN), that includes quality control and preprocessing, gene filtering, and normalization steps designed to facilitate downstream analysis of large, heterogeneous RNA-Seq data sets and we demonstrate its use with data from the Genotype-Tissue Expression (GTEx) project. An R package instantiating YARN is available at http://bioconductor.org/packages/yarn .

Anti-inflammatory and anti-oxidative effects of alpha-lipoic acid in experimentally induced acute otitis media.

PubMed

Tatar, A; Korkmaz, M; Yayla, M; Gozeler, M S; Mutlu, V; Halici, Z; Uslu, H; Korkmaz, H; Selli, J

2016-07-01

To investigate the anti-inflammatory, anti-oxidative and tissue protective effects, as well as the potential therapeutic role, of alpha-lipoic acid in experimentally induced acute otitis media. Twenty-five guinea pigs were assigned to one of five groups: a control (non-otitis) group, and otitis-induced groups treated with saline, penicillin G, alpha-lipoic acid, or alpha-lipoic acid plus penicillin G. Tissue samples were histologically analysed, and oxidative parameters in tissue samples were measured and compared between groups. The epithelial integrity was better preserved, and histological signs of inflammation and secretory metaplasia were decreased, in all groups compared to the saline treated otitis group. In the alpha-lipoic acid plus penicillin G treated otitis group, epithelial integrity was well preserved and histological findings of inflammation were significantly decreased compared to the saline, penicillin G and alpha-lipoic acid treated otitis groups. The most favourable oxidative parameters were observed in the control group, followed by the alpha-lipoic acid plus penicillin G treated otitis group. Alpha-lipoic acid, with its antioxidant, anti-inflammatory and tissue protective properties, may decrease the clinical sequelae and morbidity associated with acute otitis media.

Sensing in tissue bioreactors

NASA Astrophysics Data System (ADS)

Rolfe, P.

2006-03-01

Specialized sensing and measurement instruments are under development to aid the controlled culture of cells in bioreactors for the fabrication of biological tissues. Precisely defined physical and chemical conditions are needed for the correct culture of the many cell-tissue types now being studied, including chondrocytes (cartilage), vascular endothelial cells and smooth muscle cells (blood vessels), fibroblasts, hepatocytes (liver) and receptor neurones. Cell and tissue culture processes are dynamic and therefore, optimal control requires monitoring of the key process variables. Chemical and physical sensing is approached in this paper with the aim of enabling automatic optimal control, based on classical cell growth models, to be achieved. Non-invasive sensing is performed via the bioreactor wall, invasive sensing with probes placed inside the cell culture chamber and indirect monitoring using analysis within a shunt or a sampling chamber. Electroanalytical and photonics-based systems are described. Chemical sensing for gases, ions, metabolites, certain hormones and proteins, is under development. Spectroscopic analysis of the culture medium is used for measurement of glucose and for proteins that are markers of cell biosynthetic behaviour. Optical interrogation of cells and tissues is also investigated for structural analysis based on scatter.

Speckle-type POZ (pox virus and zinc finger protein) protein gene deletion in ovarian cancer: Fluorescence in situ hybridization analysis of a tissue microarray.

PubMed

Hu, Xiaoyu; Yang, Zhu; Zeng, Manman; Liu, Y I; Yang, Xiaotao; Li, Yanan; Li, X U; Yu, Qiubo

2016-07-01

The aim of the present study was to investigate the status of speckle-type POZ (pox virus and zinc finger protein) protein (SPOP) gene located on chromosome 17q21 in ovarian cancer (OC). The present study evaluated a tissue microarray, which contained 90 samples of ovarian cancer and 10 samples of normal ovarian tissue, using fluorescence in situ hybridization (FISH). FISH is a method where a SPOP-specific DNA red fluorescence probe was used for the experimental group and a centromere-specific DNA green fluorescence probe for chromosome 17 was used for the control group. The present study demonstrated that a deletion of the SPOP gene was observed in 52.27% (46/88) of the ovarian cancer tissues, but was not identified in normal ovarian tissues. Simultaneously, monosomy 17 was frequently identified in the ovarian cancer tissues, but not in the normal ovarian tissues. Furthermore, the present data revealed that the ovarian cancer histological subtype and grade were significantly associated with a deletion of the SPOP gene, which was assessed by the appearance of monosomy 17 in the ovarian cancer samples; the deletion of the SPOP gene was observed in a large proportion of serous epithelial ovarian cancer (41/61; 67.21%), particularly in grade 3 (31/37; 83.78%). In conclusion, deletion of the SPOP gene on chromosome 17 in ovarian cancer samples, which results from monosomy 17, indicates that the SPOP gene may serve as a tumor suppressor gene in ovarian cancer.

Rapid detection of Mannheimia haemolytica in lung tissues of sheep and from bacterial culture.

PubMed

Kumar, Jyoti; Dixit, Shivendra Kumar; Kumar, Rajiv

2015-09-01

This study was aimed to detect Mannheimia haemolytica in lung tissues of sheep and from a bacterial culture. M. haemolytica is one of the most important and well-established etiological agents of pneumonia in sheep and other ruminants throughout the world. Accurate diagnosis of M. haemolytica primarily relies on bacteriological examination, biochemical characteristics and, biotyping and serotyping of the isolates. In an effort to facilitate rapid M. haemolytica detection, polymerase chain reaction assay targeting Pasteurella haemolytica serotype-1 specific antigens (PHSSA), Rpt2 and 12S ribosomal RNA (rRNA) genes were used to detect M. haemolytica directly from lung tissues and from bacterial culture. A total of 12 archived lung tissues from sheep that died of pneumonia on an organized farm were used. A multiplex polymerase chain reaction (mPCR) based on two-amplicons targeted PHSSA and Rpt2 genes of M. haemolytica were used for identification of M. haemolytica isolates in culture from the lung samples. All the 12 lung tissue samples were tested for the presence M. haemolytica by PHSSA and Rpt2 genes based PCR and its confirmation by sequencing of the amplicons. All the 12 lung tissue samples tested for the presence of PHSSA and Rpt2 genes of M. haemolytica by mPCR were found to be positive. Amplification of 12S rRNA gene fragment as internal amplification control was obtained with each mPCR reaction performed from DNA extracted directly from lung tissue samples. All the M. haemolytica were also positive for mPCR. No amplified DNA bands were observed for negative control reactions. All the three nucleotide sequences were deposited in NCBI GenBank (Accession No. KJ534629, KJ534630 and KJ534631). Sequencing of the amplified products revealed the identity of 99-100%, with published sequence of PHSSA and Rpt2 genes of M. haemolytica available in the NCBI database. Sheep specific mitochondrial 12S rRNA gene sequence also revealed the identity of 98% with published sequences in the NCBI database. The present study emphasized the PCR as a valuable tool for rapid detection of M. haemolytica in clinical samples from animals. In addition, it offers the opportunity to perform large-scale epidemiological studies regarding the role of M. haemolytica in clinical cases of pneumonia and other disease manifestations in sheep and other ruminants, thereby providing the basis for effective preventive strategies.

Diagnosis and follow-up of Whipple's disease by amplification of the 16S rRNA gene of Tropheryma whippelii.

PubMed

Pron, B; Poyart, C; Abachin, E; Fest, T; Belanger, C; Bonnet, C; Capelle, P; Bretagne, J F; Fabianek, A; Girard, L; Hagège, H; Berche, P

1999-01-01

Amplification of the 16S rRNA gene of Tropheryma whippelii was performed in eight patients with Whipple's disease and 34 control patients to confirm a diagnosis of Whipple's disease and to monitor the course of disease. Polymerase chain reaction (PCR) tests were positive before treatment in 13 of 15 tissue samples from Whipple's disease patients (gut 8/8; lymph nodes 2/2; bone marrow 1/2; peripheral blood 2/3), in contrast to none of 54 tissue samples from controls. PCR tests converted to negative within 4-6 months in six of the Whipple's disease patients undergoing therapy. These results show that PCR is a reliable and useful tool for diagnosis of Whipple's disease and for monitoring bacterial elimination during antibiotic therapy.

Association between infection of virulence cagA gene Helicobacter pylori and laryngeal squamous cell carcinoma

PubMed Central

Burduk, Paweł Krzysztof

2013-01-01

Background The aim of the study was to evaluate the presence of cagA gene Helicobacter pylori in etiopathogenesis of initiation and development of larynx squamous cell carcinoma (LSCC) and its predictable role as a prognostic factor. Material/Methods The prospective, controlled study involved a series of 75 patients (65 male, 10 female, mean age 59.1 years, range 43 to 79 years) with larynx cancer. Samples of larynx cancerous tissue, each of 10–15 mg, were obtained from fresh tissues and were used for nucleic acid purification. DNA was extracted from 225 samples (larynx tumor – I (75), margin of tumor and normal tissue – II (75) and normal larynx tissue from opposite side to the tumor – III). All samples were subjected to H. pylori ureA detection by the PCR H. pylori diagnostic test. Samples that were positive for ureA H. pylori gene were evaluated for cagA H. pylori gene. Results Presence of H. pylori cagA gene was identified in 46,7% to 49,3% of 75 H. pylori ureA gene-positive larynx cancer depending of tissue location. There was a correlation of high incidence of positive cagA gene in larynx cancer tissue in supraglottic versus subglottic and glottic location. We observed a predominance of cagA gene in LSCC in patients with positive cervical lymph nodes and clinical stage T3 and T4. Conclusions H. pylori is present in larynx tissue and may be a possible carcinogen or co-carcinogen in LSCC development, but that must be addressed by future investigations. The presence of cagA gene in larynx cancer tissues significantly decreases survival rate and increases the disease recurrence possibilities. PMID:23860397

Association between infection of virulence cagA gene Helicobacter pylori and laryngeal squamous cell carcinoma.

PubMed

Burduk, Paweł Krzysztof

2013-07-17

The aim of the study was to evaluate the presence of cagA gene Helicobacter pylori in etiopathogenesis of initiation and development of larynx squamous cell carcinoma (LSCC) and its predictable role as a prognostic factor. The prospective, controlled study involved a series of 75 patients (65 male, 10 female, mean age 59.1 years, range 43 to 79 years) with larynx cancer. Samples of larynx cancerous tissue, each of 10-15 mg, were obtained from fresh tissues and were used for nucleic acid purification. DNA was extracted from 225 samples (larynx tumor - I (75), margin of tumor and normal tissue - II (75) and normal larynx tissue from opposite side to the tumor - III). All samples were subjected to H. pylori ureA detection by the PCR H. pylori diagnostic test. Samples that were positive for ureA H. pylori gene were evaluated for cagA H. pylori gene. Presence of H. pylori cagA gene was identified in 46,7% to 49,3% of 75 H. pylori ureA gene-positive larynx cancer depending of tissue location. There was a correlation of high incidence of positive cagA gene in larynx cancer tissue in supraglottic versus subglottic and glottic location. We observed a predominance of cagA gene in LSCC in patients with positive cervical lymph nodes and clinical stage T3 and T4. H. pylori is present in larynx tissue and may be a possible carcinogen or co-carcinogen in LSCC development, but that must be addressed by future investigations. The presence of cagA gene in larynx cancer tissues significantly decreases survival rate and increases the disease recurrence possibilities.

System and method for controlling depth of imaging in tissues using fluorescence microscopy under ultraviolet excitation following staining with fluorescing agents

DOE Office of Scientific and Technical Information (OSTI.GOV)

Levenson, Richard; Demos, Stavros

A method is disclosed for analyzing a thin tissue sample and adapted to be supported on a slide. The tissue sample may be placed on a slide and exposed to one or more different exogenous fluorophores excitable in a range of about 300 nm-200 nm, and having a useful emission band from about 350 nm-900 nm, and including one or more fluorescent dyes or fluorescently labeled molecular probes that accumulate in tissue or cellular components. The fluorophores may be excited with a first wavelength of UV light between about 200 nm-290 nm. An optical system collects emissions from the fluorophoresmore » at a second wavelength, different from the first wavelength, which are generated in response to the first wavelength of UV light, to produce an image for analysis.« less

Cell and Tissue Microarray Technologies for Protein and Nucleic Acid Expression Profiling

PubMed Central

Cardano, Marina; Diaferia, Giuseppe R.; Falavigna, Maurizio; Spinelli, Chiara C.; Sessa, Fausto; DeBlasio, Pasquale

2013-01-01

Tissue microarray (TMA) and cell microarray (CMA) are two powerful techniques that allow for the immunophenotypical characterization of hundreds of samples simultaneously. In particular, the CMA approach is particularly useful for immunophenotyping new stem cell lines (e.g., cardiac, neural, mesenchymal) using conventional markers, as well as for testing the specificity and the efficacy of newly developed antibodies. We propose the use of a tissue arrayer not only to perform protein expression profiling by immunohistochemistry but also to carry out molecular genetics studies. In fact, starting with several tissues or cell lines, it is possible to obtain the complete signature of each sample, describing the protein, mRNA and microRNA expression, and DNA mutations, or eventually to analyze the epigenetic processes that control protein regulation. Here we show the results obtained using the Galileo CK4500 TMA platform. PMID:23172795

Upregulation of Interleukin 21 and Interleukin 21 Receptor in Patients with Dermatomyositis and Polymyositis.

PubMed

Liu, Tao; Hou, Ying; Dai, Ting-Jun; Yan, Chuan-Zhu

2017-09-05

The immunopathologic mechanism underlying dermatomyositis (DM) and polymyositis (PM) remains poorly understood. Many cytokines play a pathogenic role in DM and PM. Interleukin 21 (IL-21) has a pleiotropic effect on inflammation regulation. This study aimed to detect the serum IL-21 level and investigate the expression of IL-21 and IL-21 receptor (IL-21R) in muscle tissues of patients with DM and PM. Biopsied muscle samples were obtained from 11 patients with DM, 12 with PM, and six controls; mRNA levels of IL-21 and IL-21R were analyzed by real-time quantitative reverse transcription-polymerase chain reaction; and immunohistochemical staining was used to evaluate the protein expression of IL-21 and IL-21R. Serum samples were obtained from 36 patients with DM, 19 with PM, and 20 healthy controls. The serum IL-21 level was detected by enzyme-linked immunosorbent assay. The expression of IL-21 was upregulated in patients with DM and PM. The IL-21 mRNA level was significantly increased in muscle tissues of patients with DM and PM (DM vs. control, P= 0.001; PM vs. control, P= 0.001), whereas IL-21R mRNA level in patients with DM/PM was not statistically different from that of healthy controls. Immunohistochemical staining showed both IL-21 and IL-21R were significantly expressed in the inflammatory cells in muscle tissues of patients with DM and PM. The serum IL-21 level was also significantly higher in patients with DM/PM than in controls (DM vs. control, 49.12 [45.28, 60.07] pg/ml vs. 42.54 [38.69, 48.85] pg/ml, P= 0.001; PM vs. control, 50.77 [44.19, 60.62] pg/ml vs. 42.54 [38.69, 48.85] pg/ml, P= 0.005). IL-21 expression is upregulated in patients with DM and PM in both muscle tissue and serum. In addition, IL-21R protein is highly expressed in affected muscle tissues of patients with DM and PM. IL-21 may play a pathogenic role through IL-21R in patients with DM and PM.

Detection frequency of human herpesviruses-6A, -6B, and -7 genomic sequences in central nervous system DNA samples from post-mortem individuals with unspecified encephalopathy.

PubMed

Chapenko, Svetlana; Roga, Silvija; Skuja, Sandra; Rasa, Santa; Cistjakovs, Maksims; Svirskis, Simons; Zaserska, Zane; Groma, Valerija; Murovska, Modra

2016-08-01

In this autopsy-based study, human herpesvirus-6 (HHV-6) and -7 (HHV-7) genomic sequence frequency, HHV-6 variants, HHV-6 load and the expression of HHV-6 antigens in brain samples from the individuals, with and without unspecified encephalopathy (controls), using nested and real-time polymerase chain reactions, restriction endonuclease, and immunohistochemical analysis were examined. GraphPad Prism 6.0 Mann-Whitney nonparametric and chi-square test and Fisher's exact test were used for statistical analysis. The encephalopathy diagnoses were shown by magnetic resonance imaging made during their lifetime and macro- and microscopically studied autopsy tissue materials. Widespread HHV-6 and/or HHV-7 positivity was detected in the brain tissue of various individuals with encephalopathy, as well as in controls (51/57, 89.4 % and 35/51, 68.6 %, respectively; p = 0.009). Significantly higher detection frequency of single HHV-6 and concurrent HHV-6 + HHV-7 DNA was found in pia mater meninges, frontal lobe, temporal lobe, and olfactory tract DNAs in individuals with encephalopathy compared to the control group. HHV-6 load and higher frequency of the viral load >10 copies/10(6) cells significantly differed in samples from individuals with and without encephalopathy. The expression of HHV-6 antigens was revealed in different neural cell types with strong predominance in the encephalopathy group. In all HHV-6-positive autopsy samples of individuals with and without encephalopathy, HHV-6B was revealed. Significantly higher detection frequency of beta-herpesvirus DNA, more often detected HHV-6 load >10 copies/10(6) cells, as well as the expression of HHV-6 antigens in different brain tissue samples from individuals with encephalopathy in comparison with control group indicate on potential involvement of these viruses in encephalopathy development.

Excretory-secretory antigens: a suitable candidate for immunization against ocular toxoplasmosis in a murine model.

PubMed

Norouzpour Deilami, Kiumars; Daryani, Ahmad; Ahmadpour, Ehsan; Sharif, Mehdi; Dadimoghaddam, Yousef; Sarvi, Shahabeddin; Alizadeh, Ahad

2014-12-01

Toxoplasmosis, responsible for ocular impairment, is caused by Toxoplasma gondii. We investigated the effect of Toxoplasma excretory-secretory antigens (ESA) on parasite load and distribution in the eye tissue of a murine model. Case and control groups were immunized with ESA and PBS, respectively. Two weeks after the second immunization, the mice were challenged intraperitoneally with virulent RH strain of Toxoplasma; eye tissue samples of both groups were collected daily (days 1, 2, 3, and the last day before death). Parasite load was determined using real-time quantitative PCR targeted at the B1 gene. Compared to the control group, infected mice that received ESA vaccine presented a considerable decrease in parasite load in the eye tissue, demonstrating the effect of ESA on parasite load and distribution. Diminution of parasite load in mouse eye tissue indicated that ESA might help control disease-related complications and could be a valuable immunization candidate against ocular toxoplasmosis. Copyright © 2014 Elsevier Ltd. All rights reserved.

A high-resolution optical imaging system for obtaining the serial transverse section images of biologic tissue

NASA Astrophysics Data System (ADS)

Wu, Li; Zhang, Bin; Wu, Ping; Liu, Qian; Gong, Hui

2007-05-01

A high-resolution optical imaging system was designed and developed to obtain the serial transverse section images of the biologic tissue, such as the mouse brain, in which new knife-edge imaging technology, high-speed and high-sensitive line-scan CCD and linear air bearing stages were adopted and incorporated with an OLYMPUS microscope. The section images on the tip of the knife-edge were synchronously captured by the reflection imaging in the microscope while cutting the biologic tissue. The biologic tissue can be sectioned at interval of 250 nm with the same resolution of the transverse section images obtained in x and y plane. And the cutting job can be automatically finished based on the control program wrote specially in advance, so we save the mass labor of the registration of the vast images data. In addition, by using this system a larger sample can be cut than conventional ultramicrotome so as to avoid the loss of the tissue structure information because of splitting the tissue sample to meet the size request of the ultramicrotome.

Effects of vitrification cryopreservation on follicular morphology and stress relaxation behaviors of human ovarian tissues: sucrose versus trehalose as the non-permeable protective agent.

PubMed

Tian, Ting; Zhao, Gang; Han, Dan; Zhu, Kaixuan; Chen, Dawei; Zhang, Zhiguo; Wei, Zhaolian; Cao, Yunxia; Zhou, Ping

2015-04-01

Is sucrose more effective than trehalose in human ovarian tissue cryopreservation? The effect of sucrose as a cryoprotective agent (CPA) was not significantly different from that of trehalose in human ovarian tissue cryopreservation. Sugars have the ability to keep the cell membrane intact and can decrease the toxicity of CPAs. Sucrose is the most commonly used non-permeable CPA, while trehalose is rarely used in human ovarian tissue cryopreservation. Although various methods are utilized to evaluate the efficiency of human ovarian tissue cryopreservation, few studies have evaluated the effect of cryopreservation from the viewpoint of biomechanics. A total of 15 ovarian tissue samples were collected from 15 patients (20-41 years old) with benign ovarian tumors or malignancies, and each was dissected into six slices. Two slices were taken as the fresh control group. The remaining four slices were vitrified using different vitrification protocols. After warming, samples in each group were either fixed for histological evaluation or destined for stress relaxation test. The CPA solutions for the control and vitrified groups were composed of EDS and EDT (E, ethylene glycol; D, dimethylsulphoxide; S, sucrose; T, trehalose). The stress relaxation experiments were carried out at room temperature using a dynamic mechanical analyzer. Ovarian tissue samples were assessed for both their morphology and viscoelasticity. Stress relaxation data (SRD) were calculated as a percentage, representing the ability to maintain the initial stress after stretching. The percentage of morphologically normal follicles was compared between groups, which was represented by morphologic preservation ratio. The morphologic preservation ratio of the primordial follicles in the fresh control group (87.58%) was higher than that in group S (72.33%) (P = 0.000) and group T (79.56%) (P = 0.002). Although not statistically significant, compared with the S group, vitrification with T suggested a trend toward a higher morphologic preservation ratio of the primordial follicles. The SRD in the fresh control group (0.6433 ± 0.7233) was significantly different from that in group S (0.5200 ± 0.8331, P = 0.000) or in group T (0.5667 ± 0.6415, P = 0.000). However, no significant difference was found between groups S and T. Experimental samples were directly exposed to the air, which will result in a discrepancy in the viscoelastic properties between experimental tissues and in vivo tissues. Our study suggested a trend toward a higher morphologic preservation ratio of the primordial follicles after vitrification in trehalose compared with sucrose, which may provide a basis for further optimizing human ovarian tissue vitrification. In addition, it was possible to evaluate the effect of ovarian tissue cryopreservation from a biomechanics perspective. This study was supported by the grants from the Medical Scientific Research Subject, Health Ministry of Anhui Province (2010B014) and National Basic Research Program of China (973 Program) (2012CB944704), and the National Natural Science Foundation of China (Nos. 51276179 and 51476160). The authors declare that there is no conflict of interests regarding the publication of this original paper. © The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

A SELDI mass spectrometry study of experimental autoimmune encephalomyelitis: sample preparation, reproducibility, and differential protein expression patterns.

PubMed

Azzam, Sausan; Broadwater, Laurie; Li, Shuo; Freeman, Ernest J; McDonough, Jennifer; Gregory, Roger B

2013-05-01

Experimental autoimmune encephalomyelitis (EAE) is an autoimmune, inflammatory disease of the central nervous system that is widely used as a model of multiple sclerosis (MS). Mitochondrial dysfunction appears to play a role in the development of neuropathology in MS and may also play a role in disease pathology in EAE. Here, surface enhanced laser desorption ionization mass spectrometry (SELDI-MS) has been employed to obtain protein expression profiles from mitochondrially enriched fractions derived from EAE and control mouse brain. To gain insight into experimental variation, the reproducibility of sub-cellular fractionation, anion exchange fractionation as well as spot-to-spot and chip-to-chip variation using pooled samples from brain tissue was examined. Variability of SELDI mass spectral peak intensities indicates a coefficient of variation (CV) of 15.6% and 17.6% between spots on a given chip and between different chips, respectively. Thinly slicing tissue prior to homogenization with a rotor homogenizer showed better reproducibility (CV = 17.0%) than homogenization of blocks of brain tissue with a Teflon® pestle (CV = 27.0%). Fractionation of proteins with anion exchange beads prior to SELDI-MS analysis gave overall CV values from 16.1% to 18.6%. SELDI mass spectra of mitochondrial fractions obtained from brain tissue from EAE mice and controls displayed 39 differentially expressed proteins (p≤ 0.05) out of a total of 241 protein peaks observed in anion exchange fractions. Hierarchical clustering analysis showed that protein fractions from EAE animals with severe disability clearly segregated from controls. Several components of electron transport chain complexes (cytochrome c oxidase subunit 6b1, subunit 6C, and subunit 4; NADH dehydrogenase flavoprotein 3, alpha subcomplex subunit 2, Fe-S protein 4, and Fe-S protein 6; and ATP synthase subunit e) were identified as possible differentially expressed proteins. Myelin Basic Protein isoform 8 (MBP8) (14.2 kDa) levels were lower in EAE samples with advanced disease relative to controls, while an MBP fragment (12. 4kDa), likely due to calpain digestion, was increased in EAE relative to controls. The appearance of MBP in mitochondrially enriched fractions is due to tissue freezing and storage, as MBP was not found associated with mitochondria obtained from fresh tissue. SELDI mass spectrometry can be employed to explore the proteome of a complex tissue (brain) and obtain protein profiles of differentially expressed proteins from protein fractions. Appropriate homogenization protocols and protein fractionation using anion exchange beads can be employed to reduce sample complexity without introducing significant additional variation into the SELDI mass spectra beyond that inherent in the SELDI- MS method itself. SELDI-MS coupled with principal component analysis and hierarchical cluster analysis provides protein patterns that can clearly distinguish the disease state from controls. However, identification of individual differentially expressed proteins requires a separate purification of the proteins of interest by polyacrylamide electrophoresis prior to trypsin digestion and peptide mass fingerprint analysis, and unambiguous identification of differentially expressed proteins can be difficult if protein bands consist of several proteins with similar molecular weights.

A SELDI mass spectrometry study of experimental autoimmune encephalomyelitis: sample preparation, reproducibility, and differential protein expression patterns

PubMed Central

2013-01-01

Background Experimental autoimmune encephalomyelitis (EAE) is an autoimmune, inflammatory disease of the central nervous system that is widely used as a model of multiple sclerosis (MS). Mitochondrial dysfunction appears to play a role in the development of neuropathology in MS and may also play a role in disease pathology in EAE. Here, surface enhanced laser desorption ionization mass spectrometry (SELDI-MS) has been employed to obtain protein expression profiles from mitochondrially enriched fractions derived from EAE and control mouse brain. To gain insight into experimental variation, the reproducibility of sub-cellular fractionation, anion exchange fractionation as well as spot-to-spot and chip-to-chip variation using pooled samples from brain tissue was examined. Results Variability of SELDI mass spectral peak intensities indicates a coefficient of variation (CV) of 15.6% and 17.6% between spots on a given chip and between different chips, respectively. Thinly slicing tissue prior to homogenization with a rotor homogenizer showed better reproducibility (CV = 17.0%) than homogenization of blocks of brain tissue with a Teflon® pestle (CV = 27.0%). Fractionation of proteins with anion exchange beads prior to SELDI-MS analysis gave overall CV values from 16.1% to 18.6%. SELDI mass spectra of mitochondrial fractions obtained from brain tissue from EAE mice and controls displayed 39 differentially expressed proteins (p≤ 0.05) out of a total of 241 protein peaks observed in anion exchange fractions. Hierarchical clustering analysis showed that protein fractions from EAE animals with severe disability clearly segregated from controls. Several components of electron transport chain complexes (cytochrome c oxidase subunit 6b1, subunit 6C, and subunit 4; NADH dehydrogenase flavoprotein 3, alpha subcomplex subunit 2, Fe-S protein 4, and Fe-S protein 6; and ATP synthase subunit e) were identified as possible differentially expressed proteins. Myelin Basic Protein isoform 8 (MBP8) (14.2 kDa) levels were lower in EAE samples with advanced disease relative to controls, while an MBP fragment (12. 4kDa), likely due to calpain digestion, was increased in EAE relative to controls. The appearance of MBP in mitochondrially enriched fractions is due to tissue freezing and storage, as MBP was not found associated with mitochondria obtained from fresh tissue. Conclusions SELDI mass spectrometry can be employed to explore the proteome of a complex tissue (brain) and obtain protein profiles of differentially expressed proteins from protein fractions. Appropriate homogenization protocols and protein fractionation using anion exchange beads can be employed to reduce sample complexity without introducing significant additional variation into the SELDI mass spectra beyond that inherent in the SELDI- MS method itself. SELDI-MS coupled with principal component analysis and hierarchical cluster analysis provides protein patterns that can clearly distinguish the disease state from controls. However, identification of individual differentially expressed proteins requires a separate purification of the proteins of interest by polyacrylamide electrophoresis prior to trypsin digestion and peptide mass fingerprint analysis, and unambiguous identification of differentially expressed proteins can be difficult if protein bands consist of several proteins with similar molecular weights. PMID:23635033

Telomerase activity as a marker for malignancy in feline tissues.

PubMed

Cadile, C D; Kitchell, B E; Biller, B J; Hetler, E R; Balkin, R G

2001-10-01

To establish the diagnostic significance of the telomeric repeat amplification protocol (TRAP) assay in detecting feline malignancies. Solid tissue specimens collected from 33 client-owned cats undergoing diagnostic or therapeutic procedures at the University of Illinois Veterinary Medical Teaching Hospital between July 1997 and September 1999 and an additional 20 tissue samples were collected from 3 clinically normal control cats euthanatized at the conclusion of an unrelated study. The TRAP assay was used for detection of telomerase activity. Each result was compared to its respective histopathologic diagnosis. Twenty-nine of 31 malignant and 1 of 22 benign or normal tissue samples had telomerase activity, indicating 94% sensitivity and 95% specificity of the TRAP assay in our laboratory. The diagnostic significance of telomerase activity has been demonstrated in humans and recently in dogs by our laboratory. We tested feline samples to determine whether similar patterns of telomerase activity exist. On the basis of our results, the TRAP assay may be clinically useful in providing a rapid diagnosis of malignancy in cats. The telomerase enzyme may also serve as a therapeutic target in feline tumors.

Detection of pesticides and dioxins in tissue fats and rendering oils using laser-induced breakdown spectroscopy (LIBS).

PubMed

Multari, Rosalie A; Cremers, David A; Scott, Thomas; Kendrick, Peter

2013-03-13

In laser-induced breakdown spectroscopy (LIBS), a series of powerful laser pulses are directed at a surface to form microplasmas from which light is collected and spectrally analyzed to identify the surface material. In most cases, no sample preparation is needed, and results can be automated and made available within seconds to minutes. Advances in LIBS spectral data analysis using multivariate regression techniques have led to the ability to detect organic chemicals in complex matrices such as foods. Here, the use of LIBS to differentiate samples contaminated with aldrin, 1,2,3,4,6,7,8-heptachlorodibenzo-p-dioxin, chlorpyrifos, and dieldrin in the complex matrices of tissue fats and rendering oils is described. The pesticide concentrations in the samples ranged from 0.005 to 0.1 μg/g. All samples were successfully differentiated from each other and from control samples. Sample concentrations could also be differentiated for all of the pesticides and the dioxin included in this study. The results presented here provide first proof-of-principle data for the ability to create LIBS-based instrumentation for the rapid analysis of pesticide and dioxin contamination in tissue fat and rendered oils.

Application of FTA technology for sampling, recovery and molecular characterization of viral pathogens and virus-derived transgenes from plant tissues.

PubMed

Ndunguru, Joseph; Taylor, Nigel J; Yadav, Jitender; Aly, Haytham; Legg, James P; Aveling, Terry; Thompson, Graham; Fauquet, Claude M

2005-05-18

Plant viral diseases present major constraints to crop production. Effective sampling of the viruses infecting plants is required to facilitate their molecular study and is essential for the development of crop protection and improvement programs. Retaining integrity of viral pathogens within sampled plant tissues is often a limiting factor in this process, most especially when sample sizes are large and when operating in developing counties and regions remote from laboratory facilities. FTA is a paper-based system designed to fix and store nucleic acids directly from fresh tissues pressed into the treated paper. We report here the use of FTA as an effective technology for sampling and retrieval of DNA and RNA viruses from plant tissues and their subsequent molecular analysis. DNA and RNA viruses were successfully recovered from leaf tissues of maize, cassava, tomato and tobacco pressed into FTA Classic Cards. Viral nucleic acids eluted from FTA cards were found to be suitable for diagnostic molecular analysis by PCR-based techniques and restriction analysis, and for cloning and nucleotide sequencing in a manner equivalent to that offered by tradition isolation methods. Efficacy of the technology was demonstrated both from sampled greenhouse-grown plants and from leaf presses taken from crop plants growing in farmer's fields in East Africa. In addition, FTA technology was shown to be suitable for recovery of viral-derived transgene sequences integrated into the plant genome. Results demonstrate that FTA is a practical, economical and sensitive method for sampling, storage and retrieval of viral pathogens and plant genomic sequences, when working under controlled conditions and in the field. Application of this technology has the potential to significantly increase ability to bring modern analytical techniques to bear on the viral pathogens infecting crop plants.

Osteoinduction by Ca-P biomaterials implanted into the muscles of mice*

PubMed Central

Yang, Rui-na; Ye, Feng; Cheng, Li-jia; Wang, Jin-jing; Lu, Xiao-feng; Shi, Yu-jun; Fan, Hong-song; Zhang, Xing-dong; Bu, Hong

2011-01-01

The osteoinduction of porous biphasic calcium phosphate ceramics (BCP) has been widely reported and documented, but little research has been performed on rodent animals, e.g., mice. In this study, we report osteoinduction in a mouse model. Thirty mice were divided into two groups. BCP materials (Sample A) and control ceramics (Sample B) were implanted into the leg muscle, respectively. Five mice in each group were killed at 15, 30, and 45 d after surgery. Sample A and Sample B were harvested and used for hematoxylin and eosin (HE) staining, immunohistochemistry (IHC) staining, and Alizarin Red S staining to check bone formation in the biomaterials. Histological analysis showed that no bone tissue was formed 15 d after implantation (0/5) in either of the two groups. Newly-formed bone tissues were observed in Sample A at 30 d (5/5) and 45 d (5/5) after implantation; the average amounts of newly-formed bone tissues were approximately 5.2% and 8.6%, respectively. However, we did not see any bone tissue in Sample B until 45 d after implantation. Bone-related molecular makers such as bone morphogenesis protein-2 (BMP-2), collagen type I, and osteopontin were detected by IHC staining in Sample A 30 d after implantation. In addition, the newly-formed bone was also confirmed by Alizarin Red S staining. Because this is the report of osteoinduction in the rodent animal on which all the biotechnologies were available, our results may contribute to further mechanism research. PMID:21726066

Effect of Ankaferd Blood Stopper on Skin Superoxide Dismutase and Catalase Activities in Warfarin-Treated Rats.

PubMed

Aktop, Sertaç; Emekli-Alturfan, Ebru; Gönül, Onur; Göçmen, Gökhan; Garip, Hasan; Yarat, Ayşen; Göker, Kamil

2017-03-01

Ankaferd Blood Stopper (ABS) is a new promising local hemostatic agent, and its mechanism on hemostasis has been shown by many studies. However, the effects of ABS on skin superoxide dismutase (SOD) and catalase (CAT) activities have not been investigated before. The aim of this study was to evaluate the effects of this new generation local hemostatic agent on warfarin-treated rats focusing on its the antioxidant potential in short-term soft tissue healing. Twelve systemically warfarin treated (warfarin group) and 12 none treated Wistar Albino rats (control group) were selected for the trial. Rats in the warfarin group were treated intraperitonally with 0.1 mg/kg warfarin, and rats in the control group were given 1 mL/kg saline 3 days earlier to surgical procedure and continued until killing. All rats had incisions on dorsal dermal tissue, which was applied ABS or no hemostatic agent before suturing. Six of each group were killed on day 4, and the other 6 were killed on day 8. Blood and skin samples were taken. Prothrombin time (PT) in blood samples, CAT, and SOD activities in skin samples were determined. Warfarin treatment dose was found to be convenient and warfarin treatment increased the PT levels as expected. Warfarin treatment decreased CAT activity significantly compared to the control group. The ABS treatment significantly increased SOD activities in the warfarin group at the end of the eighth day. Ankaferd Blood Stopper acted positively in short-term tissue healing by increasing SOD activity in warfarin-treated rats. Therefore, ABS may be suggeted as a promoting factor in tissue healing.

Curcumin and dexmedetomidine prevents oxidative stress and renal injury in hind limb ischemia/reperfusion injury in a rat model.

PubMed

Karahan, M A; Yalcin, S; Aydogan, H; Büyükfirat, E; Kücük, A; Kocarslan, S; Yüce, H H; Taskın, A; Aksoy, N

2016-06-01

Curcumin and dexmedetomidine have been shown to have protective effects in ischemia-reperfusion injury on various organs. However, their protective effects on kidney tissue against ischemia-reperfusion injury remain unclear. We aimed to determine whether curcumin or dexmedetomidine prevents renal tissue from injury that was induced by hind limb ischemia-reperfusion in rats. Fifty rats were divided into five groups: sham, control, curcumin (CUR) group (200 mg/kg curcumin, n = 10), dexmedetomidine (DEX) group (25 μg/kg dexmedetomidine, n = 10), and curcumin-dexmedetomidine (CUR-DEX) group (200 mg/kg curcumin and 25 μg/kg dexmedetomidine). Curcumin and dexmedetomidine were administered intraperitoneally immediately after the end of 4 h ischemia, just 5 min before reperfusion. The extremity re-perfused for 2 h and then blood samples were taken and total antioxidant capacity (TAC), total oxidative status (TOS) levels, and oxidative stress index (OSI) were measured, and renal tissue samples were histopathologically examined. The TAC activity levels in blood samples were significantly lower in the control than the other groups (p < 0.01 for all comparisons). The TOS activity levels in blood samples were significantly higher in Control group and than the other groups (p <  0.01 for all comparison). The OSI were found to be significantly increased in the control group compared to others groups (p < 0.001 for all comparisons). Histopathological examination revealed less severe lesions in the sham, CUR, DEX, and CUR-DEX groups, compared with the control group (p < 0.01). Rat hind limb ischemia-reperfusion causes histopathological changes in the kidneys. Curcumin and dexmedetomidine administered intraperitoneally was effective in reducing oxidative stress and renal histopathologic injury in an acute hind limb I/R rat model.

Tissue and organ donation for research in forensic pathology: the MRC Sudden Death Brain and Tissue Bank.

PubMed

Millar, T; Walker, R; Arango, J-C; Ironside, J W; Harrison, D J; MacIntyre, D J; Blackwood, D; Smith, C; Bell, J E

2007-12-01

Novel methodological approaches to the investigation of brain and non-central nervous system disorders have led to increased demand for well-characterized, high quality human tissue samples, particularly from control cases. In the setting of the new Human Tissue legislation, we sought to determine whether relatives who have been suddenly bereaved are willing to grant authorization for research use of post mortem tissue samples and organs in sufficient numbers to support the establishment of a brain and tissue bank based in the forensic service. Research authorization was sought from families on the day prior to forensic post mortem examination followed up by written confirmation. We have to date selected individuals who have died suddenly (age range 1-89 years) and who were likely to have normal brains or who had displayed symptoms of a CNS disorder of interest to researchers, including psychiatric disorders. One hundred and eleven families have been approached during the first 2 years of this project. Research use of tissue samples was authorized by 96% of families and 17% agreed to whole brain donation. Audit of families' experience does not suggest that they are further distressed by being approached. Respondents expressed a clear view that the opportunity for research donation should be open to all bereaved families. Despite the sometimes long post mortem intervals, the quality of tissue samples is good, as assessed by a range of markers including Agilent BioAnalyzer quantification of RNA integrity (mean value 6.4). We conclude that the vast majority of families are willing to support research use of post mortem tissues even in the context of sudden bereavement and despite previous adverse publicity. The potential for acquisition of normal CNS and non-CNS tissues and of various hard-to-get CNS disorders suggests that efforts to access the forensic post mortem service for research material are eminently worthwhile. (c) 2007 Pathological Society of Great Britain and Ireland

Optimal molecular profiling of tissue and tissue components: defining the best processing and microdissection methods for biomedical applications.

PubMed

Bova, G Steven; Eltoum, Isam A; Kiernan, John A; Siegal, Gene P; Frost, Andra R; Best, Carolyn J M; Gillespie, John W; Emmert-Buck, Michael R

2005-01-01

Isolation of well-preserved pure cell populations is a prerequisite for sound studies of the molecular basis of pancreatic malignancy and other biological phenomena. This chapter reviews current methods for obtaining anatomically specific signals from molecules isolated from tissues, a basic requirement for productive linking of phenotype and genotype. The quality of samples isolated from tissue and used for molecular analysis is often glossed-over or omitted from publications, making interpretation and replication of data difficult or impossible. Fortunately, recently developed techniques allow life scientists to better document and control the quality of samples used for a given assay, creating a foundation for improvement in this area. Tissue processing for molecular studies usually involves some or all of the following steps: tissue collection, gross dissection/identification, fixation, processing/embedding, storage/archiving, sectioning, staining, microdissection/annotation, and pure analyte labeling/identification. High-quality tissue microdissection does not necessarily mean high-quality samples to analyze. The quality of biomaterials obtained for analysis is highly dependent on steps upstream and downstream from tissue microdissection. We provide protocols for each of these steps, and encourage you to improve upon these. It is worth the effort of every laboratory to optimize and document its technique at each stage of the process, and we provide a starting point for those willing to spend the time to optimize. In our view, poor documentation of tissue and cell type of origin and the use of nonoptimized protocols is a source of inefficiency in current life science research. Even incremental improvement in this area will increase productivity significantly.

Early arthritis induces disturbances at bone nanostructural level reflected in decreased tissue hardness in an animal model of arthritis

PubMed Central

Cascão, Rita; Finnilä, Mikko A. J.; Lopes, Inês P.; Saarakkala, Simo; Zioupos, Peter; Canhão, Helena; Fonseca, João E.

2018-01-01

Introduction Arthritis induces joint erosions and skeletal bone fragility. Objectives The main goal of this work was to analyze the early arthritis induced events at bone architecture and mechanical properties at tissue level. Methods Eighty-eight Wistar rats were randomly housed in experimental groups, as follows: adjuvant induced arthritis (AIA) (N = 47) and a control healthy group (N = 41). Rats were monitored during 22 days for the inflammatory score, ankle perimeter and body weight and sacrificed at different time points (11 and 22 days post disease induction). Bone samples were collected for histology, micro computed tomography (micro-CT), 3-point bending and nanoindentation. Blood samples were also collected for bone turnover markers and systemic cytokine quantification. Results At bone tissue level, measured by nanoindentation, there was a reduction of hardness in the arthritic group, associated with an increase of the ratio of bone concentric to parallel lamellae and of the area of the osteocyte lacuna. In addition, increased bone turnover and changes in the microstructure and mechanical properties were observed in arthritic animals, since the early phase of arthritis, when compared with healthy controls. Conclusion We have shown in an AIA rat model that arthritis induces very early changes at bone turnover, structural degradation and mechanical weakness. Bone tissue level is also affected since the early phase of arthritis, characterized by decreased tissue hardness associated with changes in bone lamella organization and osteocyte lacuna surface. These observations highlight the pertinence of immediate control of inflammation in the initial stages of arthritis. PMID:29315314

Characterization of canine mitochondrial protein expression in natural and induced forms of idiopathic dilated cardiomyopathy.

PubMed

Lopes, Rosana; Solter, Philip F; Sisson, D David; Oyama, Mark A; Prosek, Robert

2006-06-01

To map canine mitochondrial proteins and identify qualitative and quantitative differences in heart mitochondrial protein expression between healthy dogs and dogs with naturally occurring and induced dilated cardiomyopathy (DCM). Left ventricle samples were obtained from 7 healthy dogs, 7 Doberman Pinschers with naturally occurring DCM, and 7 dogs with induced DCM. Fresh and frozen mitochondrial fractions were isolated from the left ventricular free wall and analyzed by 2-dimensional electrophoresis. Protein spots that increased or decreased in density by >or= 2-fold between groups were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry or quadrupole selecting, quadrupole collision cell, time-of-flight mass spectrometry. Within narrow pH gradients of control canine heart mitochondrial samples, a total of 1,528 protein spots were revealed. Forty subunits of heart mitochondrial proteins that differ significantly from control tissues were altered in tissue specimens from dogs with naturally occurring and induced forms of DCM. The most affected heart mitochondrial proteins in both groups were those of oxidative phosphorylation (55%). Upregulation of manganese superoxide dismutase was suggestive of heart oxidative injury in tissue specimens from dogs with both forms of DCM. Evidence of apoptosis was associated with overexpression of the heart mitochondrial voltage-dependent anion channel-2 protein and endonuclease G in tissue specimens from dogs with induced DCM. Alterations of heart mitochondrial proteins related to oxidative phosphorylation dysfunction were more prevalent in tissue specimens from dogs with induced or naturally occurring DCM, compared with those of control dogs.

An Automated Platform for High-Resolution Tissue Imaging Using Nanospray Desorption Electrospray Ionization Mass Spectrometry

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lanekoff, Ingela T.; Heath, Brandi S.; Liyu, Andrey V.

2012-10-02

An automated platform has been developed for acquisition and visualization of mass spectrometry imaging (MSI) data using nanospray desorption electrospray ionization (nano-DESI). The new system enables robust operation of the nano-DESI imaging source over many hours. This is achieved by controlling the distance between the sample and the probe by mounting the sample holder onto an automated XYZ stage and defining the tilt of the sample plane. This approach is useful for imaging of relatively flat samples such as thin tissue sections. Custom software called MSI QuickView was developed for visualization of large data sets generated in imaging experiments. MSImore » QuickView enables fast visualization of the imaging data during data acquisition and detailed processing after the entire image is acquired. The performance of the system is demonstrated by imaging rat brain tissue sections. High resolution mass analysis combined with MS/MS experiments enabled identification of lipids and metabolites in the tissue section. In addition, high dynamic range and sensitivity of the technique allowed us to generate ion images of low-abundance isobaric lipids. High-spatial resolution image acquired over a small region of the tissue section revealed the spatial distribution of an abundant brain metabolite, creatine, in the white and gray matter that is consistent with the literature data obtained using magnetic resonance spectroscopy.« less

Analysis of human tissue management models for medical research: preparation for implementation of the 2012 revision of the Bioethics and Safety Act of Korea.

PubMed

Ryu, Young-Joon; Kim, Hankyeom; Jang, Sejin; Koo, Young-Mo

2013-06-01

Efficient management of human tissue samples is a critical issue; the supply of samples is unable to satisfy the current demands for research. Lack of informed consent is also an ethical problem. One of the goals of the 2012 revision of Korea's Bioethics and Safety Act was to implement regulations that govern the management of human tissue samples. To remain competitive, medical institutions must prepare for these future changes. In this report, we review two tissue management models that are currently in use; model 1 is the most common system utilized by hospitals in Korea and model 2 is implemented by some of the larger institutions. We also propose three alternative models that offer advantages over the systems currently in use. Model 3 is a multi-bank model that protects the independence of physicians and pathologists. Model 4 utilizes a comprehensive single bioresource bank; although in this case, the pathologists gain control of the samples, which may make it difficult to implement. Model 5, which employs a bioresource utilization steering committee (BUSC), is viable to implement and still maintains the advantages of Model 4. To comply with the upcoming law, we suggest that physicians and pathologists in an institution should collaborate to choose one of the improved models of tissue management system that best fits for their situation.

In vitro fabrication of a tissue engineered human cardiovascular patch for future use in cardiovascular surgery.

PubMed

Yang, Chao; Sodian, Ralf; Fu, Ping; Lüders, Cora; Lemke, Thees; Du, Jing; Hübler, Michael; Weng, Yuguo; Meyer, Rudolf; Hetzer, Roland

2006-01-01

One approach to tissue engineering has been the development of in vitro conditions for the fabrication of functional cardiovascular structures intended for implantation. In this experiment, we developed a pulsatile flow system that provides biochemical and biomechanical signals in order to regulate autologous, human patch-tissue development in vitro. We constructed a biodegradable patch scaffold from porous poly-4-hydroxy-butyrate (P4HB; pore size 80 to 150 microm). The scaffold was seeded with pediatric aortic cells. The cell-seeded patch constructs were placed in a self-developed bioreactor for 7 days to observe potential tissue formation under dynamic cell culture conditions. As a control, cell-seeded scaffolds were not conditioned in the bioreactor system. After maturation in vitro, the analysis of the tissue engineered constructs included biochemical, biomechanical, morphologic, and immunohistochemical examination. Macroscopically, all tissue engineered constructs were covered by cells. After conditioning in the bioreactor, the cells were mostly viable, had grown into the pores, and had formed tissue on the patch construct. Electron microscopy showed confluent smooth surfaces. Additionally, we demonstrated the capacity to generate collagen and elastin under in vitro pulsatile flow conditions in biochemical examination. Biomechanical testing showed mechanical properties of the tissue engineered human patch tissue without any statistical differences in strength or resistance to stretch between the static controls and the conditioned patches. Immunohistochemical examination stained positive for alpha smooth muscle actin, collagen type I, and fibronectin. There was minor tissue formation in the nonconditioned control samples. Porous P4HB may be used to fabricate a biodegradable patch scaffold. Human vascular cells attached themselves to the polymeric scaffold, and extracellular matrix formation was induced under controlled biomechanical and biodynamic stimuli in a self-developed pulsatile bioreactor system.

AT1 expression in human urethral stricture tissue.

PubMed

Siregar, Safendra; Parardya, Aga; Sibarani, Jupiter; Romdan, Tjahjodjati; Adi, Kuncoro; Hernowo, Bethy S; Yantisetiasti, Anglita

2017-01-01

Urethral stricture has a high recurrence rate. There is a common doctrine stating that "once a stricture, always a stricture". This fibrotic disease pathophysiology, pathologically characterized by excessive production, deposition and contraction of extracellular matrix is unknown. Angiotensin II type 1 (AT 1 ) receptor primarily induces angiogenesis, cellular proliferation and inflammatory responses. AT 1 receptors are also expressed in the fibroblasts of hypertrophic scars, whereas angiotensin II (AngII) regulates DNA synthesis in hypertrophic scar fibroblasts through a negative cross talk between AT 1 and angiotensin II type 2 (AT 2 ) receptors, which might contribute to the formation and maturation of human hypertrophic scars. This study was conducted to determine the expression of AT 1 receptors in urethral stricture tissues. Urethral stricture tissues were collected from patients during anastomotic urethroplasty surgery. There were 24 tissue samples collected in this study with 2 samples of normal urethra for the control group. Immunohistochemistry study was performed to detect the presence of AT 1 receptor expression. Data were analyzed using Mann-Whitney U test, and statistical analysis was performed with SPSS version 20. This study showed that positive staining of AT 1 receptor was found in all urethral stricture tissues (n=24). A total of 8.33% patients had low intensity, 41.67% had moderate intensity and 50% had high intensity of AT 1 receptors, while in the control group, 100% patients had no intensity of AT 1 receptors. Using the Mann-Whitney U test, it was found that urethral stricture tissue had a higher intensity of AT 1 receptors than normal urethral tissue with a p -value = 0.012. The results showed that AT 1 receptor had a higher intensity in the urethral stricture tissue and that AT 1 receptor may play an important role in the development of urethral stricture.

Comparison of two modalities: a novel technique, 'chromohysteroscopy', and blind endometrial sampling for the evaluation of abnormal uterine bleeding.

PubMed

Alay, Asli; Usta, Taner A; Ozay, Pinar; Karadugan, Ozgur; Ates, Ugur

2014-05-01

The objective of this study was to compare classical blind endometrial tissue sampling with hysteroscopic biopsy sampling following methylene blue dyeing in premenopausal and postmenopausal patients with abnormal uterine bleeding. A prospective case-control study was carried out in the Office Hysteroscopy Unit. Fifty-four patients with complaints of abnormal uterine bleeding were evaluated. Data of 38 patients were included in the statistical analysis. Three groups were compared by examining samples obtained through hysteroscopic biopsy before and after methylene blue dyeing, and classical blind endometrial tissue sampling. First, uterine cavity was evaluated with office hysteroscopy. Methylene blue dye was administered through the hysteroscopic inlet. Tissue samples were obtained from stained and non-stained areas. Blind endometrial sampling was performed in the same patients immediately after the hysteroscopy procedure. The results of hysteroscopic biopsy from methylene blue stained and non-stained areas and blind biopsy were compared. No statistically significant differences were determined in the comparison of biopsy samples obtained from methylene-blue stained, non-stained areas and blind biopsy (P > 0.05). We suggest that chromohysteroscopy is not superior to endometrial sampling in cases of abnormal uterine bleeding. Further studies with greater sample sizes should be performed to assess the validity of routine use of endometrial dyeing. © 2014 The Authors. Journal of Obstetrics and Gynaecology Research © 2014 Japan Society of Obstetrics and Gynecology.

How anonymous is ‘anonymous’? Some suggestions towards a coherent universal coding system for genetic samples

PubMed Central

Schmidt, Harald; Callier, Shawneequa

2012-01-01

So-called ‘anonymous’ tissue samples are widely used in research. Because they lack externally identifying information, they are viewed as useful in reconciling conflicts between the control, privacy and confidentiality interests of those from whom the samples originated and the public (or commercial) interest in carrying out research, as reflected in ‘consent or anonymise’ policies. High level guidance documents suggest that withdrawal of consent and samples and the provision of feedback are impossible in the case of anonymous samples. In view of recent developments in science and consumer-driven genomics the authors argue that such statements are misleading and only muddle complex ethical questions about possible entitlements to control over samples. The authors therefore propose that terms such as ‘anonymised’, ‘anonymous’ or ‘non-identifiable’ be removed entirely from documents describing research samples, especially from those aimed at the public. This is necessary as a matter of conceptual clarity and because failure to do so may jeopardise public trust in the governance of large scale databases. As there is wide variation in the taxonomy for tissue samples and no uniform national or international standards, the authors propose that a numeral-based universal coding system be implemented that focuses on specifying incremental levels of identifiability, rather than use terms that imply that the reidentification of research samples and associated actions are categorically impossible. PMID:22345546

How anonymous is 'anonymous'? Some suggestions towards a coherent universal coding system for genetic samples.

PubMed

Schmidt, Harald; Callier, Shawneequa

2012-05-01

So-called 'anonymous' tissue samples are widely used in research. Because they lack externally identifying information, they are viewed as useful in reconciling conflicts between the control, privacy and confidentiality interests of those from whom the samples originated and the public (or commercial) interest in carrying out research, as reflected in 'consent or anonymise' policies. High level guidance documents suggest that withdrawal of consent and samples and the provision of feedback are impossible in the case of anonymous samples. In view of recent developments in science and consumer-driven genomics the authors argue that such statements are misleading and only muddle complex ethical questions about possible entitlements to control over samples. The authors therefore propose that terms such as 'anonymised', 'anonymous' or 'non-identifiable' be removed entirely from documents describing research samples, especially from those aimed at the public. This is necessary as a matter of conceptual clarity and because failure to do so may jeopardise public trust in the governance of large scale databases. As there is wide variation in the taxonomy for tissue samples and no uniform national or international standards, the authors propose that a numeral-based universal coding system be implemented that focuses on specifying incremental levels of identifiability, rather than use terms that imply that the reidentification of research samples and associated actions are categorically impossible.

Tc17 Cells in Patients with Uterine Cervical Cancer

PubMed Central

Zhang, Yan; Hou, Fei; Liu, Xin; Ma, Daoxin; Zhang, Youzhong; Kong, Beihua; Cui, Baoxia

2014-01-01

Background The existence of Tc17 cells was recently shown in several types of infectious and autoimmune diseases, but their distribution and functions in uterine cervical cancer (UCC) have not been fully elucidated. Methods The frequency of Tc17 cells in peripheral blood samples obtained from UCC patients, cervical intraepithelial neoplasia (CIN) patients and healthy controls was determined by flow cytometry. Besides, the prevalence of Tc17 cells and their relationships to Th17 cells and Foxp3-expressing T cells as well as microvessels in tissue samples of the patients were assessed by immunohistochemistry staining. Results Compared to controls, patients with UCC or CIN had a higher proportion of Tc17 cells in both peripheral blood and cervical tissues, but the level of Tc17 cells in UCC tissues was significantly higher than that in CIN tissues. Besides, the increased level of Tc17 in UCC patients was associated with the status of pelvic lymph node metastases and increased microvessel density. Finally, significant correlations of infiltration between Tc17 cells and Th17 cells or Foxp3-expressing T cells were observed in UCC and CIN tissues. Conclusions This study indicates that Tc17 cell infiltration in cervical cancers is associated with cancer progression accompanied by increased infiltrations of Th17 cells and regulatory T cells as well as promoted tumor vasculogenesis. PMID:24523865

Spatially-Resolved Proteomics: Rapid Quantitative Analysis of Laser Capture Microdissected Alveolar Tissue Samples

DOE Office of Scientific and Technical Information (OSTI.GOV)

Clair, Geremy; Piehowski, Paul D.; Nicola, Teodora

Global proteomics approaches allow characterization of whole tissue lysates to an impressive depth. However, it is now increasingly recognized that to better understand the complexity of multicellular organisms, global protein profiling of specific spatially defined regions/substructures of tissues (i.e. spatially-resolved proteomics) is essential. Laser capture microdissection (LCM) enables microscopic isolation of defined regions of tissues preserving crucial spatial information. However, current proteomics workflows entail several manual sample preparation steps and are challenged by the microscopic mass-limited samples generated by LCM, and that impact measurement robustness, quantification, and throughput. Here, we coupled LCM with a fully automated sample preparation workflow thatmore » with a single manual step allows: protein extraction, tryptic digestion, peptide cleanup and LC-MS/MS analysis of proteomes from microdissected tissues. Benchmarking against the current state of the art in ultrasensitive global proteomic analysis, our approach demonstrated significant improvements in quantification and throughput. Using our LCM-SNaPP proteomics approach, we characterized to a depth of more than 3,400 proteins, the ontogeny of protein changes during normal lung development in laser capture microdissected alveolar tissue containing ~4,000 cells per sample. Importantly, the data revealed quantitative changes for 350 low abundance transcription factors and signaling molecules, confirming earlier transcript-level observations and defining seven modules of coordinated transcription factor/signaling molecule expression patterns, suggesting that a complex network of temporal regulatory control directs normal lung development with epigenetic regulation fine-tuning pre-natal developmental processes. Our LCM-proteomics approach facilitates efficient, spatially-resolved, ultrasensitive global proteomics analyses in high-throughput that will be enabling for several clinical and biological applications.« less

Proteoglycan depletion and size reduction in lesions of early grade chondromalacia of the patella.

PubMed

Väätäinen, U; Häkkinen, T; Kiviranta, I; Jaroma, H; Inkinen, R; Tammi, M

1995-10-01

To determine the content and molecular size of proteoglycans (PGs) in patellar chondromalacia (CM) and control cartilages as a first step in investigating the role of matrix alterations in the pathogenesis of this disease. Chondromalacia tissue from 10 patients was removed with a surgical knife. Using identical techniques, apparently healthy cartilage of the same site was obtained from 10 age matched cadavers (mean age 31 years in both groups). Additional pathological cartilage was collected from 67 patients with grades II-IV CM (classified according to Outerbridge) using a motorised shaver under arthroscopic control. The shaved cartilage chips were collected with a dense net from the irrigation fluid of the shaver. The content of tissue PGs was determined by Safranin O precipitation or uronic acid content, and the molecular size by mobility on agarose gel electrophoresis. The mean PG content of the CM tissue samples with a knife was dramatically reduced, being only 15% of that in controls. The cartilage chips collected from shaving operations of grades II, III, and IV CM showed a decreasing PG content: 9%, 5%, and 1% of controls, respectively. Electrophoretic analysis of PGs extracted with guanidium chloride from the shaved tissue samples suggested a significantly reduced size of aggrecans in the mild (grade II) lesions. These data show that there is already a dramatic and progressive depletion of PGs in CM grade II lesions. This explains the softening of cartilage, a typical finding in the arthroscopic examination of CM. The PG size reduction observed in grade II implicates proteolytic attack as a factor in the pathogenesis of CM.

A flow-proteometric platform for analyzing protein concentration (FAP): Proof of concept for quantification of PD-L1 protein in cells and tissues.

PubMed

Chou, Chao-Kai; Huang, Po-Jung; Tsou, Pei-Hsiang; Wei, Yongkun; Lee, Heng-Huan; Wang, Ying-Nai; Liu, Yen-Liang; Shi, Colin; Yeh, Hsin-Chih; Kameoka, Jun; Hung, Mien-Chie

2018-05-29

Protein expression level is critically related to the cell physiological function. However, current methodologies such as Western blot (WB) and Immunohistochemistry (IHC) in analyzing the protein level are rather semi-quantitative and without the information of actual protein concentration. We have developed a microfluidic technique termed a "flow-proteometric platform for analyzing protein concentration (FAP)" that can measure the concentration of a target protein in cells or tissues without the requirement of a calibration standard, e.g., the purified target molecules. To validate our method, we tested a number of control samples with known target protein concentrations and showed that the FAP measurement resulted in concentrations that well matched the actual concentrations in the control samples (coefficient of determination [R 2 ], 0.998), demonstrating a dynamic range of concentrations from 0.13 to 130 pM of a detection for 2 min. We successfully determined a biomarker protein (for predicting the treatment response of cancer immune check-point therapy) PD-L1 concentration in cancer cell lines (HeLa PD-L1 and MDA-MB-231) and breast cancer patient tumor tissues without any prior process of sample purification and standard line construction. Therefore, FAP is a simple, faster, and reliable method to measure the protein concentration in cells and tissues, which can support the conventional methods such as WB and IHC to determine the actual protein level. Copyright © 2018 Elsevier B.V. All rights reserved.

Microjet-assisted dye-enhanced diode laser ablation of cartilaginous tissue

NASA Astrophysics Data System (ADS)

Pohl, John; Bell, Brent A.; Motamedi, Massoud; Frederickson, Chris J.; Wallace, David B.; Hayes, Donald J.; Cowan, Daniel

1994-08-01

Recent studies have established clinical application of laser ablation of cartilaginous tissue. The goal of this study was to investigate removal of cartilaginous tissue using diode laser. To enhance the interaction of laser light with tissue, improve the ablation efficiency and localize the extent of laser-induced thermal damage in surrounding tissue, we studied the use of a novel delivery system developed by MicroFab Technologies to dispense a known amount of Indocyanine Green (ICG) with a high spatial resolution to alter the optical properties of the tissue in a controlled fashion. Canine intervertebral disks were harvested and used within eight hours after collection. One hundred forty nL of ICG was topically applied to both annulus and nucleus at the desired location with the MicroJet prior to each irradiation. Fiber catheters (600 micrometers ) were used and positioned to irradiate the tissue with a 0.8 mm spot size. Laser powers of 3 - 10 W (Diomed, 810 nm) were used to irradiate the tissue with ten pulses (200 - 500 msec). Discs not stained with ICG were irradiated as control samples. Efficient tissue ablation (80 - 300 micrometers /pulse) was observed using ICG to enhance light absorption and confine thermal damage while there was no observable ablation in control studied. The extent of tissue damage observed microscopically was limited to 50 - 100 micrometers . The diode laser/Microjet combination showed promise for applications involving removal of cartilaginous tissue. This procedure can be performed using a low power compact diode laser, is efficient, and potentially more economical compared to procedures using conventional lasers.

Characterization of PD-L1 expression in Chinese non-small cell lung cancer patients with PTEN expression as a means for tissue quality screening.

PubMed

Zhang, Xu-Chao; Cao, Xu; Sun, Chun; Xie, Zhi; Guo, Jian-Jun; Yang, Jin-Ji; Yang, Xue-Ning; Dai, Hang-Jun; Li, Su-Chun; Xu, Xin-Ran; Zuo, Yun-Xia; Chen, Meng; Koeppen, Hartmut; He, Jing; Kiermaier, Astrid; Shames, David; Cheng, Gang; Wu, Yi-Long

2018-03-01

The goal of this study is to evaluate PD-L1 prevalence and its association with major clinical characteristics in Chinese non-small cell lung cancer (NSCLC) patients to inform the clinical development of anti-PD1/PD-L1 agents in this population. We used phosphatase and tensin homolog (PTEN) expression through IHC as a surrogate tissue quality marker to screen surgical NSCLC samples in tissue microarray (TMA; 172 cases) or whole-section (268 cases) format. The samples were then analyzed with a clinically validated PD-L1 IHC assay. The results were correlated with baseline characteristics and clinical outcomes. PTEN IHC showed that 108 TMA samples and 105 whole-section samples qualified for PD-L1 IHC. With a clinically relevant cutoff, 41.7% of the TMA samples were PD-L1 positive. PD-L1 level was much lower in EGFR-mutant patients and seemed to be a favorable prognostic factor for both overall survival (OS) and recurrence-free survival (RFS). These findings were confirmed in the whole-section samples except that their survival data were not mature enough for correlation analysis. In summary, PD-L1 expression was detected in approximately 40% of PTEN-qualified Chinese NSCLC samples, negatively correlated with EGFR mutation and seemed to be a favorable prognostic factor for both OS and RFS. Notably, the different results from PTEN-qualified and PTEN-disqualified samples underscore the importance of tissue quality control prior to biomarker testing.

Circulating testosterone and prostate-specific antigen in nipple aspirate fluid and tissue are associated with breast cancer.

PubMed Central

Sauter, Edward R; Tichansky, David S; Chervoneva, Inna; Diamandis, Eleftherios P

2002-01-01

Preliminary evidence has associated testosterone and prostate-specific antigen (PSA) with breast cancer. Our objective was to determine whether a) testosterone levels in nipple aspirate fluid (NAF), serum, or breast tissue are associated with breast cancer; b) testosterone levels in serum are associated with levels in NAF; c) PSA in NAF, serum, or breast tissue is associated with breast cancer; and d) serum PSA is associated with NAF PSA levels. We obtained 342 NAF specimens from 171 women by means of a modified breast pump. Additionally, we collected 201 blood samples from 99 women and 51 tissue samples from 41 subjects who underwent surgical resection for suspected disease. Women currently using birth control pills or hormone replacement therapy were excluded from the study. Controlling for age and menopausal status, serum testosterone was significantly increased in women with breast cancer (p = 0.002). NAF and serum testosterone levels were not associated. Neither NAF nor tissue testosterone was associated with breast cancer. Controlling for menopausal status and age, NAF PSA was significantly decreased in women with breast cancer (p < 0.001). We did not find serum PSA to be associated with breast cancer, although we found an indication that, in postmenopausal women, its levels were lower in women with cancer. Serum PSA was associated with NAF PSA in postmenopausal women (p < 0.001). PSA levels in cancerous tissue were significantly lower than in benign breast specimens from subjects without cancer (p = 0.011), whereas levels of PSA in histologically benign specimens from subjects with cancer were intermediate. Our results suggest that serum testosterone is increased and NAF PSA is decreased in women with breast cancer, with PSA expression being higher in normal than in cancerous breast tissues. NAF and serum PSA levels in postmenopausal women are correlated, suggesting that as laboratory assessment of PSA becomes more sensitive, serum PSA may become useful in identifying women with breast cancer. PMID:11882474

[The role of telomerase activity in non-invasive diagnostics of bladder cancer].

PubMed

Glybochko, P V; Alyaev, J G; Potoldykova, N V; Polyakovsky, K A; Vinarov, A Z; Glukhov, A I; Gordeev, S A

2016-08-01

To evaluate the potentials of determining the telomerase activity (TA) in the cellular material of the urine for noninvasive diagnosis of bladder cancer (BC). Evaluation of TA was performed in the urine of 48 patients with bladder cancer (study group) before and after transurethral resection of the bladder wall (n=38), an open resection of the bladder (n=4), and cystectomy (n=6). TA was also evaluated in 48 tumor tissue samples obtained from these patients during removal of the bladder tumor. Each sample of the tumor tissue was separated into two parts, one of which was subjected to histological examination, and the latter was used to determine the telomerase activity. In all cases, the diagnosis of bladder cancer was confirmed morphologically. Determination of TA in the samples was performed by the modified TRAP-method (telomerase repeat amplification protocol), RT-PCR, PCR, and electrophoresis. As a control, cell material of the urine and tissue in 12 patients with chronic cystitis was investigated. TA before surgery was found in 45 (93.75%) of 48 samples of cellular material of the urine from patients with suspected bladder cancer. BC was histologically verified in all patients in this group. In the postoperative period, TA was not observed in the 48 samples of cellular material of the urine from patients with BC. In the control group of patients with histologically verified cystitis, weak TA was determined only in one sample of cellular material of the urine. The analysis indicates statistically significant predominance of patients with bladder cancer in case of TA in the urine (P=0.001). TA was detected in all samples of tumor tissue. We also analyzed the dependence of TA levels in urine and tissue on the degree of BC differentiation. In patients with highly differentiated BC, mean AT in the cellular materials of the urine was 0,61% (n=15), in patients with moderately differentiated BC - 0.95% (n=23), in patients with low-grade bladder cancer - 1.33% (n=10); in other words, increase in the TA levels with decreasing the degree of differentiation was observed. This finding can be used in the prognosis of the course of disease based on determining the TA level in these patients. Preliminary data indicate the possibility of use of determining the TA in cellular material of the urine for the diagnosis and monitoring of bladder cancer recurrence.

Healing potential of transplanted allogeneic chondrocytes of three different sources in lesions of the avascular zone of the meniscus: a pilot study.

PubMed

Weinand, Christian; Peretti, Giuseppe M; Adams, Samuel B; Randolph, Mark A; Savvidis, Estafios; Gill, Thomas J

2006-11-01

Successful treatment of tears to the avascular region of the meniscus remains a challenge. Current repair techniques, such as sutures and anchors, are effective in stabilizing the peripheral, vascularized regions of the meniscus, but are not adequate for promoting healing in the avascular region. The purpose of this study was to demonstrate the healing ability of a tissue-engineered repair technique using allogenic chondrocytes from three different sources for the avascular zone of the meniscus. Articular, auricular, and costal chondrocytes were harvested from 3-month-old Yorkshire swine. A 1-cm bucket-handle lesion was created in the avascular zone of each three swine. A cell-scaffold construct, composed of a single chondrocyte cell type and Vicryl mesh, was implanted into the lesion and secured with two vertical mattress sutures. Controls consisted of each three sutured unseeded mesh implants, suture only, and untreated lesions. The swine were allowed immediate post-operative full weight bearing. Menisci and controls were harvested after 12 weeks. In all experimental samples, lesion closure was observed. Gross mechanical testing with two Adson forceps demonstrated bonding of the lesion. Histological analysis showed formation of new tissue in all three experimental samples. None of the control samples demonstrated closure and formation of new matrix. We present preliminary data that demonstrates the potential of a tissue-engineered, allogenic cellular repair to provide successful healing of lesions in the avascular zone in a large animal model.

Organochlorine pesticide levels in adipose tissue of pregnant women in Veracruz, Mexico.

PubMed

Herrero-Mercado, Margarita; Waliszewski, S M; Valencia-Quintana, R; Caba, M; Hernández-Chalate, F; García-Aguilar, E; Villalba, R

2010-06-01

DDT and Lindane (gamma-HCH) which were used until 1999 in Mexico, have provided great benefits in the combat of vectors that spread infection-borne diseases and in agriculture for crop protection. The persistence in the environment and their accumulative properties results in bioconcentration in lipid rich tissues of the human body that reflect the extent of environmental pollution. Human adipose tissue samples were taken during 2009 from abdominal cavities of 69 pregnant women by cesarean surgery and from 34 samples of control donors by autopsy in Veracruz State. The samples were analyzed by gas chromatography with ECD. The results of mean levels (mg/kg on fat basis) were higher in controls compared to pregnant women beta-HCH 0.064 vs 0.027; pp'DDE 1.187 vs. 0.745; op'DDT 0.016 vs. 0.011; pp'DDT 0.117 vs. 0.099 and Sigma-DDT 1.337 vs. 0.854. The pregnant women group was divided according to age: up to 20, 20-30, and more than 30 years, and presented an increase for the more persistent pesticides with age in terms of mean concentrations and a more pronounced higher correlation in medians levels. Pairing Body Mass Index to organochlorine pesticide mean levels revealed no correlation between these factors in pregnant women.

Assessment of test method variables for in vitro skin irritation testing of medical device extracts.

PubMed

Olsen, Daniel S; Lee, Michelle; Turley, Audrey P

2018-08-01

Skin irritation is an important component of the biological safety evaluation of medical devices. This testing has typically been performed using in vivo models. However, in an effort to reduce the need for in vivo testing, alternative methods for assessing skin irritation potential in vitro have been developed using a Reconstructed Human Epidermis (RhE) model. During the development of the protocol for the round robin validation of in vitro irritation testing for medical device extracts, it became clear that there were three points in the procedure where different options may be validated within each laboratory for routine testing: sample exposure time (18 vs 24h), SDS positive control concentration, and cytokine (IL-1α) release testing. The goal of our study was to evaluate the effect of these variables. EpiDerm™ tissues were exposed to extracts of three plain polymer samples, and four polymers embedded with known irritant chemicals. Exposures were performed for 18 and 24h. Resulting tissue viability was assessed by MTT reduction and IL-1α release was assessed by ELISA. Testing was also performed using various concentrations of SDS ranging from 0.5 to 1% (w/v). Overall, results were similar for samples tested and 18 and 24h, but the 18h exposure time has the potential to have an impact on the results of some sample types. IL-1α testing was shown to be useful to clarify conflicting tissue viability results. Use of a lower concentration of SDS as a positive control can help prevent issues that arise from excessive tissue damage often caused by 1% SDS. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

Effects of geared motor characteristics on tactile perception of tissue stiffness.

PubMed

Longnion, J; Rosen, J; Sinanan, M; Hannaford, B

2001-01-01

Endoscopic haptic surgical devices have shown promise in addressing the loss of tactile sensation associated with minimally invasive surgery. However, these devices must be capable of generating forces and torques similar to those applied on the tissue with a standard endoscopic tool. Geared motors are a possible solution for actuation; however, they possess mechanical characteristics that could potentially interfere with tactile perception of tissue qualities. The aim of the current research was to determine how the characteristics of a geared motor suitable for a haptic surgical device affect a user's perception of stiffness. The experiment involved six blindfolded subjects who were asked to discriminate the stiffness of six distinct silicone rubber samples whose mechanical properties are similar to those of soft tissue. Using a novel testing device whose dimensions approximated those of an endoscopic grasper, each subject palpated 30 permutations of sample pairs for each of three types of mechanical loads; the motor (friction and inertia), a flywheel (with the same inertia as motor), and a control (no significant mechanical interference). One factor ANOVA of the error scores and palpation time showed that no significant difference existed among error scores, but mean palpation time for the control was significantly less than for the other two methods. These results indicated that the mechanical characteristics of a geared motor chosen for application in a haptic surgical device did not interfere with the subjects' perception of the silicone samples' stiffness, but these characteristics may significantly affect the energy expenditure and time required for tissue palpation. Therefore, before geared motors can be considered for use in haptic surgical devices, consideration should be given to factors such as palpation speed and fatigue.

Biochemical and histopathologic analysis of the effects of periodontitis on left ventricular heart tissues of rats.

PubMed

Köse, O; Arabacı, T; Gedikli, S; Eminoglu, D Ö; Kermen, E; Kızıldağ, A; Kara, A; Ozkanlar, S; Yemenoglu, H

2017-04-01

Current epidemiological works have suggested that chronic infections, such as periodontitis, are associated with an increased risk of cardiovascular diseases, including hypertrophy and heart failure. However, mechanisms behind the association are not known. The aim of this study was to evaluate the effects of periodontitis on the serum lipid levels, inflammatory marker levels and left ventricular heart muscle tissues of rats. Eighteen male Sprague-Dawley rats were randomly divided into two groups: control (without ligature) and experimental periodontitis (EP; ligatured). Periodontitis was induced by placing ligatures (3.0 silk) at a submarginal position of the lower first molar teeth for 5 wk. Serum samples were collected for biochemical studies (C-reactive protein, interleukin-1β, tumor necrosis factor-α and serum lipids), after which the rats were killed and heart tissue samples were obtained for histopathological and immunological studies (nuclear factor kappa B and β-myosin heavy chain). Significant increases in C-reactive protein and interleukin-1β levels and no statistically significant increase in tumor necrosis factor-α level were observed in the EP group compared to the control group. In addition, total cholesterol, low-density lipoprotein cholesterol and triglyceride levels were significantly higher in the EP group. Stereological and immunological findings showed that the number of nuclear factor kappa B-p65- and β-myosin heavy chain-positive cardiomyocytes increased significantly in the left ventricular tissue samples of the rats with periodontitis. Early chronic phase effects of periodontitis on heart tissue are in the form of degenerative and hypotrophic changes. Prolonging the exposure to systemic inflammatory stress may increase the risk of occurrence of hypertrophic changes. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

RCL2, a potential formalin substitute for tissue fixation in routine pathological specimens.

PubMed

Masir, Noraidah; Ghoddoosi, Mahdiieh; Mansor, Suhada; Abdul-Rahman, Faridah; Florence, Chandramaya S; Mohamed-Ismail, Nor Azlin; Tamby, Mohammad-Rafaee; Md-Latar, Nani Harlina

2012-04-01

To investigate RCL2 as a fixative for tissue fixation in routine histopathological examination and to assess tissue suitability for ancillary investigations.   Forty-nine samples from 36 fresh specimens were cut into three equal pieces and fixed in RCL2 diluted in 100% ethanol, RCL2 in 95% ethanol, or neutral buffered formalin as control. Suitability for microtomy, quality of histomorphology, histochemistry, immunohistochemistry, fluorescent and silver in-situ hybridization analysis and extracted genomic DNA were assessed. Microtomy was straightforward in most tissue blocks, but there was difficulty in cutting in approximately a quarter of samples, which required careful handling by an experienced technician. There were no significant differences in tissue morphology between RCL2- and formalin-fixed tissues (P=0.08). Generally, the quality of histochemical staining, immunohistochemistry and in-situ hybridization were comparable to that of formalin-fixed tissues. Inconsistent immunoreactivity was noted, however, with antibodies against pan-cytokeratin and progesterone receptor. Genomic DNA concentration was higher in RCL2-fixed tissues. Using RCL2 diluted in 95% ethanol did not affect fixation quality. RCL2 is a potential formalin substitute suitable as a fixative for use in routine histopathological examination; however, difficulty in microtomy and occasional discrepancies in immunohistochemical reactivity require further optimization of the methodology. © 2012 Blackwell Publishing Ltd.

Spectral grading and Gleason grading of malignant prostate tissue using Stokes shift spectra

NASA Astrophysics Data System (ADS)

Al Salhi, M.; Masilamani, V.; Rabah, D.; Farhat, K.; Liu, C. H.; Pu, Y.; Alfano, R. R.

2012-01-01

Gleason score is the most common method of grading the virulence of prostate malignancy and is based on the pathological assessment of morphology of cellular matrix. Since this involves the excision of the tissue, we are working on a new, minimally invasive, non contact, procedure of spectral diagnosis of prostate malignancy. In this preliminary in vitro study reported here, we have analyzed 27 tissue samples (normal control =7: benign=8: malignant =12) by Stokes' shift spectra (SSS) to establish a one- to- one correlation between spectral grading and Gleason grading.

Hydrofocusing Bioreactor for Three-Dimensional Cell Culture

NASA Technical Reports Server (NTRS)

Gonda, Steve R.; Spaulding, Glenn F.; Tsao, Yow-Min D.; Flechsig, Scott; Jones, Leslie; Soehnge, Holly

2003-01-01

The hydrodynamic focusing bioreactor (HFB) is a bioreactor system designed for three-dimensional cell culture and tissue-engineering investigations on orbiting spacecraft and in laboratories on Earth. The HFB offers a unique hydrofocusing capability that enables the creation of a low-shear culture environment simultaneously with the "herding" of suspended cells, tissue assemblies, and air bubbles. Under development for use in the Biotechnology Facility on the International Space Station, the HFB has successfully grown large three-dimensional, tissuelike assemblies from anchorage-dependent cells and grown suspension hybridoma cells to high densities. The HFB, based on the principle of hydrodynamic focusing, provides the capability to control the movement of air bubbles and removes them from the bioreactor without degrading the low-shear culture environment or the suspended three-dimensional tissue assemblies. The HFB also provides unparalleled control over the locations of cells and tissues within its bioreactor vessel during operation and sampling.

Validation of an LC-MS/MS method to measure tacrolimus in rat kidney and liver tissue and its application to human kidney biopsies.

PubMed

Noll, Benjamin D; Coller, Janet K; Somogyi, Andrew A; Morris, Raymond G; Russ, Graeme R; Hesselink, Dennis A; Van Gelder, Teun; Sallustio, Benedetta C

2013-10-01

Tacrolimus (TAC) has a narrow therapeutic index and high interindividual and intraindividual pharmacokinetic variability, necessitating therapeutic drug monitoring to individualize dosage. Recent evidence suggests that intragraft TAC concentrations may better predict transplant outcomes. This study aimed to develop a method for the quantification of TAC in small biopsy-sized samples of rat kidney and liver tissue, which could be applied to clinical biopsy samples from kidney transplant recipients. Kidneys and livers were harvested from Mrp2-deficient TR- Wistar rats administered TAC (4 mg·kg·d for 14 days, n = 8) or vehicle (n = 10). Tissue samples (0.20-1.00 mg of dry weight) were solubilized enzymatically and underwent liquid-liquid extraction before analysis by liquid chromatography tandem mass spectrometry method. TAC-free tissue was used in the calibrator and quality control samples. Analyte detection was accomplished using positive electrospray ionization (TAC: m/z 821.5 → 768.6; internal standard ascomycin m/z 809.3 → 756.4). Calibration curves (0.04-2.6 μg/L) were linear (R > 0.99, n = 10), with interday and intraday calibrator coefficients of variation and bias <17% at the lower limit of quantification and <15% at all other concentrations (n = 6-10). Extraction efficiencies for TAC and ascomycin were approximately 70%, and matrix effects were minimal. Rat kidney TAC concentrations were higher (range 109-190 pg/mg tissue) than those in the liver (range 22-53 pg/mg of tissue), with median tissue/blood concentrations ratios of 72.0 and 17.6, respectively. In 2 transplant patients, kidney TAC concentrations ranged from 119 to 285 pg/mg of tissue and were approximately 20 times higher than whole blood trough TAC concentrations. The method displayed precision and accuracy suitable for application to TAC measurement in human kidney biopsy tissue.

Monochromatic computed microtomography using laboratory and synchrotron sources and X-ray fluorescence analysis for comprehensive analysis of structural changes in bones.

PubMed

Buzmakov, Alexey; Chukalina, Marina; Nikolaev, Dmitry; Gulimova, Victoriya; Saveliev, Sergey; Tereschenko, Elena; Seregin, Alexey; Senin, Roman; Zolotov, Denis; Prun, Victor; Shaefer, Gerald; Asadchikov, Victor

2015-06-01

A combination of X-ray tomography at different wavelengths and X-ray fluorescence analysis was applied in the study of two types of bone tissue changes: prolonged presence in microgravity conditions and age-related bone growth. The proximal tail vertebrae of geckos were selected for investigation because they do not bear the supporting load in locomotion, which allows them to be considered as an independent indicator of gravitational influence. For the vertebrae of geckos no significant differences were revealed in the elemental composition of the flight samples and the synchronous control samples. In addition, the gecko bone tissue samples from the jaw apparatus, spine and shoulder girdle were measured. The dynamics of structural changes in the bone tissue growth was studied using samples of a human fetal hand. The hands of human fetuses of 11-15 weeks were studied. Autonomous zones of calcium accumulation were found not only in individual fingers but in each of the investigated phalanges. The results obtained are discussed.

Monochromatic computed microtomography using laboratory and synchrotron sources and X-ray fluorescence analysis for comprehensive analysis of structural changes in bones1

PubMed Central

Buzmakov, Alexey; Chukalina, Marina; Nikolaev, Dmitry; Gulimova, Victoriya; Saveliev, Sergey; Tereschenko, Elena; Seregin, Alexey; Senin, Roman; Zolotov, Denis; Prun, Victor; Shaefer, Gerald; Asadchikov, Victor

2015-01-01

A combination of X-ray tomography at different wavelengths and X-ray fluorescence analysis was applied in the study of two types of bone tissue changes: prolonged presence in microgravity conditions and age-related bone growth. The proximal tail vertebrae of geckos were selected for investigation because they do not bear the supporting load in locomotion, which allows them to be considered as an independent indicator of gravitational influence. For the vertebrae of geckos no significant differences were revealed in the elemental composition of the flight samples and the synchronous control samples. In addition, the gecko bone tissue samples from the jaw apparatus, spine and shoulder girdle were measured. The dynamics of structural changes in the bone tissue growth was studied using samples of a human fetal hand. The hands of human fetuses of 11–15 weeks were studied. Autonomous zones of calcium accumulation were found not only in individual fingers but in each of the investigated phalanges. The results obtained are discussed. PMID:26089762

Measuring optical properties of a blood vessel model using optical coherence tomography

NASA Astrophysics Data System (ADS)

Levitz, David; Hinds, Monica T.; Tran, Noi; Vartanian, Keri; Hanson, Stephen R.; Jacques, Steven L.

2006-02-01

In this paper we develop the concept of a tissue-engineered optical phantom that uses engineered tissue as a phantom for calibration and optimization of biomedical optics instrumentation. With this method, the effects of biological processes on measured signals can be studied in a well controlled manner. To demonstrate this concept, we attempted to investigate how the cellular remodeling of a collagen matrix affected the optical properties extracted from optical coherence tomography (OCT) images of the samples. Tissue-engineered optical phantoms of the vascular system were created by seeding smooth muscle cells in a collagen matrix. Four different optical properties were evaluated by fitting the OCT signal to 2 different models: the sample reflectivity ρ and attenuation parameter μ were extracted from the single scattering model, and the scattering coefficient μ s and root-mean-square scattering angle θ rms were extracted from the extended Huygens-Fresnel model. We found that while contraction of the smooth muscle cells was clearly evident macroscopically, on the microscopic scale very few cells were actually embedded in the collagen. Consequently, no significant difference between the cellular and acellular samples in either set of measured optical properties was observed. We believe that further optimization of our tissue-engineering methods is needed in order to make the histology and biochemistry of the cellular samples sufficiently different from the acellular samples on the microscopic level. Once these methods are optimized, we can better verify whether the optical properties of the cellular and acellular collagen samples differ.

Do anesthetics and sampling strategies affect transcription analysis of fish tissues?

PubMed Central

Olsvik, Pål A; Lie, Kai K; Hevrøy, Ernst M

2007-01-01

Background The aim of the current examination was to evaluate if sedation and anesthetic treatment techniques affect the quality of RNA extracted from liver, gill, head kidney and brain tissues in Atlantic salmon Salmo salar L. Blood parameters were measured and tissue specimens sampled in six groups of fish; one control group (0 minutes), two groups kept in pure seawater in 90 liter tanks for 30 and 120 minutes, two groups treated with the anesthetic isoeugenol for 30 and 120 minutes, and one group kept in pure seawater for 105 minutes and then anaesthetized with metacaine for 15 minutes. RNA quality was assessed with the NanoDrop ND-1000 spectrophotometer (260/280 and 260/230 nm ratios) and with the Agilent Bioanalyzer (28S/18S ratio and RIN data) in samples either preserved in liquefied nitrogen (N2) or in RNAlater. In addition, the transcriptional levels of two fast-responding genes were quantified in gill and brain tissues. Results The results show that physiological stress during sampling does not affect the quality of RNA extracted from fish specimens. However, prolonged sedation (2 hours) resulted in a metabolic alkalosis that again affected the transcriptional levels of genes involved in ionoregulation and respiration. In gills, Na+-K+-ATPase α1b was significantly downregulated and hypoxia inducible factor 1 (HIF1) significantly upregulated after two hours of treatment with isoeugenol, suggesting that this commonly used sedative affects osmo-regulation and respiration in the fish. The results also suggest that for tissue preservation in general it is better to flash-freeze fish specimens in liquefied N2 than to use RNAlater. Conclusion Prolonged sedation may affect the transcription of fast-responding genes in tissues of fish. Two hours of sedation with isoeugenol resulted in downregulation of the Na+-K+-ATPase α1b gene and upregulation of the HIF1 gene in gills of Atlantic salmon. The quality of RNA extracted from tissue specimens, however, was not affected by sedation treatment. Flash-freezing of tissue specimens seems to be the preferred preservation technique, when sampling fish tissue specimens for RNA extraction. PMID:17559653

Immunohistochemical Analysis of the Role Connective Tissue Growth Factor in Drug-induced Gingival Overgrowth in Response to Phenytoin, Cyclosporine, and Nifedipine

PubMed Central

Anand, A. J.; Gopalakrishnan, Sivaram; Karthikeyan, R.; Mishra, Debasish; Mohapatra, Shreeyam

2018-01-01

Objective: To evaluate for the presence of connective tissue growth factor (CTGF) in drug (phenytoin, cyclosporine, and nifedipine)-induced gingival overgrowth (DIGO) and to compare it with healthy controls in the absence of overgrowth. Materials and Methods: Thirty-five patients were chosen for the study and segregated into study (25) and control groups (10). The study group consisted of phenytoin-induced (10), cyclosporine-induced (10), and nifedipine-induced (5) gingival overgrowth. After completing necessary medical evaluations, biopsy was done. The tissue samples were fixed in 10% formalin and then immunohistochemically evaluated for the presence of CTGF. The statistical analysis of the values was done using statistical package SPSS PC+ (Statistical Package for the Social Sciences, version 4.01). Results: The outcome of immunohistochemistry shows that DIGO samples express more CTGF than control group and phenytoin expresses more CTGF followed by nifedipine and cyclosporine. Conclusion: The study shows that there is an increase in the levels of CTGF in patients with DIGO in comparison to the control group without any gingival overgrowth. In the study, we compared the levels of CTGF in DIGO induced by three most commonly used drugs phenytoin, cyclosporine, and nifedipine. By comparing the levels of CTGF, we find that cyclosporine induces the production of least amount of CTGF. Therefore, it might be a more viable drug choice with reduced side effects. PMID:29629324

The impact of luteal phase support on endometrial estrogen and progesterone receptor expression: a randomized control trial

PubMed Central

2012-01-01

Background To assess the impact of luteal phase support on the expression of estrogen receptor (ER) alpha and progesterone receptors B (PR-B) on the endometrium of oocyte donors undergoing controlled ovarian hyperstimulation (COH). Methods A prospective, randomized study was conducted in women undergoing controlled ovarian hyperstimulation for oocyte donation. Participants were randomized to receive no luteal support, vaginal progesterone alone, or vaginal progesterone plus orally administered 17 Beta estradiol. Endometrial biopsies were obtained at 4 time points in the luteal phase and evaluated by tissue microarray for expression of ER alpha and PR-B. Results One-hundred and eight endometrial tissue samples were obtained from 12 patients. No differences were found in expression of ER alpha and PR-B among all the specimens with the exception of one sample value. Conclusions The administration of progesterone during the luteal phase of COH for oocyte donor cycles, either with or without estrogen, does not significantly affect the endometrial expression of ER alpha and PR. PMID:22360924

Association Between Mast Cells and Collagen Maturation in Chronic Periodontitis in Humans.

PubMed

E Ribeiro, Lívia S F; Dos Santos, Jean N; Rocha, Clarissa A G; Cury, Patricia R

2018-03-01

Mast cells (MCs) can influence the maturation of collagen fibers. This study evaluated the relationship between the distribution and degranulation of MCs and collagen maturation in human gingival tissue in chronic periodontitis. A total of 16 specimens of patients clinically diagnosed as periodontitis and 18 controls clinically diagnosed as healthy or gingivitis were included. Immunohistochemistry and Picrosirius staining were performed to identify MCs and assess collagen fibers, respectively. Chi-square, t test, and Pearson's correlation test ( p<0.05) were used. In control specimens, there was a positive association between MCs in the connective tissue and the presence of immature collagen ( p=0.001); in periodontitis samples, this association was not confirmed ( p≥0.12). There was no significant relationship between periodontal diagnosis and collagen maturation or MC degranulation ( p≥0.35). MC density was significantly higher ( p=0.04) in periodontitis tissue (339.01 ± 188.94 MCs/mm 2 ) than in control tissue (211.14 ± 131.13 MCs/mm 2 ) in the area of connective tissue containing inflammatory infiltrate. There was a correlation between the number of MCs and probing depth ( r = 0.34, p=0.04). MCs are involved in the pathogenesis of periodontal diseases and might be associated with collagen maturation in periodontal tissue during the early stages of periodontal disease pathogenesis.

Circulating tumor DNA profiling reveals clonal evolution and real-time disease progression in advanced hepatocellular carcinoma.

PubMed

Cai, Zhi-Xiong; Chen, Geng; Zeng, Yong-Yi; Dong, Xiu-Qing; Lin, Min-Jie; Huang, Xin-Hui; Zhang, Da; Liu, Xiao-Long; Liu, Jing-Feng

2017-09-01

Circulating tumor DNA (ctDNA) provides a potential non-invasive biomarker for cancer diagnosis and prognosis, but whether it could reflect tumor heterogeneity and monitor therapeutic responses in hepatocellular carcinoma (HCC) is unclear. Focusing on 574 cancer genes known to harbor actionable mutations, we identified the mutation repertoire of HCC tissues, and monitored the corresponding ctDNA features in blood samples to evaluate its clinical significance. Analysis of 3 HCC patients' mutation profiles revealed that ctDNA could overcome tumor heterogeneity and provide information of tumor burden and prognosis. Further analysis was conducted on the 4th HCC case with multiple lesion samples and sequential plasma samples. We identified 160 subclonal SNVs in tumor tissues as well as matched peritumor tissues with PBMC as control. 96.9% of this patient's tissue mutations could be also detected in plasma samples. These subclonal SNVs were grouped into 9 clusters according to their trends of cellular prevalence shift in tumor tissues. Two clusters constituted of tumor stem somatic mutations showed circulating levels relating with cancer progression. Analysis of tumor somatic mutations revealed that circulating level of such tumor stem somatic mutations could reflect tumor burden and even predict prognosis earlier than traditional strategies. Furthermore, HCK (p.V174M), identified as a recurrent/metastatic related mutation site, could promote migration and invasion of HCC cells. Taken together, study of mutation profiles in biopsy and plasma samples in HCC patients showed that ctDNA could overcome tumor heterogeneity and real-time track the therapeutic responses in the longitudinal monitoring. © 2017 UICC.

Distribution in the Western United States on Alfalfa and Cultivar Reaction to Mixed Populations of Ditylenchus dipsaci and Aphelenchoides ritzemabosi

PubMed Central

Gray, F. A.; Williams, J. L.; Griffin, G. D.; Wilson, T. E.

1994-01-01

Ditylenchus dipsaci and Aphelenchoides ritzemabosi were extracted from 29 of 40 plant samples (72.5%) collected from Arizona, California, Colorado, Idaho, Montana, Oregon, South Dakota, Utah, Washington, and Wyoming. Percentages of A. ritzemabosi in tissue of the 29 samples ranged from 1.77 to 67.82%. Only Ditylenchus dipsaci was recovered from the remaining 11 samples. All of the 16 fields sampled in Wyoming contained both nematodes. Percentages of A. ritzemabosi in the Wyoming samples ranged from 0.7-30.0%, with an overall mean of 10.3%. Individual plants collected from a field in Big Horn, Wyoming, all contained both nematodes. Percentages of A. ritzemabosi in tissue ranged from 5-70%. Alfalfa stem nematode symptomatic plants in 17 of 18 alfalfa cultivars collected from a screening nursery in California contained both nematodes, of which 10-94% were A. ritzemabosi. Only one cultivar had D. dipsaci only, and no entries had A. ritzemabosi only. Under environmentally controlled conditions, A. ritzemabosi reproduced in all nine alfalfa cultivars tested at 6 weeks of age with a mean reproductive factor (final population/initial population) of 4.1. There were more (P ≤ 0.05) A. ritzemabosi in stem and bud tissue of the susceptible cultivars at harvest than in the resistant cultivars with combined cultivar means of 238, 42, 78, and 4 A. ritzemabosi/g tissue for the susceptible, moderately resistant, resistant, and highly resistant cultivars, respectively. Percentage A. ritzemabosi in tissues decreased over time in seedlings but increased in older plants. PMID:19279952

Preservation of Multiple Mammalian Tissues to Maximize Science Return from Ground Based and Spaceflight Experiments.

PubMed

Choi, Sungshin; Ray, Hami E; Lai, San-Huei; Alwood, Joshua S; Globus, Ruth K

2016-01-01

Even with recent scientific advancements, challenges posed by limited resources and capabilities at the time of sample dissection continue to limit the collection of high quality tissues from experiments that can be conducted only infrequently and at high cost, such as in space. The resources and time it takes to harvest tissues post-euthanasia, and the methods and duration of long duration storage, potentially have negative impacts on sample quantity and quality, thereby limiting the scientific outcome that can be achieved. The goals of this study were to optimize methods for both sample recovery and science return from rodent experiments, with possible relevance to both ground based and spaceflight studies. The first objective was to determine the impacts of tissue harvest time post-euthanasia, preservation methods, and storage duration, focusing on RNA quality and enzyme activities in liver and spleen as indices of sample quality. The second objective was to develop methods that will maximize science return by dissecting multiple tissues after long duration storage in situ at -80°C. Tissues of C57Bl/6J mice were dissected and preserved at various time points post-euthanasia and stored at -80°C for up to 11 months. In some experiments, tissues were recovered from frozen carcasses which had been stored at -80°C up to 7 months. RNA quantity and quality was assessed by measuring RNA Integrity Number (RIN) values using an Agilent Bioanalyzer. Additionally, the quality of tissues was assessed by measuring activities of hepatic enzymes (catalase, glutathione reductase and GAPDH). Fresh tissues were collected up to one hour post-euthanasia, and stored up to 11 months at -80°C, with minimal adverse effects on the RNA quality of either livers or RNAlater-preserved spleens. Liver enzyme activities were similar to those of positive controls, with no significant effect observed at any time point. Tissues dissected from frozen carcasses that had been stored for up to 7 months at -80°C had variable results, depending on the specific tissue analyzed. RNA quality of liver, heart, and kidneys were minimally affected after 6-7 months of storage at -80°C, whereas RNA degradation was evident in tissues such as small intestine, bone, and bone marrow when they were collected from the carcasses frozen for 2.5 months. These results demonstrate that 1) the protocols developed for spaceflight experiments with on-orbit dissections support the retrieval of high quality samples for RNA expression and some protein analyses, despite delayed preservation post-euthanasia or prolonged storage, and 2) many additional tissues for gene expression analysis can be obtained by dissection even following prolonged storage of the tissue in situ at -80°C. These findings have relevance both to high value, ground-based experiments when sample collection capability is severely constrained, and to spaceflight experiments that entail on-orbit sample recovery by astronauts.

Cytoskeletal and functional changes in bioreactor assembled thyroid tissue organoids exposed to gamma radiation

NASA Technical Reports Server (NTRS)

Green, Lora M.; Patel, Zarana; Murray, Deborah K.; Rightnar, Steven; Burell, Cheryl G.; Gridley, Daila S.; Nelson, Gregory A.

2002-01-01

Fischer rat thyroid cells were grown under low-shear stress in a bioreactor to a stage of organization composed of integrated follicles resembling small thyroid glands prior to exposure to 3 Gray-gamma radiation. Bioreactor tissues and controls (both irradiated and non-irradiated) were harvested at 24, 48, 96 and 144 hours post-exposure. Tissue samples were fixed and fluorescently labeled for actin and microtubules. Tissues were assessed for changes in cytoskeletal components induced by radiation and quantified by laser scanning cytometry. ELISA's were used to quantify transforming growth factor-beta and thyroxin released from cells to the culture supernatant. Tissue architecture was disrupted by exposure to radiation with the structural organization of actin and loss of follicular content the most obviously affected. With time post-irradiation the actin appeared disordered and the levels of fluorescence associated with filamentous-actin and microtubules cycled in the tissue analogs, but not in the flask-grown cultures. Active transforming growth factor-beta was higher in supernatants from the irradiated bioreactor tissue. Thyroxin release paralleled cell survival in the bioreactors and control cultures. Thus, the engineered tissue responses to radiation differed from those of conventional tissue culture making it a potentially better mimic of the in vivo situation.

Robotic Prostate Biopsy in Closed MRI Scanner

DTIC Science & Technology

2009-02-01

radioactive seeds or diagnosis by harvesting tissue samples inside the mag- net bore, under remote control of the physician without mov- ing the patient out...and allows fast removal for reloading brachytherapy needles or col- lecting harvested biopsy tissue. The primary actuated motions of the robot...include two prismatic motions and two rotational motions for aligning the needle axis. In addition to these base motions, application-specific motions are

[A study on pathology and immunoglobulin of orbital tissues from cases with Grave's ophthalmopathy].

PubMed

Zhou, X; Luo, Q; Xia, R

1999-07-01

To investigate pathological changes and the expression, localization of immunoglobulins (IgA & IgE) in orbital tissues from patients with Grave's ophthalmopathy. Orbital tissues were obtained at surgery from 19 patients with Grave's ophthalmopathy and 17 control subjects. Immunohistochemical studies were carried out by using anti-IgA, anti-IgE antibody and a streptavidin-biotin-peroxidase detection system. In addition, hematoxylin-eosin, alcian blue and Masson trichrome stains were employed to demonstrate the pathologic changes in the tissues. 12 of 17 samples from patients exhibited IgA positive staining for the connective tissue associated with the extraocular muscles and orbicularis oculi, however only 2 control muscles reacted minimally. IgE positive material was found in association with the majority of leukocytes and with muscle fibers in 13 patients, but only 4 control subjects displayed mild reactions. Significant expression of IgE was more often demonstrated in patients with eye disease of short course than in those of longer course. These results support the notion that the eye muscle fiber and fibroblast are important targets of the orbital autoimmune reaction in Graves' ophthalmopathy in which IgA and IgE both play important roles.

Cardiac effects of magnesium sulfate pretreatment on acute dichlorvos-induced organophosphate poisoning: an experimental study in rats.

PubMed

Gunay, Nurullah; Kekec, Zeynep; Demiryurek, Seniz; Kose, Ataman; Namiduru, Emine S; Gunay, Nahide E; Sari, Ibrahim; Demiryurek, Abdullah T

2010-02-01

Although atropine and oximes are traditionally used in the management of organophosphate poisoning, investigations have been directed to finding additional therapeutic approaches. Thus, the aim of this study was to evaluate the cardiac effects of magnesium sulfate pretreatment on dichlorvos intoxication in rats. Rats were randomly divided into three groups as control, dichlorvos, and magnesium sulfate groups. After 6 h of dichlorvos or corn oil (as a vehicle) injection, venous blood samples were collected, and cardiac tissue samples were obtained. Biochemical analyses were performed to measure some parameters on serum and cardiac tissue. Immunohistochemical analyses of apoptosis and inducible nitric oxide (NO) synthase showed no change in cardiac tissue. Serum cholinesterase levels were markedly depressed with dichlorvos, and further suppressed markedly with magnesium sulfate pretreatment. Although we have demonstrated that serum NO levels in dichlorvos and magnesium sulfate groups were lower than the control group, cardiac tissue NO levels in magnesium sulfate group were higher than the other two groups. Mortality was not significantly affected with magnesium sulfate pretreatment. Uncertainty still persists on the right strategies for the treatment of organophosphate acute poisoning; however, it was concluded that our results do not suggest that magnesium sulfate therapy is beneficial in the management of acute dichlorvos-induced organophosphate poisoning, and also further studies are required.

A method for measuring total thiaminase activity in fish tissues

USGS Publications Warehouse

Zajicek, James L.; Tillitt, Donald E.; Honeyfield, Dale C.; Brown, Scott B.; Fitzsimons, John D.

2005-01-01

An accurate, quantitative, and rapid method for the measurement of thiaminase activity in fish samples is required to provide sufficient information to characterize the role of dietary thiaminase in the onset of thiamine deficiency in Great Lakes salmonines. A radiometric method that uses 14C-thiamine was optimized for substrate and co-substrate (nicotinic acid) concentrations, incubation time, and sample dilution. Total thiaminase activity was successfully determined in extracts of selected Great Lakes fishes and invertebrates. Samples included whole-body and selected tissues of forage fishes. Positive control material prepared from frozen alewives Alosa pseudoharengus collected in Lake Michigan enhanced the development and application of the method. The method allowed improved discrimination of thiaminolytic activity among forage fish species and their tissues. The temperature dependence of the thiaminase activity observed in crude extracts of Lake Michigan alewives followed a Q10 = 2 relationship for the 1-37??C temperature range, which is consistent with the bacterial-derived thiaminase I protein. ?? Copyright by the American Fisheries Society 2005.

Development and evaluation of a device for simultaneous uniaxial compression and optical imaging of cartilage samples in vitro

DOE Office of Scientific and Technical Information (OSTI.GOV)

Steinert, Marian; Kratz, Marita; Jones, David B.

2014-10-15

In this paper, we present a system that allows imaging of cartilage tissue via optical coherence tomography (OCT) during controlled uniaxial unconfined compression of cylindrical osteochondral cores in vitro. We describe the system design and conduct a static and dynamic performance analysis. While reference measurements yield a full scale maximum deviation of 0.14% in displacement, force can be measured with a full scale standard deviation of 1.4%. The dynamic performance evaluation indicates a high accuracy in force controlled mode up to 25 Hz, but it also reveals a strong effect of variance of sample mechanical properties on the tracking performancemore » under displacement control. In order to counterbalance these disturbances, an adaptive feed forward approach was applied which finally resulted in an improved displacement tracking accuracy up to 3 Hz. A built-in imaging probe allows on-line monitoring of the sample via OCT while being loaded in the cultivation chamber. We show that cartilage topology and defects in the tissue can be observed and demonstrate the visualization of the compression process during static mechanical loading.« less

Viscoelastic characterization of soft biological materials

NASA Astrophysics Data System (ADS)

Nayar, Vinod Timothy

Progressive and irreversible retinal diseases are among the primary causes of blindness in the United States, attacking the cells in the eye that transform environmental light into neural signals for the optic pathway. Medical implants designed to restore visual function to afflicted patients can cause mechanical stress and ultimately damage to the host tissues. Research shows that an accurate understanding of the mechanical properties of the biological tissues can reduce damage and lead to designs with improved safety and efficacy. Prior studies on the mechanical properties of biological tissues show characterization of these materials can be affected by environmental, length-scale, time, mounting, stiffness, size, viscoelastic, and methodological conditions. Using porcine sclera tissue, the effects of environmental, time, and mounting conditions are evaluated when using nanoindentation. Quasi-static tests are used to measure reduced modulus during extended exposure to phosphate-buffered saline (PBS), as well as the chemical and mechanical analysis of mounting the sample to a solid substrate using cyanoacrylate. The less destructive nature of nanoindentation tests allows for variance of tests within a single sample to be compared to the variance between samples. The results indicate that the environmental, time, and mounting conditions can be controlled for using modified nanoindentation procedures for biological samples and are in line with averages modulus values from previous studies but with increased precision. By using the quasi-static and dynamic characterization capabilities of the nanoindentation setup, the additional stiffness and viscoelastic variables are measured. Different quasi-static control methods were evaluated along with maximum load parameters and produced no significant difference in reported reduced modulus values. Dynamic characterization tests varied frequency and quasi-static load, showing that the agar could be modeled as a linearly elastic material. The effects of sample stiffness were evaluated by testing both the quasi-static and dynamic mechanical properties of different concentration agar samples, ranging from 0.5% to 5.0%. The dynamic nanoindentation protocol showed some sensitivity to sample stiffness, but characterization remained consistently applicable to soft biological materials. Comparative experiments were performed on both 0.5% and 5.0% agar as well as porcine eye tissue samples using published dynamic macrocompression standards. By comparing these new tests to those obtained with nanoindentation, the effects due to length-scale, stiffness, size, viscoelastic, and methodological conditions are evaluated. Both testing methodologies can be adapted for the environmental and mounting conditions, but the limitations of standardized macro-scale tests are explored. The factors affecting mechanical characterization of soft and thin viscoelastic biological materials are researched and a comprehensive protocol is presented. This work produces material mechanical properties for use in improving future medical implant designs on a wide variety of biological tissue and materials.

Bioaccumulation of chemical warfare agents, energetic materials, and metals in deep-sea shrimp from discarded military munitions sites off Pearl Harbor

NASA Astrophysics Data System (ADS)

Koide, Shelby; Silva, Jeff A. K.; Dupra, Vilma; Edwards, Margo

2016-06-01

The bioaccumulation of munitions-related chemicals at former military deep-water disposal sites is poorly understood. This paper presents the results of human-food-item biota sampling to assess the potential for bioaccumulation of chemical warfare agents, energetic materials, arsenic, and additional munitions-related metals in deep-sea shrimp tissue samples collected during the Hawai'i Undersea Military Munitions Assessment (HUMMA) project to date. The HUMMA investigation area is located within a former munitions sea-disposal site located south of Pearl Harbor on the island of O'ahu, Hawai'i, designated site Hawaii-05 (HI-05) by the United States Department of Defense. Indigenous deep-sea shrimp (Heterocarpus ensifer) were caught adjacent to discarded military munitions (DMM) and at control sites where munitions were absent. Tissue analysis results showed that chemical warfare agents and their degradation products were not present within the edible portions of these samples at detectable concentrations, and energetic materials and their degradation products were detected in only a few samples at concentrations below the laboratory reporting limits. Likewise, arsenic, copper, and lead concentrations were below the United States Food and Drug Administration's permitted concentrations of metals in marine biota tissue (if defined), and their presence within these samples could not be attributed to the presence of DMM within the study area based on a comparative analysis of munitions-adjacent and control samples collected. Based on this current dataset, it can be concluded that DMM existing within the HUMMA study area is not contributing to the bioaccumulation of munitions-related chemicals for the biota species investigated to date.

NASA Bioreactor tissue culture

NASA Technical Reports Server (NTRS)

1998-01-01

Dr. Lisa E. Freed of the Massachusetts Institute of Technology and her colleagues have reported that initially disc-like specimens tend to become spherical in space, demonstrating that tissues can grow and differentiate into distinct structures in microgravity. The Mir Increment 3 (Sept. 16, 1996 - Jan. 22, 1997) samples were smaller, more spherical, and mechanically weaker than Earth-grown control samples. These results demonstrate the feasibility of microgravity tissue engineering and may have implications for long human space voyages and for treating musculoskeletal disorders on earth. The work is sponsored by NASA's Office of Biological and Physical Research. The bioreactor is managed by the Biotechnology Cell Science Program at NASA's Johnson Space Center (JSC). NASA-sponsored bioreactor research has been instrumental in helping scientists to better understand normal and cancerous tissue development. In cooperation with the medical community, the bioreactor design is being used to prepare better models of human colon, prostate, breast and ovarian tumors. Cartilage, bone marrow, heart muscle, skeletal muscle, pancreatic islet cells, liver and kidney are just a few of the normal tissues being cultured in rotating bioreactors by investigators.

Frozen and fresh ovarian tissue require different culture media to promote in vitro development of bovine preantral follicles.

PubMed

Castro, Simone Vieira; Carvalho, Adeline Andrade; Silva, Cleidson Manoel Gomes; Santos, Francielli Weber; Campello, Cláudio Cabral; de Figueiredo, José Ricardo; Rodrigues, Ana Paula Ribeiro

2014-10-01

The aim of this study was to evaluate the efficiency of different media in the in vitro culture of bovine preantral follicles that were used either fresh or following slow freezing treatment. Frozen and fresh noncultured or cultured ovarian fragments were processed for histological, viability, and cell proliferation analyses. For cryopreservation, a solution containing 1.5 M ethylene glycol was frozen in a programmable biological freezer. After thawing, a portion of the samples was destined for frozen controls. The remainder were cultured in vitro for 5 days in three media: α-MEM, McCoy, or M199. Samples from these culture media were collected on days 1 and 5 for quantification of reactive oxygen species (ROS) and for hormonal assays. In fresh-cultured tissues, the percentage of morphologically normal follicles was significantly higher when cultured in M199 compared to that in the other media. In frozen-cultured tissues, McCoy medium was significantly superior to the other media, and was the only treatment that helped in maintaining the viability similar to fresh and frozen controls. Upon quantification of the nucleolus organizer region, we observed greater proliferation of granulosa cells in the frozen-cultured tissues with McCoy medium, and lesser proliferation in fresh-cultured tissues only with α-MEM. In frozen-cultured tissues, ROS levels were highest at day 1 and progressively reduced during culture, independent of the media used. In conclusion, under the conditions used in this study, the M199 and McCoy media are recommended for the culture of follicles derived from fresh and frozen ovarian tissues, respectively.

Evaluating the efficacy of osteopontin expression as a prognostic marker in oral squamous cell carcinoma in the Indian subpopulation.

PubMed

Ingale, Yashwant; Routray, Samapika; Kheur, Supriya M; Kheur, Mohit; Mohanty, Neeta

2014-09-01

This study aimed to correlate the prognostic value of osteopontin (OPN) expression using both tissue and plasma samples from patients with clinically and histologically confirmed oral squamous cell carcinoma (OSCC). The study group comprised of sixty patients (n = 60), which were clinically and histologically diagnosed for oral squamous cell carcinoma (OSCC). The Control group comprised of ten (n = 10) healthy volunteers. Plasma OPN levels were assayed using a quantitative enzyme-linked immunosorbent assay (OPN ELISA). Expression of OPN was also identified and evaluated by immunohistochemistry in tissue sections. These OPN expressions were then correlated with different parameters like age, sex, site, clinical presentation, tumor node metastasis (TNM) staging, histopathological grading and lymph node metastasis. One-way analysis of variance (ANOVA) was used to evaluate the difference in tissue intensity and plasma OPN levels between the OSCC and the normal control groups. The distribution of the plasma OPN levels and tissue OPN intensity in OSCC cohorts were compared to histopathological grades and analyzed. When evaluated OPN expression in tissue had higher intensity observed in OSCC (95% +ve) cases. And the mean plasma OPN concentration in OSCC cohort was more in comparison to the normal cohort. The results clearly showed that the plasma OPN levels and intensity grading in tissue correlated with tumor grades. The study highlights OPN as a biomarker for prognosis in OSCC in both plasma and tissue samples. We would like to emphasize on the evaluation of plasma OPN as a protocol of blood examination for all cancer patient, as it may serve as an indicator for tumor progression and potential risk of metastasis.

Immunohistochemical and Biochemical Expression Patterns of TTF-1, RAGE, GLUT-1 and SOX2 in HCV-Associated Hepatocellular Carcinomas

PubMed Central

Aboushousha, Tarek; Mamdouh, Samah; Hamdy, Hussam; Helal, Noha; Khorshed, Fatma; Safwat, Gehan; Seleem, Mohamed

2018-01-01

Objective: To investigate the expression of TTF-1, RAGE, GLUT1 and SOX2 in HCV-associated HCCs and in surrounding non-tumorous liver tissue. Material and Methods: Tissue material from partial hepatectomy cases for HCC along with corresponding serum samples and 30 control serum samples from healthy volunteers were studied. Biopsies were classified into: non-tumor hepatic tissue (36 sections); HCC (33 sections) and liver cell dysplasia (LCD) (15 sections). All cases were positive for HCV. Immunohistochemistry (IHC), gene extraction and quantitative real-time reverse-transcription assays (qRT-PCR) were applied. Results: By IHC, LCD and HCC showed significantly high percentages of positive cases with all markers. SOX2 showed significant increase with higher HCC grades, while RAGE demonstrated an inverse relation and GLUT-1 and TTF-1 lacked any correlation. In nontumorous-HCV tissue, we found significantly high TTF-1, low RAGE and negative SOX2 expression. RAGE, GLUT-1 and SOX2 show non-significant elevation positivity in high grade HCV compared to low grade lesions. TTF-1, RAGE and SOX2 exhibited low expression in cirrhosis compared to fibrosis. Biochemical studies on serum and tissue extracts revealed significant down-regulation of RAGE, GLUT-1 and SOX2 genes, as well as significant up-regulation of the TTF-1 gene in HCC cases compared to controls. All studied genes show significant correlation with HCC grade. In non-tumor tissue, only TTF-1 gene expression had a significant correlation with the fibrosis score. Conclusion: Higher expression of TTF-1, RAGE, GLUT-1 and SOX2 in HCC and dysplasia compared to non-tumor tissues indicates up-regulation of these markers as early events during the development of HCV-associated HCC. PMID:29373917

Immunohistochemical and Biochemical Expression Patterns of TTF-1, RAGE, GLUT-1 and SOX2 in HCV-Associated Hepatocellular Carcinomas

PubMed

Aboushousha, Tarek; Mamdouh, Samah; Hamdy, Hussam; Helal, Noha; Khorshed, Fatma; Safwat, Gehan; Seleem, Mohamed

2018-01-27

Objective: To investigate the expression of TTF-1, RAGE, GLUT1 and SOX2 in HCV-associated HCCs and in surrounding non-tumorous liver tissue. Material and Methods: Tissue material from partial hepatectomy cases for HCC along with corresponding serum samples and 30 control serum samples from healthy volunteers were studied. Biopsies were classified into: non-tumor hepatic tissue (36 sections); HCC (33 sections) and liver cell dysplasia (LCD) (15 sections). All cases were positive for HCV. Immunohistochemistry (IHC), gene extraction and quantitative real-time reverse-transcription assays (qRT-PCR) were applied. Results: By IHC, LCD and HCC showed significantly high percentages of positive cases with all markers. SOX2 showed significant increase with higher HCC grades, while RAGE demonstrated an inverse relation and GLUT-1 and TTF-1 lacked any correlation. In nontumorous-HCV tissue, we found significantly high TTF-1, low RAGE and negative SOX2 expression. RAGE, GLUT-1 and SOX2 show non-significant elevation positivity in high grade HCV compared to low grade lesions. TTF-1, RAGE and SOX2 exhibited low expression in cirrhosis compared to fibrosis. Biochemical studies on serum and tissue extracts revealed significant down-regulation of RAGE, GLUT-1 and SOX2 genes, as well as significant up-regulation of the TTF-1 gene in HCC cases compared to controls. All studied genes show significant correlation with HCC grade. In non-tumor tissue, only TTF-1 gene expression had a significant correlation with the fibrosis score. Conclusion: Higher expression of TTF-1, RAGE, GLUT-1 and SOX2 in HCC and dysplasia compared to non-tumor tissues indicates up-regulation of these markers as early events during the development of HCV-associated HCC. Creative Commons Attribution License

A PEG-Based Hydrogel for Effective Wound Care Management

PubMed Central

Chen, Sen-Lu; Fu, Ru-Huei; Liao, Shih-Fei; Liu, Shih-Ping; Lin, Shinn-Zong; Wang, Yu-Chi

2018-01-01

It is extremely challenging to achieve strong adhesion in soft tissues while minimizing toxicity, tissue damage, and other side effects caused by wound sealing materials. In this study, flexible synthetic hydrogel sealants were prepared based on polyethylene glycol (PEG) materials. PEG is a synthetic material that is nontoxic and inert and, thus, suitable for use in medical products. We evaluated the in vitro biocompatibility tests of the dressings to assess cytotoxicity and irritation, sensitization, pyrogen toxicity, and systemic toxicity following the International Organization for Standardization 10993 standards and the in vivo effects of the hydrogel samples using Coloskin liquid bandages as control samples for potential in wound closure. PMID:29637814

Fetal Tissues Tested for Microbial Sterility by Culture- and PCR-Based Methods Can be Safely Used in Clinics.

PubMed

Vitrenko, Yakov; Kostenko, Iryna; Kulebyakina, Kateryna; Duda, Alla; Klunnyk, Mariya; Sorochynska, Khrystyna

2017-02-16

Cell preparations to be used in clinical practice must be free of infectious agents. Safety concerns are especially elevated upon the use of human fetal tissues, which are otherwise highly advantageous in cell therapy. We demonstrate that treating fetal samples with antibiotic, extensive washing, and homogenization prior to cryoconservation efficiently removes microbes in general. Screening a large collection by an automatic culture system showed that 89.2% fetal tissue samples were sterile, while contamination was detected in 10.8% samples. Liver and chorion were contaminated more than the brain, kidney, lung, and soft tissues. Broad-range PCR from the bacterial 16s rRNA gene was adopted as a confirmatory assay; however, the concordance between the culture-based and PCR assays was weak. Taxonomic identification was done for contaminated samples by bacteriological methods and sequencing 16s rRNA PCR products. The two approaches revealed different spectra of taxonomic groups sharing only Lactobacillus, the most frequently found genus. In addition, other representatives of vaginal microbiota were detected by culture-based identification, while PCR product sequencing has also revealed a subset of nosocomial microorganisms. Importantly, species known to cause sepsis were identified by both techniques, arguing for their indispensability and mutual complementarity. We suggest that most contaminations are taken up during collection of fetal material rather than originating from an in utero infection. In conclusion, a rigorous microbiological control by culture and PCR is a prerequisite for safe clinical use of fetal tissue suspensions.

Transcriptome profiling of Kentucky bluegrass (Poa pratensis L.) accessions in response to salt stress.

PubMed

Bushman, B Shaun; Amundsen, Keenan L; Warnke, Scott E; Robins, Joseph G; Johnson, Paul G

2016-01-13

Kentucky bluegrass (Poa pratensis L.) is a prominent turfgrass in the cool-season regions, but it is sensitive to salt stress. Previously, a relatively salt tolerant Kentucky bluegrass accession was identified that maintained green colour under consistent salt applications. In this study, a transcriptome study between the tolerant (PI 372742) accession and a salt susceptible (PI 368233) accession was conducted, under control and salt treatments, and in shoot and root tissues. Sample replicates grouped tightly by tissue and treatment, and fewer differentially expressed transcripts were detected in the tolerant PI 372742 samples compared to the susceptible PI 368233 samples, and in root tissues compared to shoot tissues. A de novo assembly resulted in 388,764 transcripts, with 36,587 detected as differentially expressed. Approximately 75 % of transcripts had homology based annotations, with several differences in GO terms enriched between the PI 368233 and PI 372742 samples. Gene expression profiling identified salt-responsive gene families that were consistently down-regulated in PI 372742 and unlikely to contribute to salt tolerance in Kentucky bluegrass. Gene expression profiling also identified sets of transcripts relating to transcription factors, ion and water transport genes, and oxidation-reduction process genes with likely roles in salt tolerance. The transcript assembly represents the first such assembly in the highly polyploidy, facultative apomictic Kentucky bluegrass. The transcripts identified provide genetic information on how this plant responds to and tolerates salt stress in both shoot and root tissues, and can be used for further genetic testing and introgression.

Alkyl Phenols and Diethylhexyl Phthalate in Tissues of Sheep Grazing Pastures Fertilized with Sewage Sludge or Inorganic Fertilizer

PubMed Central

Rhind, Stewart M.; Kyle, Carol E.; Telfer, Gillian; Duff, Elizabeth I.; Smith, Alistair

2005-01-01

We studied selected tissues from ewes and their lambs that were grazing pastures fertilized with either sewage sludge (treated) or inorganic fertilizer (control) and determined concentrations of alkylphenols and phthalates in these tissues. Mean tissue concentrations of alkylphenols were relatively low (< 10–400 μg/kg) in all animals and tissues. Phthalates were detected in tissues of both control and treated animals at relatively high concentrations (> 20,000 μg/kg in many tissue samples). The use of sludge as a fertilizer was not associated with consistently increased concentrations of either alkylphenols or phthalates in the tissues of animals grazing treated pastures relative to levels in control animal tissues. Concentrations of the two classes of chemicals differed but were of a similar order of magnitude in liver and muscle as well as in fat. Concentrations of each class of compound were broadly similar in tissues derived from ewes and lambs. Although there were significant differences (p < 0.01 or p < 0.001) between years (cohorts) in mean tissue concentrations of both nonylphenol (NP) and phthalate in each of the tissues from both ewes and lambs, the differences were not attributable to either the age (6 months or 5 years) of the animal or the duration of exposure to treatments. Octylphenol concentrations were generally undetectable. There was no consistent cumulative outcome of prolonged exposure on the tissue concentrations of either class of pollutant in any ewe tissue. Mean tissue concentrations of phthalate were higher (p < 0.001) in the liver and kidney fat of male compared with female lambs. We suggest that the addition of sewage sludge to pasture is unlikely to cause large increases in tissue concentrations of NP and phthalates in sheep and other animals with broadly similar diets and digestive systems (i.e., domestic ruminants) grazing such pasture. PMID:15811823

Liquid Microjunction Surface Sampling Coupled with High-Pressure Liquid Chromatography-Electrospray Ionization-Mass Spectrometry for Analysis of Drugs and Metabolites in Whole-Body Thin Tissue Sections

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kertesz, Vilmos; Van Berkel, Gary J

2010-01-01

In this work, a commercially available autosampler was adapted to perform direct liquid microjunction (LMJ) surface sampling followed by a high-pressure liquid chromatography (HPLC) separation of the extract components and detection with electrospray ionization mass spectrometry (ESI-MS). To illustrate the utility of coupling a separation with this direct liquid extraction based surface sampling approach, four different organs (brain, lung, kidney, and liver) from whole-body thin tissue sections of propranolol dosed and control mice were examined. The parent drug was observed in the chromatograms of the surface sampling extracts from all the organs of the dosed mouse examined. In addition, twomore » isomeric phase II metabolites of propranolol (an aliphatic and an aromatic hydroxypropranolol glucuronide) were observed in the chromatograms of the extracts from lung, kidney, and liver. Confirming the presence of one or the other or both of these glucuronides in the extract from the various organs was not possible without the separation. These drug and metabolite data obtained using the LMJ surface sampling/HPLC-MS method and the results achieved by analyzing similar samples by conventional extraction of the tissues and subsequent HPLC-MS analysis were consistent.« less

MLF1 interacting protein: a potential gene therapy target for human prostate cancer?

PubMed

Zhang, Lei; Ji, Guoqing; Shao, Yuzhang; Qiao, Shaoyi; Jing, Yuming; Qin, Rongliang; Sun, Huiming; Shao, Chen

2015-02-01

Here, we investigated the role of one gene that has been previously associated with human prostate carcinoma cells-myelodysplasia/myeloid leukemia factor 1 interacting protein (MLF1IP)-in order to better ascertain its role in human prostate carcinogenesis. The prostate cancer cell line PC-3 was lentivirally transfected to silence endogenous MLF1IP gene expression, which was confirmed by real-time quantitative PCR (RT-qPCR). Cellomics ArrayScan VTI imaging and MTT assays were conducted to assess cell proliferation. Cell cycle phase arrest and apoptosis were assayed by flow cytometry. Colony formation was assessed by fluorescence microscopy. MLF1IP gene expression was also analyzed by RT-qPCR in sixteen prostate cancer tissue samples and six healthy control prostate tissue samples from human patients. Cell proliferation was significantly inhibited in MLF1IP-silenced cells relative to control cells. G1 phase, S and G2/M phase cell counts were not significantly changed in MLF1IP-silenced cells relative to control cells. Apoptosis was significantly increased in MLF1IP-silenced cells, while MLF1IP-silenced cells displayed a significantly reduced number of cell colonies, compared to control cells. The 16 human prostate cancer tissue samples revealed no clear upregulation or downregulation in MLF1IP gene expression. MLF1IP significantly promotes prostate cancer cell proliferation and colony formation and significantly inhibits apoptosis without affecting cell cycle phase arrest. Further study is required to conclusively determine whether MLF1IP is upregulated in human prostate cancer tumors and to determine the precise cellular mechanism(s) for MLF1IP in prostate carcinogenesis.

Comparison of candidate vCJD in vitro diagnostic assays using identical sample sets.

PubMed

Cooper, J K; Ladhani, K; Minor, P

2012-02-01

With four transfusion related transmissions of variant Creutzfeldt-Jakob Disease (vCJD), three of which developed clinical disease and the other died of other causes but was positive for markers of infection, there is an increased urgency to identify and implement a test for blood donor screening. With limited amounts of blood samples from vCJD cases available test evaluation is challenging. Alternative approaches are therefore needed. Control and vCJD tissues homogenates, where levels of markers of infectivity are known, were sequentially diluted in pooled human plasma. Identical sets of samples were provided blind to research groups developing diagnostic tests for vCJD; identical sample sets allows for direct comparisons of sensitivity to be made. Control and vCJD tissue homogenates were sequentially diluted in pooled human plasma (detergent solvent treated or cryo-depleted) supplied by commercial fractionators. Dilutions of vCJD tissues were within and beyond the limits of detection previously determined by the conformation-dependent immunoassay (Cooper et al.: Vox Sang 2007;92:302-310; Bellon et al.: J Gen Virol 2003;84: 1921-1925). A number of methods were used for the analysis of the blinded panels; with background signal from the normal prion protein (PrP) being removed by digestion with proteinase, epitope protection or selective capture of PrP(tse). Assay sensitivities were directly compared using identical sample sets. This approach identified several transmissible spongiform encephalopathies (TSE) diagnostic tests, based on different principles, high in analytical sensitivity that reproducibly detected markers of vCJD infectivity in tissue homogenates. The approach outlined has successfully compared in vitro diagnostics assays for their sensitivity and reproducibility and is a first step toward the evaluation of an assay suitable for blood donor screening/diagnosis of vCJD. © 2011 The Author(s). Vox Sanguinis © 2011 International Society of Blood Transfusion.

Polonium-210 in the environment around a radioactive waste disposal area and phosphate ore processing plant.

PubMed

Arthur, W J; Markham, O D

1984-04-01

Polonium-210 concentrations were determined for soil, vegetation and small mammal tissues collected at a solid radioactive waste disposal area, near a phosphate ore processing plant and at two rural areas in southeastern Idaho. Polonium concentrations in media sampled near the radioactive waste disposal facility were equal to or less than values from rural area samples, indicating that disposal of solid radioactive waste at the Idaho National Engineering Laboratory Site has not resulted in increased environmental levels of polonium. Concentrations of 210Po in soils, deer mice hide and carcass samples collected near the phosphate processing plant were statistically (P less than or equal to 0.05) greater than the other sampling locations; however, the mean 210Po concentration in soils and small mammal tissues from sampling areas near the phosphate plant were only four and three times greater, respectively, than control values. No statistical (P greater than 0.05) difference was observed for 210Po concentrations in vegetation among any of the sampling locations.

Method And Apparatus For Examining A Tissue Using The Spectral Wing Emission Therefrom Induced By Visible To Infrared Photoexcitation.

DOEpatents

Alfano, Robert R.; Demos, Stavros G.; Zhang, Gang

2003-12-16

Method and an apparatus for examining a tissue using the spectral wing emission therefrom induced by visible to infrared photoexcitation. In one aspect, the method is used to characterize the condition of a tissue sample and comprises the steps of (a) photoexciting the tissue sample with substantially monochromatic light having a wavelength of at least 600 nm; and (b) using the resultant far red and near infrared spectral wing emission (SW) emitted from the tissue sample to characterize the condition of the tissue sample. In one embodiment, the substantially monochromatic photoexciting light is a continuous beam of light, and the resultant steady-state far red and near infrared SW emission from the tissue sample is used to characterize the condition of the tissue sample. In another embodiment, the substantially monochromatic photoexciting light is a light pulse, and the resultant time-resolved far red and near infrared SW emission emitted from the tissue sample is used to characterize the condition of the tissue sample. In still another embodiment, the substantially monochromatic photoexciting light is a polarized light pulse, and the parallel and perpendicular components of the resultant polarized time-resolved SW emission emitted from the tissue sample are used to characterize the condition of the tissue sample.

Application of allflex conservation buffer in illumina genotyping.

PubMed

de Groot, M; Ras, T; van Haeringen, W A

2016-12-01

This experiment was designed to study if liquid conservation buffer used in the novel Tissue Sampling Technology (TST) from Allflex can be used for Illumina BeadChip genotyping. Ear punches were collected from 6 bovine samples, using both the Tissue Sampling Unit (TSU) as well as the Total Tagger Universal (TTU) collection system. The stability of the liquid conservation buffer was tested by genotyping samples on Illumina BeadChips, incubated at 0, 3, 15, 24, 48, 72, 168, 336, 720 h after sample collection. Additionally, a replenishment study was designed to test how often the liquid conservation buffer could be completely replenished before a significant call rate drop could be observed. Results from the stability study showed an average call rate of 0.993 for samples collected with the TSU system and 0.953 for samples collected with the TTU system, both exceeding the inclusion threshold call rate of 0.85. As an additional control, the identity of the individual animals was confirmed using the International Society of Animal Genetics (ISAG) recommended SNP panel. The replenishment study revealed a slight drop in the sample call rate after replenishing the conservation buffer for the fourth time for the TSU as well as the TTU samples. In routine analysis, this application allows for multiple experiments to be performed on the liquid conservation buffer, while maintaining the tissue samples for future use. The data collected in this study shows that the liquid conservation buffer used in the TST system can be used for Illumina BeadChip genotyping applications.

Growth characteristics of fibroblasts isolated from the trunk and distal aspect of the limb of horses and ponies.

PubMed

Miller, C B; Wilson, D A; Keegan, K G; Kreeger, J M; Adelstein, E H; Ganjam, V K

2000-01-01

To determine if there is a difference in in vitro growth of fibroblasts isolated from the trunk and distal aspect of the limb of horses and ponies. To determine the effects of a corticosteroid and monokine on in vitro growth of fibroblasts isolated from the trunk and distal aspect of the limb of horses and ponies. Growth of fibroblasts from tissues harvested from the trunk and limb were compared from horse and pony samples grown in control media and control media with triamcinolone or monokine added. Dermal and subcutaneous tissue from 22 horses and 17 ponies of various ages and breeds. Fibroblast growth was assessed by tritiated thymidine uptake using standard cell culture techniques. The effect of a monokine or triamcinolone plus control media were compared with control media for fibroblast growth. Fibroblast growth from tissues isolated from the horse limb was significantly less than growth from the horse trunk and the limb and trunk of ponies. Monokine was more effective than triamcinolone in suppressing fibroblast growth from tissues isolated from the trunk and limb in both horses and ponies. There are growth differences in fibroblasts isolated from the limb of horses compared with those isolated from the trunk and from the limb and trunk of ponies. The difference in fibroblast growth from tissues isolated from the trunk and limb of horses and ponies may provide evidence for the difference reported in the healing characteristics of limb wounds in horses and ponies. Influencing fibroblast growth may provide a key to controlling the development of exuberant granulation tissue in horses and ponies.

Biological evaluation of partially stabilized zirconia added HA/HDPE composites with osteoblast and fibroblast cell lines.

PubMed

Yari Sadi, Amir; Shokrgozar, Mohammad Ali; Homaeigohar, Seyed Shahin; Khavandi, Alireza

2008-06-01

In the present study, the biocompatibility of partially stabilized zirconia (PSZ) added hydroxyapatite (HA)--high density polyethylene (HDPE) composites was evaluated by proliferation and cell attachment assays on two osteoblast cell lines (G-292, Saos-2) and a type of fibroblast cell isolated from bone tissue namely HBF in different time intervals. Cell-material interactions on the surface of the composites were observed by scanning electron microscopy (SEM). The effect of composites on the behavior of osteoblast and fibroblast cells was compared with those of HDPE and Tissue Culture Poly Styrene (TPS) (as negative control) samples. Results showed that the composite samples supported a higher proliferation rate of osteoblast cells in the presence of composite samples as compared to the HDPE and TPS samples after 3, 7 and 14 days of incubation period. It was showed that an equal or in some cases an even higher proliferation rate of G-292 and Saos-2 osteoblast cells on composite samples in compare to negative controls in culture period (P < 0.05). The number of adhered cells on the composite samples was equal and in some cases higher than the number adhered on the HDPE and TPS samples after the above mentioned incubation periods (P < 0.05). Adhered cells presented a normal morphology by SEM and many of the cells were seen to be undergoing cell division.

Myocardial concentrations of fatty acids in dogs with dilated cardiomyopathy.

PubMed

Smith, Caren E; Freeman, Lisa M; Meydani, Mohsen; Rush, John E

2005-09-01

To compare myocardial concentrations of fatty acids in dogs with dilated cardiomyopathy (DCM) with concentrations in control dogs. Myocardial tissues from 7 dogs with DCM and 16 control dogs. Myocardial tissues were homogenized, and total fatty acids were extracted and converted to methyl esters. Myocardial concentrations of fatty acids were analyzed by use of gas chromatography and reported as corrected percentages. The amount of docosatetraenoic acid (C22:4 n-6) was significantly higher in myocardial samples from dogs with DCM (range, 0.223% to 0.774%; median, 0.451%), compared with the amount in samples obtained from control dogs (range, 0.166% to 0.621%; median, 0.280%). There were no significant differences between DCM and control dogs for concentrations of any other myocardial fatty acids. Although concentrations of most myocardial fatty acids did not differ significantly between dogs with DCM and control dogs, the concentration of docosatetraenoic acid was significantly higher in dogs with DCM. Additional investigation in a larger population is warranted to determine whether this is a primary or secondary effect of the underlying disease and whether alterations in fatty acids may be a target for intervention in dogs with DCM.

Complete validation of a unique digestion assay to detect Trichinella larvae in horse meat demonstrates the reliability of this assay for meeting food safety and trade requirements.

PubMed

Forbes, L B; Hill, D E; Parker, S; Tessaro, S V; Gamble, H R; Gajadhar, A A

2008-03-01

A tissue digestion assay using a double separatory funnel procedure for the detection of Trichinella larvae in horse meat was validated for application in food safety programs and trade. The assay consisted of a pepsin-HCl digestion step to release larvae from muscle tissue and two sequential sedimentation steps in separatory funnels to recover and concentrate larvae for detection with a stereomicroscope. With defined critical control points, the assay was conducted within a quality assurance system compliant with International Organization for Standardization-International Electrotechnical Commission (ISO/IEC) 17025 guidelines. Samples used in the validation were obtained from horses experimentally infected with Trichinella spiralis to obtain a range of muscle larvae densities. One-, 5-, and 10-g samples of infected tissue were combined with 99, 95, and 90 g, respectively, of known negative horse tissue to create a 100-g sample for testing. Samples of 5 and 10 g were more likely to be positive than were 1-g samples when larval densities were less than three larvae per gram (lpg). This difference is important because ingested meat with 1 lpg is considered the threshold for clinical disease in humans. Using a 5-g sample size, all samples containing 1.3 to 2 lpg were detected, and 60 to 100% of samples with infected horse meat containing 0.1 to 0.7 lpg were detected. In this study, the double separatory funnel digestion assay was efficient and reliable for its intended use in food safety and trade. This procedure is the only digestion assay for Trichinella in horse meat that has been validated as consistent and effective at critical levels of sensitivity.

Human herpesvirus multiplex ddPCR detection in brain tissue from low- and high-grade astrocytoma cases and controls.

PubMed

Lin, Cheng-Te Major; Leibovitch, Emily C; Almira-Suarez, M Isabel; Jacobson, Steven

2016-01-01

Glioblastoma (GBM) is a fatal CNS malignancy, representing 50 % of all gliomas with approximately 12-18 months survival time after initial diagnosis. Recently, the human herpesvirus cytomegalovirus (CMV) has been suggested to have an oncogenic role, yet this association remains controversial. In addition, human herpesvirus 6 (HHV-6) and Epstein-Barr virus (EBV) have also been associated with low-grade gliomas, but few studies have examined HHV-6 and EBV in glioblastomas. Droplet digital PCR (ddPCR) is a highly precise diagnostic tool that enables the absolute quantification of target DNA. This study examines the association between multiple human herpesviruses and astrocytomas. This study analyzed 112 brain tissue specimens, including 45 glioblastoma, 12 astrocytoma grade III, 2 astrocytoma grade II, 4 astrocytoma grade I, and 49 controls. All brain tissue samples were de-identified and pathologically confirmed. Each tissue block was sectioned for DNA extraction and CMV, EBV, HHV-6A and HHV-6B, and a cellular housekeeping gene were amplified by ddPCR. Neither CMV nor HHV-6A were detected in any of the astrocytoma samples. However, HHV-6B (p = 0.147) and EBV (p = 0.049) had a higher positivity frequency in the GBM compared to the controls. The undetectable CMV DNA in the astrocytoma cohort does not support the observation of an increased prevalence of CMV DNA in GBM, as reported in other studies. EBV has a significantly higher positivity in the GBM cohort compared to the controls, while HHV-6B has a higher but not statistically significant positivity in the case cohort. Whether these viruses play an oncogenic role in GBM remains to be further investigated.

Proteoglycan depletion and size reduction in lesions of early grade chondromalacia of the patella.

PubMed Central

Väätäinen, U; Häkkinen, T; Kiviranta, I; Jaroma, H; Inkinen, R; Tammi, M

1995-01-01

OBJECTIVE--To determine the content and molecular size of proteoglycans (PGs) in patellar chondromalacia (CM) and control cartilages as a first step in investigating the role of matrix alterations in the pathogenesis of this disease. METHODS--Chondromalacia tissue from 10 patients was removed with a surgical knife. Using identical techniques, apparently healthy cartilage of the same site was obtained from 10 age matched cadavers (mean age 31 years in both groups). Additional pathological cartilage was collected from 67 patients with grades II-IV CM (classified according to Outerbridge) using a motorised shaver under arthroscopic control. The shaved cartilage chips were collected with a dense net from the irrigation fluid of the shaver. The content of tissue PGs was determined by Safranin O precipitation or uronic acid content, and the molecular size by mobility on agarose gel electrophoresis. RESULTS--The mean PG content of the CM tissue samples with a knife was dramatically reduced, being only 15% of that in controls. The cartilage chips collected from shaving operations of grades II, III, and IV CM showed a decreasing PG content: 9%, 5%, and 1% of controls, respectively. Electrophoretic analysis of PGs extracted with guanidium chloride from the shaved tissue samples suggested a significantly reduced size of aggrecans in the mild (grade II) lesions. CONCLUSION--These data show that there is already a dramatic and progressive depletion of PGs in CM grade II lesions. This explains the softening of cartilage, a typical finding in the arthroscopic examination of CM. The PG size reduction observed in grade II implicates proteolytic attack as a factor in the pathogenesis of CM. Images PMID:7492223

Aromatase Blockade Is Associated With Increased Mortality in Acute Illness in Male Mice.

PubMed

Connerney, Jeannette J; Spratt, Daniel I

2017-09-01

The increase in circulating estrogen levels with acute illness in humans is accompanied by increased aromatase expression in adipose tissue and increased peripheral aromatization of estrogens to androgens. Animal studies indicate that estrogen may be beneficial in acute illness. We hypothesized that blockade of aromatase in acute illness would decrease survival. Prospective sham controlled. Maine Medical Center Research Institute animal facility. Six- to 8-week-old male black 6 mice. Mice underwent cecal ligation and puncture (CLP) to induce acute illness and were administered letrozole to block aromatase or saline. Mice undergoing sham surgery with or without letrozole served as controls. Adipose and cardiovascular tissue was harvested for preliminary evaluation of aromatase expression. Survival was the main outcome measurement. Evidence for aromatase expression in tissue samples was assessed using western blot and/or immunohistochemistry. With aromatase blockade, survival in CLP mice was decreased ( P = 0.04). The presence of aromatase in adipose tissue was observed by western blot in CLP but not control mice. Similarly, the presence of aromatase was observed in cardiac tissue of CLP but not in control mice. The decreased survival during sepsis with aromatase blockade suggests that this response to acute illness may be important both physiologically and clinically. The preliminary observation of aromatase expression in adipose and cardiovascular tissue during acute illness in this mouse model indicates that this model has parallels to human physiology and may be useful for further studying the aromatase response to acute illness.

Wound and soft tissue infections of Serratia marcescens in patients receiving wound care: A health care-associated outbreak.

PubMed

Us, Ebru; Kutlu, Huseyin H; Tekeli, Alper; Ocal, Duygu; Cirpan, Sevilay; Memikoglu, Kemal O

2017-04-01

We described a health care-associated Serratia marcescens outbreak of wound and soft tissue infection lasting approximately 11 months at Ankara University Ibni Sina Hospital. After identification of S marcescens strains from the clinical and environmental samples, and their susceptibility testing to antimicrobial agents, pulsed-field gel electrophoresis (PFGE) was performed to detect molecular epidemiologic relationships among these isolates. The strains which were isolated from the saline bottles used for wound cleansing in the wound care unit were found to be 100% interrelated by PFGE to the strains from the samples of the outbreak patients. Reuse of the emptied bottles has no longer been allowed since the outbreak occurred. Besides, more efficient and frequent infection control training for hospital staff has been conducted. Copyright © 2017 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Elsevier Inc. All rights reserved.

Tissue reservoirs of antiviral T cell immunity in persistent human CMV infection

PubMed Central

Gordon, Claire L.; Thome, Joseph J.C.; Igarashi, Suzu

2017-01-01

T cell responses to viruses are initiated and maintained in tissue sites; however, knowledge of human antiviral T cells is largely derived from blood. Cytomegalovirus (CMV) persists in most humans, requires T cell immunity to control, yet tissue immune responses remain undefined. Here, we investigated human CMV-specific T cells, virus persistence and CMV-associated T cell homeostasis in blood, lymphoid, mucosal and secretory tissues of 44 CMV seropositive and 28 seronegative donors. CMV-specific T cells were maintained in distinct distribution patterns, highest in blood, bone marrow (BM), or lymph nodes (LN), with the frequency and function in blood distinct from tissues. CMV genomes were detected predominantly in lung and also in spleen, BM, blood and LN. High frequencies of activated CMV-specific T cells were found in blood and BM samples with low virus detection, whereas in lung, CMV-specific T cells were present along with detectable virus. In LNs, CMV-specific T cells exhibited quiescent phenotypes independent of virus. Overall, T cell differentiation was enhanced in sites of viral persistence with age. Together, our results suggest tissue T cell reservoirs for CMV control shaped by both viral and tissue-intrinsic factors, with global effects on homeostasis of tissue T cells over the lifespan. PMID:28130404

Tissue reservoirs of antiviral T cell immunity in persistent human CMV infection.

PubMed

Gordon, Claire L; Miron, Michelle; Thome, Joseph J C; Matsuoka, Nobuhide; Weiner, Joshua; Rak, Michael A; Igarashi, Suzu; Granot, Tomer; Lerner, Harvey; Goodrum, Felicia; Farber, Donna L

2017-03-06

T cell responses to viruses are initiated and maintained in tissue sites; however, knowledge of human antiviral T cells is largely derived from blood. Cytomegalovirus (CMV) persists in most humans, requires T cell immunity to control, yet tissue immune responses remain undefined. Here, we investigated human CMV-specific T cells, virus persistence and CMV-associated T cell homeostasis in blood, lymphoid, mucosal and secretory tissues of 44 CMV seropositive and 28 seronegative donors. CMV-specific T cells were maintained in distinct distribution patterns, highest in blood, bone marrow (BM), or lymph nodes (LN), with the frequency and function in blood distinct from tissues. CMV genomes were detected predominantly in lung and also in spleen, BM, blood and LN. High frequencies of activated CMV-specific T cells were found in blood and BM samples with low virus detection, whereas in lung, CMV-specific T cells were present along with detectable virus. In LNs, CMV-specific T cells exhibited quiescent phenotypes independent of virus. Overall, T cell differentiation was enhanced in sites of viral persistence with age. Together, our results suggest tissue T cell reservoirs for CMV control shaped by both viral and tissue-intrinsic factors, with global effects on homeostasis of tissue T cells over the lifespan. @Gordon et al.

Collecting and Studying Blood and Tissue Samples From Patients With Locally Recurrent or Metastatic Prostate or Bladder/Urothelial Cancer

ClinicalTrials.gov

2017-12-04

Healthy Control; Localized Urothelial Carcinoma of the Renal Pelvis and Ureter; Metastatic Malignant Neoplasm in the Bone; Metastatic Malignant Neoplasm in the Soft Tissues; Metastatic Urothelial Carcinoma of the Renal Pelvis and Ureter; Recurrent Bladder Carcinoma; Recurrent Prostate Carcinoma; Recurrent Urothelial Carcinoma of the Renal Pelvis and Ureter; Stage IV Bladder Cancer; Stage IV Bladder Urothelial Carcinoma; Stage IV Prostate Cancer

Blood rights: the body and information privacy.

PubMed

Alston, Bruce

2005-05-01

Genetic and other medical technology makes blood, human tissue and other bodily samples an immediate and accessible source of comprehensive personal and health information about individuals. Yet, unlike medical records, bodily samples are not subject to effective privacy protection or other regulation to ensure that individuals have rights to control the collection, use and transfer of such samples. This article examines the existing coverage of privacy legislation, arguments in favour of baseline protection for bodily samples as sources of information and possible approaches to new regulation protecting individual privacy rights in bodily samples.

Rehydration Capacities and Rates for Various Porcine Tissues after Dehydration

PubMed Central

Meyer, Jacob P.; McAvoy, Kieran E.; Jiang, Jack

2013-01-01

The biphasic effects of liquid on tissue biomechanics are well known in cartilage and vocal folds, yet not extensively in other tissue types. Past studies have shown that tissue dehydration significantly impacts biomechanical properties and that rehydration can restore these properties in certain tissue types. However, these studies failed to consider how temporal exposure to dehydrating or rehydrating agents may alter tissue rehydration capacity, as overexposure to dehydration may permanently prevent rehydration to the initial liquid volume. Select porcine tissues were dehydrated until they reached between 100% and 40% of their initial mass. Each sample was allowed to rehydrate for 5 hours in a 0.9% saline solution, and the percent change between the initial and rehydrated mass values was calculated. Spearman correlation tests indicated a greater loss in mass despite rehydration when tissues were previously exposed to greater levels of dehydration. Additionally, Pearson correlation tests indicated the total liquid mass of samples after complete rehydration decreased when previously exposed to higher levels of dehydration. Rehydration rates were found by dehydrating tissues to 40% of their initial mass followed by rehydration in a 0.9% saline solution for 60 minutes, with mass measurements occurring in 15 minute intervals. All tissues rehydrated nonlinearly, most increasing significantly in mass up to 30 minutes after initial soaking. This study suggests the ability for tissues to rehydrate is dependent on the level of initial dehydration exposure. In vitro rehydration experiments therefore require controlled dosage and temporal exposure to dehydrating and rehydrating agents to avoid incomplete rehydration, and caution should be taken when combining different tissue types in models of hydration. PMID:24023753

Quantifying thermal modifications on laser welded skin tissue

NASA Astrophysics Data System (ADS)

Tabakoglu, Hasim Ö.; Gülsoy, Murat

2011-02-01

Laser tissue welding is a potential medical treatment method especially on closing cuts implemented during any kind of surgery. Photothermal effects of laser on tissue should be quantified in order to determine optimal dosimetry parameters. Polarized light and phase contrast techniques reveal information about extend of thermal change over tissue occurred during laser welding application. Change in collagen structure in skin tissue stained with hematoxilen and eosin samples can be detected. In this study, three different near infrared laser wavelengths (809 nm, 980 nm and 1070 nm) were compared for skin welding efficiency. 1 cm long cuts were treated spot by spot laser application on Wistar rats' dorsal skin, in vivo. In all laser applications, 0.5 W of optical power was delivered to the tissue, 5 s continuously, resulting in 79.61 J/cm2 energy density (15.92 W/cm2 power density) for each spot. The 1st, 4th, 7th, 14th, and 21st days of recovery period were determined as control days, and skin samples needed for histology were removed on these particular days. The stained samples were examined under a light microscope. Images were taken with a CCD camera and examined with imaging software. 809 Nm laser was found to be capable of creating strong full-thickness closure, but thermal damage was evident. The thermal damage from 980 nm laser welding was found to be more tolerable. The results showed that 1070 nm laser welding produced noticeably stronger bonds with minimal scar formation.

Association between Randall's Plaque and Calcifying Nanoparticles

NASA Technical Reports Server (NTRS)

Citfcioglu, Neva; Vejdani, Kaveh; Lee, Olivia; Mathew, Grace; Aho, Katja M.; Kajander, Olavi; McKay, David S.; Jones, Jeffrey A.; Feiveson, Alan H.; Stoller, Marshall L.

2007-01-01

Randall initially described calcified subepithelial papillary plaques, which he hypothesized as nidi for kidney stone formation. The discovery of calcifying nanoparticles (CNP) in many calcifying processes of human tissues has raised another hypothesis about their possible involvement in urinary stone formation. This research is the first attempt to investigate the potential association of these two hypotheses. We collected renal papilla and blood samples from 17 human patients who had undergone laparoscopic nephrectomy due to neoplasia. Immunohistochemical staining (IHS) was applied on the tissue samples using monoclonal antibody 8D10 (mAb) against CNP. Homogenized papillary tissues and serum samples were cultured for CNP. Scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS) analysis were performed on fixed papillary samples. Randall's plaques were visible on gross inspection in 11 out of 17 collected samples. IHS was positive for CNP antigen in 8 of these 11 visually positive samples, but in only 1 of the remaining 6 samples. SEM revealed spherical apatite formations in 14 samples, all of which had calcium and phosphate peaks detected by EDS analysis. From this study, there was some evidence of a link between the presence of Randall's plaques and the detection of CNP, also referred to as nanobacteria. Although causality was not demonstrated, these results suggest that further studies with negative control samples should be made to explore the etiology of Randall's plaque formation, thus leading to a better understanding of the pathogenesis of stone formation.

Range of motion improves after massage in children with burns: a pilot study.

PubMed

Morien, Annie; Garrison, Diane; Smith, Nancy Keeney

2008-01-01

Little is known about the effect of massage on post-burn tissue in children. We conducted a pilot study to examine the effect of massage (3-5 days) on mood and range of motion (ROM) in eight post-burn children. Participants showed significant increases in ROM from Time 1 (pre-massage, first day) to Time 2 (post-massage, last day) in massaged tissue but not control (non-massaged) tissue. Mood was elevated throughout the study and thus did not change across time. Although massage improved ROM, we are cautious in our interpretation because of the small sample size.

Methylation of Werner syndrome protein is associated with the occurrence and development of invasive meningioma via the regulation of Myc and p53 expression.

PubMed

Li, Puxian; Hao, Shuyu; Bi, Zhiyong; Zhang, Junting; Wu, Zhen; Ren, Xiaohui

2015-08-01

The aim of the present study was to investigate the positive rate of Werner syndrome protein (WRN) methylation in meningioma patients, and further assess the association between WRN methylation and the occurrence of meningioma. A total of 56 consecutive meningioma patients and 26 healthy individuals were enrolled in the study. A methylation-specific polymerase chain reaction assay was performed to detect the positive rate of WRN methylation in the peripheral blood and tissue samples collected from the recruited subjects. In addition, western blot analysis was performed to determine the protein expression levels of WRN, Myc and p53 in the peripheral blood and tissue samples. The positive rate of WRN methylation in the peripheral blood of the meningioma group was increased when compared with the control group (P<0.05). In addition, the protein expression levels of WRN were significantly decreased in the peripheral blood and tissue samples collected from the individuals with a positive WRN methylation status (P<0.05), as compared with the samples without WRN methylation. Furthermore, the protein expression levels of Myc and p53 were increased in the peripheral blood and tissue samples that exhibited positive WRN methylation when compared with those without WRN methylation (P<0.05). Therefore, WRN methylation was demonstrated to be associated with the occurrence and development of invasive meningioma, possibly through the regulation of Myc and p53 expression.

Structural locus of transmucosal albumin efflux in canine ileum. A fluorescent study.

PubMed

Granger, D N; Cook, B H; Taylor, A E

1976-12-01

This study demonstrates the effects of elevated intestinal venous pressure on the intestinal tissue spaces and the histological locus of the transmucosal albumin flux under such conditions. The authors were able to localize albumin in the tissues using an Evans blue-albumin fluorescence technique. This technique makes use of the fluorescence properties and albumin affinity of Evans blue dye (T-1824). Evans blue dye has a high affinity for albumin and emits a red-orange fluorescence at a wavelength of 720 nm. Evans blue was mixed with a solution of bovine serum albumin at concentrations that yield negligible amounts of free dye. Control ileal samples were obtained in order to visualize the natural tissue morphology and fluorescence. The Evans blue-albumin solution was injected and tissue samples were obtained 15 and 60 min postinjection, then venous outflow was occluded and after 15 and 60 min the tissues were sampled. Each sample was immediately frozen, freeze dried, embedded in paraffin, and 7-mu sections were made. The Evans blue-albumin was demonstrated histologically with a fluorescence microscope. No leakage sites were apparent at normal venous pressures. However, after elevation of venous pressure, Evans blue-albumin was observed in the interepithelial and/or intraepithelial spaces of villus tips, but no Evans blue-albumin was observed either between or within the epithelial cells of the crypts, or within the tubular crypt lumina. These results indicate that at elevated venous pressures, the transmucosal albumin flux occurs exclusively at the villus tip region, suggesting a great vulnerability of the cells found in this region to elevations in tissue pressure as compared to the crypt epithelial cells.

Application of FTA technology for sampling, recovery and molecular characterization of viral pathogens and virus-derived transgenes from plant tissues

PubMed Central

Ndunguru, Joseph; Taylor, Nigel J; Yadav, Jitender; Aly, Haytham; Legg, James P; Aveling, Terry; Thompson, Graham; Fauquet, Claude M

2005-01-01

Background Plant viral diseases present major constraints to crop production. Effective sampling of the viruses infecting plants is required to facilitate their molecular study and is essential for the development of crop protection and improvement programs. Retaining integrity of viral pathogens within sampled plant tissues is often a limiting factor in this process, most especially when sample sizes are large and when operating in developing counties and regions remote from laboratory facilities. FTA is a paper-based system designed to fix and store nucleic acids directly from fresh tissues pressed into the treated paper. We report here the use of FTA as an effective technology for sampling and retrieval of DNA and RNA viruses from plant tissues and their subsequent molecular analysis. Results DNA and RNA viruses were successfully recovered from leaf tissues of maize, cassava, tomato and tobacco pressed into FTA® Classic Cards. Viral nucleic acids eluted from FTA cards were found to be suitable for diagnostic molecular analysis by PCR-based techniques and restriction analysis, and for cloning and nucleotide sequencing in a manner equivalent to that offered by tradition isolation methods. Efficacy of the technology was demonstrated both from sampled greenhouse-grown plants and from leaf presses taken from crop plants growing in farmer's fields in East Africa. In addition, FTA technology was shown to be suitable for recovery of viral-derived transgene sequences integrated into the plant genome. Conclusion Results demonstrate that FTA is a practical, economical and sensitive method for sampling, storage and retrieval of viral pathogens and plant genomic sequences, when working under controlled conditions and in the field. Application of this technology has the potential to significantly increase ability to bring modern analytical techniques to bear on the viral pathogens infecting crop plants. PMID:15904535

Early overfeed-induced obesity leads to brown adipose tissue hypoactivity in rats.

PubMed

de Almeida, Douglas L; Fabrício, Gabriel S; Trombini, Amanda B; Pavanello, Audrei; Tófolo, Laize P; da Silva Ribeiro, Tatiane A; de Freitas Mathias, Paulo C; Palma-Rigo, Kesia

2013-01-01

Brown adipose tissue activation has been considered a potential anti-obesity mechanism because it is able to expend energy through thermogenesis. In contrast, white adipose tissue stores energy, contributing to obesity. We investigated whether the early programming of obesity by overfeeding during lactation changes structure of interscapular brown adipose tissue in adulthood and its effects on thermogenesis. Birth of litters was considered day 0. On day 2, litter size was adjusted to normal (9 pups) and small (3 pups) litters. On day 21, the litters were weaned. A temperature transponder was implanted underneath interscapular brown adipose tissue pads of 81-day-old animals; local temperature was measured during light and dark periods between days 87 and 90. The animals were euthanized, and tissue and blood samples were collected for further analysis. The vagus and retroperitoneal sympathetic nerve activity was recorded. Small litter rats presented significant lower interscapular brown adipose tissue temperature during the light (NL 37.6°C vs. SL 37.2°C) and dark (NL 38°C vs. SL 37.6°C) periods compared to controls. Morphology of small litter brown adipose tissue showed fewer lipid droplets in the tissue center and more and larger in the periphery. The activity of vagus nerve was 19,9% greater in the small litter than in control (p<0.01), and no difference was observed in the sympathetic nerve activity. In adulthood, the small litter rats were 11,7% heavier than the controls and presented higher glycemia 13,1%, insulinemia 70% and corticosteronemia 92,6%. Early overfeeding programming of obesity changes the interscapular brown adipose tissue structure in adulthood, leading to local thermogenesis hypoactivity, which may contribute to obesity in adults. © 2013 S. Karger AG, Basel.

Qualitative tissue differentiation by analysing the intensity ratios of atomic emission lines using laser induced breakdown spectroscopy (LIBS): prospects for a feedback mechanism for surgical laser systems.

PubMed

Kanawade, Rajesh; Mahari, Fanuel; Klämpfl, Florian; Rohde, Maximilian; Knipfer, Christian; Tangermann-Gerk, Katja; Adler, Werner; Schmidt, Michael; Stelzle, Florian

2015-01-01

The research work presented in this paper focuses on qualitative tissue differentiation by monitoring the intensity ratios of atomic emissions using 'Laser Induced Breakdown Spectroscopy' (LIBS) on the plasma plume created during laser tissue ablation. The background of this study is to establish a real time feedback control mechanism for clinical laser surgery systems during the laser ablation process. Ex-vivo domestic pig tissue samples (muscle, fat, nerve and skin) were used in this experiment. Atomic emission intensity ratios were analyzed to find a characteristic spectral line for each tissue. The results showed characteristic elemental emission intensity ratios for the respective tissues. The spectral lines and intensity ratios of these specific elements varied among the different tissue types. The main goal of this study is to qualitatively and precisely identify different tissue types for tissue specific laser surgery. © 2015 The Authors. Journal of Biophotonics published by WILEY-VCH Verlag.

Tissue Multiplatform-Based Metabolomics/Metabonomics for Enhanced Metabolome Coverage.

PubMed

Vorkas, Panagiotis A; Abellona U, M R; Li, Jia V

2018-01-01

The use of tissue as a matrix to elucidate disease pathology or explore intervention comes with several advantages. It allows investigation of the target alteration directly at the focal location and facilitates the detection of molecules that could become elusive after secretion into biofluids. However, tissue metabolomics/metabonomics comes with challenges not encountered in biofluid analyses. Furthermore, tissue heterogeneity does not allow for tissue aliquoting. Here we describe a multiplatform, multi-method workflow which enables metabolic profiling analysis of tissue samples, while it can deliver enhanced metabolome coverage. After applying a dual consecutive extraction (organic followed by aqueous), tissue extracts are analyzed by reversed-phase (RP-) and hydrophilic interaction liquid chromatography (HILIC-) ultra-performance liquid chromatography coupled to mass spectrometry (UPLC-MS) and nuclear magnetic resonance (NMR) spectroscopy. This pipeline incorporates the required quality control features, enhances versatility, allows provisional aliquoting of tissue extracts for future guided analyses, expands the range of metabolites robustly detected, and supports data integration. It has been successfully employed for the analysis of a wide range of tissue types.

Bi-PROF

PubMed Central

Gries, Jasmin; Schumacher, Dirk; Arand, Julia; Lutsik, Pavlo; Markelova, Maria Rivera; Fichtner, Iduna; Walter, Jörn; Sers, Christine; Tierling, Sascha

2013-01-01

The use of next generation sequencing has expanded our view on whole mammalian methylome patterns. In particular, it provides a genome-wide insight of local DNA methylation diversity at single nucleotide level and enables the examination of single chromosome sequence sections at a sufficient statistical power. We describe a bisulfite-based sequence profiling pipeline, Bi-PROF, which is based on the 454 GS-FLX Titanium technology that allows to obtain up to one million sequence stretches at single base pair resolution without laborious subcloning. To illustrate the performance of the experimental workflow connected to a bioinformatics program pipeline (BiQ Analyzer HT) we present a test analysis set of 68 different epigenetic marker regions (amplicons) in five individual patient-derived xenograft tissue samples of colorectal cancer and one healthy colon epithelium sample as a control. After the 454 GS-FLX Titanium run, sequence read processing and sample decoding, the obtained alignments are quality controlled and statistically evaluated. Comprehensive methylation pattern interpretation (profiling) assessed by analyzing 102-104 sequence reads per amplicon allows an unprecedented deep view on pattern formation and methylation marker heterogeneity in tissues concerned by complex diseases like cancer. PMID:23803588

Ambient Mass Spectrometry in Cancer Research.

PubMed

Takats, Z; Strittmatter, N; McKenzie, J S

2017-01-01

Ambient ionization mass spectrometry was developed as a sample preparation-free alternative to traditional MS-based workflows. Desorption electrospray ionization (DESI)-MS methods were demonstrated to allow the direct analysis of a broad range of samples including unaltered biological tissue specimens. In contrast to this advantageous feature, nowadays DESI-MS is almost exclusively used for sample preparation intensive mass spectrometric imaging (MSI) in the area of cancer research. As an alternative to MALDI, DESI-MSI offers matrix deposition-free experiment with improved signal in the lower (<500m/z) range. DESI-MSI enables the spatial mapping of tumor metabolism and has been broadly demonstrated to offer an alternative to frozen section histology for intraoperative tissue identification and surgical margin assessment. Rapid evaporative ionization mass spectrometry (REIMS) was developed exclusively for the latter purpose by the direct combination of electrosurgical devices and mass spectrometry. In case of the REIMS technology, aerosol particles produced by electrosurgical dissection are subjected to MS analysis, providing spectral information on the structural lipid composition of tissues. REIMS technology was demonstrated to give real-time information on the histological nature of tissues being dissected, deeming it an ideal tool for intraoperative tissue identification including surgical margin control. More recently, the method has also been used for the rapid lipidomic phenotyping of cancer cell lines as it was demonstrated in case of the NCI-60 cell line collection. © 2017 Elsevier Inc. All rights reserved.

Plutonium and americium in the foodchain lichen-reindeer-man

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jaakkola, T.; Hakanen, M.; Keinonen, M.

1977-01-01

The atmospheric nuclear tests have produced a worldwide fallout of transuranium elements. In addition to plutonium measurable concentrations of americium are to be found in terrestrial and aquatic environments. The metabolism of plutonium in reindeer was investigated by analyzing plutonium in liver, bone, and lung collected during 1963-1976. To determine the distribution of plutonium in reindeer all tissues of four animals of different ages were analyzed. To estimate the uptake of plutonium from the gastrointestinal tract in reindeer, the tissue samples of elk were also analyzed. Elk which is of the same genus as reindeer does not feed on lichenmore » but mainly on deciduous plants, buds, young twigs, and leaves of trees and bushes. The composition of its feed corresponds fairly well to that of reindeer during the summer. Studies on behaviour of americium along the foodchain lichen-reindeer-man were started by determining the Am-241 concentrations in lichen and reindeer liver. The Am-241 results were compared with those of Pu-239,240. The plutonium contents of the southern Finns, whose diet does not contain reindeer tissues, were determined by analyzing autopsy tissue samples (liver, lung, and bone). The southern Finns form a control group to the Lapps consuming reindeer tissues. Plutonium analyses of the placenta, blood, and tooth samples of the Lapps were performed.« less

Hypermethylation of CDH13, DKK3 and FOXL2 promoters and the expression of EZH2 in ovary granulosa cell tumors.

PubMed

Xu, Yanmei; Li, Xia; Wang, Hongtao; Xie, Pengmu; Yan, Xun; Bai, Yu; Zhang, Tingguo

2016-09-01

Aberrant epigenetic modification is associated with the development and progression of cancer. Hypermethylation of tumor suppressor gene promoters and cooperative histone modification have been considered to be the primary mechanisms of epigenetic modification. Ovary granulosa cell tumors (GCTs) are relatively rare, accounting for ~3% of all ovarian malignancies. The present study assessed hypermethylation of the cadherin 13 (CDH13), dickkopf WNT signaling pathway inhibitor 3 (DKK3) and forkhead box L2 (FOXL2) promoters in 30 GCT tissues and 30 healthy control tissues using methylation-specific polymerase chain reaction analysis. The data showed that the frequencies of CDH13, DKK3 and FOXL2 promoter methylation were significantly higher in the GCT tissues, compared with the healthy control tissues (86.67, vs. 23.33%; 80, vs. 26.67% and 66.67, vs. 20%, respectively; P<0.001). Immunostaining of enhancer of zeste homolog 2 (EZH2), a histone H3K27 methyltransferase, showed that the EZH2 protein was expressed in 11 of the 30 GCT tissue samples, whereas no EZH2 protein was expressed in the 30 healthy control tissues (P<0.01). These data suggested that hypermethylation of the CDH13, DKK3 and FOXL2 gene promoters, and overexpression of the EZH2 protein were involved in the development of GCT.

Investigation of the efficacy of ultrafast laser in large bowel excision

NASA Astrophysics Data System (ADS)

Mohanan, Syam Mohan P. C.; Beck, Rainer J.; Góra, Wojciech S.; Perry, Sarah L.; Shires, Mike; Jayne, David; Hand, Duncan P.; Shephard, Jonathan D.

2017-02-01

Local resection of early stage tumors in the large bowel via colonoscopy has been a widely accepted surgical modality for colon neoplasm treatment. The conventional electrocautery techniques used for the resection of neoplasia in the mucosal or submucosal layer of colon tissue has been shown to create obvious thermal necrosis to adjacent healthy tissues and lacks accuracy in resection. Ultrafast picosecond (ps) laser ablation using a wavelength of 1030 or 515 nm is a promising surgical tool to overcome the limitations seen with conventional surgical techniques. The purpose of this initial study is to analyze the depth of ablation or the extent of coagulation deployed by the laser as a function of pulse energy and fluence in an ex-vivo porcine model. Precise control of the depth of tissue removal is of paramount importance for bowel surgery where bowel perforation can lead to morbidity or mortality. Thus we investigate the regimes that are optimal for tissue resection and coagulation through plasma mediated ablation of healthy colon tissue. The ablated tissue samples were analyzed by standard histologic methods and a three dimensional optical profilometer technique. We demonstrate that ultrafast laser resection of colonic tissue can minimize the region of collateral thermal damage (<50 μm) with a controlled ablation depth. This surgical modality allows potentially easier removal of early stage lesions and has the capability to provide more control to the surgeon in comparison with a mechanical or electrocautery device.

In vitro characterization of a novel tissue engineered based hybridized nano and micro structured collagen implant and its in vivo role on tenoinduction, tenoconduction, tenogenesis and tenointegration.

PubMed

Oryan, Ahmad; Moshiri, Ali; Meimandi-Parizi, Abdolhamid

2014-03-01

Surgical reconstruction of large tendon defects is technically demanding. Tissue engineering is a new option. We produced a novel tissue engineered, collagen based, bioimplant and in vitro characterizations of the implant were investigated. In addition, we investigated role of the collagen implant on the healing of a large tendon defect model in rabbits. A two cm length of the left rabbit's Achilles tendon was transected and discarded. The injured tendons of all the rabbits were repaired by Kessler pattern to create and maintain a 2 cm tendon gap. The collagen implant was inserted in the tendon defect of the treatment group (n = 30). The defect area was left intact in the control group (n = 30). The animals were euthanized at 60 days post injury (DPI) and the macro- micro- and nano- morphologies and the biomechanical characteristics of the tendon samples were studied. Differences of P < 0.05 were considered significant. The host graft interaction was followed at various stages of tendon healing, using pilot animals. At 60 DPI, a significant increase in number, diameter and density of the collagen fibrils, number and maturity of tenoblasts and tenocytes, alignment of the collagen fibrils and maturity of the elastic fibers were seen in the treated tendons when compared to the control ones (P < 0.05). Compared to the control lesions, number of inflammatory cells, amount of peritendinous adhesions and muscle fibrosis and atrophy, were significantly lower in the treated lesions (P < 0.05). Treatment also significantly increased load to failure, tensile strength and elastic modulus of the samples as compared with the control ones. The collagen implant properly incorporated with the healing tissue and was replaced by the new tendinous structure which was superior both ultra-structurally and physically than the loose areolar connective tissue regenerated in the control lesions. The results of this study may be valuable in the clinical practice.

A Combination Tissue Engineering Strategy for Schwann Cell-Induced Spinal Cord Repair

DTIC Science & Technology

2015-10-01

the mechanical perturbation (2-3Hz) in both samples, however, there is much more power in the PVDF-TrEE sample overall. The frequency spectra for the...aligned-fibers contain signal power above and beyond the first and second harmonics of the mechanical stimulus, unlike the control sample on the...right. This finding shows that the 8 aligned PVDF-TrFE fibers generate field potentials that show up at higher harmonics of the mechanical

Detection of toxoplasmosis in experimentally infected goats by PCR.

PubMed

Sreekumar, C; Rao, J R; Mishra, A K; Ray, D; Joshi, P; Singh, R K

2004-05-15

PCR was used to diagnose toxoplasmosis in two pairs of Barbari goats infected by oral administration of doses of either 10(4) or 10(5) oocysts of Toxoplasma gondii. Blood and lymph node aspirates were collected from the infected goats and control goat at intervals, and tissues were also collected from a fetus that was aborted and a doe that died during the trial. Both processed and unprocessed samples were used for the PCR, using primers directed to the multicopy B1 gene. None of the blood samples was positive, but a specific signal was obtained from the lymph node aspirates after partial DNA extraction. Direct PCR of the lung, muscle and mesenteric lymph node of the doe and lung tissue of the aborted fetus yielded the target fragment. The simplified PCR protocols, including partial DNA extraction and direct assay of lung tissue, were effective for the diagnosis of toxoplasmosis.

Design and evaluation of a new Peltier-cooled laser ablation cell with on-sample temperature control.

PubMed

Konz, Ioana; Fernández, Beatriz; Fernández, M Luisa; Pereiro, Rosario; Sanz-Medel, Alfredo

2014-01-27

A new custom-built Peltier-cooled laser ablation cell is described. The proposed cryogenic cell combines a small internal volume (20 cm(3)) with a unique and reliable on-sample temperature control. The use of a flexible temperature sensor, directly located on the sample surface, ensures a rigorous sample temperature control throughout the entire analysis time and allows instant response to any possible fluctuation. In this way sample integrity and, therefore, reproducibility can be guaranteed during the ablation. The refrigeration of the proposed cryogenic cell combines an internal refrigeration system, controlled by a sensitive thermocouple, with an external refrigeration system. Cooling of the sample is directly carried out by 8 small (1 cm×1 cm) Peltier elements placed in a circular arrangement in the base of the cell. These Peltier elements are located below a copper plate where the sample is placed. Due to the small size of the cooling electronics and their circular allocation it was possible to maintain a peephole under the sample for illumination allowing a much better visualization of the sample, a factor especially important when working with structurally complex tissue sections. The analytical performance of the cryogenic cell was studied using a glass reference material (SRM NIST 612) at room temperature and at -20°C. The proposed cell design shows a reasonable signal washout (signal decay within less than 10 s to background level), high sensitivity and good signal stability (in the range 6.6-11.7%). Furthermore, high precision (0.4-2.6%) and accuracy (0.3-3.9%) in the isotope ratio measurements were also observed operating the cell both at room temperature and at -20°C. Finally, experimental results obtained for the cell application to qualitative elemental imaging of structurally complex tissue samples (e.g. eye sections from a native frozen porcine eye and fresh flower leaves) demonstrate that working in cryogenic conditions is critical in such type of direct sample analysis. Copyright © 2013 Elsevier B.V. All rights reserved.

Evidence of presence of Mycobacterium tuberculosis in bovine tissue samples by multiplex PCR: possible relevance to reverse zoonosis.

PubMed

Mittal, M; Chakravarti, S; Sharma, V; Sanjeeth, B S; Churamani, C P; Kanwar, N S

2014-04-01

Bovine tuberculosis, caused by Mycobacterium bovis, remains one of the most important zoonotic health concerns worldwide. The transmission of Mycobacterium tuberculosis from humans to animals also occurs especially in countries where there is close interaction of humans with the animals. In the present study, thirty bovine lung tissue autopsy samples from an organized dairy farm located in North India were screened for the presence of Mycobacterium tuberculosis complex by smear microscopy, histopathological findings and PCR. Differential diagnosis of M. tuberculosis and M. bovis was made based on the deletion of mce-3 operon in M. bovis. The present study found eight of these samples positive for M. tuberculosis by multiplex PCR. Sequencing was performed on two PCR-positive representative samples and on annotation, and BLAST analysis confirmed the presence of gene fragment specific to Mycobacterium tuberculosis. The presence of M. tuberculosis in all the positive samples raises the possibility of human-to-cattle transmission and possible adaptation of this organism in bovine tissues. This study accentuates the importance of screening and differential diagnosis of Mycobacterium tuberculosis complex in humans and livestock for adopting effective TB control and eradication programmes. © 2014 Blackwell Verlag GmbH.

Determination of the phenoxyacid herbicides MCPA, mecoprop and 2,4-D in kidney tissue using liquid chromatography with electrospray tandem mass spectrometry.

PubMed

Charlton, Andrew J A; Stuckey, Vicki; Sykes, Mark D

2009-06-01

An analytical method was developed to determine the phenoxyacid herbicides 2,4-D, MCPA and mecoprop in kidney tissue from animals where poisoning is suspected. Samples were Soxhlet extracted using diethyl ether and the extracts cleaned-up using anion exchange solid phase extraction cartridges. Analysis was performed using liquid chromatography with negative-ion electrospray tandem mass spectrometry (LC-MS/MS). The method was evaluated by analysing control kidney samples fortified at 1 and 5 mg/kg. Mean recoveries ranged from 82 to 93% with relative standard deviations from 3.2 to 19%. The limit of detection was estimated to be 0.02 mg/kg.

Care during freeze-drying of bovine pericardium tissue to be used as a biomaterial: a comparative study.

PubMed

Polak, Roberta; Pitombo, Ronaldo N M

2011-10-01

Bovine pericardium (BP) tissue is widely used in the manufacture of bioprosthetics. The effects of freeze-drying on the BP tissue have been studied by some researchers in order to decrease their cytotoxicity due to preservation in formaldehyde solution, and to increase the lifetime of the product in storage. This study was undertaken in order to study the effect of freeze-drying in the structure of BP. To perform this study BP samples were freeze-dried in two different types of freeze-dryers available in our laboratory: a laboratory freeze-dryer, in which it was not possible to control parameters and a pilot freeze-dryer, wherein all parameters during freezing and drying were controlled. After freeze-drying processes, samples were analyzed by SEM, Raman spectroscopy, tensile strength, water uptake tests and TEM. In summary, it has been demonstrated that damages occur in collagen fibers by the loss of bulk water of collagen structure implicating in a drastic decreasing of BP mechanical properties due to its structural alterations. Moreover, it was proven that the collagen fibrils suffered breakage at some points, which can be attributed to the uncontrolled parameters during drying. Copyright © 2011 Elsevier Inc. All rights reserved.

Preparation of A Spaceflight: Apoptosis Search in Sutured Wound Healing Models.

PubMed

Riwaldt, Stefan; Monici, Monica; Graver Petersen, Asbjørn; Birk Jensen, Uffe; Evert, Katja; Pantalone, Desiré; Utpatel, Kirsten; Evert, Matthias; Wehland, Markus; Krüger, Marcus; Kopp, Sascha; Frandsen, Sofie; Corydon, Thomas; Sahana, Jayashree; Bauer, Johann; Lützenberg, Ronald; Infanger, Manfred; Grimm, Daniela

2017-12-03

To prepare the ESA (European Space Agency) spaceflight project "Wound healing and Sutures in Unloading Conditions", we studied mechanisms of apoptosis in wound healing models based on ex vivo skin tissue cultures, kept for 10 days alive in serum-free DMEM/F12 medium supplemented with bovine serum albumin, hydrocortisone, insulin, ascorbic acid and antibiotics at 32 °C. The overall goal is to test: (i) the viability of tissue specimens; (ii) the gene expression of activators and inhibitors of apoptosis and extracellular matrix components in wound and suture models; and (iii) to design analytical protocols for future tissue specimens after post-spaceflight download. Hematoxylin-Eosin and Elastica-van-Gieson staining showed a normal skin histology with no signs of necrosis in controls and showed a normal wound suture. TdT-mediated dUTP-biotin nick end labeling for detecting DNA fragmentation revealed no significant apoptosis. No activation of caspase-3 protein was detectable. FASL , FADD , CASP3 , CASP8 , CASP10 , BAX , BCL2 , CYC1 , APAF1 , LAMA3 and SPP1 mRNAs were not altered in epidermis and dermis samples with and without a wound compared to 0 day samples (specimens investigated directly post-surgery). BIRC5 , CASP9 , and FN1 mRNAs were downregulated in epidermis/dermis samples with and/or without a wound compared to 0 day samples. BIRC2 , BIRC3 were upregulated in 10 day wound samples compared to 0 day samples in epidermis/dermis. RELA/FAS mRNAs were elevated in 10 day wound and no wound samples compared to 0 day samples in dermis. In conclusion, we demonstrate that it is possible to maintain live skin tissue cultures for 10 days. The viability analysis showed no significant signs of cell death in wound and suture models. The gene expression analysis demonstrated the interplay of activators and inhibitors of apoptosis and extracellular matrix components, thereby describing important features in ex vivo sutured wound healing models. Collectively, the performed methods defining analytical protocols proved to be applicable for post-flight analyzes of tissue specimens after sample return.

Triglyceride dependent differentiation of obesity in adipose tissues by FTIR spectroscopy coupled with chemometrics.

PubMed

Kucuk Baloglu, Fatma; Baloglu, Onur; Heise, Sebastian; Brockmann, Gudrun; Severcan, Feride

2017-10-01

The excess deposition of triglycerides in adipose tissue is the main reason of obesity and causes excess release of fatty acids to the circulatory system resulting in obesity and insulin resistance. Body mass index and waist circumference are not precise measure of obesity and obesity related metabolic diseases. Therefore, in the current study, it was aimed to propose triglyceride bands located at 1770-1720 cm -1 spectral region as a more sensitive obesity related biomarker using the diagnostic potential of Fourier Transform Infrared (FTIR) spectroscopy in subcutaneous (SCAT) and visceral (VAT) adipose tissues. The adipose tissue samples were obtained from 10 weeks old male control (DBA/2J) (n = 6) and four different obese BFMI mice lines (n = 6 per group). FTIR spectroscopy coupled with hierarchical cluster analysis (HCA) and principal component analysis (PCA) was applied to the spectra of triglyceride bands as a diagnostic tool in the discrimination of the samples. Successful discrimination of the obese, obesity related insulin resistant and control groups were achieved with high sensitivity and specificity. The results revealed the power of FTIR spectroscopy coupled with chemometric approaches in internal diagnosis of abdominal obesity based on the spectral differences in the triglyceride region that can be used as a spectral marker. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Principles, Techniques, and Applications of Tissue Microfluidics

NASA Technical Reports Server (NTRS)

Wade, Lawrence A.; Kartalov, Emil P.; Shibata, Darryl; Taylor, Clive

2011-01-01

The principle of tissue microfluidics and its resultant techniques has been applied to cell analysis. Building microfluidics to suit a particular tissue sample would allow the rapid, reliable, inexpensive, highly parallelized, selective extraction of chosen regions of tissue for purposes of further biochemical analysis. Furthermore, the applicability of the techniques ranges beyond the described pathology application. For example, they would also allow the posing and successful answering of new sets of questions in many areas of fundamental research. The proposed integration of microfluidic techniques and tissue slice samples is called tissue microfluidics because it molds the microfluidic architectures in accordance with each particular structure of each specific tissue sample. Thus, microfluidics can be built around the tissues, following the tissue structure, or alternatively, the microfluidics can be adapted to the specific geometry of particular tissues. By contrast, the traditional approach is that microfluidic devices are structured in accordance with engineering considerations, while the biological components in applied devices are forced to comply with these engineering presets. The proposed principles represent a paradigm shift in microfluidic technology in three important ways: Microfluidic devices are to be directly integrated with, onto, or around tissue samples, in contrast to the conventional method of off-chip sample extraction followed by sample insertion in microfluidic devices. Architectural and operational principles of microfluidic devices are to be subordinated to suit specific tissue structure and needs, in contrast to the conventional method of building devices according to fluidic function alone and without regard to tissue structure. Sample acquisition from tissue is to be performed on-chip and is to be integrated with the diagnostic measurement within the same device, in contrast to the conventional method of off-chip sample prep and subsequent insertion into a diagnostic device. A more advanced form of tissue integration with microfluidics is tissue encapsulation, wherein the sample is completely encapsulated within a microfluidic device, to allow for full surface access. The immediate applications of these approaches lie with diagnostics of tissue slices and biopsy samples e.g. for cancer but the approaches would also be very useful in comparative genomics and other areas of fundamental research involving heterogeneous tissue samples.

Accurate and sensitive liquid chromatography/tandem mass spectrometry simultaneous assay of seven steroids in monkey brain.

PubMed

Bertin, Jonathan; Dury, Alain Y; Ke, Yuyong; Ouellet, Johanne; Labrie, Fernand

2015-06-01

Following its secretion mainly by the adrenal glands, dehydroepiandrosterone (DHEA) acts primarily in the cells/tissues which express the enzymes catalyzing its intracellular conversion into sex steroids by the mechanisms of intracrinology. Although reliable assays of endogenous serum steroids are now available using mass spectrometry (MS)-based technology, sample preparation from tissue matrices remains a challenge. This is especially the case with high lipid-containing tissues such as the brain. With the combination of a UPLC system with a sensitive tandem MS, it is now possible to measure endogenous unconjugated steroids in monkey brain tissue. A Shimadzu UPLC LC-30AD system coupled to a tandem MS AB Sciex Qtrap 6500 system was used. The lower limits of quantifications are achieved at 250 pg/mL for DHEA, 200 pg/mL for 5-androstenediol (5-diol), 12 pg/mL for androstenedione (4-dione), 50 pg/mL for testosterone (Testo), 10 pg/mL for dihydrotestosterone (DHT), 4 pg/mL for estrone (E1) and 1 pg/mL for estradiol (E2). The linearity and accuracy of quality controls (QCs) and endogenous quality controls (EndoQCs) are according to the guidelines of the regulatory agencies for all seven compounds. We describe a highly sensitive, specific and robust LC-MS/MS method for the simultaneous measurement of seven unconjugated steroids in monkey brain tissue. The single and small amount of sample required using a relatively simple preparation method should be useful for steroid assays in various peripheral tissues and thus help analysis of the role of locally-made sex steroids in the regulation of specific physiological functions. Copyright © 2015 Elsevier Inc. All rights reserved.

Linagliptin prevents atrial electrical and structural remodeling in a canine model of atrial fibrillation.

PubMed

Igarashi, Tazuru; Niwano, Shinichi; Niwano, Hiroe; Yoshizawa, Tomoharu; Nakamura, Hironori; Fukaya, Hidehira; Fujiishi, Tamami; Ishizue, Naruya; Satoh, Akira; Kishihara, Jun; Murakami, Masami; Ako, Junya

2018-05-02

Dipeptidyl peptidase 4 (DPP-4) inhibitors have recently been reported to exhibit additional cardioprotective effects; however, their effect in atrial remodeling, such as in atrial fibrillation (AF), remains unclear. In this study, the effect of linagliptin on atrial electrical and structural remodeling was evaluated in a canine AF model. Sixteen beagle dogs with 3-week atrial rapid stimulation were divided into the linagliptin group (9 mg/kg/day, n = 8) and pacing control group (n = 8). Three additional dogs without rapid pacing were assigned into non-pacing group, which was used as sham in this study. In the dogs with rapid pacing, the atrial effective refractory period (AERP), conduction velocity (CV), and AF inducibility were evaluated and blood was sampled every week. After the entire protocol, atrial tissue was sampled for histological examinations using HE, Azan, and dihydroethidium (DHE) staining to evaluate any tissue damage or oxidative stress. The pacing control group exhibited a gradual AERP shortening and CV decrease along the time course as previously reported. In the linagliptin group, the AERP shortening was not affected, but the CV decrease was suppressed in comparison to the control group (p < 0.05). The AF inducibility was increased in the control group and suppressed in the linagliptin group (p < 0.05). The control group exhibited tissue fibrosis, the degree of which was suppressed in the linagliptin group. DHE staining exhibited suppression of the reactive oxygen species expression in the linagliptin group in comparison to the pacing control group. Linagliptin, a DPP-4-inhibitor, suppressed the AF inducibility, CV decrease, and overexpression of oxidative stress in the canine AF model. Such suppressive effects of linagliptin on AF in the canine model may possibly be related to the anti-oxidative effect.

Selective reactivation of human herpesvirus 6 in patients with autoimmune connective tissue diseases.

PubMed

Broccolo, Francesco; Drago, Francesco; Cassina, Giulia; Fava, Andrea; Fusetti, Lisa; Matteoli, Barbara; Ceccherini-Nelli, Luca; Sabbadini, Maria Grazia; Lusso, Paolo; Parodi, Aurora; Malnati, Mauro S

2013-11-01

Viral infections have been associated with autoimmune connective tissue diseases. To evaluate whether active infection by Epstein-Barr virus (EBV), cytomegalovirus (CMV), human herpesvirus (HHV)-6, -7, -8, as well as parvovirus B19 (B19V) occur in patients with autoimmune connective tissue diseases, viral DNA loads were assessed in paired samples of serum and peripheral blood mononuclear cells (PBMCs) of 115 patients affected by different disorders, including systemic sclerosis, systemic, and discoid lupus erythematosus, rheumatoid arthritis, and dermatomyositis. Two additional groups, patients affected by inflammatory diseases (n=51) and healthy subjects (n=58) were studied as controls. The titers of anti-HHV-6 and anti-EBV antibodies were also evaluated. Cell-free HHV-6 serum viremia was detected in a significantly higher proportion of connective tissue diseases patients compared to controls (P<0.0002); a significant association between HHV-6 reactivation and the active disease state was found only for lupus erythematosus (P=0.021). By contrast, the rate of cell-free EBV viremia was similar in patients and controls groups. Cell-free CMV, HHV-8, and B19V viremia was not detected in any subject. Anti-HHV-6 and anti-EBV early antigen IgG titers were both significantly higher in autoimmune diseases patients as compared to healthy controls, although they were not associated with the presence of viremia. EBV, HHV-6, -7 prevalence and viral load in PBMCs of patients with connective tissue diseases and controls were similar. These data suggest that HHV-6 may act as a pathogenic factor predisposing patients to the development of autoimmune connective tissue diseases or, conversely, that these disorders may predispose patients to HHV-6 reactivation. © 2013 Wiley Periodicals, Inc.

Monitoring soft tissue coagulation by optical spectroscopy

NASA Astrophysics Data System (ADS)

Lihachev, A.; Lihacova, I.; Heinrichs, H.; Spigulis, J.; Trebst, T.; Wehner, M.

2017-12-01

Laser tissue welding (LTW) or laser tissue soldering (LTS) is investigated since many years for treatment of incisions, wound closure and anastomosis of vessels [1, 2]. Depending on the process, a certain temperature in the range between 65 °C to 85 °C must be reached and held for a few seconds. Care has to be taken not to overheat the tissue, otherwise necrosis or tissue carbonization may occur and will impair wound healing. Usually the temperature is monitored during the process to control the laser power [3]. This requires either bulky equipment or expensive and fragile infrared fibers to feed the temperature signal to an infrared detector. Alternatively, changes in tissue morphology can be directly observed by analysis of spectral reflectance. We investigate spectral changes in the range between 400 nm to 900 nm wavelength. Characteristic spectral changes occur when the temperature of tissue samples increase above 70 °C which is a typical setpoint value for temperature control of coagulation. We conclude that simple spectroscopy in the visible range can provide valuable information during LTS and LTW and probably replace the delicate measurement of temperature. A major advantage is that optical measurements can be performed using standard optical fibers and can be easily integrated into a surgical tool.

DNA Repair Alterations in Children With Pediatric Malignancies: Novel Opportunities to Identify Patients at Risk for High-Grade Toxicities

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ruebe, Claudia E., E-mail: claudia.ruebe@uks.e; Fricke, Andreas; Schneider, Ruth

Purpose: To evaluate, in a pilot study, the phosphorylated H2AX ({gamma}H2AX) foci approach for identifying patients with double-strand break (DSB) repair deficiencies, who may overreact to DNA-damaging cancer therapy. Methods and Materials: The DSB repair capacity of children with solid cancers was analyzed compared with that of age-matched control children and correlated with treatment-related normal-tissue responses (n = 47). Double-strand break repair was investigated by counting {gamma}H2AX foci in blood lymphocytes at defined time points after irradiation of blood samples. Results: Whereas all healthy control children exhibited proficient DSB repair, 3 children with tumors revealed clearly impaired DSB repair capacities,more » and 2 of these repair-deficient children developed life-threatening or even lethal normal-tissue toxicities. The underlying mutations affecting regulatory factors involved in DNA repair pathways were identified. Moreover, significant differences in mean DSB repair capacity were observed between children with tumors and control children, suggesting that childhood cancer is based on genetic alterations affecting DSB repair function. Conclusions: Double-strand break repair alteration in children may predispose to cancer formation and may affect children's susceptibility to normal-tissue toxicities. Phosphorylated H2AX analysis of blood samples allows one to detect DSB repair deficiencies and thus enables identification of children at risk for high-grade toxicities.« less

Cryopreservation of Cell/Scaffold Tissue-Engineered Constructs

PubMed Central

Costa, Pedro F.; Dias, Ana F.; Reis, Rui L.

2012-01-01

The aim of this work was to study the effect of cryopreservation over the functionality of tissue-engineered constructs, analyzing the survival and viability of cells seeded, cultured, and cryopreserved onto 3D scaffolds. Further, it also evaluated the effect of cryopreservation over the properties of the scaffold material itself since these are critical for the engineering of most tissues and in particular, tissues such as bone. For this purpose, porous scaffolds, namely fiber meshes based on a starch and poly(caprolactone) blend were seeded with goat bone marrow stem cells (GBMSCs) and cryopreserved for 7 days. Discs of the same material seeded with GBMSCs were also used as controls. After this period, these samples were analyzed and compared to samples collected before the cryopreservation process. The obtained results demonstrate that it is possible to maintain cell viability and scaffolds properties upon cryopreservation of tissue-engineered constructs based on starch scaffolds and goat bone marrow mesenchymal cells using standard cryopreservation methods. In addition, the outcomes of this study suggest that the greater porosity and interconnectivity of scaffolds favor the retention of cellular content and cellular viability during cryopreservation processes, when compared with nonporous discs. These findings indicate that it might be possible to prepare off-the-shelf engineered tissue substitutes and preserve them to be immediately available upon request for patients' needs. PMID:22676448

Efficacy of a Blend of Sulfuric Acid and Sodium Sulfate against Shiga Toxin-Producing Escherichia coli, Salmonella, and Nonpathogenic Escherichia coli Biotype I on Inoculated Prerigor Beef Surface Tissue.

PubMed

Scott-Bullard, Britteny R; Geornaras, Ifigenia; Delmore, Robert J; Woerner, Dale R; Reagan, James O; Morgan, J Bred; Belk, Keith E

2017-12-01

A study was conducted to investigate the efficacy of a sulfuric acid-sodium sulfate blend (SSS) against Escherichia coli O157:H7, non-O157 Shiga toxin-producing E. coli (STEC), Salmonella, and nonpathogenic E. coli biotype I on prerigor beef surface tissue. The suitability of using the nonpathogenic E. coli as a surrogate for in-plant validation studies was also determined by comparing the data obtained for the nonpathogenic inoculum with those for the pathogenic inocula. Prerigor beef tissue samples (10 by 10 cm) were inoculated (ca. 6 log CFU/cm 2 ) on the adipose side in a laboratory-scale spray cabinet with multistrain mixtures of E. coli O157:H7 (5 strains), non-O157 STEC (12 strains), Salmonella (6 strains), or E. coli biotype I (5 strains). Treatment parameters evaluated were two SSS pH values (1.5 and 1.0) and two spray application pressures (13 and 22 lb/in 2 ). Untreated inoculated beef tissue samples served as controls for initial bacterial populations. Overall, the SSS treatments lowered inoculated (6.1 to 6.4 log CFU/cm 2 ) bacterial populations by 0.6 to 1.5 log CFU/cm 2 (P < 0.05), depending on inoculum type and recovery medium. There were no main effects (P ≥ 0.05) of solution pH or spray application pressure when SSS was applied to samples inoculated with any of the tested E. coli inocula; however, solution pH did have a significant effect (P < 0.05) when SSS was applied to samples inoculated with Salmonella. Results indicated that the response of the nonpathogenic E. coli inoculum to the SSS treatments was similar (P ≥ 0.05) to that of the pathogenic inocula tested, making the E. coli biotype I strains viable surrogate organisms for in-plant validation of SSS efficacy on beef. The application of SSS at the tested parameters to prerigor beef surface tissue may be an effective intervention for controlling pathogens in a commercial beef harvest process.

Induction of angiogenesis and neovascularization in adjacent tissue of plasma-collagen-coated silicone implants.

PubMed

Ring, Andrej; Langer, Stefan; Tilkorn, Daniel; Goertz, Ole; Henrich, Lena; Stricker, Ingo; Steinau, Hans-Ulrich; Steinstraesser, Lars; Hauser, Joerg

2010-09-28

Formation of encapsulating, avascular fibrous tissue is deemed to decrease implant's biocompatibility and versatility. We investigated whether plasma-mediated collagen coating possesses the ability to enhance neovascularization in the vicinity of silicone implants. Plasma-treated collagen-I-coated silicone samples were placed into the dorsal skinfold chambers of female balb/c mice (n = 10). Conventional silicone served as control (n = 10). Intravital microscopy was performed within implant's surrounding tissue on days 1, 5, and 10. Functional vessel density, intervascular distance, vessel diameter, microvascular permeability, red blood cell velocity, and leukocyte-endothelium interaction were determined. Enhanced angiogenesis in the tissue surrounding plasma-pretreated collagen-coated implants was noted. Significant increase of functional vessel density due to vascular new development was observed (t test, P < .05). Analyses of microvascular permeability and red blood cell velocity displayed stable perfusion of the vascular network neighboring the surface-modified implants. Intensified vascularity due to induced angiogenesis and neovascularization in the tissue surrounding plasma-collagen-coated samples were observed. These results indicate that plasma-mediated collagen coating might be a promising technology in order to improve the biocompatibility and versatility of silicone implants.

Induction of Angiogenesis and Neovascularization in Adjacent Tissue of Plasma-Collagen–Coated Silicone Implants

PubMed Central

Ring, Andrej; Langer, Stefan; Tilkorn, Daniel; Goertz, Ole; Henrich, Lena; Stricker, Ingo; Steinau, Hans-Ulrich; Steinstraesser, Lars; Hauser, Joerg

2010-01-01

Objective: Formation of encapsulating, avascular fibrous tissue is deemed to decrease implant's biocompatibility and versatility. We investigated whether plasma-mediated collagen coating possesses the ability to enhance neovascularization in the vicinity of silicone implants. Methods: Plasma-treated collagen-I–coated silicone samples were placed into the dorsal skinfold chambers of female balb/c mice (n = 10). Conventional silicone served as control (n = 10). Intravital microscopy was performed within implant's surrounding tissue on days 1, 5, and 10. Functional vessel density, intervascular distance, vessel diameter, microvascular permeability, red blood cell velocity, and leukocyte-endothelium interaction were determined. Results: Enhanced angiogenesis in the tissue surrounding plasma-pretreated collagen-coated implants was noted. Significant increase of functional vessel density due to vascular new development was observed (t test, P < .05). Analyses of microvascular permeability and red blood cell velocity displayed stable perfusion of the vascular network neighboring the surface-modified implants. Conclusion: Intensified vascularity due to induced angiogenesis and neovascularization in the tissue surrounding plasma-collagen–coated samples were observed. These results indicate that plasma-mediated collagen coating might be a promising technology in order to improve the biocompatibility and versatility of silicone implants. PMID:20936137

Determination of oxidation state of iron in normal and pathologically altered human aortic valves

NASA Astrophysics Data System (ADS)

Czapla-Masztafiak, J.; Lis, G. J.; Gajda, M.; Jasek, E.; Czubek, U.; Bolechała, F.; Borca, C.; Kwiatek, W. M.

2015-12-01

In order to investigate changes in chemical state of iron in normal and pathologically altered human aortic valves X-ray absorption spectroscopy was applied. Since Fe is suspected to play detrimental role in aortic valve stenosis pathogenesis the oxidation state of this element has been determined. The experimental material consisted of 10 μm sections of valves excised during routine surgery and from autopsies. The experiment was performed at the MicroXAS beamline of the SLS synchrotron facility in Villigen (Switzerland). The Fe K-edge XANES spectra obtained from tissue samples were carefully analyzed and compared with the spectra of reference compounds containing iron in various chemical structures. The analysis of absorption edge position and shape of the spectra revealed that both chemical forms of iron are presented in valve tissue but Fe3+ is the predominant form. Small shift of the absorption edge toward higher energy in the spectra from stenotic valve samples indicates higher content of the Fe3+ form in pathological tissue. Such a phenomenon suggests the role of Fenton reaction and reactive oxygen species in the etiology of aortic valve stenosis. The comparison of pre-edge regions of XANES spectra for control and stenotic valve tissue confirmed no differences in local symmetry or spin state of iron in analyzed samples.

Quantitative Tissue Proteomics Analysis Reveals Versican as Potential Biomarker for Early-Stage Hepatocellular Carcinoma.

PubMed

Naboulsi, Wael; Megger, Dominik A; Bracht, Thilo; Kohl, Michael; Turewicz, Michael; Eisenacher, Martin; Voss, Don Marvin; Schlaak, Jörg F; Hoffmann, Andreas-Claudius; Weber, Frank; Baba, Hideo A; Meyer, Helmut E; Sitek, Barbara

2016-01-04

Hepatocellular carcinoma (HCC) is one of the most aggressive tumors, and the treatment outcome of this disease is improved when the cancer is diagnosed at an early stage. This requires biomarkers allowing an accurate and early tumor diagnosis. To identify potential markers for such applications, we analyzed a patient cohort consisting of 50 patients (50 HCC and 50 adjacent nontumorous tissue samples as controls) using two independent proteomics approaches. We performed label-free discovery analysis on 19 HCC and corresponding tissue samples. The data were analyzed considering events known to take place in early events of HCC development, such as abnormal regulation of Wnt/b-catenin and activation of receptor tyrosine kinases (RTKs). 31 proteins were selected for verification experiments. For this analysis, the second set of the patient cohort (31 HCC and corresponding tissue samples) was analyzed using selected (multiple) reaction monitoring (SRM/MRM). We present the overexpression of ATP-dependent RNA helicase (DDX39), Fibulin-5 (FBLN5), myristoylated alanine-rich C-kinase substrate (MARCKS), and Serpin H1 (SERPINH1) in HCC for the first time. We demonstrate Versican core protein (VCAN) to be significantly associated with well differentiated and low-stage HCC. We revealed for the first time the evidence of VCAN as a potential biomarker for early-HCC diagnosis.

Pathobiological investigation of naturally infected canine rabies cases from Sri Lanka.

PubMed

Beck, S; Gunawardena, P; Horton, D L; Hicks, D J; Marston, D A; Ortiz-Pelaez, A; Fooks, A R; Núñez, A

2017-04-12

The recommended screening of rabies in 'suspect' animal cases involves testing fresh brain tissue. The preservation of fresh tissue however can be difficult under field conditions and formalin fixation provides a simple alternative that may allow a confirmatory diagnosis. The occurrence and location of histopathological changes and immunohistochemical (IHC) labelling for rabies in formalin fixed paraffin embedded (FFPE) canine brain is described in samples from 57 rabies suspect cases from Sri-Lanka. The presence of Negri bodies and immunohistochemical detection of rabies virus antigen were evaluated in the cortex, hippocampus, cerebellum and brainstem. The effect of autolysis and artefactual degeneration of the tissue was also assessed. Rabies was confirmed in 53 of 57 (93%) cases by IHC. IHC labelling was statistically more abundant in the brainstem. Negri bodies were observed in 32 of 53 (60.4%) of the positive cases. Although tissue degradation had no effect on IHC diagnosis, it was associated with an inability to detect Negri bodies. In 13 cases, a confirmatory Polymerase chain reaction (PCR) testing for rabies virus RNA was undertaken by extracting RNA from fresh frozen tissue, and also attempted using FFPE samples. PCR detection using fresh frozen samples was in agreement with the IHC results. The PCR method from FFPE tissues was suitable for control material but unsuccessful in our field cases. Histopathological examination of the brain is essential to define the differential diagnoses of behaviour modifying conditions in rabies virus negative cases, but it is unreliable as the sole method for rabies diagnosis, particularly where artefactual change has occurred. Formalin fixation and paraffin embedding does not prevent detection of rabies virus via IHC labelling even where artefactual degeneration has occurred. This could represent a pragmatic secondary assay for rabies diagnosis in the field because formalin fixation can prevent sample degeneration. The brain stem was shown to be the site with most viral immunoreactivity; supporting recommended sampling protocols in favour of improved necropsy safety in the field. PCR testing of formalin fixed tissue may be successful in certain circumstances as an alternative test.

Stable phenotype of B-cell subsets following cryopreservation and thawing of normal human lymphocytes stored in a tissue biobank.

PubMed

Rasmussen, Simon Mylius; Bilgrau, Anders Ellern; Schmitz, Alexander; Falgreen, Steffen; Bergkvist, Kim Steve; Tramm, Anette Mai; Baech, John; Jacobsen, Chris Ladefoged; Gaihede, Michael; Kjeldsen, Malene Krag; Bødker, Julie Støve; Dybkaer, Karen; Bøgsted, Martin; Johnsen, Hans Erik

2015-01-01

Cryopreservation is an acknowledged procedure to store vital cells for future biomarker analyses. Few studies, however, have analyzed the impact of the cryopreservation on phenotyping. We have performed a controlled comparison of cryopreserved and fresh cellular aliquots prepared from individual healthy donors. We studied circulating B-cell subset membrane markers and global gene expression, respectively by multiparametric flow cytometry and microarray data. Extensive statistical analysis of the generated data tested the concept that "overall, there are no phenotypic differences between cryopreserved and fresh B-cell subsets." Subsequently, we performed an uncontrolled comparison of tonsil tissue samples. By multiparametric flow analysis, we documented no significant changes following cryopreservation of subset frequencies or membrane intensity for the differentiation markers CD19, CD20, CD22, CD27, CD38, CD45, and CD200. By gene expression profiling following cryopreservation, across all samples, only 16 out of 18708 genes were significantly up or down regulated, including FOSB, KLF4, RBP7, ANXA1 or CLC, DEFA3, respectively. Implementation of cryopreserved tissue in our research program allowed us to present a performance analysis, by comparing cryopreserved and fresh tonsil tissue. As expected, phenotypic differences were identified, but to an extent that did not affect the performance of the cryopreserved tissue to generate specific B-cell subset associated gene signatures and assign subset phenotypes to independent tissue samples. We have confirmed our working concept and illustrated the usefulness of vital cryopreserved cell suspensions for phenotypic studies of the normal B-cell hierarchy; however, storage procedures need to be delineated by tissue-specific comparative analysis. © 2014 Clinical Cytometry Society.

Stable Phenotype Of B-Cell Subsets Following Cryopreservation and Thawing of Normal Human Lymphocytes Stored in a Tissue Biobank.

PubMed

Rasmussen, Simon Mylius; Bilgrau, Anders Ellern; Schmitz, Alexander; Falgreen, Steffen; Bergkvist, Kim Steve; Tramm, Anette Mai; Baech, John; Jacobsen, Chris Ladefoged; Gaihede, Michael; Kjeldsen, Malene Krag; Bødker, Julie Støve; Dybkaer, Karen; Bøgsted, Martin; Johnsen, Hans Erik

2014-09-20

Background Cryopreservation is an acknowledged procedure to store vital cells for future biomarker analyses. Few studies, however, have analyzed the impact of the cryopreservation on phenotyping. Methods We have performed a controlled comparison of cryopreserved and fresh cellular aliquots prepared from individual healthy donors. We studied circulating B-cell subset membrane markers and global gene expression, respectively by multiparametric flow cytometry and microarray data. Extensive statistical analysis of the generated data tested the concept that "overall, there are phenotypic differences between cryopreserved and fresh B-cell subsets". Subsequently, we performed a consecutive uncontrolled comparison of tonsil tissue samples. Results By multiparametric flow analysis, we documented no significant changes following cryopreservation of subset frequencies or membrane intensity for the differentiation markers CD19, CD20, CD22, CD27, CD38, CD45, and CD200. By gene expression profiling following cryopreservation, across all samples, only 16 out of 18708 genes were significantly up or down regulated, including FOSB, KLF4, RBP7, ANXA1 or CLC, DEFA3, respectively. Implementation of cryopreserved tissue in our research program allowed us to present a performance analysis, by comparing cryopreserved and fresh tonsil tissue. As expected, phenotypic differences were identified, but to an extent that did not affect the performance of the cryopreserved tissue to generate specific B-cell subset associated gene signatures and assign subset phenotypes to independent tissue samples. Conclusions We have confirmed our working concept and illustrated the usefulness of vital cryopreserved cell suspensions for phenotypic studies of the normal B-cell hierarchy; however, storage procedures need to be delineated by tissue specific comparative analysis. © 2014 Clinical Cytometry Society. Copyright © 2014 Clinical Cytometry Society.

A naive Bayes algorithm for tissue origin diagnosis (TOD-Bayes) of synchronous multifocal tumors in the hepatobiliary and pancreatic system.

PubMed

Jiang, Weiqin; Shen, Yifei; Ding, Yongfeng; Ye, Chuyu; Zheng, Yi; Zhao, Peng; Liu, Lulu; Tong, Zhou; Zhou, Linfu; Sun, Shuo; Zhang, Xingchen; Teng, Lisong; Timko, Michael P; Fan, Longjiang; Fang, Weijia

2018-01-15

Synchronous multifocal tumors are common in the hepatobiliary and pancreatic system but because of similarities in their histological features, oncologists have difficulty in identifying their precise tissue clonal origin through routine histopathological methods. To address this problem and assist in more precise diagnosis, we developed a computational approach for tissue origin diagnosis based on naive Bayes algorithm (TOD-Bayes) using ubiquitous RNA-Seq data. Massive tissue-specific RNA-Seq data sets were first obtained from The Cancer Genome Atlas (TCGA) and ∼1,000 feature genes were used to train and validate the TOD-Bayes algorithm. The accuracy of the model was >95% based on tenfold cross validation by the data from TCGA. A total of 18 clinical cancer samples (including six negative controls) with definitive tissue origin were subsequently used for external validation and 17 of the 18 samples were classified correctly in our study (94.4%). Furthermore, we included as cases studies seven tumor samples, taken from two individuals who suffered from synchronous multifocal tumors across tissues, where the efforts to make a definitive primary cancer diagnosis by traditional diagnostic methods had failed. Using our TOD-Bayes analysis, the two clinical test cases were successfully diagnosed as pancreatic cancer (PC) and cholangiocarcinoma (CC), respectively, in agreement with their clinical outcomes. Based on our findings, we believe that the TOD-Bayes algorithm is a powerful novel methodology to accurately identify the tissue origin of synchronous multifocal tumors of unknown primary cancers using RNA-Seq data and an important step toward more precision-based medicine in cancer diagnosis and treatment. © 2017 UICC.

Evaluation of salivary tumor necrosis factor-alpha in patients with the chronic periodontitis: A case-control study.

PubMed

Yousefimanesh, Hojatollah; Maryam, Robati; Mahmoud, Jahangirnezhad; Mehri, Ghafourian Boroujerdnia; Mohsen, Taghipour

2013-11-01

Periodontitis is a chronic infectious disease that leads to inflammation of the tissues supporting the teeth, bone loss, attachment loss progressively. In chronic periodontitis for starting the host response and inflammatory reaction, the presence of the infectious agent is necessary. One of inflammatory factors is tumor necrosis factor-alpha (TNF-α) that appear to be important in the destruction of periodontal tissues that were examined in this study. This study was performed in the laboratory and case-control study. The samples of study collected from 30 individuals with chronic periodontitis and 30 healthy controls that matched for age and sex, together. Unstimulated saliva samples were collected from patients and then TNF-α level were measured by enzyme-linked immunosorbent assay and were compared with the control group. In this study for statistical analysis, Mann-Whitney was used. There were differences in mean salivary concentrations of TNF-α in controls and patients. The average concentration in the case group was 9.1 (pg/ml) and the control group was 8.7 (pg/ml), but there was no significant difference between case and control groups (P > 0.05). The results of this analysis showed no significant relationship between two groups TNF-α concentration. This biomarker can not seem to be a good index to evaluate or predict periodontal disease.

Improving accuracy of DNA diet estimates using food tissue control materials and an evaluation of proxies for digestion bias.

PubMed

Thomas, Austen C; Jarman, Simon N; Haman, Katherine H; Trites, Andrew W; Deagle, Bruce E

2014-08-01

Ecologists are increasingly interested in quantifying consumer diets based on food DNA in dietary samples and high-throughput sequencing of marker genes. It is tempting to assume that food DNA sequence proportions recovered from diet samples are representative of consumer's diet proportions, despite the fact that captive feeding studies do not support that assumption. Here, we examine the idea of sequencing control materials of known composition along with dietary samples in order to correct for technical biases introduced during amplicon sequencing and biological biases such as variable gene copy number. Using the Ion Torrent PGM(©) , we sequenced prey DNA amplified from scats of captive harbour seals (Phoca vitulina) fed a constant diet including three fish species in known proportions. Alongside, we sequenced a prey tissue mix matching the seals' diet to generate tissue correction factors (TCFs). TCFs improved the diet estimates (based on sequence proportions) for all species and reduced the average estimate error from 28 ± 15% (uncorrected) to 14 ± 9% (TCF-corrected). The experimental design also allowed us to infer the magnitude of prey-specific digestion biases and calculate digestion correction factors (DCFs). The DCFs were compared with possible proxies for differential digestion (e.g. fish protein%, fish lipid%) revealing a strong relationship between the DCFs and percent lipid of the fish prey, suggesting prey-specific corrections based on lipid content would produce accurate diet estimates in this study system. These findings demonstrate the value of parallel sequencing of food tissue mixtures in diet studies and offer new directions for future research in quantitative DNA diet analysis. © 2013 John Wiley & Sons Ltd.

Experimentally reduced root-microbe interactions reveal limited plasticity in functional root traits in Acer and Quercus.

PubMed

Lee, Mei-Ho; Comas, Louise H; Callahan, Hilary S

2014-02-01

Interactions between roots and soil microbes are critical components of below-ground ecology. It is essential to quantify the magnitude of root trait variation both among and within species, including variation due to plasticity. In addition to contextualizing the magnitude of plasticity relative to differences between species, studies of plasticity can ascertain if plasticity is predictable and whether an environmental factor elicits changes in traits that are functionally advantageous. To compare functional traits and trait plasticities in fine root tissues with natural and reduced levels of colonization by microbial symbionts, trimmed and surface-sterilized root segments of 2-year-old Acer rubrum and Quercus rubra seedlings were manipulated. Segments were then replanted into satellite pots filled with control or heat-treated soil, both originally derived from a natural forest. Mycorrhizal colonization was near zero in roots grown in heat-treated soil; roots grown in control soil matched the higher colonization levels observed in unmanipulated root samples collected from field locations. Between-treatment comparisons revealed negligible plasticity for root diameter, branching intensity and nitrogen concentration across both species. Roots from treated soils had decreased tissue density (approx. 10-20 %) and increased specific root length (approx. 10-30 %). In contrast, species differences were significant and greater than treatment effects in traits other than tissue density. Interspecific trait differences were also significant in field samples, which generally resembled greenhouse samples. The combination of experimental and field approaches was useful for contextualizing trait plasticity in comparison with inter- and intra-specific trait variation. Findings that root traits are largely species dependent, with the exception of root tissue density, are discussed in the context of current literature on root trait variation, interactions with symbionts and recent progress in standardization of methods for quantifying root traits.

Increased levels of dioxin-like substances in adipose tissue in patients with deep infiltrating endometriosis.

PubMed

Martínez-Zamora, M A; Mattioli, L; Parera, J; Abad, E; Coloma, J L; van Babel, B; Galceran, M T; Balasch, J; Carmona, F

2015-05-01

Are the levels of biologically active and the most toxic dioxin-like substances in adipose tissue of patients with deep infiltrating endometriosis (DIE) higher than in a control group without endometriosis? DIE patients have higher levels of dioxins and polychlorinated biphenyls (PCBs) in adipose tissue compared with controls without endometriosis. Some studies have investigated the levels of dioxin-like substances, in serum samples, in patients with endometriosis, with inconsistent results. Case-control study including two groups of patients. The study group (DIE group) consisted of 30 patients undergoing laparoscopic surgery because of DIE. In all patients, an extensive preoperative work-up was performed including clinical exploration, magnetic resonance imaging (MRI) and transvaginal sonography. All patients with DIE underwent a confirmatory histological study for DIE after surgery. The non-endometriosis control group (control group), included the next consecutive patient undergoing laparoscopic surgery in our center due to adnexal benign gynecological disease (ovarian or tubal procedures other than endometriosis) after each DIE patient, and who did not present any type of endometriosis. During the surgical procedure 1-2 g of adipose tissue from the omentum were obtained. Dioxin-like substances were analyzed in adipose tissue in DIE patients and controls without endometriosis. The total toxic equivalence and concentrations of both dioxins and PCBs were significantly higher in patients with DIE in comparison with the control group (P < 0.05), mainly due to the significantly higher values of the two most toxic dioxins (2,3,7,8-tetrachlorodibenzo-p-dioxin [2,3,7,8-TCDD] and 1,2,3,7,8-pentachlorodibenzo-p-dioxin [1,2,3,7,8-PeCDD]) (P < 0.01 for each compound). The levels of furan 2,3,4,7,8-PeCDF were statistically higher in the DIE group compared with controls. Only four congeners of PCBs had toxic equivalence values and concentrations that were statistically higher in patients with DIE, but these included the most toxic and carcinogenic PCB-126 (PCB-114 P < 0.05; PCB-156 P < 0.05; PCB-189 P = 0.04; PCB-126 P < 0.01). Since few patients were recruited, the study is only exploratory. Our results need to be confirmed in larger and more heterogeneous population studies since environmental and even genetic factors involved in determining dioxins and PCBs widely vary in different countries. Furthermore, the strict eligibility criteria used may preclude generalization of the results to other populations and the surgery-based sampling frame may induce a selection bias. Finally, adipose tissue was obtained only from the omentum, and not from other adipose tissue of the body. Our results suggest a potential role of dioxin-like substances in the pathogenesis of DIE. Further studies are warranted to confirm our findings. None. Not applicable. © The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Bioactive glass 13-93 as a subchondral substrate for tissue-engineered osteochondral constructs: a pilot study.

PubMed

Jayabalan, Prakash; Tan, Andrea R; Rahaman, Mohammed N; Bal, B Sonny; Hung, Clark T; Cook, James L

2011-10-01

Replacement of diseased areas of the joint with tissue-engineered osteochondral grafts has shown potential in the treatment of osteoarthritis. Bioactive glasses are candidates for the osseous analog of these grafts. (1) Does Bioactive Glass 13-93 (BG 13-93) as a subchondral substrate improve collagen and glycosaminoglycan production in a tissue-engineered cartilage layer? (2) Does BG 13-93 as a culture medium supplement increase the collagen and glycosaminoglycan production and improve the mechanical properties in a tissue-engineered cartilage layer? In Study 1, bioactive glass samples (n = 4) were attached to a chondrocyte-seeded agarose layer to form an osteochondral construct, cultured for 6 weeks, and compared to controls. In Study 2, bioactive glass samples (n = 5) were cocultured with cell-seeded agarose for 6 weeks. The cell-seeded agarose layer was exposed to BG 13-93 either continuously or for the first or last 2 weeks in culture or had no exposure. Osteochondral constructs with a BG 13-93 base had improved glycosaminoglycan deposition but less collagen II content. Agarose scaffolds that had a temporal exposure to BG 13-93 within the culture medium had improved mechanical and biochemical properties compared to continuous or no exposure. When used as a subchondral substrate, BG 13-93 did not improve biochemical properties compared to controls. However, as a culture medium supplement, BG 13-93 improved the biochemical and mechanical properties of a tissue-engineered cartilage layer. BG 13-93 may not be suitable in osteochondral constructs but could have potential as a medium supplement for neocartilage formation.

Determination of niclosamide residues in rainbow trout (Oncorhynchus mykiss) and channel catfish (Ictalurus punctatus) fillet tissue by high-performance liquid chromatography

USGS Publications Warehouse

Schreier, Theresa M.; Dawson, V.K.; Cho, Yirang; Spanjers, N.J.; Boogaard, M.A.

2000-01-01

Bayluscide [the ethanolamine salt of niclosamide (NIC)] is a registered piscicide used in combination with 3-(trifluoromethyl)-4-nitrophenol (TFM) to control sea lamprey populations in streams tributary to the Great Lakes. A high-performance liquid chromatography (HPLC) method was developed for the determination of NIC residues in muscle fillet tissues of fish exposed to NIC and TFM during sea lamprey control treatments. NIC was extracted from fortified channel catfish and rainbow trout fillet tissue with a series of acetone extractions and cleaned up on C-18 solid-phase extraction cartridges. NIC concentrations were determined by HPLC with detection at 360 and 335 nm for rainbow trout and catfish, respectively. Recovery of NIC from rainbow trout (n = 7) fortified at 0.04 mu g/g was 77 +/- 6.5% and from channel catfish (n = 7) fortified at 0.02 mu g/g was 113 +/- 11%. NIC detection limit was 0.0107 mu g/g for rainbow trout and 0.0063 mu g/g for catfish. Percent recovery of incurred radioactive residues by this method from catfish exposed to [C-14]NIC was 89.3 +/- 4.1%. Percent recoveries of NIC from fortified storage stability tissue samples for rainbow trout (n = 3) analyzed at 5 and 7.5 month periods were 78 +/- 5.1 and 68 +/- 2.4%, respectively. Percent recoveries of NIC from fortified storage stability tissue samples for channel catfish (n = 3) analyzed at 5 and 7.5 month periods were 88 +/- 13 and 76 +/- 21%, respectively.

Analysis of Lutein, Zeaxanthin, and Meso-Zeaxanthin in the Organs of Carotenoid-Supplemented Chickens.

PubMed

Phelan, David; Prado-Cabrero, Alfonso; Nolan, John M

2018-02-03

The macular carotenoids (i.e., lutein (L), zeaxanthin (Z) and meso -zeaxanthin (MZ)) exhibit anti-inflammatory, antioxidant and optical properties that are believed to support human health and function. Studying the accumulation and distribution of these nutrients in tissues and organs, in addition to the eye, is an important step in understanding how these nutrients might support global human function and health (e.g., heart and brain). Chicken is an appropriate animal model with which to study the accumulation of these carotenoids in organs, as the relevant transport molecules and carotenoid binding proteins for L, Z and MZ are present in both humans and chickens. In this experiment, a sample of 3 chickens that were supplemented with L and MZ diacetate (active group) and a sample of 3 chickens that received a standard diet (control group) were analysed. Both groups were analysed for L, Z and MZ concentrations in the brain, eyes, heart, lung, duodenum/pancreas, jejunum/ileum, kidney and breast tissue. L, Z and MZ were identified in all the organs/tissues analysed from the active group. L and Z were identified in all of the organs/tissues analysed from the control group; while, MZ was identified in the eyes of these animals only. The discovery that MZ is accumulated in the tissues and organs of chickens supplemented with this carotenoid is important, given that it is known that a combination of L, Z and MZ exhibits superior antioxidant capacity when compared to any of these carotenoids in isolation.

Life sciences research in space: The requirement for animal models

NASA Technical Reports Server (NTRS)

Fuller, C. A.; Philips, R. W.; Ballard, R. W.

1987-01-01

Use of animals in NASA space programs is reviewed. Animals are needed because life science experimentation frequently requires long-term controlled exposure to environments, statistical validation, invasive instrumentation or biological tissue sampling, tissue destruction, exposure to dangerous or unknown agents, or sacrifice of the subject. The availability and use of human subjects inflight is complicated by the multiple needs and demands upon crew time. Because only living organisms can sense, integrate and respond to the environment around them, the sole use of tissue culture and computer models is insufficient for understanding the influence of the space environment on intact organisms. Equipment for spaceborne experiments with animals is described.

Serum and tissue angiotensin-converting enzyme in patients with alopecia areata.

PubMed

Fahim, Shabnam; Montazer, Fatemeh; Tohidinik, Hamid Reza; Naraghi, Zahra Safaei; Abedini, Robabeh; Nasimi, Maryam; Ghandi, Narges

2018-03-27

Alopecia areata is an immune-dependent disorder characterized by the interaction of T-lymphocytes with follicular antigens. Recent studies have shown the existence of a local renin-angiotensin system in the skin, where angiotensin-converting enzyme (ACE) plays a role in autoimmunity and inflammation. The objective of this study was to evaluate serum and tissue ACE activity in patients with alopecia areata. This case-control study was conducted on patients with alopecia areata and healthy controls. Serum and tissue ACE activity were assessed and compared between the two groups. Twenty-five alopecia areata patients (60% male, mean age 32.1 ± 9.9 years) and 24 controls (50% male, mean age 37.4 ± 8.8 years) were included. Mean serum ACE activity was 52.1 ± 9 U/L in cases and 55.3 ± 14.7 U/L in controls (P = 0.37). Tissue ACE activity was significantly lower in cases in all parts of the skin i.e. epidermis (P = 0.016), follicular epithelium (P = 0.004), and endothelium (P = 0.037). Among cases, serum ACE activity was significantly higher in patients with more severe disease (P = 0.030), nonpatchy alopecia areata (alopecia universalis; ophiasis, patchy and ophiasis, diffuse) (P = 0.029), and with nail involvement (P = 0.027). The sample size was too small to draw definite conclusions. Further, most of the patients had only mild or moderate alopecia areata. Unlike in some other inflammatory diseases, the tissue level of ACE seems to be significantly lower in alopecia areata compared to normal controls. Serum ACE was significantly higher in patients with more severe disease.

Novel strategies of Raman imaging for brain tumor research.

PubMed

Anna, Imiela; Bartosz, Polis; Lech, Polis; Halina, Abramczyk

2017-10-17

Raman diagnostics and imaging have been shown to be an effective tool for the analysis and discrimination of human brain tumors from normal structures. Raman spectroscopic methods have potential to be applied in clinical practice as they allow for identification of tumor margins during surgery. In this study, we investigate medulloblastoma (grade IV WHO) (n= 5), low-grade astrocytoma (grades I-II WHO) (n =4), ependymoma (n=3) and metastatic brain tumors (n= 1) and the tissue from the negative margins used as normal controls. We compare a high grade medulloblastoma, low grade astrocytoma and non-tumor samples from human central nervous system (CNS) tissue. Based on the properties of the Raman vibrational features and Raman images we provide a real-time feedback method that is label-free to monitor tumor metabolism that reveals reprogramming of biosynthesis of lipids, proteins, DNA and RNA. Our results indicate marked metabolic differences between low and high grade brain tumors. We discuss molecular mechanisms causing these metabolic changes, particularly lipid alterations in malignant medulloblastoma and low grade gliomas that may shed light on the mechanisms driving tumor recurrence thereby revealing new approaches for the treatment of malignant glioma. We have found that the high-grade tumors of central nervous system (medulloblastoma) exhibit enhanced level of β-sheet conformation and down-regulated level of α-helix conformation when comparing against normal tissue. We have found that almost all tumors studied in the paper have increased Raman signals of nucleic acids. This increase can be interpreted as increased DNA/RNA turnover in brain tumors. We have shown that the ratio of Raman intensities I 2930 /I 2845 at 2930 and 2845 cm -1 is a good source of information on the ratio of lipid and protein contents. We have found that the ratio reflects the different lipid and protein contents of cancerous brain tissue compared to the non-tumor tissue. We found that levels of the saturated fatty acids were significantly reduced in the high grade medulloblastoma samples compared with non-tumor brain samples and low grade astrocytoma. Differences were also noted in the n-6/n-3 polyunsaturated fatty acids (PUFA) content between medulloblastoma and non-tumor brain samples. The content of the oleic acid (OA) was significantly smaller in almost all brain high grade brain tumors than that observed in the control samples. It indicates that the fatty acid composition of human brain tumors differs from that found in non-tumor brain tissue. The iodine number N I for the normal brain tissue is 60. For comparison OA has 87, docosahexaenoic acid (DHA) 464, α-linolenic acid (ALA) 274. The high grade tumors have the iodine numbers between that for palmitic acid, stearic acid, arachidic acid (N I =0) and oleic acid (N I =87). Most low grade tumors have N I similar to that of OA. The iodine number for arachidonic acid (AA) (N I =334) is much higher than those observed for all studied samples.

Novel strategies of Raman imaging for brain tumor research

PubMed Central

Anna, Imiela; Bartosz, Polis; Lech, Polis; Halina, Abramczyk

2017-01-01

Raman diagnostics and imaging have been shown to be an effective tool for the analysis and discrimination of human brain tumors from normal structures. Raman spectroscopic methods have potential to be applied in clinical practice as they allow for identification of tumor margins during surgery. In this study, we investigate medulloblastoma (grade IV WHO) (n= 5), low-grade astrocytoma (grades I-II WHO) (n =4), ependymoma (n=3) and metastatic brain tumors (n= 1) and the tissue from the negative margins used as normal controls. We compare a high grade medulloblastoma, low grade astrocytoma and non-tumor samples from human central nervous system (CNS) tissue. Based on the properties of the Raman vibrational features and Raman images we provide a real–time feedback method that is label-free to monitor tumor metabolism that reveals reprogramming of biosynthesis of lipids, proteins, DNA and RNA. Our results indicate marked metabolic differences between low and high grade brain tumors. We discuss molecular mechanisms causing these metabolic changes, particularly lipid alterations in malignant medulloblastoma and low grade gliomas that may shed light on the mechanisms driving tumor recurrence thereby revealing new approaches for the treatment of malignant glioma. We have found that the high-grade tumors of central nervous system (medulloblastoma) exhibit enhanced level of β-sheet conformation and down-regulated level of α-helix conformation when comparing against normal tissue. We have found that almost all tumors studied in the paper have increased Raman signals of nucleic acids. This increase can be interpreted as increased DNA/RNA turnover in brain tumors. We have shown that the ratio of Raman intensities I2930/I2845 at 2930 and 2845 cm-1 is a good source of information on the ratio of lipid and protein contents. We have found that the ratio reflects the different lipid and protein contents of cancerous brain tissue compared to the non-tumor tissue. We found that levels of the saturated fatty acids were significantly reduced in the high grade medulloblastoma samples compared with non-tumor brain samples and low grade astrocytoma. Differences were also noted in the n-6/n-3 polyunsaturated fatty acids (PUFA) content between medulloblastoma and non-tumor brain samples. The content of the oleic acid (OA) was significantly smaller in almost all brain high grade brain tumors than that observed in the control samples. It indicates that the fatty acid composition of human brain tumors differs from that found in non-tumor brain tissue. The iodine number NI for the normal brain tissue is 60. For comparison OA has 87, docosahexaenoic acid (DHA) 464, α-linolenic acid (ALA) 274. The high grade tumors have the iodine numbers between that for palmitic acid, stearic acid, arachidic acid (NI=0) and oleic acid (NI=87). Most low grade tumors have NI similar to that of OA. The iodine number for arachidonic acid (AA) (NI=334) is much higher than those observed for all studied samples. PMID:29156720

Investigation of ultrashort-pulsed laser on dental hard tissue

NASA Astrophysics Data System (ADS)

Uchizono, Takeyuki; Awazu, Kunio; Igarashi, Akihiro; Kato, Junji; Hirai, Yoshito

2007-02-01

Ultrashort-pulsed laser (USPL) can ablate various materials with precious less thermal effect. In laser dentistry, to solve the problem that were the generation of crack and carbonized layer by irradiating with conventional laser such as Er:YAG and CO II laser, USPL has been studied to ablate dental hard tissues by several researchers. We investigated the effectiveness of ablation on dental hard tissues by USPL. In this study, Ti:sapphire laser as USPL was used. The laser parameter had the pulse duration of 130 fsec, 800nm wavelength, 1KHz of repetition rate and the average power density of 90~360W/cm2. Bovine root dentin plates and crown enamel plates were irradiated with USPL at 1mm/sec using moving stage. The irradiated samples were analyzed by SEM, EDX, FTIR and roughness meter. In all irradiated samples, the cavity margin and wall were sharp and steep, extremely. In irradiated dentin samples, the surface showed the opened dentin tubules and no smear layer. The Ca/P ratio by EDX measurement and the optical spectrum by FTIR measurement had no change on comparison irradiated samples and non-irradiated samples. These results confirmed that USPL could ablate dental hard tissue, precisely and non-thermally. In addition, the ablation depths of samples were 10μm, 20μm, and 60μm at 90 W/cm2, 180 W/cm2, and 360 W/cm2, approximately. Therefore, ablation depth by USPL depends on the average power density. USPL has the possibility that can control the precision and non-thermal ablation with depth direction by adjusting the irradiated average power density.

Dietary quebracho tannins are not absorbed, but increase the antioxidant capacity of liver and plasma in sheep.

PubMed

López-Andrés, Patricia; Luciano, Giuseppe; Vasta, Valentina; Gibson, Trevor M; Biondi, Luisa; Priolo, Alessandro; Mueller-Harvey, Irene

2013-08-01

A total of sixteen lambs were divided into two groups and fed two different diets. Of these, eight lambs were fed a control diet (C) and eight lambs were fed the C diet supplemented with quebracho tannins (C+T). The objective of the present study was to assess whether dietary quebracho tannins can improve the antioxidant capacity of lamb liver and plasma and if such improvement is due to a direct transfer of phenolic compounds or their metabolites, to the animal tissues. Feed, liver and plasma samples were purified by solid-phase extraction (SPE) and analysed by liquid chromatography-MS for phenolic compounds. Profisitinidin compounds were identified in the C+T diet. However, no phenolic compounds were found in lamb tissues. The liver and the plasma from lambs fed the C+T diet displayed a greater antioxidant capacity than tissues from lambs fed the C diet, but only when samples were not purified with SPE. Profisetinidin tannins from quebracho seem not to be degraded or absorbed in the gastrointestinal tract. However, they induced antioxidant effects in animal tissues.

Microgravity

NASA Image and Video Library

1998-01-01

Dr. Lisa E. Freed of the Massachusetts Institute of Technology and her colleagues have reported that initially disc-like specimens tend to become spherical in space, demonstrating that tissues can grow and differentiate into distinct structures in microgravity. The Mir Increment 3 (Sept. 16, 1996 - Jan. 22, 1997) samples were smaller, more spherical, and mechanically weaker than Earth-grown control samples. These results demonstrate the feasibility of microgravity tissue engineering and may have implications for long human space voyages and for treating musculoskeletal disorders on earth. The work is sponsored by NASA's Office of Biological and Physical Research. The bioreactor is managed by the Biotechnology Cell Science Program at NASA's Johnson Space Center (JSC). NASA-sponsored bioreactor research has been instrumental in helping scientists to better understand normal and cancerous tissue development. In cooperation with the medical community, the bioreactor design is being used to prepare better models of human colon, prostate, breast and ovarian tumors. Cartilage, bone marrow, heart muscle, skeletal muscle, pancreatic islet cells, liver and kidney are just a few of the normal tissues being cultured in rotating bioreactors by investigators.

Evaluation of residual radioactivity in human tissues associated with weapons testing at the nevada test site. Technical report, 1 June 1981-31 March 1989

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wrenn, M.E.; Singh, N.P.; Paschoa, A.S.

1992-11-01

Residual radioactivity consisting of 239 Pu were measured by radiochemistry and alpha spectrometry in samples of bone and soft tissues from 100 autopsies or surgeries from northern and southwestern Utah and from control areas in Colorado and Pennsylvania. Based upon the isotopic ratio 240 Pu/239 Pu contamination was attributable to atmospheric weapons tests at the Nevada Test Site (NTS). In addition, 110 thyroid tissue samples obtained from tissue blocks made at autopsy of veterans dying at the VA hospital in Salt Lake City in the 1940's and 1950's were measured for 129 I (half life 16 million year) and 127more » I (stable) by neutron activation. The results were analyzed by year of death, by periods before and during atmospheric nuclear weapons testing at the NTS and by origin of usual residence. A model was developed to relate thyroid dose from 131 I to the measured 124/127 I ratios, and thyroid dose estimates were made based upon the measured ratios.« less

A novel approach to non-biased systematic random sampling: a stereologic estimate of Purkinje cells in the human cerebellum.

PubMed

Agashiwala, Rajiv M; Louis, Elan D; Hof, Patrick R; Perl, Daniel P

2008-10-21

Non-biased systematic sampling using the principles of stereology provides accurate quantitative estimates of objects within neuroanatomic structures. However, the basic principles of stereology are not optimally suited for counting objects that selectively exist within a limited but complex and convoluted portion of the sample, such as occurs when counting cerebellar Purkinje cells. In an effort to quantify Purkinje cells in association with certain neurodegenerative disorders, we developed a new method for stereologic sampling of the cerebellar cortex, involving calculating the volume of the cerebellar tissues, identifying and isolating the Purkinje cell layer and using this information to extrapolate non-biased systematic sampling data to estimate the total number of Purkinje cells in the tissues. Using this approach, we counted Purkinje cells in the right cerebella of four human male control specimens, aged 41, 67, 70 and 84 years, and estimated the total Purkinje cell number for the four entire cerebella to be 27.03, 19.74, 20.44 and 22.03 million cells, respectively. The precision of the method is seen when comparing the density of the cells within the tissue: 266,274, 173,166, 167,603 and 183,575 cells/cm3, respectively. Prior literature documents Purkinje cell counts ranging from 14.8 to 30.5 million cells. These data demonstrate the accuracy of our approach. Our novel approach, which offers an improvement over previous methodologies, is of value for quantitative work of this nature. This approach could be applied to morphometric studies of other similarly complex tissues as well.

A practical tissue sampling method using ordinary paper for molecular detection of infectious bursal disease virus RNA by RT-PCR.

PubMed

Maw, Min Thein; Yamaguchi, Tsuyoshi; Kasanga, Christopher J; Terasaki, Kaori; Fukushi, Hideto

2006-12-01

A practical sampling method for bursal tissue using ordinary paper for molecular diagnosis of infectious bursal disease (IBD) was established. IBD virus-infected bursa was directly smeared on chromatography paper, filter paper, or stationery copy paper and was then fixed with absolute ethanol, Tris-HCl-saturated phenol, or phenol:chloroform:isoamyl alcohol (25:24:1). Flinders Technology Associates (FTA) card, which is designed for the collection of biological samples for molecular detection, was also used. After storage at 37 C for up to 30 days, total RNA directly extracted from the tissue fixed on the papers and FTA card were subjected to reverse transcriptase-polymerase chain reaction (RT-PCR) for detection of IBD virus (IBDV) RNA. In addition, the ability of each chemical used in the fixation and the FTA card to inactivate IBDV was evaluated. Regardless of the paper quality, storage period, and fixation method, IBDV RNA was consistently detected in all of the samples. IBDV in the bursal tissue was inactivated with phenol but not with ethanol or the unknown chemicals in FTA card. These results show that ordinary papers sustain the viral RNA, as does FTA card, but phenol fixation is superior to FTA card in inactivating IBDV. The new sampling method using ordinary paper with phenol fixation is safe, inexpensive, simple, and easy, and is thus suitable for conducting a global survey of IBD even where laboratory resources are limited. This practical method should contribute to the control of IBD worldwide.

A novel approach to non-biased systematic random sampling: A stereologic estimate of Purkinje cells in the human cerebellum

PubMed Central

Agashiwala, Rajiv M.; Louis, Elan D.; Hof, Patrick R.; Perl, Daniel P.

2010-01-01

Non-biased systematic sampling using the principles of stereology provides accurate quantitative estimates of objects within neuroanatomic structures. However, the basic principles of stereology are not optimally suited for counting objects that selectively exist within a limited but complex and convoluted portion of the sample, such as occurs when counting cerebellar Purkinje cells. In an effort to quantify Purkinje cells in association with certain neurodegenerative disorders, we developed a new method for stereologic sampling of the cerebellar cortex, involving calculating the volume of the cerebellar tissues, identifying and isolating the Purkinje cell layer and using this information to extrapolate non-biased systematic sampling data to estimate the total number of Purkinje cells in the tissues. Using this approach, we counted Purkinje cells in the right cerebella of four human male control specimens, aged 41, 67, 70 and 84 years, and estimated the total Purkinje cell number for the four entire cerebella to be 27.03, 19.74, 20.44 and 22.03 million cells, respectively. The precision of the method is seen when comparing the density of the cells within the tissue: 266,274, 173,166, 167,603 and 183,575 cells/cm3, respectively. Prior literature documents Purkinje cell counts ranging from 14.8 to 30.5 million cells. These data demonstrate the accuracy of our approach. Our novel approach, which offers an improvement over previous methodologies, is of value for quantitative work of this nature. This approach could be applied to morphometric studies of other similarly complex tissues as well. PMID:18725208

Impact of Neurodegenerative Diseases on Drug Binding to Brain Tissues: From Animal Models to Human Samples.

PubMed

Ugarte, Ana; Corbacho, David; Aymerich, María S; García-Osta, Ana; Cuadrado-Tejedor, Mar; Oyarzabal, Julen

2018-04-19

Drug efficacy in the central nervous system (CNS) requires an additional step after crossing the blood-brain barrier. Therapeutic agents must reach their targets in the brain to modulate them; thus, the free drug concentration hypothesis is a key parameter for in vivo pharmacology. Here, we report the impact of neurodegeneration (Alzheimer's disease (AD) and Parkinson's disease (PD) compared with healthy controls) on the binding of 10 known drugs to postmortem brain tissues from animal models and humans. Unbound drug fractions, for some drugs, are significantly different between healthy and injured brain tissues (AD or PD). In addition, drugs binding to brain tissues from AD and PD animal models do not always recapitulate their binding to the corresponding human injured brain tissues. These results reveal potentially relevant implications for CNS drug discovery.

Diagnosis of Swine Toxoplasmosis by PCR and Genotyping of Toxoplasma gondii from pigs in Henan, Central China.

PubMed

Wang, Haiyan; Zhang, Longxian; Ren, Qinge; Yu, Fuchang; Yang, Yurong

2017-05-31

Toxoplasma gondii, a widely prevalent protozoan parasite, causes serious toxoplasmosis infections in humans and other animals. Among livestock, pigs are susceptible to T. gondii infection. Despite Henan being one of the biggest pig-raising provinces in China, little information exists on the epidemiology of toxoplasmosis in this location. Therefore, we molecularly characterized DNA samples from pigs in Henan. A total of 1647 samples, including 952 from dead piglets, 478 from seriously sick fattening pigs and 217 from abortion sows, were collected from different animal hospitals or pig farms from 10 different cities in Henan (2006-2008). Each pig corresponded to a separate pig farm. DNA was extracted from 3 to 5 g of the most severely affected pig tissue (liver, spleen, lung, hilar lymph nodes and amniotic fluid) after postmortem examination. The presence of the T. gondii B1 gene was detected using nested polymerase chain reactions (PCR). Genotyping was performed directly on DNA from the PCR-positive tissue samples using 11 PCR restriction fragment length polymorphism markers (SAG1, 5'- and 3'-SAG2, alternative SAG2, SAG3, BTUB, GRA6, L358, PK1, c22-8, c29-2, and Apico). Of all samples, thirty-four were positive for the T. gondii B1 gene (2.06%, 95% CI: 1.86%-2.26%) from four cities, including 31 from NanYang city, one (PgXY 1) from Xinyang City, one (PgZZ 1) from Zhengzhou City and one (PgZK1) from Zhoukou City. The prevalence was found to be highest in piglets than in fattening pigs and sows. And the difference was statistically significant (P<0.01). The following 32 samples were genotyped with complete data: 13 hilar lymph node tissue samples, seven liver tissue samples, seven lung tissue samples, four spleen tissue samples, and one amniotic fluid sample. Only one genotype, belonging to ToxoDB Genotype #9, was identified. This is the first large-scale survey molecularly characterizing T. gondii from pigs in Henan. The results of the present study revealed that T. gondii infection is present in swine in Henan and is a potential source of foodborne toxoplasmosis in the investigated areas. Implementation of effective control measures for T. gondii to reduce the chance of zoonotic toxoplasmosis spreading from pig farms may be warranted. The results show that the ToxoDB #9 genotype may be the dominant T. gondii lineage in mainland China. These findings strengthen the limited Chinese T. gondii epidemiology database.

Effects of dietary zinc oxide nanoparticles on growth performance and antioxidative status in broilers.

PubMed

Zhao, Cui-Yan; Tan, Shu-Xian; Xiao, Xi-Yu; Qiu, Xian-Shuai; Pan, Jia-Qiang; Tang, Zhao-Xin

2014-09-01

Broilers in four groups were fed a basal diet supplemented with 60 mg/kg zinc oxide (60-ZnO; control), or 20, 60, or 100 mg/kg ZnO nanoparticles (20-, 60-, and 100-nano-ZnO, respectively). Compared with the controls, after 14 days, birds in the 20- and 60-nano-ZnO groups had significantly greater weight gains and better feed conversion ratios. However, the body weight of birds in the 100-nano-ZnO group was dramatically reduced after 28 days. Relative to the control group, the total antioxidant capability (T-AOC) in serum and liver tissue was significantly higher in the 20-nano-ZnO group at all time points and also significantly higher in the 60- and 100-nano-ZnO groups in serum on days 28 and 35 and in liver tissues on days 21 and 28. Compared with the controls, the activity of copper-zinc superoxide dismutase (Cu-Zn-SOD) was significantly greater in the 60- and 100-nano-ZnO groups in serum on days 28 and 35 and in liver tissues after 21 days. Catalase activity in serum samples was significantly higher in the 20- and 60-nano-ZnO groups relative to the control and 100-nano-ZnO birds, but catalase activity in liver tissue was not affected by different nano-ZnO levels. Malondialdehyde content in serum and liver tissues was significantly reduced in the 20-, 60-, and 100-nano-ZnO groups compared with that in the control group at all time points except day 42. Taken together, our data indicate that appropriate concentration of dietary ZnO nanoparticles improves growth performance and antioxidative capabilities in broilers, and 20 mg/kg nano-ZnO is the optimal concentration.

Pathological effects of cyanobacteria on sea fans in southeast Florida.

PubMed

Kiryu, Y; Landsberg, J H; Peters, E C; Tichenor, E; Burleson, C; Perry, N

2015-07-01

In early August 2008, observations by divers indicated that sea fans, particularly Gorgonia ventalina, Gorgonia flabellum, and Iciligorgia schrammi, were being covered by benthic filamentous cyanobacteria. From August 2008 through January 2009 and again in April 2009, tissue samples from a targeted G. ventalina colony affected by cyanobacteria and from a nearby, apparently healthy (without cyanobacteria) control colony, were collected monthly for histopathological examination. The primary cellular response of the sea fan to overgrowth by cyanobacteria was an increase in the number of acidophilic amoebocytes (with their granular contents dispersed) that were scattered throughout the coenenchyme tissue. Necrosis of scleroblasts and zooxanthellae and infiltration of degranulated amoebocytes were observed in the sea fan surface tissues at sites overgrown with cyanobacteria. Fungal hyphae in the axial skeleton were qualitatively more prominent in cyanobacteria-affected sea fans than in controls. Copyright © 2015 Elsevier Inc. All rights reserved.

MicroRNA Expression in Laser Micro-dissected Breast Cancer Tissue Samples - a Pilot Study.

PubMed

Seclaman, Edward; Narita, Diana; Anghel, Andrei; Cireap, Natalia; Ilina, Razvan; Sirbu, Ioan Ovidiu; Marian, Catalin

2017-10-28

Breast cancer continues to represent a significant public health burden despite outstanding research advances regarding the molecular mechanisms of cancer biology, biomarkers for diagnostics and prognostic and therapeutic management of this disease. The studies of micro RNAs in breast cancer have underlined their potential as biomarkers and therapeutic targets; however most of these studies are still done on largely heterogeneous whole breast tissue samples. In this pilot study we have investigated the expression of four micro RNAs (miR-21, 145, 155, 92) known to be involved in breast cancer, in homogenous cell populations collected by laser capture microdissection from breast tissue section slides. Micro RNA expression was assessed by real time PCR, and associations with clinical and pathological characteristics were also explored. Our results have confirmed previous associations of miR-21 expression with poor prognosis characteristics of breast cancers such as high stage, large and highly proliferative tumors. No statistically significant associations were found with the other micro RNAs investigated, possibly due to the small sample size of our study. Our results also suggest that miR-484 could be a suitable endogenous control for data normalization in breast tissues, these results needing further confirmation by future studies. In summary, our pilot study showed the feasibility of detecting micro RNAs expression in homogenous laser captured microdissected invasive breast cancer samples, and confirmed some of the previously reported associations with poor prognostic characteristics of breast tumors.

Oxidative stress induced by 1.8 GHz radio frequency electromagnetic radiation and effects of garlic extract in rats.

PubMed

Avci, Bahattin; Akar, Ayşegül; Bilgici, Birşen; Tunçel, Özgür Korhan

2012-11-01

We aimed to study the oxidative damage induced by radiofrequency electromagnetic radiation (RF-EMR) emitted by mobile telephones and the protective effect of garlic extract used as an anti-oxidant against this damage. A total of 66 albino Wistar rats were divided into three groups. The first group of rats was given 1.8 GHz, 0.4 W/kg specific absorption rate (SAR) for 1 h a day for three weeks. The second group was given 500 mg/kg garlic extract in addition to RF-EMR. The third group of rats was used as the control group. At the end of the study, blood and brain tissue samples were collected from the rats. After the RF-EMR exposed, the advanced oxidation protein product (AOPP) levels of brain tissue increased compared with the control group (p < 0.001). Garlic administration accompanying the RF-EMR, on the other hand, significantly reduced AOPP levels in brain tissue (p < 0.001). The serum nitric oxide (NO) levels significantly increased both in the first and second group (p < 0.001). However, in the group for which garlic administration accompanied that of RF-EMR, there was no difference in serum NO levels compared with the RF-EMR exposed group (p > 0.05). There was no significant difference among the groups with respect to malondialdehyde (MDA) levels in brain tissue and blood samples (p > 0.05). Similarly, no difference was detected among the groups regarding serum paroxonase (PON) levels (p > 0.05). We did not detect any PON levels in the brain tissue. The exposure of RF-EMR similar to 1.8 GHz Global system for mobile communication (GSM) leads to protein oxidation in brain tissue and an increase in serum NO. We observed that garlic administration reduced protein oxidation in brain tissue and that it did not have any effects on serum NO levels.

Gadolinium Deposition in Human Brain Tissues after Contrast-enhanced MR Imaging in Adult Patients without Intracranial Abnormalities.

PubMed

McDonald, Robert J; McDonald, Jennifer S; Kallmes, David F; Jentoft, Mark E; Paolini, Michael A; Murray, David L; Williamson, Eric E; Eckel, Laurence J

2017-11-01

Purpose To determine whether gadolinium deposits in neural tissues of patients with intracranial abnormalities following intravenous gadolinium-based contrast agent (GBCA) exposure might be related to blood-brain barrier integrity by studying adult patients with normal brain pathologic characteristics. Materials and Methods After obtaining antemortem consent and institutional review board approval, the authors compared postmortem neuronal tissue samples from five patients who had undergone four to 18 gadolinium-enhanced magnetic resonance (MR) examinations between 2005 and 2014 (contrast group) with samples from 10 gadolinium-naive patients who had undergone at least one MR examination during their lifetime (control group). All patients in the contrast group had received gadodiamide. Neuronal tissues from the dentate nuclei, pons, globus pallidus, and thalamus were harvested and analyzed with inductively coupled plasma mass spectrometry (ICP-MS), transmission electron microscopy with energy-dispersive x-ray spectroscopy, and light microscopy to quantify, localize, and assess the effects of gadolinium deposition. Results Tissues from the four neuroanatomic regions of gadodiamide-exposed patients contained 0.1-19.4 μg of gadolinium per gram of tissue in a statistically significant dose-dependent relationship (globus pallidus: ρ = 0.90, P = .04). In contradistinction, patients in the control group had undetectable levels of gadolinium with ICP-MS. All patients had normal brain pathologic characteristics at autopsy. Three patients in the contrast group had borderline renal function (estimated glomerular filtration rate <45 mL/min/1.73 m 2 ) and hepatobiliary dysfunction at MR examination. Gadolinium deposition in the contrast group was localized to the capillary endothelium and neuronal interstitium and, in two cases, within the nucleus of the cell. Conclusion Gadolinium deposition in neural tissues after GBCA administration occurs in the absence of intracranial abnormalities that might affect the permeability of the blood-brain barrier. These findings challenge current understanding of the biodistribution of these contrast agents and their safety. © RSNA, 2017.

Impairment of adipose tissue in Prader-Willi syndrome rescued by growth hormone treatment.

PubMed

Cadoudal, T; Buléon, M; Sengenès, C; Diene, G; Desneulin, F; Molinas, C; Eddiry, S; Conte-Auriol, F; Daviaud, D; Martin, P G P; Bouloumié, A; Salles, J-P; Tauber, M; Valet, P

2014-09-01

Prader-Willi syndrome (PWS) results from abnormalities in the genomic imprinting process leading to hypothalamic dysfunction with an alteration of growth hormone (GH) secretion. PWS is associated with early morbid obesity and short stature which can be efficiently improved with GH treatment. Our aims were to highlight adipose tissue structural and functional impairments in children with PWS and to study the modifications of those parameters on GH treatment. Plasma samples and adipose tissue biopsies were obtained from 23 research centers in France coordinated by the reference center for PWS in Toulouse, France. Lean controls (n=33), non-syndromic obese (n=53), untreated (n=26) and GH-treated PWS (n=43) children were enrolled in the study. Adipose tissue biopsies were obtained during scheduled surgeries from 15 lean control, 7 untreated and 8 GH-treated PWS children. Children with PWS displayed higher insulin sensitivity as shown by reduced glycemia, insulinemia and HOMA-IR compared with non-syndromic obese children. In contrast, plasma inflammatory cytokines such as TNF-α, MCP-1 and IL-8 were increased in PWS. Analysis of biopsies compared with control children revealed decreased progenitor cell content in the stromal vascular fraction of adipose tissue and an impairment of lipolytic response to β-adrenergic agonist in PWS adipocytes. Interestingly, both of these alterations in PWS seem to be ameliorated on GH treatment. Herein, we report adipose tissue dysfunctions in children with PWS which may be partially restored by GH treatment.

Microbiome and Culture Based Analysis of Chronic Rhinosinusitis Compared to Healthy Sinus Mucosa.

PubMed

Koeller, Kerstin; Herlemann, Daniel P R; Schuldt, Tobias; Ovari, Attila; Guder, Ellen; Podbielski, Andreas; Kreikemeyer, Bernd; Olzowy, Bernhard

2018-01-01

The role of bacteria in chronic rhinosinusitis (CRS) is still not well understood. Whole microbiome analysis adds new aspects to our current understanding that is mainly based on isolated bacteria. It is still unclear how the results of microbiome analysis and the classical culture based approaches interrelate. To address this, middle meatus swabs and tissue samples were obtained during sinus surgery in 5 patients with CRS with nasal polyps (CRSwNP), 5 patients with diffuse CRS without nasal polyps (CRSsNP), 5 patients with unilateral purulent maxillary CRS (upm CRS) and 3 patients with healthy sinus mucosa. Swabs were cultured, and associated bacteria were identified. Additionally, parts of each tissue sample also underwent culture approaches, and in parallel DNA was extracted for 16S rRNA gene amplicon-based microbiome analysis. From tissue samples 4.2 ± 1.2 distinct species per patient were cultured, from swabs 5.4 ± 1.6. The most frequently cultured species from the swabs were Propionibacterium acnes, Staphylococcus epidermidis, Corynebacterium spp. and Staphylococcus aureus . The 16S-RNA gene analysis revealed no clear differentiation of the bacterial community of healthy compared to CRS samples of unilateral purulent maxillary CRS and CRSwNP. However, the bacterial community of CRSsNP differed significantly from the healthy controls. In the CRSsNP samples Flavobacterium, Pseudomonas, Pedobacter, Porphyromonas, Stenotrophomonas , and Brevundimonas were significantly enriched compared to the healthy controls. Species isolated from culture did not generally correspond with the most abundant genera in microbiome analysis. Only Fusobacteria, Parvimonas , and Prevotella found in 2 unilateral purulent maxillary CRS samples by the cultivation dependent approach were also found in the cultivation independent approach in high abundance, suggesting a classic infectious pathogenesis of odontogenic origin in these two specific cases. Alterations of the bacterial community might be a more crucial factor for the development of CRSsNP compared to CRSwNP. Further studies are needed to investigate the relation between bacterial community characteristics and the development of CRSsNP.

Microbiome and Culture Based Analysis of Chronic Rhinosinusitis Compared to Healthy Sinus Mucosa

PubMed Central

Koeller, Kerstin; Herlemann, Daniel P. R.; Schuldt, Tobias; Ovari, Attila; Guder, Ellen; Podbielski, Andreas; Kreikemeyer, Bernd; Olzowy, Bernhard

2018-01-01

The role of bacteria in chronic rhinosinusitis (CRS) is still not well understood. Whole microbiome analysis adds new aspects to our current understanding that is mainly based on isolated bacteria. It is still unclear how the results of microbiome analysis and the classical culture based approaches interrelate. To address this, middle meatus swabs and tissue samples were obtained during sinus surgery in 5 patients with CRS with nasal polyps (CRSwNP), 5 patients with diffuse CRS without nasal polyps (CRSsNP), 5 patients with unilateral purulent maxillary CRS (upm CRS) and 3 patients with healthy sinus mucosa. Swabs were cultured, and associated bacteria were identified. Additionally, parts of each tissue sample also underwent culture approaches, and in parallel DNA was extracted for 16S rRNA gene amplicon-based microbiome analysis. From tissue samples 4.2 ± 1.2 distinct species per patient were cultured, from swabs 5.4 ± 1.6. The most frequently cultured species from the swabs were Propionibacterium acnes, Staphylococcus epidermidis, Corynebacterium spp. and Staphylococcus aureus. The 16S-RNA gene analysis revealed no clear differentiation of the bacterial community of healthy compared to CRS samples of unilateral purulent maxillary CRS and CRSwNP. However, the bacterial community of CRSsNP differed significantly from the healthy controls. In the CRSsNP samples Flavobacterium, Pseudomonas, Pedobacter, Porphyromonas, Stenotrophomonas, and Brevundimonas were significantly enriched compared to the healthy controls. Species isolated from culture did not generally correspond with the most abundant genera in microbiome analysis. Only Fusobacteria, Parvimonas, and Prevotella found in 2 unilateral purulent maxillary CRS samples by the cultivation dependent approach were also found in the cultivation independent approach in high abundance, suggesting a classic infectious pathogenesis of odontogenic origin in these two specific cases. Alterations of the bacterial community might be a more crucial factor for the development of CRSsNP compared to CRSwNP. Further studies are needed to investigate the relation between bacterial community characteristics and the development of CRSsNP. PMID:29755418

Bilateral sinus elevation evaluating plasma rich in growth factors technology: a report of five cases.

PubMed

Anitua, Eduardo; Prado, Roberto; Orive, Gorka

2012-03-01

The purpose of this study was to evaluate the potential effects of plasma rich in growth factors (PRGF) technology and its autologous formulations in five consecutive patients in which bilateral sinus lift augmentation was carried out. Five consecutive patients received bilateral sinus floor augmentation. All patients presented a residual bone height of class D (1-3 mm). The effects of PRGF combined with bovine anorganic bone (one side) were compared with the biomaterial alone (contralateral side). The effects of using liquid PRGF to maintain the bone window and autologous fibrin membrane to seal the defect were evaluated. A complete histological and histomorphometrical analysis was performed 5 months after surgery. One patient was excluded from the study as the Schneiderian membrane of the control side was perforated during the surgery. In two patients, the biopsies obtained from the control sides 5 months postsurgery were not acceptable for processing. PRGF technology facilitated the surgical approach of sinus floor elevation. The control area was more inflamed than the area treated with PRGF technology. Patients referred also to an increased sensation of pain in the control area. PRGF-treated samples had more new vital bone than controls. In patient number 1, image processing revealed 21.4% new vital bone in the PRGF area versus 8.4% in the control area, whereas in patient number 2, 28.4% new vital bone was quantified in the PRGF area compared with the 8.2% of the control side. The immunohistochemical processing of the biopsies revealed that the number of blood vessels per square millimeter of connective tissue was 116 vessels in the PRGF sample versus 7 in the control biopsy. These preliminary results suggest that from a practical point of view, PRGF may present a role in reducing tissue inflammation after surgery, increasing new bone formation and promoting the vascularization of bone tissue. © 2010 Wiley Periodicals, Inc.

Determination of scattering properties and damage thresholds in tissue using ultrafast laser ablation

NASA Astrophysics Data System (ADS)

Martin, Chris; Ben-Yakar, Adela

2016-11-01

Ultrafast laser surgery of tissue requires precise knowledge of the tissue's optical properties to control the extent of subsurface ablation. Here, we present a method to determine the scattering lengths, ℓs, and fluence thresholds, Fth, in multilayered and turbid tissue by finding the input energies required to initiate ablation at various depths in each tissue layer. We validated the method using tissue-mimicking phantoms and applied it to porcine vocal folds, which consist of an epithelial (ep) layer and a superficial lamina propia (SLP) layer. Across five vocal fold samples, we found ℓ=51.0±3.9 μm, F=1.78±0.08 J/cm2, ℓ=26.5±1.6 μm, and F=1.14±0.12 J/cm2. Our method can enable personalized determination of tissue optical properties in a clinical setting, leading to less patient-to-patient variability and more favorable outcomes in operations, such as femto-LASIK surgery.

Enhancement of ultraweak photon emission with 3 MHz ultrasonic irradiation on transplanted tumor tissues of mice.

PubMed

Kim, Hongbae; Ahn, Saeyoung; Kim, Jungdae; Soh, Kwang-Sup

2008-07-01

We investigated photon emissions of various bio-samples which were induced by ultrasonic stimulation. It has been reported that ultrasonic stimulations induced the thermal excitation of the bio-tissues. After ultrasonic stimulation, any measurement of photon radiation in the visible spectral range has not been carried out yet. The instruments consisted of electronic devices for an ultrasonic generator of the frequency 3 MHz and a photomultiplier tube (PMT) system counting photons from bio-tissues. The transplanted tumor tissues of mice were prepared for the experiments and their liver and spleen tissues were also used for the controls. It was found that the continuous ultrasonic stimulations with the electrical power 2300 mW induced ultraweak photon emissions from the tumor tissues. The number of induced photon was dependent of the type of the tissues and the stimulation time intervals. The level of photon emission was increased from the mouse tumor exposed to the ultrasonic stimulations, and the changes were discriminated from those of the spleens and livers.

Improved method for analysis of RNA present in long-term preserved thyroid cancer tissue of atomic bomb survivors.

PubMed

Hamatani, Kiyohiro; Eguchi, Hidetaka; Mukai, Mayumi; Koyama, Kazuaki; Taga, Masataka; Ito, Reiko; Hayashi, Yuzo; Nakachi, Kei

2010-01-01

Since many thyroid cancer tissue samples from atomic bomb (A-bomb) survivors have been preserved for several decades as unbuffered formalin-fixed, paraffin-embedded specimens, molecular oncological analysis of such archival specimens is indispensable for clarifying the mechanisms of thyroid carcinogenesis in A-bomb survivors. Although RET gene rearrangements are the most important targets, it is a difficult task to examine all of the 13 known types of RET gene rearrangements with the use of the limited quantity of RNA that has been extracted from invaluable paraffin-embedded tissue specimens of A-bomb survivors. In this study, we established an improved 5' rapid amplification of cDNA ends (RACE) method using a small amount of RNA extracted from archival thyroid cancer tissue specimens. Three archival thyroid cancer tissue specimens from three different patients were used as in-house controls to determine the conditions for an improved switching mechanism at 5' end of RNA transcript (SMART) RACE method; one tissue specimen with RET/PTC1 rearrangement and one with RET/PTC3 rearrangement were used as positive samples. One other specimen, used as a negative sample, revealed no detectable expression of the RET gene tyrosine kinase domain. We established a 5' RACE method using an amount of RNA as small as 10 ng extracted from long-term preserved, unbuffered formalin-fixed, paraffin-embedded thyroid cancer tissue by application of SMART technology. This improved SMART RACE method not only identified common RET gene rearrangements, but also isolated a clone containing a 93-bp insert of rare RTE/PTC8 in RNA extracted from formalin-fixed, paraffin-embedded thyroid cancer specimens from one A-bomb survivor who had been exposed to a high radiation dose. In addition, in the papillary thyroid cancer of another high-dose A-bomb survivor, this method detected one novel type of RET gene rearrangement whose partner gene is acyl coenzyme A binding domain 5, located on chromosome 10p. We conclude that our improved SMART RACE method is expected to prove useful in molecular analyses using archival formalin-fixed, paraffin-embedded tissue samples of limited quantity.

Life Sciences Research Facility automation requirements and concepts for the Space Station

NASA Technical Reports Server (NTRS)

Rasmussen, Daryl N.

1986-01-01

An evaluation is made of the methods and preliminary results of a study on prospects for the automation of the NASA Space Station's Life Sciences Research Facility. In order to remain within current Space Station resource allocations, approximately 85 percent of planned life science experiment tasks must be automated; these tasks encompass specimen care and feeding, cage and instrument cleaning, data acquisition and control, sample analysis, waste management, instrument calibration, materials inventory and management, and janitorial work. Task automation will free crews for specimen manipulation, tissue sampling, data interpretation and communication with ground controllers, and experiment management.

[Experimental study on the treatment of serious soft tissue injuries with strengthening the spleen and replenishing qi].

PubMed

Chen, Xun-wen; Zhu, Yong-zhan; Chen, Zhi-wei; Wu, Zheng-jie; He, Li-lei

2008-09-01

To study the effects of Chinese drugs based on strengthening the spleen and replenishing qi treatment rule on neoformative capillaries and fibroblast during the soft tissue repair after serious trauma in rats, so as to explore the biological basis of the TCM theory "the spleen dominate extremities and muscles" applied to the treatment of soft tissue injuries. The model rats were established by bleeding from femoral artery and lancing method, and the rats were randomly divided into the control group, strengthening the spleen group and activating blood and resolving stasis group. The samples were got from the tissue of the wounded area at the 5th, 10th and 15th days after oral administration of the traditional Chinese medicine. After fixation and section, the tissues were stained by CD31 and PCNA staining. The amount of the capillaries and fibroblasts in the tissue of the wounded area were observed through multi-purpose microscope (ZEISS Axioskop2). Quantitative analysis was carried out on Image-ProPlus image analyzer. The amount of the capillaries and fibroblasts in the wounded tissue in the strengthening the spleen group were larger than that in the control group at the 5th, 10th and 15th day. And the proliferation speed of capillaries and fibroblasts was faster than those in the control group or the activating blood and resolving stasis group. The Chinese drugs according to strengthening the spleen and replenishing qi treatment rule were effective to promote growth of the granulation tissue and facilitate healing of the wounded area. And it has better effect than the treatment of promoting blood circulation and removing stasis.

Identification of differentially expressed genes induced by energy restriction using annealing control primer system from the liver and adipose tissues of broilers.

PubMed

Wang, J W; Chen, W; Kang, X T; Huang, Y Q; Tian, Y D; Wang, Y B

2012-04-01

Female Arbor Acre broilers were divided into 2 groups at 18 d of age. One group of chickens had free access to feed (AL), and the other group of chickens had 30% energy restriction (ER). Adipose and hepatic RNA samples were collected at 48 d of age. We employed an accurate reverse-transcription (RT) PCR method that involves annealing control primers to identify the differentially expressed genes (DEG) between ER and AL groups. Using 20 annealing control primers, 43 differentially expressed bands (40 downregulated and 3 upregulated in the ER group) were detected from the hepatic tissue, whereas no differentially expressed bands were detected from the adipose tissue. It seems that energy restriction could induce more DEG in hepatic tissue than that in adipose tissue and could result in more gene-expression downregulation in hepatic tissue. Eight DEG (6 known and 2 unknown genes) were gained from hepatic tissue and confirmed by RT-PCR, which were all supported by released expressed sequence tag sequences. Their expressions were all downregulated by energy restriction in hepatic tissues. Six known genes are RPL7, RPLP1, FBXL12, ND1, ANTXR2, and SLC22A18, respectively, which seem to play essential roles in the protein translation, energy metabolism, and tumor inhibition. The alterations of gene expression in 3 selected genes, including ND1 (P < 0.01), FBXL12 (P < 0.01), and RPLP1 (P < 0.05), were supported by real-time quantitative RT-PCR reaction. Our data provide new insights on the metabolic state of broilers changed by energy restriction.

Expression of SLP-2 gene and CCBE1 are associated with prognosis of rectal cancer.

PubMed

Zhang, L; Liu, F-J

2017-03-01

This study aims to investigate the clinical significance of SLP-2 gene for patients with rectal cancer. To analyze the effect of CCBE1 (Collagen and calcium-binding EGF domain-containing protein 1) on rectal cancer tissue and lymph vessels of para-carcinoma tissue. A total of 50 samples of rectal cancer tissues were enrolled in the experimental group, confirmed by pathological examination. 50 samples of para-carcinoma normal tissues were collected as control group. Protein expression of SLP-2 and CCBE1 was examined with immunohistochemical staining. mRNA expression of SLP-2 was examined with RT-PCR. Lymphatic vessel density (LVD) was evaluated with LYVE-1 immunohistochemical staining. Correlation analysis was performed to assess the relationship between patient survival data and clinical pathological features of rectal cancer. Immunohistochemical staining showed that, compared with the control group, a positive expression rate of SLP-2 in the experimental group was significantly higher (68.0% vs. 24.0%, p<0.05), and mRNA of SLP-2 was also significantly increased (p<0.05). Compared with the control group, protein expression of CCBE1 in the experimental group was significantly higher (p<0.05). Moreover, the expression level of SLP-2 was remarkably associated with TNM classification and lymphatic metastasis. Further analysis demonstrated that a positive expression of CCBE1 was associated with lymphatic metastasis, LVD and Ducks classification, and had a negative correlation with survival rate. Increased expression of SLP-2 promoted the formation of lymph vessels and exacerbated lymphatic metastasis of rectal cancer via up-regulating CCBE1. As a risk factor related to lymphatic metastasis, CCBE1 could be a novel biomarker for diagnosis and prognosis of rectal cancer.

Investigations on the potential of a low power diode pumped Er:YAG laser system for oral surgery

NASA Astrophysics Data System (ADS)

Stock, Karl; Wurm, Holger; Hausladen, Florian; Wagner, Sophia; Hibst, Raimund

2015-02-01

Flash lamp pumped Er:YAG-lasers are used in clinical practice for dental applications successfully. As an alternative, several diode pumped Er:YAG laser systems (Pantec Engineering AG) become available, with mean laser power of 2W, 15W, and 30W. The aim of the presented study is to investigate the potential of the 2W Er:YAG laser system for oral surgery. At first an appropriate experimental set-up was realized with a beam delivery and both, a focusing unit for non-contact tissue cutting and a fiber tip for tissue cutting in contact mode. In order to produce reproducible cuts, the samples (porcine gingiva) were moved by a computer controlled translation stage. On the fresh samples cutting depth and quality were determined by light microscopy. Afterwards histological sections were prepared and microscopically analyzed regarding cutting depth and thermal damage zone. The experiments show that low laser power ≤ 2W is sufficient to perform efficient oral soft tissue cutting with cut depth up to 2mm (sample movement 2mm/s). The width of the thermal damage zone can be controlled by the irradiation parameters within a range of about 50μm to 110μm. In general, thermal injury is more pronounced using fiber tips in contact mode compared to the focused laser beam. In conclusion the results reveal that even the low power diode pumped Er:YAG laser is an appropriate tool for oral surgery.

Preliminary evaluation of several disinfection/sterilization techniques for use with microdialysis probes.

PubMed

Huff, Jacquelyn K; Bresnahan, James F; Davies, Malonne I

2003-06-06

This study evaluated the suitability of some disinfection and sterilization methods for use with microdialysis probes. Disinfection or sterilization should minimize the tissue inflammatory reaction and improve the long-term health of rats on study and ensure the quality of data obtained by microdialysis sampling. Furthermore, the treatment should not negatively impact probe integrity or sampling performance. The techniques chosen for evaluation included two disinfection methods (70% ethanol and a commercial contact lens solution) and two sterilization methods (hydrogen peroxide plasma, and e-beam radiation). Linear microdialysis probes treated by these processes were compared to untreated probes removed from the manufacturer's packaging as if sterile (the control group). The probes were aseptically implanted in the livers of rats and monitored for 72 hours. The parameters chosen to evaluate probe performance were relative sample mass recovery and the relative in vivo extraction efficiency of the probe for caffeine. Post mortem bacterial counts and histopathology examination of liver tissue were also conducted. The probes remained intact and functional for the entire study period. The methods tested did not acutely alter the probes although hydrogen peroxide plasma and contact lens solution groups showed reduced extraction efficiencies. Minimal tissue damage was observed surrounding the probes and acute inflammatory reaction was mild to moderate. Low numbers of bacterial colonies from the implantation sites indicates that the health of animals in this study was not impaired. This was also true for the control group (untreated probe).

Iodine-131 dose-dependent gene expression: alterations in both normal and tumour thyroid tissues of post-Chernobyl thyroid cancers.

PubMed

Abend, M; Pfeiffer, R M; Ruf, C; Hatch, M; Bogdanova, T I; Tronko, M D; Hartmann, J; Meineke, V; Mabuchi, K; Brenner, A V

2013-10-15

A strong, consistent association between childhood irradiation and subsequent thyroid cancer provides an excellent model for studying radiation carcinogenesis. We evaluated gene expression in 63 paired RNA specimens from frozen normal and tumour thyroid tissues with individual iodine-131 (I-131) doses (0.008-8.6 Gy, no unirradiated controls) received from Chernobyl fallout during childhood (Ukrainian-American cohort). Approximately half of these randomly selected samples (32 tumour/normal tissue RNA specimens) were hybridised on 64 whole-genome microarrays (Agilent, 4 × 44 K). Associations between I-131 dose and gene expression were assessed separately in normal and tumour tissues using Kruskal-Wallis and linear trend tests. Of 155 genes significantly associated with I-131 after Bonferroni correction and with ≥2-fold increase per dose category, we selected 95 genes. On the remaining 31 RNA samples these genes were used for validation purposes using qRT-PCR. Expression of eight genes (ABCC3, C1orf9, C6orf62, FGFR1OP2, HEY2, NDOR1, STAT3, and UCP3) in normal tissue and six genes (ANKRD46, CD47, HNRNPH1, NDOR1, SCEL, and SERPINA1) in tumour tissue was significantly associated with I-131. PANTHER/DAVID pathway analyses demonstrated significant over-representation of genes coding for nucleic acid binding in normal and tumour tissues, and for p53, EGF, and FGF signalling pathways in tumour tissue. The multistep process of radiation carcinogenesis begins in histologically normal thyroid tissue and may involve dose-dependent gene expression changes.

Downregulated Kynurenine 3-Monooxygenase Gene Expression and Enzyme Activity in Schizophrenia and Genetic Association With Schizophrenia Endophenotypes

PubMed Central

Wonodi, Ikwunga; Stine, O. Colin; Sathyasaikumar, Korrapati V.; Roberts, Rosalinda C.; Mitchell, Braxton D.; Hong, L. Elliot; Kajii, Yasushi; Thaker, Gunvant K.; Schwarcz, Robert

2013-01-01

Context Kynurenic acid, a metabolite of the kynurenine pathway of tryptophan degradation, is an antagonist at N-methyl-d-aspartate and α7 nicotinic acetylcholine receptors and modulates glutamate, dopamine, and acetylcholine signaling. Cortical kynurenic acid concentrations are elevated in the brain and cerebrospinal fluid of schizophrenia patients. The proximal cause may be an impairment of kynurenine 3-monooxygenase (KMO), a rate-limiting enzyme at the branching point of the kynurenine pathway. Objectives To examine KMO messenger RNA expression and KMO enzyme activity in postmortem tissue from the frontal eye field (FEF; Brodmann area 6) obtained from schizophrenia individuals compared with healthy control individuals and to explore the relationship between KMO single-nucleotide polymorphisms and schizophrenia oculomotor endophenotypes. Design Case-control postmortem and clinical study. Setting Maryland Brain Collection, outpatient clinics. Participants Postmortem specimens from schizophrenia patients (n=32) and control donors (n=32) and a clinical sample of schizophrenia patients (n=248) and healthy controls (n=228). Main Outcome Measures Comparison of quantitative KMO messenger RNA expression and KMO enzyme activity in postmortem FEF tissue between schizophrenia patients and controls and association of KMO single-nucleotide polymorphisms with messenger RNA expression in postmortem FEF and schizophrenia and oculomotor endophenotypes (ie, smooth pursuit eye movements and oculomotor delayed response). Results In postmortem tissue, we found a significant and correlated reduction in KMO gene expression and KMO enzyme activity in the FEF in schizophrenia patients. In the clinical sample, KMO rs2275163 was not associated with a diagnosis of schizophrenia but showed modest effects on predictive pursuit and visuospatial working memory endophenotypes. Conclusion Our results provide converging lines of evidence implicating reduced KMO activity in the etiopathophysiology of schizophrenia and related neurocognitive deficits. PMID:21727251

Downregulated kynurenine 3-monooxygenase gene expression and enzyme activity in schizophrenia and genetic association with schizophrenia endophenotypes.

PubMed

Wonodi, Ikwunga; Stine, O Colin; Sathyasaikumar, Korrapati V; Roberts, Rosalinda C; Mitchell, Braxton D; Hong, L Elliot; Kajii, Yasushi; Thaker, Gunvant K; Schwarcz, Robert

2011-07-01

Kynurenic acid, a metabolite of the kynurenine pathway of tryptophan degradation, is an antagonist at N-methyl-d-aspartate and α7 nicotinic acetylcholine receptors and modulates glutamate, dopamine, and acetylcholine signaling. Cortical kynurenic acid concentrations are elevated in the brain and cerebrospinal fluid of schizophrenia patients. The proximal cause may be an impairment of kynurenine 3-monooxygenase (KMO), a rate-limiting enzyme at the branching point of the kynurenine pathway. To examine KMO messenger RNA expression and KMO enzyme activity in postmortem tissue from the frontal eye field (FEF; Brodmann area 6) obtained from schizophrenia individuals compared with healthy control individuals and to explore the relationship between KMO single-nucleotide polymorphisms and schizophrenia oculomotor endophenotypes. Case-control postmortem and clinical study. Maryland Brain Collection, outpatient clinics. Postmortem specimens from schizophrenia patients (n = 32) and control donors (n = 32) and a clinical sample of schizophrenia patients (n = 248) and healthy controls (n = 228). Comparison of quantitative KMO messenger RNA expression and KMO enzyme activity in postmortem FEF tissue between schizophrenia patients and controls and association of KMO single-nucleotide polymorphisms with messenger RNA expression in postmortem FEF and schizophrenia and oculomotor endophenotypes (ie, smooth pursuit eye movements and oculomotor delayed response). In postmortem tissue, we found a significant and correlated reduction in KMO gene expression and KMO enzyme activity in the FEF in schizophrenia patients. In the clinical sample, KMO rs2275163 was not associated with a diagnosis of schizophrenia but showed modest effects on predictive pursuit and visuospatial working memory endophenotypes. Our results provide converging lines of evidence implicating reduced KMO activity in the etiopathophysiology of schizophrenia and related neurocognitive deficits.

Enantiomer fractions of polychlorinated biphenyls in three selected Standard Reference Materials.

PubMed

Morrissey, Joshua A; Bleackley, Derek S; Warner, Nicholas A; Wong, Charles S

2007-01-01

The enantiomer composition of six chiral polychlorinated biphenyls (PCBs) were measured in three different certified Standard Reference Materials (SRMs) from the US National Institute of Standards and Technology (NIST): SRM 1946 (Lake Superior fish tissue), SRM 1939a (PCB Congeners in Hudson River Sediment), and SRM 2978 (organic contaminants in mussel tissue--Raritan Bay, New Jersey) to aid in quality assurance/quality control methodologies in the study of chiral pollutants in sediments and biota. Enantiomer fractions (EFs) of PCBs 91, 95, 136, 149, 174, and 183 were measured using a suite of chiral columns by gas chromatography/mass spectrometry. Concentrations of target analytes were in agreement with certified values. Target analyte EFs in reference materials were measured precisely (<2% relative standard deviation), indicating the utility of SRM in quality assurance/control methodologies for analyses of chiral compounds in environmental samples. Measured EFs were also in agreement with previously published analyses of similar samples, indicating that similar enantioselective processes were taking place in these environmental matrices.

MMP-7 and TIMP-1, new targets in predicting poor wound healing in apical periodontitis.

PubMed

Letra, Ariadne; Ghaneh, Ghazaleh; Zhao, Min; Ray, Herbert; Francisconi, Carolina Favaro; Garlet, Gustavo Pompermaier; Silva, Renato Menezes

2013-09-01

Matrix metalloproteinases (MMPs) and the tissue inhibitors of metalloproteinases (TIMPs) are strongly associated with tissue destruction because of inflammation. In this study, we investigated the expression of MMPs and TIMPs messenger RNA and protein levels in apical periodontitis lesions. Tissue samples from patients presenting clinical signs of chronic apical abscess (CAA) or asymptomatic apical periodontitis (AAP) were collected postoperatively and used for gene expression analysis of MMP-2, -3, -7, -9, -14, -16, and -25; TIMP-1; and TIMP-2 in real-time polymerase chain reaction. Immunohistochemistry was also performed to detect the expression of MMP-7 and TIMP-1 proteins. Lastly, U-937 cells were induced to terminal differentiation into macrophages, infected with purified Escherichia coli lipopolysaccharide, and assessed for the expression of MMP-7 and TIMP-1 using immunocytochemistry and confocal microscopy. Significantly higher messenger RNA levels were found for all genes in AAP and CAA samples when compared with healthy control samples (P < .001). AAP cases exhibited significantly higher TIMP-1 when compared with CAA cases, whereas CAA cases showed higher MMP-2, MMP-7, and MMP-9 messenger RNA levels (P < .05). We also detected positive the expression of MMP-7 and TIMP-1 proteins in the tissue samples. The expression of both MMP-7 and TIMP-1 were increased in lipopolysaccharide-stimulated cells compared with nonstimulated cells and appear to colocalize in the Golgi apparatus. MMPs appear to have an influential role in CAA cases in which ongoing tissue destruction is observed. TIMPs are preferentially associated with AAP, perhaps as a subsequent defense mechanism against excessive destruction. Taken together, our findings implicate MMP and TIMP molecules in the dynamics of inflammatory periapical lesion development. Copyright © 2013 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Concordance of anaplastic lymphoma kinase (ALK) gene rearrangements between circulating tumor cells and tumor in non-small cell lung cancer

PubMed Central

Lim, Tony KH; Tan, Daniel Shao-Weng; Chua, Yong Wei; Ang, Mei Kim; Pang, Brendan; Lim, Chwee Teck; Takano, Angela; Lim, Alvin Soon-Tiong; Leong, Man Chun; Lim, Wan-Teck

2016-01-01

Anaplastic lymphoma kinase (ALK) gene rearrangement in non-small cell lung cancer (NSCLC) is routinely evaluated by fluorescent in-situ hybridization (FISH) testing on biopsy tissues. Testing can be challenging however, when suitable tissue samples are unavailable. We examined the relevance of circulating tumor cells (CTC) as a surrogate for biopsy-based FISH testing. We assessed paired tumor and CTC samples from patients with ALK rearranged lung cancer (n = 14), ALK-negative lung cancer (n = 12), and healthy controls (n = 5) to derive discriminant CTC counts, and to compare ALK rearrangement patterns. Blood samples were enriched for CTCs to be used for ALK FISH testing. ALK-positive CTCs counts were higher in ALK-positive NSCLC patients (3–15 cells/1.88 mL of blood) compared with ALK-negative NSCLC patients and healthy donors (0–2 cells/1.88 mL of blood). The latter range was validated as the ‘false positive’ cutoff for ALK FISH testing of CTCs. ALK FISH signal patterns observed on tumor biopsies were recapitulated in CTCs in all cases. Sequential CTC counts in an index case of lung cancer with no evaluable tumor tissue treated with crizotinib showed six, three and eleven ALK-positive CTCs per 1.88 mL blood at baseline, partial response and post-progression time points, respectively. Furthermore, ALK FISH rearrangement suggestive of gene copy number increase was observed in CTCs following progression. Recapitulation of ALK rearrangement patterns in the tumor on CTCs, suggested that CTCs might be used to complement tissue-based ALK testing in NSCLC to guide ALK-targeted therapy when suitable tissue biopsy samples are unavailable for testing. PMID:26993609

Concordance of anaplastic lymphoma kinase (ALK) gene rearrangements between circulating tumor cells and tumor in non-small cell lung cancer.

PubMed

Tan, Chye Ling; Lim, Tse Hui; Lim, Tony Kh; Tan, Daniel Shao-Weng; Chua, Yong Wei; Ang, Mei Kim; Pang, Brendan; Lim, Chwee Teck; Takano, Angela; Lim, Alvin Soon-Tiong; Leong, Man Chun; Lim, Wan-Teck

2016-04-26

Anaplastic lymphoma kinase (ALK) gene rearrangement in non-small cell lung cancer (NSCLC) is routinely evaluated by fluorescent in-situ hybridization (FISH) testing on biopsy tissues. Testing can be challenging however, when suitable tissue samples are unavailable. We examined the relevance of circulating tumor cells (CTC) as a surrogate for biopsy-based FISH testing. We assessed paired tumor and CTC samples from patients with ALK rearranged lung cancer (n = 14), ALK-negative lung cancer (n = 12), and healthy controls (n = 5) to derive discriminant CTC counts, and to compare ALK rearrangement patterns. Blood samples were enriched for CTCs to be used for ALK FISH testing. ALK-positive CTCs counts were higher in ALK-positive NSCLC patients (3-15 cells/1.88 mL of blood) compared with ALK-negative NSCLC patients and healthy donors (0-2 cells/1.88 mL of blood). The latter range was validated as the 'false positive' cutoff for ALK FISH testing of CTCs. ALK FISH signal patterns observed on tumor biopsies were recapitulated in CTCs in all cases. Sequential CTC counts in an index case of lung cancer with no evaluable tumor tissue treated with crizotinib showed six, three and eleven ALK-positive CTCs per 1.88 mL blood at baseline, partial response and post-progression time points, respectively. Furthermore, ALK FISH rearrangement suggestive of gene copy number increase was observed in CTCs following progression. Recapitulation of ALK rearrangement patterns in the tumor on CTCs, suggested that CTCs might be used to complement tissue-based ALK testing in NSCLC to guide ALK-targeted therapy when suitable tissue biopsy samples are unavailable for testing.

Tissue and serum expression of TGM-3 may be prognostic marker in patients of oral squamous cell carcinoma undergoing chemo-radiotherapy.

PubMed

Nayak, Seema; Bhatt, M L B; Goel, Madhu Mati; Gupta, Seema; Mahdi, Abbas Ali; Mishra, Anupam; Mehrotra, Divya

2018-01-01

Radioresistance is one of the main determinants of treatment outcome in oral squamous cell carcinoma (OSCC), but its prediction is difficult. Several authors aimed to establish radioresistant OSCC cell lines to identify genes with altered expression in response to radioresistance. The development of OSCC is a multistep carcinogenic process that includes activation of several oncogenes and inactivation of tumour suppressor genes. TGM-3 is a tumour suppressor gene and contributes to carcinogenesis process. The aim of this study was to estimate serum and tissue expression of TGM-3 and its correlation with clinico-pathological factors and overall survival in patients of OSCC undergoing chemo-radiotherapy. Tissue expression was observed in formalin fixed tissue biopsies of 96 cases of OSCC and 32 healthy controls were subjected to immunohistochemistry (IHC) by using antibody against TGM-3 and serum level was estimated by ELISA method. mRNA expression was determined by using Real-Time PCR. Patients were followed for 2 year for chemo radiotherapy response. In OSCC, 76.70% cases and in controls 90.62% were positive for TGM-3 IHC expression. TGM-3 expression was cytoplasmic and nuclear staining expressed in keratinized layer, stratum granulosum and stratum spinosum in controls and tumour cells. Mean serum TGM-3 in pre chemo-radiotherapy OSCC cases were 1304.83±573.55, post chemo-radiotherapy samples were 1530.64±669.33 and controls were 1869.16±1377.36, but difference was significant in pre chemo-radiotherapy samples as compared to controls (p<0.018). This finding was also confirmed by real- time PCR analysis in which down regulation (-7.92 fold change) of TGM-3 in OSCC as compared to controls. TGM-3 expression was significantly associated with response to chemo-radiotherapy treatment (p<0.007) and overall survival (p<0.015). Patents having higher level of TGM-3 expression have good response to chemo-radiotherapy and also have better overall survival. TGM-3 may serve as a candidate biomarker for responsiveness to chemo-radiotherapy treatment in OSCC patients.

Microbiome Heterogeneity Characterizing Intestinal Tissue and Inflammatory Bowel Disease Phenotype.

PubMed

Tyler, Andrea D; Kirsch, Richard; Milgrom, Raquel; Stempak, Joanne M; Kabakchiev, Boyko; Silverberg, Mark S

2016-04-01

Inflammatory bowel disease has been associated with differential abundance of numerous organisms when compared to healthy controls (HCs); however, few studies have investigated variability in the microbiome across intestinal locations and how this variability might be related to disease location and phenotype. In this study, we have analyzed the microbiome of a large cohort of individuals recruited at Mount Sinai Hospital in Toronto, Canada. Biopsies were taken from subjects with Crohn's disease, ulcerative colitis, and HC, and also individuals having undergone ileal pouch-anal anastomosis for treatment of ulcerative colitis or familial adenomatous polyposis. Microbial 16S rRNA was sequenced using the Illumina MiSeq platform. We observed a great deal of variability in the microbiome characterizing different sampling locations. Samples from pouch and afferent limb were comparable in microbial composition. When comparing sigmoid and terminal ileum samples, more differences were observed. The greatest number of differentially abundant microbes was observed when comparing either pouch or afferent limb samples to sigmoid or terminal ileum. Despite these differences, we were able to observe modest microbial variability between inflammatory bowel disease phenotypes and HCs, even when controlling for sampling location and additional experimental factors. Most detected associations were observed between HCs and Crohn's disease, with decreases in specific genera in the families Ruminococcaceae and Lachnospiraceae characterizing tissue samples from individuals with Crohn's disease. This study highlights important considerations when analyzing the composition of the microbiome and also provides useful insight into differences in the microbiome characterizing these seemingly related phenotypes.

In-depth proteomic profiling of left ventricular tissues in human end-stage dilated cardiomyopathy.

PubMed

Liu, Shanshan; Xia, Yan; Liu, Xiaohui; Wang, Yi; Chen, Zhangwei; Xie, Juanjuan; Qian, Juying; Shen, Huali; Yang, Pengyuan

2017-07-18

Dilated cardiomyopathy (DCM) is caused by reduced left ventricular (LV) myocardial function, which is one of the most common causes of heart failure (HF). We performed iTRAQ-coupled 2D-LC-MS/MS to profile the cardiac proteome of LV tissues from healthy controls and patients with end-stage DCM. We identified 4263 proteins, of which 125 were differentially expressed in DCM tissues compared to LV controls. The majority of these were membrane proteins related to cellular junctions and neuronal metabolism. In addition, these proteins were involved in membrane organization, mitochondrial organization, translation, protein transport, and cell death process. Four key proteins involved in the cell death process were also detected by western blotting, indicated that cell death was activated in DCM tissues. Furthermore, S100A1 and eEF2 were enriched in the "cellular assembly and organization" and "cell cycle" networks, respectively. We verified decreases in these two proteins in end-stage DCM LV samples through multiple reaction monitoring (MRM). These observations demonstrate that our understanding of the mechanisms underlying DCM can be deepened through comparison of the proteomes of normal LV tissues with that from end-stage DCM in humans.

Identification and validation of differentially expressed transcripts by RNA-sequencing of formalin-fixed, paraffin-embedded (FFPE) lung tissue from patients with Idiopathic Pulmonary Fibrosis.

PubMed

Vukmirovic, Milica; Herazo-Maya, Jose D; Blackmon, John; Skodric-Trifunovic, Vesna; Jovanovic, Dragana; Pavlovic, Sonja; Stojsic, Jelena; Zeljkovic, Vesna; Yan, Xiting; Homer, Robert; Stefanovic, Branko; Kaminski, Naftali

2017-01-12

Idiopathic Pulmonary Fibrosis (IPF) is a lethal lung disease of unknown etiology. A major limitation in transcriptomic profiling of lung tissue in IPF has been a dependence on snap-frozen fresh tissues (FF). In this project we sought to determine whether genome scale transcript profiling using RNA Sequencing (RNA-Seq) could be applied to archived Formalin-Fixed Paraffin-Embedded (FFPE) IPF tissues. We isolated total RNA from 7 IPF and 5 control FFPE lung tissues and performed 50 base pair paired-end sequencing on Illumina 2000 HiSeq. TopHat2 was used to map sequencing reads to the human genome. On average ~62 million reads (53.4% of ~116 million reads) were mapped per sample. 4,131 genes were differentially expressed between IPF and controls (1,920 increased and 2,211 decreased (FDR < 0.05). We compared our results to differentially expressed genes calculated from a previously published dataset generated from FF tissues analyzed on Agilent microarrays (GSE47460). The overlap of differentially expressed genes was very high (760 increased and 1,413 decreased, FDR < 0.05). Only 92 differentially expressed genes changed in opposite directions. Pathway enrichment analysis performed using MetaCore confirmed numerous IPF relevant genes and pathways including extracellular remodeling, TGF-beta, and WNT. Gene network analysis of MMP7, a highly differentially expressed gene in both datasets, revealed the same canonical pathways and gene network candidates in RNA-Seq and microarray data. For validation by NanoString nCounter® we selected 35 genes that had a fold change of 2 in at least one dataset (10 discordant, 10 significantly differentially expressed in one dataset only and 15 concordant genes). High concordance of fold change and FDR was observed for each type of the samples (FF vs FFPE) with both microarrays (r = 0.92) and RNA-Seq (r = 0.90) and the number of discordant genes was reduced to four. Our results demonstrate that RNA sequencing of RNA obtained from archived FFPE lung tissues is feasible. The results obtained from FFPE tissue are highly comparable to FF tissues. The ability to perform RNA-Seq on archived FFPE IPF tissues should greatly enhance the availability of tissue biopsies for research in IPF.

A postoperative anti-adhesion barrier based on photoinduced imine-crosslinking hydrogel with tissue-adhesive ability.

PubMed

Yang, Yunlong; Liu, Xiaolin; Li, Yan; Wang, Yang; Bao, Chunyan; Chen, Yunfeng; Lin, Qiuning; Zhu, Linyong

2017-10-15

Postoperative adhesion is a serious complication that can further lead to morbidity and/or mortality. Polymer anti-adhesion barrier material provides an effective precaution to reduce the probability of postoperative adhesion. Clinical application requires these materials to be easily handled, biocompatible, biodegradable, and most importantly tissue adherent to provide target sites with reliable isolation. However, currently there is nearly no polymer barrier material that can fully satisfy these requirements. In this study, based on the photoinduced imine-crosslinking (PIC) reaction, we had developed a photo-crosslinking hydrogel (CNG hydrogel) that composed of o-nitrobenzyl alcohol (NB) modified carboxymethyl cellulose (CMC-NB) and glycol chitosan (GC) as an anti-adhesion barrier material. Under light irradiation, CMC-NB generated aldehyde groups which subsequently reacted with amino groups distributed on GC or tissue surface to form a hydrogel barrier that covalently attached to tissue surface. Rheological analysis demonstrated that CNG hydrogel (30mg/mL polymer content) could be formed in 30s upon light irradiation. Tissue adhesive tests showed that the tissue adhesive strength of CNG hydrogel (30mg/mL) was about 8.32kPa-24.65kPa which increased with increasing CMC-NB content in CNG hydrogel. Toxicity evaluation by L929 cells demonstrated that CNG hydrogel was cytocompatible. Furthermore, sidewall defect-cecum abrasion model of rat was employed to evaluate the postoperative anti-adhesion efficacy of CNG hydrogel. And a significantly reduction of tissue adhesion (20% samples with low score adhesion) was found in CNG hydrogel treated group, compared with control group (100% samples with high score adhesion). In addition, CNG hydrogel could be degraded in nearly 14days and showed no side effect on wound healing. These findings indicated that CNG hydrogel can effectively expanded the clinical treatments of postoperative tissue adhesion. In this study, a tissue adhesive photo-crosslinking hydrogel (CNG) was developed based on photo-induced imine crosslinking reaction (PIC) for postoperative anti-adhesion. CNG hydrogel showed the features of easy and convenient operation, fast and controllable gelation, suitable gel strength, good biocompatibility, and most importantly strong tissue adhesiveness. Therefore, it shows very high performance to prevent postoperative tissue adhesion. Overall, our study provides a more suitable hydrogel barrier material that can overcome the shortcomings of current barriers for clinical postoperative anti-adhesion. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Molecular diagnosis of Eimeria stiedae in hepatic tissue of experimentally infected rabbits.

PubMed

Hassan, Khaled M; Arafa, Waleed M; Mousa, Waheed M; Shokier, Khaled A M; Shany, Salama A; Aboelhadid, Shawky M

2016-10-01

The early detection of Eimeria stiedae in the hepatic tissue of experimentally infected rabbits was investigated using molecular assay. Forty 6-week-old male New Zealand rabbits were divided into two groups. Group A (30 animals) was infected with 2.5 × 10(4) sporulated oocysts of E. stiedae per animal on Day 0 and Group B (10 animals) was used as the uninfected controls. Three animals from Group A and one from Group B were sacrificed at 0, 3, 6, 9, 12, 15, 18, 21, 24 and 27 days post infection (PI). Gross and microscopic post-mortem findings were recorded. Polymerase chain reaction (PCR) of the E. stiedae internal transcribed spacer 1 genomic region was conducted on blood, liver tissue, and feces from the Group A experimentally infected animals. Macroscopically, the liver showed irregular yellowish white nodules pathognomonic to E. stiedae infection beginning on Day 15 PI. Hepatomegaly and ascites were obvious from Day 21-24 PI. The presence of different E. stiedae schizonts and gametocytes in the histopathological sections of the biliary epithelium were evident on Day 15 PI. The E. stiedae PCR was first positive in liver tissues on Day 12 and in fecal samples on Day 18 PI, but the blood samples were negative. In conclusion, the PCR can be used for early diagnosis and control of E. stiedae schizonts before shedding of the oocysts in feces. Copyright © 2016 Elsevier Inc. All rights reserved.

Expression and clinical significance of PIWIL2 in hilar cholangiocarcinoma tissues and cell lines.

PubMed

Chen, Y J; Xiong, X F; Wen, S Q; Tian, L; Cheng, W L; Qi, Y Q

2015-06-26

The objective of this study was to explore the relationship between PIWI-like protein 2 (PIWIL2) and clinicopathological charac-teristics and prognosis after radical resection. To accomplish this, we analyzed PIWIL2 expression in hilar cholangiocarcinoma tissues and cell lines. PIWIL2 expression was detected by immunohistochemistry in 41 hilar cholangiocarcinoma samples and 10 control tissues. Western blotting and immunocytofluorescence were used to investigate PIWIL2 expression in the cholangiocarcinoma cell line QBC939 and the bile duct epithelial cell line HIBEpic. Univariate and multivariate surviv-al analyses were performed using the Kaplan-Meier method for hilar cholangiocarcinoma patients who underwent radical resection. PIWIL2 expression was significantly higher in the hilar cholangiocarcinoma tissues and QBC939 cells than in control tissues and HIBEpic cells, respectively (P < 0.05). Poorly and moderately differentiated cholan-giocarcinoma tissues had significantly higher PIWIL2 expression than well-differentiated tissues (P < 0.05). Univariate analysis demonstrated that high PIWIL2 expression was associated with shorter survival time after radical resection (P < 0.05). Multivariate analysis showed that PI-WIL2 expression was an independent prognostic factor after radical re-section of hilar cholangiocarcinoma (P < 0.05). PIWIL2 expression was also associated with tumor-node-metastasis stage and differentiation. PIWIL2 was an independent prognostic factor after radical resection of hilar cholangiocarcinoma.

Upregulation of angiogenesis in oral lichen planus.

PubMed

Al-Hassiny, A; Friedlander, L T; Parachuru, V P B; Seo, B; Hussaini, H M; Rich, A M

2018-02-01

As angiogenesis is fundamental to the pathogenesis of many chronic inflammatory disorders, this study investigated the expression of various vascular markers in oral lichen planus and non-specific oral mucosal inflammatory tissues. Archival specimens of oral lichen planus (n = 15) and inflamed tissues (n = 13) were stained using immunohistochemistry with antibodies to CD34, vascular endothelial growth factor, vascular endothelial growth factor receptor and vasohibin. Nine representative sites at the epithelial-connective tissue junction and through the fibrous connective tissue were selected, and automated analysis techniques were used to determine the extent of positivity expressed as the percentage of positive cells. Significance was denoted when P < .05. The expression of pro-angiogenic factors was higher in lichen planus samples compared with inflamed controls. A higher level of CD34 was observed in the deeper parts of the connective tissue of Oral lichen planus (OLP) (P = .04), whereas VEGF and VEGFR2 expressions were higher all through the tissues (respectively, P < .02 and P < .01). The expression of the anti-angiogenic VASH1 was higher in inflamed tissue compared with lichen planus in all sites evaluated (P < .01). The findings indicate that angiogenic factors are differentially expressed in oral lichen planus compared with inflamed controls, with increased expression of pro-angiogenic factors and decreased anti-angiogenic expression. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

The Acid-Secreting Parietal Cell as an Endocrine Source of Sonic Hedgehog During Gastric Repair

PubMed Central

Engevik, Amy C.; Feng, Rui; Yang, Li

2013-01-01

Sonic Hedgehog (Shh) has been shown to regulate wound healing in various tissues. Despite its known function in tissue regeneration, the role of Shh secreted from the gastric epithelium during tissue repair in the stomach remains unknown. Here we tested the hypothesis that Shh secreted from the acid-secreting parietal cell is a fundamental circulating factor that drives gastric repair. A mouse model expressing a parietal cell-specific deletion of Shh (PC-ShhKO) was generated using animals bearing loxP sites flanking exon 2 of the Shh gene (Shhflx/flx) and mice expressing a Cre transgene under the control of the H+,K+-ATPase β-subunit promoter. Shhflx/flx, the H+,K+-ATPase β-subunit promoter, and C57BL/6 mice served as controls. Ulcers were induced via acetic acid injury. At 1, 2, 3, 4, 5, and 7 days after the ulcer induction, gastric tissue and blood samples were collected. Parabiosis experiments were used to establish the effect of circulating Shh on ulcer repair. Control mice exhibited an increased expression of Shh in the gastric tissue and plasma that correlated with the repair of injury within 7 days after surgery. PC-ShhKO mice showed a loss of ulcer repair and reduced Shh tissue and plasma concentrations. In a parabiosis experiment whereby a control mouse was paired with a PC-ShhKO littermate and both animals subjected to gastric injury, a significant increase in the circulating Shh was measured in both parabionts. Elevated circulating Shh concentrations correlated with the repair of gastric ulcers in the PC-ShhKO parabionts. Therefore, the acid-secreting parietal cell within the stomach acts as an endocrine source of Shh during repair. PMID:24092639

Sport- and sample-specific features of trace elements in adolescent female field hockey players and fencers.

PubMed

Nabatov, Alexey A; Troegubova, Natalya A; Gilmutdinov, Ruslan R; Sereda, Andrey P; Samoilov, Alexander S; Rylova, Natalya V

2017-09-01

Active physical exercises and growth are associated with mineral imbalances in young athletes. The purpose of this study was to examine the impact of sport-related factors on tissue mineral status in adolescent female athletes. Saliva and hair samples were used for the analysis of immediate and more permanent tissue mineral status, respectively. Samples taken from a control non-athletic female group and two groups of female athletes (field hockey and fencing) were analyzed for seven essential minerals: calcium, chromium, iron, potassium, magnesium, selenium and zinc. Inductively-coupled plasma mass spectrometry was used for the quantification of elements having very low concentration range in samples (Se, Cr and Zn) whereas inductively coupled plasma optical emission spectrometry was used for quantification of more ubiquitous elements (Mg, К, Са, Fe). The obtained results for athletic groups were compared with control. Female athletes had increased levels of selenium in both saliva and hair as well as chromium in saliva. Field hockey players had the higher level of zinc in hair whereas fencers had the lower levels of salivary calcium. Strong negative correlation between potassium levels in saliva and hair was identified. Iron and magnesium did not differ between the studied groups. In conclusion, novel sport-specific features of chromium tissue levels in female athletes were found. The studied sport disciplines have different impact on the distribution of osteoporosis-related minerals (calcium and zinc). Our finding can help in the development of osteoporosis preventive trainings and in the proper nutrient supplementation to correct mineral imbalances in female athletes. Copyright © 2016 Elsevier GmbH. All rights reserved.

NF-κB involvement in hyperoxia-induced myocardial damage in newborn rat hearts.

PubMed

Zara, Susi; De Colli, Marianna; Rapino, Monica; Di Valerio, Valentina; Marconi, Guya Diletta; Cataldi, Amelia; Macchi, Veronica; De Caro, Raffaele; Porzionato, Andrea

2013-11-01

Premature newborns are frequently exposed to hyperoxia ventilation and some literature data indicate the possibility of hyperoxia-induced myocardial damage. Since nuclear factor κB (NF-κB) is a crucial signaling molecule involved in physiological response to hyperoxia in different cell types as well as in various tissues, our attention has been focused on the role played by NF-κB pathway in response to moderate and severe hyperoxia exposure in rat neonatal heart tissue. Akt and IκBα levels, involved in NF-κB activation, along with the balance between apoptotic and survival pathways have also been investigated. Experimental design of the study has involved exposure of newborn rats to room air (controls), 60 % O2 (moderate hyperoxia), or 95 % O2 (severe hyperoxia) for the first two postnatal weeks. Morphological analysis shows a less compact tissue in rat heart exposed to moderate hyperoxia and a decreased number of nuclei in samples exposed to severe hyperoxia. A significant increase of NF-κB positive nuclei percentage and p-IκBα expression in samples exposed to 95 % hyperoxia compared to control and to 60 % hyperoxia is evidenced; in parallel, an increase of pAkt/Akt ratio in both samples exposed to 95 and 60 % hyperoxia is shown. Furthermore, a more evident cytochrome c/Apaf-1 immunocomplex and a decreased Bcl2 expression in 95 % hyperoxia-exposed sample compared to 60 % exposed one is evidenced. In conclusion, our findings suggest the involvement of the NF-κB pathway and Akt signaling in the mechanisms of myocardial hyperoxic damage in the newborns, with particular reference to the induction of oxidative stress-related apoptosis.

The use of embryonic cells in the treatment of osteochondral defects of the knee: an ovine in vivo study.

PubMed

Manunta, Andrea Fabio; Zedde, Pietro; Pilicchi, Susanna; Rocca, Stefano; Pool, Roy R; Dattena, Maria; Masala, Gerolamo; Mara, Laura; Casu, Sara; Sanna, Daniela; Manunta, Maria Lucia; Passino, Eraldo Sanna

2016-01-01

the aim of this study was to determine whether local delivery of embryonic stem-like (ESL) cells into osteochondral defects in the femoral condyles of sheep would enhance regeneration of hyaline articular cartilage. male ESL cells embedded in fibrin glue were engrafted into osteochondral defects in the medial condyles (ESL-M) of the left femur in 22 ewes. An identical defect was created in the medial condyle of the contralateral stifle joint and left untreated as a control (empty defect, ED). The ewes were divided into 5 groups. Four sheep each were euthanized at 1, 2, 6, and 12 months from surgery, and 6 ewes were euthanized 24 months post-implantation. To study the effect of varying loads on the long-term regeneration process, an identical defect was also created and ESL cell engraftment performed in the lateral condyle (ESL-L) of the left stifle joint of the animals in the 12- and 24-month groups. The evaluation of regenerated tissue was performed by biomechanical, macroscopic, histological, immunohistochemical (collagen type II) and fluorescent in situ hybridization (FISH) assays. no significant differences were found between treated and control sites in the biomechanical assays at any time point. ESL cell grafts showed significantly greater macroscopic evidence of regeneration as compared to controls at 24 months after surgery; significantly better histological evidence of repair in ESL-M samples versus controls was found throughout the considered period. At 24 months from surgery there was significantly improved integration of graft edges with the host tissue in the ESL-M as compared to the ESL-L samples, demonstrating that load bearing positively affects the long-term regeneration process. ESL cells enhanced the regeneration of hyaline cartilage. FISH confirmed that the regenerative tissue originated from ESL cells. ESL cells are able to self-renew for prolonged periods without differentiation and, most importantly, to differentiate into a large variety of tissues.

The use of embryonic cells in the treatment of osteochondral defects of the knee: an ovine in vivo study

PubMed Central

MANUNTA, ANDREA FABIO; ZEDDE, PIETRO; PILICCHI, SUSANNA; ROCCA, STEFANO; POOL, ROY R.; DATTENA, MARIA; MASALA, GEROLAMO; MARA, LAURA; CASU, SARA; SANNA, DANIELA; MANUNTA, MARIA LUCIA; PASSINO, ERALDO SANNA

2016-01-01

Purpose the aim of this study was to determine whether local delivery of embryonic stem-like (ESL) cells into osteochondral defects in the femoral condyles of sheep would enhance regeneration of hyaline articular cartilage. Methods male ESL cells embedded in fibrin glue were engrafted into osteochondral defects in the medial condyles (ESL-M) of the left femur in 22 ewes. An identical defect was created in the medial condyle of the contralateral stifle joint and left untreated as a control (empty defect, ED). The ewes were divided into 5 groups. Four sheep each were euthanized at 1, 2, 6, and 12 months from surgery, and 6 ewes were euthanized 24 months post-implantation. To study the effect of varying loads on the long-term regeneration process, an identical defect was also created and ESL cell engraftment performed in the lateral condyle (ESL-L) of the left stifle joint of the animals in the 12- and 24-month groups. The evaluation of regenerated tissue was performed by biomechanical, macroscopic, histological, immunohistochemical (collagen type II) and fluorescent in situ hybridization (FISH) assays. Results no significant differences were found between treated and control sites in the biomechanical assays at any time point. ESL cell grafts showed significantly greater macroscopic evidence of regeneration as compared to controls at 24 months after surgery; significantly better histological evidence of repair in ESL-M samples versus controls was found throughout the considered period. At 24 months from surgery there was significantly improved integration of graft edges with the host tissue in the ESL-M as compared to the ESL-L samples, demonstrating that load bearing positively affects the long-term regeneration process. Conclusions ESL cells enhanced the regeneration of hyaline cartilage. FISH confirmed that the regenerative tissue originated from ESL cells. Clinical Relevance ESL cells are able to self-renew for prolonged periods without differentiation and, most importantly, to differentiate into a large variety of tissues. PMID:27602346

Methylation of L1RE1, RARB, and RASSF1 function as possible biomarkers for the differential diagnosis of lung cancer.

PubMed

Walter, R F H; Rozynek, P; Casjens, S; Werner, R; Mairinger, F D; Speel, E J M; Zur Hausen, A; Meier, S; Wohlschlaeger, J; Theegarten, D; Behrens, T; Schmid, K W; Brüning, T; Johnen, G

2018-01-01

Lung cancer is the major cause of cancer-related deaths worldwide. Differential diagnosis can be difficult, especially when only small samples are available. Epigenetic changes are frequently tissue-specific events in carcinogenesis and hence may serve as diagnostic biomarkers. 138 representative formalin-fixed, paraffin-embedded (FFPE) tissues (116 lung cancer cases and 22 benign controls) were used for targeted DNA methylation analysis via pyrosequencing of ten literature-derived methylation markers (APC, CDH1, CDKN2A, EFEMP1, FHIT, L1RE1, MGMT, PTEN, RARB, and RASSF1). Methylation levels were analyzed with the Classification and Regression Tree Algorithm (CART), Conditional Interference Trees (ctree) and ROC. Validation was performed with additional 27 lung cancer cases and 38 benign controls. TCGA data for 282 lung cancer cases was included in the analysis. CART and ctree analysis identified the combination of L1RE1 and RARB as well as L1RE1 and RASSF1 as independent methylation markers with high discriminative power between tumor and benign tissue (for each combination, 91% specificity and 100% sensitivity). L1RE1 methylation associated significantly with tumor type and grade (p<0.001) with highest methylation in the control group. The opposite was found for RARB (p<0.001). RASSF1 methylation increased with tumor type and grade (p<0.001) with strongest methylation in neuroendocrine tumors (NET). Hypomethylation of L1RE1 is frequent in tumors compared to benign controls and associates with higher grade, whereas increasing methylation of RARB is an independent marker for tumors and higher grade. RASSF1 hypermethylation was frequent in tumors and most prominent in NET making it an auxiliary marker for separation of NSCLC and NET. L1RE1 in combination with either RARB or RASSF1 could function as biomarkers for separating lung cancer and non-cancerous tissue and could be useful for samples of limited size such as biopsies.

Array-based comparative genomic hybridization-guided identification of reference genes for normalization of real-time quantitative polymerase chain reaction assay data for lymphomas, histiocytic sarcomas, and osteosarcomas of dogs.

PubMed

Tsai, Pei-Chien; Breen, Matthew

2012-09-01

To identify suitable reference genes for normalization of real-time quantitative PCR (RT-qPCR) assay data for common tumors of dogs. Malignant lymph node (n = 8), appendicular osteosarcoma (9), and histiocytic sarcoma (12) samples and control samples of various nonneoplastic canine tissues. Array-based comparative genomic hybridization (aCGH) data were used to guide selection of 9 candidate reference genes. Expression stability of candidate reference genes and 4 commonly used reference genes was determined for tumor samples with RT-qPCR assays and 3 software programs. LOC611555 was the candidate reference gene with the highest expression stability among the 3 tumor types. Of the commonly used reference genes, expression stability of HPRT was high in histiocytic sarcoma samples, and expression stability of Ubi and RPL32 was high in osteosarcoma samples. Some of the candidate reference genes had higher expression stability than did the commonly used reference genes. Data for constitutively expressed genes with high expression stability are required for normalization of RT-qPCR assay results. Without such data, accurate quantification of gene expression in tumor tissue samples is difficult. Results of the present study indicated LOC611555 may be a useful RT-qPCR assay reference gene for multiple tissue types. Some commonly used reference genes may be suitable for normalization of gene expression data for tumors of dogs, such as lymphomas, osteosarcomas, or histiocytic sarcomas.

COMPARISON OF TISSUE CULTURE AND ANIMAL MODELS FOR ASSESSMENT OF CRYPTOSPRIDIUM PARVUM INFECTION

EPA Science Inventory

Data from three different disinfection studies using both cell culture and mouse infectivity to assess Cryptosporidium parvum inactivation were evaluated in a total of 35 comparison including process controls and treated samples. C. parvum infectivity in the in vitro FDM-MPN assa...

Method for sampling and analysis of volatile biomarkers in process gas from aerobic digestion of poultry carcasses using time-weighted average SPME and GC-MS.

PubMed

Koziel, Jacek A; Nguyen, Lam T; Glanville, Thomas D; Ahn, Heekwon; Frana, Timothy S; Hans van Leeuwen, J

2017-10-01

A passive sampling method, using retracted solid-phase microextraction (SPME) - gas chromatography-mass spectrometry and time-weighted averaging, was developed and validated for tracking marker volatile organic compounds (VOCs) emitted during aerobic digestion of biohazardous animal tissue. The retracted SPME configuration protects the fragile fiber from buffeting by the process gas stream, and it requires less equipment and is potentially more biosecure than conventional active sampling methods. VOC concentrations predicted via a model based on Fick's first law of diffusion were within 6.6-12.3% of experimentally controlled values after accounting for VOC adsorption to the SPME fiber housing. Method detection limits for five marker VOCs ranged from 0.70 to 8.44ppbv and were statistically equivalent (p>0.05) to those for active sorbent-tube-based sampling. The sampling time of 30min and fiber retraction of 5mm were found to be optimal for the tissue digestion process. Copyright © 2017 Elsevier Ltd. All rights reserved.

Evaluation of a method for determining concentrations of isoeugenol, an AQUI-S residue, in fillet tissue from freshwater fish species.

USGS Publications Warehouse

Meinertz, J.R.; Schreier, Theresa M.; Bernardy, J.A.

2008-01-01

AQUI-S is a fish anesthetic/sedative that is approved for use in a number of countries throughout the world and has the potential for use in the United States. The active ingredient in AQUI-S is isoeugenol. A method for determining isoeugenol concentrations in edible fillet tissue is needed for regulatory purposes, including surveillance and potential use in studies fulfilling human food safety data requirements if U.S. Food and Drug Administration approval is pursued. A method was developed and evaluated for determining isoeugenol concentrations in fillet tissue using relatively common procedures and equipment. The method produced accurate and precise results with fillet tissue from 10 freshwater fish species. The percentage of isoeugenol recovered from samples fortified with isoeugenol at nominal concentrations of 1, 50, and 100 microg/g for all species was always >80 and <97%. Within-day precision for samples fortified at those same concentrations was < or =10%, and day-to-day precision was < or =4.0%. Method precision with fillet tissue containing biologically incurred isoeugenol was < or =8.1%. There were no or minimal chromatographic interferences in control fillet tissue extracts from 9 of the 10 species. The method detection limits for all but one species ranged from 0.004 to 0.014 microg/g, and the quantitation limits ranged from 0.012 to 0.048 microg/g.

Metabolic Pathway Signatures Associated with Urinary Metabolite Biomarkers Differentiate Bladder Cancer Patients from Healthy Controls.

PubMed

Kim, Won Tae; Yun, Seok Joong; Yan, Chunri; Jeong, Pildu; Kim, Ye Hwan; Lee, Il Seok; Kang, Ho Won; Park, Sunghyouk; Moon, Sung Kwon; Choi, Yung Hyun; Choi, Young Deuk; Kim, Isaac Yi; Kim, Jayoung; Kim, Wun Jae

2016-07-01

Our previous high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry study identified bladder cancer (BCA)-specific urine metabolites, including carnitine, acylcarnitines, and melatonin. The objective of the current study was to determine which metabolic pathways are perturbed in BCA, based on our previously identified urinary metabolome. A total of 135 primary BCA samples and 26 control tissue samples from healthy volunteers were analyzed. The association between specific urinary metabolites and their related encoding genes was analyzed. Significant alterations in the carnitine-acylcarnitine and tryptophan metabolic pathways were detected in urine specimens from BCA patients compared to those of healthy controls. The expression of eight genes involved in the carnitine-acylcarnitine metabolic pathway (CPT1A, CPT1B, CPT1C, CPT2, SLC25A20, and CRAT) or tryptophan metabolism (TPH1 and IDO1) was assessed by RT-PCR in our BCA cohort (n=135). CPT1B, CPT1C, SLC25A20, CRAT, TPH1, and IOD1 were significantly downregulated in tumor tissues compared to normal bladder tissues (p<0.05 all) of patients with non-muscle invasive BCA, whereas CPT1B, CPT1C, CRAT, and TPH1 were downregulated in those with muscle invasive BCA (p<0.05), with no changes in IDO1 expression. Alterations in the expression of genes associated with the carnitine-acylcarnitine and tryptophan metabolic pathways, which were the most perturbed pathways in BCA, were determined.

Relative IGF-1 and IGF-2 gene expression in maternal and fetal tissues from diabetic swine

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wolverton, C.K.; Leaman, D.W.; White, M.E.

1990-02-26

Fourteen pregnant, crossbred gilts were utilized in this study. Seven gilts were injected with alloxan (50 mg/kg) at day 75 of gestation to induce diabetes. Gilts underwent caesarean section on day 105 of gestation. Samples were collected from maternal skeletal muscle, adipose tissue, uterus and endometrium; and from fetal skeletal muscle, adipose tissue, placenta, liver, lung, kidney, heart, brain and spleen. Tissues were frozen in liquid nitrogen for later analysis of IGF-1 and IGF-2 gene expression. Samples were pooled and total RNA was isolated using the guanidine isothiocynate method. Total mRNA was analyzed by dot blot hybridization. Blots were probedmore » with {sup 32}P-cDNA for porcine IGF-1 and rat IGF-2. IGF-1 gene expression in maternal tissues was unaffected by diabetes. Maternal diabetes increased IGF-2 mRNA in maternal adipose tissue but exhibited no effect in muscle or uterus. Expression of IGF-2 by maternal endometrium was decreased by diabetes. Maternal diabetes induced an increase in IGF-1 gene expression in muscle and placenta while causing an increase in IGF-2 expression in fetal liver and placenta. IGF-2 mRNA was lower in lung from fetuses of diabetic mothers than in controls. These results suggest that maternal diabetes alters IGF-1 and IGF-2 gene expression in specific tissues and differential regulation of these genes appears to exist in the mother and developing fetus.« less

Effects of Hyperbaric Oxygen Treatment on Renal System.

PubMed

Tezcan, Orhan; Caliskan, Ahmet; Demirtas, Sinan; Yavuz, Celal; Kuyumcu, Mahir; Nergiz, Yusuf; Guzel, Abdulmenap; Karahan, Oguz; Ari, Seyhmus; Soker, Sevda; Yalinkilic, Ibrahim; Turkdogan, Kenan Ahmet

2017-01-01

Hyperbaric oxygen (HBO) treatment is steadily increasing as a therapeutic modality for various types of diseases. Although good clinical outcomes were reported with HBO treatment for various diseases, the multisystemic effects of this modality are still unclear. This study aimed to investigate the renal effects of HBO experimentally. Fourteen New Zealand White rabbits were divided into 2 groups randomly as the control group and the study group. The study group received HBO treatment for 28 days (100% oxygen at 2.5 atmospheres for 90 minutes daily) and the control group was used to obtain normal renal tissue of the animal genus. After the intervention period, venous blood samples were obtained, and renal tissue samples were harvested for comparisons. Normal histological morphology was determined with Masson trichrome staining and periodic acid-Schiff staining in the control group. Atrophic glomerular structures, vacuolated tubule cells, and degeneration were detected in the renal samples of the study group with Masson trichrome staining. Additionally, flattening was observed on the brush borders of the proximal tubules, and tubular dilatation was visualized with periodic acid-Schiff staining. The histopathologic disruption of renal morphology was verified with detection of significantly elevated kidney function laboratory biomarkers in the study group. Our findings suggests that HBO has adverse effects on renal glomerulus and proximal tubules. However, the functional effects of this alteration should be investigated with further studies.

Trace-Element Concentrations in Tissues of Aquatic Organisms from Rivers and Streams of the United States, 1992-1999

USGS Publications Warehouse

DeWeese, Lawrence R.; Stephens, Verlin C.; Short, Terry M.; Dubrovsky, Neil M.

2007-01-01

The U.S. Geological Survey National Water-Quality Assessment Program collected tissue samples from a variety of aquatic organisms during 1992-1999 within 47 study units across the United States. These tissue samples were collected to determine the occurrence and distribution of 20 major and minor trace elements in aquatic organisms. This report presents the tissue trace-element concentration data, sample summaries, and concentration statistics for 1,457 tissue samples representing 76 species or groups of fish, aquatic invertebrates, and plants were collected at 824 sampling sites.

Effect of noise pollution on testicular tissue and hormonal assessment in rat.

PubMed

Farzadinia, P; Bigdeli, M; Akbarzadeh, S; Mohammadi, M; Daneshi, A; Bargahi, A

2016-11-01

Many studies have focused on the effect of noise stress on the health. So far, few studies have been conducted on the effect of noise on reproductive system. The aim of study was to investigate the effect of noise pollution on morphometric parameters of testicular tissue and hormonal assessment (ACTH, cortisol and testosterone). In this study, 40 male rats were exposed to control, 95, 105 and 115 dB noise intensity for sixty days. At the end of study, blood sampling was performed and ACTH, cortisol and testosterone concentrations were assessed. The results showed that noise stress decreased testosterone levels in the 115 dB-treated group, while it increased the ACTH and cortisol levels. Histological sections of testis showed that the mean diameter of the seminiferous tubules and thickness of the germinal epithelium reduced compared to the control group. Also the ratio of the interstitial tissue area to the total testicular tissue area was increased significantly. Our study shows that noise stress may have negative influences on male fertility. © 2016 Blackwell Verlag GmbH.

Nanobacteria may be linked to calcification in placenta.

PubMed

Lu, He; Guo, Ya-nan; Liu, Sheng-nan; Zhang, De-chun

2012-05-01

Placental calcification is a common pathologic condition in obstetrics. To detect the bacteria infection mechanisms for calcification, an experiment was performed to isolate, culture, and identify the nanobacteria in placental calcification. Sixteen cases of placental calcification of pregnant women were collected for the purpose of the isolation of nanobacteria, cultivation, and identification of 16S rDNA sequence. Under transmission electron microscope, novel oval-shape nanobacteria-like particles (NLP) in extracellular matrix of calcified placenta tissues were found with 50-500 nm in diameter, and among hydroxyapatite crystals aggregation existed. After about 4 weeks of culturing and isolating NLP from these calcified tissues, all calcified placental tissue samples and one adjacent tissue of calcified placental tissue samples showed white granular depositions, which were firmly attached to the bottom of the culture tubes and visible to the naked eyes. In the control group they could not be seen. After PCR was amplified a 1407-bp fragment was obtained and submitted to GenBank after sequencing with accession number JN029830. The 16S rDNA sequence homology between the isolation strain and strain nanobacteria (X98418) was 92% in GenBank. For the first time isolated, cultured, and identified nanobacteria in placental calcification indicated that nanobacteria infection is related to placental calcification.

The direct analysis of drug distribution of rotigotine-loaded microspheres from tissue sections by LESA coupled with tandem mass spectrometry.

PubMed

Xu, Li-Xiao; Wang, Tian-Tian; Geng, Yin-Yin; Wang, Wen-Yan; Li, Yin; Duan, Xiao-Kun; Xu, Bin; Liu, Charles C; Liu, Wan-Hui

2017-09-01

The direct analysis of drug distribution of rotigotine-loaded microspheres (RoMS) from tissue sections by liquid extraction surface analysis (LESA) coupled with tandem mass spectrometry (MS/MS) was demonstrated. The RoMS distribution in rat tissues assessed by the ambient LESA-MS/MS approach without extensive or tedious sample pretreatment was compared with that obtained by a conventional liquid chromatography tandem mass spectrometry (LC-MS/MS) method in which organ excision and subsequent solvent extraction were commonly employed before analysis. Results obtained from the two were well correlated for a majority of the organs, such as muscle, liver, stomach, and hippocampus. The distribution of RoMS in the brain, however, was found to be mainly focused in the hippocampus and striatum regions as shown by the LESA-imaged profiles. The LESA approach we developed is sensitive enough, with an estimated LLOQ at 0.05 ng/mL of rotigotine in brain tissue, and information-rich with minimal sample preparation, suitable, and promising in assisting the development of new drug delivery systems for controlled drug release and protection. Graphical abstract Workflow for the LESA-MS/MS imaging of brain tissue section after intramuscular RoMS administration.

Selective laser vaporization of polypropylene sutures and mesh

NASA Astrophysics Data System (ADS)

Burks, David; Rosenbury, Sarah B.; Kennelly, Michael J.; Fried, Nathaniel M.

2012-02-01

Complications from polypropylene mesh after surgery for female stress urinary incontinence (SUI) may require tedious surgical revision and removal of mesh materials with risk of damage to healthy adjacent tissue. This study explores selective laser vaporization of polypropylene suture/mesh materials commonly used in SUI. A compact, 7 Watt, 647-nm, red diode laser was operated with a radiant exposure of 81 J/cm2, pulse duration of 100 ms, and 1.0-mm-diameter laser spot. The 647-nm wavelength was selected because its absorption by water, hemoglobin, and other major tissue chromophores is low, while polypropylene absorption is high. Laser vaporization of ~200-μm-diameter polypropylene suture/mesh strands, in contact with fresh urinary tissue samples, ex vivo, was performed. Non-contact temperature mapping of the suture/mesh samples with a thermal camera was also conducted. Photoselective vaporization of polypropylene suture and mesh using a single laser pulse was achieved with peak temperatures of 180 and 232 °C, respectively. In control (safety) studies, direct laser irradiation of tissue alone resulted in only a 1 °C temperature increase. Selective laser vaporization of polypropylene suture/mesh materials is feasible without significant thermal damage to tissue. This technique may be useful for SUI procedures requiring surgical revision.

Exposure assessment for workers applying DDT to control malaria in Veracruz, Mexico.

PubMed Central

Rivero-Rodriguez, L; Borja-Aburto, V H; Santos-Burgoa, C; Waliszewskiy, S; Rios, C; Cruz, V

1997-01-01

DDT has systematically been used in sanitation campaigns against malaria in Mexico. To assess chronic occupational exposure, we studied a group of workers dedicated to spraying houses to control malaria vectors in the state of Veracruz. Exposure was directly estimated for a subgroup of 40 workers by measuring DDT metabolites in adipose tissue samples and indirectly estimated for 331 workers by using a questionnaire to determine their occupational history. Participants ranged in age from 20 to 70 years, and 80% of the workers had been employed in the sanitation campaign for at least 20 years. The mean concentrations of extractable lipids found in adipose tissue samples were as follows: total DDT, 104.48 micrograms/g; p,p'-DDE, 60.98 micrograms/g; p,p'-DDT, 31.0 micrograms/g; o,p'-DDT, 2.10 micrograms/g; and p,p'-DDD, 0.95 microgram/g. The DDT metabolite p,p'-DDE was selected as the indicator of chronic exposure. An index of chronic occupational exposure was constructed according to worker position and based on the historical duration and intensity of DDT application. A linear model including this index, the use of protective gear, and recent weight loss explained 55% of the variation of p,p'-DDE concentrations in adipose tissue. By this model, the predicted values of p,p'-DDE concentration in adipose tissue for the 331 workers are between 9.56 micrograms/g and 298.4 micrograms/g of fat, with a geometric mean of 67.41 micrograms/g. These high levels of DDT in adipose tissue call for exposure prevention programs and the promotion of more secure application measures and hygiene. We also discuss the use of indirect measures of DDT exposure in epidemiological studies of health effects. PMID:9074888

Reusable bi-directional 3ω sensor to measure thermal conductivity of 100-μm thick biological tissues

NASA Astrophysics Data System (ADS)

Lubner, Sean D.; Choi, Jeunghwan; Wehmeyer, Geoff; Waag, Bastian; Mishra, Vivek; Natesan, Harishankar; Bischof, John C.; Dames, Chris

2015-01-01

Accurate knowledge of the thermal conductivity (k) of biological tissues is important for cryopreservation, thermal ablation, and cryosurgery. Here, we adapt the 3ω method—widely used for rigid, inorganic solids—as a reusable sensor to measure k of soft biological samples two orders of magnitude thinner than conventional tissue characterization methods. Analytical and numerical studies quantify the error of the commonly used "boundary mismatch approximation" of the bi-directional 3ω geometry, confirm that the generalized slope method is exact in the low-frequency limit, and bound its error for finite frequencies. The bi-directional 3ω measurement device is validated using control experiments to within ±2% (liquid water, standard deviation) and ±5% (ice). Measurements of mouse liver cover a temperature ranging from -69 °C to +33 °C. The liver results are independent of sample thicknesses from 3 mm down to 100 μm and agree with available literature for non-mouse liver to within the measurement scatter.

Novel Multiplex MethyLight Protocol for Detection of DNA Methylation in Patient Tissues and Bodily Fluids

PubMed Central

Olkhov-Mitsel, Ekaterina; Zdravic, Darko; Kron, Ken; van der Kwast, Theodorus; Fleshner, Neil; Bapat, Bharati

2014-01-01

Aberrant DNA methylation is a hallmark of cancer and is an important potential biomarker. Particularly, combined analysis of a panel of hypermethylated genes shows the most promising clinical performance. Herein, we developed, optimized and standardized a multiplex MethyLight assay to simultaneously detect hypermethylation of APC, HOXD3 and TGFB2 in DNA extracted from prostate cancer (PCa) cell lines, archival tissue specimens, and urine samples. We established that the assay is capable of discriminating between fully methylated and unmethylated alleles with 100% specificity and demonstrated the assay as highly accurate and reproducible as the singleplex approach. For proof of principle, we analyzed the methylation status of these genes in tissue and urine samples of PCa patients as well as PCa-free controls. These data show that the multiplex MethyLight assay offers a significant advantage when working with limited quantities of DNA and has potential applications in research and clinical settings. PMID:24651255

Acoustic technology for high-performance disruption and extraction of plant proteins.

PubMed

Toorchi, Mahmoud; Nouri, Mohammad-Zaman; Tsumura, Makoto; Komatsu, Setsuko

2008-07-01

Acoustic technology shows the capability of protein pellet homogenization from different tissue samples of soybean and rice in a manner comparable to the ordinary mortar/pestle method and far better than the vortex/ultrasonic method with respect to the resolution of the protein pattern through two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). With acoustic technology, noncontact tissue disruption and protein pellet homogenization can be carried out in a computer-controlled manner, which ultimately increases the efficiency of the process for a large number of samples. A lysis buffer termed the T-buffer containing TBP, thiourea, and CHAPS yields an excellent result for the 2D-PAGE separation of soybean plasma membrane proteins followed by the 2D-PAGE separation of crude protein of soybean and rice tissues. For this technology, the T-buffer is preferred because protein quantification is possible by eliminating the interfering compound 2-mercaptoethanol and because of the high reproducibility of 2D-PAGE separation.

Reusable bi-directional 3ω sensor to measure thermal conductivity of 100-μm thick biological tissues.

PubMed

Lubner, Sean D; Choi, Jeunghwan; Wehmeyer, Geoff; Waag, Bastian; Mishra, Vivek; Natesan, Harishankar; Bischof, John C; Dames, Chris

2015-01-01

Accurate knowledge of the thermal conductivity (k) of biological tissues is important for cryopreservation, thermal ablation, and cryosurgery. Here, we adapt the 3ω method-widely used for rigid, inorganic solids-as a reusable sensor to measure k of soft biological samples two orders of magnitude thinner than conventional tissue characterization methods. Analytical and numerical studies quantify the error of the commonly used "boundary mismatch approximation" of the bi-directional 3ω geometry, confirm that the generalized slope method is exact in the low-frequency limit, and bound its error for finite frequencies. The bi-directional 3ω measurement device is validated using control experiments to within ±2% (liquid water, standard deviation) and ±5% (ice). Measurements of mouse liver cover a temperature ranging from -69 °C to +33 °C. The liver results are independent of sample thicknesses from 3 mm down to 100 μm and agree with available literature for non-mouse liver to within the measurement scatter.

Increased Expression of Cathelicidin by Direct Activation of Protease-Activated Receptor 2: Possible Implications on the Pathogenesis of Rosacea

PubMed Central

Kim, Ji Young; Kim, Yoon Jee; Lim, Beom Jin; Sohn, Hyo Jung; Shin, Dongyun

2014-01-01

Purpose Recent findings of increased cathelicidin protein and its proteolytic fragments in rosacea suggest a pathogenic role for cathelicidin in this disease. The relationship between cathelicidin and protease-activated receptor 2 (PAR-2) is therefore of interest, as PAR-2, expressed principally in keratinocytes, regulates pro-inflammatory cytokine expression in the skin. The purpose of this study was to determine the relationship between expression of PAR-2 and cathelicidin in rosacea and to test the effect of direct PAR-2 activation on cathelicidin expression in keratinocytes. Materials and Methods Samples from 40 patients with clinicopathologic diagnosis of rosacea and facial skin tissue samples from 20 patients with no specific findings or milium without inflammation were retrieved. Intensities of immunohistochemical staining for PAR-2 and cathelicidin were compared between normal and rosacea-affected skin tissues. Additionally, correlations between PAR-2 and cathelicidin staining intensities within rosacea patients were analyzed. In cultured keratinocytes, changes in PAR-2, cathelicidin, and vascular endothelial growth factor (VEGF) mRNA and protein were analyzed after treatment with PAR-2 activating peptide (AP). Results Cathelicidin expression was significantly higher in rosacea skin tissues than in normal tissues (p<0.001), while PAR-2 expression was not significantly higher in rosacea tissues than in normal skin tissues. A positive correlation between PAR-2 and cathelicidin within rosacea samples was observed (R=0.330, p=0.037). After treatment of PAR-2 AP, both mRNA and protein levels for PAR-2, cathelicidin, and VEGF significantly increased in cultured keratinocytes, compared with PAR-2 control peptide treatment. Conclusion PAR-2 may participate in the pathogenesis of rosacea through activation of cathelicidin LL-37, a mediator of innate immune responses in the skin. PMID:25323904

Extraction of tamoxifen and its metabolites from formalin-fixed, paraffin-embedded tissues: an innovative quantitation method using liquid chromatography and tandem mass spectrometry.

PubMed

Ng, Ella S M; Kangarloo, S Bill; Konno, Mie; Paterson, Alexander; Magliocco, Anthony M

2014-03-01

Tamoxifen is a key therapeutic option for breast cancer treatment. Understanding its complex metabolism and pharmacokinetics is important for dose optimization. We examined the possibility of utilizing archival formalin-fixed paraffin-embedded (FFPE) tissue as an alternative sample source for quantification since well-annotated retrospective samples were always limited. Six 15 μm sections of FFPE tissues were deparaffinized with xylene and purified using solid-phase extraction. Tamoxifen and its metabolites were separated and detected by liquid chromatography-tandem mass spectrometry using multiple-reaction monitoring. This method was linear between 0.4 and 200 ng/g for 4-hydroxy-tamoxifen and endoxifen, and 4-2,000 ng/g for tamoxifen and N-desmethyl-tamoxifen. Inter- and intra-assay precisions were <9 %, and mean accuracies ranged from 81 to 106 %. Extraction recoveries were between 83 and 88 %. The validated method was applied to FFPE tissues from two groups of patients, who received 20 mg/day of tamoxifen for >6 months, and were classified into breast tumor recurrence and non-recurrence. Our preliminary data show that levels of tamoxifen metabolites were significantly lower in patients with recurrent cancer, suggesting that inter-individual variability in tamoxifen metabolism might partly account for the development of cancer recurrence. Nevertheless, other causes such as non-compliance or stopping therapy of tamoxifen could possibly lead to the concentration differences. The ability to successfully study tamoxifen metabolism in such tissue samples will rapidly increase our knowledge of how tamoxifen's action, metabolism and tissue distribution contribute to breast cancer control. However, larger population studies are required to understand the underlying mechanism of tamoxifen metabolism for optimization of its treatment.

Blood and tissue tocopherol levels in rats following intraperitoneally administered alpha-tocopheryl acetate.

PubMed

McGee, C D; Greenwood, C E; Jeejeebhoy, K N

1990-01-01

The correction or maintenance of blood and tissue alpha-tocopherol (alpha-Toc) levels by intraperitoneally administered all-rac-alpha-tocopheryl acetate (alpha-Tac) was compared with RRR- alpha-tocopherol (alpha-Toc) in vitamin E-depleted and control rats. Rats received 1.3 TE vitamin E daily for 7 days. alpha-Tac was detected in plasma of one-third of alpha-Tac-treated rats 24 hr after the first treatment, although not in subsequent samplings. Both alpha-Tac and alpha-Toc increased tocopherol levels in plasma and liver of E-deprived rats, while little or no change was observed in adipose tissue and brain. Similarly, control rats treated with alpha-Tac or alpha-Toc had significantly greater (p less than 0.05) plasma and liver alpha-Toc levels at day 3 and day 7 than did saline-treated rats. There was no significant difference in adipose alpha-Toc levels among treatment groups of control rats. The results of this study suggest that alpha-Tac is rapidly hydrolyzed to its biologically active alcohol form and results in similar effects to that of intraperitoneally administered alpha-Toc.

Quantification of HCV RNA in Liver Tissue by bDNA Assay.

PubMed

Dailey, P J; Collins, M L; Urdea, M S; Wilber, J C

1999-01-01

With this statement, Sherlock and Dooley have described two of the three major challenges involved in quantitatively measuring any analyte in tissue samples: the distribution of the analyte in the tissue; and the standard of reference, or denominator, with which to make comparisons between tissue samples. The third challenge for quantitative measurement of an analyte in tissue is to ensure reproducible and quantitative recovery of the analyte on extraction from tissue samples. This chapter describes a method that can be used to measure HCV RNA quantitatively in liver biopsy and tissue samples using the bDNA assay. All three of these challenges-distribution, denominator, and recovery-apply to the measurement of HCV RNA in liver biopsies.

A highly sensitive method for analysis of 7-dehydrocholesterol for the study of Smith-Lemli-Opitz syndrome[S

PubMed Central

Liu, Wei; Xu, Libin; Lamberson, Connor; Haas, Dorothea; Korade, Zeljka; Porter, Ned A.

2014-01-01

We describe a highly sensitive method for the detection of 7-dehydrocholesterol (7-DHC), the biosynthetic precursor of cholesterol, based on its reactivity with 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD) in a Diels-Alder cycloaddition reaction. Samples of biological tissues and fluids with added deuterium-labeled internal standards were derivatized with PTAD and analyzed by LC-MS. This protocol permits fast processing of samples, short chromatography times, and high sensitivity. We applied this method to the analysis of cells, blood, and tissues from several sources, including human plasma. Another innovative aspect of this study is that it provides a reliable and highly reproducible measurement of 7-DHC in 7-dehydrocholesterol reductase (Dhcr7)-HET mouse (a model for Smith-Lemli-Opitz syndrome) samples, showing regional differences in the brain tissue. We found that the levels of 7-DHC are consistently higher in Dhcr7-HET mice than in controls, with the spinal cord and peripheral nerve showing the biggest differences. In addition to 7-DHC, sensitive analysis of desmosterol in tissues and blood was also accomplished with this PTAD method by assaying adducts formed from the PTAD “ene” reaction. The method reported here may provide a highly sensitive and high throughput way to identify at-risk populations having errors in cholesterol biosynthesis. PMID:24259532

An evaluation of the histological effects of intra-articular methadone in the canine model.

PubMed

Jones, Timothy A; Hand, Walter R; Ports, Michael D; Unger, Daniel V; Herbert, Daniel; Pellegrini, Joseph E

2003-02-01

Methadone hydrochloride is an opiate that has pharmacodynamic and pharmacokinetic properties that suggest it may provide longer analgesia than morphine when administered via the intra-articular route. However, no studies to date have been conducted examining the effects of intra-articular methadone hydrochloride on local tissues. Therefore, the purpose of this study was to determine the histopathologic effects of intra-articular methadone hydrochloride on local tissues in the canine knee. Nine canines, 1 to 4 years old, weighing between 20 kg and 23 kg were used. All canines had their knees randomized to receive either bupivacaine, 0.5% with epinephrine 1:200,000 (4.5 mL), and 5 mg methadone hydrochloride (0.5 mL) for the study knee, or bupivacaine, 0.5% with epinephrine 1:200,000 (4.5 mL), and 0.5 mL normal saline for the control knee. Serum methadone hydrochloride levels were obtained on all canines at 6 and 24 hours. Canines were randomly assigned to 1 of 3 groups to be euthanized at either 24 hours, 14 days, or 28 days. Following euthanization and necropsy, synovial fluid levels and tissue samples were obtained and examined for histopathologic changes. Synovial fluid samples noted a few white blood cells at 24 hours and none at 14 and 28 days. Tissue samples showed no histopathologic changes, and serum concentration levels of methadone hydrochloride were negligible.

Detection and a possible link between parvovirus B19 and thyroid cancer.

PubMed

Etemadi, Ashkan; Mostafaei, Shayan; Yari, Kheirollah; Ghasemi, Amir; Minaei Chenar, Hamzeh; Moghoofei, Mohsen

2017-06-01

Human parvovirus B19 (B19) is a small, non-enveloped virus and belongs to Parvoviridae family. B19 persists in many tissues such as thyroid tissue and even thyroid cancer. The main aim of this study was to determine the presence of B19, its association with increased inflammation in thyroid tissue, and thus its possible role in thyroid cancer progression. Studies have shown that virus replication in non-permissive tissue leads to overexpression of non-structural protein and results in upregulation of proinflammatory cytokines such as interleukin 6 and tumor necrosis factor alpha. A total of 36 paraffin-embedded thyroid specimens and serum were collected from patients and 12 samples were used as control. Various methods were employed, including polymerase chain reaction, real-time polymerase chain reaction, and enzyme-linked immunosorbent assay. The results have shown the presence of B19 DNA in 31 of 36 samples (86.11%). Almost in all samples, the levels of non-structural protein 1, nuclear factor kappa B, tumor necrosis factor alpha, and interleukin 6 were simultaneously high. The presence of parvovirus B19 has a significant positive correlation with nuclear factor kappa B, tumor necrosis factor alpha, and interleukin 6 levels. This study suggests that B19 infection may play an important role in tumorigenesis and thyroid cancer development via the inflammatory mechanisms.

Validation of reference genes aiming accurate normalization of qRT-PCR data in Dendrocalamus latiflorus Munro.

PubMed

Liu, Mingying; Jiang, Jing; Han, Xiaojiao; Qiao, Guirong; Zhuo, Renying

2014-01-01

Dendrocalamus latiflorus Munro distributes widely in subtropical areas and plays vital roles as valuable natural resources. The transcriptome sequencing for D. latiflorus Munro has been performed and numerous genes especially those predicted to be unique to D. latiflorus Munro were revealed. qRT-PCR has become a feasible approach to uncover gene expression profiling, and the accuracy and reliability of the results obtained depends upon the proper selection of stable reference genes for accurate normalization. Therefore, a set of suitable internal controls should be validated for D. latiflorus Munro. In this report, twelve candidate reference genes were selected and the assessment of gene expression stability was performed in ten tissue samples and four leaf samples from seedlings and anther-regenerated plants of different ploidy. The PCR amplification efficiency was estimated, and the candidate genes were ranked according to their expression stability using three software packages: geNorm, NormFinder and Bestkeeper. GAPDH and EF1α were characterized to be the most stable genes among different tissues or in all the sample pools, while CYP showed low expression stability. RPL3 had the optimal performance among four leaf samples. The application of verified reference genes was illustrated by analyzing ferritin and laccase expression profiles among different experimental sets. The analysis revealed the biological variation in ferritin and laccase transcript expression among the tissues studied and the individual plants. geNorm, NormFinder, and BestKeeper analyses recommended different suitable reference gene(s) for normalization according to the experimental sets. GAPDH and EF1α had the highest expression stability across different tissues and RPL3 for the other sample set. This study emphasizes the importance of validating superior reference genes for qRT-PCR analysis to accurately normalize gene expression of D. latiflorus Munro.

Effect of glucose on fatigue-induced changes in the microstructure and mechanical properties of demineralized bovine cortical bone.

PubMed

Trębacz, Hanna; Zdunek, Artur; Wlizło-Dyś, Ewa; Cybulska, Justyna; Pieczywek, Piotr

2015-10-16

The aim of this study was to test a hypothesis that fatigue-induced weakening of cortical bone was intensified in bone incubated in glucose and that this weakening is revealed in the microstructure and mechanical competence of the bone matrix. Cubic specimens of bovine femoral shaft were incubated in glucose solution (G) or in buffer (NG). One half of G samples and one half of NG were axially loaded in 300 cycles (30 mm/min) at constant deformation (F); the other half was a control (C). Samples from each group (GF, NGF, GC, NGC) were completely demineralized. Slices from demineralized samples were used for microscopic image analysis. A combined effect of glycation and fatigue on demineralized bone was tested in compression (10 mm/min). Damage of samples during the test was examined in terms of acoustic emission analysis (AE). During the fatigue procedure, resistance to loading in glycated samples decreased by 14.5% but only by 8.1% in nonglycated samples. In glycated samples fatigue resulted in increased porosity with pores significantly larger than in the other groups. Under compression, strain at failure in demineralized bone was significantly affected by glucose and fatigue. AE from demineralized bone matrix was considerably related to the largest pores in the tissue. The results confirm the hypothesis that the effect of fatigue on cortical bone tissue was intensified after incubation in glucose, both in the terms of the mechanical competence of bone tissue and the structural changes in the collagenous matrix of bone.

[Forensic medical implications of histomorphological changes in the bone and cartilage tissues under effect of radiation].

PubMed

Osipenkova-Vichtomova, T K

2013-01-01

The objective of the present work was to study roentgenological, microscopic, and histomorphological changes in the bone and cartilage tissues under effect of different doses of gamma-ray radiation from Gammatron-2 (GUT Co 400) and betatron bremsstrahlung radiation (25 MeV). The total radiation dose varied from 9.6 Gy to 120 Gy per unit area during 5-8 weeks. The study included 210 patients at the age from 7 to 82 years (97 men and 113 women). Histomorphological studies were carried out using samples of bone and cartilage tissues taken from different body regions immediately after irradiation and throughout the follow-up period of up to 4 years 6 months. Control samples were the unexposed bone and cartilage tissues from the same subjects (n = 14). The tissues were stained either with eosin and hematoxylin or by Van Gieson's and Mallory's methods. Gomori's nonspecific staining was used to detect acid and alkaline phosphatase activities. Moreover, argyrophilic substance was identified in the cartilaginous tissue. Best's carmine was used for glycogen staining and Weigert's stain for elastic fibers. Metachromasia was revealed by toluidine blue staining and fat by the sudan III staining technique. In addition, the ultrastructure of cartilaginous tissue was investigated. Taken together, these methods made it possible to identify the signs of radiation-induced damage to the bone and cartilage tissues in conjunction with complications that are likely to develop at different periods after irradiation including such ones as spontaneous fractures, deforming arthrosis and radiation-induced tumours.

NASA Robotic Neurosurgery Testbed

NASA Technical Reports Server (NTRS)

Mah, Robert

1997-01-01

The detection of tissue interface (e.g., normal tissue, cancer, tumor) has been limited clinically to tactile feedback, temperature monitoring, and the use of a miniature ultrasound probe for tissue differentiation during surgical operations, In neurosurgery, the needle used in the standard stereotactic CT or MRI guided brain biopsy provides no information about the tissue being sampled. The tissue sampled depends entirely upon the accuracy with which the localization provided by the preoperative CT or MRI scan is translated to the intracranial biopsy site. In addition, no information about the tissue being traversed by the needle (e.g., a blood vessel) is provided. Hemorrhage due to the biopsy needle tearing a blood vessel within the brain is the most devastating complication of stereotactic CT/MRI guided brain biopsy. A robotic neurosurgery testbed has been developed at NASA Ames Research Center as a spin-off of technologies from space, aeronautics and medical programs. The invention entitled "Robotic Neurosurgery Leading to Multimodality Devices for Tissue Identification" is nearing a state ready for commercialization. The devices will: 1) improve diagnostic accuracy and precision of general surgery, with near term emphasis on stereotactic brain biopsy, 2) automate tissue identification, with near term emphasis on stereotactic brain biopsy, to permit remote control of the procedure, and 3) reduce morbidity for stereotactic brain biopsy. The commercial impact from this work is the potential development of a whole new generation of smart surgical tools to increase the safety, accuracy and efficiency of surgical procedures. Other potential markets include smart surgical tools for tumor ablation in neurosurgery, general exploratory surgery, prostate cancer surgery, and breast cancer surgery.

NASA Robotic Neurosurgery Testbed

NASA Technical Reports Server (NTRS)

Mah, Robert

1997-01-01

The detection of tissue interface (e.g., normal tissue, cancer, tumor) has been limited clinically to tactile feedback, temperature monitoring, and the use of a miniature ultrasound probe for tissue differentiation during surgical operations. In neurosurgery, the needle used in the standard stereotactic CT (Computational Tomography) or MRI (Magnetic Resonance Imaging) guided brain biopsy provides no information about the tissue being sampled. The tissue sampled depends entirely upon the accuracy with which the localization provided by the preoperative CT or MRI scan is translated to the intracranial biopsy site. In addition, no information about the tissue being traversed by the needle (e.g., a blood vessel) is provided. Hemorrhage due to the biopsy needle tearing a blood vessel within the brain is the most devastating complication of stereotactic CT/MRI guided brain biopsy. A robotic neurosurgery testbed has been developed at NASA Ames Research Center as a spin-off of technologies from space, aeronautics and medical programs. The invention entitled 'Robotic Neurosurgery Leading to Multimodality Devices for Tissue Identification' is nearing a state ready for commercialization. The devices will: 1) improve diagnostic accuracy and precision of general surgery, with near term emphasis on stereotactic brain biopsy, 2) automate tissue identification, with near term emphasis on stereotactic brain biopsy, to permit remote control of the procedure, and 3) reduce morbidity for stereotactic brain biopsy. The commercial impact from this work is the potential development of a whole new generation of smart surgical tools to increase the safety, accuracy and efficiency of surgical procedures. Other potential markets include smart surgical tools for tumor ablation in neurosurgery, general exploratory surgery, prostate cancer surgery, and breast cancer surgery.

Visible effects of rapamycin (sirolimus) on human skin explants in vitro.

PubMed

Peramo, Antonio; Marcelo, Cynthia L

2013-03-01

In this manuscript, we report observations of the effects of rapamycin in an organotypic culture of human skin explants. The tissues were cultured for 5 days at the air-liquid interface or in submersed conditions with media with and without rapamycin at 2 nM concentration. Histological analysis of tissue sections indicated that rapamycin-treated samples maintained a better epidermal structure in the upper layers of the tissue than untreated samples, mostly evident when skin was cultured in submersed conditions. A significant decrease in the number of positive proliferative cells using the Ki67 antigen was observed when specimens were treated with rapamycin, in both air-liquid and submersed conditions but apoptosis differences between treated and untreated specimens, as seen by cleaved caspase-3 positive cells, were only observed in submersed specimens. Finally, a decrease and variability in the location in the expression of the differentiation marker involucrin and in E-cadherin were also evident in submersed samples. These results suggest that the development of topical applications containing rapamycin, instead of systemic delivery, may be a useful tool in the treatment of skin diseases that require reduction of proliferation and modulation or control of keratinocyte differentiation.

[Mechanism of action for oligomeric proanthocyaniclins in pava qnat-induced acute lung injury].

PubMed

Liu, P; Zhou, Y S; Qin, Y L; Li, L; Liu, Y; Xu, B; Huang, K; Ji, C C; Lin, F; Wang, Y G; Li, K; Chen, S H; Shao, L F; Mu, J S

2017-11-20

Objective: The present study was designed to evaluate the protective effects of oligomeric proanthocyanidins (OPC) in mice exposed to paraquat (PQ) , and to explore the molecular mechanism. Methods: Four experimental groups were designed. 10 BALB/c mice were intraperitoneally injected with normal saline) . PQ group: 10 BALB/c mice were intraperitoneally injected with PQ (100 mg/kg) . PQ+OPC group: 10 BALB/c mice were administered with OPC (100 mg/kg) for 1 h before PQ (100 mg/kg) expo-sure. OPC group: 10 BALB/c mice were intraperitoneally injected with OPC (100 mg/kg) . The peripheral blood samples or lung tissue samples were collected at the designed time points for measuring the levels of oxi-dative stress indicators, the related protein levels of nuclear factor-kappa B (NF-κB) pathway and nuclear fac-tor erythroid related factor-2 (Nrf2) pathway. Results: Compared with the control group, the level of reactive oxygen species (ROS) , the content of malondialdehyde (MDA) in the PQ group were significantly induced, and the activity of superoxide dismutase (SOD) in the PQ group was decreased in the peripheral blood. As com-pared with the PQ group, the level of ROS and the content of MDA in the PQ+OPC group were significantly re-duced, the activity SOD in the PQ+OPC group was increased in the peripheral blood; the level of ROS and the content of MDA were also reduced in lung tissues in the PQ+OPC group. Moreover, compared with the con-trol group, the phosphorylation of IκBα and the expression of NF-κB p65 were increased in lung tissues in the PQ group. The phosphorylation of IκBα and the expression of NF-κB p65 were decreased in lung tissues in the PQ+OPC group as compared with the PQ group. In addition, compared with the control group, the expressions of HO-1 and Nrf2 were increased in lung tissues in OPC group, and these were decreased in lung tissues in PQ groups. Furthermore, the expressions of HO-1 and Nrf2 were also increased in lung tissues in PQ+OPC as com-pared with the PQ group. Conclusion: OPC could alleviate PQ-induced systemic toxicity in mice by regulating oxidative stress via NF-κB and Nrf2 pathway.

Mechanical characterization of matrix-induced autologous chondrocyte implantation (MACI®) grafts in an equine model at 53 weeks.

PubMed

Griffin, Darvin J; Bonnevie, Edward D; Lachowsky, Devin J; Hart, James C A; Sparks, Holly D; Moran, Nance; Matthews, Gloria; Nixon, Alan J; Cohen, Itai; Bonassar, Lawrence J

2015-07-16

There has been much interest in using autologous chondrocytes in combination with scaffold materials to aid in cartilage repair. In the present study, a total of 27 animals were used to compare the performance of matrix-assisted chondrocyte implantation (MACI®) using a collagen sponge as a chondrocyte delivery vehicle, the sponge membrane alone, and empty controls. A total of three distinct types of mechanical analyses were performed on repaired cartilage harvested from horses after 53 weeks of implantation: (1) compressive behavior of samples to measure aggregate modulus (HA) and hydraulic permeability (k) in confined compression; (2) local and global shear modulus using confocal strain mapping; and (3) boundary friction coefficient using a custom-built tribometer. Cartilage defects receiving MACI® implants had equilibrium modulus values that were 70% of normal cartilage, and were not statistically different than normal tissue. Defects filled with Maix™ membrane alone or left empty were only 46% and 51-63% of control, respectively. The shear modulus of tissue from all groups of cartilage defects were between 4 and 10 times lower than control tissue, and range from 0.2 to 0.4 MPa. The average values of boundary mode friction coefficients of control tissue from all groups ranged from 0.42 to 0.52. This study represents an extensive characterization of the mechanical performance of the MACI® grafts implant in a large animal model at 53 weeks. Collectively, these data demonstrate a range of implant performance, revealing similar compressive and frictional properties to native tissue, with inferior shear properties. Copyright © 2015. Published by Elsevier Ltd.

Recent progress in tissue optical clearing for spectroscopic application

NASA Astrophysics Data System (ADS)

Sdobnov, A. Yu.; Darvin, M. E.; Genina, E. A.; Bashkatov, A. N.; Lademann, J.; Tuchin, V. V.

2018-05-01

This paper aims to review recent progress in optical clearing of the skin and over naturally turbid biological tissues and blood using this technique in vivo and in vitro with multiphoton microscopy, confocal Raman microscopy, confocal microscopy, NIR spectroscopy, optical coherence tomography, and laser speckle contrast imaging. Basic principles of the technique, its safety, advantages and limitations are discussed. The application of optical clearing agent on a tissue allows for controlling the optical properties of tissue. Optical clearing-induced reduction of tissue scattering significantly facilitates the observation of deep-located tissue regions, at the same time improving the resolution and image contrast for a variety of optical imaging methods suitable for clinical applications, such as diagnostics and laser treatment of skin diseases, mucosal tumor imaging, laser disruption of pathological abnormalities, etc. Structural images of different skin layers obtained ex vivo for porcine ear skin samples at application of Omnipaque™ and glycerol solutions during 60 min. Red color corresponds to TPEAF signal channel. Green color corresponds to SHG signal channel.

Imaging of the interaction of low frequency electric fields with biological tissues by optical coherence tomography

NASA Astrophysics Data System (ADS)

Peña, Adrian F.; Devine, Jack; Doronin, Alexander; Meglinski, Igor

2014-03-01

We report the use of conventional Optical Coherence Tomography (OCT) for visualization of propagation of low frequency electric field in soft biological tissues ex vivo. To increase the overall quality of the experimental images an adaptive Wiener filtering technique has been employed. Fourier domain correlation has been subsequently applied to enhance spatial resolution of images of biological tissues influenced by low frequency electric field. Image processing has been performed on Graphics Processing Units (GPUs) utilizing Compute Unified Device Architecture (CUDA) framework in the frequencydomain. The results show that variation in voltage and frequency of the applied electric field relates exponentially to the magnitude of its influence on biological tissue. The magnitude of influence is about twice more for fresh tissue samples in comparison to non-fresh ones. The obtained results suggest that OCT can be used for observation and quantitative evaluation of the electro-kinetic changes in biological tissues under different physiological conditions, functional electrical stimulation, and potentially can be used non-invasively for food quality control.

Optical coherence tomography can assess skeletal muscle tissue from mouse models of muscular dystrophy by parametric imaging of the attenuation coefficient

PubMed Central

Klyen, Blake R.; Scolaro, Loretta; Shavlakadze, Tea; Grounds, Miranda D.; Sampson, David D.

2014-01-01

We present the assessment of ex vivo mouse muscle tissue by quantitative parametric imaging of the near-infrared attenuation coefficient µt using optical coherence tomography. The resulting values of the local total attenuation coefficient µt (mean ± standard error) from necrotic lesions in the dystrophic skeletal muscle tissue of mdx mice are higher (9.6 ± 0.3 mm−1) than regions from the same tissue containing only necrotic myofibers (7.0 ± 0.6 mm−1), and significantly higher than values from intact myofibers, whether from an adjacent region of the same sample (4.8 ± 0.3 mm−1) or from healthy tissue of the wild-type C57 mouse (3.9 ± 0.2 mm−1) used as a control. Our results suggest that the attenuation coefficient could be used as a quantitative means to identify necrotic lesions and assess skeletal muscle tissue in mouse models of human Duchenne muscular dystrophy. PMID:24761302

Combined Bisulfite Restriction Analysis for brain tissue identification.

PubMed

Samsuwan, Jarunya; Muangsub, Tachapol; Yanatatsaneejit, Pattamawadee; Mutirangura, Apiwat; Kitkumthorn, Nakarin

2018-05-01

According to the tissue-specific methylation database (doi: 10.1016/j.gene.2014.09.060), methylation at CpG locus cg03096975 in EML2 has been preliminarily proven to be specific to brain tissue. In this study, we enlarged sample size and developed a technique for identifying brain tissue in aged samples. Combined Bisulfite Restriction Analysis-for EML2 (COBRA-EML2) technique was established and validated in various organ samples obtained from 108 autopsies. In addition, this technique was also tested for its reliability, minimal DNA concentration detected, and use in aged samples and in samples obtained from specific brain compartments and spinal cord. COBRA-EML2 displayed 100% sensitivity and specificity for distinguishing brain tissue from other tissues, showed high reliability, was capable of detecting minimal DNA concentration (0.015ng/μl), could be used for identifying brain tissue in aged samples. In summary, COBRA-EML2 is a technique to identify brain tissue. This analysis is useful in criminal cases since it can identify the vital organ tissues from small samples acquired from criminal scenes. The results from this analysis can be counted as a medical and forensic marker supporting criminal investigations, and as one of the evidences in court rulings. Copyright © 2018 Elsevier B.V. All rights reserved.

Polymorphism and Expression Profile of Cholecystokinin Type A Receptor in Relation to Gallstone Disease Susceptibility.

PubMed

Kazmi, Hasan Raza; Chandra, Abhijit; Nigam, Jaya; Baghel, Kavita; Srivastava, Meenu; Maurya, Shailendra S; Parmar, Devendra

2016-10-01

In the present study, we investigated expression pattern of Cholecystokinin type A receptor (CCKAR) in relation to its commonly studied polymorphism (rs1800857, T/C) in gallstone disease (GSD) patients and controls. A total of 502 subjects (272 GSD and 230 controls) were enrolled, and genotyping was performed by evaluating restriction fragments of PstI digested DNA. For analyzing expression pattern of CCKAR in relation to polymorphism, gallbladder tissue samples from 80 subjects (GSD-55; control-25) were studied. Expression of CCKAR mRNA was evaluated by reverse transcriptase-PCR and confirmed using real-time PCR. Protein expression was evaluated by enzyme-linked immunosorbent assay. We observed significantly (p < 0.0001) lower expression of CCKAR mRNA and protein in GSD tissues as compared with control. Significantly higher frequency of A1/A1 genotype (C/T transition) (p = 0.0005) was observed for GSD as compared with control. Expression of CCKAR protein was found to be significantly lower (p < 0.0001) in A1/A1 genotype as compared with other genotypes for GSD patients. Perhaps, this is the first report providing evidence of alteration in CCKAR expression in relation to its polymorphism elucidating the molecular pathway of the disease. Additional investigations with lager sample size are needed to confirm these findings.

Detection of IS6110 and HupB gene sequences of Mycobacterium tuberculosis and bovis in the aortic tissue of patients with Takayasu’s arteritis

PubMed Central

2012-01-01

Background Takayasu’s arteritis (TA) is a chronic inflammatory disease affecting the large arteries and their branches; its etiology is still unknown. In individuals suffering from TA, arterial inflammation progresses to stenosis and/or occlusion, leading to organ damage and affecting survival. Relation of TA with Mycobacterium tuberculosis has been known, but there have been only a few systematic studies focusing on this association. The IS6110 sequence identifies the Mycobacterium tuberculosis complex and the HupB establishes the differences between M. tuberculosis and M. bovis. Our objective was to search the presence of IS6110 and HupB genes in aorta of patients with TA. Methods We analyzed aorta tissues embedded in paraffin from 5760 autopsies obtained from our institution, we divided the selected samples as cases and controls; Cases: aortic tissues of individuals with Takayasu’s arteritis. Control positive: aortic tissues (with tuberculosis disease confirmed) and control negative with other disease aortic (atherosclerosis). Results Of 181 selected aorta tissues, 119 fulfilled the corresponding criteria for TA, TB or atherosclerosis. Thus 33 corresponded to TA, 33 to tuberculosis (TB) and 53 to atherosclerosis. The mean age was 22 ± 13, 41 ± 19, and 57 ± 10, respectively. IS6110 and HupB sequences were detected in 70% of TA tissues, 82% in tuberculosis, and in 32% with atherosclerosis. Important statistical differences between groups with TA, tuberculosis versus atherosclerosis (p = 0.004 and 0.0001, respectively) were found. Conclusion We identified a higher frequency of IS6110 and HupB genes in aortic tissues of TA patients. This data suggests that arterial damage could occur due to previous infection with M. tuberculosis. PMID:22905864

Tissue spray ionization mass spectrometry for rapid recognition of human lung squamous cell carcinoma

NASA Astrophysics Data System (ADS)

Wei, Yiping; Chen, Liru; Zhou, Wei; Chingin, Konstantin; Ouyang, Yongzhong; Zhu, Tenggao; Wen, Hua; Ding, Jianhua; Xu, Jianjun; Chen, Huanwen

2015-05-01

Tissue spray ionization mass spectrometry (TSI-MS) directly on small tissue samples has been shown to provide highly specific molecular information. In this study, we apply this method to the analysis of 38 pairs of human lung squamous cell carcinoma tissue (cancer) and adjacent normal lung tissue (normal). The main components of pulmonary surfactants, dipalmitoyl phosphatidylcholine (DPPC, m/z 757.47), phosphatidylcholine (POPC, m/z 782.52), oleoyl phosphatidylcholine (DOPC, m/z 808.49), and arachidonic acid stearoyl phosphatidylcholine (SAPC, m/z 832.43), were identified using high-resolution tandem mass spectrometry. Monte Carlo sampling partial least squares linear discriminant analysis (PLS-LDA) was used to distinguish full-mass-range mass spectra of cancer samples from the mass spectra of normal tissues. With 5 principal components and 30 - 40 Monte Carlo samplings, the accuracy of cancer identification in matched tissue samples reached 94.42%. Classification of a tissue sample required less than 1 min, which is much faster than the analysis of frozen sections. The rapid, in situ diagnosis with minimal sample consumption provided by TSI-MS is advantageous for surgeons. TSI-MS allows them to make more informed decisions during surgery.

An informatics model for tissue banks--lessons learned from the Cooperative Prostate Cancer Tissue Resource.

PubMed

Patel, Ashokkumar A; Gilbertson, John R; Parwani, Anil V; Dhir, Rajiv; Datta, Milton W; Gupta, Rajnish; Berman, Jules J; Melamed, Jonathan; Kajdacsy-Balla, Andre; Orenstein, Jan; Becich, Michael J

2006-05-05

Advances in molecular biology and growing requirements from biomarker validation studies have generated a need for tissue banks to provide quality-controlled tissue samples with standardized clinical annotation. The NCI Cooperative Prostate Cancer Tissue Resource (CPCTR) is a distributed tissue bank that comprises four academic centers and provides thousands of clinically annotated prostate cancer specimens to researchers. Here we describe the CPCTR information management system architecture, common data element (CDE) development, query interfaces, data curation, and quality control. Data managers review the medical records to collect and continuously update information for the 145 clinical, pathological and inventorial CDEs that the Resource maintains for each case. An Access-based data entry tool provides de-identification and a standard communication mechanism between each group and a central CPCTR database. Standardized automated quality control audits have been implemented. Centrally, an Oracle database has web interfaces allowing multiple user-types, including the general public, to mine de-identified information from all of the sites with three levels of specificity and granularity as well as to request tissues through a formal letter of intent. Since July 2003, CPCTR has offered over 6,000 cases (38,000 blocks) of highly characterized prostate cancer biospecimens, including several tissue microarrays (TMA). The Resource developed a website with interfaces for the general public as well as researchers and internal members. These user groups have utilized the web-tools for public query of summary data on the cases that were available, to prepare requests, and to receive tissues. As of December 2005, the Resource received over 130 tissue requests, of which 45 have been reviewed, approved and filled. Additionally, the Resource implemented the TMA Data Exchange Specification in its TMA program and created a computer program for calculating PSA recurrence. Building a biorepository infrastructure that meets today's research needs involves time and input of many individuals from diverse disciplines. The CPCTR can provide large volumes of carefully annotated prostate tissue for research initiatives such as Specialized Programs of Research Excellence (SPOREs) and for biomarker validation studies and its experience can help development of collaborative, large scale, virtual tissue banks in other organ systems.

Application of ribonucleoside vanadyl complex (RVC) for developing a multifunctional tissue preservative solution

PubMed Central

Yu, Cheng-Chia; Chen, Chin-Chuan

2018-01-01

The quality of biological samples greatly affects the accuracy of scientific results. However, RNA in cryopreserved tissues gradually degrades during storage, leading to errors in the results of subsequent experiments. A suitable sample preservative solution can prolong storage and enhance the research value of samples. Here, we developed a sample preservative solution using the properties of the ribonucleoside vanadyl complex (RVC) and compared its effects on RNA and DNA quality, protein activity, and tissue morphology with the commercially available and widely used RNAlater® Stabilization Solution. The results showed that both the RVC-based preservative solution and RNAlater can effectively delay RNA degradation in tissue samples stored at 4°C or −80°C compared with samples stored without any preservative solution. In contrast to RNAlater, the RVC-based preservative solution did not result in damage to the tissue morphology or a loss of protein activity. Additionally, the RVC-based preservative solution did not affect the RNA and genomic DNA contents of the tissue samples or the results of subsequent experimental analyses. An RVC-based reagent can be used as a multifunctional yet relatively inexpensive tissue preservative solution to provide a comprehensive and cost-effective method for preserving samples for tissue banks. PMID:29538436

Effects of Electromagnetic Radiation Use on Oxidant/Antioxidant Status and DNA Turn-over Enzyme Activities in Erythrocytes and Heart, Kidney, Liver, and Ovary Tissues From Rats: Possible Protective Role of Vitamin C.

PubMed

Devrim, Erdinç; Ergüder, Imge B; Kılıçoğlu, Bülent; Yaykaşlı, Emine; Cetin, Recep; Durak, Ilker

2008-01-01

ABSTRACT In this study, the aim was to investigate possible effects of Electromagnetic Radiation (EMR) use on oxidant and antioxidant status in erythrocytes and kidney, heart, liver, and ovary tissues from rats, and possible protective role of vitamin C. For this aim, 40 Wistar albino female rats were used throughout the study. The treatment group was exposed to EMR in a frequency of 900 MHz, the EMR plus vitamin C group was exposed to the same EMR frequency and given vitamin C (250 mg/kg/day) orally for 4 weeks. There were 10 animals in each group including control and vitamin C groups. At the end of the study period, blood samples were obtained from the animals to get erythrocyte sediments. Then the animals were sacrificed and heart, kidney, liver, and ovary tissues were removed. Malondialdehyde (MDA) levels and superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), xanthine oxidase (XO), and adenosine deaminase (ADA) enzyme activities were measured in the tissues and erythrocytes. It was observed that MDA level, XO, and GSH-Px activities significantly increased in the EMR group as compared with those of the control group in the erythrocytes. In the kidney tissues, it was found that MDA level and CAT activity significantly increased, whereas XO and ADA activities decreased in the cellular phone group as compared with those of the control group. However, in the heart tissues it was observed that MDA level, ADA, and XO activities significantly decreased in the cellular phone group as compared with those of the control group. The results suggest that EMR at the frequency generated by a cell phone causes oxidative stress and peroxidation in the erythrocytes and kidney tissues from rats. In the erythrocytes, vitamin C seems to make partial protection against the oxidant stress.

Estimation of pentraxin-3 levels in the gingival tissues of chronic and aggressive periodontitis participants: an in vivo study.

PubMed

Lakshmanan, Reema; Jayakumar, N D; Sankari, Malaiappan; Padmalatha, Ogoti; Varghese, Sheeja

2014-02-01

Pentraxins are acute-phase proteins that belong to a family of evolutionarily conserved proteins, and they are considered markers of inflammation. Pentraxin-3 (PTX3) is a prototype of the long pentraxin group. It is suggested to play an important role in innate resistance against pathogens, regulation of inflammation, and clearance of apoptotic cells. The aim of this study is to estimate the level of PTX3 in gingival tissues of individuals with chronic (CP) and aggressive (AgP) periodontitis and control participants and further correlate the level of PTX3 with clinical parameters. The study population consisted of 50 participants ranging in age from 20 to 55 years and attending the outpatient section of Department of Periodontics, Saveetha Dental College and Hospital, Chennai, India. The study groups included the following: 1) group A, patients with generalized CP (n = 20); 2) group B (n = 20), patients with generalized AgP (GAgP); and 3) group C (n = 10), healthy controls. Tissue samples from participants were assayed for PTX3 levels using enzyme-linked immunosorbent assay. Gingival tissues from patients with GAgP (8.349 ± 5.076 ng/mL) had a higher mean concentration of PTX3 than tissues from patients with generalized CP (5.068 ± 3.274 ng/mL) and controls (0.251 ± 0.277). The PTX3 levels in the gingival tissues correlated positively with clinical parameters in all the groups. Among the parameters, probing depth was the most significant predictor variable associated with PTX3 in cases with periodontitis. PTX3 concentration in gingival tissues of patients with GAgP was higher than in tissues from patients with CP, and the levels correlated positively with clinical parameters. Hence, tissue PTX3 level can be considered a marker of inflammation in periodontal disease.

Histology and Biaxial Mechanical Behavior of Abdominal Aortic Aneurysm Tissue Samples.

PubMed

Pancheri, Francesco Q; Peattie, Robert A; Reddy, Nithin D; Ahamed, Touhid; Lin, Wenjian; Ouellette, Timothy D; Iafrati, Mark D; Luis Dorfmann, A

2017-03-01

Abdominal aortic aneurysms (AAAs) represent permanent, localized dilations of the abdominal aorta that can be life-threatening if progressing to rupture. Evaluation of risk of rupture depends on understanding the mechanical behavior of patient AAA walls. In this project, a series of patient AAA wall tissue samples have been evaluated through a combined anamnestic, mechanical, and histopathologic approach. Mechanical properties of the samples have been characterized using a novel, strain-controlled, planar biaxial testing protocol emulating the in vivo deformation of the aorta. Histologically, the tissue ultrastructure was highly disrupted. All samples showed pronounced mechanical stiffening with stretch and were notably anisotropic, with greater stiffness in the circumferential than the axial direction. However, there were significant intrapatient variations in wall stiffness and stress. In biaxial tests in which the longitudinal stretch was held constant at 1.1 as the circumferential stretch was extended to 1.1, the maximum average circumferential stress was 330 ± 70 kPa, while the maximum average axial stress was 190 ± 30 kPa. A constitutive model considering the wall as anisotropic with two preferred directions fit the measured data well. No statistically significant differences in tissue mechanical properties were found based on patient gender, age, maximum bulge diameter, height, weight, body mass index, or smoking history. Although a larger patient cohort is merited to confirm these conclusions, the project provides new insight into the relationships between patient natural history, histopathology, and mechanical behavior that may be useful in the development of accurate methods for rupture risk evaluation.

Histological and Thermometric Examination of Soft Tissue De-Epithelialization Using Digitally Controlled Er:YAG Laser Handpiece: An Ex Vivo Study.

PubMed

Grzech-Leśniak, Kinga; Matys, Jacek; Jurczyszyn, Kamil; Ziółkowski, Piotr; Dominiak, Marzena; Brugnera Junior, Aldo; Romeo, Umberto

2018-06-01

The purpose of this study was histological and thermometric examination of soft tissue de-epithelialization using digitally controlled laser handpiece (DCLH) - X-Runner. Commonly used techniques for de-epithelialization include scalpel, abrasion with diamond bur, or a combination of the two. Despite being simple, inexpensive and effective, these techniques are invasive and may produce unwanted side effects. It is important to look for alternative techniques using novel tools, which are minimally invasive and effective. 114 porcine samples sized 6 × 6 mm were collected from the attached gingiva (AG) of the alveolar process of the mandible using 15C scalpel blade. The samples were irradiated by means of Er:YAG laser (LightWalker, Fotona, Slovenia), using X-Runner and HO 2 handpieces at different parameters; 80, 100, and 140 mJ/20 Hz in time of 6 or 16 sec, respectively. The temperature was measured with a K-type thermocouple. For the histopathological analysis of efficiency of epithelium removal and thermal injury, 3 random samples were de-epithelialized with an HO 2 handpiece, and 9 random samples with an X-Runner handpiece with different parameters. For the samples irradiated with DCLH, we have used three different settings, which resulted in removing 1 to 3 layers of the soft tissue. The efficiency of epithelium removal and the rise of temperature were analyzed. DCLH has induced significantly lower temperature increase compared with HO 2 at each energy to frequency ratio. The histological examination revealed total epithelium removal when HO 2 handpiece was used at 100 and 140 mJ/20 Hz and when DCLH was used for two- and threefold lasing at 80, 100, and 140 mJ/20 Hz. Er:YAG laser with DCLH handpiece may be an efficient tool in epithelium removal without excessive thermal damage.

Effects of sterilization on poly(ethylene glycol) hydrogels.

PubMed

Kanjickal, Deenu; Lopina, Stephanie; Evancho-Chapman, M Michelle; Schmidt, Steven; Donovan, Duane

2008-12-01

The past few decades have witnessed a dramatic increase in the development of polymeric biomaterials. These biomaterials have to undergo a sterilization procedure before implantation. However, many sterilization procedures have been shown to profoundly affect polymer properties. Poly(ethylene glycol) hydrogels have gained increasing importance in the controlled delivery of therapeutics and in tissue engineering. We evaluated the effect of ethylene oxide (EtO), hydrogen peroxide (H(2)O(2)), and gamma sterilization of poly(ethylene glycol) hydrogels on properties relevant to controlled drug delivery and tissue engineering. We observed that the release of cyclosporine (CyA) (an immunosuppressive drug that is effective in combating tissue rejection following organ transplantation) was significantly affected by the type of sterilization. However, that was not the case with rhodamine B, a dye. Hence, the drug release characteristics were observed to be dependent not only on the sterilization procedure but also on the type of agent that needs to be delivered. In addition, differences in the swelling ratios for the sterilized and unsterilized hydrogels were statistically significant for 1:1 crosslinked hydrogels derived from the 8000 MW polymer. Significant differences were also observed for gamma sterilization for 1:1 crosslinked hydrogels derived from the 3350 MW polymer and also the 2:1 crosslinked hydrogels derived from the 8000 MW polymer. Atomic force microscopy (AFM) studies revealed that the roughness parameter for the unsterilized and EtO-sterilized PEG hydrogels remained similar. However, a statistically significant reduction of the roughness parameter was observed for the H(2)O(2) and gamma-sterilized samples. Electron spin resonance (ESR) studies on the unsterilized and the sterilized samples revealed the presence of the peroxy and the triphenyl methyl carbon radical in the samples. The gamma and the H(2)O(2)-sterilized samples were observed to have a much higher concentration of the radical pecies when compared with the EtO and the unsterilized samples. (c) 2008 Wiley Periodicals, Inc. J Biomed Mater Res, 2008.

Integrative analysis for identification of shared markers from various functional cells/tissues for rheumatoid arthritis.

PubMed

Xia, Wei; Wu, Jian; Deng, Fei-Yan; Wu, Long-Fei; Zhang, Yong-Hong; Guo, Yu-Fan; Lei, Shu-Feng

2017-02-01

Rheumatoid arthritis (RA) is a systemic autoimmune disease. So far, it is unclear whether there exist common RA-related genes shared in different tissues/cells. In this study, we conducted an integrative analysis on multiple datasets to identify potential shared genes that are significant in multiple tissues/cells for RA. Seven microarray gene expression datasets representing various RA-related tissues/cells were downloaded from the Gene Expression Omnibus (GEO). Statistical analyses, testing both marginal and joint effects, were conducted to identify significant genes shared in various samples. Followed-up analyses were conducted on functional annotation clustering analysis, protein-protein interaction (PPI) analysis, gene-based association analysis, and ELISA validation analysis in in-house samples. We identified 18 shared significant genes, which were mainly involved in the immune response and chemokine signaling pathway. Among the 18 genes, eight genes (PPBP, PF4, HLA-F, S100A8, RNASEH2A, P2RY6, JAG2, and PCBP1) interact with known RA genes. Two genes (HLA-F and PCBP1) are significant in gene-based association analysis (P = 1.03E-31, P = 1.30E-2, respectively). Additionally, PCBP1 also showed differential protein expression levels in in-house case-control plasma samples (P = 2.60E-2). This study represented the first effort to identify shared RA markers from different functional cells or tissues. The results suggested that one of the shared genes, i.e., PCBP1, is a promising biomarker for RA.

Use of Sequenom Sample ID Plus® SNP Genotyping in Identification of FFPE Tumor Samples

PubMed Central

Miller, Jessica K.; Buchner, Nicholas; Timms, Lee; Tam, Shirley; Luo, Xuemei; Brown, Andrew M. K.; Pasternack, Danielle; Bristow, Robert G.; Fraser, Michael; Boutros, Paul C.; McPherson, John D.

2014-01-01

Short tandem repeat (STR) analysis, such as the AmpFlSTR® Identifiler® Plus kit, is a standard, PCR-based human genotyping method used in the field of forensics. Misidentification of cell line and tissue DNA can be costly if not detected early; therefore it is necessary to have quality control measures such as STR profiling in place. A major issue in large-scale research studies involving archival formalin-fixed paraffin embedded (FFPE) tissues is that varying levels of DNA degradation can result in failure to correctly identify samples using STR genotyping. PCR amplification of STRs of several hundred base pairs is not always possible when DNA is degraded. The Sample ID Plus® panel from Sequenom allows for human DNA identification and authentication using SNP genotyping. In comparison to lengthy STR amplicons, this multiplexing PCR assay requires amplification of only 76–139 base pairs, and utilizes 47 SNPs to discriminate between individual samples. In this study, we evaluated both STR and SNP genotyping methods of sample identification, with a focus on paired FFPE tumor/normal DNA samples intended for next-generation sequencing (NGS). The ability to successfully validate the identity of FFPE samples can enable cost savings by reducing rework. PMID:24551080

Use of Sequenom sample ID Plus® SNP genotyping in identification of FFPE tumor samples.

PubMed

Miller, Jessica K; Buchner, Nicholas; Timms, Lee; Tam, Shirley; Luo, Xuemei; Brown, Andrew M K; Pasternack, Danielle; Bristow, Robert G; Fraser, Michael; Boutros, Paul C; McPherson, John D

2014-01-01

Short tandem repeat (STR) analysis, such as the AmpFlSTR® Identifiler® Plus kit, is a standard, PCR-based human genotyping method used in the field of forensics. Misidentification of cell line and tissue DNA can be costly if not detected early; therefore it is necessary to have quality control measures such as STR profiling in place. A major issue in large-scale research studies involving archival formalin-fixed paraffin embedded (FFPE) tissues is that varying levels of DNA degradation can result in failure to correctly identify samples using STR genotyping. PCR amplification of STRs of several hundred base pairs is not always possible when DNA is degraded. The Sample ID Plus® panel from Sequenom allows for human DNA identification and authentication using SNP genotyping. In comparison to lengthy STR amplicons, this multiplexing PCR assay requires amplification of only 76-139 base pairs, and utilizes 47 SNPs to discriminate between individual samples. In this study, we evaluated both STR and SNP genotyping methods of sample identification, with a focus on paired FFPE tumor/normal DNA samples intended for next-generation sequencing (NGS). The ability to successfully validate the identity of FFPE samples can enable cost savings by reducing rework.

Rapid molecular detection of invasive species in ballast and harbor water by integrating environmental DNA and light transmission spectroscopy.

PubMed

Egan, Scott P; Grey, Erin; Olds, Brett; Feder, Jeffery L; Ruggiero, Steven T; Tanner, Carol E; Lodge, David M

2015-04-07

Invasive species introduced via the ballast water of commercial ships cause enormous environmental and economic damage worldwide. Accurate monitoring for these often microscopic and morphologically indistinguishable species is challenging but critical for mitigating damages. We apply eDNA sampling, which involves the filtering and subsequent DNA extraction of microscopic bits of tissue suspended in water, to ballast and harbor water sampled during a commercial ship's 1400 km voyage through the North American Great Lakes. Using a lab-based gel electrophoresis assay and a rapid, field-ready light transmission spectroscopy (LTS) assay, we test for the presence of two invasive species: quagga (Dreissena bugensis) and zebra (D. polymorpha) mussels. Furthermore, we spiked a set of uninfested ballast and harbor samples with zebra mussel tissue to further test each assay's detection capabilities. In unmanipulated samples, zebra mussel was not detected, while quagga mussel was detected in all samples at a rate of 85% for the gel assay and 100% for the LTS assay. In the spiked experimental samples, both assays detected zebra mussel in 94% of spiked samples and 0% of negative controls. Overall, these results demonstrate that eDNA sampling is effective for monitoring ballast-mediated invasions and that LTS has the potential for rapid, field-based detection.

Development of a new method for the determination of residues of the neonictinoid insecticide imidacloprid in juvenile Chinook (Oncorhynchus tyshawytscha) using ELISA detection

USGS Publications Warehouse

Frew, John A.; Grue, Christian E.

2012-01-01

The neonicotinoid insecticide imidacloprid (IMI) has been proposed as an alternative to carbaryl for controlling indigenous burrowing shrimp on commercial oyster beds in Willapa Bay and Grays Harbor, Washington. A focus of concern over the use of this insecticide in an aquatic environment is the potential for adverse effects from exposure to non-target species residing in the Bay, such as juvenile Chinook (Oncorhynchus tshawytscha) and cutthroat trout (O. clarki). Federal registration and State permiting approval for the use of IMI will require confirmation that the compound does not adversely impact these salmonids following field applications. This will necessitate an environmental monitoring program for evaluating exposure in salmonids following the treatment of beds. Quantification of IMI residues in tissue can be used for determining salmonid exposure to the insecticide. Refinement of an existing protocol using liquid-chromatography mass spectrometry (LC-MS) detection would provide the low limits of quantification, given the relatively small tissue sample sizes, necessary for determining exposure in individual fish. Such an approach would not be viable for the environmental monitoring effort in Willapa Bay and Grays Harbor due to the high costs associated with running multiple analyses, however. A new sample preparation protocol was developed for use with a commercially available enzyme-linked immunosorbent assay (ELISA) for the quantification of IMI, thereby providing a low-cost alternative to LC-MS for environmental monitoring in Willapa Bay and Grays Harbor. Extraction of the analyte from the salmonid brain tissue was achieved by Dounce homogenization in 4.0 mL of 20.0 mM Triton X-100, followed by a 6 h incubation at 50–55 °C. Centrifugal ultrafiltration and reversed phase solid phase extraction were used for sample cleanup. The limit of quantification for an average 77.0 mg whole brain sample was calculated at 18.2 μg kg-1 (ppb) with an average recovery of 79%. This relatively low limit of quantification allows for the analysis of individual fish. Using controlled laboratory studies, a curvelinear relationship was found between the measured IMI residue concentrations in brain tissue and exposure concentrations in seawater. Additonally, a range of IMI brain residue concentrations was associated with an overt effect; illustrating the utility of the IMI tissue residue quantification approach for linking exposure with defined effects.

[Oral mucosa analog allografts in non-consanguineous rats].

PubMed

González, Luis; Padrón, Karla; Salmen, Siham; Jerez, Elsy; Dávila, Lorena; Solórzano, Eduvigis

2017-01-24

Although there are therapeutic options for the treatment of oral mucosa defects, the need for functional, anatomical and aesthetically similar substitutes persists, as well as for solutions to reduce autologous grafts morbidity. To determine clinical and histological compatibility of equivalent oral mucosa allografts generated through tissue engineering in non-consanguineous rats. We used a sample of oral mucosa from Sprague Dawley rats to obtain a fibroblast culture and a keratinocytes and fibroblasts co-culture. In both cases, we used a commercial collagen membrane as "scaffold". After ten weeks of culture, we grafted the resulting membranes into four Wistar rats. The first phase of the study was the development of the oral mucosa equivalents generated by tissue engineering. Then, we implanted them in immunocompetent Wistar rats, and finallywe evaluated the clinical and histological features of the allografts. In vivo evaluation of mucosal substitutes showed a correct integration of artificial oral mucosa in immunocompetent hosts, with an increase in periodontal biotype and the creation of a zone with increased keratinization. Histologically, the tissue was similar to the control oral mucosa sample with no inflammatory reaction nor clinical or histological rejection signs. The equivalent oral mucosa allografts generated by tissue engineering showed clinical and histological compatibility.

Enhanced peroxidase activity and tumour tissue visualization by cobalt-doped magnetoferritin nanoparticles

NASA Astrophysics Data System (ADS)

Zhang, Tongwei; Cao, Changqian; Tang, Xu; Cai, Yao; Yang, Caiyun; Pan, Yongxin

2017-01-01

Magnetoferritin (M-HFn) is a biomimetic magnetic nanoparticle with a human heavy-chain ferritin (HFn) shell, trapping a magnetite (Fe3O4) core that has inherited peroxidase-like activity. In this study, cobalt-doped M-HFn nanoparticles (M-HFn-Co x Fe3-x O4) with different amounts of cobalt were successfully synthesized. Experimental results indicate that the controlled doping of a certain amount of cobalt into the magnetite cores of M-HFn nanoparticles enhances its peroxidase-like catalytic activity and efficacy for visualizing tumour tissues. For example, compared with sample Co0 (without cobalt doping), the peroxidase-like activity of the cobalt-doped nanoparticle sample Co60 (with a cobalt doping molar percentage of ˜34.2%) increases 1.7 times, and has the maximal reaction velocity (V max) values. Moreover, after a one-step incubation with Co60 nanoparticles, and using the peroxidase substrate 3,3‧-diaminobenzidine tetrahydrochloride (DAB) for colour development, the tumour tissues of breast, colorectal, stomach and pancreas tumours showed a deeper brown colour with clear boundaries between the healthy and tumourous cells. Therefore, this suggests that the cobalt-doped magnetoferritin nanoparticles enhance peroxidase activity and tumour tissue visualization.

Sensitivity and specificity of univariate MRI analysis of experimentally degraded cartilage

PubMed Central

Lin, Ping-Chang; Reiter, David A.; Spencer, Richard G.

2010-01-01

MRI is increasingly used to evaluate cartilage in tissue constructs, explants, and animal and patient studies. However, while mean values of MR parameters, including T1, T2, magnetization transfer rate km, apparent diffusion coefficient ADC, and the dGEMRIC-derived fixed charge density, correlate with tissue status, the ability to classify tissue according to these parameters has not been explored. Therefore, the sensitivity and specificity with which each of these parameters was able to distinguish between normal and trypsin- degraded, and between normal and collagenase-degraded, cartilage explants were determined. Initial analysis was performed using a training set to determine simple group means to which parameters obtained from a validation set were compared. T1 and ADC showed the greatest ability to discriminate between normal and degraded cartilage. Further analysis with k-means clustering, which eliminates the need for a priori identification of sample status, generally performed comparably. Use of fuzzy c-means (FCM) clustering to define centroids likewise did not result in improvement in discrimination. Finally, a FCM clustering approach in which validation samples were assigned in a probabilistic fashion to control and degraded groups was implemented, reflecting the range of tissue characteristics seen with cartilage degradation. PMID:19705467

Enhanced peroxidase activity and tumour tissue visualization by cobalt-doped magnetoferritin nanoparticles.

PubMed

Zhang, Tongwei; Cao, Changqian; Tang, Xu; Cai, Yao; Yang, Caiyun; Pan, Yongxin

2017-01-27

Magnetoferritin (M-HFn) is a biomimetic magnetic nanoparticle with a human heavy-chain ferritin (HFn) shell, trapping a magnetite (Fe 3 O 4 ) core that has inherited peroxidase-like activity. In this study, cobalt-doped M-HFn nanoparticles (M-HFn-Co x Fe 3-x O 4 ) with different amounts of cobalt were successfully synthesized. Experimental results indicate that the controlled doping of a certain amount of cobalt into the magnetite cores of M-HFn nanoparticles enhances its peroxidase-like catalytic activity and efficacy for visualizing tumour tissues. For example, compared with sample Co0 (without cobalt doping), the peroxidase-like activity of the cobalt-doped nanoparticle sample Co60 (with a cobalt doping molar percentage of ∼34.2%) increases 1.7 times, and has the maximal reaction velocity (V max ) values. Moreover, after a one-step incubation with Co60 nanoparticles, and using the peroxidase substrate 3,3'-diaminobenzidine tetrahydrochloride (DAB) for colour development, the tumour tissues of breast, colorectal, stomach and pancreas tumours showed a deeper brown colour with clear boundaries between the healthy and tumourous cells. Therefore, this suggests that the cobalt-doped magnetoferritin nanoparticles enhance peroxidase activity and tumour tissue visualization.

Differentiating oral lesions in different carcinogenesis stages with optical coherence tomography

NASA Astrophysics Data System (ADS)

Tsai, Meng-Tsan; Lee, Cheng-Kuang; Lee, Hsiang-Chieh; Chen, Hsin-Ming; Chiang, Chun-Pin; Wang, Yih-Ming; Yang, Chih-Chung

2009-07-01

A swept-source optical coherence tomography (SS-OCT) system is used to clinically scan oral lesions in different oral carcinogenesis stages, including normal oral mucosa control, mild dysplasia (MiD), moderate dysplasia (MoD), early-stage squamous cell carcinoma (ES-SCC), and well-developed SCC (WD-SCC), for diagnosis purpose. On the basis of the analyses of the SS-OCT images, the stages of dysplasia (MiD and MoD), and SCC (ES-SCC and WD-SCC) can be differentiated from normal control by evaluating the depth-dependent standard deviation (SD) values of lateral variations. In the dysplasia stage, the boundary between the epithelium (EP) and lamina propria (LP) layers can still be identified and the EP layer becomes significantly thicker than that of normal control. Also, in a certain range of the EP layer above the EP/LP boundary, the SD value becomes larger than a certain percentage of the maximum level, which is observed around the EP/LP boundary. On the other hand, in the ES-SCC and WD-SCC stages, the EP/LP boundary disappears. Because of the higher density of connective tissue papillae in the ES-SCC stage, the SD values of the slowly varying lateral scan profiles in the ES-SCC samples are significantly larger than those in the WD-SCC sample. Also, ES-SCC can be differentiated from WD-SCC by comparing the exponential decay constants of averaged A-mode scan profiles. Because of the higher tissue absorption in the WD-SCC lesion, the decay constants in the WD-SCC samples are significantly higher than those in the ES-SCC samples.

Fiber laser-microscope system for femtosecond photodisruption of biological samples

PubMed Central

Yavaş, Seydi; Erdogan, Mutlu; Gürel, Kutan; Ilday, F. Ömer; Eldeniz, Y. Burak; Tazebay, Uygar H.

2012-01-01

We report on the development of a ultrafast fiber laser-microscope system for femtosecond photodisruption of biological targets. A mode-locked Yb-fiber laser oscillator generates few-nJ pulses at 32.7 MHz repetition rate, amplified up to ∼125 nJ at 1030 nm. Following dechirping in a grating compressor, ∼240 fs-long pulses are delivered to the sample through a diffraction-limited microscope, which allows real-time imaging and control. The laser can generate arbitrary pulse patterns, formed by two acousto-optic modulators (AOM) controlled by a custom-developed field-programmable gate array (FPGA) controller. This capability opens the route to fine optimization of the ablation processes and management of thermal effects. Sample position, exposure time and imaging are all computerized. The capability of the system to perform femtosecond photodisruption is demonstrated through experiments on tissue and individual cells. PMID:22435105

Investigation of real tissue water equivalent path lengths using an efficient dose extinction method

NASA Astrophysics Data System (ADS)

Zhang, Rongxiao; Baer, Esther; Jee, Kyung-Wook; Sharp, Gregory C.; Flanz, Jay; Lu, Hsiao-Ming

2017-07-01

For proton therapy, an accurate conversion of CT HU to relative stopping power (RSP) is essential. Validation of the conversion based on real tissue samples is more direct than the current practice solely based on tissue substitutes and can potentially address variations over the population. Based on a novel dose extinction method, we measured water equivalent path lengths (WEPL) on animal tissue samples to evaluate the accuracy of CT HU to RSP conversion and potential variations over a population. A broad proton beam delivered a spread out Bragg peak to the samples sandwiched between a water tank and a 2D ion-chamber detector. WEPLs of the samples were determined from the transmission dose profiles measured as a function of the water level in the tank. Tissue substitute inserts and Lucite blocks with known WEPLs were used to validate the accuracy. A large number of real tissue samples were measured. Variations of WEPL over different batches of tissue samples were also investigated. The measured WEPLs were compared with those computed from CT scans with the Stoichiometric calibration method. WEPLs were determined within  ±0.5% percentage deviation (% std/mean) and  ±0.5% error for most of the tissue surrogate inserts and the calibration blocks. For biological tissue samples, percentage deviations were within  ±0.3%. No considerable difference (<1%) in WEPL was observed for the same type of tissue from different sources. The differences between measured WEPLs and those calculated from CT were within 1%, except for some bony tissues. Depending on the sample size, each dose extinction measurement took around 5 min to produce ~1000 WEPL values to be compared with calculations. This dose extinction system measures WEPL efficiently and accurately, which allows the validation of CT HU to RSP conversions based on the WEPL measured for a large number of samples and real tissues.

Micro-organisms isolated from cadaveric samples of allograft musculoskeletal tissue.

PubMed

Varettas, Kerry

2013-12-01

Allograft musculoskeletal tissue is commonly used in orthopaedic surgical procedures. Cadaveric donors of musculoskeletal tissue supply multiple allografts such as tendons, ligaments and bone. The microbiology laboratory of the South Eastern Area Laboratory Services (SEALS, Australia) has cultured cadaveric allograft musculoskeletal tissue samples for bacterial and fungal isolates since 2006. This study will retrospectively review the micro-organisms isolated over a 6-year period, 2006-2011. Swab and tissue samples were received for bioburden testing and were inoculated onto agar and/or broth culture media. Growth was obtained from 25.1 % of cadaveric allograft musculoskeletal tissue samples received. The predominant organisms isolated were coagulase-negative staphylococci and coliforms, with the heaviest bioburden recovered from the hemipelvis. The rate of bacterial and fungal isolates from cadaveric allograft musculoskeletal tissue samples is higher than that from living donors. The type of organism isolated may influence the suitability of the allograft for transplant.

CODIFI (Concordance in Diabetic Foot Ulcer Infection): a cross-sectional study of wound swab versus tissue sampling in infected diabetic foot ulcers in England.

PubMed

Nelson, Andrea; Wright-Hughes, Alexandra; Backhouse, Michael Ross; Lipsky, Benjamin A; Nixon, Jane; Bhogal, Moninder S; Reynolds, Catherine; Brown, Sarah

2018-01-31

To determine the extent of agreement and patterns of disagreement between wound swab and tissue samples in patients with an infected diabetic foot ulcer (DFU). Multicentre, prospective, cross-sectional study. Primary and secondary care foot ulcer/diabetic outpatient clinics and hospital wards across England. Inclusion criteria: consenting patients aged ≥18 years; diabetes mellitus; suspected infected DFU. clinically inappropriate to take either sample. Wound swab obtained using Levine's technique; tissue samples collected using a sterile dermal curette or scalpel. Coprimary: reported presence, and number, of pathogens per sample; prevalence of resistance to antimicrobials among likely pathogens. Secondary: recommended change in antibiotic therapy based on blinded clinical review; adverse events; sampling costs. 400 consenting patients (79% male) from 25 centres.Most prevalent reported pathogens were Staphylococcus aureus (43.8%), Streptococcus (16.7%) and other aerobic Gram-positive cocci (70.6%). At least one potential pathogen was reported from 70.1% of wound swab and 86.1% of tissue samples. Pathogen results differed between sampling methods in 58% of patients, with more pathogens and fewer contaminants reported from tissue specimens.The majority of pathogens were reported significantly more frequently in tissue than wound swab samples (P<0.01), with equal disagreement for S. aureus and Pseudomonas aeruginosa. Blinded clinicians more often recommended a change in antibiotic regimen based on tissue compared with wound swab results (increase of 8.9%, 95% CI 2.65% to 15.3%). Ulcer pain and bleeding occurred more often after tissue collection versus wound swabs (pain: 9.3%, 1.3%; bleeding: 6.8%, 1.5%, respectively). Reports of tissue samples more frequently identified pathogens, and less frequently identified non-pathogens compared with wound swab samples. Blinded clinicians more often recommended changes in antibiotic therapy based on tissue compared with wound swab specimens. Further research is needed to determine the effect of the additional information provided by tissue samples. ISRCTN52608451. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

A comparative proteomic analysis of bile for biomarkers of cholangiocarcinoma.

PubMed

Laohaviroj, Marut; Potriquet, Jeremy; Jia, Xinying; Suttiprapa, Sutas; Chamgramol, Yaovalux; Pairojkul, Chawalit; Sithithaworn, Paiboon; Mulvenna, Jason; Sripa, Banchob

2017-06-01

Cholangiocarcinoma is a primary malignant tumor of the bile duct epithelium. Cholangiocarcinoma is usually detected at an advanced stage when successful treatment is no longer possible. As the tumor originates from the bile duct epithelium, bile is an ideal source of tumor biomarkers for cholangiocarcinoma. In this study, we used a quantitative proteomics approach to identify potential tumor-associated proteins in the bile fluid of six cholangiocarcinoma patients. Three different gross-appearance tumor types were used in the analysis: mass-forming type ( n = 2), periductal infiltrating type ( n = 2), and intraductal growth type ( n = 2). Two bile samples from non-cancerous patients were used as controls. Isobaric labeling, coupled with Tandem mass spectrometry, was used to quantify protein levels in the bile of cholangiocarcinoma and control patients. In all, 63 proteins were significantly increased in cholangiocarcinoma bile compared to normal bile. Alpha-1-antitrypsin was one of the overexpressed proteins that increased in cholangiocarcinoma bile samples. Immunohistochemical analysis revealed that alpha-1-antitrypsin was detected in 177 (50%) of 354 cholangiocarcinoma tissues from our Tissue Bank. Immunoblotting of 54 cholangiocarcinoma bile samples showed that alpha-1-antitrypsin was positive in 38 (70%) samples. Fecal enzyme-linked immunosorbent assay showed that alpha-1-antitrypsin level was able to distinguish cholangiocarcinoma patients from normal individuals. In conclusion, alpha-1-antitrypsin is a potential marker for early diagnosis of cholangiocarcinoma.

Cell block samples from malignant pleural effusion might be valid alternative samples for anaplastic lymphoma kinase detection in patients with advanced non-small-cell lung cancer.

PubMed

Zhou, Jianya; Yao, Hongtian; Zhao, Jing; Zhang, Shumeng; You, Qihan; Sun, Ke; Zou, Yinying; Zhou, Caicun; Zhou, Jianying

2015-06-01

To evaluate the clinical value of cell block samples from malignant pleural effusion (MPE) as alternative samples to tumour tissue for anaplastic lymphoma kinase (ALK) detection in patients with advanced non-small-cell lung cancer (NSCLC). Fifty-two matched samples were eligible for analysis. ALK status was detected by Ventana immunohistochemistry (IHC) (with the D5F3 clone), reverse transcription polymerase chain reaction (RT-PCR) and fluorescence in-situ hybridization (FISH) in MPE cell block samples, and by FISH in tumour tissue block samples. In total, ALK FISH results were obtained for 52 tumour tissue samples and 41 MPE cell block samples. Eight cases (15.4%) were ALK-positive in tumour tissue samples by FISH, and among matched MPE cell block samples, five were ALK-positive by FISH, seven were ALK-positive by RT-PCR, and eight were ALK-positive by Ventana IHC. The ALK status concordance rates between tumour tissue and MPE cell block samples were 78.9% by FISH, 98.1% by RT-PCR, and 100% by Ventana IHC. In MPE cell block samples, the sensitivity and specificity of Ventana IHC (100% and 100%) and RT-PCR (87.5% and 100%) were higher than those of FISH (62.5% and 100%). Malignant pleural effusion cell block samples had a diagnostic performance for ALK detection in advanced NSCLC that was comparable to that of tumour tissue samples. MPE cell block samples might be valid alternative samples for ALK detection when tissue is not available. Ventana IHC could be the most suitable method for ALK detection in MPE cell block samples. © 2014 John Wiley & Sons Ltd.

Proteolytic enzymes from Bromelia antiacantha as tools for controlled tissue hydrolysis in entomology.

PubMed

Macció, Laura; Vallés, Diego; Cantera, Ana Maria

2013-12-01

A crude extract with high proteolytic activity (78.1 EU/mL), prepared from ripe fruit of Bromelia antiacantha was used to hydrolyze and remove soft tissues from the epigyne of Apopyllus iheringi. This enzymatic extract presented four actives isoforms which have a broad substrate specificity action. Enzyme action on samples was optimized after evaluation under different conditions of pH, enzyme-substrate ratio and time (parameters selected based on previous studies) of treatment (pH 4.0, 6.0 and 8.0 at 42°C with different amount of enzyme). Scanning electron microscopy was used to evaluate conditions resulting in complete digestion of epigyne soft tissues. Optimal conditions for soft tissue removal were 15.6 total enzyme units, pH 6.0 for 18 h at 42°C.

Wavelength-dependent backscattering measurements for quantitative real-time monitoring of apoptosis in living cells

NASA Astrophysics Data System (ADS)

Mulvey, Christine S.; Sherwood, Carly A.; Bigio, Irving J.

2009-11-01

Apoptosis-programmed cell death-is a cellular process exhibiting distinct biochemical and morphological changes. An understanding of the early morphological changes that a cell undergoes during apoptosis can provide the opportunity to monitor apoptosis in tissue, yielding diagnostic and prognostic information. There is avid interest regarding the involvement of apoptosis in cancer. The initial response of a tumor to successful cancer treatment is often massive apoptosis. Current apoptosis detection methods require cell culture disruption. Our aim is to develop a nondisruptive optical method to monitor apoptosis in living cells and tissues. This would allow for real-time evaluation of apoptotic progression of the same cell culture over time without alteration. Elastic scattering spectroscopy (ESS) is used to monitor changes in light-scattering properties of cells in vitro due to apoptotic morphology changes. We develop a simple instrument capable of wavelength-resolved ESS measurements from cell cultures in the backward direction. Using Mie theory, we also develop an algorithm that extracts the size distribution of scatterers in the sample. The instrument and algorithm are validated with microsphere suspensions. For cell studies, Chinese hamster ovary (CHO) cells are cultured to confluence on plates and are rendered apoptotic with staurosporine. Backscattering measurements are performed on pairs of treated and control samples at a sequence of times up to 6-h post-treatment. Initial results indicate that ESS is capable of discriminating between treated and control samples as early as 10- to 15-min post-treatment, much earlier than is sensed by standard assays for apoptosis. Extracted size distributions from treated and control samples show a decrease in Rayleigh and 150-nm scatterers, relative to control samples, with a corresponding increase in 200-nm particles. Work continues to correlate these size distributions with underlying morphology. To our knowledge, this is the first report of the use of backscattering spectral measurements to quantitatively monitor apoptosis in viable cell cultures in vitro.

Local Lymphocytes and Nitric Oxide Synthase in the Uterine Cervical Stroma of Patients with Grade III Cervical Intraepithelial Neoplasia

PubMed Central

da Silva, Cléber Sergioda; Michelin, Marcia Antoniazi; Etchebehere, Renata Margarida; Adad, Sheila Jorge; Murta, Eddie Fernando Candido

2010-01-01

OBJECTIVES: Precancerous and cancerous cells can trigger an immune response that may limit tumor development and can be used as a prognostic marker. The aims of the present study were to quantify the presence of B and T lymphocytes, macrophages and cells expressing inducible nitric oxide synthase (iNOS) in the cervical stroma of women with grade III cervical intraepithelial neoplasia (CIN III) or in the intratumoral and peritumoral tissue of women with stage I invasive carcinoma. METHODS: Cervical tissue specimens were obtained from 60 women (20 each from control tissues, CIN III and invasive carcinomas). The average ages in the control, CIN III and invasive groups were 43.9 (± 4.3), 35.5 (± 9.5), and 50 (± 11.2) years, respectively. The specimens were immunohistochemically labeled with antibodies to identify T lymphocytes (CD3), cytotoxic lymphocytes (CD8), B lymphocytes (CD20), macrophages (CD68) and iNOS. We evaluated the markers in the stroma above the squamocolumnar junction (control), at the intraepithelial lesion (CIN cases), and in the nfiltrating tumor. Two independent observers performed the immunohistochemical analysis. RESULTS: T lymphocytes, B lymphocytes, macrophages and iNOS were present more frequently (P<0.05) in the stroma of peritumoral invasive tumors compared to the controls and intratumoral invasive cancer samples. CD3+ and CD20+ lymphocytes were present more frequently in CIN III patients compared to samples from patients with intratumoral invasive cancer (P<0.05). CONCLUSION: High numbers of T and B lymphocytes, macrophages and iNOS-expressing cells in the peritumoral stroma of the invasive tumors were observed. Cell migration appeared to be proportional to the progression of the lesion. PMID:20613932

Topical Application of Aloe vera Accelerated Wound Healing, Modeling, and Remodeling: An Experimental Study.

PubMed

Oryan, Ahmad; Mohammadalipour, Adel; Moshiri, Ali; Tabandeh, Mohammad Reza

2016-01-01

Treatment of large wounds is technically demanding and several attempts have been taken to improve wound healing. Aloe vera has been shown to have some beneficial roles on wound healing but its mechanism on various stages of the healing process is not clear. This study was designed to investigate the effect of topical application of A. vera on cutaneous wound healing in rats. A rectangular 2 × 2-cm cutaneous wound was created in the dorsum back of rats. The animals were randomly divided into 3 groups of control (n = 20), low-dose (n = 20), and high-dose (n = 20) A. vera. The control and treated animals were treated daily with topical application of saline, low-dose (25 mg/mL), and high-dose (50 mg/mL) A. vera gel, up to 10 days, respectively. The wound surface, wound contraction, and epithelialization were monitored. In each group, the animals were euthanized at 10 (n = 5), 20 (n = 5), and 30 (n = 10) days post injury (DPI). At 10, 20, and 30 DPI, the skin samples were used for histopathological and biochemical investigations; and at 30 DPI, the skin samples were also subjected for biomechanical studies. Aloe vera modulated the inflammation, increased wound contraction and epithelialization, decreased scar tissue size, and increased alignment and organization of the regenerated scar tissue. A dose-dependent increase in the tissue level of dry matter, collagen, and glycosaminoglycans' content was seen in the treated lesions, compared to the controls. The treated lesions also demonstrated greater maximum load, ultimate strength, and modulus of elasticity compared to the control ones (P < 0.05). Topical application of A. vera improved the biochemical, morphological, and biomechanical characteristics of the healing cutaneous wounds in rats. This treatment option may be valuable in clinical practice.

A workflow to preserve genome-quality tissue samples from plants in botanical gardens and arboreta.

PubMed

Gostel, Morgan R; Kelloff, Carol; Wallick, Kyle; Funk, Vicki A

2016-09-01

Internationally, gardens hold diverse living collections that can be preserved for genomic research. Workflows have been developed for genomic tissue sampling in other taxa (e.g., vertebrates), but are inadequate for plants. We outline a workflow for tissue sampling intended for two audiences: botanists interested in genomics research and garden staff who plan to voucher living collections. Standard herbarium methods are used to collect vouchers, label information and images are entered into a publicly accessible database, and leaf tissue is preserved in silica and liquid nitrogen. A five-step approach for genomic tissue sampling is presented for sampling from living collections according to current best practices. Collecting genome-quality samples from gardens is an economical and rapid way to make available for scientific research tissue from the diversity of plants on Earth. The Global Genome Initiative will facilitate and lead this endeavor through international partnerships.

Ghrelin O-acyltransferase (GOAT) enzyme is overexpressed in prostate cancer, and its levels are associated with patient's metabolic status: Potential value as a non-invasive biomarker.

PubMed

Hormaechea-Agulla, Daniel; Gómez-Gómez, Enrique; Ibáñez-Costa, Alejandro; Carrasco-Valiente, Julia; Rivero-Cortés, Esther; L-López, Fernando; Pedraza-Arevalo, Sergio; Valero-Rosa, José; Sánchez-Sánchez, Rafael; Ortega-Salas, Rosa; Moreno, María M; Gahete, Manuel D; López-Miranda, José; Requena, María J; Castaño, Justo P; Luque, Raúl M

2016-12-01

Ghrelin-O-acyltransferase (GOAT) is the key enzyme regulating ghrelin activity, and has been proposed as a potential therapeutic target for obesity/diabetes and as a biomarker in some endocrine-related cancers. However, GOAT presence and putative role in prostate-cancer (PCa) is largely unknown. Here, we demonstrate, for the first time, that GOAT is overexpressed (mRNA/protein-level) in prostatic tissues (n = 52) and plasma/urine-samples (n = 85) of PCa-patients, compared with matched controls [healthy prostate tissues (n = 12) and plasma/urine-samples from BMI-matched controls (n = 28), respectively]. Interestingly, GOAT levels in PCa-patients correlated with aggressiveness and metabolic conditions (i.e. diabetes). Actually, GOAT expression was regulated by metabolic inputs (i.e. In1-ghrelin, insulin/IGF-I) in cultured normal prostate cells and PCa-cell lines. Importantly, ROC-curve analysis unveiled a valuable diagnostic potential for GOAT to discriminate PCa at the tissue/plasma/urine-level with high sensitivity/specificity, particularly in non-diabetic individuals. Moreover, we discovered that GOAT is secreted by PCa-cells, and that its levels are higher in urine samples from a stimulated post-massage vs. pre-massage prostate-test. In conclusion, plasmatic GOAT levels exhibit high specificity/sensitivity to predict PCa-presence compared with other PCa-biomarkers, especially in non-diabetic individuals, suggesting that GOAT holds potential as a novel non-invasive PCa-biomarker. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Molecular effects of leptin on peroxisome proliferator activated receptor gamma (PPAR-γ) mRNA expression in rat's adipose and liver tissue.

PubMed

Abbasi, A; Moghadam, A A; Kahrarian, Z; Abbsavaran, R; Yari, K; Alizadeh, E

2017-08-15

Leptin is a 16-kDa peptide hormone secreted by adipose tissue that participates in the regulation of energy homeostasis. The aim of this study was to determine the effect of leptin injection on mRNA expression of peroxisome proliferator-activated receptor gamma (PPAR-γ) and comparison of PPAR-γ mRNA expression in rat's adipose and liver tissue. Twenty adult male rats were divided into the following groups: Group 1asa control (n=10) that did not receive any treatment. Group 2as a treatment (n=10) that received leptin (30 µg ⁄ kg BW) intraperitoneally (ip) for two successive days. Blood samples were taken before and one day after second leptin injection for triglyceride (TG), Free Fatty Acid (FFA), HLD-cholesterol, and LDL-cholesterol measurement. Total RNA was extractedfrom the adipose tissue and liver tissues of rats.  Adipose and liver tissue cells' cDNA was synthesized to characterize the expression of PPAR-γ. Gene expression of PPAR-γ mRNA was tested by RT- PCR technique. Results show leptin decreases expression of PPAR-γ on rat. Low levels of PPAR-γ mRNA were detected in adipose and liver tissues of treatment rats in comparison to control group. In treatment group, the level of PPAR-γ mRNA in liver tissue was very lower than the adipose tissue. The levels of HDL and FFA in treatment rats were increased whereas serum levels TG, VLDL and LDL were not changed. It is concluded that leptin signal with suppressing of PPAR-γ mRNA expression in rat's adipose and liver tissues can result in lipolysis instead of lipogenesis.

[Expression of high mobility group box-1 in the lung tissue and serum of patients with pulmonary tuberculosis].

PubMed

Yang, Xiao-min; Yang, Hua

2013-07-01

To explore the expression of high mobility group box-1 (HMGB1) in the lung tissue and serum of patients with pulmonary tuberculosis and to explore its relationship with tumor necrosis factor (TNF)-α and interleukin(IL)-1β. Sixty samples of lung tissues were obtained from patients with pulmonary tuberculosis who had underwent pneumonectomy in Department of Chest Surgery, First Affiliated Hospital of Zunyi Medical College from June 2010 to December 2011. At the same period, 40 normal lung samples were also obtained from patients with pulmonary contusion and lung cancer by surgical resections as the control group. The mRNA expressions of HMGB1 was detected by reverse transcription-polymerase chain reaction (RT-PCR), and the protein level of HMGB1 was measured by immunohistochemical staining of tissue microarrays in lung tissue. Blood samples were taken from 89 patients with active pulmonary tuberculosis (pulmonary tuberculosis group), including hematogenous disseminated pulmonary tuberculosis (type II) in 35 cases and secondary pulmonary tuberculosis (type III) in 54 cases, and 50 healthy volunteers (control group). Furthermore, the 54 patients with secondary pulmonary tuberculosis were divided into different subgroups according to cavity formation and the lung fields involved: patients without lung cavity (35 cases) vs those with lung cavity (19 cases), patients with involvement of <2 lung fields (31 cases) vs ≥ 2 lung fields (23 cases). Serum concentration of HMGB1, TNF-α and IL-1β were detected by ELISA. Two sample t-test was used to compare date among groups, liner correlation analysis was established for correlation analysis. The average optical density of HMGB1 in pulmonary tuberculosis (69 ± 29) was significantly higher than that in normal lung tissue (22 ± 12) (t = 2.389, P < 0.05). The mRNA relative transcript levels of HMGB1 in pulmonary tuberculosis (786 ± 86) was significantly higher than that in normal lung tissue (202 ± 60) (t = 3.872, P < 0.01). The serum concentration of HMGB1, TNF-α and IL-1β in the pulmonary tuberculosis group were (5.0 ± 3.2) µg/L, (118 ± 77) ng/L and (33 ± 20) ng/L, respectively, which were significantly higher than those in the control group [(1.7 ± 1.0) µg/L, (40 ± 11) ng/L and (18 ± 12) ng/L, respectively], the respective t values being -0.928, 4.268 and 11.064, all P < 0.01. In the subgroup of patients with hematogenous disseminated pulmonary tuberculosis, the serum concentration of HMGB1 and TNF-α[ (6.4 ± 3.3) µg/L, (147 ± 89) ng/L] were significantly higher than those in patients with secondary pulmonary tuberculosis [(4.1 ± 2.7) µg/L, (85 ± 37) ng/L] (t = 3.643 and t = 3.111, both P < 0.01). HMGB1 were correlated positively with TNF-α and IL-1β (r = 0.722 and r = 0.620, P < 0.01, respectively, n = 89) in the pulmonary tuberculosis group. Overexpression of HMGB1 in the lung tissue and serum of patients with pulmonary tuberculosis may play an important role in the inflammatory response of pulmonary tuberculosis. The measurement of serum HMGB1 is useful to evaluate the severity of disease.

Mechanisms for the control of local tissue blood flow during thermal interventions: influence of temperature‐dependent ATP release from human blood and endothelial cells

PubMed Central

Chiesa, Scott T.; Trangmar, Steven J.; Ali, Leena; Lotlikar, Makrand D.; González‐Alonso, José

2017-01-01

New Findings What is the central question of this study? Skin and muscle blood flow increases with heating and decreases with cooling, but the temperature‐sensitive mechanisms underlying these responses are not fully elucidated. What is the main finding and its importance? We found that local tissue hyperaemia was related to elevations in ATP release from erythrocytes. Increasing intravascular ATP augmented skin and tissue perfusion to levels equal or above thermal hyperaemia. ATP release from isolated erythrocytes was altered by heating and cooling. Our findings suggest that erythrocytes are involved in thermal regulation of blood flow via modulation of ATP release. Local tissue perfusion changes with alterations in temperature during heating and cooling, but the thermosensitivity of the vascular ATP signalling mechanisms for control of blood flow during thermal interventions remains unknown. Here, we tested the hypotheses that the release of the vasodilator mediator ATP from human erythrocytes, but not from endothelial cells or other blood constituents, is sensitive to both increases and reductions in temperature and that increasing intravascular ATP availability with ATP infusion would potentiate thermal hyperaemia in limb tissues. We first measured blood temperature, brachial artery blood flow and plasma [ATP] during passive arm heating and cooling in healthy men and found that they increased by 3.0 ± 1.2°C, 105 ± 25 ml min−1 °C−1 and twofold, respectively, (all P < 0.05) with heating, but decreased or remained unchanged with cooling. In additional men, infusion of ATP into the brachial artery increased skin and deep tissue perfusion to levels equal or above thermal hyperaemia. In isolated erythrocyte samples exposed to different temperatures, ATP release increased 1.9‐fold from 33 to 39°C (P < 0.05) and declined by ∼50% at 20°C (P < 0.05), but no changes were observed in cultured human endothelial cells, plasma or serum samples. In conclusion, increases in plasma [ATP] and skin and deep tissue perfusion with limb heating are associated with elevations in ATP release from erythrocytes, but not from endothelial cells or other blood constituents. Erythrocyte ATP release is also sensitive to temperature reductions, suggesting that erythrocytes may function as thermal sensors and ATP signalling generators for control of tissue perfusion during thermal interventions. PMID:27859767

The decrease in silicon concentration of the connective tissues with age in rats is a marker of connective tissue turnover.

PubMed

Jugdaohsingh, Ravin; Watson, Abigail I E; Pedro, Liliana D; Powell, Jonathan J

2015-06-01

Silicon may be important for bone and connective tissue health. Higher concentrations of silicon are suggested to be associated with bone and the connective tissues, compared with the non-connective soft tissues. Moreover, in connective tissues it has been suggested that silicon levels may decrease with age based upon analyses of human aorta. These claims, however, have not been tested under controlled conditions. Here connective and non-connective tissues were collected and analysed for silicon levels from female Sprague-Dawley rats of different ages (namely, 3, 5, 8, 12, 26 and 43 weeks; n=8-10 per age group), all maintained on the same feed source and drinking water, and kept in the same environment from weaning to adulthood. Tissues (696 samples) were digested in nitric acid and analysed by inductively coupled plasma optical emission spectrometry for total silicon content. Fasting serum samples were also collected, diluted and analysed for silicon. Higher concentrations of silicon (up to 50-fold) were found associated with bone and the connective tissues compared with the non-connective tissues. Although total silicon content increased with age in all tissues, the highest connective tissue silicon concentrations (up to 9.98 μg/g wet weight) were found in young weanling rats, decreasing thereafter with age (by 2-6 fold). Fasting serum silicon concentrations reflected the pattern of connective tissue silicon concentrations and, both measures, when compared to collagen data from a prior experiment in Sprague-Dawley rats, mirrored type I collagen turnover with age. Our findings confirm the link between silicon and connective tissues and would imply that young growing rats have proportionally higher requirements for dietary silicon than mature adults, for bone and connective tissue development, although this was not formally investigated here. However, estimation of total body silicon content suggested that actual Si requirements may be substantially lower than previously estimated which could explain why absolute silicon deficiency is difficult to achieve but, when it is achieved in young growing animals, it results in stunted growth and abnormal development of bone and other connective tissues. Copyright © 2015. Published by Elsevier Inc.

The decrease in silicon concentration of the connective tissues with age in rats is a marker of connective tissue turnover☆

PubMed Central

Jugdaohsingh, Ravin; Watson, Abigail I.E.; Pedro, Liliana D.; Powell, Jonathan J.

2015-01-01

Silicon may be important for bone and connective tissue health. Higher concentrations of silicon are suggested to be associated with bone and the connective tissues, compared with the non-connective soft tissues. Moreover, in connective tissues it has been suggested that silicon levels may decrease with age based upon analyses of human aorta. These claims, however, have not been tested under controlled conditions. Here connective and non-connective tissues were collected and analysed for silicon levels from female Sprague–Dawley rats of different ages (namely, 3, 5, 8, 12, 26 and 43 weeks; n = 8–10 per age group), all maintained on the same feed source and drinking water, and kept in the same environment from weaning to adulthood. Tissues (696 samples) were digested in nitric acid and analysed by inductively coupled plasma optical emission spectrometry for total silicon content. Fasting serum samples were also collected, diluted and analysed for silicon. Higher concentrations of silicon (up to 50-fold) were found associated with bone and the connective tissues compared with the non-connective tissues. Although total silicon content increased with age in all tissues, the highest connective tissue silicon concentrations (up to 9.98 μg/g wet weight) were found in young weanling rats, decreasing thereafter with age (by 2–6 fold). Fasting serum silicon concentrations reflected the pattern of connective tissue silicon concentrations and, both measures, when compared to collagen data from a prior experiment in Sprague–Dawley rats, mirrored type I collagen turnover with age. Our findings confirm the link between silicon and connective tissues and would imply that young growing rats have proportionally higher requirements for dietary silicon than mature adults, for bone and connective tissue development, although this was not formally investigated here. However, estimation of total body silicon content suggested that actual Si requirements may be substantially lower than previously estimated which could explain why absolute silicon deficiency is difficult to achieve but, when it is achieved in young growing animals, it results in stunted growth and abnormal development of bone and other connective tissues. PMID:25687224

A Well-Controlled Nucleus Pulposus Tissue Culture System with Injection Port for Evaluating Regenerative Therapies.

PubMed

Arkesteijn, Irene T M; Mouser, Vivian H M; Mwale, Fackson; van Dijk, Bart G M; Ito, Keita

2016-05-01

In vitro evaluation of nucleus pulposus (NP) tissue regeneration would be useful, but current systems for NP culture are not ideal for injections. The aim of this study was to develop a long-term culture system for NP tissue that allows injections of regenerative agents. Bovine caudal NPs were harvested and placed in the newly designed culture system. After equilibration of the tissue to 0.3 MPa the volume was fixed and the tissue was cultured for 28 days. The cell viability and extracellular matrix composition remained unchanged during the culture period and gene expression profiles were similar to those obtained in earlier studies. Furthermore, to test the responsiveness of bovine caudal NPs in the system, samples were cultured for 4 days and injected twice (day 1 and 3) with (1) PBS, (2) Link-N, for regeneration, and (3) TNF-α, for degeneration. It was shown that TNF-α increased COX2 gene expression, whereas no effect of Link-N was detected. In conclusion, the newly designed system allows long-term culture of NP tissue, wherein tissue reactions to injected stimulants can be observed.

The effects of corrosive substances on human bone, teeth, hair, nails, and soft tissue.

PubMed

Hartnett, Kristen M; Fulginiti, Laura C; Di Modica, Frank

2011-07-01

This research investigates the effects of household chemicals on human tissues. Five different human tissues (bone, tooth, hair, fingernails, and skin/muscle/fat) were immersed into six different corrosive agents. These agents consisted of hydrochloric acid, sulfuric acid, lye, bleach, organic septic cleaner, and Coca-Cola(®) soda. Tap water was used as a control. Tissue samples were cut to consistent sizes and submerged in the corrosive liquids. Over time, the appearance, consistency, and weight were documented. Hydrochloric acid was the most destructive agent in this study, consuming most tissues within 24 h. Sulfuric acid was the second most destructive agent in this study. Bleach, lye, and cola had no structural effects on the hard tissues of the body, but did alter the appearance or integrity of the hair, nails, or flesh in some way. The organic septic cleaner and tap water had no effect on any of the human tissue tested during the timeframe of the study. 2011 American Academy of Forensic Sciences. Published 2011. This article is a U.S. Government work and is in the public domain in the U.S.A.

Study of melanin bleaching after immunohistochemistry of melanin-containing tissues.

PubMed

Shen, Hongwu; Wu, Wenqiao

2015-04-01

Melanin may interfere with immunohistochemical staining. The goal of this study was to investigate the effects of trichloroisocyanuric acid (TCCA) bleaching, potassium permanganate bleaching, and potassium dichromate bleaching on melanin, tissue antigen, and 3,3'-diaminobenzidine (DAB) using melanin-containing and melanin-free tissue samples. Our results demonstrated that all 3 bleaching methods efficiently bleached melanin and partially destroyed tissue antigen. In addition, potassium permanganate bleaching and potassium dichromate bleaching clearly destroyed DAB, whereas TCCA bleaching had no significant effect on DAB. Therefore, neither potassium permanganate nor potassium dichromate is an ideal solution, whereas TCCA might be an ideal solution for melanin bleaching after the immunohistochemical staining of melanin-containing tissues. After immunostaining followed by TCCA bleaching, the melanin could be completely removed in all 120 malignant melanoma tissue sections. Compared with the control, the DAB intensity was clear, and the tissue structure and cellular nuclei were well maintained. It is worth noting that TCCA should be freshly prepared before each experiment, and used within 2 hours of its preparation. In addition, sections should not be incubated with TCCA for over 30 minutes.

Dietary (n-6 : n-3) Fatty Acids Alter Plasma and Tissue Fatty Acid Composition in Pregnant Sprague Dawley Rats

PubMed Central

Kassem, Amira Abdulbari; Abu Bakar, Md Zuki; Yong Meng, Goh; Mustapha, Noordin Mohamed

2012-01-01

The objective of this paper is to study the effects of varying dietary levels of n-6 : n-3 fatty acid ratio on plasma and tissue fatty acid composition in rat. The treatment groups included control rats fed chow diet only, rats fed 50% soybean oil (SBO): 50% cod liver oil (CLO) (1 : 1), 84% SBO: 16% CLO (6 : 1), 96% SBO: 4% CLO (30 : 1). Blood samples were taken at day 15 of pregnancy, and the plasma and tissue were analyzed for fatty acid profile. The n-3 PUFA in plasma of Diet 1 : 1 group was significantly higher than the other diet groups, while the total n-6 PUFA in plasma was significantly higher in Diet 30 : 1 group as compared to the control and Diet 1 : 1 groups. The Diet 1 : 1 group showed significantly greater percentages of total n-3 PUFA and docosahexaenoic acid in adipose and liver tissue, and this clearly reflected the contribution of n-3 fatty acids from CLO. The total n-6 PUFA, linoleic acid, and arachidonic acid were significantly difference in Diet 30 : 1 as compared to Diet 1 : 1 and control group. These results demonstrated that the dietary ratio of n-6 : n-3 fatty acid ratio significantly affected plasma and tissue fatty acids profile in pregnant rat. PMID:22489205

Parallel telomere shortening in multiple body tissues owing to malaria infection.

PubMed

Asghar, Muhammad; Palinauskas, Vaidas; Zaghdoudi-Allan, Nadège; Valkiūnas, Gediminas; Mukhin, Andrey; Platonova, Elena; Färnert, Anna; Bensch, Staffan; Hasselquist, Dennis

2016-08-17

Several studies have shown associations between shorter telomere length in blood and weakened immune function, susceptibility to infections, and increased risk of morbidity and mortality. Recently, we have shown that malaria accelerates telomere attrition in blood cells and shortens lifespan in birds. However, the impact of infections on telomere attrition in different body tissues within an individual is unknown. Here, we tested whether malarial infection leads to parallel telomere shortening in blood and tissue samples from different organs. We experimentally infected siskins (Spinus spinus) with the avian malaria parasite Plasmodium ashfordi, and used real-time quantitative polymerase chain reaction (PCR) to measure telomere length in control and experimentally infected siskins. We found that experimentally infected birds showed faster telomere attrition in blood over the course of infection compared with control individuals (repeatedly measured over 105 days post-infection (DPI)). Shorter telomeres were also found in the tissue of all six major organs investigated (liver, lungs, spleen, heart, kidney, and brain) in infected birds compared with controls at 105 DPI. To the best of our knowledge, this is the first study showing that an infectious disease results in synchronous telomere shortening in the blood and tissue cells of internal organs within individuals, implying that the infection induces systemic stress. Our results have far-reaching implications for understanding how the short-term effects of an infection can translate into long-term costs, such as organ dysfunction, degenerative diseases, and ageing. © 2016 The Author(s).

Primary cilia are increased in number and demonstrate structural abnormalities in human cancer.

PubMed

Yasar, Binnaz; Linton, Kim; Slater, Christian; Byers, Richard

2017-07-01

Primary cilia play an important role in the regulation of cell signalling pathways and are thought to have a role in cancer but have seldom been studied in human cancer samples. Primary cilia were visualised by dual immunofluorescence for anti-CROCC (ciliary rootlet coiled-coil) and anti-tubulin in a range of human cancers (including carcinomas of stomach, pancreas, prostate, lung and colon, lobular and ductal breast cancers and follicular lymphoma) and in matched normal tissue (stomach, pancreas, lung, large and small intestines, breast and reactive lymph nodes) samples using a tissue microarray; their frequency, association with proliferation, was measured by Ki-67 staining and their structure was analysed. Compared with normal tissues, primary cilia frequency was significantly elevated in adenocarcinoma of the lung (2.75% vs 1.85%, p=0.016), adenocarcinoma of the colon (3.80% vs 2.43%, respectively, p=0.017), follicular lymphoma (1.18% vs 0.83%, p=0.003) and pancreatic adenocarcinoma (7.00% vs 5.26%, p=0.002); there was no statistically significant difference compared with normal control tissue for gastric and prostatic adenocarcinomas or for lobular and ductal breast cancers. Additionally, structural abnormalities of primary cilia were identified in cancer tissues, including elongation of the axoneme, multiple basal bodies and branching of the axoneme. Ki-67 scores ranged from 0.7% to 78.4% and showed no statistically significant correlation with primary cilia frequency across all tissues (p=0.1501). The results show upregulation of primary cilia and the presence of structural defects in a wide range of human cancer tissue samples demonstrating association of dysregulation of primary cilia with human cancer. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

Differential Expression of Glypican-3 and Insulin–Like Growth Factor-II mRNAs and Alpha-Fetoprotein and Ki-67 Markers in HCV Related Hepatocellular Carcinomas In Egyptian Patients

PubMed

Saber, Mohamed A; MM AbdelHafiz, Samah; Khorshed, Fatma E; Aboushousha, Tarek S; Hamdy, Hussam EM; Seleem, Mohamed I; Soliman, Amira H

2017-01-01

Background: Increasing evidence indicates that in hepatocellular carcinomas (HCCs) abnormal gene expression, for example of glypican-3 (GPC-3) and insulin-like growth factor-II (IGF-II), are associated with the occurrence and progression of HCC. The objective of this study was to evaluate the differential expression of GPC-3 and IGF-II mRNAs in HCC tissues with a background of chronic hepatitis C virus (HCV) genotype 4 cirrhosis, in relation to Ki-67 and alpha-feto protein (AFP) tissue markers. Methods: One hundred and five patients with HCCs who had undergone hepatectomy, were included, after obtaining informed consent. Total RNA was extracted from malignant and corresponding peri-malignant liver tissues, and GPC-3 and IGF-II mRNAs in addition to beta-actin mRNA as an internal control, were evaluated in all samples by reverse transcriptase-polymerase chain reactions (RT-PCR). Routine histopathological diagnosis as well as immunohistochemical (IHC) staining using monoclonal antibodies for Ki-67 and AFP were also performed. Result: Expression of GPC-3 mRNA was positive in all HCC malignant tissue, with overexpression in 86/105 (81.9%); in respect to the grade of the tumor (1-3 grades), while in peri-malignant tissue it was over expressed only in 20/105 (19%). The IGF-II mRNA was over expressed in only 10/105 (9.5%) malignant and peri-malignant samples. AFP was expressed in 33.3% of malignant samples but absent in peri-malignant tissues. Ki-67 expression was significantly increased in malignant compared to peri-malignant tissue. Conclusion: GPC-3 and IGF II mRNAs may be good molecular markers for HCC, especially with a background of cirrhosis due to chronic HCV infection. Significant correlations were noted with the pattern of AFP and Ki-67 expression. Creative Commons Attribution License

Nigella sativa relieves the deleterious effects of ischemia reperfusion injury on liver

PubMed Central

Yildiz, Fahrettin; Coban, Sacit; Terzi, Alpaslan; Ates, Mustafa; Aksoy, Nurten; Cakir, Hale; Ocak, Ali Riza; Bitiren, Muharrem

2008-01-01

AIM: To determine whether Nigella sativa prevents hepatic ischemia-reperfusion injury to the liver. METHODS: Thirty rats were divided into three groups as sham (Group 1), control (Group 2), and Nigella sativa (NS) treatment group (Group 3). All rats underwent hepatic ischemia for 45 min followed by 60 min period of reperfusion. Rats were intraperitoneally infused with only 0.9% saline solution in group 2. Rats in group 3 received NS (0.2 mL/kg) intraperitoneally, before ischemia and before reperfusion. Blood samples and liver tissues were harvested from the rats, and then the rats were sacrificed. Serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and lactate dehydrogenase (LDH) levels were determined. Total antioxidant capacity (TAC), catalase (CAT), total oxidative status (TOS), oxidative stress index (OSI) and myeloperoxidase (MPO) in hepatic tissue were measured. Also liver tissue histopathology was evaluated by light microscopy. RESULTS: The levels of liver enzymes in group 3 were significantly lower than those in the group 2. TAC in liver tissue was significantly higher in group 3 than in group 2. TOS, OSI and MPO in hepatic tissue were significantly lower in group 3 than the group 2. Histological tissue damage was milder in the NS treatment group than that in the control group. CONCLUSION: Our results suggest that Nigella sativa treatment protects the rat liver against to hepatic ischemia-reperfusion injury. PMID:18777598

The sequential appearance of sperm abnormalities after scrotal insulation or dexamethasone treatment in bulls.

PubMed Central

Barth, A D; Bowman, P A

1994-01-01

Scrotal insulation and dexamethasone treatment were used as a model to compare the effect of testicular heating and stress on spermatogenesis. Insulation was applied to the scrotum of eight bulls (insulated) for a period of four days, eight bulls were treated daily for seven days with 20 mg dexamethasone injected intramuscularly, and four bulls were untreated controls. Semen from four bulls in each group was collected and evaluated over a six-week period after treatment. Blood samples for testosterone analysis were taken hourly for eight hours at the beginning and the end of the six-week period from the control bulls and before and after treatment from the four insulated and four dexamethasone-treated bulls that were not used for semen collection. At the end of the last blood sampling period, the four bulls in each group were castrated for the collection of testicular tissue for the determination of testosterone concentrations. Basal, peak episodic, and mean serum testosterone concentrations among control bulls, pre and postinsulated bulls, and pretreatment samples of dexamethasone-treated bulls were not different (p > 0.05); however, bulls that had received dexamethasone treatments had significantly lower basal, peak episodic, and mean testosterone concentrations (p < 0.05). Tissue concentrations of testosterone in control, insulated, and dexamethasone-treated bulls were not significantly different but tended to be lower in dexamethasone-treated bulls (p > 0.13). The spermiograms of the control bulls varied insignificantly over the six-week sampling period; however, there was a marked increase in sperm defects in insulated and dexamethasone-treated bulls. The types of sperm defects and the temporal relationships of rises and declines of sperm defects were quite similar for both treatments. All bulls recovered to approximately pretreatment levels of sperm defects by six weeks after the initiation of treatment. Results indicate that two of the most common types of insults to spermatogenesis in bulls, heat and stress, result in similar spermiograms. PMID:8069831

Update on Controls for Isolation and Quantification Methodology of Extracellular Vesicles Derived from Adipose Tissue Mesenchymal Stem Cells

PubMed Central

Franquesa, Marcella; Hoogduijn, Martin J.; Ripoll, Elia; Luk, Franka; Salih, Mahdi; Betjes, Michiel G. H.; Torras, Juan; Baan, Carla C.; Grinyó, Josep M.; Merino, Ana Maria

2014-01-01

The research field on extracellular vesicles (EV) has rapidly expanded in recent years due to the therapeutic potential of EV. Adipose tissue human mesenchymal stem cells (ASC) may be a suitable source for therapeutic EV. A major limitation in the field is the lack of standardization of the challenging techniques to isolate and characterize EV. The aim of our study was to incorporate new controls for the detection and quantification of EV derived from ASC and to analyze the applicability and limitations of the available techniques. ASC were cultured in medium supplemented with 5% of vesicles-free fetal bovine serum. The EV were isolated from conditioned medium by differential centrifugation with size filtration (0.2 μm). As a control, non-conditioned culture medium was used (control medium). To detect EV, electron microscopy, conventional flow cytometry, and western blot were used. The quantification of the EV was by total protein quantification, ExoELISA immunoassay, and Nanosight. Cytokines and growth factors in the EV samples were measured by multiplex bead array kit. The EV were detected by electron microscope. Total protein measurement was not useful to quantify EV as the control medium showed similar protein contents as the EV samples. The ExoELISA kits had technical troubles and it was not possible to quantify the concentration of exosomes in the samples. The use of Nanosight enabled quantification and size determination of the EV. It is, however, not possible to distinguish protein aggregates from EV with this method. The technologies for quantification and characterization of the EV need to be improved. In addition, we detected protein contaminants in the EV samples, which make it difficult to determine the real effect of EV in experimental models. It will be crucial in the future to optimize design novel methods for purification and characterization of EV. PMID:25374572

SNP rs16906252C>T is an expression and methylation quantitative trait locus associated with an increased risk of developing MGMT-methylated colorectal cancer

PubMed Central

Kuroiwa-Trzmielina, Joice; Wang, Fan; Rapkins, Robert W.; Ward, Robyn L.; Buchanan, Daniel D.; Win, Aung Ko; Clendenning, Mark; Rosty, Christophe; Southey, Melissa C.; Winship, Ingrid M.; Hopper, John L.; Jenkins, Mark A.; Olivier, Jake; Hawkins, Nicholas J.; Hitchins, Megan P.

2016-01-01

Purpose Methylation of the MGMT promoter is the major cause of O6-methylguanine methyltransferase deficiency in cancer and has been associated with the T variant of the promoter-enhancer SNP rs16906252C>T. We sought evidence for an association between the rs16906252C>T genotype and increased risk of developing a subtype of colorectal cancer (CRC) featuring MGMT methylation, mediated by genotype-dependent epigenetic silencing within normal tissues. Experimental design By applying a molecular pathological epidemiology case-control study design, associations between rs16906252C>T and risk for CRC overall, and CRC stratified by MGMT methylation status, were estimated using multinomial logistic regression in two independent retrospective series of CRC cases and controls. The test sample comprised 1054 CRC cases and 451 controls from Sydney, Australia. The validation sample comprised 612 CRC cases and 245 controls from the Australasian Colon Cancer Family Registry (ACCFR). To determine if rs16906252C>T was linked to a constitutively altered epigenetic state, quantitative allelic expression and methylation analyses were performed in normal tissues. Results An association between rs16906252C>T and increased risk of developing MGMT-methylated CRC in the Sydney sample was observed (OR 3.3; 95%CI=2.0–5.3; P<0.0001), which was replicated in the ACCFR sample (OR 4.0; 95%CI=2.4–6.8; P<0.0001). The T allele demonstrated ~2.5-fold reduced transcription in normal colorectal mucosa from cases and controls, and was selectively methylated in a minority of normal cells, indicating rs16906252C>T represents an expression and methylation quantitative trait locus. Conclusions We provide evidence that rs16906252C>T is associated with elevated risk for MGMT-methylated CRC, likely mediated by constitutive epigenetic repression of the T allele. PMID:27267851

Technologies for Protein Analysis and Tissue Engineering, with Applications in Cancer

NASA Astrophysics Data System (ADS)

Vermesh, Udi Benjamin

The first part of this thesis describes electrolyte transport through an array of 20 nm wide, 20 mum long SiO2 nanofluidic transistors. At sufficiently low ionic strength, the Debye screening length exceeds the channel width, and ion transport is limited by the negatively charged channel surfaces. At source-drain biases > 5 V, the current exhibits a sharp, nonlinear increase, with a 20 - 50-fold conductance enhancement. This behavior is attributed to a breakdown of the zero-slip condition. Implications for peptide sequencing as well as energy conversion devices are discussed. The next part describes a technology for the detection of the highly aggressive brain cancer glioblastoma multiforme (GBM). In this study, we used an antibody-based microarray to compare plasma samples from glioblastoma patients and healthy controls with respect to the plasma levels of 35 different proteins known to be generally associated with tumor growth, survival, invasion, migration, and immune regulation. Average-linkage hierarchical clustering of the patient data stratified the two groups effectively, permitting accurate assignment of test samples into either GBM or healthy control groups with a sensitivity and specificity as high as 90 % and 94 %, respectively. Using the same 35-protein panel, we then analyzed plasma samples from GBM patients who were treated with the chemotherapeutic drug Avastin (Bevacizumab) and were able to effectively stratify patients based on treatment-responsiveness. Finally, single-cell resolution patterning of tissue engineered structures is demonstrated. The proper functioning of engineered constructs for tissue and organ transplantation requires positioning different cell types in anatomically precise arrangements that mimic their configurations in native tissues. Toward this end, we have developed a technique that involves two microfluidic-patterning steps run perpendicularly to each other using "anchor" and "bridge" DNA oligomers to create dense arrays of DNA grids which can then be converted into cell arrays. As a proof-of-concept, both a neuron-astrocyte construct and a pancreatic islet construct containing 2 distinct islet cell types were patterned separately as a dense array of cell grids. Once fixed in a hydrogel matrix, layers of patterned cells were then stacked to form 3-D tissue engineered constructs.

The effect of oyster mushroom β-1.3/1.6-D-glucan and oxytetracycline antibiotic on biometrical, haematological, biochemical, and immunological indices, and histopathological changes in common carp (Cyprinus carpio L.).

PubMed

Dobšíková, Radka; Blahová, Jana; Mikulíková, Ivana; Modrá, Helena; Prášková, Eva; Svobodová, Zdeňka; Skorič, Mišo; Jarkovský, Jiří; Siwicki, Andrzej-Krzysztof

2013-12-01

The aim of the study was to evaluate the effect of micronized β-1.3/1.6-D-glucan (BG) derived from the oyster mushroom Pleurotus ostreatus Hiratake and tetracycline antibiotic oxytetracycline (OTC) on biometrical, haematological, biochemical, and immunological indices, and histopathological changes in tissues of one- to two-year-old common carp (Cyprinus carpio L.). The fish tested were divided into five experimental groups and one control. Carp in the control group were fed commercial carp feed pellets. Fish in the five experimental groups were fed the same pellets supplemented with either OTC, a combination of OTC and BG, or BG as follows: 75 mg oxytetracycline kg(-1) bw (OTC group), 75 mg oxytetracycline kg(-1) bw and 0.5% β-glucan (OTC + 0.5% BG group), 75 mg oxytetracycline kg(-1) bw and 2.0% β-glucan (OTC + 2.0% BG group), 0.5% β-glucan (0.5% BG group), and 2.0% β-glucan (2.0% BG group). OTC- and BG-supplemented diets and the control diet were administered to experimental and control carp for 50 days (i.e. samplings 1-3, the exposure period); for the following 14 days, fish were fed only control feed pellets with no OTC or BG supplementation (i.e. sampling 4, the recovery period). Blood and tissue samples were collected both during, and at the end of the study. No significant changes in biometrical indices (i.e. total length, standard length, total weight, hepatosomatic and spleen somatic index, and Fulton's condition factor) were found in experimental carp compared to control in any sampling. In haematological indices, significant changes were found only in sampling 2, in which shifts in PCV (P < 0.01), Hb (P < 0.01), and WBC (P < 0.01), and in the counts of lymphocytes (P < 0.01), monocytes (P < 0.01), and neutrophil granulocytes-segments (P < 0.05) were revealed. As for biochemical profiling, plasma concentrations of glucose, albumins, cholesterol, natrium, and chlorides (all P < 0.01), and total proteins, lactate, phosphorus, and potassium (all P < 0.05) as well as the catalytic activity of ALP (P < 0.05) were altered in common carp. A significant change in induced (opsonizedzymosan particles, OZP) chemiluminescence (P < 0.05) in sampling 3 and no shifts in serum immunoglobulins concentration were found in the immunological analysis. Histopathological examination of skin, gills, liver, spleen, and cranial and caudal kidneys revealed no obvious specific changes in any tissue analysed. The use of β-glucans in clinically healthy aquaculture remains an issue. Nevertheless, their use in breeding endangered by stress stimuli, infectious disease, or adverse environmental factors is defensible. Copyright © 2013 Elsevier Ltd. All rights reserved.

Protective effect of Spirulina platensis against cell damage and apoptosis in hepatic tissue caused by high fat diet.

PubMed

Yigit, F; Gurel-Gurevin, E; Isbilen-Basok, B; Esener, O B B; Bilal, T; Keser, O; Altiner, A; Yilmazer, N; Ikitimur-Armutak, E I

2016-01-01

Spirulina platensis is a microalga that may be a source of antioxidants that can reduce body fat deposition. Consumption of a high fat diet produces elevated blood lipid levels, inflammation and apoptosis. We investigated the possible effects of S. platensis on the blood lipid profile, and liver inflammation and apoptosis in rats fed a high fat diet. Sixty-four young male rats were divided into eight equal groups. The control group was fed a basic diet. The experimental groups were fed a diet for 60 days that was prepared by mixing variable amounts of 43% vegetable oil and 10% cholesterol with or without 3% S. platensis mixed with the basal diet. Blood and liver tissue samples were collected from each animal. Serum samples were used to analyze lipid parameters, total antioxidant status and total oxidant status. iNOS and eNOS were determined by immunohistochemistry. TUNEL staining was used to detect apoptosis to investigate a possible connection between inflammation and apoptosis in the liver tissue. The relations between fat deposition and liver degeneration were assessed by Sirius red staining and alpha-smooth muscle actin immunostaining. S. platensis reduced serum HDL-C, LDL-C and triglyceride, increased HDL-C levels in rats fed a high fat diet to near control levels, and reduced iNOS levels and increased eNOS levels in the liver tissue compared to vegetable oil and cholesterol treated groups. The apoptotic index was reduced in the groups that were fed a high fat or a basic diet when supplemented with S. platensis.

Advances in Raman spectroscopy for the diagnosis of Alzheimer's disease

NASA Astrophysics Data System (ADS)

Sudworth, Caroline D.; Archer, John K. J.; Black, Richard A.; Mann, David

2006-02-01

Within the next 50 years Alzheimer's disease is expected to affect 100 million people worldwide. The progressive decline in the mental health of the patient is caused by severe brain atrophy generated by the breakdown and aggregation of proteins, resulting in β-amyloid plaques and neurofibrillary tangles. The greatest challenge to Alzheimer's disease lies in the pursuit of an early and definitive diagnosis, in order that suitable treatment can be administered. At the present time, definitive diagnosis is restricted to post-mortem examination. Alzheimer's disease also remains without a long-term cure. This research demonstrates the potential role of Raman spectroscopy, combined with principle components analysis (PCA), as a diagnostic method. Analyses of ethically approved ex vivo post-mortem brain tissues (originating from frontal and occipital lobes) from control (3 normal elderly subjects and 3 Huntingdon's disease subjects) and Alzheimer's disease (12 subjects) brain sections, and a further set of 12 blinded samples are presented. Spectra originating from these tissues are highly reproducible, and initial results indicate a vital difference in protein content and conformation, relating to the abnormally high levels of aggregated proteins in the diseased tissues. Further examination of these spectra using PCA allows for the separation of control from diseased tissues. The validation of the PCA models using blinded samples also displays promise for the identification of Alzheimer's disease, in conjunction with secondary information regarding other brain diseases and dementias. These results provide a route for Raman spectroscopy as a possible non-invasive, non-destructive tool for the early diagnosis of Alzheimer's disease.

ERic Acute StrokE Recanalization: A study using predictive analytics to assess a new device for mechanical thrombectomy.

PubMed

Siemonsen, Susanne; Forkert, Nils D; Bernhardt, Martina; Thomalla, Götz; Bendszus, Martin; Fiehler, Jens

2017-08-01

Aim and hypothesis Using a new study design, we investigate whether next-generation mechanical thrombectomy devices improve clinical outcomes in ischemic stroke patients. We hypothesize that this new methodology is superior to intravenous tissue plasminogen activator therapy alone. Methods and design ERic Acute StrokE Recanalization is an investigator-initiated prospective single-arm, multicenter, controlled, open label study to compare the safety and effectiveness of a new recanalization device and distal access catheter in acute ischemic stroke patients with symptoms attributable to acute ischemic stroke and vessel occlusion of the internal cerebral artery or middle cerebral artery. Study outcome The primary effectiveness endpoint is the volume of saved tissue. Volume of saved tissue is defined as difference of the actual infarct volume and the brain volume that is predicted to develop infarction by using an optimized high-level machine learning model that is trained on data from a historical cohort treated with IV tissue plasminogen activator. Sample size estimates Based on own preliminary data, 45 patients fulfilling all inclusion criteria need to complete the study to show an efficacy >38% with a power of 80% and a one-sided alpha error risk of 0.05 (based on a one sample t-test). Discussion ERic Acute StrokE Recanalization is the first prospective study in interventional stroke therapy to use predictive analytics as primary and secondary endpoint. Such trial design cannot replace randomized controlled trials with clinical endpoints. However, ERic Acute StrokE Recanalization could serve as an exemplary trial design for evaluating nonpivotal neurovascular interventions.

Role of sediment size and biostratinomy on the development of biofilms in recent avian vertebrate remains

NASA Astrophysics Data System (ADS)

Peterson, Joseph E.; Lenczewski, Melissa E.; Clawson, Steven R.; Warnock, Jonathan P.

2017-04-01

Microscopic soft tissues have been identified in fossil vertebrate remains collected from various lithologies. However, the diagenetic mechanisms to preserve such tissues have remained elusive. While previous studies have described infiltration of biofilms in Haversian and Volkmann’s canals, biostratinomic alteration (e.g., trampling), and iron derived from hemoglobin as playing roles in the preservation processes, the influence of sediment texture has not previously been investigated. This study uses a Kolmogorov Smirnov Goodness-of-Fit test to explore the influence of biostratinomic variability and burial media against the infiltration of biofilms in bone samples. Controlled columns of sediment with bone samples were used to simulate burial and subsequent groundwater flow. Sediments used in this study include clay-, silt-, and sand-sized particles modeled after various fluvial facies commonly associated with fossil vertebrates. Extant limb bone samples obtained from Gallus gallus domesticus (Domestic Chicken) buried in clay-rich sediment exhibit heavy biofilm infiltration, while bones buried in sands and silts exhibit moderate levels. Crushed bones exhibit significantly lower biofilm infiltration than whole bone samples. Strong interactions between biostratinomic alteration and sediment size are also identified with respect to biofilm development. Sediments modeling crevasse splay deposits exhibit considerable variability; whole-bone crevasse splay samples exhibit higher frequencies of high-level biofilm infiltration, and crushed-bone samples in modeled crevasse splay deposits display relatively high frequencies of low-level biofilm infiltration. These results suggest that sediment size, depositional setting, and biostratinomic condition play key roles in biofilm infiltration in vertebrate remains, and may influence soft tissue preservation in fossil vertebrates.

Comparison of hard tissues that are useful for DNA analysis in forensic autopsy.

PubMed

Kaneko, Yu; Ohira, Hiroshi; Tsuda, Yukio; Yamada, Yoshihiro

2015-11-01

Forensic analysis of DNA from hard tissues can be important when investigating a variety of cases resulting from mass disaster or criminal cases. This study was conducted to evaluate the most suitable tissues, method and sample size for processing of hard tissues prior to DNA isolation. We also evaluated the elapsed time after death in relation to the quantity of DNA extracted. Samples of hard tissues (37 teeth, 42 skull, 42 rib, and 39 nails) from 42 individuals aged between 50 and 83 years were used. The samples were taken from remains following forensic autopsy (from 2 days to 2 years after death). To evaluate the integrity of the nuclear DNA isolated, the percentage of allele calls for short tandem repeat profiles were compared between the hard tissues. DNA typing results indicated that until 1 month after death, any of the four hard tissue samples could be used as an alternative to teeth, allowing analysis of all of the loci. However, in terms of the sampling site, collection method and sample size adjustment, the rib appeared to be the best choice in view of the ease of specimen preparation. Our data suggest that the rib could be an alternative hard tissue sample for DNA analysis of human remains. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Implementation of a non-lethal biopsy punch monitoring program for mercury in smallmouth bass, Micropterus dolomieu Lacepède, from the Eleven Point River, Missouri

USGS Publications Warehouse

Ackerson, J.R.; Schmitt, C.J.; McKee, M.J.; Brumbaugh, W.G.

2013-01-01

A non-lethal biopsy method for monitoring mercury (Hg) concentrations in smallmouth bass (Micropterus dolomieu; smallmouth) from the Eleven Point River in southern Missouri USA was evaluated. A biopsy punch was used to remove a muscle tissue plug from the area immediately below the anterior dorsal fin of 31 smallmouth. An additional 35 smallmouth (controls) were held identically except that no tissue plug was removed. After sampling, all fish were held in a concrete hatchery raceway for 6 weeks. Mean survival at the end of the holding period was 97 % for both groups. Smallmouth length, weight and Fulton’s condition factor at the end of the holding period were also similar between plugged and non-plugged controls, indicating that the biopsy procedure had minimal impact on growth under these conditions. Tissue plug Hg concentrations were similar to smallmouth Hg data obtained in previous years by removing the entire fillet for analysis.

Implementation of a non-lethal biopsy punch monitoring program for mercury in smallmouth bass, Micropterus dolomieu Lacepede, from the Eleven Point River, Missouri

USGS Publications Warehouse

Ackerson, R.J.; McKee, J.M.; Schmitt, C.J.; Brumbaugh, William G.

2014-01-01

A non-lethal biopsy method for monitoring mercury (Hg) concentrations in smallmouth bass (Micropterus dolomieu; smallmouth) from the Eleven Point River in southern Missouri USA was evaluated. A biopsy punch was used to remove a muscle tissue plug from the area immediately below the anterior dorsal fin of 31 smallmouth. An additional 35 smallmouth (controls) were held identically except that no tissue plug was removed. After sampling, all fish were held in a concrete hatchery raceway for 6 weeks. Mean survival at the end of the holding period was 97 % for both groups. Smallmouth length, weight and Fulton’s condition factor at the end of the holding period were also similar between plugged and non-plugged controls, indicating that the biopsy procedure had minimal impact on growth under these conditions. Tissue plug Hg concentrations were similar to smallmouth Hg data obtained in previous years by removing the entire fillet for analysis.

Normalization of gene expression measurement of tissue samples obtained by transurethral resection of bladder tumors.

PubMed

Pop, Laura A; Pileczki, Valentina; Cojocneanu-Petric, Roxana M; Petrut, Bogdan; Braicu, Cornelia; Jurj, Ancuta M; Buiga, Rares; Achimas-Cadariu, Patriciu; Berindan-Neagoe, Ioana

2016-01-01

Sample processing is a crucial step for all types of genomic studies. A major challenge for researchers is to understand and predict how RNA quality affects the identification of transcriptional differences (by introducing either false-positive or false-negative errors). Nanotechnologies help improve the quality and quantity control for gene expression studies. The study was performed on 14 tumor and matched normal pairs of tissue from patients with bladder urothelial carcinomas. We assessed the RNA quantity by using the NanoDrop spectrophotometer and the quality by nano-microfluidic capillary electrophoresis technology provided by Agilent 2100 Bioanalyzer. We evaluated the amplification status of three housekeeping genes and one small nuclear RNA gene using the ViiA 7 platform, with specific primers. Every step of the sample handling protocol, which begins with sample harvest and ends with the data analysis, is of utmost importance due to the fact that it is time consuming, labor intensive, and highly expensive. High temperature of the surgical procedure does not affect the small nucleic acid sequences in comparison with the mRNA. Gene expression is clearly affected by the RNA quality, but less affected in the case of small nuclear RNAs. We proved that the high-temperature, highly invasive transurethral resection of bladder tumor procedure damages the tissue and affects the integrity of the RNA from biological specimens.

Normalization of gene expression measurement of tissue samples obtained by transurethral resection of bladder tumors

PubMed Central

Pop, Laura A; Pileczki, Valentina; Cojocneanu-Petric, Roxana M; Petrut, Bogdan; Braicu, Cornelia; Jurj, Ancuta M; Buiga, Rares; Achimas-Cadariu, Patriciu; Berindan-Neagoe, Ioana

2016-01-01

Background Sample processing is a crucial step for all types of genomic studies. A major challenge for researchers is to understand and predict how RNA quality affects the identification of transcriptional differences (by introducing either false-positive or false-negative errors). Nanotechnologies help improve the quality and quantity control for gene expression studies. Patients and methods The study was performed on 14 tumor and matched normal pairs of tissue from patients with bladder urothelial carcinomas. We assessed the RNA quantity by using the NanoDrop spectrophotometer and the quality by nano-microfluidic capillary electrophoresis technology provided by Agilent 2100 Bioanalyzer. We evaluated the amplification status of three housekeeping genes and one small nuclear RNA gene using the ViiA 7 platform, with specific primers. Results Every step of the sample handling protocol, which begins with sample harvest and ends with the data analysis, is of utmost importance due to the fact that it is time consuming, labor intensive, and highly expensive. High temperature of the surgical procedure does not affect the small nucleic acid sequences in comparison with the mRNA. Conclusion Gene expression is clearly affected by the RNA quality, but less affected in the case of small nuclear RNAs. We proved that the high-temperature, highly invasive transurethral resection of bladder tumor procedure damages the tissue and affects the integrity of the RNA from biological specimens. PMID:27330317

Residual antibiotics in decontaminated human cardiovascular tissues intended for transplantation and risk of falsely negative microbiological analyses.

PubMed

Buzzi, Marina; Guarino, Anna; Gatto, Claudio; Manara, Sabrina; Dainese, Luca; Polvani, Gianluca; Tóthová, Jana D'Amato

2014-01-01

We investigated the presence of antibiotics in cryopreserved cardiovascular tissues and cryopreservation media, after tissue decontamination with antibiotic cocktails, and the impact of antibiotic residues on standard tissue bank microbiological analyses. Sixteen cardiovascular tissues were decontaminated with bank-prepared cocktails and cryopreserved by two different tissue banks according to their standard operating procedures. Before and after decontamination, samples underwent microbiological analysis by standard tissue bank methods. Cryopreserved samples were tested again with and without the removal of antibiotic residues using a RESEP tube, after thawing. Presence of antibiotics in tissue homogenates and processing liquids was determined by a modified agar diffusion test. All cryopreserved tissue homogenates and cryopreservation media induced important inhibition zones on both Staphylococcus aureus- and Pseudomonas aeruginosa-seeded plates, immediately after thawing and at the end of the sterility test. The RESEP tube treatment markedly reduced or totally eliminated the antimicrobial activity of tested tissues and media. Based on standard tissue bank analysis, 50% of tissues were found positive for bacteria and/or fungi, before decontamination and 2 out of 16 tested samples (13%) still contained microorganisms after decontamination. After thawing, none of the 16 cryopreserved samples resulted positive with direct inoculum method. When the same samples were tested after removal of antibiotic residues, 8 out of 16 (50%) were contaminated. Antibiotic residues present in tissue allografts and processing liquids after decontamination may mask microbial contamination during microbiological analysis performed with standard tissue bank methods, thus resulting in false negatives.

Residual Antibiotics in Decontaminated Human Cardiovascular Tissues Intended for Transplantation and Risk of Falsely Negative Microbiological Analyses

PubMed Central

Gatto, Claudio; Manara, Sabrina; Dainese, Luca; Polvani, Gianluca; Tóthová, Jana D'Amato

2014-01-01

We investigated the presence of antibiotics in cryopreserved cardiovascular tissues and cryopreservation media, after tissue decontamination with antibiotic cocktails, and the impact of antibiotic residues on standard tissue bank microbiological analyses. Sixteen cardiovascular tissues were decontaminated with bank-prepared cocktails and cryopreserved by two different tissue banks according to their standard operating procedures. Before and after decontamination, samples underwent microbiological analysis by standard tissue bank methods. Cryopreserved samples were tested again with and without the removal of antibiotic residues using a RESEP tube, after thawing. Presence of antibiotics in tissue homogenates and processing liquids was determined by a modified agar diffusion test. All cryopreserved tissue homogenates and cryopreservation media induced important inhibition zones on both Staphylococcus aureus- and Pseudomonas aeruginosa-seeded plates, immediately after thawing and at the end of the sterility test. The RESEP tube treatment markedly reduced or totally eliminated the antimicrobial activity of tested tissues and media. Based on standard tissue bank analysis, 50% of tissues were found positive for bacteria and/or fungi, before decontamination and 2 out of 16 tested samples (13%) still contained microorganisms after decontamination. After thawing, none of the 16 cryopreserved samples resulted positive with direct inoculum method. When the same samples were tested after removal of antibiotic residues, 8 out of 16 (50%) were contaminated. Antibiotic residues present in tissue allografts and processing liquids after decontamination may mask microbial contamination during microbiological analysis performed with standard tissue bank methods, thus resulting in false negatives. PMID:25397402

Computational deconvolution of genome wide expression data from Parkinson's and Huntington's disease brain tissues using population-specific expression analysis

PubMed Central

Capurro, Alberto; Bodea, Liviu-Gabriel; Schaefer, Patrick; Luthi-Carter, Ruth; Perreau, Victoria M.

2015-01-01

The characterization of molecular changes in diseased tissues gives insight into pathophysiological mechanisms and is important for therapeutic development. Genome-wide gene expression analysis has proven valuable for identifying biological processes in neurodegenerative diseases using post mortem human brain tissue and numerous datasets are publically available. However, many studies utilize heterogeneous tissue samples consisting of multiple cell types, all of which contribute to global gene expression values, confounding biological interpretation of the data. In particular, changes in numbers of neuronal and glial cells occurring in neurodegeneration confound transcriptomic analyses, particularly in human brain tissues where sample availability and controls are limited. To identify cell specific gene expression changes in neurodegenerative disease, we have applied our recently published computational deconvolution method, population specific expression analysis (PSEA). PSEA estimates cell-type-specific expression values using reference expression measures, which in the case of brain tissue comprises mRNAs with cell-type-specific expression in neurons, astrocytes, oligodendrocytes and microglia. As an exercise in PSEA implementation and hypothesis development regarding neurodegenerative diseases, we applied PSEA to Parkinson's and Huntington's disease (PD, HD) datasets. Genes identified as differentially expressed in substantia nigra pars compacta neurons by PSEA were validated using external laser capture microdissection data. Network analysis and Annotation Clustering (DAVID) identified molecular processes implicated by differential gene expression in specific cell types. The results of these analyses provided new insights into the implementation of PSEA in brain tissues and additional refinement of molecular signatures in human HD and PD. PMID:25620908

The Hit and Away technique: optimal usage of the ultrasonic scalpel in laparoscopic gastrectomy.

PubMed

Irino, Tomoyuki; Hiki, Naoki; Ohashi, Manabu; Nunobe, Souya; Sano, Takeshi; Yamaguchi, Toshiharu

2016-01-01

Thermal injury and unexpected bleeding caused by ultrasonic scalpels can lead to fatal complications in laparoscopic gastrectomy (LG), such as postoperative pancreatic fistulas (POPF). In this study, we developed the "Hit and Away" protocol for optimal usage of the ultrasonic scalpel, which in essence involves dividing tissues and vessels in batches using the tip of the scalpel to control tissue temperature. To assess the effectiveness of the technique, the surface temperature of the mesocolon of female swine after ultrasonic scalpel activations was measured, and tissue samples were collected to evaluate microscopic thermal injury to the pancreas. In parallel, we retrospectively surveyed 216 patients who had undergone LG before or after the introduction of this technique and assessed the ability of this technique to reduce POPF. The tissue temperature of the swine mesocolon reached 43 °C, a temperature at which adipose tissue melted but fibrous tissue, including vessels, remained intact. The temperature returned to baseline within 3 s of turning off the ultrasonic scalpel, demonstrating the advantage of using ultrasonic scalpel in a pulsatile manner. Tissue samples from the pancreas demonstrated that the extent of thermal injury post-procedure was limited to the capsule of the pancreas. Moreover, with respect to the clinical outcomes before and after the introduction of this technique, POPF incidence decreased significantly from 7.8 to 1.0% (p = 0.021). The "Hit and Away" technique can reduce blood loss and thermal injury to the pancreas and help to ensure the safety of lymph node dissection in LG.

Morphometric study of uninvolved rectal mucosa 10 cm and 20 cm away from the malignant tumor.

PubMed

Despotović, Sanja Z; Milićević, Novica M; Milosević, Dragoslav P; Despotović, Nebojsa; Erceg, Predrag; Bojić, Bozidar; Bojić, Danijela; Svorcan, Petar; Mihajlović, Gordana; Dorđević, Jelena; Lalić, Ivana M; Milićević, Zivana

2014-02-01

Recently, many details of the interplay between tumor cells and tumor-associated stromal elements leading to the progression of malignant disease were elucidated. In contrast, little is known about the role of uninvolved stromal tissue in the remote surrounding of the malignant tumor. Therefore, we performed a computer-aided morphometric study of rectal mucosa in samples taken 10 cm and 20 cm away from the malignant tumor during endoscopic examination of 23 patients older than 60 years. The samples of rectal mucosa from 10 healthy persons of corresponding age subjected to diagnostic rectoscopy during active screening for asymptomatic cancer were used as control. All structural elements of the rectal mucosa were studied and the number of nucleated cells in the lamina propria per 0.1 mm² of tissue was assessed. Our study revealed a reduced number of cells in the lamina propria of the rectal mucosa 10 cm and 20 cm away from the tumor lesion in both male and female patients. The decreased mucosal height and increased crypt number were registered in female patients 10 cm away from the tumor. The connective tissue of lamina propria showed a disorderly organization: the collagen fibers were frail, loosely arranged and signs of tissue edema were present. Small blood vessels and capillaries were much more frequently seen than in healthy tissue. Our results demonstrate the complex interactions between the cancer and remote mucosal tissue of the affected organ.

Tissue-Specific Methylation of Long Interspersed Nucleotide Element-1 of Homo Sapiens (L1Hs) During Human Embryogenesis and Roles in Neural Tube Defects.

PubMed

Wang, L; Chang, S; Guan, J; Shangguan, S; Lu, X; Wang, Z; Wu, L; Zou, J; Zhao, H; Bao, Y; Qiu, Z; Niu, B; Zhang, T

2015-01-01

Epigenetic regulation of long interspersed nucleotide element-1 (LINE-1) retrotransposition events plays crucial roles during early development. Previously we showed that LINE-1 hypomethylation in neuronal tissues is associated with pathogenesis of neural tube defect (NTD). Herein, we further evaluated LINE-1 Homo sapiens (L1Hs) methylation in tissues derived from three germ layers of stillborn NTD fetuses, to define patterns of tissue specific methylation and site-specific hypomethylation at CpG sites within an L1Hs promoter region. Stable, tissue-specific L1Hs methylation patterns throughout three germ layer lineages of the fetus, placenta, and maternal peripheral blood were observed. Samples from maternal peripheral blood exhibited the highest level of L1Hs methylation (64.95%) and that from placenta showed the lowest (26.82%). Between samples from NTDs and controls, decrease in L1Hs methylation was only significant in NTD-affected brain tissue at 7.35%, especially in females (8.98%). L1Hs hypomethylation in NTDs was also associated with a significant increase in expression level of an L1Hs-encoded transcript in females (r = -0.846, p = 0.004). This could be due to genomic DNA instability and alternation in chromatins accessibility resulted from abnormal L1Hs hypomethylation, as showed in this study with HCT-15 cells treated with methylation inhibitor 5-Aza.

Optimal molecular profiling of tissue and tissue components: defining the best processing and microdissection methods for biomedical applications.

PubMed

Bova, G Steven; Eltoum, Isam A; Kiernan, John A; Siegal, Gene P; Frost, Andra R; Best, Carolyn J M; Gillespie, John W; Su, Gloria H; Emmert-Buck, Michael R

2005-02-01

Isolation of well-preserved pure cell populations is a prerequisite for sound studies of the molecular basis of any tissue-based biological phenomenon. This article reviews current methods for obtaining anatomically specific signals from molecules isolated from tissues, a basic requirement for productive linking of phenotype and genotype. The quality of samples isolated from tissue and used for molecular analysis is often glossed over or omitted from publications, making interpretation and replication of data difficult or impossible. Fortunately, recently developed techniques allow life scientists to better document and control the quality of samples used for a given assay, creating a foundation for improvement in this area. Tissue processing for molecular studies usually involves some or all of the following steps: tissue collection, gross dissection/identification, fixation, processing/embedding, storage/archiving, sectioning, staining, microdissection/annotation, and pure analyte labeling/identification and quantification. We provide a detailed comparison of some current tissue microdissection technologies, and provide detailed example protocols for tissue component handling upstream and downstream from microdissection. We also discuss some of the physical and chemical issues related to optimal tissue processing, and include methods specific to cytology specimens. We encourage each laboratory to use these as a starting point for optimization of their overall process of moving from collected tissue to high quality, appropriately anatomically tagged scientific results. In optimized protocols is a source of inefficiency in current life science research. Improvement in this area will significantly increase life science quality and productivity. The article is divided into introduction, materials, protocols, and notes sections. Because many protocols are covered in each of these sections, information relating to a single protocol is not contiguous. To get the greatest benefit from this article, readers are advised to read through the entire article first, identify protocols appropriate to their laboratory for each step in their workflow, and then reread entries in each section pertaining to each of these single protocols.

Optimal molecular profiling of tissue and tissue components: defining the best processing and microdissection methods for biomedical applications.

PubMed

Rodriguez-Canales, Jaime; Hanson, Jeffrey C; Hipp, Jason D; Balis, Ulysses J; Tangrea, Michael A; Emmert-Buck, Michael R; Bova, G Steven

2013-01-01

Isolation of well-preserved pure cell populations is a prerequisite for sound studies of the molecular basis of any tissue-based biological phenomenon. This updated chapter reviews current methods for obtaining anatomically specific signals from molecules isolated from tissues, a basic requirement for productive linking of phenotype and genotype. The quality of samples isolated from tissue and used for molecular analysis is often glossed over or omitted from publications, making interpretation and replication of data difficult or impossible. Fortunately, recently developed techniques allow life scientists to better document and control the quality of samples used for a given assay, creating a foundation for improvement in this area. Tissue processing for molecular studies usually involves some or all of the following steps: tissue collection, gross dissection/identification, fixation, processing/embedding, storage/archiving, sectioning, staining, microdissection/annotation, and pure analyte labeling/identification and quantification. We provide a detailed comparison of some current tissue microdissection technologies and provide detailed example protocols for tissue component handling upstream and downstream from microdissection. We also discuss some of the physical and chemical issues related to optimal tissue processing and include methods specific to cytology specimens. We encourage each laboratory to use these as a starting point for optimization of their overall process of moving from collected tissue to high-quality, appropriately anatomically tagged scientific results. Improvement in this area will significantly increase life science quality and productivity. The chapter is divided into introduction, materials, protocols, and notes subheadings. Because many protocols are covered in each of these sections, information relating to a single protocol is not contiguous. To get the greatest benefit from this chapter, readers are advised to read through the entire chapter first, identify protocols appropriate to their laboratory for each step in their workflow, and then reread entries in each section pertaining to each of these single protocols.

United States Transuranium and Uranium Registries. Annual Report, October 1, 1993--September 30, 1994

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kathren, R.L.; Harwick, L.A.

1995-08-01

This report summarizes the salient activities and progress of the United States Transuranium. and Uranium Registries for the period October 1, 1993 through September 30, 1994, along with details of specific programs areas including the National Human Radiobiology Tissue Repository (NHRTR) and tissue radiochemistry analysis project. Responsibility for tissue radioanalysis was transferred from Los Alamos National Laboratory to Washington State University in February 1994. The University of Washington was selected as the Quality Assurance/Quality Control laboratory and a three way intercomparison with them and LANL has been initiated. The results of the initial alpha spectrometry intercomparison showed excellent agreement amongmore » the laboratories and are documented in full in the Appendices to the report. The NHRTR serves as the initial point of receipt for samples received from participants in the USTUR program. Samples are weighed, divided, and reweighed, and a portion retained by the NHRTR as backup or for use in other studies. Tissue specimens retained in the NHRTR are maintained frozen at -70 C and include not only those from USTUR registrants but also those from the radium dial painter and thorium worker studies formerly conducted by Argonne National Laboratory. In addition, there are fixed tissues and a large collection of histopathology slides from all the studies, plus about 20,000 individual solutions derived from donated tissues. These tissues and tissue related materials are made available to other investigators for legitimate research purposes. Ratios of the concentration of actinides in various tissues have been used to evaluate the biokinetics, and retention half times of plutonium and americium. Retention half times for plutonium in various soft tissues range from 10-20 y except for the testes for which a retention half time of 58 y was observed. For americium, the retention half time in various soft tissues studied was 2.2-3.5 y.« less

Death-associated protein kinase promoter methylation correlates with clinicopathological and prognostic features in nonsmall cell lung cancer patients: A cohort study.

PubMed

Yang, Xiao-Yu; Zhang, Jun; Yu, Xiao-Ling; Zheng, Guo-Feng; Zhao, Fei; Jia, Xiao-Jing

2018-01-01

The objective was to study the correlation between death-associated protein kinase (DAPK) promoter methylation and the clinicopathological and prognostic features in nonsmall cell lung cancer (NSCLC) patients. A total of 117 NSCLC patients were recruited into our study between December 2012 and December 2014. Methylation-specific polymerase chain reaction was employed to detect the methylation status of DAPK in cancer tissues, peficancerous tissues, and serum samples of 117 NSCLC patients. In addition, serum samples of 115 healthy subjects were analyzed as controls. A literature search of English and Chinese databases, based on predefined criteria, identified published studies closely related to this study. Data were extracted, and meta-analysis was performed using STATA 12.0 software (STATA Corporation, College Station, TX, USA). Our study results showed that DAPK promoter methylation frequency was significantly higher in NSCLC tissues compared to peficancerous normal tissues (58.1% vs. 12.8%, χ 2 = 52.45, P < 0.001). When serum samples were compared, DAPK methylation frequency in NSCLC patients was higher than the control group (27.4% vs. 0, χ 2 = 37.07, P < 0.001). Our meta-analysis results demonstrated that DAPK methylation frequency was lower in tumor node metastasis (TNM) stage I-II compared to TNM stage III-IV (relative risk [RR] =0.87, 95% confidence interval [CI] =0.76-0.99, P = 0.041). DAPK promoter methylation frequency in NSCLC patients with lymph node metastasis was significantly higher compared to the patients with no metastases (RR = 1.26, 95% CI = 1.04-1.52, P = 0.020). Finally, the 5-year survival rate was lower in NSCLC patient group with high frequency of DAPK methylation, compared to the patient group with unmethylated DAPK (RR = 0.71, 95% CI = 0.56-0.89, P = 0.004). Our results showed that DAPK promoter methylation is tightly correlated with clinicopathological features of NSCLC and is associated with poor prognosis in patients.

Experimentally reduced root–microbe interactions reveal limited plasticity in functional root traits in Acer and Quercus

PubMed Central

Lee, Mei-Ho; Comas, Louise H.; Callahan, Hilary S.

2014-01-01

Background and Aims Interactions between roots and soil microbes are critical components of below-ground ecology. It is essential to quantify the magnitude of root trait variation both among and within species, including variation due to plasticity. In addition to contextualizing the magnitude of plasticity relative to differences between species, studies of plasticity can ascertain if plasticity is predictable and whether an environmental factor elicits changes in traits that are functionally advantageous. Methods To compare functional traits and trait plasticities in fine root tissues with natural and reduced levels of colonization by microbial symbionts, trimmed and surface-sterilized root segments of 2-year-old Acer rubrum and Quercus rubra seedlings were manipulated. Segments were then replanted into satellite pots filled with control or heat-treated soil, both originally derived from a natural forest. Mycorrhizal colonization was near zero in roots grown in heat-treated soil; roots grown in control soil matched the higher colonization levels observed in unmanipulated root samples collected from field locations. Key Results Between-treatment comparisons revealed negligible plasticity for root diameter, branching intensity and nitrogen concentration across both species. Roots from treated soils had decreased tissue density (approx. 10–20 %) and increased specific root length (approx. 10–30 %). In contrast, species differences were significant and greater than treatment effects in traits other than tissue density. Interspecific trait differences were also significant in field samples, which generally resembled greenhouse samples. Conclusions The combination of experimental and field approaches was useful for contextualizing trait plasticity in comparison with inter- and intra-specific trait variation. Findings that root traits are largely species dependent, with the exception of root tissue density, are discussed in the context of current literature on root trait variation, interactions with symbionts and recent progress in standardization of methods for quantifying root traits. PMID:24363335

Potential for autoimmune pathogenesis of Rift Valley Fever virus retinitis.

PubMed

Newman-Gerhardt, Shoshana; Muiruri, Samuel; Muchiri, Eric; Peters, Clarence J; Morrill, John; Lucas, Alexander H; King, Charles H; Kazura, James; LaBeaud, Angelle Desiree

2013-09-01

Rift Valley Fever (RVF) is a significant threat to human health because it can progress to retinitis, encephalitis, and hemorrhagic fever. The timing of onset of Rift Valley Fever virus (RVFV) retinitis suggests an autoimmune origin. To determine whether RVFV retinitis is associated with increased levels of IgG against retinal tissue, we measured and compared levels of IgG against healthy human eye tissue by immunohistochemical analysis. We found that serum samples from RVFV-exposed Kenyans with retinitis (n = 8) were slightly more likely to have antibodies against retinal tissue than control populations, but the correlation was not statistically significant. Further investigation into the possible immune pathogenesis of RVFV retinitis could lead to improved therapies to prevent or treat this severe complication.

Potential for Autoimmune Pathogenesis of Rift Valley Fever Virus Retinitis

PubMed Central

Newman-Gerhardt, Shoshana; Muiruri, Samuel; Muchiri, Eric; Peters, Clarence J.; Morrill, John; Lucas, Alexander H.; King, Charles H.; Kazura, James; LaBeaud, Angelle Desiree

2013-01-01

Rift Valley Fever (RVF) is a significant threat to human health because it can progress to retinitis, encephalitis, and hemorrhagic fever. The timing of onset of Rift Valley Fever virus (RVFV) retinitis suggests an autoimmune origin. To determine whether RVFV retinitis is associated with increased levels of IgG against retinal tissue, we measured and compared levels of IgG against healthy human eye tissue by immunohistochemical analysis. We found that serum samples from RVFV-exposed Kenyans with retinitis (n = 8) were slightly more likely to have antibodies against retinal tissue than control populations, but the correlation was not statistically significant. Further investigation into the possible immune pathogenesis of RVFV retinitis could lead to improved therapies to prevent or treat this severe complication. PMID:23918215

Robotic Surgery

ERIC Educational Resources Information Center

Childress, Vincent W.

2007-01-01

The medical field has many uses for automated and remote-controlled technology. For example, if a tissue sample is only handled in the laboratory by a robotic handling system, then it will never come into contact with a human. Such a system not only helps to automate the medical testing process, but it also helps to reduce the chances of…

[Role of biometric analysis in the retrospective assessment of exposure to asbestos].

PubMed

Pairon, J C; Dumortier, P

1999-12-01

Despite intrinsic limitations due to differences in the bio-persistence of the various asbestos types, in the definition of control populations and in analytical techniques used by the laboratories, mineralogical analysis of biological samples is useful in the assessment of past exposure to asbestos. It provides additional information to occupational and environmental questionnaires, particularly when exposure to asbestos is doubtful, unknown or forgotten by a subject. Results should be interpreted taking into account clinical information. A positive result does not mean existence of asbestos-related disease. A negative result does not exclude previous significant asbestos exposure, clearly identified by an occupational questionnaire (particularly for exposure to chrysotile). Threshold values indicative of a high probability of previous asbestos exposure have been established for bronchoalveolar lavage fluid (BALF) samples and lung tissue samples. Quantification of asbestos bodies by light microscopy is easy to perform. Sensitivity and specificity of this analysis towards the total pulmonary asbestos fiber burden is good. Therefore this analysis should be performed first. Mineralogical analysis in BALF or lung tissue should be considered only when sampling is supported by diagnostic or therapeutic implications.

Determination of friction coefficient in unconfined compression of brain tissue.

PubMed

Rashid, Badar; Destrade, Michel; Gilchrist, Michael D

2012-10-01

Unconfined compression tests are more convenient to perform on cylindrical samples of brain tissue than tensile tests in order to estimate mechanical properties of the brain tissue because they allow homogeneous deformations. The reliability of these tests depends significantly on the amount of friction generated at the specimen/platen interface. Thus, there is a crucial need to find an approximate value of the friction coefficient in order to predict a possible overestimation of stresses during unconfined compression tests. In this study, a combined experimental-computational approach was adopted to estimate the dynamic friction coefficient μ of porcine brain matter against metal platens in compressive tests. Cylindrical samples of porcine brain tissue were tested up to 30% strain at variable strain rates, both under bonded and lubricated conditions in the same controlled environment. It was established that μ was equal to 0.09±0.03, 0.18±0.04, 0.18±0.04 and 0.20±0.02 at strain rates of 1, 30, 60 and 90/s, respectively. Additional tests were also performed to analyze brain tissue under lubricated and bonded conditions, with and without initial contact of the top platen with the brain tissue, with different specimen aspect ratios and with different lubricants (Phosphate Buffer Saline (PBS), Polytetrafluoroethylene (PTFE) and Silicone). The test conditions (lubricant used, biological tissue, loading velocity) adopted in this study were similar to the studies conducted by other research groups. This study will help to understand the amount of friction generated during unconfined compression of brain tissue for strain rates of up to 90/s. Copyright © 2012 Elsevier Ltd. All rights reserved.

A system for the rapid detection of bacterial contamination in cell-based therapeutica

NASA Astrophysics Data System (ADS)

Bolwien, Carsten; Erhardt, Christian; Sulz, Gerd; Thielecke, Hagen; Johann, Robert; Pudlas, Marieke; Mertsching, Heike; Koch, Steffen

2010-02-01

Monitoring the sterility of cell or tissue cultures is of major concern, particularly in the fields of regenerative medicine and tissue engineering when implanting cells into the human body. Our sterility-control system is based on a Raman micro-spectrometer and is able to perform fast sterility testing on microliters of liquid samples. In conventional sterility control, samples are incubated for weeks to proliferate the contaminants to concentrations above the detection limit of conventional analysis. By contrast, our system filters particles from the liquid sample. The filter chip fabricated in microsystem technology comprises a silicon nitride membrane with millions of sub-micrometer holes to retain particles of critical sizes and is embedded in a microfluidic cell specially suited for concomitant microscopic observation. After filtration, identification is carried out on the single particle level: image processing detects possible contaminants and prepares them for Raman spectroscopic analysis. A custom-built Raman-spectrometer-attachment coupled to the commercial microscope uses 532nm or 785nm Raman excitation and records spectra up to 3400cm-1. In the final step, the recorded spectrum of a single particle is compared to an extensive library of GMP-relevant organisms, and classification is carried out based on a support vector machine.

Autoinflammation Around AES Total Ankle Replacement Implants.

PubMed

Koivu, Helka; Takakubo, Yuya; Mackiewicz, Zygmunt; Al-Samadi, Ahmed; Soininen, Antti; Peled, Nitai; Kukis, Modestas; Trokovic, Nina; Konttinen, Yrjö T

2015-12-01

Failure of total ankle replacement (TAR) can be characterized by early peri-implant osteolysis even in the presence of very modest numbers of wear particles. The hypothesis of the study was that this reaction is in part mediated by autoinflammatory responses mediated via damage-associated molecular patterns (DAMPs, danger signals) and pattern-recognizing danger signal receptors (PRRs). Peri-implant tissue and control samples from 10 patients with AES implants were immunostained for hypoxia inducible factor-1α (HIF-1α), activated caspase-3, high-mobility group box 1 (HMGB1), receptor for advanced glycation end product (RAGE), and toll-like receptors TLR2 and TLR4. Results were evaluated on a 0 to 4 scale (from 0% to >50% stained area). Peri-implant tissue around failed TAR implants had a relatively high mean HIF-1α score of 3 on a scale, which however was similar in control samples. HMGB1 (a DAMP) was seen to be mobilized from nuclei to cellular cytoplasm, and the active caspase-3(+) cells were increased. All PRRs were increased in revision samples. Increased expression of HMGB1 and other danger signals together with increased PRR-dependent responsiveness could contribute to autoinflammatory peri-implantitis, multilocular cyst formation, and osteolysis in failed TAR implants. Level IV, case series. © The Author(s) 2015.

HIF-1 Alpha and Placental Growth Factor in Pregnancies Complicated With Preeclampsia: A Qualitative and Quantitative Analysis.

PubMed

Rath, Gayatri; Aggarwal, Ruby; Jawanjal, Poonam; Tripathi, Richa; Batra, Aruna

2016-01-01

The pathophysiology of preeclampsia is not clearly understood worldwide. Hypoxia inducible factor 1α (HIF-1α) is thought to be the preliminary factor for the hypoxic conditions prevailing in preeclampsia, which causes imbalance in the expression of angiogenic proteins. A proangiogenic protein, placental growth factor (PIGF), is reported to be dysregulated in preeclampsia. Therefore, this study focuses on the investigation of HIF-1α and PIGF in preeclamptic conditions and a possible molecular association between them. Placental tissue (n = 45 + 45) and serum samples (n = 80 + 80) of preeclamptic patients and healthy control were collected and processed for the analysis of HIF-1α and PIGF by immunohistochemistry and enzyme-linked immunosorbent assay (ELISA). In preeclamptic group, the significant nuclear and cytoplasmic expression of HIF-1α was noticed in syncytiotrophoblast (P = 0.0001) but in control placenta, it was localized to cytoplasm (P = 0.0001). The intensity of PIGF expression was lower in syncytiotrophoblast cytoplasm (P = 0.0001) in preeclamptic cases as compared with control. Also, the significant upregulated concentration of HIF-1α and downregulated PIGF was observed in serum samples of preeclamptic woman (P = 0.0001). Thus, there was a significant direct negative correlation between HIF-1α and PIGF both at tissue and serum level (P < 0.01). The direct inverse association between HIF-1α and PIGF in serum and placental tissues may be responsible for the low oxidative stress and endothelial dysfunction, leading to the pathogenesis of preeclampsia. © 2014 Wiley Periodicals, Inc.

Development and validation of a protocol for optimizing the use of paraffin blocks in molecular epidemiological studies: The example from the HPV-AHEAD study.

PubMed

Mena, Marisa; Lloveras, Belen; Tous, Sara; Bogers, Johannes; Maffini, Fausto; Gangane, Nitin; Kumar, Rekha Vijay; Somanathan, Thara; Lucas, Eric; Anantharaman, Devasena; Gheit, Tarik; Castellsagué, Xavier; Pawlita, Michael; de Sanjosé, Silvia; Alemany, Laia; Tommasino, Massimo

2017-01-01

Worldwide use of formalin-fixed paraffin-embedded blocks (FFPE) is extensive in diagnosis and research. Yet, there is a lack of optimized/standardized protocols to process the blocks and verify the quality and presence of the targeted tissue. In the context of an international study on head and neck cancer (HNC)-HPV-AHEAD, a standardized protocol for optimizing the use of FFPEs in molecular epidemiology was developed and validated. First, a protocol for sectioning the FFPE was developed to prevent cross-contamination and distributed between participating centers. Before processing blocks, all sectioning centers underwent a quality control to guarantee a satisfactory training process. The first and last sections of the FFPEs were used for histopathological assessment. A consensus histopathology evaluation form was developed by an international panel of pathologists and evaluated for four indicators in a pilot analysis in order to validate it: 1) presence/type of tumor tissue, 2) identification of other tissue components that could affect the molecular diagnosis and 3) quality of the tissue. No HPV DNA was found in sections from empty FFPE generated in any histology laboratories of HPV-AHEAD consortium and all centers passed quality assurance for processing after quality control. The pilot analysis to validate the histopathology form included 355 HNC cases. The form was filled by six pathologists and each case was randomly assigned to two of them. Most samples (86%) were considered satisfactory. Presence of >50% of invasive carcinoma was observed in all sections of 66% of cases. Substantial necrosis (>50%) was present in <2% of samples. The concordance for the indicators targeted to validate the histopathology form was very high (kappa > 0.85) between first and last sections and fair to high between pathologists (kappa/pabak 0.21-0.72). The protocol allowed to correctly process without signs of contamination all FFPE of the study. The histopathology evaluation of the cases assured the presence of the targeted tissue, identified the presence of other tissues that could disturb the molecular diagnosis and allowed the assessment of tissue quality.

MCT1 and MCT4 kinetic of mRNA expression in different tissues after aerobic exercise at maximal lactate steady state workload.

PubMed

de Araujo, G G; Gobatto, C A; de Barros Manchado-Gobatto, F; Teixeira, L Fm; Dos Reis, I Gm; Caperuto, L C; Papoti, M; Bordin, S; Cavaglieri, C R; Verlengia, R

2015-01-01

We evaluate the mRNA expression of monocarboxylate transporters 1 and 4 (MCT1 and MCT4) in skeletal muscle (soleus, red and white gastrocnemius), heart and liver tissues in mice submitted to a single bout of swimming exercise at the maximal lactate steady state workload (MLSSw). After 72 h of MLSS test, the animals were submitted to a swimming exercise session for 25 min at individual MLSSw. Tissues and muscle samples were obtained at rest (control, n=5), immediately (n=5), 5 h (n=5) and 10 h (n=5) after exercise for determination of the MCT1 and MCT4 mRNA expression (RT-PCR). The MCT1 mRNA expression in liver increased after 10 h in relation to the control, immediate and 5 h groups, but the MCT4 remained unchanged. The MCT1 mRNA expression in heart increased by 31 % after 10 h when compared to immediate, but no differences were observed in relation to the control group. No significant differences were observed for red gastrocnemius in MCT1 and MCT4 mRNA expression. However, white gastrocnemius increased MCT1 mRNA expression immediately when compared to rest, 5 and 10 h test groups. In soleus muscle, the MCT1 mRNA expression increased immediately, 5 and 10 h after exercise when compared to the control. In relation to MCT4 mRNA expression, the soleus increased immediately and 10 h after acute exercise when compared to the control group. The soleus, liver and heart were the main tissues that showed improved the MCT1 mRNA expression, indicating its important role in controlling MLSS concentration in mice.

CYP19 gene expression in subcutaneous adipose tissue is associated with blood pressure in women with polycystic ovary syndrome.

PubMed

Lecke, Sheila B; Morsch, Débora M; Spritzer, Poli M

2011-11-01

In polycystic ovary syndrome (PCOS), hypertension has been linked to androgen excess and insulin resistance. Aromatase, an enzyme encoded by the CYP19 gene, affects androgen metabolism and estrogen synthesis, influencing the androgen to estrogen balance. We characterized CYP19 gene expression in subcutaneous adipose tissue of women with PCOS and normal controls and evaluated the association between subcutaneous fat CYP19 mRNA, circulating hormone levels, and blood pressure. This case-control study was carried out with 31 PCOS patients and 27 BMI-matched normotensive non-hirsute women with regular cycles. Participants underwent anthropometric measurements, collection of blood samples, and adipose tissue biopsy (28 PCOS and 19 controls). Hypertension was defined as systolic blood pressure ≥ 130 mmHg and/or diastolic blood pressure ≥ 85 mmHg. PCOS patients were divided into normotensive and hypertensive. Main outcome measures were serum estrogen and androgen levels, estrogen-to-androgen ratio, and CYP19 gene expression in subcutaneous fat. Subcutaneous CYP19 mRNA was higher in hypertensive PCOS than in control and normotensive PCOS women (p = 0.014). Estrogen-to-androgen ratio was lower in hypertensive PCOS than controls (p < 0.003). Estrogen-to-androgen ratio ≤ 0.06 (median for the three groups) was observed in 91% of hypertensive PCOS women, vs. 37% and 61% in the control and normotensive PCOS groups (p = 0.011). CYP19 gene expression in subcutaneous fat of PCOS patient correlated positively with systolic (p = 0.006) and diastolic blood pressure (p = 0.009). Androgen excess and hyperinsulinemia may play a role in the molecular mechanisms that activate aromatase mRNA transcription in abdominal fat tissue. Copyright © 2011 Elsevier Inc. All rights reserved.

Polydopamine deposition with anodic oxidation for better connective tissue attachment to transmucosal implants.

PubMed

Teng, F; Chen, H; Xu, Y; Liu, Y; Ou, G

2018-04-01

Nowadays, most designs for the transmucosal surface of implants are machined-smooth. However, connective tissue adhered to the smooth surface of an implant has poor mechanical resistance, which can render separation of tissue from the implant interface and induce epithelial downgrowth. Modification of the transmucosal surface of implants, which can help form a good seal of connective tissue, is therefore desired. We hypothesized that anodic oxidation (AO) and polydopamine (PD) deposition could be used to enhance the attachment between an implant and peri-implant connective tissue. We tested this hypothesis in the mandibles of Beagle dogs. AO and PD were used to modify the transmucosal region of transmucosal implants (implant neck). The surface microstructure, surface roughness and elemental composition were investigated in vitro. L929 mouse fibroblasts were cultured to test the effect of PD on cell adhesion. Six Beagle dogs were used for the in vivo experiment (n = 6 dogs per group). Three months after building the edentulous animal model, four groups of implants (control, AO, PD and AO + PD) were inserted. After 4 months of healing, samples were harvested for histometric analyses. The surfaces of anodized implant necks were overlaid with densely distributed pores, 2-7 μm in size. On the PD-modified surfaces, N1s, the chemical bond of nitrogen in PD, was detected using X-ray photoelectron spectroscopy. L929 developed pseudopods more quickly on the PD-modified surfaces than on the surfaces of the control group. The in vivo experiment showed a longer connective tissue seal and a more coronally located peri-implant soft-tissue attachment in the AO + PD group than in the control group (P < .05). The modification of AO + PD on the implant neck yielded better attachment between the implant and peri-implant connective tissue. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

OCT Amplitude and Speckle Statistics of Discrete Random Media.

PubMed

Almasian, Mitra; van Leeuwen, Ton G; Faber, Dirk J

2017-11-01

Speckle, amplitude fluctuations in optical coherence tomography (OCT) images, contains information on sub-resolution structural properties of the imaged sample. Speckle statistics could therefore be utilized in the characterization of biological tissues. However, a rigorous theoretical framework relating OCT speckle statistics to structural tissue properties has yet to be developed. As a first step, we present a theoretical description of OCT speckle, relating the OCT amplitude variance to size and organization for samples of discrete random media (DRM). Starting the calculations from the size and organization of the scattering particles, we analytically find expressions for the OCT amplitude mean, amplitude variance, the backscattering coefficient and the scattering coefficient. We assume fully developed speckle and verify the validity of this assumption by experiments on controlled samples of silica microspheres suspended in water. We show that the OCT amplitude variance is sensitive to sub-resolution changes in size and organization of the scattering particles. Experimentally determined and theoretically calculated optical properties are compared and in good agreement.

A new model for ancient DNA decay based on paleogenomic meta-analysis

PubMed Central

Ware, Roselyn; Smith, Oliver; Collins, Matthew

2017-01-01

Abstract The persistence of DNA over archaeological and paleontological timescales in diverse environments has led to a revolutionary body of paleogenomic research, yet the dynamics of DNA degradation are still poorly understood. We analyzed 185 paleogenomic datasets and compared DNA survival with environmental variables and sample ages. We find cytosine deamination follows a conventional thermal age model, but we find no correlation between DNA fragmentation and sample age over the timespans analyzed, even when controlling for environmental variables. We propose a model for ancient DNA decay wherein fragmentation rapidly reaches a threshold, then subsequently slows. The observed loss of DNA over time may be due to a bulk diffusion process in many cases, highlighting the importance of tissues and environments creating effectively closed systems for DNA preservation. This model of DNA degradation is largely based on mammal bone samples due to published genomic dataset availability. Continued refinement to the model to reflect diverse biological systems and tissue types will further improve our understanding of ancient DNA breakdown dynamics. PMID:28486705

MR-guided transcranial brain HIFU in small animal models

PubMed Central

Larrat, Benoît; Pernot, Mathieu; Aubry, Jean-François; Dervishi, Elvis; Sinkus, Ralph; Seilhean, Danielle; Marie, Yannick; Boch, Anne-Laure; Fink, Mathias; Tanter, Mickaël

2010-01-01

Recent studies have demonstrated the feasibility of transcranial High Intensity Focused Ultrasound (HIFU) therapy in the brain using adaptive focusing techniques. However, the complexity of the procedures imposes to provide an accurate targeting, monitoring and control of this emerging therapeutic modality in order to ensure the safety of the treatment and avoid potential damaging effects of ultrasound on healthy tissues. For these purposes, a complete workflow and setup for HIFU treatment under Magnetic Resonance (MR) guidance is proposed and implemented in rats. For the first time, tissue displacements induced by the acoustic radiation force are detected in vivo in brain tissues and measured quantitatively using motion-sensitive MR sequences. Such a valuable target control prior to treatment assesses the quality of the focusing pattern in situ and enables to estimate the acoustic intensity at focus. This MR-Acoustic radiation force imaging is then correlated with conventional MR-Thermometry sequences which are used to follow the temperature changes during the HIFU therapeutic session. Last, pre and post treatment Magnetic Resonance Elastography (MRE) datasets are acquired and evaluated as a new potential way to non invasively control the stiffness changes due to the presence of thermal necrosis. As a proof of concept, MRguided HIFU is performed in vitro in turkey breast samples and in vivo in transcranial rat brain experiments. The experiments are conducted using a dedicated MR compatible HIFU setup in a high field MRI scanner (7T). Results obtained on rats confirmed that both the MR localization of the US focal point and the pre and post HIFU measurement of the tissue stiffness, together with temperature control during HIFU are feasible and valuable techniques for an efficient monitoring of HIFU in the brain. Brain elasticity appears to be more sensitive to the presence of oedema than to tissue necrosis. PMID:20019400

High-Throughput Multi-Analyte Luminex Profiling Implicates Eotaxin-1 in Ulcerative Colitis

PubMed Central

Coburn, Lori A.; Horst, Sara N.; Chaturvedi, Rupesh; Brown, Caroline T.; Allaman, Margaret M.; Scull, Brooks P.; Singh, Kshipra; Piazuelo, M. Blanca; Chitnavis, Maithili V.; Hodges, Mallary E.; Rosen, Michael J.; Williams, Christopher S.; Slaughter, James C.; Beaulieu, Dawn B.; Schwartz, David A.; Wilson, Keith T.

2013-01-01

Accurate and high-throughput technologies are needed for identification of new therapeutic targets and for optimizing therapy in inflammatory bowel disease. Our aim was to assess multi-analyte protein-based assays of cytokines/chemokines using Luminex technology. We have reported that Luminex-based profiling was useful in assessing response to L-arginine therapy in the mouse model of dextran sulfate sodium colitis. Therefore, we studied prospectively collected samples from ulcerative colitis (UC) patients and control subjects. Serum, colon biopsies, and clinical information were obtained from subjects undergoing colonoscopy for evaluation of UC or for non-UC indications. In total, 38 normal controls and 137 UC cases completed the study. Histologic disease severity and the Mayo Disease Activity Index (DAI) were assessed. Serum and colonic tissue cytokine/chemokine profiles were measured by Luminex-based multiplex testing of 42 analytes. Only eotaxin-1 and G-CSF were increased in serum of patients with histologically active UC vs. controls. While 13 cytokines/chemokines were increased in active UC vs. controls in tissues, only eotaxin-1 was increased in all levels of active disease in both serum and tissue. In tissues, eotaxin-1 correlated with the DAI and with eosinophil counts. Increased eotaxin-1 levels were confirmed by real-time PCR. Tissue eotaxin-1 levels were also increased in experimental murine colitis induced by dextran sulfate sodium, oxazolone, or Citrobacter rodentium, but not in murine Helicobacter pylori infection. Our data implicate eotaxin-1 as an etiologic factor and therapeutic target in UC, and indicate that Luminex-based assays may be useful to assess IBD pathogenesis and to select patients for anti-cytokine/chemokine therapies. PMID:24367513

Carprofen pharmacokinetics in plasma and in control and inflamed canine tissue fluid using in vivo ultrafiltration.

PubMed

Messenger, K M; Wofford, J A; Papich, M G

2016-02-01

Measurement of unbound drug concentrations at their sites of action is necessary for accurate PK/PD modeling. The objective of this study was to determine the unbound concentration of carprofen in canine interstitial fluid (ISF) using in vivo ultrafiltration and to compare pharmacokinetic parameters of free carprofen concentrations between inflamed and control tissue sites. We hypothesized that active concentrations of carprofen would exhibit different dispositions in ISF between inflamed vs. normal tissues. Bilateral ultrafiltration probes were placed subcutaneously in six healthy Beagle dogs 12 h prior to induction of inflammation. Two milliliters of either 2% carrageenan or saline control was injected subcutaneously at each probe site, 12 h prior to intravenous carprofen (4 mg/kg) administration. Plasma and ISF samples were collected at regular intervals for 72 h, and carprofen concentrations were determined using HPLC. Prostaglandin E2 (PGE2 ) concentrations were quantified in ISF using ELISA. Unbound carprofen concentrations were higher in ISF compared with predicted unbound plasma drug concentrations. Concentrations were not significantly higher in inflamed ISF compared with control ISF. Compartmental modeling was used to generate pharmacokinetic parameter estimates, which were not significantly different between sites. Terminal half-life (T½) was longer in the ISF compared with plasma. PGE2 in ISF decreased following administration of carprofen. In vivo ultrafiltration is a reliable method to determine unbound carprofen in ISF, and that disposition of unbound drug into tissue is much higher than predicted from unbound drug concentration in plasma. However, concentrations and pharmacokinetic parameter estimates are not significantly different in inflamed vs. un-inflamed tissues. © 2015 John Wiley & Sons Ltd.

Epstein-Barr virus infection and breast invasive ductal carcinoma in Egyptian women: A single center experience.

PubMed

El-Naby, Noha Ed Hassab; Hassan Mohamed, Hameda; Mohamed Goda, Asmaa; El Sayed Mohamed, Ahmed

2017-06-01

A controversy of the role of Epstein-Barr virus (EBV) infection in breast carcinomas has been reported in the literature. We carried on this research to explore possible association between EBV infection and breast invasive ductal carcinoma (IDC) in Egyptian women attending our center. This study carried out at Sohag university hospital on 84 paraffin embedded samples of breast tissue, of them 42 breast IDC as the case group and 42 breast fibroadenomas as the control group. Nested PCRand immunohistochemistry (IHC) done separately for all samples to identify the Epstein-Barr nuclear antigen-1 (EBNA-1) gene and EBV latent membrane protein-1 (LMP-1) respectively, in breast cancer cells and controls. Specimen considered positive when both (EBNA-1) gene and LMP-1 were detected using PCR and IHC separately for the same sample, this was achieved by 10/42 (23.81%) of breast IDC (case group) and 6/42 (14.29%) of breast fibro-adenomas (control group) (P-value=0.4). Nodal involvement was the only parameter that demonstrated a significant statistical relationship with EBV presence in cancerous tissue with p-value=0.003. Our research could not find a significant statistical association between EBV infection and breast IDC in Egyptian women attending our center, but, there might be an association between the existence of EBV and tumor aggressiveness. Copyright © 2017 National Cancer Institute, Cairo University. Production and hosting by Elsevier B.V. All rights reserved.

An empirically derived dietary pattern associated with breast cancer risk is validated in a nested case-control cohort from a randomized primary prevention trial.

PubMed

Hidaka, Brandon H; Kimler, Bruce F; Fabian, Carol J; Carlson, Susan E

2017-02-01

We reported an association between cytologic atypia, a reversible biomarker of breast cancer risk, and lower omega-3/omega-6 fatty acid ratio in blood and breast tissue. Our goal was to develop and validate a dietary pattern index in this high-risk sample of U.S. women, and test its capacity to predict incidence in a nested case-control cohort of Canadian women from a randomized trial of a low-fat dietary intervention for primary prevention of breast cancer. Food intake was measured by food frequency questionnaire in the U.S. sample (n = 65) and multiple dietary recalls in the Canadian sample (n = 220 cases; 440 controls). Principal component analysis identified a dietary pattern associated with atypia. We measured differences among dietary pattern tertiles in (a) fatty acid composition in blood lipids and breast tissue in the U.S. sample, and (b) risk of breast cancer subtypes in the Canadian cohort. Registered under ClinicalTrials.gov Identifier: NCT00148057. A Modern diet was characterized as consuming more grains, dairy, and sugar and less vegetables, fish and poultry; these women had lower tissue omega-3 fatty acids and higher omega-6 and trans fatty acids. The low-fat intervention increased the likelihood of a Modern diet after randomization. A Modern diet at baseline and post-randomization was associated with estrogen-receptor negative (ER-) breast cancer risk among those at least 160 cm tall. A Traditional diet (the reciprocal of Modern) at baseline was associated with lower ER-positive (ER+) risk in the comparison group, but not the low-fat intervention group. A Modern diet (high in grains, dairy, and sugar and low in vegetables, fish, and poultry) is associated with ER- breast cancer risk among taller women. Recommending dietary fat reduction may have untoward effects on breast cancer risk. Copyright © 2016 European Society for Clinical Nutrition and Metabolism. Published by Elsevier Ltd. All rights reserved.

Fluorescence spectroscopy for neoplasms control

NASA Astrophysics Data System (ADS)

Bratchenko, I. A.; Kristoforova, Yu. A.; Myakinin, O. O.; Artemyev, D. N.; Kozlov, S. V.; Moryatov, A. A.; Zakharov, V. P.

2016-04-01

Investigation of malignant skin tumors diagnosis was performed involving two setups for native tissues fluorescence control in visible and near infrared regions. Combined fluorescence analysis for skin malignant melanomas and basal cell carcinomas was performed. Autofluorescence spectra of normal skin and oncological pathologies stimulated by 457 nm and 785 nm lasers were registered for 74 skin tissue samples. Spectra of 10 melanomas and 27 basal cell carcinomas were registered ex vivo. Skin tumors analysis was made on the basis of autofluorescence spectra intensity and curvature for analysis of porphyrins, lipo-pigments, flavins and melanin. Separation of melanomas and basal cell carcinomas was performed on the basis of discriminant analysis. Overall accuracy of basal cell carcinomas and malignant melanomas separation in current study reached 86.5% with 70% sensitivity and 92.6% specificity.

Trends in the development of microfluidic cell biochips for in vitro hepatotoxicity.

PubMed

Baudoin, Régis; Corlu, Anne; Griscom, Laurent; Legallais, Cécile; Leclerc, Eric

2007-06-01

Current developments in the technological fields of liver tissue engineering, bioengineering, biomechanics, microfabrication and microfluidics have lead to highly complex and pertinent new tools called "cell biochips" for in vitro toxicology. The purpose of "cell biochips" is to mimic organ tissues in vitro in order to partially reduce the amount of in vivo testing. These "cell biochips" consist of microchambers containing engineered tissue and living cell cultures interconnected by a microfluidic network, which allows the control of microfluidic flows for dynamic cultures, by continuous feeding of nutrients to cultured cells and waste removal. Cell biochips also allow the control of physiological contact times of diluted molecules with the tissues and cells, for rapid testing of sample preparations or specific addressing. Cell biochips can be situated between in vitro and in vivo testing. These types of systems can enhance functionality of cells by mimicking the tissue architecture complexities when compared to in vitro analysis but at the same time present a more rapid and simple process when compared to in vivo testing procedures. In this paper, we first introduce the concepts of microfluidic and biochip systems based on recent progress in microfabrication techniques used to mimic liver tissue in vitro. This includes progress and understanding in biomaterials science (cell culture substrate), biomechanics (dynamic cultures conditions) and biology (tissue engineering). The development of new "cell biochips" for chronic toxicology analysis of engineered tissues can be achieved through the combination of these research domains. Combining these advanced research domains, we then present "cell biochips" that allow liver chronic toxicity analysis in vitro on engineered tissues. An extension of the "cell biochip" idea has also allowed "organ interactions on chip", which can be considered as a first step towards the replacement of animal testing using a combined liver/lung organ model.

A Microfluidic Interface for the Culture and Sampling of Adiponectin from Primary Adipocytes

PubMed Central

Godwin, Leah A.; Brooks, Jessica C.; Hoepfner, Lauren D.; Wanders, Desiree; Judd, Robert L.; Easley, Christopher J.

2014-01-01

Secreted from adipose tissue, adiponectin is a vital endocrine hormone that acts in glucose metabolism, thereby establishing its crucial role in diabetes, obesity, and other metabolic disease states. Insulin exposure to primary adipocytes cultured in static conditions has been shown to stimulate adiponectin secretion. However, conventional, static methodology for culturing and stimulating adipocytes falls short of truly mimicking physiological environments. Along with decreases in experimental costs and sample volume, and increased temporal resolution, microfluidic platforms permit small-volume flowing cell culture systems, which more accurately represent the constant flow conditions through vasculature in vivo. Here, we have integrated a customized primary tissue culture reservoir into a passively operated microfluidic device made of polydimethylsiloxane (PDMS). Fabrication of the reservoir was accomplished through unique PDMS “landscaping” above sampling channels, with a design strategy targeted to primary adipocytes to overcome issues of positive cell buoyancy. This reservoir allowed three-dimensional culture of primary murine adipocytes, accurate control over stimulants via constant perfusion, and sampling of adipokine secretion during various treatments. As the first report of primary adipocyte culture and sampling within microfluidic systems, this work sets the stage for future studies in adipokine secretion dynamics. PMID:25423362

Whole Transcriptome Sequencing Enables Discovery and Analysis of Viruses in Archived Primary Central Nervous System Lymphomas

PubMed Central

DeBoever, Christopher; Reid, Erin G.; Smith, Erin N.; Wang, Xiaoyun; Dumaop, Wilmar; Harismendy, Olivier; Carson, Dennis; Richman, Douglas; Masliah, Eliezer; Frazer, Kelly A.

2013-01-01

Primary central nervous system lymphomas (PCNSL) have a dramatically increased prevalence among persons living with AIDS and are known to be associated with human Epstein Barr virus (EBV) infection. Previous work suggests that in some cases, co-infection with other viruses may be important for PCNSL pathogenesis. Viral transcription in tumor samples can be measured using next generation transcriptome sequencing. We demonstrate the ability of transcriptome sequencing to identify viruses, characterize viral expression, and identify viral variants by sequencing four archived AIDS-related PCNSL tissue samples and analyzing raw sequencing reads. EBV was detected in all four PCNSL samples and cytomegalovirus (CMV), JC polyomavirus (JCV), and HIV were also discovered, consistent with clinical diagnoses. CMV was found to express three long non-coding RNAs recently reported as expressed during active infection. Single nucleotide variants were observed in each of the viruses observed and three indels were found in CMV. No viruses were found in several control tumor types including 32 diffuse large B-cell lymphoma samples. This study demonstrates the ability of next generation transcriptome sequencing to accurately identify viruses, including DNA viruses, in solid human cancer tissue samples. PMID:24023918