Technique for examining biological materials using diffuse reflectance spectroscopy and the kubelka-munk function

DOEpatents

Alfano, Robert R.; Yang, Yuanlong

2003-09-02

Method and apparatus for examining biological materials using diffuse reflectance spectroscopy and the Kubelka-Munk function. In one aspect, the method is used to determine whether a tissue sample is cancerous or not and comprises the steps of (a) measuring the diffuse reflectance from the tissue sample at a first wavelength and at a second wavelength, wherein the first wavelength is a wavelength selected from the group consisting of 255-265 nm and wherein the second wavelength is a wavelength selected from the group consisting of 275-285 nm; (b) using the Kubelka-Munk function to transform the diffuse reflectance measurement obtained at the first and second wavelengths; and (c) comparing a ratio or a difference of the transformed Kubelka-Munk measurements at the first and second wavelengths to appropriate standards determine whether or not the tissue sample is cancerous. One can use the spectral profile of KMF between 250 nm to 300 nm to determine whether or not the tissue sample is cancerous or precancerous. According to the value at the first and second wavelengths determine whether or not the malignant tissue is invasive or mixed invasive and in situ or carcinoma in situ.

Biobanking of fresh frozen tissue from clinical surgical specimens: transport logistics, sample selection, and histologic characterization.

PubMed

Botling, Johan; Micke, Patrick

2011-01-01

Access to high-quality fresh frozen tissue is critical for translational cancer research and molecular -diagnostics. Here we describe a workflow for the collection of frozen solid tissue samples derived from fresh human patient specimens after surgery. The routines have been in operation at Uppsala University Hospital since 2001. We have integrated cryosection and histopathologic examination of each biobank sample into the biobank manual. In this way, even small, macroscopically ill-defined lesions can be -procured without a diagnostic hazard due to the removal of uncharacterized tissue from a clinical -specimen. Also, knowledge of the histomorphology of the frozen tissue sample - tumor cell content, stromal components, and presence of necrosis - is pivotal before entering a biobank case into costly molecular profiling studies.

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.

Organochlorine contaminant concentrations in multiple tissues of free-ranging Steller sea lions (Eumetopias jubatus) in Alaska.

PubMed

Beckmen, Kimberlee B; Keogh, Mandy J; Burek-Huntington, Kathleen A; Ylitalo, Gina M; Fadely, Brian S; Pitcher, Kenneth W

2016-01-15

The relationships of selected organochlorine (OC) contaminants between blubber, blood, feces, and milk of young, free-ranging Steller sea lions (Eumetopias jubatus) were examined. Both between and within each tissue there was considerable individual variation. In spite of the variation, similar patterns were observed across the tissues for most of the selected PCB congeners. In all four tissues, the major PCB congeners were PCB101, PCB118, PCB138, and PCB153. The most prominent congener, both as a weight (ng/g lipid) and as a percentage of summed PCBs (∑PCBs), was PCB 153. Comparisons between paired tissues showed that ∑DDTs in blubber samples were related to concentrations in blood, feces, and milk. The ∑PCBs in blubber were related to concentrations in milk and fecal samples, though the relationship with feces was weak. Our findings show milk samples, in particular, are useful for assessing OCs in young sea lions. Blubber concentrations of PCB101, PCB118, and PCB138 were an order of magnitude higher than those in milk, supporting the biomagnification of these PCB congeners in SSL tissues. The findings indicate alternative tissues may be used as indicators of relative contaminant exposure in lieu of surgical blubber biopsy. Copyright © 2015 Elsevier B.V. All rights reserved.

CLOSED-LOOP STRIPPING ANALYSIS (CLSA) OF SYNTHETIC MUSK COMPOUNDS FROM FISH TISSUES WITH MEASUREMENT BY GAS CHROMATOGRAPHY-MASS SPECTROMETRY WITH SELECTED-ION MONITORING

EPA Science Inventory

Synthetic musk compounds have been found in surface water, fish tissues, and human breast milk. Current techniques for separating these compounds from fish tissues require tedious sample clean-upprocedures A simple method for the deterrnination of these compounds in fish tissues ...

A mixture model-based approach to the clustering of microarray expression data.

PubMed

McLachlan, G J; Bean, R W; Peel, D

2002-03-01

This paper introduces the software EMMIX-GENE that has been developed for the specific purpose of a model-based approach to the clustering of microarray expression data, in particular, of tissue samples on a very large number of genes. The latter is a nonstandard problem in parametric cluster analysis because the dimension of the feature space (the number of genes) is typically much greater than the number of tissues. A feasible approach is provided by first selecting a subset of the genes relevant for the clustering of the tissue samples by fitting mixtures of t distributions to rank the genes in order of increasing size of the likelihood ratio statistic for the test of one versus two components in the mixture model. The imposition of a threshold on the likelihood ratio statistic used in conjunction with a threshold on the size of a cluster allows the selection of a relevant set of genes. However, even this reduced set of genes will usually be too large for a normal mixture model to be fitted directly to the tissues, and so the use of mixtures of factor analyzers is exploited to reduce effectively the dimension of the feature space of genes. The usefulness of the EMMIX-GENE approach for the clustering of tissue samples is demonstrated on two well-known data sets on colon and leukaemia tissues. For both data sets, relevant subsets of the genes are able to be selected that reveal interesting clusterings of the tissues that are either consistent with the external classification of the tissues or with background and biological knowledge of these sets. EMMIX-GENE is available at http://www.maths.uq.edu.au/~gjm/emmix-gene/

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.

Improved selenium recovery from tissue with modified sample decomposition

USGS Publications Warehouse

Brumbaugh, W. G.; Walther, M.J.

1991-01-01

The present paper describes a simple modification of a recently reported decomposition method for determination of selenium in biological tissue by hydride generation atomic absorption. The modified method yielded slightly higher selenium recoveries (3-4%) for selected reference tissues and fish tissue spiked with selenomethionine. Radiotracer experiments indicated that the addition of a small volume of hydrochloric acid to the wet digestate mixture reduced slight losses of selenium as the sample initially went to dryness before ashing. With the modified method, selenium spiked as selenomethionine behaved more like the selenium in reference tissues than did the inorganic spike forms when this digestion modification was used.

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.

Wavelet analysis in two-dimensional tomography

NASA Astrophysics Data System (ADS)

Burkovets, Dimitry N.

2002-02-01

The diagnostic possibilities of wavelet-analysis of coherent images of connective tissue in its pathological changes diagnostics. The effectiveness of polarization selection in obtaining wavelet-coefficients' images is also shown. The wavelet structures, characterizing the process of skin psoriasis, bone-tissue osteoporosis have been analyzed. The histological sections of physiological normal and pathologically changed samples of connective tissue of human skin and spongy bone tissue have been analyzed.

Measurement of stable isotopic enrichment and concentration of long-chain fatty acyl-carnitines in tissue by HPLC-MS.

PubMed

Sun, Dayong; Cree, Melanie G; Zhang, Xiao-Jun; Bøersheim, Elisabet; Wolfe, Robert R

2006-02-01

We have developed a new method for the simultaneous measurements of stable isotopic tracer enrichments and concentrations of individual long-chain fatty acyl-carnitines in muscle tissue using ion-pairing high-performance liquid chromatography-electrospray ionization quadrupole mass spectrometry in the selected ion monitoring (SIM) mode. Long-chain fatty acyl-carnitines were extracted from frozen muscle tissue samples by acetonitrile/methanol. Baseline separation was achieved by reverse-phase HPLC in the presence of the volatile ion-pairing reagent heptafluorobutyric acid. The SIM capability of a single quadrupole mass analyzer allows further separation of the ions of interest from the sample matrixes, providing very clean total and selected ion chromatograms that can be used to calculate the stable isotopic tracer enrichment and concentration of long-chain fatty acyl-carnitines in a single analysis. The combination of these two separation techniques greatly simplifies the sample preparation procedure and increases the detection sensitivity. Applying this protocol to biological muscle samples proves it to be a very sensitive, accurate, and precise analytical tool.

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.

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.

Sample Preparation for Mass Spectrometry Imaging of Plant Tissues: A Review

PubMed Central

Dong, Yonghui; Li, Bin; Malitsky, Sergey; Rogachev, Ilana; Aharoni, Asaph; Kaftan, Filip; Svatoš, Aleš; Franceschi, Pietro

2016-01-01

Mass spectrometry imaging (MSI) is a mass spectrometry based molecular ion imaging technique. It provides the means for ascertaining the spatial distribution of a large variety of analytes directly on tissue sample surfaces without any labeling or staining agents. These advantages make it an attractive molecular histology tool in medical, pharmaceutical, and biological research. Likewise, MSI has started gaining popularity in plant sciences; yet, information regarding sample preparation methods for plant tissues is still limited. Sample preparation is a crucial step that is directly associated with the quality and authenticity of the imaging results, it therefore demands in-depth studies based on the characteristics of plant samples. In this review, a sample preparation pipeline is discussed in detail and illustrated through selected practical examples. In particular, special concerns regarding sample preparation for plant imaging are critically evaluated. Finally, the applications of MSI techniques in plants are reviewed according to different classes of plant metabolites. PMID:26904042

Large-scale atlas of microarray data reveals the distinct expression landscape of different tissues in Arabidopsis

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

He, Fei; Maslov, Sergei; Yoo, Shinjae

Here, transcriptome datasets from thousands of samples of the model plant Arabidopsis thaliana have been collectively generated by multiple individual labs. Although integration and meta-analysis of these samples has become routine in the plant research community, it is often hampered by the lack of metadata or differences in annotation styles by different labs. In this study, we carefully selected and integrated 6,057 Arabidopsis microarray expression samples from 304 experiments deposited to NCBI GEO. Metadata such as tissue type, growth condition, and developmental stage were manually curated for each sample. We then studied global expression landscape of the integrated dataset andmore » found that samples of the same tissue tend to be more similar to each other than to samples of other tissues, even in different growth conditions or developmental stages. Root has the most distinct transcriptome compared to aerial tissues, but the transcriptome of cultured root is more similar to those of aerial tissues as the former samples lost their cellular identity. Using a simple computational classification method, we showed that the tissue type of a sample can be successfully predicted based on its expression profile, opening the door for automatic metadata extraction and facilitating re-use of plant transcriptome data. As a proof of principle we applied our automated annotation pipeline to 708 RNA-seq samples from public repositories and verified accuracy of our predictions with samples’ metadata provided by authors.« less

Large-scale atlas of microarray data reveals the distinct expression landscape of different tissues in Arabidopsis

DOE PAGES

He, Fei; Maslov, Sergei; Yoo, Shinjae; ...

2016-05-25

Here, transcriptome datasets from thousands of samples of the model plant Arabidopsis thaliana have been collectively generated by multiple individual labs. Although integration and meta-analysis of these samples has become routine in the plant research community, it is often hampered by the lack of metadata or differences in annotation styles by different labs. In this study, we carefully selected and integrated 6,057 Arabidopsis microarray expression samples from 304 experiments deposited to NCBI GEO. Metadata such as tissue type, growth condition, and developmental stage were manually curated for each sample. We then studied global expression landscape of the integrated dataset andmore » found that samples of the same tissue tend to be more similar to each other than to samples of other tissues, even in different growth conditions or developmental stages. Root has the most distinct transcriptome compared to aerial tissues, but the transcriptome of cultured root is more similar to those of aerial tissues as the former samples lost their cellular identity. Using a simple computational classification method, we showed that the tissue type of a sample can be successfully predicted based on its expression profile, opening the door for automatic metadata extraction and facilitating re-use of plant transcriptome data. As a proof of principle we applied our automated annotation pipeline to 708 RNA-seq samples from public repositories and verified accuracy of our predictions with samples’ metadata provided by authors.« 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...

Assessment of heavy metals in seawater and fish tissues at Pulau Indah, Selangor, Malaysia

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

Md Yunus, Sabarina, E-mail: sabarina2020@salam.uitm.edu.m; Hamzah, Zaini; Wood, Ab. Khalik

This study focuses on the levels of heavy metals in seawater and selected fish tissue at Pulau Indah, Selangor, Malaysia. Pulau Indah primarily contains Westport (Malaysia’s major port), and a host of full scales factories. Therefore, it is important to monitor the levels of pollution in this water body and the aquatic organisms in the long term effects, due to the human activities in this area. Water samples in this study were taken from 8 locations along the coastal area. The water samples were collected using water sampler and sampling locations were determined using a Global Positioning system (GPS). Similarly,more » in situ water quality parameters including temperature, dissolve oxygen (DO), salinity, total suspended solid (TSS), pH and turbidity were measured by using portable multi probes meter. Then, the samples were acidified until pH 2 and filtered. Fish samples were purchased from local fisherman along the Pulau Indah coastal area and samples were digested using concentrated nitric acid in wet digestion method. The levels of selected heavy metals in four species of fish and seawater from Pulau Indah coastal area were determined using Inductive Coupled Plasma Mass Spectrometer (ICPMS) after dilution to the samples. In general, the quality of water at Pulau Indah is lower than Malaysia Marine Water Quality Standard except for few locations were higher than the maximum permissible levels. The concentration of heavy metals which are lead (Pb), copper (Cu), zinc (Zn) and uranium (U) in water samples are in the range of 0.36-5.43 µg/L, 75.20-621.58 µg/L, 11.92-30.52 µg/L, and 4.00-4.65 µg/L respectively. While the results of the four selected fish showed the following order of abundance Zn> U> Cu> Pb. Transfer factor (TF) of heavy metals in fish tissue of selected fish species from the water was discussed. However, the observed metal concentration in the sample tissue did not exceed the allowable limit of Malaysian Food Act (1983) and Regulation (1985). Therefore, it is safe for human consumption. Moreover, the pollution levels of these heavy metals were also compared with other studies. This present study can also be used to evaluate the safety dose uptake level of marine biota as well as to monitor environmental health.« less

Assessment of heavy metals in seawater and fish tissues at Pulau Indah, Selangor, Malaysia

NASA Astrophysics Data System (ADS)

Md Yunus, Sabarina; Hamzah, Zaini; Wood, Ab. Khalik; Ahmad

2015-04-01

This study focuses on the levels of heavy metals in seawater and selected fish tissue at Pulau Indah, Selangor, Malaysia. Pulau Indah primarily contains Westport (Malaysia's major port), and a host of full scales factories. Therefore, it is important to monitor the levels of pollution in this water body and the aquatic organisms in the long term effects, due to the human activities in this area. Water samples in this study were taken from 8 locations along the coastal area. The water samples were collected using water sampler and sampling locations were determined using a Global Positioning system (GPS). Similarly, in situ water quality parameters including temperature, dissolve oxygen (DO), salinity, total suspended solid (TSS), pH and turbidity were measured by using portable multi probes meter. Then, the samples were acidified until pH 2 and filtered. Fish samples were purchased from local fisherman along the Pulau Indah coastal area and samples were digested using concentrated nitric acid in wet digestion method. The levels of selected heavy metals in four species of fish and seawater from Pulau Indah coastal area were determined using Inductive Coupled Plasma Mass Spectrometer (ICPMS) after dilution to the samples. In general, the quality of water at Pulau Indah is lower than Malaysia Marine Water Quality Standard except for few locations were higher than the maximum permissible levels. The concentration of heavy metals which are lead (Pb), copper (Cu), zinc (Zn) and uranium (U) in water samples are in the range of 0.36-5.43 µg/L, 75.20-621.58 µg/L, 11.92-30.52 µg/L, and 4.00-4.65 µg/L respectively. While the results of the four selected fish showed the following order of abundance Zn> U> Cu> Pb. Transfer factor (TF) of heavy metals in fish tissue of selected fish species from the water was discussed. However, the observed metal concentration in the sample tissue did not exceed the allowable limit of Malaysian Food Act (1983) and Regulation (1985). Therefore, it is safe for human consumption. Moreover, the pollution levels of these heavy metals were also compared with other studies. This present study can also be used to evaluate the safety dose uptake level of marine biota as well as to monitor environmental health.

Histological Evaluation of Selected Organs of the Eurasian Beavers (Castor fiber) Inhabiting Poland.

PubMed

Dolka, I; Giżejewska, A; Giżejewski, Z; Kluciński, W; Kołodziejska, J

2015-10-01

There is a general scarcity of data on the histological structure of major organs in the Eurasian beaver (Castor fiber). This study presents the histological characteristics of beaver organs such as the liver, spleen, cardiac muscle, lungs and kidneys. Tissue samples were collected from 21 beavers and analysed. Selected samples of tail tissue were additionally examined. Tissue samples were placed in neutral buffered formalin and embedded in paraffin. 4-μm-thick sections were stained with haematoxylin and eosin, and other staining techniques were also used. Scant amounts of inter-lobular connective tissue were found in the liver. Ion or copper deposition was not observed, but scattered cytoplasmic glycogen deposits were present in hepatocytes. Our results suggest that beavers have defensive rather than storage spleens. Interestingly, the presence of melanin in splenic red pulp was noted. The histological structure of the examined organs closely resembled that of other rodent species. According to our knowledge, this is the first study evaluating the histological structure of beaver organs. Nevertheless, precise characterization of the evaluated organs requires further work with the involvement of accurate and reliable techniques, such as molecular biology or electron microscopy methods. © 2014 Blackwell Verlag GmbH.

Typeability of PowerPlex Y (Promega) profiles in selected tissue samples incubated in various environments.

PubMed

Niemcunowicz-Janica, Anna; Pepiński, Witold; Janica, Jacek Robert; Janica, Jerzy; Skawrońska, Małgorzata; Koc-Zórawska, Ewa

2007-01-01

In cases of decomposed bodies, Y chromosomal STR markers may be useful in identification of a male relative. The authors assessed typeability of PowerPlex Y (Promega) loci in post mortem tissue material stored in various environments. Kidney, spleen and pancreas specimens were collected during autopsies of five persons aged 20-30 years, whose time of death was determined within the limit of 14 hours. Tissue material was incubated at 21 degrees C and 4 degrees C in various environmental conditions. DNA was extracted by the organic method from tissue samples collected in 7-day intervals and subsequently typed using the PowerPlexY-STR kit and ABI 310. A fast decrease in the typeability rate was seen in specimens incubated in peat soil and in sand. Kidney tissue samples were typeable in all PowerPlexY-STR loci within 63 days of incubation at 4 degrees C. Faster DNA degradation was recorded in spleen and pancreas specimens. In samples with negative genotyping results, no DNA was found by fluorometric quantitation. Decomposed soft tissues are a potential material for DNA typing.

Assessment of Sample Preparation Bias in Mass Spectrometry-Based Proteomics.

PubMed

Klont, Frank; Bras, Linda; Wolters, Justina C; Ongay, Sara; Bischoff, Rainer; Halmos, Gyorgy B; Horvatovich, Péter

2018-04-17

For mass spectrometry-based proteomics, the selected sample preparation strategy is a key determinant for information that will be obtained. However, the corresponding selection is often not based on a fit-for-purpose evaluation. Here we report a comparison of in-gel (IGD), in-solution (ISD), on-filter (OFD), and on-pellet digestion (OPD) workflows on the basis of targeted (QconCAT-multiple reaction monitoring (MRM) method for mitochondrial proteins) and discovery proteomics (data-dependent acquisition, DDA) analyses using three different human head and neck tissues (i.e., nasal polyps, parotid gland, and palatine tonsils). Our study reveals differences between the sample preparation methods, for example, with respect to protein and peptide losses, quantification variability, protocol-induced methionine oxidation, and asparagine/glutamine deamidation as well as identification of cysteine-containing peptides. However, none of the methods performed best for all types of tissues, which argues against the existence of a universal sample preparation method for proteome analysis.

Concentrations of selected organochlorine compounds in fish tissue in the Mississippi Embayment Study Unit : Arkansas, Kentucky, Louisiana, Mississippi, Missouri, and Tennessee, 1995-99

USGS Publications Warehouse

Femmer, Suzanne R.; Coupe, Richard H.; Justus, B.G.; Kleiss, Barbara A.

2004-01-01

Whole fish were collected at 52 sites during 1995-99 to evaluate the occurrence and distribution of selected organochlorine compounds in the Mississippi Embayment Study Unit. Samples were collected as part of the U.S. Geological Survey National Water Quality Assessment Program. From 5 to 8 fish were collected at each site; the fish were composited, and an aliquot of the tissue was analyzed for 28 organo-chlorine compounds, which included pesticides, pesticide degradates, and polychlorinated biphenyls. The use of these organochlorine compounds has been discontinued or severely restricted within the United States, but the continued detection of these compounds or their degradates in the air, water, soil, and biota in national surveys, coupled with known environmental problems associated with these compounds (such as a long half-life and the propensity to accumulate in living tissue), is cause for continued interest in their environmental fate. At least one organochlorine compound was detected in every fish-tissue sample, and as many as 15 different compounds were detected in some. The most frequently detected com-pounds were the degradates of p,p'-dichlorodiphenyltrichlo-roethane (p,p'-DDT); p,p'-dichlorodiphenyldichloroethylene (p,p'-DDE) was detected in every sample above the method reporting limit, and p,p'-dichlorodiphenyldichloroethane (p,p'-DDD), was detected in 94 percent of the samples. Polychlorinated biphenyl compounds and dieldrin were detected in 83 and 78 percent of fish-tissue samples, respectively. Because these were whole fish samples, the results are not directly comparable to human health standards, which are based on fish fillets. Comparison of these results, however, to the guidelines for the protection of fish-eating wildlife indicates that concentrations of the p,p'-DDT degradates and toxaphene continue to be of environmental concern.

Spectral analysis of tissues from patients with cancer using a portable spectroscopic diagnostic ratiometer unit

NASA Astrophysics Data System (ADS)

Sordillo, Laura A.; Pu, Yang; Sordillo, Peter P.; Budansky, Yury; Alfano, R. R.

2014-05-01

Spectral profiles of tissues from patients with breast carcinoma, malignant carcinoid and non-small cell lung carcinoma were acquired using native fluorescence spectroscopy. A novel spectroscopic ratiometer device (S3-LED) with selective excitation wavelengths at 280 nm and 335 nm was used to produce the emission spectra of the key biomolecules, tryptophan and NADH, in the tissue samples. In each of the samples, analysis of emission intensity peaks from biomolecules showed increased 340 nm/440 nm and 340 nm/460 nm ratios in the malignant samples compared to their paired normal samples. This most likely represented increased tryptophan to NADH ratios in the malignant tissue samples compared to their paired normal samples. Among the non-small cell lung carcinoma and breast carcinomas, it appeared that tumors of very large size or poor differentiation had an even greater increase in the 340 nm/440 nm and 340 nm/460 nm ratios. In the samples of malignant carcinoid, which is known to be a highly metabolically active tumor, a marked increase in these ratios was also seen.

Rapid Column-Free Enrichment of Mononuclear Cells from Solid Tissues

PubMed Central

Scoville, Steven D.; Keller, Karen A.; Cheng, Stephanie; Zhang, Michael; Zhang, Xiaoli; Caligiuri, Michael A.; Freud, Aharon G.

2015-01-01

We have developed a rapid negative selection method to enrich rare mononuclear cells from human tissues. Unwanted and antibody-tethered cells are selectively depleted during a Ficoll separation step, and there is no need for magnetic-based reagents and equipment. The new method is fast, customizable, inexpensive, remarkably efficient, and easy to perform, and per sample the overall cost is less than one-tenth the cost associated with a magnetic column-based method. PMID:26223896

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

Wolfe, M.; Norman, D.

Birds and mammals exposed to waterborne mercury (Hg) and methylmercury (MeHg) were collected and/or sampled at Clear Lake, California, USA, to field test the predictive wildlife criteria model developed for the Great Lakes Water Quality Initiative (GLWQI). Tissue samples collected from sampled animals were analyzed for Hg and organochlorine residues, and for selected physiologic parameters known to be affected by Hg. All mammalian organ tissues analyzed contained less than 12 ppm total Hg, wet weight. All avian tissue samples analyzed contained less than 3 ppm total Hg, wet weight. No evidence of Hg-associated health effects was found. Tissue Hg residuesmore » were compared with water, sediment, and animal food samples to characterize bioaccumulation of mercury in the Clear Lake food web. Total Hg bioaccumulation factors for the Clear Lake site closest to the Hg source were: TL-2: 11,100; TL-3: 31,200; TL-4, 190,000. The results support the final wildlife criterion and suggest that the GLWQI model, with site-specific modifications, is predictive for other Hg-bearing aquatic systems.« less

Fit for purpose frozen tissue collections by RNA integrity number-based quality control assurance at the Erasmus MC tissue bank.

PubMed

Kap, Marcel; Oomen, Monique; Arshad, Shazia; de Jong, Bas; Riegman, Peter

2014-04-01

About 5000 frozen tissue samples are collected each year by the Erasmus Medical Center tissue bank. Two percent of these samples are randomly selected annually for RNA isolation and RNA Integrity Number (RIN) measurement. A similar quality assessment was conducted during centralization of a 20-year-old tissue collection from the cancer institute, a 15-year-old liver sample archive (-80°C), and a 13-year-old clinical pathology frozen biopsy archive (Liquid Nitrogen). Samples were divided into either high-quality (RIN ≥6.5) or low-quality overall categories, or into four "fit-for-purpose" quality groups: RIN <5: not reliable for demanding downstream analysis; 5 ≤RIN <6: suitable for RT-qPCR; 6 ≤RIN <8: suitable for gene array analysis; and RIN ≥8: suitable for all downstream techniques. In general, low RIN values were correlated with fatty, fibrous, pancreatic, or necrotic tissue. When the percentage of samples with RIN ≥6.5 is higher than 90%, the tissue bank performance is adequate. The annual 2011 quality control assessment showed that 90.3% (n=93) of all samples had acceptable RIN values; 97.4% (n=39) of the cancer institute collection had RIN values above 6.5; and 88.6% (n=123) of samples from the liver sample archive collection had RIN values higher than 6.5. As the clinical pathology biopsy collection contained only 58.8% (n=24) acceptable samples, the procurement protocols used for these samples needed immediate evaluation. When the distribution of RIN values of the different collections were compared, no significant differences were found, despite differences in average storage time and temperature. According to the principle of "fit-for-purpose" distribution, the vast majority of samples are considered good enough for most downstream techniques. In conclusion, an annual tissue bank quality control procedure provides useful information on tissue sample quality and sheds light on where and if improvements need to be made.

Method development and subsequent survey analysis of biological tissues for platinum, lead, and manganese content.

PubMed Central

Yoakum, A M; Stewart, P L; Sterrett, J E

1975-01-01

An emission spectrochemical method is described for the determination of trace quantities of platinum, lead, and manganese in biological tissues. Total energy burns in an argon-oxygen atmosphere are employed. Sample preparation, conditions of analysis, and preparation of standards are discussed. The precision of the method is consistently better than +/- 15%, and comparative analyses indicate comparable accuracies. Data obtained for experimental rat tissues and for selected autopsy tissues are presented. PMID:1157798

Single-molecule RNA detection at depth by hybridization chain reaction and tissue hydrogel embedding and clearing.

PubMed

Shah, Sheel; Lubeck, Eric; Schwarzkopf, Maayan; He, Ting-Fang; Greenbaum, Alon; Sohn, Chang Ho; Lignell, Antti; Choi, Harry M T; Gradinaru, Viviana; Pierce, Niles A; Cai, Long

2016-08-01

Accurate and robust detection of mRNA molecules in thick tissue samples can reveal gene expression patterns in single cells within their native environment. Preserving spatial relationships while accessing the transcriptome of selected cells is a crucial feature for advancing many biological areas - from developmental biology to neuroscience. However, because of the high autofluorescence background of many tissue samples, it is difficult to detect single-molecule fluorescence in situ hybridization (smFISH) signals robustly in opaque thick samples. Here, we draw on principles from the emerging discipline of dynamic nucleic acid nanotechnology to develop a robust method for multi-color, multi-RNA imaging in deep tissues using single-molecule hybridization chain reaction (smHCR). Using this approach, single transcripts can be imaged using epifluorescence, confocal or selective plane illumination microscopy (SPIM) depending on the imaging depth required. We show that smHCR has high sensitivity in detecting mRNAs in cell culture and whole-mount zebrafish embryos, and that combined with SPIM and PACT (passive CLARITY technique) tissue hydrogel embedding and clearing, smHCR can detect single mRNAs deep within thick (0.5 mm) brain slices. By simultaneously achieving ∼20-fold signal amplification and diffraction-limited spatial resolution, smHCR offers a robust and versatile approach for detecting single mRNAs in situ, including in thick tissues where high background undermines the performance of unamplified smFISH. © 2016. Published by The Company of Biologists Ltd.

DDT contamination in selected estuarine and coastal marine finfish and shellfish of New Jersey.

PubMed

Kennish, M J; Ruppel, B E

1996-08-01

Analysis of DDT contamination in selected finfish and shellfish species from estuarine and coastal marine waters of New Jersey (USA) reveals consistently highest organochlorine pesticide levels in samples from the north and northeast regions of the state. The mean concentrations of DDT and its metabolites, DDE and DDD, in bluefish, striped bass, weakfish, and blue crabs collected at 27 stations throughout the state between 1988 and 1991 ranged from <25 to >300 microg/kg wet weight (wet wt). Gas chromatographic analysis of 175 tissue samples from these four species showed that the highest levels of DDTs (DDT plus DDE and DDD), exceeding 300 microg/kg wet wt, occurred in blue crabs (hepatopancreas) from the Hudson-Newark-Raritan Bay complex in the northeast region. Lower mean concentrations of DDTs (<200 microg/kg wet wt) were recorded in tissue samples of these species from all other regions of the state. The lowest levels of DDT contamination (mean <110 microg/kg wet wt) were found in samples from the south coast region. The greatest impact of DDT contamination is nearby metropolitan areas of the state, although the total concentrations of DDT in tissue samples from these areas are far less than the U.S. Food and Drug Administration action level of 5,000 microg/kg wet wt for this contaminant.

Spatial cluster analysis of nanoscopically mapped serotonin receptors for classification of fixed brain tissue

NASA Astrophysics Data System (ADS)

Sams, Michael; Silye, Rene; Göhring, Janett; Muresan, Leila; Schilcher, Kurt; Jacak, Jaroslaw

2014-01-01

We present a cluster spatial analysis method using nanoscopic dSTORM images to determine changes in protein cluster distributions within brain tissue. Such methods are suitable to investigate human brain tissue and will help to achieve a deeper understanding of brain disease along with aiding drug development. Human brain tissue samples are usually treated postmortem via standard fixation protocols, which are established in clinical laboratories. Therefore, our localization microscopy-based method was adapted to characterize protein density and protein cluster localization in samples fixed using different protocols followed by common fluorescent immunohistochemistry techniques. The localization microscopy allows nanoscopic mapping of serotonin 5-HT1A receptor groups within a two-dimensional image of a brain tissue slice. These nanoscopically mapped proteins can be confined to clusters by applying the proposed statistical spatial analysis. Selected features of such clusters were subsequently used to characterize and classify the tissue. Samples were obtained from different types of patients, fixed with different preparation methods, and finally stored in a human tissue bank. To verify the proposed method, samples of a cryopreserved healthy brain have been compared with epitope-retrieved and paraffin-fixed tissues. Furthermore, samples of healthy brain tissues were compared with data obtained from patients suffering from mental illnesses (e.g., major depressive disorder). Our work demonstrates the applicability of localization microscopy and image analysis methods for comparison and classification of human brain tissues at a nanoscopic level. Furthermore, the presented workflow marks a unique technological advance in the characterization of protein distributions in brain tissue sections.

Deep UV autofluorescence microscopy for cell biology and tissue histology.

PubMed

Jamme, Frédéric; Kascakova, Slavka; Villette, Sandrine; Allouche, Fatma; Pallu, Stéphane; Rouam, Valérie; Réfrégiers, Matthieu

2013-07-01

Autofluorescence spectroscopy is a powerful tool for molecular histology and for following metabolic processes in biological samples as it does not require labelling. However, at the microscopic scale, it is mostly limited to visible and near infrared excitation of the samples. Several interesting and naturally occurring fluorophores can be excited in the UV and deep UV (DUV), but cannot be monitored in cellulo nor in vivo due to a lack of available microscopic instruments working in this wavelength range. To fulfil this need, we have developed a synchrotron-coupled DUV microspectrofluorimeter which is operational since 2010. An extended selection of endogenous autofluorescent probes that can be excited in DUV, including their spectral characteristics, is presented. The distribution of the probes in various biological samples, including cultured cells, soft tissues, bone sections and maize stems, is shown to illustrate the possibilities offered by this system. In this work we demonstrate that DUV autofluorescence is a powerful tool for tissue histology and cell biology. To fulfil this need, we have developed a synchrotron-coupled DUV microspectrofluorimeter which is operational since 2010. An extended selection of endogenous autofluorescent probes that can be excited in DUV, including their spectral characteristics, is presented. The distribution of the probes in various biological samples, including cultured cells, soft tissues, bone sections and maize stems, is shown to illustrate the possibilities offered by this system. In this work we demonstrate that DUV autofluorescence is a powerful tool for tissue histology and cell biology. In this work we demonstrate that DUV autofluorescence is a powerful tool for tissue histology and cell biology. © 2013 Société Française des Microscopies and Société de Biologie Cellulaire de France. Published by John Wiley & Sons Ltd.

Presence of archaea and selected bacteria in infected root canal systems.

PubMed

Brzezińska-Błaszczyk, Ewa; Pawłowska, Elżbieta; Płoszaj, Tomasz; Witas, Henryk; Godzik, Urszula; Agier, Justyna

2018-05-01

Infections of the root canal have polymicrobial etiology. The main group of microflora in the infected pulp is bacteria. There is limited data that archaea may be present in infected pulp tissue. The aim of this study was to check the prevalence of archaea in necrotic root canal samples obtained from patients with primary or post-treatment infection. The prevalence of selected bacteria species (Prevotella intermedia, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, Synergistes sp.) in necrotic samples was evaluated as well. Sixty-four samples from root canal were collected for DNA and RNA extraction. A PCR assay based on the 16S rRNA gene was used to determine the presence of archaea and selected bacteria. Of the 64 samples, 6 were analyzed by semiquantitative reverse transcription PCR to estimate expression profiles of 16S rRNA, and another 9 were selected for direct sequencing. Archaea were detected in 48.4% samples. Statistical analysis indicated a negative association in coexistence between archaea and Treponema denticola (P < 0.05; Pearson's χ 2 test). The main representative of the Archaea domain found in infected pulp tissue was Methanobrevibacter oralis. Archaea 16S rRNA gene expression was significantly lower than Synergistes sp., Porphyromonas gingivalis, and Tannerella forsythia (P < 0.05; Student's t test). Thus, it can be hypothesized that archaea may participate in the endodontic microbial community.

Monitoring of Au(iii) species in plants using a selective fluorescent probe.

PubMed

Li, Zhen; Xu, Yuqing; Fu, Jie; Zhu, Hailiang; Qian, Yong

2018-01-23

A colorimetric and ratiometric probe with a push-pull chromophore dicyanoisophorone system, AuP, has been developed for the detection of Au(iii) species with highly sensitive and selective response to real-water samples and living tissues of Arabidopsis thaliana.

Organic compounds and trace elements in fish tissue and bed sediment from streams in the Yellowstone River basin, Montana and Wyoming, 1998

USGS Publications Warehouse

Peterson, David A.; Boughton, Gregory K.

2000-01-01

A comprehensive water-quality investigation of the Yellowstone River Basin began in 1997, under the National Water-Quality Assessment (NAWQA) Program. Twenty-four sampling sites were selected for sampling of fish tissue and bed sediment during 1998. Organic compounds analyzed included organochlorine insecticides and their metabolites and total polychlorinated biphenyls (PCBs) from fish-tissue and bed-sediment samples, and semivolatile organic compounds from bed-sediment samples. A broad suite of trace elements was analyzed from both fish-tissue and bed-sediment samples, and a special study related to mercury also was conducted. Of the 12 organochlorine insecticides and metabolites detected in the fish-tissue samples, the most compounds per site were detected in samples from integrator sites which represent a mixture of land uses. The presence of DDT, and its metabolites DDD and DDE, in fish collected in the Yellowstone Park area likely reflects long-term residual effects from historical DDT-spraying programs for spruce budworm. Dieldrin, chlordane, and other organic compounds also were detected in the fish-tissue samples. The compound p, p'-DDE was detected at 71 percent of the sampling sites, more than any other compound. The concentrations of total DDT in fish samples were low, however, compared to concentrations from historical data from the study area, other NAWQA studies in the Rocky Mountains, and national baseline concentrations. Only 2 of the 27 organochlorine insecticides and metabolites and total PCBs analyzed in bed sediment were detected. Given that 12 of the compounds were detected in fish-tissue samples, fish appeared to be more sensitive indicators of contamination than bed sediment.Concentrations of some trace elements in fish and bed sediment were higher at sites in mineralized areas than at other sites. Concentrations of selenium in fish tissue from some sites were above background levels. Concentrations of arsenic, chromium, copper, and lead in some of the bed-sediment samples potentially exceeded criteria for the protection of aquatic life.

Solid tissue culture for cytogenetic analysis: a collaborative survey for the Association of Clinical Cytogeneticists.

PubMed Central

Rodgers, C S; Creasy, M R; Fitchett, M; Maliszewska, C T; Pratt, N R; Waters, J J

1996-01-01

AIMS: To survey the diagnostic service provided by UK laboratories for the culture of solid tissue samples (excluding tumours) and in particular to examine the variation in culture success rates and the problems of maternal cell overgrowth. METHODS: Twenty seven laboratories took part in a collaborative survey during 1992. Each laboratory submitted data on up to a maximum of 60 consecutive specimens (n = 1361) over a six month period. RESULTS: Skin specimens, the largest category received (n = 520), were the most problematic (51% success rate). Culture success rates were significantly lower (43%) when skin specimens (n = 140) were transported dry to the laboratory. Success rates for skin specimens also varied, depending on the origin of the specimen, from 18% for intra-uterine deaths (IUD) (n = 94) to 85% for neonatal deaths (n = 33) and 83% for live patients (n = 54). Culture of selected extra-fetal tissues from IUD, stillbirths and following elective termination of pregnancy (TOP) gave comparable success rates to those achieved for skin samples from neonatal deaths and live births. Skewed sex ratios, female > male, were identified for products of conception (POC) (n = 298) and placental biopsy specimens (n = 97). CONCLUSIONS: By appropriate selection, transport and processing of tissues, and in particular by avoiding relying solely on skin samples from IUD, stillbirths and TOP, an increase in culture success rates for solid tissue samples submitted for cytogenetic analysis could be achieved. The high risk of maternal cell contamination from POC and placental biopsy specimens was also identified in this survey. PMID:8881913

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.

Pattern Genes Suggest Functional Connectivity of Organs

NASA Astrophysics Data System (ADS)

Qin, Yangmei; Pan, Jianbo; Cai, Meichun; Yao, Lixia; Ji, Zhiliang

2016-05-01

Human organ, as the basic structural and functional unit in human body, is made of a large community of different cell types that organically bound together. Each organ usually exerts highly specified physiological function; while several related organs work smartly together to perform complicated body functions. In this study, we present a computational effort to understand the roles of genes in building functional connection between organs. More specifically, we mined multiple transcriptome datasets sampled from 36 human organs and tissues, and quantitatively identified 3,149 genes whose expressions showed consensus modularly patterns: specific to one organ/tissue, selectively expressed in several functionally related tissues and ubiquitously expressed. These pattern genes imply intrinsic connections between organs. According to the expression abundance of the 766 selective genes, we consistently cluster the 36 human organs/tissues into seven functional groups: adipose & gland, brain, muscle, immune, metabolism, mucoid and nerve conduction. The organs and tissues in each group either work together to form organ systems or coordinate to perform particular body functions. The particular roles of specific genes and selective genes suggest that they could not only be used to mechanistically explore organ functions, but also be designed for selective biomarkers and therapeutic targets.

Structure-Preserving Color Normalization and Sparse Stain Separation for Histological Images.

PubMed

Vahadane, Abhishek; Peng, Tingying; Sethi, Amit; Albarqouni, Shadi; Wang, Lichao; Baust, Maximilian; Steiger, Katja; Schlitter, Anna Melissa; Esposito, Irene; Navab, Nassir

2016-08-01

Staining and scanning of tissue samples for microscopic examination is fraught with undesirable color variations arising from differences in raw materials and manufacturing techniques of stain vendors, staining protocols of labs, and color responses of digital scanners. When comparing tissue samples, color normalization and stain separation of the tissue images can be helpful for both pathologists and software. Techniques that are used for natural images fail to utilize structural properties of stained tissue samples and produce undesirable color distortions. The stain concentration cannot be negative. Tissue samples are stained with only a few stains and most tissue regions are characterized by at most one effective stain. We model these physical phenomena that define the tissue structure by first decomposing images in an unsupervised manner into stain density maps that are sparse and non-negative. For a given image, we combine its stain density maps with stain color basis of a pathologist-preferred target image, thus altering only its color while preserving its structure described by the maps. Stain density correlation with ground truth and preference by pathologists were higher for images normalized using our method when compared to other alternatives. We also propose a computationally faster extension of this technique for large whole-slide images that selects an appropriate patch sample instead of using the entire image to compute the stain color basis.

Relationship of In Vivo MR Parameters to Histopathological and Molecular Characteristics of Newly Diagnosed, Nonenhancing Lower-Grade Gliomas.

PubMed

Luks, Tracy L; McKnight, Tracy Richmond; Jalbert, Llewellyn E; Williams, Aurelia; Neill, Evan; Lobo, Khadjia A; Persson, Anders I; Perry, Arie; Phillips, Joanna J; Molinaro, Annette M; Chang, Susan M; Nelson, Sarah J

2018-06-05

The goal of this research was to elucidate the relationship between WHO 2016 molecular classifications of newly diagnosed, nonenhancing lower grade gliomas (LrGG), tissue sample histopathology, and magnetic resonance (MR) parameters derived from diffusion, perfusion, and 1 H spectroscopic imaging from the tissue sample locations and the entire tumor. A total of 135 patients were scanned prior to initial surgery, with tumor cellularity scores obtained from 88 image-guided tissue samples. MR parameters were obtained from corresponding sample locations, and histograms of normalized MR parameters within the T2 fluid-attenuated inversion recovery lesion were analyzed in order to evaluate differences between subgroups. For tissue samples, higher tumor scores were related to increased normalized apparent diffusion coefficient (nADC), lower fractional anisotropy (nFA), lower cerebral blood volume (nCBV), higher choline (nCho), and lower N-acetylaspartate (nNAA). Within the T2 lesion, higher tumor grade was associated with higher nADC, lower nFA, and higher Cho to NAA index. Pathological analysis confirmed that diffusion and metabolic parameters increased and perfusion decreased with tumor cellularity. This information can be used to select targets for tissue sampling and to aid in making decisions about treating residual disease. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Trace elements and organic compounds in streambed sediment and fish tissue of coastal New England streams, 1998-99

USGS Publications Warehouse

Chalmers, Ann

2002-01-01

Streambed sediment and fish tissue were collected at 14 river sites in eastern New England during low-flow conditions in 1998 and 1999 as part of the New England Coastal Basins (NECB) study of the U.S. Geological Survey National Water-Quality Assessment (NAWQA) Program. Sampling sites were selected over a range of urban settings. Population densities at selected sites ranged from 26 to 3,585 people per square mile, and urban land use ranged from 1 to 68 percent. The streambed sediment samples were analyzed for a total of 141 contaminants, including 45 trace elements, 32 organochlorine compounds, and 64 semi-volatile organic compounds. The fish tissue samples were analyzed for 22 trace elements and 28 organochlorine compounds. Concentrations of selected contaminants in both streambed sediment and fish tissue correlated more strongly with population density than with other watershed characteristics. Cadmium, copper, lead, mercury, zinc, total polycyclic aromatic hydrocarbons (PAHs), total polychlorinated biphenyls (PCBs), dichloro diphenyl trichloroethane and metabolites (DDTM), and total chlordane in streambed sediment all showed strong positive correlations with population density (rho = 0.71 to 0.85, p value = 0.005 to <0.001). Correlations between population density and selected contaminants in fish tissue were less significant than with streambed sediment (rho = 0.62 to 0.72, p value = 0.03 to 0.008). Organic carbon concentrations were correlated with concentrations of arsenic, selenium, total PAHs, total PCBs, and DDTM in streambed sediment. The relation between concentrations of contaminants in streambed sediment and fish tissue was stronger for organochlorine compounds (rho = 0.75 to 0.55, p = 0.005 to 0.065) than for trace elements (rho = 0.63 to 0.53, p = 0.029 to 0.069). The NECB study area had the highest median concentrations of lead, mercury, total PAHs, total PCBs, and DDTM in streambed sediment and the highest median concentration of PCBs in fish tissue compared to 45 other NAWQA study units across the Nation. Concentrations of many of these constituents in streambed sediment also were frequently above the consensus-based Sediment-Quality Guidelines for the protection of wildlife, suggesting they are a threat to the health of aquatic biota in New England.

Epstein-Barr Virus and Gastric Cancer Risk: A Meta-analysis With Meta-regression of Case-control Studies.

PubMed

Bae, Jong-Myon; Kim, Eun Hee

2016-03-01

Research on how the risk of gastric cancer increases with Epstein-Barr virus (EBV) infection is lacking. In a systematic review that investigated studies published until September 2014, the authors did not calculate the summary odds ratio (SOR) due to heterogeneity across studies. Therefore, we include here additional studies published until October 2015 and conduct a meta-analysis with meta-regression that controls for the heterogeneity among studies. Using the studies selected in the previously published systematic review, we formulated lists of references, cited articles, and related articles provided by PubMed. From the lists, only case-control studies that detected EBV in tissue samples were selected. In order to control for the heterogeneity among studies, subgroup analysis and meta-regression were performed. In the 33 case-control results with adjacent non-cancer tissue, the total number of test samples in the case and control groups was 5280 and 4962, respectively. In the 14 case-control results with normal tissue, the total number of test samples in case and control groups was 1393 and 945, respectively. Upon meta-regression, the type of control tissue was found to be a statistically significant variable with regard to heterogeneity. When the control tissue was normal tissue of healthy individuals, the SOR was 3.41 (95% CI, 1.78 to 6.51; I-squared, 65.5%). The results of the present study support the argument that EBV infection increases the risk of gastric cancer. In the future, age-matched and sex-matched case-control studies should be conducted.

Determination of d-limonene in adipose tissue by gas chromatography-mass spectrometry

PubMed Central

Miller, Jessica A.; Hakim, Iman A.; Thomson, Cynthia; Thompson, Patricia; Chow, H-H. Sherry

2008-01-01

We developed a novel method for analyzing d-limonene levels in adipose tissue. Fat samples were subjected to saponification followed by solvent extraction. d-Limonene in the sample extract was analyzed using gas chromatography-mass spectrometry (GC-MS) with selected ion monitoring. Linear calibration curves were established over the mass range of 79.0-2,529 ng d-limonene per 0.1 grams of adipose tissue. Satisfactory within day precision (RSD 6.7 to 9.6%) and accuracy (% difference of −2.7 to 3.8%) and between day precision (RSD 6.0 to 10.7%) and accuracy (% difference of 1.8 to 2.6%) were achieved. The assay was successfully applied to human fat biopsy samples from a d-limonene feeding trial. PMID:18571481

Prospective Evaluation of Intraprostatic Inflammation and Focal Atrophy as a Predictor of Risk of High-Grade Prostate Cancer and Recurrence after Prostatectomy

DTIC Science & Technology

2014-07-01

the two trials. The expected sample size for this work was 100 cases and 200 controls. Tissue was sufficient for 291 of the men (Task 2 completed in...not collected in SELECT), physical activity (PCPT [not collected in SELECT), cigarette smoking status at randomization (SELECT), use of aspirin

[Study of near infrared spectral preprocessing and wavelength selection methods for endometrial cancer tissue].

PubMed

Zhao, Li-Ting; Xiang, Yu-Hong; Dai, Yin-Mei; Zhang, Zhuo-Yong

2010-04-01

Near infrared spectroscopy was applied to measure the tissue slice of endometrial tissues for collecting the spectra. A total of 154 spectra were obtained from 154 samples. The number of normal, hyperplasia, and malignant samples was 36, 60, and 58, respectively. Original near infrared spectra are composed of many variables, for example, interference information including instrument errors and physical effects such as particle size and light scatter. In order to reduce these influences, original spectra data should be performed with different spectral preprocessing methods to compress variables and extract useful information. So the methods of spectral preprocessing and wavelength selection have played an important role in near infrared spectroscopy technique. In the present paper the raw spectra were processed using various preprocessing methods including first derivative, multiplication scatter correction, Savitzky-Golay first derivative algorithm, standard normal variate, smoothing, and moving-window median. Standard deviation was used to select the optimal spectral region of 4 000-6 000 cm(-1). Then principal component analysis was used for classification. Principal component analysis results showed that three types of samples could be discriminated completely and the accuracy almost achieved 100%. This study demonstrated that near infrared spectroscopy technology and chemometrics method could be a fast, efficient, and novel means to diagnose cancer. The proposed methods would be a promising and significant diagnosis technique of early stage cancer.

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

A New Method for Endoscopic Sampling of Submucosal Tissue in the Gastrointestinal Tract: A Comparison of the Biopsy Forceps and a New Drill Instrument.

PubMed

Walther, Charles; Jeremiasen, Martin; Rissler, Pehr; Johansson, Jan L M; Larsson, Marie S; Walther, Bruno S C S

2016-12-01

Background Sampling of submucosal lesions in the gastrointestinal tract through a flexible endoscope is a well-recognized clinical problem. One technique often used is endoscopic ultrasound-guided fine-needle aspiration, but it does not provide solid tissue biopsies with preserved architecture for histopathological evaluation. To obtain solid tissue biopsies from submucosal lesions, we have constructed a new endoscopic biopsy tool and compared it in a crossover study with the standard double cupped forceps. Methods Ten patients with endoscopically verified submucosal lesions were sampled. The endoscopist selected the position for the biopsies and used the instrument selected by randomization. After a biopsy was harvested, the endoscopist chose the next site for a biopsy and again used the instrument picked by randomization. A total of 6 biopsies, 3 with the forceps and 3 with the drill instrument, were collected in every patient. Results The drill instrument resulted in larger total size biopsies (mm 2 ; Mann-Whitney U test, P = .048) and larger submucosal part (%) of the biopsies (Mann-Whitney U test, P = .003) than the forceps. Two patients were observed because of chest pain and suspicion of bleeding in 24 hours. No therapeutic measures were necessary to be taken. Conclusion The new drill instrument for flexible endoscopy can safely deliver submucosal tissue samples from submucosal lesions in the upper gastrointestinal tract. © The Author(s) 2016.

75 FR 74003 - Endangered Species; Permit No. 15677

Federal Register 2010, 2011, 2012, 2013, 2014

2010-11-30

... measured, weighed, sampled for genetic tissue analysis, and PIT tagged. Additionally, selected adults and..., shortnose sturgeon sex would be determined from a sample of fish annually by either laparoscopy or tubular... disrupters in the environment. Manual tracking and passive detections of telemetered fish at fixed receiver...

Comparison of methods for the extraction of DNA from formalin-fixed, paraffin-embedded archival tissues.

PubMed

Sengüven, Burcu; Baris, Emre; Oygur, Tulin; Berktas, Mehmet

2014-01-01

Discussing a protocol involving xylene-ethanol deparaffinization on slides followed by a kit-based extraction that allows for the extraction of high quality DNA from FFPE tissues. DNA was extracted from the FFPE tissues of 16 randomly selected blocks. Methods involving deparaffinization on slides or tubes, enzyme digestion overnight or for 72 hours and isolation using phenol chloroform method or a silica-based commercial kit were compared in terms of yields, concentrations and the amplifiability. The highest yield of DNA was produced from the samples that were deparaffinized on slides, digested for 72 hours and isolated with a commercial kit. Samples isolated with the phenol-chloroform method produced DNA of lower purity than the samples that were purified with kit. The samples isolated with the commercial kit resulted in better PCR amplification. Silica-based commercial kits and deparaffinized on slides should be considered for DNA extraction from FFPE.

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.

Electrochemical imaging of cells and tissues

PubMed Central

Lin, Tzu-En; Rapino, Stefania; Girault, Hubert H.

2018-01-01

The technological and experimental progress in electrochemical imaging of biological specimens is discussed with a view on potential applications for skin cancer diagnostics, reproductive medicine and microbial testing. The electrochemical analysis of single cell activity inside cell cultures, 3D cellular aggregates and microtissues is based on the selective detection of electroactive species involved in biological functions. Electrochemical imaging strategies, based on nano/micrometric probes scanning over the sample and sensor array chips, respectively, can be made sensitive and selective without being affected by optical interference as many other microscopy techniques. The recent developments in microfabrication, electronics and cell culturing/tissue engineering have evolved in affordable and fast-sampling electrochemical imaging platforms. We believe that the topics discussed herein demonstrate the applicability of electrochemical imaging devices in many areas related to cellular functions. PMID:29899947

Antidepressant pharmaceuticals in two U.S. effluent-impacted streams: Occurrence and fate in water and sediment and selective uptake in fish neural tissue

USGS Publications Warehouse

Schultz, M.M.; Furlong, E.T.; Kolpin, D.W.; Werner, S.L.; Schoenfuss, H.L.; Barber, L.B.; Blazer, V.S.; Norris, D.O.; Vajda, A.M.

2010-01-01

Antidepressant pharmaceuticals are widely prescribed in the United States; release of municipal wastewater effluent is a primary route introducing them to aquatic environments, where little is known about their distribution and fate. Water, bed sediment, and brain tissue from native white suckers (Catostomus commersoni)were collected upstream and atpoints progressively downstream from outfalls discharging to two effluentimpacted streams, Boulder Creek (Colorado) and Fourmile Creek (Iowa). A liquid chromatography/tandem mass spectrometry method was used to quantify antidepressants, including fluoxetine, norfluoxetine (degradate), sertraline, norsertraline (degradate), paroxetine, Citalopram, fluvoxamine, duloxetine, venlafaxine, and bupropion in all three sample matrices. Antidepressants were not present above the limit of quantitation in water samples upstream from the effluent outfalls but were present at points downstream at ng/L concentrations, even at the farthest downstream sampling site 8.4 km downstream from the outfall. The antidepressants with the highest measured concentrations in both streams were venlafaxine, bupropion, and Citalopram and typically were observed at concentrations of at least an order of magnitude greater than the more commonly investigated antidepressants fluoxetine and sertraline. Concentrations of antidepressants in bed sediment were measured at ng/g levels; venlafaxine and fluoxetine were the predominant chemicals observed. Fluoxetine, sertraline, and their degradates were the principal antidepressants observed in fish brain tissue, typically at low ng/g concentrations. Aqualitatively different antidepressant profile was observed in brain tissue compared to streamwater samples. This study documents that wastewater effluent can be a point source of antidepressants to stream ecosystems and that the qualitative composition of antidepressants in brain tissue from exposed fish differs substantially from the compositions observed in streamwater and sediment, suggesting selective uptake. ?? 2010 American Chemical Society.

DNA methylation markers for diagnosis and prognosis of common cancers

PubMed Central

Hao, Xiaoke; Luo, Huiyan; Krawczyk, Michal; Wei, Wei; Wang, Wenqiu; Wang, Juan; Flagg, Ken; Hou, Jiayi; Zhang, Heng; Yi, Shaohua; Jafari, Maryam; Lin, Danni; Chung, Christopher; Caughey, Bennett A.; Li, Gen; Dhar, Debanjan; Shi, William; Zheng, Lianghong; Hou, Rui; Zhu, Jie; Zhao, Liang; Fu, Xin; Zhang, Edward; Zhang, Charlotte; Zhu, Jian-Kang; Karin, Michael; Xu, Rui-Hua; Zhang, Kang

2017-01-01

The ability to identify a specific cancer using minimally invasive biopsy holds great promise for improving the diagnosis, treatment selection, and prediction of prognosis in cancer. Using whole-genome methylation data from The Cancer Genome Atlas (TCGA) and machine learning methods, we evaluated the utility of DNA methylation for differentiating tumor tissue and normal tissue for four common cancers (breast, colon, liver, and lung). We identified cancer markers in a training cohort of 1,619 tumor samples and 173 matched adjacent normal tissue samples. We replicated our findings in a separate TCGA cohort of 791 tumor samples and 93 matched adjacent normal tissue samples, as well as an independent Chinese cohort of 394 tumor samples and 324 matched adjacent normal tissue samples. The DNA methylation analysis could predict cancer versus normal tissue with more than 95% accuracy in these three cohorts, demonstrating accuracy comparable to typical diagnostic methods. This analysis also correctly identified 29 of 30 colorectal cancer metastases to the liver and 32 of 34 colorectal cancer metastases to the lung. We also found that methylation patterns can predict prognosis and survival. We correlated differential methylation of CpG sites predictive of cancer with expression of associated genes known to be important in cancer biology, showing decreased expression with increased methylation, as expected. We verified gene expression profiles in a mouse model of hepatocellular carcinoma. Taken together, these findings demonstrate the utility of methylation biomarkers for the molecular characterization of cancer, with implications for diagnosis and prognosis. PMID:28652331

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.

NS3 protease resistance-associated substitutions in liver tissue and plasma samples from patients infected by hepatitis C virus genotype 1A or 1B.

PubMed

Morsica, Giulia; Andolina, Andrea; Merli, Marco; Messina, Emanuela; Hasson, Hamid; Lazzarin, Adriano; Uberti-Foppa, Caterina; Bagaglio, Sabrina

2017-08-01

The presence of naturally occurring resistance-associated substitutions (RASs) in the HCV-protease domain has been poorly investigated in the liver, the main site of HCV replication. We evaluated the natural resistance of the virus to NS3 protease inhibitors in liver tissue and plasma samples taken from HCV-infected patients. RASs were investigated by means of viral population sequencing in liver tissue samples from 18 HCV-infected patients harbouring genotype 1a or genotype 1b; plasma samples from 12 of these patients were also available for virological investigation. A discordant genotype was found in two of the 12 patients (16.6%) who provided samples from both compartments. Sequence analysis of the NS3 protease domain showed the presence of RASs in four of the 18 liver tissue samples (22.2%), two of which showed cross-resistance to protease inhibitors in clinical use or phase 2-3 trials. The analysis of the 12 paired tissues and plasma samples excluded the presence of RASs in the plasma compartment. The dominance of discordant genotypes in the paired liver and plasma samples of some HCV-infected patients suggests mixed infection possibly leading to the selective advantage of different genotype in the two compartments. The presence of RASs at intra-hepatic level is not uncommon and may lead to the early emergence of cross-resistant strains.

Development and Validation of an Inductively Coupled Plasma Mass Spectrometry (ICP-MS) Method for Quantitative Analysis of Platinum in Plasma, Urine, and Tissues.

PubMed

Zhang, Ti; Cai, Shuang; Forrest, Wai Chee; Mohr, Eva; Yang, Qiuhong; Forrest, M Laird

2016-09-01

Cisplatin, a platinum chemotherapeutic, is one of the most commonly used chemotherapeutic agents for many solid tumors. In this work, we developed and validated an inductively coupled plasma mass spectrometry (ICP-MS) method for quantitative determination of platinum levels in rat urine, plasma, and tissue matrices including liver, brain, lungs, kidney, muscle, heart, spleen, bladder, and lymph nodes. The tissues were processed using a microwave accelerated reaction system (MARS) system prior to analysis on an Agilent 7500 ICP-MS. According to the Food and Drug Administration guidance for industry, bioanalytical validation parameters of the method, such as selectivity, accuracy, precision, recovery, and stability were evaluated in rat biological samples. Our data suggested that the method was selective for platinum without interferences caused by other presenting elements, and the lower limit of quantification was 0.5 ppb. The accuracy and precision of the method were within 15% variation and the recoveries of platinum for all tissue matrices examined were determined to be 85-115% of the theoretical values. The stability of the platinum-containing solutions, including calibration standards, stock solutions, and processed samples in rat biological matrices was investigated. Results indicated that the samples were stable after three cycles of freeze-thaw and for up to three months. © The Author(s) 2016.

Development and Validation of an Inductively Coupled Plasma Mass Spectrometry (ICP-MS) Method for Quantitative Analysis of Platinum in Plasma, Urine, and Tissues

PubMed Central

Zhang, Ti; Cai, Shuang; Forrest, Wai Chee; Mohr, Eva; Yang, Qiuhong; Forrest, M. Laird

2016-01-01

Cisplatin, a platinum chemotherapeutic, is one of the most commonly used chemotherapeutic agents for many solid tumors. In this work, we developed and validated an inductively coupled plasma mass spectrometry (ICP-MS) method for quantitative determination of platinum levels in rat urine, plasma, and tissue matrices including liver, brain, lungs, kidney, muscle, heart, spleen, bladder, and lymph nodes. The tissues were processed using a microwave accelerated reaction system (MARS) system prior to analysis on an Agilent 7500 ICP-MS. According to the Food and Drug Administration guidance for industry, bioanalytical validation parameters of the method, such as selectivity, accuracy, precision, recovery, and stability were evaluated in rat biological samples. Our data suggested that the method was selective for platinum without interferences caused by other presenting elements, and the lower limit of quantification was 0.5 ppb. The accuracy and precision of the method were within 15% variation and the recoveries of platinum for all tissue matrices examined were determined to be 85–115% of the theoretical values. The stability of the platinum-containing solutions, including calibration standards, stock solutions, and processed samples in rat biological matrices was investigated. Results indicated that the samples were stable after three cycles of freeze–thaw and for up to three months. PMID:27527103

Measurement of indicator genes using global complementary DNA (cDNA) amplification, by polyadenylic acid reverse transcriptase polymerase chain reaction (poly A RT-PCR): A feasibility study using paired samples from tissue and ductal juice in patients undergoing pancreatoduodenectomy.

PubMed

Sanyal, Sudip; Siriwardena, Ajith K; Byers, Richard

2018-06-01

The aim of this study is to compare gene expression profiles in RNA isolated from pancreatic ductal juice with the RNA expression profiles of the same genes from matched intra-operative tissue samples from pancreatic tumours. Intra-operative sampling of pancreatic juice and collection of matched tissue samples was undertaken in patients undergoing pancreatoduodenectomy for clinically suspected pancreatic cancer and a precursor lesion, main-duct intraductal papillary mucinous neoplasm. RNA was isolated and Poly A PCR was used to globally amplify the RNA. Real-time polymerase chain reaction (RT-PCR) was used to measure expression levels of 17 genes selected from microarray studies. Spearman's rank correlation test was used to examine the relationship of gene expression between pancreatic juice and tissue. The study was approved by Regional Ethics Committee. Mesothelin (MSLN) showed significant correlation (p < 0.008) in expression levels between paired pancreatic juice and tissue samples in pancreas cancer. In intraductal papillary mucinous neoplasms (IPMN), Matrix Metalloproteinase 7 (MMP7), showed significant correlation (p < 0.01) in the expression levels between paired pancreatic juice and tissue samples. This study confirms that RNA analysis of paired pancreatic juice and tissue samples and establishment of cDNA using poly A PCR is technically feasible. Application of the technique to non-invasively obtained pancreatic juice during endoscopic assessment of tumours and the use of gene arrays of cancer indicator genes are the next steps in development of this technique. Copyright © 2018 IAP and EPC. Published by Elsevier B.V. All rights reserved.

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.

Dynamical properties of the brain tissue under oscillatory shear stresses at large strain range

NASA Astrophysics Data System (ADS)

Boudjema, F.; Khelidj, B.; Lounis, M.

2017-01-01

In this experimental work, we study the viscoelastic behaviour of in vitro brain tissue, particularly the white matter, under oscillatory shear strain. The selective vulnerability of this tissue is the anisotropic mechanical properties of theirs different regions lead to a sensitivity to the angular shear rate and magnitude of strain. For this aim, shear storage modulus (G‧) and loss modulus (G″) were measured over a range of frequencies (1 to 100 Hz), for different levels of strain (1 %, to 50 %). The mechanical responses of the brain matter samples showed a viscoelastic behaviour that depend on the correlated strain level and frequency range and old age sample. The samples have been showed evolution behaviour by increasing then decreasing the strain level. Also, the stiffness anisotropy of brain matter was showed between regions and species.

Genotype, development and tissue-derived variation of cell-wall properties in the lignocellulosic energy crop Miscanthus

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

da Costa, Ricardo M. F.; Lee, Scott J.; Allison, Gordon G.

Species and hybrids of the genus Miscanthus contain attributes that make them front-runners among current selections of dedicated bioenergy crops. A key trait for plant biomass conversion to biofuels and biomaterials is cell-wall quality; however, knowledge of cell-wall composition and biology in Miscanthus species is limited. This study presents data on cell-wall compositional changes as a function of development and tissue type across selected genotypes, and considers implications for the development of miscanthus as a sustainable and renewable bioenergy feedstock. Cell-wall biomass was analysed for 25 genotypes, considering different developmental stages and stem vs. leaf compositional variability, by Fourier transformmore » mid-infrared spectroscopy and lignin determination. In addition, a Clostridium phytofermentans bioassay was used to assess cell-wall digestibility and conversion to ethanol. Important cell-wall compositional differences between miscanthus stem and leaf samples were found to be predominantly associated with structural carbohydrates. Lignin content increased as plants matured and was higher in stem tissues. Although stem lignin concentration correlated inversely with ethanol production, no such correlation was observed for leaves. Leaf tissue contributed significantly to total above-ground biomass at all stages, although the extent of this contribution was genotype-dependent. In conclusion, it is hypothesized that divergent carbohydrate compositions and modifications in stem and leaf tissues are major determinants for observed differences in cell-wall quality. The findings indicate that improvement of lignocellulosic feedstocks should encompass tissue-dependent variation as it affects amenability to biological conversion. For gene-trait associations relating to cell-wall quality, the data support the separate examination of leaf and stem composition, as tissue-specific traits may be masked by considering only total above-ground biomass samples, and sample variability could be mostly due to varying tissue contributions to total biomass.« less

Genotype, development and tissue-derived variation of cell-wall properties in the lignocellulosic energy crop Miscanthus

DOE PAGES

da Costa, Ricardo M. F.; Lee, Scott J.; Allison, Gordon G.; ...

2014-04-15

Species and hybrids of the genus Miscanthus contain attributes that make them front-runners among current selections of dedicated bioenergy crops. A key trait for plant biomass conversion to biofuels and biomaterials is cell-wall quality; however, knowledge of cell-wall composition and biology in Miscanthus species is limited. This study presents data on cell-wall compositional changes as a function of development and tissue type across selected genotypes, and considers implications for the development of miscanthus as a sustainable and renewable bioenergy feedstock. Cell-wall biomass was analysed for 25 genotypes, considering different developmental stages and stem vs. leaf compositional variability, by Fourier transformmore » mid-infrared spectroscopy and lignin determination. In addition, a Clostridium phytofermentans bioassay was used to assess cell-wall digestibility and conversion to ethanol. Important cell-wall compositional differences between miscanthus stem and leaf samples were found to be predominantly associated with structural carbohydrates. Lignin content increased as plants matured and was higher in stem tissues. Although stem lignin concentration correlated inversely with ethanol production, no such correlation was observed for leaves. Leaf tissue contributed significantly to total above-ground biomass at all stages, although the extent of this contribution was genotype-dependent. In conclusion, it is hypothesized that divergent carbohydrate compositions and modifications in stem and leaf tissues are major determinants for observed differences in cell-wall quality. The findings indicate that improvement of lignocellulosic feedstocks should encompass tissue-dependent variation as it affects amenability to biological conversion. For gene-trait associations relating to cell-wall quality, the data support the separate examination of leaf and stem composition, as tissue-specific traits may be masked by considering only total above-ground biomass samples, and sample variability could be mostly due to varying tissue contributions to total biomass.« less

Genotype, development and tissue-derived variation of cell-wall properties in the lignocellulosic energy crop Miscanthus.

PubMed

da Costa, Ricardo M F; Lee, Scott J; Allison, Gordon G; Hazen, Samuel P; Winters, Ana; Bosch, Maurice

2014-10-01

Species and hybrids of the genus Miscanthus contain attributes that make them front-runners among current selections of dedicated bioenergy crops. A key trait for plant biomass conversion to biofuels and biomaterials is cell-wall quality; however, knowledge of cell-wall composition and biology in Miscanthus species is limited. This study presents data on cell-wall compositional changes as a function of development and tissue type across selected genotypes, and considers implications for the development of miscanthus as a sustainable and renewable bioenergy feedstock. Cell-wall biomass was analysed for 25 genotypes, considering different developmental stages and stem vs. leaf compositional variability, by Fourier transform mid-infrared spectroscopy and lignin determination. In addition, a Clostridium phytofermentans bioassay was used to assess cell-wall digestibility and conversion to ethanol. Important cell-wall compositional differences between miscanthus stem and leaf samples were found to be predominantly associated with structural carbohydrates. Lignin content increased as plants matured and was higher in stem tissues. Although stem lignin concentration correlated inversely with ethanol production, no such correlation was observed for leaves. Leaf tissue contributed significantly to total above-ground biomass at all stages, although the extent of this contribution was genotype-dependent. It is hypothesized that divergent carbohydrate compositions and modifications in stem and leaf tissues are major determinants for observed differences in cell-wall quality. The findings indicate that improvement of lignocellulosic feedstocks should encompass tissue-dependent variation as it affects amenability to biological conversion. For gene-trait associations relating to cell-wall quality, the data support the separate examination of leaf and stem composition, as tissue-specific traits may be masked by considering only total above-ground biomass samples, and sample variability could be mostly due to varying tissue contributions to total biomass. © The Author 2014. Published by Oxford University Press on behalf of the Annals of Botany Company.

Spatial mapping of metals in tissue-sections using combination of mass-spectrometry and histology through image registration

NASA Astrophysics Data System (ADS)

Anyz, Jiri; Vyslouzilova, Lenka; Vaculovic, Tomas; Tvrdonova, Michaela; Kanicky, Viktor; Haase, Hajo; Horak, Vratislav; Stepankova, Olga; Heger, Zbynek; Adam, Vojtech

2017-01-01

We describe a new procedure for the parallel mapping of selected metals in histologically characterized tissue samples. Mapping is achieved via image registration of digital data obtained from two neighbouring cryosections by scanning the first as a histological sample and subjecting the second to laser ablation inductively coupled plasma mass spectrometry. This computer supported procedure enables determination of the distribution and content of metals of interest directly in the chosen histological zones and represents a substantial improvement over the standard approach, which determines these values in tissue homogenates or whole tissue sections. The potential of the described procedure was demonstrated in a pilot study that analysed Zn and Cu levels in successive development stages of pig melanoma tissue using MeLiM (Melanoma-bearing-Libechov-Minipig) model. We anticipate that the procedure could be useful for a complex understanding of the role that the spatial distribution of metals plays within tissues affected by pathological states including cancer.

Spatial mapping of metals in tissue-sections using combination of mass-spectrometry and histology through image registration

PubMed Central

Anyz, Jiri; Vyslouzilova, Lenka; Vaculovic, Tomas; Tvrdonova, Michaela; Kanicky, Viktor; Haase, Hajo; Horak, Vratislav; Stepankova, Olga; Heger, Zbynek; Adam, Vojtech

2017-01-01

We describe a new procedure for the parallel mapping of selected metals in histologically characterized tissue samples. Mapping is achieved via image registration of digital data obtained from two neighbouring cryosections by scanning the first as a histological sample and subjecting the second to laser ablation inductively coupled plasma mass spectrometry. This computer supported procedure enables determination of the distribution and content of metals of interest directly in the chosen histological zones and represents a substantial improvement over the standard approach, which determines these values in tissue homogenates or whole tissue sections. The potential of the described procedure was demonstrated in a pilot study that analysed Zn and Cu levels in successive development stages of pig melanoma tissue using MeLiM (Melanoma-bearing-Libechov-Minipig) model. We anticipate that the procedure could be useful for a complex understanding of the role that the spatial distribution of metals plays within tissues affected by pathological states including cancer. PMID:28071735

[Differential expression genes of bone tissues surrounding implants in diabetic rats by gene chip].

PubMed

Wang, Xin-xin; Ma, Yue; Li, Qing; Jiang, Bao-qi; Lan, Jing

2012-10-01

To compare mRNA expression profiles of bone tissues surrounding implants between normal rats and rats with diabetes using microarray technology. Six Wistar rats were randomly selected and divided into normal model group and diabetic group. Diabetic model condition was established by injecting Streptozotocin into peritoneal space. Titanium implants were implanted into the epiphyseal end of the rats' tibia. Bone tissues surrounding implant were harvested and sampled after 3 months to perform comprehensive RNA gene expression profiling, including 17983 for genome-wide association study.GO analysis was used to compare different gene expression and real-time PCR was used to confirm the results on core samples. The results indicated that there were 1084 differential gene expression. In the diabetic model, there were 352 enhanced expression genes, 732 suppressed expression genes. GO analysis involved 1154 different functional type. Osteoblast related gene expressions in bone tissue samples of diabetic rats were decreased, and lipid metabolism pathway related gene expression was increased.

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.

Data on occurrence of selected trace metals, organochlorines, and semivolatile organic compounds in edible fish tissues from Lake Worth, Fort Worth, Texas, 1999

USGS Publications Warehouse

Moring, J. Bruce

2002-01-01

A public-health assessment conducted for the Texas Department of Health and the Agency for Toxic Substances and Disease Registry concluded that exposure to contaminants through the aquatic food chain is an indeterminate human-health hazard in Lake Worth, Fort Worth, Texas. In 1999, the U.S. Geological Survey, in cooperation with the U.S. Air Force and in collaboration with the Texas Department of Health, collected samples of edible fish tissues from Lake Worth for analysis of selected trace metals, organochlorines, and semivolatile organic compounds to support a human-health risk assessment. Left-side, skin-off fillet samples were collected from 10 individuals each of channel catfish, common carp, freshwater drum (gaspergou), largemouth bass, and white crappie but only from five smallmouth buffalo. The U.S. Geological Survey National Water Quality Laboratory analyzed the samples for 22 trace metals, 40 organochlorine pesticides and polychlorinated biphenyls, and 75 semivolatile organic compounds.

A DNA methylation map of human cancer at single base-pair resolution.

PubMed

Vidal, E; Sayols, S; Moran, S; Guillaumet-Adkins, A; Schroeder, M P; Royo, R; Orozco, M; Gut, M; Gut, I; Lopez-Bigas, N; Heyn, H; Esteller, M

2017-10-05

Although single base-pair resolution DNA methylation landscapes for embryonic and different somatic cell types provided important insights into epigenetic dynamics and cell-type specificity, such comprehensive profiling is incomplete across human cancer types. This prompted us to perform genome-wide DNA methylation profiling of 22 samples derived from normal tissues and associated neoplasms, including primary tumors and cancer cell lines. Unlike their invariant normal counterparts, cancer samples exhibited highly variable CpG methylation levels in a large proportion of the genome, involving progressive changes during tumor evolution. The whole-genome sequencing results from selected samples were replicated in a large cohort of 1112 primary tumors of various cancer types using genome-scale DNA methylation analysis. Specifically, we determined DNA hypermethylation of promoters and enhancers regulating tumor-suppressor genes, with potential cancer-driving effects. DNA hypermethylation events showed evidence of positive selection, mutual exclusivity and tissue specificity, suggesting their active participation in neoplastic transformation. Our data highlight the extensive changes in DNA methylation that occur in cancer onset, progression and dissemination.

Histopathological findings in immunohistological staining of the granulomatous tissue reaction associated with tuberculosis.

PubMed

Karimi, Shirin; Shamaei, Masoud; Pourabdollah, Mihan; Sadr, Makan; Karbasi, Mehrdad; Kiani, Arda; Bahadori, Moslem

2014-01-01

Purpose. The histological diagnosis of Mycobacterium tuberculosis (MTB) remains a diagnostic challenge despite different methods. Immunohistochemistry (IHC) not only could confirm granulomatous tissue involvement but also can demonstrate MTB antigen immunolocalization. This study tries to clarify the details of immunohistochemical staining for MTB with pAbBCG. Materials/Methods. Twenty-three confirmed TB granulomatous tissue samples were studied by Ziehl-Neelsen and immunohistochemistry (IHC) staining with pAbBCG. Samples were selected from the archive of the Department of Pathology, National Research Institute of Tuberculosis and Lung Disease, Tehran, Iran. Results. IHC staining was positive in all samples, whereas Ziehl-Neelsen was positive in 9 cases out of 23 (39.1%). Tissue types used were pleural tissue, lymph nodes, and lung tissue. IHC showed positive coarse granular cytoplasmic and round, fragmented bacillary staining. In this study, epithelioid cells clearly showed more positive staining at the periphery of the granuloma rather than the center of granuloma. There is also positive staining in endothelial cells, fibroblasts, plasma cells, lymphocytes, and macrophages outside the granuloma. Conclusion. Considering the criteria of positive immunohistochemical staining of TB granulomatous reactions, this stain not only highlights the presence of mycobacterial antigens for tissue diagnosis, but also could morphologically localize its distribution in different cells.

Histopathological Findings in Immunohistological Staining of the Granulomatous Tissue Reaction Associated with Tuberculosis

PubMed Central

Karimi, Shirin; Pourabdollah, Mihan; Sadr, Makan; Karbasi, Mehrdad; Bahadori, Moslem

2014-01-01

Purpose. The histological diagnosis of Mycobacterium tuberculosis (MTB) remains a diagnostic challenge despite different methods. Immunohistochemistry (IHC) not only could confirm granulomatous tissue involvement but also can demonstrate MTB antigen immunolocalization. This study tries to clarify the details of immunohistochemical staining for MTB with pAbBCG. Materials/Methods. Twenty-three confirmed TB granulomatous tissue samples were studied by Ziehl-Neelsen and immunohistochemistry (IHC) staining with pAbBCG. Samples were selected from the archive of the Department of Pathology, National Research Institute of Tuberculosis and Lung Disease, Tehran, Iran. Results. IHC staining was positive in all samples, whereas Ziehl-Neelsen was positive in 9 cases out of 23 (39.1%). Tissue types used were pleural tissue, lymph nodes, and lung tissue. IHC showed positive coarse granular cytoplasmic and round, fragmented bacillary staining. In this study, epithelioid cells clearly showed more positive staining at the periphery of the granuloma rather than the center of granuloma. There is also positive staining in endothelial cells, fibroblasts, plasma cells, lymphocytes, and macrophages outside the granuloma. Conclusion. Considering the criteria of positive immunohistochemical staining of TB granulomatous reactions, this stain not only highlights the presence of mycobacterial antigens for tissue diagnosis, but also could morphologically localize its distribution in different cells. PMID:24511393

Immunochemical Studies on Selected Marine Toxins

DTIC Science & Technology

1993-05-31

6) Critical evaluation of established immunoassays was made through analysis of various naturally -occurring MCYST containing samples. We found that...hepatopancreas of toxic clams rather than the soft tissues. Microcystin could also be detected by ELISA in naturally -occurring algae bloom samples that...antibody against MCYST 13 7. Other immunochemical studies on MCYST 13 a. Analysis of MCYST in algae and various samples collected from naturally -occurring

Sample processing, protocol, and statistical analysis of the time-of-flight secondary ion mass spectrometry (ToF-SIMS) of protein, cell, and tissue samples.

PubMed

Barreto, Goncalo; Soininen, Antti; Sillat, Tarvo; Konttinen, Yrjö T; Kaivosoja, Emilia

2014-01-01

Time-of-flight secondary ion mass spectrometry (ToF-SIMS) is increasingly being used in analysis of biological samples. For example, it has been applied to distinguish healthy and osteoarthritic human cartilage. This chapter discusses ToF-SIMS principle and instrumentation including the three modes of analysis in ToF-SIMS. ToF-SIMS sets certain requirements for the samples to be analyzed; for example, the samples have to be vacuum compatible. Accordingly, sample processing steps for different biological samples, i.e., proteins, cells, frozen and paraffin-embedded tissues and extracellular matrix for the ToF-SIMS are presented. Multivariate analysis of the ToF-SIMS data and the necessary data preprocessing steps (peak selection, data normalization, mean-centering, and scaling and transformation) are discussed in this chapter.

Simultaneous determination of ten Aconitum alkaloids in rat tissues by UHPLC-MS/MS and its application to a tissue distribution study on the compatibility of Heishunpian and Fritillariae thunbergii Bulbus.

PubMed

Yang, Bin; Xu, Yanyan; Wu, Yuanyuan; Wu, Huanyu; Wang, Yuan; Yuan, Lei; Xie, Jiabin; Li, Yubo; Zhang, Yanjun

2016-10-15

A rapid, sensitive and selective ultra-high performance liquid chromatography with tandem mass spectrometry (UHPLC-MS/MS) method was developed and validated for simultaneous determination of ten Aconitum alkaloids in rat tissues. The tissue samples were prepared by a simple procedure protein precipitation with acetonitrile containing 0.1% acetic acid and separated on an Agilent XDB C18 column (4.6 mm×50mm, 1.8μm) using gradient elution with a mobile phase consisting of water and acetonitrile (both containing 0.1% formic acid) at a flow rate of 0.3mL/min. The quantitive determination was performed on an electrospray ionization (ESI) triple quadrupole tandem mass spectrometer using selective reaction monitoring (SRM) under positive ionization mode. The established method was fully validated according to the USA Food and Drug Administration (FDA) bioanalytical method validation guidance and the results demonstrated that the method was sensitive and selective with the lowest limits of quantification (LLOQ) at 0.025ng/mL in rat tissue homogenates. Meanwhile, the linearity, precision, accuracy, extraction recovery, matrix effect and stability were all within the required limits of biological sample analysis. After method validation, the validated method was successfully applied to the tissue distribution study on the compatibility of Heishunpian (HSP, the processed product of Aconitum carmichaelii Debx) and Fritillariae thunbergii Bulbus (Zhebeimu, ZBM). The results indicated that the distribution feature of monoester diterpenoid aconitines (MDAs), diester diterpenoid aconitines (DDAs) and non-ester alkaloids (NEAs) were inconsistency, and the compatibility of HSP and ZBM resulted in the distribution amount of DDAs increased in tissues. What's more, the results could provide the reliable basis for systematic research on the substance foundation of the compatibility of the herbal pair. Copyright © 2016 Elsevier B.V. All rights reserved.

[Methylation of selected tumor-supressor genes in benign and malignant ovarian tumors].

PubMed

Cul'bová, M; Lasabová, Z; Stanclová, A; Tilandyová, P; Zúbor, P; Fiolka, R; Danko, J; Visnovský, J

2011-09-01

To evaluate the usefullness of examination of methylation status of selected tumor-supressor genes in early diagnosis of ovarian cancer. Prospective clinical study. Department of Gynecology and Obstetrics, Department of Molecular Biology, Jessenius Medical Faculty, Commenius University, Martin, Slovak Republic. In this study we analyzed hypermethylation of 5 genes RASSF1A, GSTP, E-cadherin, p16 and APC in ovarian tumor samples from 34 patients - 13 patients with epithelial ovarian cancer, 2 patients with border-line ovarian tumors, 12 patients with benign lesions of ovaries and 7 patients with healthy ovarian tissue. The methylation status of promoter region of tumor-supressor genes was determined by Methylation Specific Polymerase Chain Reaction (MSP) using a nested two-step approach with bisulfite modified DNA template and specific primers. Gene methylation analysis revealed hypermethylation of gene RASSF1A (46%) and GSTP (8%) only in malignant ovarian tissue samples. Ecad, p16 and APC genes were methylated both in maignant and benign tissue samples. Methylation positivity in observed genes was present independently to all clinical stages of ovarian cancer and to tumor grades. However, there was observed a trend of increased number and selective involvement of methylated genes with increasing disease stages. Furthermore, there was no association between positive methylation status and histological subtypes of ovarian carcinomas. RASSF1A and GSTP promoter methylation positivity is associated with ovarian cancer. The revealed gene-selective methylation positivity and the increased number of methylated genes with advancing disease stages could be considered as a useful molecular marker for early detection of ovarian cancer. However, there is need to find diagnostic approach of specifically and frequently methylated genes to determining a methylation phenotype for early detection of ovarian malignancies.

Selective sensitivity of Mueller imaging for tissue scattering over absorption changes in cancer mimicking phantoms

NASA Astrophysics Data System (ADS)

Fathima, Adeeba; Sharma B. S., Mahima; N., Sujatha

2018-03-01

Tissue characterization using optical polarimetry, especially Mueller imaging is receiving sustained interest due to its potential in achieving optical contrast between normal and malignant variations. This is particularly important in identifying the margin of malignant growth in suspected tissue regions for accurate surgical removal, or in aiding the sampling procedure during biopsy. The sensitivity of Mueller matrix derived depolarization index to the combined effects of changes in scattering and absorption occurring in a cancerous growth is illustrated in this study. Depolarization imaging is shown to be useful in demarcating the boundary of two regions of differing optical properties using a tissue phantom, modeled according to the changes expected during cancerous growth in tissue. Tissue scattering and absorption are expected to generally increase with the nuclear size change and crowding as well as angiogenesis associated with malignancy. We have observed that there is selective sensitivity for the Mueller elements and derived depolarization index to tissue scattering over absorption in the object field. Although the scattering and absorption are expected to increase and decrease depolarization respectively, the optical contrast of Mueller images and the derived depolarization index between normal and cancerous tissue is found appreciable in this region.

Plant Tissues in 3D via X-Ray Tomography: Simple Contrasting Methods Allow High Resolution Imaging

PubMed Central

Staedler, Yannick M.; Masson, David; Schönenberger, Jürg

2013-01-01

Computed tomography remains strongly underused in plant sciences despite its high potential in delivering detailed 3D phenotypical information because of the low X-ray absorption of most plant tissues. Existing protocols to study soft tissues display poor performance, especially when compared to those used on animals. More efficient protocols to study plant material are therefore needed. Flowers of Arabidopsis thaliana and Marcgravia caudata were immersed in a selection of contrasting agents used to treat samples for transmission electron microscopy. Grayscale values for floral tissues and background were measured as a function of time. Contrast was quantified via a contrast index. The thick buds of Marcgravia were scanned to determine which contrasting agents best penetrate thick tissues. The highest contrast increase with cytoplasm-rich tissues was obtained with phosphotungstate, whereas osmium tetroxide and bismuth tatrate displayed the highest contrast increase with vacuolated tissues. Phosphotungstate also displayed the best sample penetration. Furthermore, infiltration with phosphotungstate allowed imaging of all plants parts at a high resolution of 3 µm, which approaches the maximum resolution of our equipment: 1.5 µm. The high affinity of phosphotungstate for vasculature, cytoplasm-rich tissue, and pollen causes these tissues to absorb more X-rays than the surrounding tissues, which, in turn, makes these tissues appear brighter on the scan data. Tissues with different brightness can then be virtually dissected from each other by selecting the bracket of grayscale to be visualized. Promising directions for the future include in silico phenotyping and developmental studies of plant inner parts (e.g., ovules, vasculature, pollen, and cell nuclei) via virtual dissection as well as correlations of quantitative phenotypes with omics datasets. Therefore, this work represents a crucial improvement of previous methods, allowing new directions of research to be undertaken in areas ranging from morphology to systems biology. PMID:24086499

Comparison of Methods for the Extraction of DNA from Formalin-Fixed, Paraffin-Embedded Archival Tissues

PubMed Central

Sengüven, Burcu; Baris, Emre; Oygur, Tulin; Berktas, Mehmet

2014-01-01

Aim: Discussing a protocol involving xylene-ethanol deparaffinization on slides followed by a kit-based extraction that allows for the extraction of high quality DNA from FFPE tissues. Methods: DNA was extracted from the FFPE tissues of 16 randomly selected blocks. Methods involving deparaffinization on slides or tubes, enzyme digestion overnight or for 72 hours and isolation using phenol chloroform method or a silica-based commercial kit were compared in terms of yields, concentrations and the amplifiability. Results: The highest yield of DNA was produced from the samples that were deparaffinized on slides, digested for 72 hours and isolated with a commercial kit. Samples isolated with the phenol-chloroform method produced DNA of lower purity than the samples that were purified with kit. The samples isolated with the commercial kit resulted in better PCR amplification. Conclusion: Silica-based commercial kits and deparaffinized on slides should be considered for DNA extraction from FFPE. PMID:24688314

Determination of catechins and catechin gallates in tissues by liquid chromatography with coulometric array detection and selective solid phase extraction.

PubMed

Chu, Kai On; Wang, Chi Chiu; Chu, Ching Yan; Rogers, Michael Scott; Choy, Kwong Wai; Pang, Chi Pui

2004-10-25

Catechins levels in organ tissues, particularly liver, determined by published methods are unexpectedly low, probably due to the release of oxidative enzymes, metal ions and reactive metabolites from tissue cells during homogenization and to the pro-oxidant effects of ascorbic acid during sample processing in the presence of metal ions. We describe a new method for simultaneous analysis of eight catechins in tissue: (+)-catechin (C), (-)-epicatechin (EC), (-)-gallocatechin (GC), (-)-epigallocatechin (EGC), (-)-catechin gallate (CG), (-)-epicatechin gallate (ECG), (-)-gallocatechin gallate (GCG) and (-)-epigallocatechin gallate (EGCG) (Fig. 1). The new extraction procedure utilized a methanol/ethylacetate/dithionite (2:1:3) mixture during homogenization for simultaneous enzyme precipitation and antioxidant protection. Selective solid phase extraction was used to remove most interfering bio-matrices. Reversed phase HPLC with CoulArray detection was used to determine the eight catechins simultaneously within 25 min. Good linearity (>0.9922) was obtained in the range 20-4000 ng/g. The coefficients of variance (CV) were less than 5%. Absolute recovery ranged from 62 to 96%, accuracy 92.5 +/- 4.5 to 104.9 +/- 6%. The detection limit was 5 ng/g. This method is capable for determining catechins in rat tissues of liver, brain, spleen, and kidney. The method is robust, reproducible, with high recovery, and has been validated for both in vitro and in vivo sample analysis.

Cancer Liquid Biopsy: Is It Ready for Clinic?

PubMed

Pan, Ying; Ji, John S; Jin, Jason Gang; Kuo, Winston Patrick; Kang, Hongjun

2017-01-01

The management of cancer relies on a combination of imaging and tissue biopsy for diagnosis, monitoring, and molecular classification-based patient stratification to ensure appropriate treatment. Conventional tissue biopsy harvests tumor samples with invasive procedures, which are often difficult for patients with advanced disease. Given the well-recognized intratumor genetic heterogeneity [1], the biopsy of small tumor fragments does not necessarily represent all the genetic aberrations in the tumor, but sampling the entire tumor in each patient is not realistic. Moreover, tumors evolve all the time from local to advanced disease and by adapting to selective pressure from treatment.

Nonnegative constraint analysis of key fluorophores within human breast cancer using native fluorescence spectroscopy excited by selective wavelength of 300 nm

NASA Astrophysics Data System (ADS)

Pu, Yang; Sordillo, Laura A.; Alfano, Robert R.

2015-03-01

Native fluorescence spectroscopy offers an important role in cancer discrimination. It is widely acknowledged that the emission spectrum of tissue is a superposition of spectra of various salient fluorophores. In this study, the native fluorescence spectra of human cancerous and normal breast tissues excited by selected wavelength of 300 nm are used to investigate the key building block fluorophores: tryptophan and reduced nicotinamide adenine dinucleotide (NADH). The basis spectra of these key fluorophores' contribution to the tissue emission spectra are obtained by nonnegative constraint analysis. The emission spectra of human cancerous and normal tissue samples are projected onto the fluorophore spectral subspace. Since previous studies indicate that tryptophan and NADH are key fluorophores related with tumor evolution, it is essential to obtain their information from tissue fluorescence but discard the redundancy. To evaluate the efficacy of for cancer detection, linear discriminant analysis (LDA) classifier is used to evaluate the sensitivity, and specificity. This research demonstrates that the native fluorescence spectroscopy measurements are effective to detect changes of fluorophores' compositions in tissues due to the development of cancer.

Chlordane contamination in selected estuarine and coastal marine finfish and shellfish of New Jersey, USA.

PubMed

Kennish, M J; Ruppel, B E

1996-01-01

Analysis of chlordane contamination in selected finfish and shellfish species from estuarine and coastal marine waters of New Jersey, USA, indicates consistently highest organochlorine pesticide levels in samples from the north and northeast regions of the state. Gas chromatographic analysis of tissue samples from four finfish species (American eel, Anguilla rostrata; bluefish, Pomatomus saltatrix; striped bass, Morone saxatilis; and weakfish, Cynoscion regalis) and one shellfish species (blue crab, Callinectes sapidus) collected at 28 stations throughout the state during the 1986-1987 sampling period and 42 stations during the 1988-1991 sampling period revealed chlordane concentrations ranging from 5 to 2150 ppb wet weight (wet wt). Highest mean concentrations of chlordane (300 ppb wet wt) occurred in American eel from the Camden region, with secondary maxima (100 ppb wet wt) observed in the northeast region. Lower mean concentrations of chlordane (< 100 ppb wet wt) were recorded in tissue samples of other species collected elsewhere in the state. In general, the lowest levels of chlordane contamination (mean < 50 ppb wet wt) were found in samples from the south coast and Delaware regions. A major conclusion of this study is that some commercially and recreationally important finfish and shellfish species in New Jersey waters, especially those which are lipid-rich, have continued to accumulate chlordane from the environment long after restrictive regulations were first placed on its use in the US. The greatest impact of chlordane contamination is nearby metropolitan centers of the state (i.e., New York, Newark, Camden), although the total concentrations of chlordane in tissue samples from these areas are currently far less than the US Food and Drug Administration action level of 300 ppb wet wt.

Selection of reliable reference genes for quantitative real-time PCR gene expression analysis in Jute (Corchorus capsularis) under stress treatments

PubMed Central

Niu, Xiaoping; Qi, Jianmin; Zhang, Gaoyang; Xu, Jiantang; Tao, Aifen; Fang, Pingping; Su, Jianguang

2015-01-01

To accurately measure gene expression using quantitative reverse transcription PCR (qRT-PCR), reliable reference gene(s) are required for data normalization. Corchorus capsularis, an annual herbaceous fiber crop with predominant biodegradability and renewability, has not been investigated for the stability of reference genes with qRT-PCR. In this study, 11 candidate reference genes were selected and their expression levels were assessed using qRT-PCR. To account for the influence of experimental approach and tissue type, 22 different jute samples were selected from abiotic and biotic stress conditions as well as three different tissue types. The stability of the candidate reference genes was evaluated using geNorm, NormFinder, and BestKeeper programs, and the comprehensive rankings of gene stability were generated by aggregate analysis. For the biotic stress and NaCl stress subsets, ACT7 and RAN were suitable as stable reference genes for gene expression normalization. For the PEG stress subset, UBC, and DnaJ were sufficient for accurate normalization. For the tissues subset, four reference genes TUBβ, UBI, EF1α, and RAN were sufficient for accurate normalization. The selected genes were further validated by comparing expression profiles of WRKY15 in various samples, and two stable reference genes were recommended for accurate normalization of qRT-PCR data. Our results provide researchers with appropriate reference genes for qRT-PCR in C. capsularis, and will facilitate gene expression study under these conditions. PMID:26528312

Technical issues affecting the implementation of US Environmental Protection Agency's proposed fish tissue-based aquatic criterion for selenium.

PubMed

Lemly, A Dennis; Skorupa, Joseph P

2007-10-01

The US Environmental Protection Agency is developing a national water quality criterion for selenium that is based on concentrations of the element in fish tissue. Although this approach offers advantages over the current water-based regulations, it also presents new challenges with respect to implementation. A comprehensive protocol that answers the "what, where, and when" is essential with the new tissue-based approach in order to ensure proper acquisition of data that apply to the criterion. Dischargers will need to understand selenium transport, cycling, and bioaccumulation in order to effectively monitor for the criterion and, if necessary, develop site-specific standards. This paper discusses 11 key issues that affect the implementation of a tissue-based criterion, ranging from the selection of fish species to the importance of hydrological units in the sampling design. It also outlines a strategy that incorporates both water column and tissue-based approaches. A national generic safety-net water criterion could be combined with a fish tissue-based criterion for site-specific implementation. For the majority of waters nationwide, National Pollution Discharge Elimination System permitting and other activities associated with the Clean Water Act could continue without the increased expense of sampling and interpreting biological materials. Dischargers would do biotic sampling intermittently (not a routine monitoring burden) on fish tissue relative to the fish tissue criterion. Only when the fish tissue criterion is exceeded would a full site-specific analysis including development of intermedia translation factors be necessary.

High axial resolution imaging system for large volume tissues using combination of inclined selective plane illumination and mechanical sectioning

PubMed Central

Zhang, Qi; Yang, Xiong; Hu, Qinglei; Bai, Ke; Yin, Fangfang; Li, Ning; Gang, Yadong; Wang, Xiaojun; Zeng, Shaoqun

2017-01-01

To resolve fine structures of biological systems like neurons, it is required to realize microscopic imaging with sufficient spatial resolution in three dimensional systems. With regular optical imaging systems, high lateral resolution is accessible while high axial resolution is hard to achieve in a large volume. We introduce an imaging system for high 3D resolution fluorescence imaging of large volume tissues. Selective plane illumination was adopted to provide high axial resolution. A scientific CMOS working in sub-array mode kept the imaging area in the sample surface, which restrained the adverse effect of aberrations caused by inclined illumination. Plastic embedding and precise mechanical sectioning extended the axial range and eliminated distortion during the whole imaging process. The combination of these techniques enabled 3D high resolution imaging of large tissues. Fluorescent bead imaging showed resolutions of 0.59 μm, 0.47μm, and 0.59 μm in the x, y, and z directions, respectively. Data acquired from the volume sample of brain tissue demonstrated the applicability of this imaging system. Imaging of different depths showed uniform performance where details could be recognized in either the near-soma area or terminal area, and fine structures of neurons could be seen in both the xy and xz sections. PMID:29296503

An improved method for assaying phosphocholine and glycerophosphocholine in mouse tissue.

PubMed

Murai, S; Saito, H; Shirato, R; Kawaguchi, T

2001-01-01

To measure the levels of phosphocholine (PCh) and glycerophosphocholine (GPCh) in the tissues and organs of mice, we developed a simple and rapid method using high-performance liquid chromatography (HPLC) with electrochemical detection (ECD) and an immobilized enzyme column. Under our modifications of the separation procedure of Klein et al. [Neurochem. Int. 2 (1993) 293], PCh and GPCh in the hydrophilic phase of the homogenate samples were selectively hydrolyzed into free choline by alkaline phosphatase and a 0.4-N perchloric acid solution, respectively, and the resulting hydrolyzed mixtures were directly injected into the HPLC system for analysis. The present method permits PCh or GPCh assay within 5 min in one chromatographic run. Recoveries from tissue samples were 97% for PCh and 101% for GPCh. The percentages of the crossover reaction to the authentic PCh and GPCh were 0.4% and 3.8%, respectively. The within-run coefficients of variation for choline derived from PCh and GPCh in the tissue samples were 1.2% and 1.4%, respectively. The method is effective and has been applied to the measurement of PCh and GPCh levels in several tissues of mice.

CLOSED-LOOP STRIPPING ANALYSIS (CLSA) OF ...

EPA Pesticide Factsheets

Synthetic musk compounds have been found in surface water, fish tissues, and human breast milk. Current techniques for separating these compounds from fish tissues require tedious sample clean-upprocedures A simple method for the deterrnination of these compounds in fish tissues has been developed. Closed-loop stripping of saponified fish tissues in a I -L Wheaton purge-and-trap vessel is used to strip compounds with high vapor pressures such as synthetic musks from the matrix onto a solid sorbent (Abselut Nexus). This technique is useful for screening biological tissues that contain lipids for musk compounds. Analytes are desorbed from the sorbent trap sequentially with polar and nonpolar solvents, concentrated, and directly analyzed by high resolution gas chromatography coupled to a mass spectrometer operating in the selected ion monitoring mode. In this paper, we analyzed two homogenized samples of whole fish tissues with spiked synthetic musk compounds using closed-loop stripping analysis (CLSA) and pressurized liquid extraction (PLE). The analytes were not recovered quantitatively but the extraction yield was sufficiently reproducible for at least semi-quantitative purposes (screening). The method was less expensive to implement and required significantly less sample preparation than the PLE technique. The research focused on in the subtasks is the development and application of state-of the-art technologies to meet the needs of the public, Office of Water,

Bayesian Modeling of NMR Data: Quantifying Longitudinal Relaxation in Vivo, and in Vitro with a Tissue-Water-Relaxation Mimic (Crosslinked Bovine Serum Albumin).

PubMed

Meinerz, Kelsey; Beeman, Scott C; Duan, Chong; Bretthorst, G Larry; Garbow, Joel R; Ackerman, Joseph J H

2018-01-01

Recently, a number of MRI protocols have been reported that seek to exploit the effect of dissolved oxygen (O 2 , paramagnetic) on the longitudinal 1 H relaxation of tissue water, thus providing image contrast related to tissue oxygen content. However, tissue water relaxation is dependent on a number of mechanisms, and this raises the issue of how best to model the relaxation data. This problem, the model selection problem, occurs in many branches of science and is optimally addressed by Bayesian probability theory. High signal-to-noise, densely sampled, longitudinal 1 H relaxation data were acquired from rat brain in vivo and from a cross-linked bovine serum albumin (xBSA) phantom, a sample that recapitulates the relaxation characteristics of tissue water in vivo . Bayesian-based model selection was applied to a cohort of five competing relaxation models: (i) monoexponential, (ii) stretched-exponential, (iii) biexponential, (iv) Gaussian (normal) R 1 -distribution, and (v) gamma R 1 -distribution. Bayesian joint analysis of multiple replicate datasets revealed that water relaxation of both the xBSA phantom and in vivo rat brain was best described by a biexponential model, while xBSA relaxation datasets truncated to remove evidence of the fast relaxation component were best modeled as a stretched exponential. In all cases, estimated model parameters were compared to the commonly used monoexponential model. Reducing the sampling density of the relaxation data and adding Gaussian-distributed noise served to simulate cases in which the data are acquisition-time or signal-to-noise restricted, respectively. As expected, reducing either the number of data points or the signal-to-noise increases the uncertainty in estimated parameters and, ultimately, reduces support for more complex relaxation models.

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.

Optimized approaches for quantification of drug transporters in tissues and cells by MRM proteomics.

PubMed

Prasad, Bhagwat; Unadkat, Jashvant D

2014-07-01

Drug transporter expression in tissues (in vivo) usually differs from that in cell lines used to measure transporter activity (in vitro). Therefore, quantification of transporter expression in tissues and cell lines is important to develop scaling factor for in vitro to in vivo extrapolation (IVIVE) of transporter-mediated drug disposition. Since traditional immunoquantification methods are semiquantitative, targeted proteomics is now emerging as a superior method to quantify proteins, including membrane transporters. This superiority is derived from the selectivity, precision, accuracy, and speed of analysis by liquid chromatography tandem mass spectrometry (LC-MS/MS) in multiple reaction monitoring (MRM) mode. Moreover, LC-MS/MS proteomics has broader applicability because it does not require selective antibodies for individual proteins. There are a number of recent research and review papers that discuss the use of LC-MS/MS for transporter quantification. Here, we have compiled from the literature various elements of MRM proteomics to provide a comprehensive systematic strategy to quantify drug transporters. This review emphasizes practical aspects and challenges in surrogate peptide selection, peptide qualification, peptide synthesis and characterization, membrane protein isolation, protein digestion, sample preparation, LC-MS/MS parameter optimization, method validation, and sample analysis. In particular, bioinformatic tools used in method development and sample analysis are discussed in detail. Various pre-analytical and analytical sources of variability that should be considered during transporter quantification are highlighted. All these steps are illustrated using P-glycoprotein (P-gp) as a case example. Greater use of quantitative transporter proteomics will lead to a better understanding of the role of drug transporters in drug disposition.

Characterization of HBV integration patterns and timing in liver cancer and HBV-infected livers.

PubMed

Furuta, Mayuko; Tanaka, Hiroko; Shiraishi, Yuichi; Unida, Takuro; Imamura, Michio; Fujimoto, Akihiro; Fujita, Masahi; Sasaki-Oku, Aya; Maejima, Kazuhiro; Nakano, Kaoru; Kawakami, Yoshiiku; Arihiro, Koji; Aikata, Hiroshi; Ueno, Masaki; Hayami, Shinya; Ariizumi, Shun-Ichi; Yamamoto, Masakazu; Gotoh, Kunihito; Ohdan, Hideki; Yamaue, Hiroki; Miyano, Satoru; Chayama, Kazuaki; Nakagawa, Hidewaki

2018-05-18

Integration of Hepatitis B virus (HBV) into the human genome can cause genetic instability, leading to selective advantages for HBV-induced liver cancer. Despite the large number of studies for HBV integration into liver cancer, little is known about the mechanism of initial HBV integration events owing to the limitations of materials and detection methods. We conducted an HBV sequence capture, followed by ultra-deep sequencing, to screen for HBV integrations in 111 liver samples from human-hepatocyte chimeric mice with HBV infection and human clinical samples containing 42 paired samples from non-tumorous and tumorous liver tissues. The HBV infection model using chimeric mice verified the efficiency of our HBV-capture analysis and demonstrated that HBV integration could occur 23 to 49 days after HBV infection via microhomology-mediated end joining and predominantly in mitochondrial DNA. Overall HBV integration sites in clinical samples were significantly enriched in regions annotated as exhibiting open chromatin, a high level of gene expression, and early replication timing in liver cells. These data indicate that HBV integration in liver tissue was biased according to chromatin accessibility, with additional selection pressures in the gene promoters of tumor samples. Moreover, an integrative analysis using paired non-tumorous and tumorous samples and HBV-related transcriptional change revealed the involvement of TERT and MLL4 in clonal selection. We also found frequent and non-tumorous liver-specific HBV integrations in FN1 and HBV-FN1 fusion transcript. Extensive survey of HBV integrations facilitates and improves the understanding of the timing and biology of HBV integration during infection and HBV-related hepatocarcinogenesis.

Computational selection of antibody-drug conjugate targets for breast cancer

PubMed Central

Fauteux, François; Hill, Jennifer J.; Jaramillo, Maria L.; Pan, Youlian; Phan, Sieu; Famili, Fazel; O'Connor-McCourt, Maureen

2016-01-01

The selection of therapeutic targets is a critical aspect of antibody-drug conjugate research and development. In this study, we applied computational methods to select candidate targets overexpressed in three major breast cancer subtypes as compared with a range of vital organs and tissues. Microarray data corresponding to over 8,000 tissue samples were collected from the public domain. Breast cancer samples were classified into molecular subtypes using an iterative ensemble approach combining six classification algorithms and three feature selection techniques, including a novel kernel density-based method. This feature selection method was used in conjunction with differential expression and subcellular localization information to assemble a primary list of targets. A total of 50 cell membrane targets were identified, including one target for which an antibody-drug conjugate is in clinical use, and six targets for which antibody-drug conjugates are in clinical trials for the treatment of breast cancer and other solid tumors. In addition, 50 extracellular proteins were identified as potential targets for non-internalizing strategies and alternative modalities. Candidate targets linked with the epithelial-to-mesenchymal transition were identified by analyzing differential gene expression in epithelial and mesenchymal tumor-derived cell lines. Overall, these results show that mining human gene expression data has the power to select and prioritize breast cancer antibody-drug conjugate targets, and the potential to lead to new and more effective cancer therapeutics. PMID:26700623

Next generation sequencing techniques in liquid biopsy: focus on non-small cell lung cancer patients.

PubMed

Malapelle, Umberto; Pisapia, Pasquale; Rocco, Danilo; Smeraglio, Riccardo; di Spirito, Maria; Bellevicine, Claudio; Troncone, Giancarlo

2016-10-01

The advent of genomic based personalized medicine has led to multiple advances in the molecular characterization of many tumor types, such as non-small cell lung cancer (NSCLC). NSCLC is diagnosed in most cases on small tissue samples that may be not always sufficient for EGFR mutational assessment to select patients for first and second generations' tyrosine kinase inhibitors (TKIs) therapy. In patients without tissue availability at presentation, the analysis of cell free DNA (cfDNA) derived from liquid biopsy samples, in particular from plasma, represent an established alternative to provide EGFR mutational testing for treatment decision making. In addition, a new paradigm for TKIs resistance management was recently approved by Food and Drug Administration, supporting the liquid biopsy based genotyping prior to tissue based genotyping for the detection of T790M mutation to select patients for third generation TKIs. In these settings, real time PCR (RT-PCR) and digital PCR 'targeted' methods, which detect known mutations by specific probes, have extensively been adopted. Taking into account the restricted reference range and the limited multiplexing power of these targeted methods, the performance of liquid biopsy analyses may be further improved by next generation sequencing (NGS). While most tissue based NGS genotyping is well established, liquid biopsy NGS application is challenging, requiring a careful validation of the whole process, from blood collection to variant calling. Here we review this evolving field, highlighting those methodological points that are crucial to accurately select NSCLC patients for TKIs treatment administration by NGS on cfDNA.

Discovery of Novel Gene Elements Associated with Prostate Cancer Progression

DTIC Science & Technology

2014-12-01

consent under an Institutional Review Board (IRB) approved protocol at the University of Michigan [SPORE in Prostate Cancer (Tissue/Serum/Urine) Bank IRB...1994-0481]. For the Weill Cornell Medical College patient samples, prostate tissues were collected as part of an IRB- approved protocol at Weill...PCAT-1 or nontargeting short hairpin RNA (shRNA) lentiviral constructs for 48 hours. GFPþ cells were drug -selected using 1 mg/mL puromycin. PCAT-1

Miniature and Molecularly Specific Optical Screening Technologies for Breast Cancer

DTIC Science & Technology

2006-10-01

modeling of the heat dissipation effects of compact LEDs on tissue samples, selection of multiwavelength compact light sources, calculating bandwidth...Opto Technology also designs custom chip on board assemblies with single and multiple wavelengths of UV , Visible and IR LED die (365 – 940 nm...reflectance with high signal to noise for optical properties typical of tissue in the UV -VIS. We have furthermore investigated the potential use of LEDs as

Micro-polarimetry for pre-clinical diagnostics of pathological changes in human tissues

NASA Astrophysics Data System (ADS)

Golnik, Andrzej; Golnik, Natalia; Pałko, Tadeusz; Sołtysiński, Tomasz

2008-05-01

The paper presents a practical study of several methods of image analysis applied to polarimetric images of regular and malignant human tissues. The images of physiological and pathologically changed tissues from body and cervix of uterus, intestine, kidneys and breast were recorded in transmitted light of different polarization state. The set up of the conventional optical microscope with CCD camera and rotating polarizer's were used for analysis of the polarization state of the light transmitted through the tissue slice for each pixel of the camera image. The set of images corresponding to the different coefficients of the Stockes vectors, a 3×3 subset of the Mueller matrix as well as the maps of the magnitude and in-plane direction of the birefringent components in the sample were calculated. Then, the statistical analysis and the Fourier transform as well as the autocorrelation methods were used to analyze spatial distribution of birefringent elements in the tissue samples. For better recognition of tissue state we proposed a novel method that takes advantage of multiscale image data decomposition The results were used for selection of the optical characteristics with significantly different values for regular and malignant tissues.

MO-FG-CAMPUS-JeP2-01: 4D-MRI with 3D Radial Sampling and Self-Gating-Based K-Space Sorting: Image Quality Improvement by Slab-Selective Excitation

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

Deng, Z; Pang, J; Tuli, R

Purpose: A recent 4D MRI technique based on 3D radial sampling and self-gating-based K-space sorting has shown promising results in characterizing respiratory motion. However due to continuous acquisition and potentially drastic k-space undersampling resultant images could suffer from low blood-to-tissue contrast and streaking artifacts. In this study 3D radial sampling with slab-selective excitation (SS) was proposed in attempt to enhance blood-to-tissue contrast by exploiting the in-flow effect and to suppress the excess signal from the peripheral structures particularly in the superior-inferior direction. The feasibility of improving image quality by using this approach was investigated through a comparison with the previouslymore » developed non-selective excitation (NS) approach. Methods: Two excitation approaches SS and NS were compared in 5 cancer patients (1 lung 1 liver 2 pancreas and 1 esophagus) at 3Tesla. Image artifact was assessed in all patients on a 4-point scale (0: poor; 3: excellent). Signal-tonoise ratio (SNR) of the blood vessel (aorta) at the center of field-of-view and its nearby tissue were measured in 3 of the 5 patients (1 liver 2 pancreas) and blood-to-tissue contrast-to-noise ratio (CNR) were then determined. Results: Compared with NS the image quality of SS was visually improved with overall higher signal in all patients (2.6±0.55 vs. 3.4±0.55). SS showed an approximately 2-fold increase of SNR in the blood (aorta: 16.39±1.95 vs. 32.19±7.93) and slight increase in the surrounding tissue (liver/pancreas: 16.91±1.82 vs. 22.31±3.03). As a result the blood-totissue CNR was dramatically higher in the SS method (1.20±1.20 vs. 9.87±6.67). Conclusion: The proposed 3D radial sampling with slabselective excitation allows for reduced image artifact and improved blood SNR and blood-to-tissue CNR. The success of this technique could potentially benefit patients with cancerous tumors that have invaded the surrounding blood vessels where radiation therapy is needed to remove tumor from those regions prior to surgical resection. This work is partially supported by NIH R03CA173273; and CTSI core voucher award.« less

A DNA methylation map of human cancer at single base-pair resolution

PubMed Central

Vidal, E; Sayols, S; Moran, S; Guillaumet-Adkins, A; Schroeder, M P; Royo, R; Orozco, M; Gut, M; Gut, I; Lopez-Bigas, N; Heyn, H; Esteller, M

2017-01-01

Although single base-pair resolution DNA methylation landscapes for embryonic and different somatic cell types provided important insights into epigenetic dynamics and cell-type specificity, such comprehensive profiling is incomplete across human cancer types. This prompted us to perform genome-wide DNA methylation profiling of 22 samples derived from normal tissues and associated neoplasms, including primary tumors and cancer cell lines. Unlike their invariant normal counterparts, cancer samples exhibited highly variable CpG methylation levels in a large proportion of the genome, involving progressive changes during tumor evolution. The whole-genome sequencing results from selected samples were replicated in a large cohort of 1112 primary tumors of various cancer types using genome-scale DNA methylation analysis. Specifically, we determined DNA hypermethylation of promoters and enhancers regulating tumor-suppressor genes, with potential cancer-driving effects. DNA hypermethylation events showed evidence of positive selection, mutual exclusivity and tissue specificity, suggesting their active participation in neoplastic transformation. Our data highlight the extensive changes in DNA methylation that occur in cancer onset, progression and dissemination. PMID:28581523

Method for detection of dental caries and periodontal disease using optical imaging

DOEpatents

Nathel, Howard; Kinney, John H.; Otis, Linda L.

1996-01-01

A method for detecting the presence of active and inactive caries in teeth and diagnosing periodontal disease uses non-ionizing radiation with techniques for reducing interference from scattered light. A beam of non-ionizing radiation is divided into sample and reference beams. The region to be examined is illuminated by the sample beam, and reflected or transmitted radiation from the sample is recombined with the reference beam to form an interference pattern on a detector. The length of the reference beam path is adjustable, allowing the operator to select the reflected or transmitted sample photons that recombine with the reference photons. Thus radiation scattered by the dental or periodontal tissue can be prevented from obscuring the interference pattern. A series of interference patterns may be generated and interpreted to locate dental caries and periodontal tissue interfaces.

Detection of hydroxyapatite in calcified cardiovascular tissues.

PubMed

Lee, Jae Sam; Morrisett, Joel D; Tung, Ching-Hsuan

2012-10-01

The objective of this study is to develop a method for selective detection of the calcific (hydroxyapatite) component in human aortic smooth muscle cells in vitro and in calcified cardiovascular tissues ex vivo. This method uses a novel optical molecular imaging contrast dye, Cy-HABP-19, to target calcified cells and tissues. A peptide that mimics the binding affinity of osteocalcin was used to label hydroxyapatite in vitro and ex vivo. Morphological changes in vascular smooth muscle cells were evaluated at an early stage of the mineralization process induced by extrinsic stimuli, osteogenic factors and a magnetic suspension cell culture. Hydroxyapatite components were detected in monolayers of these cells in the presence of osteogenic factors and a magnetic suspension environment. Atherosclerotic plaque contains multiple components including lipidic, fibrotic, thrombotic, and calcific materials. Using optical imaging and the Cy-HABP-19 molecular imaging probe, we demonstrated that hydroxyapatite components could be selectively distinguished from various calcium salts in human aortic smooth muscle cells in vitro and in calcified cardiovascular tissues, carotid endarterectomy samples and aortic valves, ex vivo. Hydroxyapatite deposits in cardiovascular tissues were selectively detected in the early stage of the calcification process using our Cy-HABP-19 probe. This new probe makes it possible to study the earliest events associated with vascular hydroxyapatite deposition at the cellular and molecular levels. This target-selective molecular imaging probe approach holds high potential for revealing early pathophysiological changes, leading to progression, regression, or stabilization of cardiovascular diseases. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Detection of Hydroxyapatite in Calcified Cardiovascular Tissues

PubMed Central

Lee, Jae Sam; Morrisett, Joel D.; Tung, Ching-Hsuan

2012-01-01

Objective The objective of this study is to develop a method for selective detection of the calcific (hydroxyapatite) component in human aortic smooth muscle cells in vitro and in calcified cardiovascular tissues ex vivo. This method uses a novel optical molecular imaging contrast dye, Cy-HABP-19, to target calcified cells and tissues. Methods A peptide that mimics the binding affinity of osteocalcin was used to label hydroxyapatite in vitro and ex vivo. Morphological changes in vascular smooth muscle cells were evaluated at an early stage of the mineralization process induced by extrinsic stimuli, osteogenic factors and a magnetic suspension cell culture. Hydroxyapatite components were detected in monolayers of these cells in the presence of osteogenic factors and a magnetic suspension environment. Results Atherosclerotic plaque contains multiple components including lipidic, fibrotic, thrombotic, and calcific materials. Using optical imaging and the Cy-HABP-19 molecular imaging probe, we demonstrated that hydroxyapatite components could be selectively distinguished from various calcium salts in human aortic smooth muscle cells in vitro and in calcified cardiovascular tissues, carotid endarterectomy samples and aortic valves, ex vivo. Conclusion Hydroxyapatite deposits in cardiovascular tissues were selectively detected in the early stage of the calcification process using our Cy-HABP-19 probe. This new probe makes it possible to study the earliest events associated with vascular hydroxyapatite deposition at the cellular and molecular levels. This target-selective molecular imaging probe approach holds high potential for revealing early pathophysiological changes, leading to progression, regression, or stabilization of cardiovascular diseases. PMID:22877867

Pharmacokinetics and Tissue Distribution Study of Chlorogenic Acid from Lonicerae Japonicae Flos Following Oral Administrations in Rats

PubMed Central

Zhou, Yulu; Zhou, Ting; Pei, Qi; Liu, Shikun; Yuan, Hong

2014-01-01

Chlorogenic acid (ChA) is proposed as the major bioactive compounds of Lonicerae Japonicae Flos (LJF). Forty-two Wistar rats were randomly divided into seven groups to investigate the pharmacokinetics and tissue distribution of ChA, via oral administration of LJF extract, using ibuprofen as internal standard, employing a high performance liquid chromatography in conjunction with tandem mass spectrometry. Analytes were extracted from plasma samples and tissue homogenate by liquid–liquid extraction with acetonitrile, separated on a C 18 column by linear gradient elution, and detected by electrospray ionization mass spectrometry in negative selected multiple reaction monitoring mode. Our results successfully demonstrate that the method has satisfactory selectivity, linearity, extraction recovery, matrix effect, precision, accuracy, and stability. Using noncompartment model to study pharmacokinetics, profile revealed that ChA was rapidly absorbed and eliminated. Tissue study indicated that the highest level was observed in liver, followed by kidney, lung, heart, and spleen. In conclusion, this method was suitable for the study on pharmacokinetics and tissue distribution of ChA after oral administration. PMID:25140190

Distribution of spotted fever group rickettsiae in select tissues of experimentally infected and field-collected Gulf Coast ticks.

PubMed

Edwards, Kristine T; Goddard, Jerome; Varela-Stokes, Andrea

2011-05-01

Salivary glands, midgut, Malpighian tubules, and ovaries were dissected from infected, colony-derived Amblyomma maculatum (Gulf Coast ticks) injected as nymphs with either Rickettsia parkeri (a spotted fever group rickettsia [SFGR]; treatment) or phosphate-buffered saline (negative control). For comparison, similar tissues were dissected from hemolymph-positive, field-collected ticks. Tissues were analyzed by indirect fluorescent antibody (IFA) tests. All phosphate-buffered saline-injected ticks were IFA negative, whereas SFGR were detected by IFA in 100% of the salivary glands and ovaries and 78 and 75% of midgut and Malpighian tubule samples, respectively, of R. parkeri-injected ticks. Nearly 22% (10/46) of the field-collected ticks were hemolymph positive. Of those, SFGR were detected by IFA in 80% of the salivary glands, 67% of the ovaries, and 60% in the midgut and Malpighian tubules. This is the first study to assess the distribution of SFGR in select tissues of A. maculatum ticks.

Correlation between Infectivity and Disease Associated Prion Protein in the Nervous System and Selected Edible Tissues of Naturally Affected Scrapie Sheep

PubMed Central

Chianini, Francesca; Cosseddu, Gian Mario; Steele, Philip; Hamilton, Scott; Hawthorn, Jeremy; Síso, Sílvia; Pang, Yvonne; Finlayson, Jeanie; Eaton, Samantha L.; Reid, Hugh W.; Dagleish, Mark P.; Di Bari, Michele Angelo; D’Agostino, Claudia; Agrimi, Umberto; Terry, Linda; Nonno, Romolo

2015-01-01

The transmissible spongiform encephalopathies (TSEs) or prion diseases are a group of fatal neurodegenerative disorders characterised by the accumulation of a pathological form of a host protein known as prion protein (PrP). The validation of abnormal PrP detection techniques is fundamental to allow the use of high-throughput laboratory based tests, avoiding the limitations of bioassays. We used scrapie, a prototype TSE, to examine the relationship between infectivity and laboratory based diagnostic tools. The data may help to optimise strategies to prevent exposure of humans to small ruminant TSE material via the food chain. Abnormal PrP distribution/accumulation was assessed by immunohistochemistry (IHC), Western blot (WB) and ELISA in samples from four animals. In addition, infectivity was detected using a sensitive bank vole bioassay with selected samples from two of the four sheep and protein misfolding cyclic amplification using bank vole brain as substrate (vPMCA) was also carried out in selected samples from one animal. Lymph nodes, oculomotor muscles, sciatic nerve and kidney were positive by IHC, WB and ELISA, although at levels 100–1000 fold lower than the brain, and contained detectable infectivity by bioassay. Tissues not infectious by bioassay were also negative by all laboratory tests including PMCA. Although discrepancies were observed in tissues with very low levels of abnormal PrP, there was an overall good correlation between IHC, WB, ELISA and bioassay results. Most importantly, there was a good correlation between the detection of abnormal PrP in tissues using laboratory tests and the levels of infectivity even when the titre was low. These findings provide useful information for risk modellers and represent a first step toward the validation of laboratory tests used to quantify prion infectivity, which would greatly aid TSE risk assessment policies. PMID:25807559

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

Polarization sensitive optical coherence tomography – a review [Invited

PubMed Central

de Boer, Johannes F.; Hitzenberger, Christoph K.; Yasuno, Yoshiaki

2017-01-01

Optical coherence tomography (OCT) is now a well-established modality for high-resolution cross-sectional and three-dimensional imaging of transparent and translucent samples and tissues. Conventional, intensity based OCT, however, does not provide a tissue-specific contrast, causing an ambiguity with image interpretation in several cases. Polarization sensitive (PS) OCT draws advantage from the fact that several materials and tissues can change the light’s polarization state, adding an additional contrast channel and providing quantitative information. In this paper, we review basic and advanced methods of PS-OCT and demonstrate its use in selected biomedical applications. PMID:28663869

Quantifying the motion of magnetic particles in excised tissue: Effect of particle properties and applied magnetic field

NASA Astrophysics Data System (ADS)

Kulkarni, Sandip; Ramaswamy, Bharath; Horton, Emily; Gangapuram, Sruthi; Nacev, Alek; Depireux, Didier; Shimoji, Mika; Shapiro, Benjamin

2015-11-01

This article presents a method to investigate how magnetic particle characteristics affect their motion inside tissues under the influence of an applied magnetic field. Particles are placed on top of freshly excised tissue samples, a calibrated magnetic field is applied by a magnet underneath each tissue sample, and we image and quantify particle penetration depth by quantitative metrics to assess how particle sizes, their surface coatings, and tissue resistance affect particle motion. Using this method, we tested available fluorescent particles from Chemicell of four sizes (100 nm, 300 nm, 500 nm, and 1 μm diameter) with four different coatings (starch, chitosan, lipid, and PEG/P) and quantified their motion through freshly excised rat liver, kidney, and brain tissues. In broad terms, we found that the applied magnetic field moved chitosan particles most effectively through all three tissue types (as compared to starch, lipid, and PEG/P coated particles). However, the relationship between particle properties and their resulting motion was found to be complex. Hence, it will likely require substantial further study to elucidate the nuances of transport mechanisms and to select and engineer optimal particle properties to enable the most effective transport through various tissue types under applied magnetic fields.

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

An enhanced droplet-based liquid microjunction surface sampling system coupled with HPLC-ESI-MS/MS for spatially resolved analysis

DOE PAGES

Van Berkel, Gary J.; Weiskittel, Taylor M.; Kertesz, Vilmos

2014-11-07

Droplet-based liquid microjunction surface sampling coupled with high-performance liquid chromatography (HPLC)-electrospray ionization (ESI)-tandem mass spectrometry (MS/MS) for spatially resolved analysis provides the possibility of effective analysis of complex matrix samples and can provide a greater degree of chemical information from a single spot sample than is typically possible with a direct analysis of an extract. Described here is the setup and enhanced capabilities of a discrete droplet liquid microjunction surface sampling system employing a commercially available CTC PAL autosampler. The system enhancements include incorporation of a laser distance sensor enabling unattended analysis of samples and sample locations of dramatically disparatemore » height as well as reliably dispensing just 0.5 μL of extraction solvent to make the liquid junction to the surface, wherein the extraction spot size was confined to an area about 0.7 mm in diameter; software modifications improving the spatial resolution of sampling spot selection from 1.0 to 0.1 mm; use of an open bed tray system to accommodate samples as large as whole-body rat thin tissue sections; and custom sample/solvent holders that shorten sampling time to approximately 1 min per sample. Lastly, the merit of these new features was demonstrated by spatially resolved sampling, HPLC separation, and mass spectral detection of pharmaceuticals and metabolites from whole-body rat thin tissue sections and razor blade (“crude”) cut mouse tissue.« less

An enhanced droplet-based liquid microjunction surface sampling system coupled with HPLC-ESI-MS/MS for spatially resolved analysis

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

Van Berkel, Gary J.; Weiskittel, Taylor M.; Kertesz, Vilmos

Droplet-based liquid microjunction surface sampling coupled with high-performance liquid chromatography (HPLC)-electrospray ionization (ESI)-tandem mass spectrometry (MS/MS) for spatially resolved analysis provides the possibility of effective analysis of complex matrix samples and can provide a greater degree of chemical information from a single spot sample than is typically possible with a direct analysis of an extract. Described here is the setup and enhanced capabilities of a discrete droplet liquid microjunction surface sampling system employing a commercially available CTC PAL autosampler. The system enhancements include incorporation of a laser distance sensor enabling unattended analysis of samples and sample locations of dramatically disparatemore » height as well as reliably dispensing just 0.5 μL of extraction solvent to make the liquid junction to the surface, wherein the extraction spot size was confined to an area about 0.7 mm in diameter; software modifications improving the spatial resolution of sampling spot selection from 1.0 to 0.1 mm; use of an open bed tray system to accommodate samples as large as whole-body rat thin tissue sections; and custom sample/solvent holders that shorten sampling time to approximately 1 min per sample. Lastly, the merit of these new features was demonstrated by spatially resolved sampling, HPLC separation, and mass spectral detection of pharmaceuticals and metabolites from whole-body rat thin tissue sections and razor blade (“crude”) cut mouse tissue.« less

Review of recent advances in analytical techniques for the determination of neurotransmitters

PubMed Central

Perry, Maura; Li, Qiang; Kennedy, Robert T.

2009-01-01

Methods and advances for monitoring neurotransmitters in vivo or for tissue analysis of neurotransmitters over the last five years are reviewed. The review is organized primarily by neurotransmitter type. Transmitter and related compounds may be monitored by either in vivo sampling coupled to analytical methods or implanted sensors. Sampling is primarily performed using microdialysis, but low-flow push-pull perfusion may offer advantages of spatial resolution while minimizing the tissue disruption associated with higher flow rates. Analytical techniques coupled to these sampling methods include liquid chromatography, capillary electrophoresis, enzyme assays, sensors, and mass spectrometry. Methods for the detection of amino acid, monoamine, neuropeptide, acetylcholine, nucleoside, and soluable gas neurotransmitters have been developed and improved upon. Advances in the speed and sensitivity of these methods have enabled improvements in temporal resolution and increased the number of compounds detectable. Similar advances have enabled improved detection at tissue samples, with a substantial emphasis on single cell and other small samples. Sensors provide excellent temporal and spatial resolution for in vivo monitoring. Advances in application to catecholamines, indoleamines, and amino acids have been prominent. Improvements in stability, sensitivity, and selectivity of the sensors have been of paramount interest. PMID:19800472

Testing the use of juvenile Salmo trutta L. as biomonitors of heavy metal pollution in freshwater.

PubMed

Lamas, S; Fernández, J A; Aboal, J R; Carballeira, A

2007-02-01

Individual specimens of Salmo trutta were captured, from four sampling sites in Galician rivers (NW Spain) affected by different types of contamination: diffuse urban waste, run-off from an unrestored dump at a copper mine and waste from a fish farm. The ages of the captured trouts were established and only those belonging to the 1+ age class were selected for study. The liver and kidney were removed from each fish and analysed to determine the tissue concentrations of Cu, Fe and Zn. The results obtained showed that: (i) the use of 1+ individuals allowed differentiation of contamination scenarios on the basis of the tissue concentrations of metal; (ii) the use of 1+ individuals allowed standardization of the time of exposure, which was sufficiently long for differential uptake to have taken place; (iii) liver tissue provided the best results as, less effort was required than for processing kidney tissue, and significant differences between sampling sites were detected because the intrapopulational variability in metal levels was lower than for kidney, and (iv) the levels of elements detected were not affected by basal tissue concentrations or residual concentrations due to past contamination, which older trouts may have been exposed to. In addition, the use of 1+ trout may provide better results in annual environmental sampling surveys.

Determination of iodine in bread and fish using the iodide ion-selective electrode

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

Steiner, J.B.

The purpose of this study was to assess the potential for use of the ion-selective electrode (ISE) as a method for measuring the iodine content in bread and fish. Ashing methods, sample preparation and electrode responses were evaluated. The iodine values obtained using the iodide electrode were compared to iodine values obtained by the arsenic-cerium method (As-Ce). Ashing methods were used in preparing bread and haddock for iodine analysis by the ISE. The values were compared to unashed samples measured by the ISE. Electrode response to iodide was examined by varying the sample pH, measuring electrode equilibrium times, and comparingmore » direct measurement in ppm to iodide values obtained by the method of known addition. Oyster reference tissue with a known iodine concentration was used to determine rates of recovery. For the As-Ce procedure, an alkaline dry ash for two hour followed by colorimetric analysis at 320 nm was recommended. The study showed that the pre-treatment of bread and fish was necessary for ISE measurement. The iodine values obtained by the ISE in the analysis of oyster reference tissue, haddock and bread were not in agreement with their corresponding As-Ce values. Further work needs to be done to determine an ashing procedure that has minimal iodide loss an/or develop sample treatments that will improve the reliability and precision of iodine values obtained using the ion-selective electrode.« less

Systematic interrogation of diverse Omic data reveals interpretable, robust, and generalizable transcriptomic features of clinically successful therapeutic targets.

PubMed

Rouillard, Andrew D; Hurle, Mark R; Agarwal, Pankaj

2018-05-01

Target selection is the first and pivotal step in drug discovery. An incorrect choice may not manifest itself for many years after hundreds of millions of research dollars have been spent. We collected a set of 332 targets that succeeded or failed in phase III clinical trials, and explored whether Omic features describing the target genes could predict clinical success. We obtained features from the recently published comprehensive resource: Harmonizome. Nineteen features appeared to be significantly correlated with phase III clinical trial outcomes, but only 4 passed validation schemes that used bootstrapping or modified permutation tests to assess feature robustness and generalizability while accounting for target class selection bias. We also used classifiers to perform multivariate feature selection and found that classifiers with a single feature performed as well in cross-validation as classifiers with more features (AUROC = 0.57 and AUPR = 0.81). The two predominantly selected features were mean mRNA expression across tissues and standard deviation of expression across tissues, where successful targets tended to have lower mean expression and higher expression variance than failed targets. This finding supports the conventional wisdom that it is favorable for a target to be present in the tissue(s) affected by a disease and absent from other tissues. Overall, our results suggest that it is feasible to construct a model integrating interpretable target features to inform target selection. We anticipate deeper insights and better models in the future, as researchers can reuse the data we have provided to improve methods for handling sample biases and learn more informative features. Code, documentation, and data for this study have been deposited on GitHub at https://github.com/arouillard/omic-features-successful-targets.

Immunohistochemical findings of the granulomatous reaction associated with tuberculosis.

PubMed

Karimi, Shirin; Shamaei, Masoud; Pourabdollah, Mihan; Sadr, Makan; Karbasi, Mehrdad; Kiani, Arda; Bahadori, Moslem

2016-12-01

The histological diagnosis of Mycobacterium tuberculosis (MTB) has long been a diagnostic challenge in the anatomical pathology field despite availability of different laboratory methods. Immunohistochemistry (IHC) could not only confirm granulomatous tissue involvement but also demonstrate MTB antigen immunolocalization. This study tries to clarify the details of IHC staining for MTB with pAbBCG. A total of 50 patients undergoing simultaneous biopsy and tissue culture with positive tissue culture for MTB during 2005-2009 were selected from the MRC Department at Masih Daneshvari Hospital, Tehran, Iran. Using the archives of the Pathology Department of this hospital, which is a referral center for pathological lung lesions, hematoxylin and eosin slides of the selected patients were evaluated. Twenty-three confirmed TB granulomatous tissue samples with adequate tissue and number of granulomas were chosen and studied by Ziehl-Neelsen and IHC staining with pAbBCG. A total of 23 cases were evaluated, of which 17 (73.9%) were males. The types of tissue obtained from study cases were as follows: pleura (9 cases, 39.1%), lymph node (cervical, axillary, and thoracic [9 cases, 39.1%]), and lung tissues (5 cases, 21.7%). IHC staining was positive in all samples, whereas Ziehl-Neelsen staining was positive in nine cases of 23 (39.1%). IHC showed positive coarse granular cytoplasmic and round, fragmented bacillary staining. In this study, epithelioid cells clearly showed more positive staining at the periphery rather than at the center of granuloma. There is also positive staining in endothelial cells, fibroblasts, plasma cells, macrophages, and lymphocytes outside the granuloma. Detection of TB in tissue slides is still based on the histological pattern of the granuloma, which has several differential diagnoses with different treatments. Presence of mycobacterial antigens and tissue morphology can be evaluated using the IHC technique. Considering the criteria of positive IHC staining of TB granulomatous reactions, this stain not only highlights the presence of mycobacterial antigens for tissue diagnosis, but also could morphologically localize their distribution in different cells. Pathologists must be familiar with adequate staining pattern, elimination of background staining, and type of selected antibody. This method is especially important for application in countries with high prevalence of TB as a technique with early diagnostic value in tissue specimens. Early diagnosis using this technique can reduce related morbidity and mortality and decrease the rate of complications due to misdiagnosis and mistreatment of TB. Copyright © 2016.

Comparative NEXAFS study of the selected icefish hard tissues and hydroxyapatite

NASA Astrophysics Data System (ADS)

Petrova, O. V.; Nekipelov, S. V.; Sivkov, D. V.; Mingaleva, A. E.; Nikolaev, A.; Frank-Kamenetskaya, O. V.; Bazhenov, V. V.; Vyalikh, D. V.; Molodtsov, S. L.; Sivkov, V. N.; Ehrlich, H.

2017-11-01

The structure of native Champsocephalus gunnari icefish otoliths, scales, teeth, bones and pristine hydroxyapatite (HA) were examined using Near Edge X-ray Absorption Fine Structure (NEXAFS) spectroscopy. NEXAFS Cls-absorption spectra of the selected icefish hard tissues indicate that otoliths contain anion [CO3]2-. NEXAFS P2p-spectra clearly indicate the absence of phosphorus atoms only within otoliths and scales samples. However, the icefish teeth and bones P2p-spectra demonstrate identical spectral feature typical for the HA. NEXAFS Ca2p-spectra of the icefish hard tissues studied also shows features, which are in good correspondence with HA spectra. Interestingly, there is a red shift ≈ 0.1 eV of the 2p1/2,3/2 → 3d transition energies in NEXAFS Ca2p-spectra of teethes and bones of the C. gunnari in comparison to HA.

Method for detection of dental caries and periodontal disease using optical imaging

DOEpatents

Nathel, H.; Kinney, J.H.; Otis, L.L.

1996-10-29

A method is disclosed for detecting the presence of active and inactive caries in teeth and diagnosing periodontal disease uses non-ionizing radiation with techniques for reducing interference from scattered light. A beam of non-ionizing radiation is divided into sample and reference beams. The region to be examined is illuminated by the sample beam, and reflected or transmitted radiation from the sample is recombined with the reference beam to form an interference pattern on a detector. The length of the reference beam path is adjustable, allowing the operator to select the reflected or transmitted sample photons that recombine with the reference photons. Thus radiation scattered by the dental or periodontal tissue can be prevented from obscuring the interference pattern. A series of interference patterns may be generated and interpreted to locate dental caries and periodontal tissue interfaces. 7 figs.

Spectral triangulation: a 3D method for locating single-walled carbon nanotubes in vivo

NASA Astrophysics Data System (ADS)

Lin, Ching-Wei; Bachilo, Sergei M.; Vu, Michael; Beckingham, Kathleen M.; Bruce Weisman, R.

2016-05-01

Nanomaterials with luminescence in the short-wave infrared (SWIR) region are of special interest for biological research and medical diagnostics because of favorable tissue transparency and low autofluorescence backgrounds in that region. Single-walled carbon nanotubes (SWCNTs) show well-known sharp SWIR spectral signatures and therefore have potential for noninvasive detection and imaging of cancer tumours, when linked to selective targeting agents such as antibodies. However, such applications face the challenge of sensitively detecting and localizing the source of SWIR emission from inside tissues. A new method, called spectral triangulation, is presented for three dimensional (3D) localization using sparse optical measurements made at the specimen surface. Structurally unsorted SWCNT samples emitting over a range of wavelengths are excited inside tissue phantoms by an LED matrix. The resulting SWIR emission is sampled at points on the surface by a scanning fibre optic probe leading to an InGaAs spectrometer or a spectrally filtered InGaAs avalanche photodiode detector. Because of water absorption, attenuation of the SWCNT fluorescence in tissues is strongly wavelength-dependent. We therefore gauge the SWCNT-probe distance by analysing differential changes in the measured SWCNT emission spectra. SWCNT fluorescence can be clearly detected through at least 20 mm of tissue phantom, and the 3D locations of embedded SWCNT test samples are found with sub-millimeter accuracy at depths up to 10 mm. Our method can also distinguish and locate two embedded SWCNT sources at distinct positions.Nanomaterials with luminescence in the short-wave infrared (SWIR) region are of special interest for biological research and medical diagnostics because of favorable tissue transparency and low autofluorescence backgrounds in that region. Single-walled carbon nanotubes (SWCNTs) show well-known sharp SWIR spectral signatures and therefore have potential for noninvasive detection and imaging of cancer tumours, when linked to selective targeting agents such as antibodies. However, such applications face the challenge of sensitively detecting and localizing the source of SWIR emission from inside tissues. A new method, called spectral triangulation, is presented for three dimensional (3D) localization using sparse optical measurements made at the specimen surface. Structurally unsorted SWCNT samples emitting over a range of wavelengths are excited inside tissue phantoms by an LED matrix. The resulting SWIR emission is sampled at points on the surface by a scanning fibre optic probe leading to an InGaAs spectrometer or a spectrally filtered InGaAs avalanche photodiode detector. Because of water absorption, attenuation of the SWCNT fluorescence in tissues is strongly wavelength-dependent. We therefore gauge the SWCNT-probe distance by analysing differential changes in the measured SWCNT emission spectra. SWCNT fluorescence can be clearly detected through at least 20 mm of tissue phantom, and the 3D locations of embedded SWCNT test samples are found with sub-millimeter accuracy at depths up to 10 mm. Our method can also distinguish and locate two embedded SWCNT sources at distinct positions. Electronic supplementary information (ESI) available: Details concerning instrumental design, experimental procedures, related experiments, and triangulation computations, plus a video showing operation of the scanner. See DOI: 10.1039/c6nr01376g

Quantifying the ultrastructure of carotid arteries using high-resolution micro-diffusion tensor imaging—comparison of intact versus open cut tissue

NASA Astrophysics Data System (ADS)

Salman Shahid, Syed; Gaul, Robert T.; Kerskens, Christian; Flamini, Vittoria; Lally, Caitríona

2017-12-01

Diffusion magnetic resonance imaging (dMRI) can provide insights into the microstructure of intact arterial tissue. The current study employed high magnetic field MRI to obtain ultra-high resolution dMRI at an isotropic voxel resolution of 117 µm3 in less than 2 h of scan time. A parameter selective single shell (128 directions) diffusion-encoding scheme based on Stejskel-Tanner sequence with echo-planar imaging (EPI) readout was used. EPI segmentation was used to reduce the echo time (TE) and to minimise the susceptibility-induced artefacts. The study utilised the dMRI analysis with diffusion tensor imaging (DTI) framework to investigate structural heterogeneity in intact arterial tissue and to quantify variations in tissue composition when the tissue is cut open and flattened. For intact arterial samples, the region of interest base comparison showed significant differences in fractional anisotropy and mean diffusivity across the media layer (p  <  0.05). For open cut flat samples, DTI based directionally invariant indices did not show significant differences across the media layer. For intact samples, fibre tractography based indices such as calculated helical angle and fibre dispersion showed near circumferential alignment and a high degree of fibre dispersion, respectively. This study demonstrates the feasibility of fast dMRI acquisition with ultra-high spatial and angular resolution at 7 T. Using the optimised sequence parameters, this study shows that DTI based markers are sensitive to local structural changes in intact arterial tissue samples and these markers may have clinical relevance in the diagnosis of atherosclerosis and aneurysm.

Improved selection of cortical ovarian strips for autotransplantation of ovarian tissue using full-field optical coherence tomography (FFOCT)

NASA Astrophysics Data System (ADS)

Stegehuis, Paulien L.; Peters, Inge T. A.; Eggermont, Jeroen; Kuppen, Peter J. K.; Trimbos, J. Baptist; Lelieveldt, Boudewijn P. F.; van de Velde, Cornelis J. H.; Bosse, Tjalling; Dijkstra, Jouke; Vahrmeijer, Alexander L.

2016-02-01

Premature ovarian failure is a major concern in women of reproductive age who undergo gonadotoxic cancer treatment. Autotransplantation of frozen-thawed cortical ovarian tissue allows the immediate start of cancer treatment, but risks reintroduction of cancer. Current tumor detection methods compromise the ovarian tissue's viability and can therefore only be used to exclude the presence of metastases in the cortical ovarian strips that are not transplanted. A non-invasive method is needed that can be used to exclude metastases in the actual ovarian autografts without affecting the tissue's viability. In this study we applied FFOCT - a non-fixative technique that uses white light interferometry to make highresolution images (1μm isotropic) of fresh tissue - to study healthy and malignant ovarian tissue. We created an image atlas of healthy ovarian tissues from premenopausal patients and ovarian tissues with breast cancer metastases. To get the best possible match between hematoxylin-and-eosin stained slides and FFOCT images formalinfixed paraffin-embedded tissue samples were deparaffinized and FFOCT images were acquired within a few minutes. FFOCT images were compared with histology images. All normal structures such as follicles in all phases, inclusion cysts, blood vessels, corpora lutea, and corpora albicantia were clearly recognizable. Ovarian metastases could be well distinguished from normal ovarian tissue. FFOCT is a promising technique in the field of fertility preservation: metastases can be detected and additionally cortical ovarian strips can be selected on the basis of high follicle density.

Validation of an LC-MS/MS method for the quantification of choline-related compounds and phospholipids in foods and tissues.

PubMed

Xiong, Yeping; Zhao, Yuan-Yuan; Goruk, Sue; Oilund, Kirsten; Field, Catherine J; Jacobs, René L; Curtis, Jonathan M

2012-12-12

A hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC LC-MS/MS) method was developed and validated to simultaneously quantify six aqueous choline-related compounds and eight major phospholipids classes in a single run. HILIC chromatography was coupled to positive ion electrospray mass spectrometry. A combination of multiple scan modes including precursor ion scan, neutral loss scan and multiple reaction monitoring was optimized for the determination of each compound or class in a single LC/MS run. This work developed a simplified extraction scheme in which both free choline and related compounds along with phospholipids were extracted into a homogenized phase using chloroform/methanol/water (1:2:0.8) and diluted into methanol for the analysis of target compounds in a variety of sample matrices. The analyte recoveries were evaluated by spiking tissues and food samples with two isotope-labeled internal standards, PC-d(3) and Cho-d(3). Recoveries of between 90% and 115% were obtained by spiking a range of sample matrices with authentic standards containing all 14 of the target analytes. The precision of the analysis ranged from 1.6% to 13%. Accuracy and precision was comparable to that obtained by quantification of selected phospholipid classes using (31)P NMR. A variety of sample matrices including egg yolks, human diets and animal tissues were analyzed using the validated method. The measurements of total choline in selected foods were found to be in good agreement with values obtained from the USDA choline database. Copyright © 2012 Elsevier B.V. All rights reserved.

Transgenic zebrafish reveal tissue-specific differences in estrogen signaling in response to environmental water samples.

PubMed

Gorelick, Daniel A; Iwanowicz, Luke R; Hung, Alice L; Blazer, Vicki S; Halpern, Marnie E

2014-04-01

Environmental endocrine disruptors (EEDs) are exogenous chemicals that mimic endogenous hormones such as estrogens. Previous studies using a zebrafish transgenic reporter demonstrated that the EEDs bisphenol A and genistein preferentially activate estrogen receptors (ERs) in the larval heart compared with the liver. However, it was not known whether the transgenic zebrafish reporter was sensitive enough to detect estrogens from environmental samples, whether environmental estrogens would exhibit tissue-specific effects similar to those of BPA and genistein, or why some compounds preferentially target receptors in the heart. We tested surface water samples using a transgenic zebrafish reporter with tandem estrogen response elements driving green fluorescent protein expression (5xERE:GFP). Reporter activation was colocalized with tissue-specific expression of ER genes by RNA in situ hybridization. We observed selective patterns of ER activation in transgenic fish exposed to river water samples from the Mid-Atlantic United States, with several samples preferentially activating receptors in embryonic and larval heart valves. We discovered that tissue specificity in ER activation was due to differences in the expression of ER subtypes. ERα was expressed in developing heart valves but not in the liver, whereas ERβ2 had the opposite profile. Accordingly, subtype-specific ER agonists activated the reporter in either the heart valves or the liver. The use of 5xERE:GFP transgenic zebrafish revealed an unexpected tissue-specific difference in the response to environmentally relevant estrogenic compounds. Exposure to estrogenic EEDs in utero was associated with adverse health effects, with the potentially unanticipated consequence of targeting developing heart valves.

Transgenic zebrafish reveal tissue-specific differences in estrogen signaling in response to environmental water samples

USGS Publications Warehouse

Gorelick, Daniel A.; Iwanowicz, Luke R.; Hung, Alice L.; Blazer, Vicki; Halpern, Marnie E.

2014-01-01

Background: Environmental endocrine disruptors (EED) are exogenous chemicals that mimic endogenous hormones, such as estrogens. Previous studies using a zebrafish transgenic reporter demonstrated that the EEDs bisphenol A and genistein preferentially activate estrogen receptors (ER) in the larval heart compared to the liver. However, it was not known whether the transgenic zebrafish reporter was sensitive enough to detect estrogens from environmental samples, whether environmental estrogens would exhibit similar tissue-specific effects as BPA and genistein or why some compounds preferentially target receptors in the heart. Methods: We tested surface water samples using a transgenic zebrafish reporter with tandem estrogen response elements driving green fluorescent protein expression (5xERE:GFP). Reporter activation was colocalized with tissue-specific expression of estrogen receptor genes by RNA in situ hybridization. Results: Selective patterns of ER activation were observed in transgenic fish exposed to river water samples from the Mid-Atlantic United States, with several samples preferentially activating receptors in embryonic and larval heart valves. We discovered that tissue-specificity in ER activation is due to differences in the expression of estrogen receptor subtypes. ERα is expressed in developing heart valves but not in the liver, whereas ERβ2 has the opposite profile. Accordingly, subtype-specific ER agonists activate the reporter in either the heart valves or the liver. Conclusion: The use of 5xERE:GFP transgenic zebrafish has revealed an unexpected tissue-specific difference in the response to environmentally relevant estrogenic compounds. Exposure to estrogenic EEDs in utero is associated with adverse health effects, with the potentially unanticipated consequence of targeting developing heart valves.

Screening spectroscopy of prostate cancer

NASA Astrophysics Data System (ADS)

Yermolenko, S. B.; Voloshynskyy, D. I.; Fedoruk, O. S.

2015-11-01

The aim of the study was to establish objective parameters of the field of laser and incoherent radiation of different spectral ranges (UV, visible, IR) as a non-invasive optical method of interaction with different samples of biological tissues and fluids of patients to determine the state of prostate cancer and choosing the best personal treatment. The objects of study were selected venous blood plasma of patient with prostate cancer, histological sections of rat prostate gland in the postoperative period. As diagnostic methods have been used ultraviolet spectrometry samples of blood plasma in the liquid state, infrared spectroscopy middle range (2,5-25 microns) dry residue of plasma by spectral diagnostic technique of thin histological sections of biological tissues.

Methyl sulfonyl polychlorinated biphenyls and 2,2-bis(4-chlorophenyl)-1,1-dichlorethene in gray seal tissues determined by gas chromatography with electron capture detection and atomic emission detection

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

Janak, K.; Becker, G.; Colmisjoe, A.

1998-06-01

The presence of 24 methyl sulfonyl polychlorinated biphenyl (PCB) congeners (MeSO{sub 2}-CBs) and 3-methyl sulfonyl 2,2-bis(4-chlorophenyl)-1,1-dichlorethene (DDE) (MeSO{sub 2}-DDE), metabolites of PCB and DDE, in blubber, lung, and liver of gray seals has been determined by using atomic emission detection (AED) and electron capture detection (ECD). Selective accumulation of aryl methyl sulfones in blubber, liver, and lung tissue was also investigated. For the liver samples, a substantial and highly specific retention of PCB methyl sulfones was observed. The atomic emission technique significantly improved the determination of measured solutes compared with ECD. Atomic emission detection was also valuable for the monitoringmore » of the prefractionation and to decrease the requirements of sample clean-up. Comparing both detection techniques showed a good correlation between the results of the AED sulfur-selective line and ECD.« less

Evaluation of Protein Profiles From Treated Xenograft Tumor Models Identifies an Antibody Panel for Formalin-fixed and Paraffin-embedded (FFPE) Tissue Analysis by Reverse Phase Protein Arrays (RPPA)*

PubMed Central

Bader, Sabine; Zajac, Magdalena; Friess, Thomas; Ruge, Elisabeth; Rieder, Natascha; Gierke, Berthold; Heubach, Yvonne; Thomas, Marlene; Pawlak, Michael

2015-01-01

Reverse phase protein arrays (RPPA) are an established tool for measuring the expression and activation status of multiple proteins in parallel using only very small amounts of tissue. Several studies have demonstrated the value of this technique for signaling pathway analysis using proteins extracted from fresh frozen (FF) tissue in line with validated antibodies for this tissue type; however, formalin fixation and paraffin embedding (FFPE) is the standard method for tissue preservation in the clinical setting. Hence, we performed RPPA to measure profiles for a set of 300 protein markers using matched FF and FFPE tissue specimens to identify which markers performed similarly using the RPPA technique in fixed and unfixed tissues. Protein lysates were prepared from matched FF and FFPE tissue specimens of individual tumors taken from three different xenograft models of human cancer. Materials from both untreated mice and mice treated with either anti-HER3 or bispecific anti-IGF-1R/EGFR monoclonal antibodies were analyzed. Correlations between signals from FF and FFPE tissue samples were investigated. Overall, 60 markers were identified that produced comparable profiles between FF and FFPE tissues, demonstrating significant correlation between the two sample types. The top 25 markers also showed significance after correction for multiple testing. The panel of markers covered several clinically relevant tumor signaling pathways and both phosphorylated and nonphosphorylated proteins were represented. Biologically relevant changes in marker expression were noted when RPPA profiles from treated and untreated xenografts were compared. These data demonstrate that, using appropriately selected antibodies, RPPA analysis from FFPE tissue is well feasible and generates biologically meaningful information. The identified panel of markers that generate similar profiles in matched fixed and unfixed tissue samples may be clinically useful for pharmacodynamic studies of drug effect using FFPE tissues. PMID:26106084

Automated Liquid Microjunction Surface Sampling-HPLC-MS/MS 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

A fully automated liquid extraction-based surface sampling system utilizing a commercially available autosampler coupled to high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) detection is reported. Discrete spots selected for droplet-based sampling and automated sample queue generation for both the autosampler and MS were enabled by using in-house developed software. In addition, co-registration of spatially resolved sampling position and HPLC-MS information to generate heatmaps of compounds monitored for subsequent data analysis was also available in the software. The system was evaluated with whole-body thin tissue sections from propranolol dosed rat. The hands-free operation of the system was demonstrated by creating heatmapsmore » of the parent drug and its hydroxypropranolol glucuronide metabolites with 1 mm resolution in the areas of interest. The sample throughput was approximately 5 min/sample defined by the time needed for chromatographic separation. The spatial distributions of both the drug and its metabolites were consistent with previous studies employing other liquid extraction-based surface sampling methodologies.« less

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.

Examining the occurrence of residues of flunixin meglumine in cull dairy cows by use of the flunixin cull cow survey.

PubMed

Deyrup, Cynthia L; Southern, Kristal J; Cornett, Julie A; Shultz, Craig E; Cera, Deborah A

2012-07-15

To determine whether cull dairy cows with signs of certain clinical conditions, termed suspect, are more likely than healthy-appearing cull dairy cows to have violative concentrations of flunixin meglumine in their tissues at slaughter. Cross-sectional study. 961 cull dairy cows. Suspect cull dairy cows were selected from 21 beef slaughter establishments with a high production volume of dairy cows, and kidney and liver tissues were collected for screening. Kidney tissues were screened for antibiotics and sulfonamides with the fast antimicrobial screening test (FAST). Liver tissues were screened for flunixin meglumine with an ELISA, and quantitative analysis of ELISA-positive samples was performed with high-performance liquid chromatography. During the same time period, liver tissues from 251 healthy-appearing cull dairy cows were collected for the Food Safety and Inspection Service National Residue Program Scheduled Sampling Plan, but were screened only for flunixin meglumine. Of 710 suspect cull dairy cows, 50 (7.04%) had liver tissue flunixin concentrations higher than the flunixin tolerance concentration (0.125 ppm). Thirty-one of 168 (18.45%) FAST-positive and 19 of 542 (3.51%) FAST-negative suspect cull dairy cows had violative tissue flunixin concentrations. Two of the 251 (0.80%) healthy-appearing cull dairy cows had violative tissue flunixin concentrations. Suspect cull dairy cows, especially those that were also FAST positive, had a significantly higher incidence of violative tissue flunixin concentrations than healthy-appearing cull dairy cows at slaughter. Targeted sampling plans for flunixin meglumine in suspect dairy cows can help to support more efficient use of resources and further safeguard the nation's food supply.

A novel agar formulation for isolation and direct enumeration of Vibrio vulnificus from oyster tissue.

PubMed

Griffitt, Kimberly J; Grimes, D Jay

2013-08-01

A new selective and differential medium, Vibrio vulnificus X-Gal (VVX), was developed for direct enumeration of V. vulnificus (Vv) from oyster samples. This agar utilizes cellobiose and lactose as carbon sources, and the antibiotics colistin and polymyxin B as selective agents. Hydrolysis of 5-bromo-4-chloro-3-indolyl- beta-d-galactopyranoside (x-gal), used in the agar as a lactose analog, produces an insoluble blue dye that makes lactose positive colonies easily distinguishable from any non-lactose fermenting bacteria. Various bacterial species were spot plated onto thiosulfate-citrate-bile salts-sucrose agar (TCBS), and CHROMagar Vibrio, two vibrio-specific selective agars, non-selective agar, and VVX to compare selectivity of VVX to other widely used media. A V. vulnificus pure culture was serially diluted on VVX and non-selective agar to determine the VVX percent recovery. Water and oyster samples were spread plated on VVX agar and allowed to incubate for 16-18 h at 33 °C. Blue and white colonies from VVX agar were picked and screened by end point PCR for the Vv hemolysin vvhA. VVX agar showed a significant improvement over TCBS and CHROMagar at preventing non-target growth. There was an 87.5% recovery compared to non-selective plating and a 98% positivity rate of blue colonies picked from oyster tissue plating. The findings suggest that this new agar is a fast, distinctive, and accurate method for enumeration of V. vulnificus from the environment. Copyright © 2013 Elsevier B.V. All rights reserved.

Blood from a turnip: tissue origin of low-coverage shotgun sequencing libraries affects recovery of mitogenome sequences

USGS Publications Warehouse

Barker, F. Keith; Oyler-McCance, Sara; Tomback, Diana F.

2015-01-01

Next generation sequencing methods allow rapid, economical accumulation of data that have many applications, even at relatively low levels of genome coverage. However, the utility of shotgun sequencing data sets for specific goals may vary depending on the biological nature of the samples sequenced. We show that the ability to assemble mitogenomes from three avian samples of two different tissue types varies widely. In particular, data with coverage typical of microsatellite development efforts (∼1×) from DNA extracted from avian blood failed to cover even 50% of the mitogenome, relative to at least 500-fold coverage from muscle-derived data. Researchers should consider possible applications of their data and select the tissue source for their work accordingly. Practitioners analyzing low-coverage shotgun sequencing data (including for microsatellite locus development) should consider the potential benefits of mitogenome assembly, including internal barcode verification of species identity, mitochondrial primer development, and phylogenetics.

Modelling and characterization of photothermal effects assisted with gold nanorods in ex vivo samples and in a murine model

NASA Astrophysics Data System (ADS)

Lamela Rivera, Horacio; Rodríguez Jara, Félix; Cunningham, Vincent

2011-03-01

We discuss in this article the implementation of a laser-tissue interaction and bioheat-transfer 2-D finite-element model for Photothermal Therapy assisted with Gold Nanorods. We have selected Gold Nanorods as absorbing nanostructures in order to improve the efficiency of using compact diode lasers because of their high opto-thermal conversion efficiency at 808 and 850 nm. The goal is to model the distribution of the optical energy among the tissue including the skin absorption effects and the tissue thermal response, with and without the presence of Gold Nanorods. The heat generation due to the optical energy absorption and the thermal propagation will be computationally modeled and optimized. The model has been evaluated and compared with experimental ex-vivo data in fresh chicken muscle samples and in-vivo BALB/c mice animal model.

Concurrent Isolation of 3 Distinct Cardiac Stem Cell Populations From a Single Human Heart Biopsy.

PubMed

Monsanto, Megan M; White, Kevin S; Kim, Taeyong; Wang, Bingyan J; Fisher, Kristina; Ilves, Kelli; Khalafalla, Farid G; Casillas, Alexandria; Broughton, Kathleen; Mohsin, Sadia; Dembitsky, Walter P; Sussman, Mark A

2017-07-07

The relative actions and synergism between distinct myocardial-derived stem cell populations remain obscure. Ongoing debates on optimal cell population(s) for treatment of heart failure prompted implementation of a protocol for isolation of multiple stem cell populations from a single myocardial tissue sample to develop new insights for achieving myocardial regeneration. Establish a robust cardiac stem cell isolation and culture protocol to consistently generate 3 distinct stem cell populations from a single human heart biopsy. Isolation of 3 endogenous cardiac stem cell populations was performed from human heart samples routinely discarded during implantation of a left ventricular assist device. Tissue explants were mechanically minced into 1 mm 3 pieces to minimize time exposure to collagenase digestion and preserve cell viability. Centrifugation removes large cardiomyocytes and tissue debris producing a single cell suspension that is sorted using magnetic-activated cell sorting technology. Initial sorting is based on tyrosine-protein kinase Kit (c-Kit) expression that enriches for 2 c-Kit + cell populations yielding a mixture of cardiac progenitor cells and endothelial progenitor cells. Flowthrough c-Kit - mesenchymal stem cells are positively selected by surface expression of markers CD90 and CD105. After 1 week of culture, the c-Kit + population is further enriched by selection for a CD133 + endothelial progenitor cell population. Persistence of respective cell surface markers in vitro is confirmed both by flow cytometry and immunocytochemistry. Three distinct cardiac cell populations with individualized phenotypic properties consistent with cardiac progenitor cells, endothelial progenitor cells, and mesenchymal stem cells can be successfully concurrently isolated and expanded from a single tissue sample derived from human heart failure patients. © 2017 American Heart Association, Inc.

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.

Simultaneous isolation of high-quality DNA, RNA, miRNA and proteins from tissues for genomic applications

PubMed Central

Peña-Llopis, Samuel; Brugarolas, James

2014-01-01

Genomic technologies have revolutionized our understanding of complex Mendelian diseases and cancer. Solid tumors present several challenges for genomic analyses, such as tumor heterogeneity and tumor contamination with surrounding stroma and infiltrating lymphocytes. We developed a protocol to (i) select tissues of high cellular purity on the basis of histological analyses of immediately flanking sections and (ii) simultaneously extract genomic DNA (gDNA), messenger RNA (mRNA), noncoding RNA (ncRNA; enriched in microRNA (miRNA)) and protein from the same tissues. After tissue selection, about 12–16 extractions of DNA/RNA/protein can be obtained per day. Compared with other similar approaches, this fast and reliable methodology allowed us to identify mutations in tumors with remarkable sensitivity and to perform integrative analyses of whole-genome and exome data sets, DNA copy numbers (by single-nucleotide polymorphism (SNP) arrays), gene expression data (by transcriptome profiling and quantitative PCR (qPCR)) and protein levels (by western blotting and immunohistochemical analysis) from the same samples. Although we focused on renal cell carcinoma, this protocol may be adapted with minor changes to any human or animal tissue to obtain high-quality and high-yield nucleic acids and proteins. PMID:24136348

Transgenic Zebrafish Reveal Tissue-Specific Differences in Estrogen Signaling in Response to Environmental Water Samples

PubMed Central

Iwanowicz, Luke R.; Hung, Alice L.; Blazer, Vicki S.; Halpern, Marnie E.

2014-01-01

Background: Environmental endocrine disruptors (EEDs) are exogenous chemicals that mimic endogenous hormones such as estrogens. Previous studies using a zebrafish transgenic reporter demonstrated that the EEDs bisphenol A and genistein preferentially activate estrogen receptors (ERs) in the larval heart compared with the liver. However, it was not known whether the transgenic zebrafish reporter was sensitive enough to detect estrogens from environmental samples, whether environmental estrogens would exhibit tissue-specific effects similar to those of BPA and genistein, or why some compounds preferentially target receptors in the heart. Methods: We tested surface water samples using a transgenic zebrafish reporter with tandem estrogen response elements driving green fluorescent protein expression (5xERE:GFP). Reporter activation was colocalized with tissue-specific expression of ER genes by RNA in situ hybridization. Results: We observed selective patterns of ER activation in transgenic fish exposed to river water samples from the Mid-Atlantic United States, with several samples preferentially activating receptors in embryonic and larval heart valves. We discovered that tissue specificity in ER activation was due to differences in the expression of ER subtypes. ERα was expressed in developing heart valves but not in the liver, whereas ERβ2 had the opposite profile. Accordingly, subtype-specific ER agonists activated the reporter in either the heart valves or the liver. Conclusion: The use of 5xERE:GFP transgenic zebrafish revealed an unexpected tissue-specific difference in the response to environmentally relevant estrogenic compounds. Exposure to estrogenic EEDs in utero was associated with adverse health effects, with the potentially unanticipated consequence of targeting developing heart valves. Citation: Gorelick DA, Iwanowicz LR, Hung AL, Blazer VS, Halpern ME. 2014. Transgenic zebrafish reveal tissue-specific differences in estrogen signaling in response to environmental water samples. Environ Health Perspect 122:356–362; http://dx.doi.org/10.1289/ehp.1307329 PMID:24425189

Utility of cytopathological specimens and an automated image analysis for the evaluation of HER2 status and intratumor heterogeneity in breast carcinoma.

PubMed

Arihiro, Koji; Oda, Miyo; Ogawa, Katsunari; Kaneko, Yoshie; Shimizu, Tomomi; Tanaka, Yuna; Marubashi, Yukari; Ishida, Katsunari; Takai, Chikako; Taoka, Chie; Kimura, Shuji; Shiroma, Noriyuki

2016-12-01

Although updated HER2 testing guidelines have been improved by a collaboration between the American Society of Clinical Oncology (ASCO) and the College of American Pathologists (CAP) in 2013, HER2 evaluation is still problematic because of issues involving CEP17 polysomy, heterogeneity, and HER2 score 2+ cases. The aim of this retrospective study was to evaluate the relationship between HER2 gene heterogeneity, or so called CEP17 polysomy, using breast carcinoma cells sampled by scraping and the IHC score graded by automated image analysis using whole slide image. We randomly selected 23 breast carcinoma cases with a HER2 score 0, 24 cases with a HER2 score 1+, 24 cases with HER2 score 2+, and 23 cases with HER2 score 3+ from the records of patients with breast cancer at Hiroshima University Hospital. We compared the results of fluorescent in situ hybridization (FISH) using formalin-fixed, paraffin-embedded (FFPE) tissues and cytological samples and compared the HER2 score calculated using an automated image analysis using wholly scanned slide images and visual counting. We successfully performed the FISH assay in 78 of 94 cases (83%) using FFPE tissues and in all 94 (100%) cases using cytological samples. Frequency of both HER2 amplification and CEP17 polysomy was higher when cytological samples were used than when FFPE tissue was used. Frequency of HER2 heterogeneity using cytological samples was higher that than using FFPE tissue, except for the IHC score 3+ cases. When assessment of HER2 status based on FISH using FFPE tissue cannot be accomplished, FISH using cytological samples should be considered. When intensity of HER2 is heterogeneous in the tumor tissue, particularly in cases regarded as score 2+, they should be evaluated by automated image analysis using the whole slide image. Copyright © 2016 Elsevier GmbH. All rights reserved.

Application of solid phase microextraction followed by liquid chromatography-mass spectrometry in the determination of antibiotic drugs and their metabolites in human whole blood and tissue samples.

PubMed

Szultka-Mlynska, Malgorzata; Pomastowski, Pawel; Buszewski, Boguslaw

2018-06-01

A sensitive, rapid and specific analytical method using high performance liquid chromatography coupled with mass spectrometry (HPLC-QqQ-MS) was developed to determine selected antibiotic drugs and their metabolites (amoxicillin, cefotaxime, ciprofloxacin, clindamycin and metronidazole; amoxycilloic acid, 4-hydroxyphenyl glycyl amoxicillin, desacetyl cefotaxime, 3-desacetyl cefotaxime lactone, ciprofloxacin N-oxide, N-demethylclindamycin, clindamycin sulfoxide, and hydroxy metronidazole) in human whole blood and vascularized tissue after single oral administration. The samples were prepared by solid phase microextraction with C18 fibers (SPME C18 ) and determined on a GRACE analytical C18 column, Vision HT (50 × 2 mm, 1.5 μm) at the flow rate of 0.4 mL min -1 using water and acetonitrile (containing 0.1% formic acid) as the mobile phase. The proposed method was successfully applied in a pharmacokinetic study of the selected antibiotic drugs and their metabolites in real human samples. Additionally, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI/TOF-MS) was used for identification and qualification analysis of the target compounds. Copyright © 2018 Elsevier B.V. All rights reserved.

Bio-monitoring of Tissue Accumulation and Genotoxic Effect of Heavy Metals in Cyprinus carpio from River Kabul Khyber Pakhtunkhwa Pakistan.

PubMed

Siraj, Muhammad; Khisroon, Muhammad; Khan, Ajmal; Zaidi, Farrah; Ullah, Ahmad; Rahman, Ghani

2018-03-01

The study explored (I) the concentration of heavy metals in water samples (II) their bioaccumulation in common carp Cyprinus carpio (III) and the subsequent genotoxicity in the selected organs of carp; from river Kabul, Khyber Pakhtunkhwa Pakistan. Except for Mercury (Hg) the water samples had all the heavy metals within permissible limits of recommended dietary allowance (RDA). Nonetheless a number of heavy metals (Zn, Ni, Cr, Cd, Pb and Hg) showed bioaccumulation at levels higher than permissible. Zinc (Zn) was the most while Cadmium (Cd) was the least accumulated metal in all tissue samples analyzed. The metal burden in different organs of C. carpio was in sequence of intestine > skin > liver > gills > muscle. The Comet assay established DNA damage in selected organs to be in accordance with metal burden; the most to least damage being in sequence of blood > intestine > skin > liver > gills > muscle. In conclusion assessment of DNA damage in the organs of C. carpio appears to be a useful bio-marker to evaluate genotoxic effects of heavy metal pollution.

Contaminants in fish tissue from US lakes and reservoirs: A ...

EPA Pesticide Factsheets

An unequal probability design was used to develop national estimates for 268 persistent, bioaccumulative, and toxic chemicals in fish tissue from lakes and reservoirs of the conterminous United States (excluding the Laurentian Great Lakes and Great Salt Lake). Predator (fillet) and bottom-dweller (whole-body) composites were collected from 500 lakes selected randomly from the target population of 147,343 lakes in the lower 48 states. Each of these composite types comprised nationally representative samples whose results were extrapolated to the sampled population of an estimated 76,559 lakes for predators and 46,190 lakes for bottom dwellers. Mercury and PCBs were detected in all fish samples. Dioxins and furans were detected in 81% and 99% of predator and bottom-dweller samples, respectively. Cumulative frequency distributions showed that mercury concentrations exceeded the EPA 300 ppb mercury fish tissue criterion at nearly half of the lakes in the sampled population. Total PCB concentrations exceeded a 12 ppb human health risk-based consumption limit at nearly 17% of lakes, and dioxins and furans exceeded a 0.15 ppt (toxic equivalent or TEQ) risk-based threshold at nearly 8% of lakes in the sampled population. In contrast, 43 target chemicals were not detected in any samples. No detections were reported for nine organophosphate pesticides, one PCB congener, 16 polycyclic aromatic hydrocarbons, or 17 other semivolatile organic chemicals. An unequal prob

Contaminants in fish tissue from US lakes and reservoirs: a national probabilistic study.

PubMed

Stahl, Leanne L; Snyder, Blaine D; Olsen, Anthony R; Pitt, Jennifer L

2009-03-01

An unequal probability design was used to develop national estimates for 268 persistent, bioaccumulative, and toxic chemicals in fish tissue from lakes and reservoirs of the conterminous United States (excluding the Laurentian Great Lakes and Great Salt Lake). Predator (fillet) and bottom-dweller (whole body) composites were collected from 500 lakes selected randomly from the target population of 147,343 lakes in the lower 48 states. Each of these composite types comprised nationally representative samples whose results were extrapolated to the sampled population of an estimated 76,559 lakes for predators and 46,190 lakes for bottom dwellers. Mercury and PCBs were detected in all fish samples. Dioxins and furans were detected in 81% and 99% of predator and bottom-dweller samples, respectively. Cumulative frequency distributions showed that mercury concentrations exceeded the EPA 300 ppb mercury fish tissue criterion at nearly half of the lakes in the sampled population. Total PCB concentrations exceeded a 12 ppb human health risk-based consumption limit at nearly 17% of lakes, and dioxins and furans exceeded a 0.15 ppt (toxic equivalent or TEQ) risk-based threshold at nearly 8% of lakes in the sampled population. In contrast, 43 target chemicals were not detected in any samples. No detections were reported for nine organophosphate pesticides, one PCB congener, 16 polycyclic aromatic hydrocarbons, or 17 other semivolatile organic chemicals.

The Ecology and Evolution of Cancer: The Ultra-Microevolutionary Process.

PubMed

Wu, Chung-I; Wang, Hurng-Yi; Ling, Shaoping; Lu, Xuemei

2016-11-23

Although tumorigenesis has been accepted as an evolutionary process ( 20 , 102 ), many forces may operate differently in cancers than in organisms, as they evolve at vastly different time scales. Among such forces, natural selection, here defined as differential cellular proliferation among distinct somatic cell genotypes, is particularly interesting because its action might be thwarted in multicellular organisms ( 20 , 29 ). In this review, selection is analyzed in two stages of cancer evolution: Stage I is the evolution between tumors and normal tissues, and Stage II is the evolution within tumors. The Cancer Genome Atlas (TCGA) data show a low degree of convergent evolution in Stage I, where genetic changes are not extensively shared among cases. An equally important, albeit much less highlighted, discovery using TCGA data is that there is almost no net selection in cancer evolution. Both positive and negative selection are evident but they neatly cancel each other out, rendering total selection ineffective in the absence of recombination. The efficacy of selection is even lower in Stage II, where neutral (non-Darwinian) evolution is increasingly supported by high-density sampling studies ( 81 , 123 ). Because natural selection is not a strong deterministic force, cancers usually evolve divergently even in similar tissue environments.

Selectively Adsorptive Extraction of Phenylarsonic Acids in Chicken Tissue by Carboxymethyl α-Cyclodextrin Immobilized Fe3O4 Magnetic Nanoparticles Followed Ultra Performance Liquid Chromatography Coupled Tandem Mass Spectrometry Detection

PubMed Central

Jia, Jing; Zhang, Wei; Wang, Jing; Wang, Peilong; Zhu, Ruohua

2014-01-01

Carboxymethyl α-cyclodextrin immobilized Fe3O4 magnetic nanoparticles (CM-α-CD-Fe3O4) were synthesized for the selectively adsorptive extraction of five phenylarsonic acids including p-amino phenylarsonic acid, p-nitro phenylarsonic acid, p-hydroxy phenylarsonic acid, p-acylamino phenylarsonic acid and p-hydroxy-3-nitro phenylarsonic acid in chicken tissue. Using ultra performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS), a highly sensitive analytical method was proposed for the determination of five phenylarsonic acids. It was shown that CM-α-CD-Fe3O4 could extract the five phenylarsonic acids in complex chicken tissue samples with high extraction efficiency. Under the optimal conditions, a high enrichment factor, ranging from 349 to 606 fold, was obtained. The limits of detection (LODs) (at a signal-to-noise ratio of 3) were in the range of 0.05–0.11 µg/kg for the five phenylarsonic acids. The proposed method was applied for the determination of five target phenylarsonic acids in chicken muscle and liver samples. Recoveries for the spiked samples with 0.2 µg/kg, 2.0 µg/kg and 20 µg/kg of each phenylarsonic acids were in the range of 77.2%–110.2%, with a relative standard deviation (RSD) of less than 12.5%. PMID:25215503

Development of a new paper based nano-biosensor using the co-catalytic effect of tyrosinase from banana peel tissue (Musa Cavendish) and functionalized silica nanoparticles for voltammetric determination of l-tyrosine.

PubMed

Rahimi-Mohseni, Mohadeseh; Raoof, Jahan Bakhsh; Ojani, Reza; Aghajanzadeh, Tahereh A; Bagheri Hashkavayi, Ayemeh

2018-07-01

In this paper, a new and facile method for the electrochemical determination of l-tyrosine was designed. First, 3-mercaptopropyl trimethoxysilane-functionalized silica nanoparticles were added to a paper disc. Then, the banana peel tissue and the mediator potassium hexacyanoferrate were dropped onto the paper, respectively. The modified paper disc was placed on the top of the graphite screen printed electrode and electrochemical characterization of this biosensor was studied by cyclic voltammetry and electrochemical impedance spectroscopy methods. The effective parameters like pH, banana peel tissue percentage, and the amount of mediator loading were optimized. l-tyrosine measurements were done by differential pulse voltammetry with a little sample (3 μL) for analysis. The biosensor showed a linear response for l-tyrosine in the wide concentration range of 0.05-600 μM and a low detection limit about 0.02 μM because of the co-catalytic effect of enzyme and nanoparticles. The stability of the biosensor and its selectivity were evaluated. This biosensor was applied for the voltammetric determination of l-tyrosine in the blood plasma sample. The results of the practical application study were comparable with the standard method (HPLC). In conclusion, a simple, inexpensive, rapid, sensitive and selective technique was successfully applied to the l-tyrosine analysis of the little samples. Copyright © 2018 Elsevier B.V. All rights reserved.

Characterization of the endogenous enzymatic hydrolyses of Petroselinum crispum glycosides: determined by chromatography upon their sugar and flavonoid products.

PubMed

Boldizsár, Imre; Füzfai, Zsófia; Molnár-Perl, Ibolya

2013-06-07

The behavior of the flavonoid diglycosides, relevant constituents of parsley (Petroselinum crispum) fruit (PFr) and leaf (PLe) samples was characterized upon their enzymatic hydrolyses applying complementary liquid chromatography-ultraviolet (LC-UV) and gas chromatography mass selective (GC-MS) detections. Analyses were performed in quantitative manner, from the same extracts as a function of hydrolysis times. Both in fruit and leaf tissue extracts, in intact and in enzyme hydrolyzed ones, apigenin, chrysoeriol, their glycosides, sugars, sugar alcohols, carboxylic acids and phytosterols, in total 17 constituents were identified and quantified. Based primarily on the selective mass fragmentation properties of the trimethylsilyl (oxime) ether/ester derivatives of constituents, we confirmed several novelties to the field. (i) It was shown for the first time that in parsley tissues different types of glycosidase enzyme are active. In PFr samples, both the stepwise and disaccharide specific endogenous mechanisms were certified, quantifying simultaneously the continuous release of apigenin, chrysoeriol, 2-O-apiosyl-apiose, apiose and glucose. (ii) 2-O-Apiosyl-glucose was demonstrated as disaccharide due to its formation under derivatization conditions from parsley glycosides. (iii) Both in PFr and in PLe samples even the invertase enzyme activity was attainable: sucrose decomposition in both tissues was going on with the same intensity. Three different types of enzymatic glycosidase processes were followed with their specific hydrolysis products by means of HPLC-UV and GC-MS, simultaneously. Copyright © 2013 Elsevier B.V. All rights reserved.

The influence of the cumulated deformation energy in the measurement by the DSI method on the selected mechanical properties of bone tissues.

PubMed

Makuch, Anna M; Skalski, Konstanty R; Pawlikowski, Marek

2017-01-01

The goal of the study was to determine the influence of DSI test conditions, i.e., loading/unloading rates, hold time, and the value of the maximum loading force on selected mechanical properties of trabecular bone tissue. The test samples were resected from a femoral head of a patient qualified for a hip replacement surgery. During the DSI tests hardness (HV, HM, HIT) and elastic modulus (EIT) of trabecular bone tissue were measured using the Micro Hardness Tester (MHT, CSEM). The analysis of the results of measurements and the calculations of total energy, i.e., elastic and inelastic (Wtotal, Welastic, Winelastic) and those of hardness and elasticity made it possible to assess the impact of the process parameters (loading velocity, force and hold time) on mechanical properties of bone structures at a microscopic level. The coefficient k dependent on the EIT/HIT ratio and on the stored energy (ΔW = Wtotal - Welastic) is a measure of the material reaction to the loading and the deformation of tissue.

Identification of Differentially Expressed IGFBP5-Related Genes in Breast Cancer Tumor Tissues Using cDNA Microarray Experiments.

PubMed

Akkiprik, Mustafa; Peker, İrem; Özmen, Tolga; Amuran, Gökçe Güllü; Güllüoğlu, Bahadır M; Kaya, Handan; Özer, Ayşe

2015-11-10

IGFBP5 is an important regulatory protein in breast cancer progression. We tried to identify differentially expressed genes (DEGs) between breast tumor tissues with IGFBP5 overexpression and their adjacent normal tissues. In this study, thirty-eight breast cancer and adjacent normal breast tissue samples were used to determine IGFBP5 expression by qPCR. cDNA microarrays were applied to the highest IGFBP5 overexpressed tumor samples compared to their adjacent normal breast tissue. Microarray analysis revealed that a total of 186 genes were differentially expressed in breast cancer compared with normal breast tissues. Of the 186 genes, 169 genes were downregulated and 17 genes were upregulated in the tumor samples. KEGG pathway analyses showed that protein digestion and absorption, focal adhesion, salivary secretion, drug metabolism-cytochrome P450, and phenylalanine metabolism pathways are involved. Among these DEGs, the prominent top two genes (MMP11 and COL1A1) which potentially correlated with IGFBP5 were selected for validation using real time RT-qPCR. Only COL1A1 expression showed a consistent upregulation with IGFBP5 expression and COL1A1 and MMP11 were significantly positively correlated. We concluded that the discovery of coordinately expressed genes related with IGFBP5 might contribute to understanding of the molecular mechanism of the function of IGFBP5 in breast cancer. Further functional studies on DEGs and association with IGFBP5 may identify novel biomarkers for clinical applications in breast cancer.

Molecular detection of Toxoplasma gondii and Neospora caninum in birds from South Africa.

PubMed

Lukášová, Radka; Kobédová, Kateřina; Halajian, Ali; Bártová, Eva; Murat, Jean-Benjamin; Rampedi, Kgethedi Michael; Luus-Powell, Wilmien J

2018-02-01

There are not any records on the detection of Toxoplasma gondii and Neospora caninum in tissues of wild birds in the African continent. The aim of the study was to investigate the occurrence of DNA from these protozoan parasites in brain tissue samples collected in years 2014-2015 from 110 wild and domestic birds of 15 orders. Birds came mainly from the province of Limpopo (n=103); the other seven birds came from other five provinces of South Africa. Parasite DNAs were detected by PCR in animal brains. While all samples were negative for N. caninum, T. gondii DNA was detected in three (2.7%) birds: a Red-eyed Dove (Streptopelia semitorquata), a Laughing Dove (S. senegalensis) and a Southern-Yellow-billed Hornbill (Tockus leucomelas), all from Limpopo province. Positive samples were selected for genotyping by a 15 microsatellite markers method in a single multiplex PCR assay. Only the sample from the Red-eyed Dove was successfully genotyped and characterized as type II. This is the first detection of T. gondii in tissue of native African wild birds and the first study focusing on N. caninum in birds from South Africa. Copyright © 2017. Published by Elsevier B.V.

Micro and bulk analysis of prostate tissues classified as hyperplasia

NASA Astrophysics Data System (ADS)

Kwiatek, W. M.; Banaś, A.; Banaś, K.; Cinque, G.; Dyduch, G.; Falkenberg, G.; Kisiel, A.; Marcelli, A.; Podgórczyk, M.

2007-07-01

BPH (Benign Prostatic Hyperplasia) is the most common benign neoplasm (non cancerous enlargement of the prostate gland), whose prevalence increases with age. The gland, when increased in size, exerts pressure on the urethra, causing obstruction to urine flow. The latter may result in severe urinary tract and kidney conditions. In this work prostate samples from patients diagnosed with BPH were analyzed using synchrotron radiation. Micro-analysis of the hyperplastic samples was carried out on the L-beam line at HASYLAB, DESY (Germany), while bulk analysis on selected samples was performed at the DRX2 beamline at LNF, Frascati (Italy). Microanalysis with a mono-energetic beam 15 μm in diameter confirmed that concentrations of certain elements, such as S, Mn, Cu, Fe and Zn, are good indicators of pathological disorders in prostate tissue that may be considered effective tracers of developing compliant. The concentrations of Mn, Cu, Fe and Zn are higher in hyperplastic tissues, as compared to normal ones, while for sulphur the opposite is observed. Additionally, Fe and S K-edge XANES (X-ray Absorption Near Edge Structure) spectroscopy experiments were carried out in order to determine the chemical speciation of these elements in our samples.

Correlations of trace elements in breast human tissues: Evaluation of spatial distribution using μ-XRF

NASA Astrophysics Data System (ADS)

Silva, Marina Piacenti da; Silva, Deisy Mara da; Ribeiro-Silva, Alfredo; Poletti, Martin Eduardo

2012-05-01

The aim of this work is to investigate microscopic correlations between trace elements in breast human tissues. A synchrotron X-ray fluorescence microprobe system (μ-XRF) was used to obtain two-dimensional distribution of trace element Ca, Fe, Cu and Zn in normal (6 samples) and malignant (14 samples) breast tissues. The experiment was performed in X-ray Fluorescence beam line at Laboratório Nacional de Luz Síncrotron (LNLS), Campinas, Brazil. The white microbeam was generated with a fine conical capillary with a 20 μm output diameter. The samples were supported on a XYZ table. An optical microscope with motorized zoom was used for sample positioning and choice the area to be scanned. Automatic two-dimensional scans were programmed and performed with steps of 30 μm in each direction (x, y) on the selected area. The fluorescence signals were recorded using a Si(Li) detector, positioned at 90 degrees with respect to the incident beam, with a collection time of 10 s per point. The elemental maps obtained from each sample were overlap to observe correlation between trace elements. Qualitative results showed that the pairs of elements Ca-Zn and Fe-Cu could to be correlated in malignant breast tissues. Quantitative results, achieved by Spearman correlation tests, indicate that there is a spatial correlation between these pairs of elements (p < 0.001) suggesting the importance of these elements in metabolic processes associated with the development of the tumor.

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

Beyond laser microdissection technology: follow the yellow brick road for cancer research

PubMed Central

Legres, Luc G; Janin, Anne; Masselon, Christophe; Bertheau, Philippe

2014-01-01

Normal biological tissues harbour different populations of cells with intricate spacial distribution patterns resulting in heterogeneity of their overall cellular composition. Laser microdissection involving direct viewing and expertise by a pathologist, enables access to defined cell populations or specific region on any type of tissue sample, thus selecting near-pure populations of targeted cells. It opens the way for molecular methods directed towards well-defined populations, and provides also a powerful tool in studies focused on a limited number of cells. Laser microdissection has wide applications in oncology (diagnosis and research), cellular and molecular biology, biochemistry and forensics for tissue selection, but other areas have been gradually opened up to these new methodological approaches, such as cell cultures and cytogenetics. In clinical oncology trials, molecular profiling of microdissected samples can yield global “omics” information which, together, with the morphological analysis of cells, can provide the basis for diagnosis, prognosis and patient-tailored treatments. This remarkable technology has brought new insights in the understanding of DNA, RNA, and the biological functions and regulation of proteins to identify molecular disease signatures. We review herein the different applications of laser microdissection in a variety of fields, and we particularly focus attention on the pre-analytical steps that are crucial to successfully perform molecular-level investigations. PMID:24482735

Seasonal trends in growth and biomass accumulation of selected nutrients and metals in six species of emergent aquatic macrophytes

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

Behrends, L.L.; Bailey, E.; Bulls, M.J.

1996-05-01

Growth and biomass accumulation of selected nutrients and trace metals were monitored for six species of aquatic macrophytes during June, August and November, 1993. Plant species were cultivated in two polyculture treatments, each replicated three times. Polyculture I consisted of Scirpus acutus (hardstem bullrush), Phragmites communes (common reed), and Phalaris arundinacea (canary grass). Polyculture H consisted of Typha spp. (cattail), Scirpus atrovirens (green bullrush), and Scirpus cyperinus (wool grass). Each of the six cells (6 x 9 x 0.6 m), was operated as a gravel-substrate, subsurface-flow wetlands in a continuous recirculating mode. At six week intervals, macro, micro and tracemore » elements were dissolved and added to the sump of the recirculating system. On each of three sampling dates, replicate shoot and root samples were collected, segregated by species and tissue type (roots, rhizomes, stems and leaves), and prepared for gravimetric biomass estimates and chemical analysis. Tissue specific concentrations of N, P, K, Ca, Mg, Fe, Mn, Zn and Cu, were determined on each date for each species and tissue type. Results will be discussed with respect to species specific growth rates, biomass accumulation, and seasonal uptake and translocation of plant nutrients.« less

Potato stem cuttings to study Verticillium dahliae infection for resistance breeding and ‘omics’ studies

USDA-ARS?s Scientific Manuscript database

The use of consistent and effective methods for early discrimination of resistance to pathogens and selection of appropriate times for tissue sampling are important for experiments focused on global gene expression and metabolomics. Assays for resistance to the vascular pathogen Verticillium dahliae...

Behavior of optical properties of coagulated blood sample at 633 nm wavelength

NASA Astrophysics Data System (ADS)

Morales Cruzado, Beatriz; Vázquez y Montiel, Sergio; Delgado Atencio, José Alberto

2011-03-01

Determination of tissue optical parameters is fundamental for application of light in either diagnostics or therapeutical procedures. However, in samples of biological tissue in vitro, the optical properties are modified by cellular death or cellular agglomeration that can not be avoided. This phenomena change the propagation of light within the biological sample. Optical properties of human blood tissue were investigated in vitro at 633 nm using an optical setup that includes a double integrating sphere system. We measure the diffuse transmittance and diffuse reflectance of the blood sample and compare these physical properties with those obtained by Monte Carlo Multi-Layered (MCML). The extraction of the optical parameters: absorption coefficient μa, scattering coefficient μs and anisotropic factor g from the measurements were carried out using a Genetic Algorithm, in which the search procedure is based in the evolution of a population due to selection of the best individual, evaluated by a function that compares the diffuse transmittance and diffuse reflectance of those individuals with the experimental ones. The algorithm converges rapidly to the best individual, extracting the optical parameters of the sample. We compare our results with those obtained by using other retrieve procedures. We found that the scattering coefficient and the anisotropic factor change dramatically due to the formation of clusters.

Prevalence and molecular identification of Chlamydia abortus in commercial dairy goat farms in a hot region in Mexico.

PubMed

Campos-Hernández, Eleuterio; Vázquez-Chagoyán, Juan Carlos; Salem, Abdelfattah Z M; Saltijeral-Oaxaca, Jorge Antonio; Escalante-Ochoa, Cristina; López-Heydeck, Sandra M; de Oca-Jiménez, Roberto Montes

2014-08-01

The aim of this study was to determine the seroprevalence and presence of Chlamydia abortus in Saanen breed female goats from commercial dairy goat farms under intensive production in the municipality of Guanajuato, Mexico. Sera were collected to determine the prevalence of anti-C. abortus IgG antibodies using recombinant enzyme-linked immunosorbent assay (rELISA) and cell culture. Polymerase chain reaction (PCR) was used to prove the presence of the pathogen in swab samples collected from the vagina and rectum of selected animals. Additionally, foetal tissue samples from a sudden abortion were collected. C. abortus prevalence in female goats of commercial milking farms sampled in Guanajuato, Mexico, was 4.87% (n = 246). Seropositive animals were found in six out of nine (66.6%) dairy goat farms sampled, and prevalence among animals in individual farms ranged between 3.44 and 13.51%. C. abortus was detected using PCR in spleen tissue from the aborted foetus. PCR-based detection, as well as isolation from vaginal and rectal swabs, was not possible in the present study. Isolation through cell culture was also unsuccessful from aborted foetal tissue samples. In conclusion, the results from rELISA and PCR show that C. abortus is present in dairy goat farms in the state of Guanajuato, Mexico.

Endoscopic ultrasound guided fine needle aspiration and useful ancillary methods

PubMed Central

Tadic, Mario; Stoos-Veic, Tajana; Kusec, Rajko

2014-01-01

The role of endoscopic ultrasound (EUS) in evaluating pancreatic pathology has been well documented from the beginning of its clinical use. High spatial resolution and the close proximity to the evaluated organs within the mediastinum and abdominal cavity allow detection of small focal lesions and precise tissue acquisition from suspected lesions within the reach of this method. Fine needle aspiration (FNA) is considered of additional value to EUS and is performed to obtain tissue diagnosis. Tissue acquisition from suspected lesions for cytological or histological analysis allows, not only the differentiation between malignant and non-malignant lesions, but, in most cases, also the accurate distinction between the various types of malignant lesions. It is well documented that the best results are achieved only if an adequate sample is obtained for further analysis, if the material is processed in an appropriate way, and if adequate ancillary methods are performed. This is a multi-step process and could be quite a challenge in some cases. In this article, we discuss the technical aspects of tissue acquisition by EUS-guided-FNA (EUS-FNA), as well as the role of an on-site cytopathologist, various means of specimen processing, and the selection of the appropriate ancillary method for providing an accurate tissue diagnosis and maximizing the yield of this method. The main goal of this review is to alert endosonographers, not only to the different possibilities of tissue acquisition, namely EUS-FNA, but also to bring to their attention the importance of proper sample processing in the evaluation of various lesions in the gastrointestinal tract and other accessible organs. All aspects of tissue acquisition (needles, suction, use of stylet, complications, etc.) have been well discussed lately. Adequate tissue samples enable comprehensive diagnoses, which answer the main clinical questions, thus enabling targeted therapy. PMID:25339816

Development of Highly Sensitive and Specific mRNA Multiplex System (XCYR1) for Forensic Human Body Fluids and Tissues Identification

PubMed Central

Xu, Yan; Xie, Jianhui; Cao, Yu; Zhou, Huaigu; Ping, Yuan; Chen, Liankang; Gu, Lihua; Hu, Wei; Bi, Gang; Ge, Jianye; Chen, Xin; Zhao, Ziqin

2014-01-01

The identification of human body fluids or tissues through mRNA-based profiling is very useful for forensic investigations. Previous studies have shown mRNA biomarkers are effective to identify the origin of biological samples. In this study, we selected 16 tissue specific biomarkers to evaluate their specificities and sensitivities for human body fluids and tissues identification, including porphobilinogen deaminase (PBGD), hemoglobin beta (HBB) and Glycophorin A (GLY) for circulatory blood, protamine 2 (PRM2) and transglutaminase 4 (TGM4) for semen, mucin 4 (MUC4) and human beta defensin 1(HBD1) for vaginal secretion, matrix metalloproteinases 7 and 11 (MMP7 and MMP11) for menstrual blood, keratin 4(KRT4) for oral mucosa, loricrin (LOR) and cystatin 6 (CST6) for skin, histatin 3(HTN3) for saliva, statherin (STATH) for nasal secretion, dermcidin (DCD) for sweat and uromodulin (UMOD) for urine. The above mentioned ten common forensic body fluids or tissues were used in the evaluation. Based on the evaluation, a reverse transcription (RT) PCR multiplex assay, XCYR1, which includes 12 biomarkers (i.e., HBB, GLY, HTN3, PRM2, KRT4, MMP11, MUC4, DCD, UMOD, MMP7, TGM4, and STATH) and 2 housekeeping genes [i.e., glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and 18SrRNA], was developed. This assay was further validated with real casework samples and mock samples (with both single source and mixture) and it was approved that XCYR1 is effective to identify common body fluids or tissues (i.e., circulatory blood, saliva, semen, vaginal secretion, menstrual blood, oral mucosa, nasal secretion, sweat and urine) in forensic casework samples. PMID:24991806

Development of highly sensitive and specific mRNA multiplex system (XCYR1) for forensic human body fluids and tissues identification.

PubMed

Xu, Yan; Xie, Jianhui; Cao, Yu; Zhou, Huaigu; Ping, Yuan; Chen, Liankang; Gu, Lihua; Hu, Wei; Bi, Gang; Ge, Jianye; Chen, Xin; Zhao, Ziqin

2014-01-01

The identification of human body fluids or tissues through mRNA-based profiling is very useful for forensic investigations. Previous studies have shown mRNA biomarkers are effective to identify the origin of biological samples. In this study, we selected 16 tissue specific biomarkers to evaluate their specificities and sensitivities for human body fluids and tissues identification, including porphobilinogen deaminase (PBGD), hemoglobin beta (HBB) and Glycophorin A (GLY) for circulatory blood, protamine 2 (PRM2) and transglutaminase 4 (TGM4) for semen, mucin 4 (MUC4) and human beta defensin 1(HBD1) for vaginal secretion, matrix metalloproteinases 7 and 11 (MMP7 and MMP11) for menstrual blood, keratin 4(KRT4) for oral mucosa, loricrin (LOR) and cystatin 6 (CST6) for skin, histatin 3(HTN3) for saliva, statherin (STATH) for nasal secretion, dermcidin (DCD) for sweat and uromodulin (UMOD) for urine. The above mentioned ten common forensic body fluids or tissues were used in the evaluation. Based on the evaluation, a reverse transcription (RT) PCR multiplex assay, XCYR1, which includes 12 biomarkers (i.e., HBB, GLY, HTN3, PRM2, KRT4, MMP11, MUC4, DCD, UMOD, MMP7, TGM4, and STATH) and 2 housekeeping genes [i.e., glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and 18SrRNA], was developed. This assay was further validated with real casework samples and mock samples (with both single source and mixture) and it was approved that XCYR1 is effective to identify common body fluids or tissues (i.e., circulatory blood, saliva, semen, vaginal secretion, menstrual blood, oral mucosa, nasal secretion, sweat and urine) in forensic casework samples.

Spatial-Resolution Cell Type Proteome Profiling of Cancer Tissue by Fully Integrated Proteomics Technology.

PubMed

Xu, Ruilian; Tang, Jun; Deng, Quantong; He, Wan; Sun, Xiujie; Xia, Ligang; Cheng, Zhiqiang; He, Lisheng; You, Shuyuan; Hu, Jintao; Fu, Yuxiang; Zhu, Jian; Chen, Yixin; Gao, Weina; He, An; Guo, Zhengyu; Lin, Lin; Li, Hua; Hu, Chaofeng; Tian, Ruijun

2018-05-01

Increasing attention has been focused on cell type proteome profiling for understanding the heterogeneous multicellular microenvironment in tissue samples. However, current cell type proteome profiling methods need large amounts of starting materials which preclude their application to clinical tumor specimens with limited access. Here, by seamlessly combining laser capture microdissection and integrated proteomics sample preparation technology SISPROT, specific cell types in tumor samples could be precisely dissected with single cell resolution and processed for high-sensitivity proteome profiling. Sample loss and contamination due to the multiple transfer steps are significantly reduced by the full integration and noncontact design. H&E staining dyes which are necessary for cell type investigation could be selectively removed by the unique two-stage design of the spintip device. This easy-to-use proteome profiling technology achieved high sensitivity with the identification of more than 500 proteins from only 0.1 mm 2 and 10 μm thickness colon cancer tissue section. The first cell type proteome profiling of four cell types from one colon tumor and surrounding normal tissue, including cancer cells, enterocytes, lymphocytes, and smooth muscle cells, was obtained. 5271, 4691, 4876, and 2140 protein groups were identified, respectively, from tissue section of only 5 mm 2 and 10 μm thickness. Furthermore, spatially resolved proteome distribution profiles of enterocytes, lymphocytes, and smooth muscle cells on the same tissue slices and across four consecutive sections with micrometer distance were successfully achieved. This fully integrated proteomics technology, termed LCM-SISPROT, is therefore promising for spatial-resolution cell type proteome profiling of tumor microenvironment with a minute amount of clinical starting materials.

Optimal tumor sampling for immunostaining of biomarkers in breast carcinoma

PubMed Central

2011-01-01

Introduction Biomarkers, such as Estrogen Receptor, are used to determine therapy and prognosis in breast carcinoma. Immunostaining assays of biomarker expression have a high rate of inaccuracy; for example, estimates are as high as 20% for Estrogen Receptor. Biomarkers have been shown to be heterogeneously expressed in breast tumors and this heterogeneity may contribute to the inaccuracy of immunostaining assays. Currently, no evidence-based standards exist for the amount of tumor that must be sampled in order to correct for biomarker heterogeneity. The aim of this study was to determine the optimal number of 20X fields that are necessary to estimate a representative measurement of expression in a whole tissue section for selected biomarkers: ER, HER-2, AKT, ERK, S6K1, GAPDH, Cytokeratin, and MAP-Tau. Methods Two collections of whole tissue sections of breast carcinoma were immunostained for biomarkers. Expression was quantified using the Automated Quantitative Analysis (AQUA) method of quantitative immunofluorescence. Simulated sampling of various numbers of fields (ranging from one to thirty five) was performed for each marker. The optimal number was selected for each marker via resampling techniques and minimization of prediction error over an independent test set. Results The optimal number of 20X fields varied by biomarker, ranging between three to fourteen fields. More heterogeneous markers, such as MAP-Tau protein, required a larger sample of 20X fields to produce representative measurement. Conclusions The optimal number of 20X fields that must be sampled to produce a representative measurement of biomarker expression varies by marker with more heterogeneous markers requiring a larger number. The clinical implication of these findings is that breast biopsies consisting of a small number of fields may be inadequate to represent whole tumor biomarker expression for many markers. Additionally, for biomarkers newly introduced into clinical use, especially if therapeutic response is dictated by level of expression, the optimal size of tissue sample must be determined on a marker-by-marker basis. PMID:21592345

Mesenchymal Stem Cell Levels of Human Spinal Tissues.

PubMed

Harris, Liam; Vangsness, C Thomas

2018-05-01

Systematic review. The aim of this study was to investigate, quantify, compare, and compile the various mesenchymal stem cell (MSC) tissue sources within human spinal tissues to act as a compendium for clinical and research application. Recent years have seen a dramatic increase in academic and clinical understanding of human MSCs. Previously limited to cells isolated from bone marrow, the past decade has illicited the characterization and isolation of human MSCs from adipose, bone marrow, synovium, muscle, periosteum, peripheral blood, umbilical cord, placenta, and numerous other tissues. As researchers explore practical applications of cells in these tissues, the absolute levels of MSCs in specific spinal tissue will be critical to guide future research. The PubMED, MEDLINE, EMBASE, and Cochrane databases were searched for articles relating to the harvest, characterization, isolation, and quantification of human MSCs from spinal tissues. Selected articles were examined for relevant data, categorized according to type of spinal tissue, and when possible, standardized to facilitate comparisons between sites. Human MSC levels varied widely between spinal tissues. Yields for intervertebral disc demonstrated roughly 5% of viable cells to be positive for MSC surface markers. Cartilage endplate cells yielded 18,500 to 61,875 cells/0.8 mm thick sample of cartilage end plate. Ligamentum flavum yielded 250,000 to 500,000 cells/g of tissue. Annulus fibrosus fluorescence activated cell sorting treatment found 29% of cells positive for MSC marker Stro-1. Nucleus pulposus yielded mean tissue samples of 40,584 to 234,137 MSCs per gram of tissue. Numerous tissues within and surrounding the spine represent a consistent and reliable source for the harvest and isolation of human MSCs. Among the tissues of the spine, the annulus fibrosus and ligamentum flavum each offer considerable levels of MSCs, and may prove comparable to that of bone marrow. 5.

Identification and Validation of Reference Genes and Their Impact on Normalized Gene Expression Studies across Cultivated and Wild Cicer Species

PubMed Central

Reddy, Palakolanu Sudhakar; Sri Cindhuri, Katamreddy; Sivaji Ganesh, Adusumalli; Sharma, Kiran Kumar

2016-01-01

Quantitative Real-Time PCR (qPCR) is a preferred and reliable method for accurate quantification of gene expression to understand precise gene functions. A total of 25 candidate reference genes including traditional and new generation reference genes were selected and evaluated in a diverse set of chickpea samples. The samples used in this study included nine chickpea genotypes (Cicer spp.) comprising of cultivated and wild species, six abiotic stress treatments (drought, salinity, high vapor pressure deficit, abscisic acid, cold and heat shock), and five diverse tissues (leaf, root, flower, seedlings and seed). The geNorm, NormFinder and RefFinder algorithms used to identify stably expressed genes in four sample sets revealed stable expression of UCP and G6PD genes across genotypes, while TIP41 and CAC were highly stable under abiotic stress conditions. While PP2A and ABCT genes were ranked as best for different tissues, ABCT, UCP and CAC were most stable across all samples. This study demonstrated the usefulness of new generation reference genes for more accurate qPCR based gene expression quantification in cultivated as well as wild chickpea species. Validation of the best reference genes was carried out by studying their impact on normalization of aquaporin genes PIP1;4 and TIP3;1, in three contrasting chickpea genotypes under high vapor pressure deficit (VPD) treatment. The chickpea TIP3;1 gene got significantly up regulated under high VPD conditions with higher relative expression in the drought susceptible genotype, confirming the suitability of the selected reference genes for expression analysis. This is the first comprehensive study on the stability of the new generation reference genes for qPCR studies in chickpea across species, different tissues and abiotic stresses. PMID:26863232

Identification and Validation of Reference Genes and Their Impact on Normalized Gene Expression Studies across Cultivated and Wild Cicer Species.

PubMed

Reddy, Dumbala Srinivas; Bhatnagar-Mathur, Pooja; Reddy, Palakolanu Sudhakar; Sri Cindhuri, Katamreddy; Sivaji Ganesh, Adusumalli; Sharma, Kiran Kumar

2016-01-01

Quantitative Real-Time PCR (qPCR) is a preferred and reliable method for accurate quantification of gene expression to understand precise gene functions. A total of 25 candidate reference genes including traditional and new generation reference genes were selected and evaluated in a diverse set of chickpea samples. The samples used in this study included nine chickpea genotypes (Cicer spp.) comprising of cultivated and wild species, six abiotic stress treatments (drought, salinity, high vapor pressure deficit, abscisic acid, cold and heat shock), and five diverse tissues (leaf, root, flower, seedlings and seed). The geNorm, NormFinder and RefFinder algorithms used to identify stably expressed genes in four sample sets revealed stable expression of UCP and G6PD genes across genotypes, while TIP41 and CAC were highly stable under abiotic stress conditions. While PP2A and ABCT genes were ranked as best for different tissues, ABCT, UCP and CAC were most stable across all samples. This study demonstrated the usefulness of new generation reference genes for more accurate qPCR based gene expression quantification in cultivated as well as wild chickpea species. Validation of the best reference genes was carried out by studying their impact on normalization of aquaporin genes PIP1;4 and TIP3;1, in three contrasting chickpea genotypes under high vapor pressure deficit (VPD) treatment. The chickpea TIP3;1 gene got significantly up regulated under high VPD conditions with higher relative expression in the drought susceptible genotype, confirming the suitability of the selected reference genes for expression analysis. This is the first comprehensive study on the stability of the new generation reference genes for qPCR studies in chickpea across species, different tissues and abiotic stresses.

Imaging a Large Sample with Selective Plane Illumination Microscopy Based on Multiple Fluorescent Microsphere Tracking

NASA Astrophysics Data System (ADS)

Ryu, Inkeon; Kim, Daekeun

2018-04-01

A typical selective plane illumination microscopy (SPIM) image size is basically limited by the field of view, which is a characteristic of the objective lens. If an image larger than the imaging area of the sample is to be obtained, image stitching, which combines step-scanned images into a single panoramic image, is required. However, accurately registering the step-scanned images is very difficult because the SPIM system uses a customized sample mount where uncertainties for the translational and the rotational motions exist. In this paper, an image registration technique based on multiple fluorescent microsphere tracking is proposed in the view of quantifying the constellations and measuring the distances between at least two fluorescent microspheres embedded in the sample. Image stitching results are demonstrated for optically cleared large tissue with various staining methods. Compensation for the effect of the sample rotation that occurs during the translational motion in the sample mount is also discussed.

Distribution and chemical form of mercury in commercial fish tissues.

PubMed

Watanabe, Naoko; Tayama, Misato; Inouye, Minoru; Yasutake, Akira

2012-01-01

We analyzed total Hg concentrations in various tissue samples obtained from 7 commercially available fish species. MeHg contents were also estimated for muscle and liver samples by a selective analysis of inorganic Hg. Among the tissues, high Hg accumulations were shown in liver, muscle, heart and spleen throughout all fish species. Carnivorous fish, such as scorpion fish, sea bream and Japanese whiting, tended to show higher Hg accumulations in the muscle, with the highest Hg levels being shown by scorpion fish. Although the liver was expected to show the highest Hg accumulations among tissues throughout all fish species, the highest accumulation in the liver was observed only in scorpion fish. In contrast, the muscle level was significantly higher than the liver in Pacific saury and Japanese whiting. MeHg accumulated in fish is considered to show a sustained increase throughout the life of the fish, due to its long biological half-life. In fact, in the present study, muscle Hg levels in Japanese whiting, Japanese flying fish, and halfbeak showed good correlations with body weights. However, such correlations were not clear in scorpion fish, sea bream, Jack mackerel and Pacific saury. Selective analyses of inorganic Hg levels revealed that most of the Hg (> 95%) in fish muscle existed as MeHg, while the rates of MeHg contents in the liver varied from 56% in scorpion fish to 84% in Jack mackerel. As a result, fish muscle showed the highest MeHg accumulations in all fish species examined. These results suggest that reliable information on total Hg contents in fish muscle might be sufficient to avoid the risk of MeHg exposure caused by eating fish, even when one consumes other tissues such as fish liver.

Canary TMA — EDRN Public Portal

Cancer.gov

This protocol describes a multi-center, retrospective, case-cohort tissue microarray (TMA) study to evaluate tissue biomarkers for their ability to predict recurrent prostate cancer at the time of radical prostatectomy (RP). Candidate biomarkers will be assessed by performing tissue localization studies on TMAs containing recurrent prostate cancer and non-recurrent prostate cancer. De-identified data will be transferred to a central repository for statistical analysis. Participating institutions will use a variation of case-cohort sampling to randomly select a subset of patients from a retrospectively constructed RP cohort and/or perform selected assays on the cohort. The study endpoint is time to recurrence; of primary interest is five year recurrence free survival. Recurrent prostate cancer is defined by 1) a single serum prostate-specific antigen (PSA) level greater than 0.2 ng/mL after RP and/or 2) receipt of salvage or secondary therapy after RP and/or 3) clinical or radiological evidence of metastatic disease. Non-recurrent prostate cancer is defined as disease with no evidence of recurrence.

Determination of the neuropharmacological drug nodakenin in rat plasma and brain tissues by liquid chromatography tandem mass spectrometry: Application to pharmacokinetic studies.

PubMed

Song, Yingshi; Yan, Huiyu; Xu, Jingbo; Ma, Hongxi

2017-09-01

A rapid and sensitive liquid chromatography tandem mass spectrometry detection using selected reaction monitoring in positive ionization mode was developed and validated for the quantification of nodakenin in rat plasma and brain. Pareruptorin A was used as internal standard. A single step liquid-liquid extraction was used for plasma and brain sample preparation. The method was validated with respect to selectivity, precision, accuracy, linearity, limit of quantification, recovery, matrix effect and stability. Lower limit of quantification of nodakenin was 2.0 ng/mL in plasma and brain tissue homogenates. Linear calibration curves were obtained over concentration ranges of 2.0-1000 ng/mL in plasma and brain tissue homogenates for nodakenin. Intra-day and inter-day precisions (relative standard deviation, RSD) were <15% in both biological media. This assay was successfully applied to plasma and brain pharmacokinetic studies of nodakenin in rats after intravenous administration. Copyright © 2017 John Wiley & Sons, Ltd.

A liquid chromatography-tandem mass spectrometry-based targeted proteomics assay for monitoring P-glycoprotein levels in human breast tissue.

PubMed

Yang, Ting; Chen, Fei; Xu, Feifei; Wang, Fengliang; Xu, Qingqing; Chen, Yun

2014-09-25

P-glycoprotein (P-gp) can efflux drugs from cancer cells, and its overexpression is commonly associated with multi-drug resistance (MDR). Thus, the accurate quantification of P-gp would help predict the response to chemotherapy and for prognosis of breast cancer patients. An advanced liquid chromatography-tandem mass spectrometry (LC/MS/MS)-based targeted proteomics assay was developed and validated for monitoring P-gp levels in breast tissue. Tryptic peptide 368IIDNKPSIDSYSK380 was selected as a surrogate analyte for quantification, and immuno-depleted tissue extract was used as a surrogate matrix. Matched pairs of breast tissue samples from 60 patients who were suspected to have drug resistance were subject to analysis. The levels of P-gp were quantified. Using data from normal tissue, we suggested a P-gp reference interval. The experimental values of tumor tissue samples were compared with those obtained from Western blotting and immunohistochemistry (IHC). The result indicated that the targeted proteomics approach was comparable to IHC but provided a lower limit of quantification (LOQ) and could afford more reliable results at low concentrations than the other two methods. LC/MS/MS-based targeted proteomics may allow the quantification of P-gp in breast tissue in a more accurate manner. Copyright © 2014 Elsevier B.V. All rights reserved.

An unsupervised MVA method to compare specific regions in human breast tumor tissue samples using ToF-SIMS.

PubMed

Bluestein, Blake M; Morrish, Fionnuala; Graham, Daniel J; Guenthoer, Jamie; Hockenbery, David; Porter, Peggy L; Gamble, Lara J

2016-03-21

Imaging time-of-flight secondary ion mass spectrometry (ToF-SIMS) and principal component analysis (PCA) were used to investigate two sets of pre- and post-chemotherapy human breast tumor tissue sections to characterize lipids associated with tumor metabolic flexibility and response to treatment. The micron spatial resolution imaging capability of ToF-SIMS provides a powerful approach to attain spatially-resolved molecular and cellular data from cancerous tissues not available with conventional imaging techniques. Three ca. 1 mm(2) areas per tissue section were analyzed by stitching together 200 μm × 200 μm raster area scans. A method to isolate and analyze specific tissue regions of interest by utilizing PCA of ToF-SIMS images is presented, which allowed separation of cellularized areas from stromal areas. These PCA-generated regions of interest were then used as masks to reconstruct representative spectra from specifically stromal or cellular regions. The advantage of this unsupervised selection method is a reduction in scatter in the spectral PCA results when compared to analyzing all tissue areas or analyzing areas highlighted by a pathologist. Utilizing this method, stromal and cellular regions of breast tissue biopsies taken pre- versus post-chemotherapy demonstrate chemical separation using negatively-charged ion species. In this sample set, the cellular regions were predominantly all cancer cells. Fatty acids (i.e. palmitic, oleic, and stearic), monoacylglycerols, diacylglycerols and vitamin E profiles were distinctively different between the pre- and post-therapy tissues. These results validate a new unsupervised method to isolate and interpret biochemically distinct regions in cancer tissues using imaging ToF-SIMS data. In addition, the method developed here can provide a framework to compare a variety of tissue samples using imaging ToF-SIMS, especially where there is section-to-section variability that makes it difficult to use a serial hematoxylin and eosin (H&E) stained section to direct the SIMS analysis.

Reconstruction and feature selection for desorption electrospray ionization mass spectroscopy imagery

NASA Astrophysics Data System (ADS)

Gao, Yi; Zhu, Liangjia; Norton, Isaiah; Agar, Nathalie Y. R.; Tannenbaum, Allen

2014-03-01

Desorption electrospray ionization mass spectrometry (DESI-MS) provides a highly sensitive imaging technique for differentiating normal and cancerous tissue at the molecular level. This can be very useful, especially under intra-operative conditions where the surgeon has to make crucial decision about the tumor boundary. In such situations, the time it takes for imaging and data analysis becomes a critical factor. Therefore, in this work we utilize compressive sensing to perform the sparse sampling of the tissue, which halves the scanning time. Furthermore, sparse feature selection is performed, which not only reduces the dimension of data from about 104 to less than 50, and thus significantly shortens the analysis time. This procedure also identifies biochemically important molecules for further pathological analysis. The methods are validated on brain and breast tumor data sets.

Optimising the quantification of cytokines present at low concentrations in small human mucosal tissue samples using Luminex assays☆

PubMed Central

Staples, Emily; Ingram, Richard James Michael; Atherton, John Christopher; Robinson, Karen

2013-01-01

Sensitive measurement of multiple cytokine profiles from small mucosal tissue biopsies, for example human gastric biopsies obtained through an endoscope, is technically challenging. Multiplex methods such as Luminex assays offer an attractive solution but standard protocols are not available for tissue samples. We assessed the utility of three commercial Luminex kits (VersaMAP, Bio-Plex and MILLIPLEX) to measure interleukin-17A (IL-17) and interferon-gamma (IFNγ) concentrations in human gastric biopsies and we optimised preparation of mucosal samples for this application. First, we assessed the technical performance, limits of sensitivity and linear dynamic ranges for each kit. Next we spiked human gastric biopsies with recombinant IL-17 and IFNγ at a range of concentrations (1.5 to 1000 pg/mL) and assessed kit accuracy for spiked cytokine recovery and intra-assay precision. We also evaluated the impact of different tissue processing methods and extraction buffers on our results. Finally we assessed recovery of endogenous cytokines in unspiked samples. In terms of sensitivity, all of the kits performed well within the manufacturers' recommended standard curve ranges but the MILLIPLEX kit provided most consistent sensitivity for low cytokine concentrations. In the spiking experiments, the MILLIPLEX kit performed most consistently over the widest range of concentrations. For tissue processing, manual disruption provided significantly improved cytokine recovery over automated methods. Our selected kit and optimised protocol were further validated by measurement of relative cytokine levels in inflamed and uninflamed gastric mucosa using Luminex and real-time polymerase chain reaction. In summary, with proper optimisation Luminex kits (and for IL-17 and IFNγ the MILLIPLEX kit in particular) can be used for the sensitive detection of cytokines in mucosal biopsies. Our results should help other researchers seeking to quantify multiple low concentration cytokines in small tissue samples. PMID:23644159

Distribution and elimination of [14C] sarafloxacin hydrochloride from tissues of juvenile channel catfish (Ictalurus punctatus)

USGS Publications Warehouse

Gingerich, W.H.; Meinertz, J.R.; Dawson, V.K.; Gofus, J.E.; Delaney, L.J.; Bunnell, P.R.

1995-01-01

The distribution and loss of radioactivity from tissues were determined in 60 juvenile channel catfish (Ictalurus punctatus) following oral dosing with the candidate fish therapeutant Sarafin® ([14C] sarafloxacin hydrochloride) at 10 mg/kg for 5 consecutive days. Twelve groups of 5 fish each were sampled at selected times ranging from 3 to 240 h after the last dose was administered, The concentration and content of sarafloxacin-equivalent activity was determined in liver, gallbladder, kidney, skin, and skinless fillet by sample oxidation and liquid scintillation counting; content of sarafloxacin-equivalent activity was determined in stomach and anterior and posterior intestines, Skinless fillet tissues were also analyzed for sarafloxacin and for potential metabolites by gradient-elution high-performance liquid chromatography (HPLC) with in-line radiometric and fluorescence detection, Loss of radioactivity from the whole body conformed to a bimodal elimination pattern with a rapid initial phase (t1/2=11 h) and a slower secondary phase (t1/2=222 h). Tissue and contents of the gastrointestinal tract (i.e. stomach and anterior and posterior intestines) were a principal depot of activity during the first four sample times (3, 6, 12, and 24 h); the combined head, skeleton, and fins (i.e. residual carcass) were the principal depot of activity in samples taken after 24 h. Of those tissues sampled 3 h after the last dose, relative sarafloxacin concentration was greatest in the liver (4.06 μg equivalents/g) and least in the residual carcass (1.13 μg equivalents/g), Intermediate concentrations were found in the kidney (2.04 μg equivalents/g), skinless fillet (1.71 μg equivalents/ g), and the skin (1.51 μg equivalents/g). Concentrations of sarafloxacin-equivalent residues in edible skinless fillet were consistently among the lowest of all tissues examined. The highest mean concentration of parent-equivalent material in the fillet tissue was found 12 h after administration of the last dose (2.27 μg equivalents/g) and declined thereafter, Sarafloxacin constituted between 80 and 90% of the extractable radioactive residues from the fillet homogenates. No other peaks were resolved in any of the fillet tissue samples analyzed by HPLC with in-line radiometric detection.

Biological Sampling and Analysis in Sinclair and Dyes Inlets, Washington: Chemical Analyses for 2007 Puget Sound Biota Study

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

Brandenberger, Jill M.; Suslick, Carolynn R.; Johnston, Robert K.

2008-10-09

Evaluating spatial and temporal trends in contaminant residues in Puget Sound fish and macroinvertebrates are the objectives of the Puget Sound Ambient Monitoring Program (PSAMP). In a cooperative effort between the ENVironmental inVESTment group (ENVVEST) and Washington State Department of Fish and Wildlife, additional biota samples were collected during the 2007 PSAMP biota survey and analyzed for chemical residues and stable isotopes of carbon (δ13C) and nitrogen (δ15N). Approximately three specimens of each species collected from Sinclair Inlet, Georgia Basin, and reference locations in Puget Sound were selected for whole body chemical analysis. The muscle tissue of specimens selected formore » chemical analyses were also analyzed for δ13C and δ15N to provide information on relative trophic level and food sources. This data report summarizes the chemical residues for the 2007 PSAMP fish and macro-invertebrate samples. In addition, six Spiny Dogfish (Squalus acanthias) samples were necropsied to evaluate chemical residue of various parts of the fish (digestive tract, liver, embryo, muscle tissue), as well as, a weight proportional whole body composite (WBWC). Whole organisms were homogenized and analyzed for silver, arsenic, cadmium, chromium, copper, nickel, lead, zinc, mercury, 19 polychlorinated biphenyl (PCB) congeners, PCB homologues, percent moisture, percent lipids, δ13C, and δ15N.« less

Selective preparation of hard dental tissue: classical and laser treatments comparison

NASA Astrophysics Data System (ADS)

Dostálova, Tat'jana; Jelínkova, Helena; Němec, Michal; Koranda, Petr; Miyagi, Mitsunobu; Iwai, Katsumasa; Shi, Yi-Wei; Matsuura, Yuji

2006-02-01

For the purpose of micro-selective preparation which is part of the modern dentistry four various methods were examined: ablation by Er:YAG laser radiation (free-running or Q-switching regime), preparation of tissues by ultrasonic round ball tip, and by the classical dental drilling machine using diamond round bur. In the case of Er:YAG laser application the interaction energy 40 mJ in pulse of 200 us yielding to the interaction intensity 62 kW/cm2, and 20 mJ in pulse of 100 ns yielding to the interaction intensity 62 MW/cm2 was used for the case of free running, and Q-switch regime, respectively. For comparisson with the classical methods the ultrasound preparation tip (Sonixflex cariex TC, D - Sonicsys micro) and dental driller together with usual preparation burrs and standard handpiece were used. For the interaction experiment the samples of extracted human teeth and ebony cut into longitudinal sections and polished were used. The thickness of the prepared samples ranged from 5 to 7 mm. The methods were compared from the point of prepared cavity shape (SEM), inner surface, and possibility of selective removal of carries. The composite filling material was used to reconstruct the cavities. The dye penetrating analysis was performed.

Imaging the hard/soft tissue interface.

PubMed

Bannerman, Alistair; Paxton, Jennifer Z; Grover, Liam M

2014-03-01

Interfaces between different tissues play an essential role in the biomechanics of native tissues and their recapitulation is now recognized as critical to function. As a consequence, imaging the hard/soft tissue interface has become increasingly important in the area of tissue engineering. Particularly as several biotechnology based products have made it onto the market or are close to human trials and an understanding of their function and development is essential. A range of imaging modalities have been developed that allow a wealth of information on the morphological and physical properties of samples to be obtained non-destructively in vivo or via destructive means. This review summarizes the use of a selection of imaging modalities on interfaces to date considering the strengths and weaknesses of each. We will also consider techniques which have not yet been utilized to their full potential or are likely to play a role in future work in the area.

Tissue temperature distribution measurement by MRI and laser immunology for cancer treatment

NASA Astrophysics Data System (ADS)

Chen, Yichao; Gnyawali, Surya C.; Wu, Feng; Liu, Hong; Tesiram, Yasvir A.; Abbott, Andrew; Towner, Rheal A.; Chen, Wei R.

2007-02-01

In cancer treatment and immune response enhancement research, Magnetic Resonance Imaging (MRI) is an ideal method for non-invasive, three-dimensional temperature measurement. We used a 7.1-Tesla magnetic resonance imager for ex vivo tissues and small animal to determine temperature distribution of target tissue during laser irradiation. The feasibility of imaging is approved with high spatial resolution and high signal-noise- ratio. Tissue-simulating gel phantom gel, biological tissues, and tumor-bearing animals were used in the experiments for laser treatment and MR imaging. Thermal couple measurement of temperature in target samples was used for system calibration. An 805-nm laser was used to irradiate the samples with a laser power in the range of 1 to 2.5 watts. Using the MRI system and a specially developed processing algorithm, a clear temperature distribution matrix in the target tissue and surrounding tissue was obtained. The temperature profiles show that the selective laser photothermal effect could result in tissue temperature elevation in a range of 10 to 45 °C. The temperature resolution of the measurement was about 0.37°C including the total system error. The spatial resolution was 0.4 mm (128x128 pixels with field of view of 5.5x5.5 cm). The temperature distribution provided in vivo thermal information and future reference for optimizing dye concentration and irradiation parameters to achieve optimal thermal effects in cancer treatment.

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.

Detection of Salmonella spp. in veterinary samples by combining selective enrichment and real-time PCR.

PubMed

Goodman, Laura B; McDonough, Patrick L; Anderson, Renee R; Franklin-Guild, Rebecca J; Ryan, James R; Perkins, Gillian A; Thachil, Anil J; Glaser, Amy L; Thompson, Belinda S

2017-11-01

Rapid screening for enteric bacterial pathogens in clinical environments is essential for biosecurity. Salmonella found in veterinary hospitals, particularly Salmonella enterica serovar Dublin, can pose unique challenges for culture and testing because of its poor growth. Multiple Salmonella serovars including Dublin are emerging threats to public health given increasing prevalence and antimicrobial resistance. We adapted an automated food testing method to veterinary samples and evaluated the performance of the method in a variety of matrices including environmental samples ( n = 81), tissues ( n = 52), feces ( n = 148), and feed ( n = 29). A commercial kit was chosen as the basis for this approach in view of extensive performance characterizations published by multiple independent organizations. A workflow was established for efficiently and accurately testing veterinary matrices and environmental samples by use of real-time PCR after selective enrichment in Rappaport-Vassiliadis soya (RVS) medium. Using this method, the detection limit for S. Dublin improved by 100-fold over subculture on selective agars (eosin-methylene blue, brilliant green, and xylose-lysine-deoxycholate). Overall, the procedure was effective in detecting Salmonella spp. and provided next-day results.

Breast composition measurement with a cadmium-zinc-telluride based spectral computed tomography system

PubMed Central

Ding, Huanjun; Ducote, Justin L.; Molloi, Sabee

2012-01-01

Purpose: To investigate the feasibility of breast tissue composition in terms of water, lipid, and protein with a cadmium-zinc-telluride (CZT) based computed tomography (CT) system to help better characterize suspicious lesions. Methods: Simulations and experimental studies were performed using a spectral CT system equipped with a CZT-based photon-counting detector with energy resolution. Simulations of the figure-of-merit (FOM), the signal-to-noise ratio (SNR) of the dual energy image with respect to the square root of mean glandular dose (MGD), were performed to find the optimal configuration of the experimental acquisition parameters. A calibration phantom 3.175 cm in diameter was constructed from polyoxymethylene plastic with cylindrical holes that were filled with water and oil. Similarly, sized samples of pure adipose and pure lean bovine tissues were used for the three-material decomposition. Tissue composition results computed from the images were compared to the chemical analysis data of the tissue samples. Results: The beam energy was selected to be 100 kVp with a splitting energy of 40 keV. The tissue samples were successfully decomposed into water, lipid, and protein contents. The RMS error of the volumetric percentage for the three-material decomposition, as compared to data from the chemical analysis, was estimated to be approximately 5.7%. Conclusions: The results of this study suggest that the CZT-based photon-counting detector may be employed in the CT system to quantify the water, lipid, and protein mass densities in tissue with a relatively good agreement. PMID:22380361

Deep sequencing reveals distinct patterns of DNA methylation in prostate cancer.

PubMed

Kim, Jung H; Dhanasekaran, Saravana M; Prensner, John R; Cao, Xuhong; Robinson, Daniel; Kalyana-Sundaram, Shanker; Huang, Christina; Shankar, Sunita; Jing, Xiaojun; Iyer, Matthew; Hu, Ming; Sam, Lee; Grasso, Catherine; Maher, Christopher A; Palanisamy, Nallasivam; Mehra, Rohit; Kominsky, Hal D; Siddiqui, Javed; Yu, Jindan; Qin, Zhaohui S; Chinnaiyan, Arul M

2011-07-01

Beginning with precursor lesions, aberrant DNA methylation marks the entire spectrum of prostate cancer progression. We mapped the global DNA methylation patterns in select prostate tissues and cell lines using MethylPlex-next-generation sequencing (M-NGS). Hidden Markov model-based next-generation sequence analysis identified ∼68,000 methylated regions per sample. While global CpG island (CGI) methylation was not differential between benign adjacent and cancer samples, overall promoter CGI methylation significantly increased from ~12.6% in benign samples to 19.3% and 21.8% in localized and metastatic cancer tissues, respectively (P-value < 2 × 10(-16)). We found distinct patterns of promoter methylation around transcription start sites, where methylation occurred not only on the CGIs, but also on flanking regions and CGI sparse promoters. Among the 6691 methylated promoters in prostate tissues, 2481 differentially methylated regions (DMRs) are cancer-specific, including numerous novel DMRs. A novel cancer-specific DMR in the WFDC2 promoter showed frequent methylation in cancer (17/22 tissues, 6/6 cell lines), but not in the benign tissues (0/10) and normal PrEC cells. Integration of LNCaP DNA methylation and H3K4me3 data suggested an epigenetic mechanism for alternate transcription start site utilization, and these modifications segregated into distinct regions when present on the same promoter. Finally, we observed differences in repeat element methylation, particularly LINE-1, between ERG gene fusion-positive and -negative cancers, and we confirmed this observation using pyrosequencing on a tissue panel. This comprehensive methylome map will further our understanding of epigenetic regulation in prostate cancer progression.

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 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

Biomarkers of the Hedgehog/Smoothened pathway in healthy volunteers

PubMed Central

Kadam, Sunil K; Patel, Bharvin K R; Jones, Emma; Nguyen, Tuan S; Verma, Lalit K; Landschulz, Katherine T; Stepaniants, Sergey; Li, Bin; Brandt, John T; Brail, Leslie H

2012-01-01

The Hedgehog (Hh) pathway is involved in oncogenic transformation and tumor maintenance. The primary objective of this study was to select surrogate tissue to measure messenger ribonucleic acid (mRNA) levels of Hh pathway genes for measurement of pharmacodynamic effect. Expression of Hh pathway specific genes was measured by quantitative real time polymerase chain reaction (qRT-PCR) and global gene expression using Affymetrix U133 microarrays. Correlations were made between the expression of specific genes determined by qRT-PCR and normalized microarray data. Gene ontology analysis using microarray data for a broader set of Hh pathway genes was performed to identify additional Hh pathway-related markers in the surrogate tissue. RNA extracted from blood, hair follicle, and skin obtained from healthy subjects was analyzed by qRT-PCR for 31 genes, whereas 8 samples were analyzed for a 7-gene subset. Twelve sample sets, each with ≤500 ng total RNA derived from hair, skin, and blood, were analyzed using Affymetrix U133 microarrays. Transcripts for several Hh pathway genes were undetectable in blood using qRT-PCR. Skin was the most desirable matrix, followed by hair follicle. Whether processed by robust multiarray average or microarray suite 5 (MAS5), expression patterns of individual samples showed co-clustered signals; both normalization methods were equally effective for unsupervised analysis. The MAS5- normalized probe sets appeared better suited for supervised analysis. This work provides the basis for selection of a surrogate tissue and an expression analysis-based approach to evaluate pathway-related genes as markers of pharmacodynamic effect with novel inhibitors of the Hh pathway. PMID:22611475

Substance P expression in the gingival tissue after upper third molar extraction: effect of ketoprofen, a preliminary study.

PubMed

Abbate, G M; Mangano, A; Sacerdote, P; Amodeo, G; Moschetti, G; Levrini, L

2017-01-01

The aim of this study was to evaluate substance P (SP) levels and the effect of a non-steroidal anti-inflammatory drug (NSAID), ketoprofen, on SP in the pericoronal gingival tissue after extraction of upper third molars. A sample of 20 young non-smoking systemically healthy adults of both sexes, with a healthy upper third molar to extract for orthodontic purposes, was selected. After extraction, a sample of the gingival tissue of the pericoronal region was collected with a sterile scalpel, placed into test tubes and kept frozen at -20°C until the SP determination. SP levels were determined by using a commercially available enzyme immunoassay (ELISA) kit. The subjects were randomly divided into two groups: group 1 received a single dose of ketoprofen 30 minutes prior to the experimental procedure. The subjects of group 2 did not receive any kind of drug administration before extraction. The patients were asked to complete a diary on the postoperative pain. A relevant amount of SP was measured in all the gingival samples. No statistically significant difference could be detected in SP expression between the two groups. In group 1 pain appearance was significantly delayed (6.2±0.13 hours) in comparison with group 2 (3.95±0.2 hours). In this small selected group of subjects and limited study design, preventive administration of ketoprofen did not significantly affect the gingival levels of SP, the clinical recommendation emerging is that of NSAID administration postoperatively but before pain appearance in order to optimize the management of pain of the patient.

Automated imaging of cellular spheroids with selective plane illumination microscopy on a chip (Conference Presentation)

NASA Astrophysics Data System (ADS)

Paiè, Petra; Bassi, Andrea; Bragheri, Francesca; Osellame, Roberto

2017-02-01

Selective plane illumination microscopy (SPIM) is an optical sectioning technique that allows imaging of biological samples at high spatio-temporal resolution. Standard SPIM devices require dedicated set-ups, complex sample preparation and accurate system alignment, thus limiting the automation of the technique, its accessibility and throughput. We present a millimeter-scaled optofluidic device that incorporates selective plane illumination and fully automatic sample delivery and scanning. To this end an integrated cylindrical lens and a three-dimensional fluidic network were fabricated by femtosecond laser micromachining into a single glass chip. This device can upgrade any standard fluorescence microscope to a SPIM system. We used SPIM on a CHIP to automatically scan biological samples under a conventional microscope, without the need of any motorized stage: tissue spheroids expressing fluorescent proteins were flowed in the microchannel at constant speed and their sections were acquired while passing through the light sheet. We demonstrate high-throughput imaging of the entire sample volume (with a rate of 30 samples/min), segmentation and quantification in thick (100-300 μm diameter) cellular spheroids. This optofluidic device gives access to SPIM analyses to non-expert end-users, opening the way to automatic and fast screening of a high number of samples at subcellular resolution.

Laser dissection sampling modes for direct mass spectral analysis [using a hybrid optical microscopy/laser ablation liquid vortex capture/electrospray ionization system

DOE PAGES

Cahill, John F.; Kertesz, Vilmos; Van Berkel, Gary J.

2016-02-01

Here, laser microdissection coupled directly with mass spectrometry provides the capability of on-line analysis of substrates with high spatial resolution, high collection efficiency, and freedom on shape and size of the sampling area. Establishing the merits and capabilities of the different sampling modes that the system provides is necessary in order to select the best sampling mode for characterizing analytically challenging samples. The capabilities of laser ablation spot sampling, laser ablation raster sampling, and laser 'cut and drop' sampling modes of a hybrid optical microscopy/laser ablation liquid vortex capture electrospray ionization mass spectrometry system were compared for the analysis ofmore » single cells and tissue. Single Chlamydomonas reinhardtii cells were monitored for their monogalactosyldiacylglycerol (MGDG) and diacylglyceryltrimethylhomo-Ser (DGTS) lipid content using the laser spot sampling mode, which was capable of ablating individual cells (4-15 m) even when agglomerated together. Turbid Allium Cepa cells (150 m) having unique shapes difficult to precisely measure using the other sampling modes could be ablated in their entirety using laser raster sampling. Intact microdissections of specific regions of a cocaine-dosed mouse brain tissue were compared using laser 'cut and drop' sampling. Since in laser 'cut and drop' sampling whole and otherwise unmodified sections are captured into the probe, 100% collection efficiencies were achieved. Laser ablation spot sampling has the highest spatial resolution of any sampling mode, while laser ablation raster sampling has the highest sampling area adaptability of the sampling modes. In conclusion, laser ablation spot sampling has the highest spatial resolution of any sampling mode, useful in this case for the analysis of single cells. Laser ablation raster sampling was best for sampling regions with unique shapes that are difficult to measure using other sampling modes. Laser 'cut and drop' sampling can be used for cases where the highest sensitivity is needed, for example, monitoring drugs present in trace amounts in tissue.« less

Laser dissection sampling modes for direct mass spectral analysis [using a hybrid optical microscopy/laser ablation liquid vortex capture/electrospray ionization system

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

Cahill, John F.; Kertesz, Vilmos; Van Berkel, Gary J.

Here, laser microdissection coupled directly with mass spectrometry provides the capability of on-line analysis of substrates with high spatial resolution, high collection efficiency, and freedom on shape and size of the sampling area. Establishing the merits and capabilities of the different sampling modes that the system provides is necessary in order to select the best sampling mode for characterizing analytically challenging samples. The capabilities of laser ablation spot sampling, laser ablation raster sampling, and laser 'cut and drop' sampling modes of a hybrid optical microscopy/laser ablation liquid vortex capture electrospray ionization mass spectrometry system were compared for the analysis ofmore » single cells and tissue. Single Chlamydomonas reinhardtii cells were monitored for their monogalactosyldiacylglycerol (MGDG) and diacylglyceryltrimethylhomo-Ser (DGTS) lipid content using the laser spot sampling mode, which was capable of ablating individual cells (4-15 m) even when agglomerated together. Turbid Allium Cepa cells (150 m) having unique shapes difficult to precisely measure using the other sampling modes could be ablated in their entirety using laser raster sampling. Intact microdissections of specific regions of a cocaine-dosed mouse brain tissue were compared using laser 'cut and drop' sampling. Since in laser 'cut and drop' sampling whole and otherwise unmodified sections are captured into the probe, 100% collection efficiencies were achieved. Laser ablation spot sampling has the highest spatial resolution of any sampling mode, while laser ablation raster sampling has the highest sampling area adaptability of the sampling modes. In conclusion, laser ablation spot sampling has the highest spatial resolution of any sampling mode, useful in this case for the analysis of single cells. Laser ablation raster sampling was best for sampling regions with unique shapes that are difficult to measure using other sampling modes. Laser 'cut and drop' sampling can be used for cases where the highest sensitivity is needed, for example, monitoring drugs present in trace amounts in tissue.« less

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.

Selective enrichment and desalting of hydrophilic peptides using graphene oxide.

PubMed

Jiang, Miao; Qi, Linyu; Liu, Peiru; Wang, Zijun; Duan, Zhigui; Wang, Ying; Liu, Zhonghua; Chen, Ping

2016-08-01

The wide variety and low abundance of peptides in tissue brought great difficulties to the separation and identification of peptides, which is not in favor of the development of peptidomics. RP-HPLC, which could purify small molecules based on their hydrophobicity, has been widely used in the separation and enrichment of peptide due to its fast, good reproducibility and high resolution. However, RP-HPLC requires the instrument and expensive C18 column and its sample capacity is also limited. Recently, graphene oxide has been applied to the adsorption of amino acids. However, the enrichment efficiency and selectivity of graphene oxide for peptides remain unclear. In this study, the adsorption efficiency and selectivity of graphene oxide and RP-C18 matrix were compared on trypsinized α-actin and also on tissue extracts from pituitary gland and hippocampus. For α-actin, there exhibit similar elution peaks for total trypsinized products and those adsorpted by GO and C18 matrix. But peptides adsorbed by GO showed the higher hydrophilic peaks than which adsorbed by C18 matrix. The resulted RP-HPLC profile showed that most of peptides enriched by graphene oxide were eluted at low concentration of organic solvent, while peptides adsorbed by RP-C18 matrix were mostly eluted at relatively high concentration. Moreover, mass spectrometry analysis suggested that, in pituitary sample, there were 495 peptides enriched by graphene oxide, 447 peptides enriched by RP-C18 matrix while in hippocampus sample 333 and 243 peptides respectively. The GRAVY value analysis suggested that the graphene oxide has a stronger adsorption for highly hydrophilic peptides compared to the RP-C18 matrix. Furthermore, the combination of these two methods could notably increase the number of identification peptides but also the number of predicted protein precursors. Our study provided a new thought to the role of graphene oxide during the enrichment of peptides from tissue which should be useful for peptidomics study. Copyright © 2016 Elsevier B.V. All rights reserved.

Importance of Relating Efficacy Measures to Unbound Drug Concentrations for Anti-Infective Agents

PubMed Central

Gonzalez, Daniel; Schmidt, Stephan

2013-01-01

SUMMARY For the optimization of dosing regimens of anti-infective agents, it is imperative to have a good understanding of pharmacokinetics (PK) and pharmacodynamics (PD). Whenever possible, drug efficacy needs to be related to unbound concentrations at the site of action. For anti-infective drugs, the infection site is typically located outside plasma, and a drug must diffuse through capillary membranes to reach its target. Disease- and drug-related factors can contribute to differential tissue distribution. As a result, the assumption that the plasma concentration of drugs represents a suitable surrogate of tissue concentrations may lead to erroneous conclusions. Quantifying drug exposure in tissues represents an opportunity to relate the pharmacologically active concentrations to an observed pharmacodynamic parameter, such as the MIC. Selection of an appropriate specimen to sample and the advantages and limitations of the available sampling techniques require careful consideration. Ultimately, the goal will be to assess the appropriateness of a drug and dosing regimen for a specific pathogen and infection. PMID:23554417

Accounting for tissue heterogeneity in infrared spectroscopic imaging for accurate diagnosis of thyroid carcinoma subtypes.

PubMed

Martinez-Marin, David; Sreedhar, Hari; Varma, Vishal K; Eloy, Catarina; Sobrinho-Simões, Manuel; Kajdacsy-Balla, André; Walsh, Michael J

2017-07-01

Fourier transform infrared (FT-IR) microscopy was used to image tissue samples from twenty patients diagnosed with thyroid carcinoma. The spectral data were then used to differentiate between follicular thyroid carcinoma and follicular variant of papillary thyroid carcinoma using principle component analysis coupled with linear discriminant analysis and a Naïve Bayesian classifier operating on a set of computed spectral metrics. Classification of patients' disease type was accomplished by using average spectra from a wide region containing follicular cells, colloid, and fibrosis; however, classification of disease state at the pixel level was only possible when the extracted spectra were limited to follicular epithelial cells in the samples, excluding the relatively uninformative areas of fibrosis. The results demonstrate the potential of FT-IR microscopy as a tool to assist in the difficult diagnosis of these subtypes of thyroid cancer, and also highlights the importance of selectively and separately analyzing spectral information from different features of a tissue of interest.

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.

Hog Charm II tetracycline test screening results compared with a liquid chromatography tandem mass spectrometry 10-μg/kg method.

PubMed

Salter, Robert; Holmes, Steven; Legg, David; Coble, Joel; George, Bruce

2012-02-01

Pork tissue samples that tested positive and negative by the Charm II tetracycline test screening method in the slaughter plant laboratory were tested with the modified AOAC International liquid chromatography tandem mass spectrometry (LC-MS-MS) method 995.09 to determine the predictive value of the screening method at detecting total tetracyclines at 10 μg/kg of tissue, in compliance with Russian import regulations. There were 218 presumptive-positive tetracycline samples of 4,195 randomly tested hogs. Of these screening test positive samples, 83% (182) were positive, >10 μg/kg by LC-MS-MS; 12.8% (28) were false violative, greater than limit of detection (LOD) but <10 μg/kg; and 4.2% (8) were not detected at the LC-MS-MS LOD. The 36 false-violative and not-detected samples represent 1% of the total samples screened. Twenty-seven of 30 randomly selected tetracycline screening negative samples tested below the LC-MS-MS LOD, and 3 samples tested <3 μg/kg chlortetracycline. Results indicate that the Charm II tetracycline test is effective at predicting hogs containing >10 μg/kg total tetracyclines in compliance with Russian import regulations.

Trends in concentrations of polychlorinated biphenyls in fish tissue from selected sites in the Delaware River basin in New Jersey, New York, and Pennsylvania, 1969-98

USGS Publications Warehouse

Riva-Murray, Karen; Brightbill, Robin A.; Bilger, Michael D.

2003-01-01

Trends in concentrations of polychlorinated biphenyls in fish tissue from selected sites in the Delaware River basin in New Jersey, New York, and Pennsylvania, 1969-98 by Karen Riva-Murray, Robin A. Brightbill, and Michael D. Bilger U.S. GEOLOGICAL SURVEY Water-Resources Investigations Report 01-4066 ABSTRACT Polychlorinated biphenyl (PCB) concentrations in fish tissue collected during the 1990's from selected sites in the Delaware River Basin were compared with concentrations in fish tissue collected during 1969-88. Data collected by State and Federal agencies on concentrations in whole-body common carp (Cyprinus carpio) and white sucker (Catostomus commersoni), and edible portions of American eel (Anguilla rostrata), smallmouth bass (Micropterus dolomieu), and channel catfish (Ictalurus punctatus) during 1969-98 were compiled to define temporal trends in concentrations of PCBs in fish tissue from selected segments of the Delaware River, Lehigh River, Schuylkill River, and Brandywine Creek. The Delaware River in the vicinity of Trenton, New Jersey and Yardley, Pennsylvania (above the tidal influence) had the largest long-term data set among the sites considered for this study and was the only site with sufficient data for statistical analysis. A general pattern of decline in PCB concentrations during 1969-98 was apparent for this river segment. PCB concentrations in whole-body white sucker from this lower Delaware River segment declined during 1969-98 from a highest concentration of 7 micrograms per gram (?g/g, wet weight) in a sample collected during 1972 to 0.26 ?g/g (wet weight) in a sample collected during 1998. PCB concentration was negatively correlated with year (Spearman rank correlation -0.46, p < 0.08, n = 15); especially after removal of a sample from 1977 with an unusually low concentration (Spearman rank correlation -0.53, p = 0.05, n = 14). PCB concentrations in edible flesh of American eel declined during 1975-95, from a highest concentration of 3.8 ?g/g (wet weight) in a sample collected during 1976 to less than the reporting limit of 0.26 ?g/g (wet weight) in samples collected during 1993 and 1995. PCB concentrations in most samples (for species considered in this study) collected from the lower Delaware River exceeded the National Academy of Sciences and National Academy of Engineering (NAS/NAE) wildlife guideline level of 0.5 ?g/g during the 1970's and 1980's, and decreased to below this level during the 1990's. No samples of edible portions of game fish exceeded the U.S. Food and Drug Administration (FDA) tolerance level by the mid 1980's. However, the PCB concentration in a smallmouth bass fillet sample that was collected during 1998 (0.37 ?g/g) exceeded the Pennsylvania fish-consumption advisory level of 0.06 ?g/g, and the concentrations in whole-body common carp and white sucker collected during 1998 (1.10 ?g/g and 0.26 ?g/g, respectively) exceeded the New York State Department of Environmental Conservation wildlife criterion concentration of 0.11 ?g/g. (The concentration in carp also exceeded the 1973 NAS/NAE wildlife guideline concentration of 0.5 ?g/g.) Graphical analysis of PCB concentrations in whole white sucker and (or) edible portions of American eel from the upper Delaware River, lower Delaware River, middle Schuylkill River, and Brandywine Creek indicate a decline from the 1970's and (or) 1980's to the middle to late 1990's. Temporal trends in PCB concentrations in white sucker samples from the lower Lehigh and Schuylkill Rivers during 1979-98 are less clear; the PCB concentration (wet-weight basis) from a sample collected in 1998 from the lower Lehigh River was similar to that from a sample collected in 1979, and concentrations actually increased during 1982-98. Similarly, PCB concentrations in samples of white sucker and American eel from the lower Schuylkill River were highly variable over time. A decrease in lipid-adjusted PCB concentrations at both sites (for several whi

Usefulness of tissue autofluorescence imaging in actinic cheilitis diagnosis.

PubMed

Takahama Junior, Ademar; Kurachi, Cristina; Cosci, Alessandro; Pereira Faustino, Isabel Schausltz; Camisasca, Danielle Resende; da Costa Fontes, Karla Bianca Fernandes; Pires, Fábio Ramôa; Azevedo, Rebeca Souza

2013-07-01

Actinic cheilitis (AC) is a potentially malignant disorder of the lips. Because of its heterogeneous clinical aspect, it is difficult to indicate representative biopsy area. The purpose of this study was to evaluate the usefulness of tissue autofluorescence in AC diagnosis. The system was composed of a 405-nm light-emitting diode, sent to the sample by a dichroic, that allows the fluorescence signal to reach a camera directly plugged in the system. Fifty-seven patients with clinical diagnosis of AC and 45 normal volunteers were selected. According to clinical and fluorescence features, one or more areas were selected for biopsies in the AC group and epithelial dysplasia (ED) grades were established. The autofluorescence images were processed by a clustering algorithm for AC automated diagnosis. The tissue autofluorescence image revealed a heterogeneous pattern of loss and increase of fluorescence in patients with AC. ED was found in 93% of the cases, and most of the areas graded as moderate or severe ED were chosen with the aid of autofluorescence. The processed autofluorescence images from AC patients showed a higher number of spots in an irregular pattern. Tissue autofluorescence image system is a useful technique in association with clinical examination for AC diagnosis.

Comparison of Four Nuclear Isolation Buffers for Plant DNA Flow Cytometry

PubMed Central

LOUREIRO, JOÃO; RODRIGUEZ, ELEAZAR; DOLEŽEL, JAROSLAV; SANTOS, CONCEIÇÃO

2006-01-01

• Background and Aims DNA flow cytometry requires preparation of suspensions of intact nuclei, which are stained using a DNA-specific fluorochrome prior to analysis. Various buffer formulas were developed to preserve nuclear integrity, protect DNA from degradation and facilitate its stoichiometric staining. Although nuclear isolation buffers differ considerably in chemical composition, no systematic comparison of their performance has been made until now. This knowledge is required to select the appropriate buffer for a given species and tissue. • Methods Four common lysis buffers (Galbraith's, LB01, Otto's and Tris.MgCl2) were used to prepare samples from leaf tissues of seven plant species (Sedum burrito, Oxalis pes-caprae, Lycopersicon esculentum, Celtis australis, Pisum sativum, Festuca rothmaleri and Vicia faba). The species were selected to cover a wide range of genome sizes (1·30–26·90 pg per 2C DNA) and a variety of leaf tissue types. The following parameters were assessed: forward (FS) and side (SS) light scatters, fluorescence of propidium iodide-stained nuclei, coefficient of variation of DNA peaks, presence of debris background and the number of nuclei released from sample tissue. The experiments were performed independently by two operators and repeated on three different days. • Key Results Clear differences among buffers were observed. With the exception of O. pes-caprae, any buffer provided acceptable results for all species. LB01 and Otto's were generally the best buffers, with Otto's buffer providing better results in species with low DNA content. Galbraith's buffer led to satisfactory results and Tris.MgCl2 was generally the worst, although it yielded the best histograms in C. australis. A combined analysis of FS and SS provided a ‘fingerprint’ for each buffer. The variation between days was more significant than the variation between operators. • Conclusions Each lysis buffer tested responded to a specific problem differently and none of the buffers worked best with all species. These results expand our knowledge on nuclear isolation buffers and will facilitate selection of the most appropriate buffer depending on species, tissue type and the presence of cytosolic compounds interfering with DNA staining. PMID:16820407

Development and validation of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the quantification of tigecycline in rat brain tissues.

PubMed

Munyeza, Chiedza F; Shobo, Adeola; Baijnath, Sooraj; Bratkowska, Dominika; Naiker, Suhashni; Bester, Linda A; Singh, Sanil D; Maguire, Glenn E M; Kruger, Hendrik G; Naicker, Tricia; Govender, Thavendran

2016-06-01

Tigecycline (TIG), a derivative of minocycline, is the first in the novel class of glycylcyclines and is currently indicated for the treatment of complicated skin structure and intra-abdominal infections. A selective, accurate and reversed-phase high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was developed for the determination of TIG in rat brain tissues. Sample preparation was based on protein precipitation and solid phase extraction using Supel-Select HLB (30 mg/1 mL) cartridges. The samples were separated on a YMC Triart C18 column (150 mm x 3.0 mm. 3.0 µm) using gradient elution. Positive electrospray ionization (ESI+) was used for the detection mechanism with the multiple reaction monitoring (MRM) mode. The method was validated over the concentration range of 150-1200 ng/mL for rat brain tissue. The precision and accuracy for all brain analyses were within the acceptable limit. The mean extraction recovery in rat brain was 83.6%. This validated method was successfully applied to a pharmacokinetic study in female Sprague Dawley rats, which were given a dose of 25 mg/kg TIG intraperitoneally at various time-points. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

Evaluating differential nuclear DNA yield rates and osteocyte numbers among human bone tissue types: A synchrotron radiation micro-CT approach.

PubMed

Andronowski, Janna M; Mundorff, Amy Z; Pratt, Isaac V; Davoren, Jon M; Cooper, David M L

2017-05-01

Molecular human identification has conventionally focused on DNA sampling from dense, weight-bearing cortical bone tissue, typically from femora or tibiae. A comparison of skeletal elements from three contemporary individuals demonstrated that elements with high quantities of cancellous bone yielded nuclear DNA at the highest rates, suggesting that preferentially sampling cortical bone may be suboptimal (Mundorff & Davoren, 2014). Despite these findings, the reason for the differential DNA yields between cortical and cancellous bone tissues remains unknown. The primary goal of this work is to ascertain whether differences in bone microstructure can be used to explain differential nuclear DNA yield among bone tissue types observed by Mundorff and Davoren (2014), with a focus on osteocytes and the three-dimensional (3D) quantification of their associated lacunae. Osteocytes and other bone cells are recognized to house DNA in bone tissue, thus examining the density of their lacunae may explain why nuclear DNA yield rates differ among bone tissue types. Lacunae were visualized and quantified using synchrotron radiation-based micro-Computed Tomographic imaging (SR micro-CT). Volumes of interest (VOIs) from cortical and cancellous bone tissues (n=129) were comparatively analyzed from the three skeletons sampled for Mundorff and Davoren's (2014) study. Analyses tested the primary hypothesis that the abundance and density of osteocytes (inferred from their lacunar spaces) vary between cortical and cancellous bone tissue types. Results demonstrated that osteocyte lacunar abundance and density vary between cortical and cancellous bone tissue types, with cortical bone VOIs containing a higher lacunar abundance and density. We found that the osteocyte lacunar density values are independent of nuclear DNA yield, suggesting an alternative explanation for the higher nuclear DNA yields from bones with greater quantities of cancellous bone tissue. The use of SR micro-CT allowed for a scale of analysis that revealed a high range of variation in lacunar abundance in both tissue types. Moreover, high-resolution SR micro-CT imaging revealed potential soft tissue remnants within marrow spaces not visible macroscopically. It is hypothesized that soft tissue remnants observed among the trabeculae of skeletal elements with high quantities of cancellous bone tissue are responsible for the high nuclear DNA yields. These findings have significant implications for bone-sample selection for nuclear DNA analysis in a forensic context when skeletal remains are recovered from the ground surface. Copyright © 2017 Elsevier B.V. All rights reserved.

Tryptophan as key biomarker to detect gastrointestinal tract cancer using non-negative biochemical analysis of native fluorescence and Stokes Shift spectroscopy

NASA Astrophysics Data System (ADS)

Wang, Leana; Zhou, Yan; Liu, Cheng-hui; Zhou, Lixin; He, Yong; Pu, Yang; Nguyen, Thien An; Alfano, Robert R.

2015-03-01

The objective of this study was to find out the emission spectral fingerprints for discrimination of human colorectal and gastric cancer from normal tissue in vitro by applying native fluorescence. The native fluorescence (NFL) and Stokes shift spectra of seventy-two human cancerous and normal colorectal (colon, rectum) and gastric tissues were analyzed using three selected excitation wavelengths (e.g. 300 nm, 320 nm and 340 nm). Three distinct biomarkers, tryptophan, collagen and reduced nicotinamide adenine dinucleotide hydrate (NADH), were found in the samples of cancerous and normal tissues from eighteen subjects. The spectral profiles of tryptophan exhibited a sharp peak in cancerous colon tissues under a 300 nm excitation when compared with normal tissues. The changes in compositions of tryptophan, collagen, and NADH were found between colon cancer and normal tissues under an excitation of 300 nm by the non-negative basic biochemical component analysis (BBCA) model.

Water-quality assessment of part of the upper Mississippi River basin, Minnesota and Wisconsin: Design and implementation of water-quality studies, 1995-98

USGS Publications Warehouse

Stark, James R.; Fallon, J.D.; Fong, A.L.; Goldstein, R.M.; Hanson, P.E.; Kroening, S.E.; Lee, K.E.

1999-01-01

This report describes the design, site-selection, and implementation of the study. Methods used to collect, process, and analyze samples; characterize sites; and assess habitat are described. A comprehensive list of sample sites is provided. Sample analyses for water-quality studies included chlorophyll a, major inorganic constituents, nutrients, trace elements, tritium, radon, environmental isotopes, organic carbon, pesticides, volatile organic compounds, and other synthetic and naturallyoccurring organic compounds. Aquatic-biological samples included fish, benthic macroinvertebrates, and algal enumeration and identification, as well as synthetic-organic compounds and trace elements in fish tissue.

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.

A Versatile Panel of Reference Gene Assays for the Measurement of Chicken mRNA by Quantitative PCR

PubMed Central

Maier, Helena J.; Van Borm, Steven; Young, John R.; Fife, Mark

2016-01-01

Quantitative real-time PCR assays are widely used for the quantification of mRNA within avian experimental samples. Multiple stably-expressed reference genes, selected for the lowest variation in representative samples, can be used to control random technical variation. Reference gene assays must be reliable, have high amplification specificity and efficiency, and not produce signals from contaminating DNA. Whilst recent research papers identify specific genes that are stable in particular tissues and experimental treatments, here we describe a panel of ten avian gene primer and probe sets that can be used to identify suitable reference genes in many experimental contexts. The panel was tested with TaqMan and SYBR Green systems in two experimental scenarios: a tissue collection and virus infection of cultured fibroblasts. GeNorm and NormFinder algorithms were able to select appropriate reference gene sets in each case. We show the effects of using the selected genes on the detection of statistically significant differences in expression. The results are compared with those obtained using 28s ribosomal RNA, the present most widely accepted reference gene in chicken work, identifying circumstances where its use might provide misleading results. Methods for eliminating DNA contamination of RNA reduced, but did not completely remove, detectable DNA. We therefore attached special importance to testing each qPCR assay for absence of signal using DNA template. The assays and analyses developed here provide a useful resource for selecting reference genes for investigations of avian biology. PMID:27537060

Development of solid-phase microextraction coupled with liquid chromatography for analysis of tramadol in brain tissue using its molecularly imprinted polymer.

PubMed

Habibi-Khorasani, Monireh; Mohammadpour, Amir Hooshang; Mohajeri, Seyed Ahmad

2017-02-01

In this work, performance of a molecularly imprinted polymer (MIP) as a selective solid-phase microextraction sorbent for the extraction and enrichment of tramadol in aqueous solution and rabbit brain tissue, is described. Binding properties of MIPs were studied in comparison with their nonimprinted polymer (NIP). Ten milligrams of the optimized MIP was then evaluated as a sorbent, for preconcentration, in molecularly imprinted solid-phase microextraction (MISPME) of tramadol from aqueous solution and rabbit brain tissue. The analytical method was calibrated in the range of 0.004 ppm (4 ng mL -1 ) and 10 ppm (10 μg mL -1 ) in aqueous media and in the ranges of 0.01 and 10 ppm in rabbit brain tissue, respectively. The results indicated significantly higher binding affinity of MIPs to tramadol, in comparison with NIP. The MISPME procedure was developed and optimized with a recovery of 81.12-107.54% in aqueous solution and 76.16-91.20% in rabbit brain tissue. The inter- and intra-day variation values were <8.24 and 5.06%, respectively. Finally the calibrated method was applied for determination of tramadol in real rabbit brain tissue samples after administration of a lethal dose. Our data demonstrated the potential of MISPME for rapid, sensitive and cost-effective sample analysis. Copyright © 2016 John Wiley & Sons, Ltd.

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.

Isolation and confirmation of viral nervous necrosis (VNN) disease in golden grey mullet (Liza aurata) and leaping mullet (Liza saliens) in the Iranian waters of the Caspian Sea.

PubMed

Zorriehzahra, M E J; Ghasemi, M; Ghiasi, M; Karsidani, S Haghighi; Bovo, G; Nazari, A; Adel, M; Arizza, V; Dhama, K

2016-07-15

The present study was conducted on 428 moribund mullet fish samples to isolate and identify the causative agent of a mysterious acute mortality which recently occurred in wild mullets in Iranian waters of Caspian Sea, suspected to be due to viral nervous necrosis (VNN) disease. Disease investigation was carried out employing various diagnostic procedures such as virology, bacteriology, parasitology, haematology, histopathology, IFAT, IHC and nested RT-PCR. Brain and eye samples of affected fishes were collected in sterile conditions and then kept at -80°C for cell culture isolation and nested RT-PCR detection of the causative agent. Other tissue samples were also collected and fixed for histopathology, IHC and EM examinations. CPE was observed in cell cultures at 6days after inoculation. Nine samples were found positive with virological assay. Nested RT-PCR, performed on suspected tissues and CPE positive samples, showed that about 21 tissue samples and all the CPE positive samples were positive for VNN virus (VNNV). IFAT was selected as a confirmatory method for detecting the presence of Betanodavirus antigen, cell culture isolation results and nested RT-PCR findings. Moreover, VNNV particles with 25-30nm in diameter were also visualized in the infected brain and retina. In pathogenicity studies, guppy fishes bathed in VNNV-infected tissue culture (10(-4) TCID50) showed clinical signs similar to naturally infected mullet after 15days post infection (dpi), with mortality rates reaching up to 100% at 30dpi. Affected organ samples as examined by cell culture isolation, IFAT, IHC and histopathology, revealed the presence of VNNV in the guppy fishes. In conclusion, it was confirmed that VNNV was the main causative agent for the disease outbreak in mullet fish in the Caspian Sea, and this is such first official report of VNN disease from Iran. Copyright © 2016 Elsevier B.V. All rights reserved.

Identification of Foreign Particles in Human Tissues using Raman Microscopy.

PubMed

Campion, Alan; Smith, Kenneth J; Fedulov, Alexey V; Gregory, David; Fan, Yuwei; Godleski, John J

2018-06-12

The precise identification of foreign particles in tissue for patient care and research has been studied using polarized light microscopy, electron microscopy with X-ray analysis, and electron diffraction. The goal of this study was to unambiguously identify particles in tissues using a combina-tion of polarized light microscopy and Raman microscopy, which provides chemical composition and microstructural characterization of complex materials with submicron spatial resolution. We designed a model system of stained and unstained cells that contained birefringent talc particles, and systematically investigated the influence of slide and coverslip materials, laser wavelengths, and mounting media on the Raman spectra ob-tained. Hematoxylin and eosin stained slides did not produce useful results because of fluorescence interference from the stains. Unstained cell samples prepared with standard slides and coverslips produce high quality Raman spectra when excited at 532 nm; the spectra are uniquely as-signed to talc. We also obtain high quality Raman spectra specific for talc in unstained tissue samples (pleural tissue following talc pleurodesis and ovarian tissue following long-term perineal talc exposure). Raman microscopy is sufficiently sensitive and compositionally selective to identify particles as small as one micron in diameter. Among commonly used coverslip mounting media, Cytoseal 60 is recommended; Permount was unacceptable due to intense background interference. Raman spectra have been catalogued for thousands of substances, which suggests that this approach is likely to be successful in identifying other particles of interest in tissues, potentially making Raman microscopy a powerful new tool in pathology.

MALDI Orbitrap Mass Spectrometry Profiling of Dysregulated Sulfoglycosphingolipids in Renal Cell Carcinoma Tissues

NASA Astrophysics Data System (ADS)

Jirásko, Robert; Holčapek, Michal; Khalikova, Maria; Vrána, David; Študent, Vladimír; Prouzová, Zuzana; Melichar, Bohuslav

2017-08-01

Matrix-assisted laser desorption/ionization coupled with Orbitrap mass spectrometry (MALDI-Orbitrap-MS) is used for the clinical study of patients with renal cell carcinoma (RCC), as the most common type of kidney cancer. Significant changes in sulfoglycosphingolipid abundances between tumor and autologous normal kidney tissues are observed. First, sulfoglycosphingolipid species in studied RCC samples are identified using high mass accuracy full scan and tandem mass spectra. Subsequently, optimization, method validation, and statistical evaluation of MALDI-MS data for 158 tissues of 80 patients are discussed. More than 120 sulfoglycosphingolipids containing one to five hexosyl units are identified in human RCC samples based on the systematic study of their fragmentation behavior. Many of them are recorded here for the first time. Multivariate data analysis (MDA) methods, i.e., unsupervised principal component analysis (PCA) and supervised orthogonal partial least square discriminant analysis (OPLS-DA), are used for the visualization of differences between normal and tumor samples to reveal the most up- and downregulated lipids in tumor tissues. Obtained results are closely correlated with MALDI mass spectrometry imaging (MSI) and histologic staining. Important steps of the present MALDI-Orbitrap-MS approach are also discussed, such as the selection of best matrix, correct normalization, validation for semiquantitative study, and problems with possible isobaric interferences on closed masses in full scan mass spectra.

Data-adaptive test statistics for microarray data.

PubMed

Mukherjee, Sach; Roberts, Stephen J; van der Laan, Mark J

2005-09-01

An important task in microarray data analysis is the selection of genes that are differentially expressed between different tissue samples, such as healthy and diseased. However, microarray data contain an enormous number of dimensions (genes) and very few samples (arrays), a mismatch which poses fundamental statistical problems for the selection process that have defied easy resolution. In this paper, we present a novel approach to the selection of differentially expressed genes in which test statistics are learned from data using a simple notion of reproducibility in selection results as the learning criterion. Reproducibility, as we define it, can be computed without any knowledge of the 'ground-truth', but takes advantage of certain properties of microarray data to provide an asymptotically valid guide to expected loss under the true data-generating distribution. We are therefore able to indirectly minimize expected loss, and obtain results substantially more robust than conventional methods. We apply our method to simulated and oligonucleotide array data. By request to the corresponding author.

PCV2d-2 is the predominant type of PCV2 DNA in pig samples collected in the U.S. during 2014-2016.

PubMed

Xiao, Chao-Ting; Harmon, Karen M; Halbur, Patrick G; Opriessnig, Tanja

2016-12-25

Porcine circovirus type 2 (PCV2) vaccination was introduced in the US in 2006 and since has been adopted by most pig producers. While porcine circovirus associated disease (PCVAD) outbreaks are now relatively uncommon in the US, PCV2 remains a concern which is emphasized by increasing numbers of PCR and sequencing requests for PCV2. In the present study, randomly selected lung tissues from 586 pigs submitted in 2015 were tested for presence of PCV2 DNA. Positive samples were further characterized by sequencing and combined with available PCV2 open-reading-frame (ORF) 2 sequences from the client data base of the Iowa State University Veterinary Diagnostic Laboratory. The prevalence of PCV2 in the randomly selected lung tissues was 23% (135/586) with 11.3% PCV2a, 29% PCV2b and 71.8% for PCV2d subgroup PCV2d-2. A total of 455 ORF2 sequences obtained from 2014 through 2016 were analyzed and PCV2d accounted for 66.7% of the 2014 sequences, 71.8% of the 2015 sequences, and 72% of the 2016 sequences. Interestingly, only 1.9% (9/455) of the sequences belonged to the recently identified PCV2e genotype. The present data indicates that despite an almost 100% PCV2 vaccine coverage in the US, PCV2 DNA can still be detected in almost 1 of 4 randomly selected pig tissues. PCV2d-2 is now the predominant genotype in the USA suggesting that PCV2d-2 may have some advantage over PCV2a and PCV2b in its ability to replicate in pigs under vaccination pressure. Copyright © 2016. Published by Elsevier B.V.

Subepithelial corneal fibrosis partially due to epithelial-mesenchymal transition of ocular surface epithelium

PubMed Central

Kawashima, Motoko; Higa, Kazunari; Satake, Yoshiyuki; Omoto, Masahiro; Tsubota, Kazuo; Shimmura, Shigeto; Shimazaki, Jun

2010-01-01

Purpose To determine whether epithelial-mesenchymal transition is involved in the development of corneal subepithelial fibrosis (pannus). Methods Frozen samples of pannus tissue removed from human corneas with a diagnosis of total limbal stem cell deficiency were characterized by immunostaining for both epithelial and mesenchymal markers. We selected transformation-related protein 63 (p63) and pancytokeratin as epithelial markers and vimentin and α-smooth muscle actin (α-SMA) as mesenchymal markers. Immunostaining for β-catenin and E-cadherin was performed to determine wingless-Int (Wnt)-pathway activation. RT–PCR analysis was also performed on epithelial tissue obtained from pannus samples after dispase digestion. Results Immunohistochemistry revealed strong nuclear expression of p63 and weak intercellular expression of E-cadherin in epithelial basal cells of pannus tissue. Furthermore, translocation of β-catenin from intercellular junctions to the nucleus and cytoplasm was also observed. Double-positive cells for both p63 and α-SMA were observed in the subepithelial stroma of pannus tissue, which was supported by RT–PCR and cytospin analysis. Conclusions Epithelial-mesenchymal transition may be partially involved in the development of subepithelial corneal fibrosis due to total limbal stem cell deficiency. PMID:21179238

TMA Navigator: network inference, patient stratification and survival analysis with tissue microarray data

PubMed Central

Lubbock, Alexander L. R.; Katz, Elad; Harrison, David J.; Overton, Ian M.

2013-01-01

Tissue microarrays (TMAs) allow multiplexed analysis of tissue samples and are frequently used to estimate biomarker protein expression in tumour biopsies. TMA Navigator (www.tmanavigator.org) is an open access web application for analysis of TMA data and related information, accommodating categorical, semi-continuous and continuous expression scores. Non-biological variation, or batch effects, can hinder data analysis and may be mitigated using the ComBat algorithm, which is incorporated with enhancements for automated application to TMA data. Unsupervised grouping of samples (patients) is provided according to Gaussian mixture modelling of marker scores, with cardinality selected by Bayesian information criterion regularization. Kaplan–Meier survival analysis is available, including comparison of groups identified by mixture modelling using the Mantel-Cox log-rank test. TMA Navigator also supports network inference approaches useful for TMA datasets, which often constitute comparatively few markers. Tissue and cell-type specific networks derived from TMA expression data offer insights into the molecular logic underlying pathophenotypes, towards more effective and personalized medicine. Output is interactive, and results may be exported for use with external programs. Private anonymous access is available, and user accounts may be generated for easier data management. PMID:23761446

Current trends and challenges in sample preparation for metallic nanoparticles analysis in daily products and environmental samples: A review

NASA Astrophysics Data System (ADS)

De la Calle, Inmaculada; Menta, Mathieu; Séby, Fabienne

2016-11-01

Due to the increasing use of nanoparticles (NPs) in consumer products, it becomes necessary to develop different strategies for their detection, identification, characterization and quantification in a wide variety of samples. Since the analysis of NPs in consumer products and environmental samples is particularly troublesome, a detailed description of challenges and limitations is given here. This review mainly focuses on sample preparation procedures applied for the mostly used techniques for metallic and metal oxide NPs characterization in consumer products and most outstanding publications of biological and environmental samples (from 2006 to 2015). We summarize the procedures applied for total metal content, extraction/separation and/or preconcentration of NPs from the matrix, separation of metallic NPs from their ions or from larger particles and NPs' size fractionation. Sample preparation procedures specifically for microscopy are also described. Selected applications in cosmetics, food, other consumer products, biological tissues and environmental samples are presented. Advantages and inconveniences of those procedures are considered. Moreover, selected simplified schemes for NPs sample preparation, as well as usual techniques applied are included. Finally, promising directions for further investigations are discussed.

The Resin-Embedded Cornea Prepared Via Rapid Processing Protocol : A Good Histomorphometric Target for Clinical Investigation in Ophthalmology and Optometry

PubMed Central

Cheah, Pike See; Mohidin, Norhani; Mohd Ali, Bariah; Maung, Myint; Latif, Azian Abdul

2008-01-01

This study illustrates and quantifies the changes on corneal tissue between the paraffin-embedded and resin-embedded blocks and thus, selects a better target in investigational ophthalmology and optometry via light microscopy. Corneas of two cynomolgus monkeys (Macaca fascicularis) were used in this study. The formalin-fixed cornea was prepared in paraffin block via the conventional tissue processing protocol (4-day protocol) and stained with haematoxylin and eosin. The glutaraldehyde-fixed cornea was prepared in resin block via the rapid and modified tissue processing procedure (1.2-day protocol) and stained with toluidine blue. The paraffin-embedded sample exhibits various undesired tissue damage and artifact such as thinner epithelium (due to the substantial volumic extraction from the tissue), thicker stroma layer (due to the separation of lamellae and the presence of voids) and the distorted endothelium. In contrast, the resin-embedded corneal tissue has demonstrated satisfactory corneal ultrastructural preservation. The rapid and modified tissue processing method for preparing the resin-embedded is particularly beneficial to accelerate the microscopic evaluation in ophthalmology and optometry. PMID:22570589

Survey design for lakes and reservoirs in the United States to assess contaminants in fish tissue.

PubMed

Olsen, Anthony R; Snyder, Blaine D; Stahl, Leanne L; Pitt, Jennifer L

2009-03-01

The National Lake Fish Tissue Study (NLFTS) was the first survey of fish contamination in lakes and reservoirs in the 48 conterminous states based on a probability survey design. This study included the largest set (268) of persistent, bioaccumulative, and toxic (PBT) chemicals ever studied in predator and bottom-dwelling fish species. The U.S. Environmental Protection Agency (USEPA) implemented the study in cooperation with states, tribal nations, and other federal agencies, with field collection occurring at 500 lakes and reservoirs over a four-year period (2000-2003). The sampled lakes and reservoirs were selected using a spatially balanced unequal probability survey design from 270,761 lake objects in USEPA's River Reach File Version 3 (RF3). The survey design selected 900 lake objects, with a reserve sample of 900, equally distributed across six lake area categories. A total of 1,001 lake objects were evaluated to identify 500 lake objects that met the study's definition of a lake and could be accessed for sampling. Based on the 1,001 evaluated lakes, it was estimated that a target population of 147,343 (+/-7% with 95% confidence) lakes and reservoirs met the NLFTS definition of a lake. Of the estimated 147,343 target lakes, 47% were estimated not to be sampleable either due to landowner access denial (35%) or due to physical barriers (12%). It was estimated that a sampled population of 78,664 (+/-12% with 95% confidence) lakes met the NLFTS lake definition, had either predator or bottom-dwelling fish present, and could be sampled.

Tributyltin bioaccumulation and toxic effects in freshwater gastropods Pomacea canaliculata after a chronic exposure: field and laboratory studies.

PubMed

Martínez, María L; Piol, María N; Sbarbati Nudelman, Norma; Verrengia Guerrero, Noemí R

2017-07-01

Freshwater samples and gastropod mollusks (Pomacea canaliculata) were collected at 5 sampling stations located along the lower Río de la Plata basin, Argentina, to assess the extent of tributyltin (TBT) contamination. Determined data revealed the presence of TBT and some of its breakdown products (dibutyltin: DBT, and monobutyltin: MBT) in all freshwater samples and also in soft tissues of P. canaliculata gastropods. Chronic bioassays (6 months) were performed using female gastropods that had been reared under laboratory conditions and exposed to a similar TBT concentration than the value determined in freshwater samples (1 µg L -1 ). The aims of this study were to evaluate the extent of TBT accumulation, the tissue distribution, and the effects on selected biomarkers (activity of superoxide dismutasa: SOD, activity of catalase: CAT, levels of total glutathione: t-GSH, lipid peroxidation, and activity of acetylcholinesterase: AChE). Gonads presented the highest accumulation, followed by the cephalopedal region, albumin gland, and finally hepatopancreas. Both metabolites, DBT and MBT, were also found. All exposed female animals presented development of a penis reflecting the potential of TBT as an endocrine disrupting chemical for this gastropod species. Results on the selected biomarkers confirmed additional adverse effects induced by TBT. An increase in CAT activity and changes in t-GSH levels are indicative of alterations on the cellular redox status. The inhibition of AChE could reflect signs of neurotoxicity. Altogether, these results reveal a negative impact on the health of this gastropod population.

Attitudes and Perceptions of Cancer Patients Toward Biospecimen Donation for Cancer Research: A Cross-Sectional Survey Among Chinese Cancer Patients.

PubMed

He, Na; Guo, Yan; He, Min; Qiang, Wanmin; Li, Haixin

2017-08-01

High-quality biospecimen collection from consented patients is crucial for cancer research activities. Patients' attitudes and willingness toward specimen donation influence high-quality biospecimen collection for cancer research activities. We carried out a cross-sectional study among randomly selected patients from 11 cancer departments of Tianjin Medical University Cancer Institute and Hospital between August 2014 and August 2015. A total of 784 patients were included to complete a 30-item self-administered survey. We evaluated the patients' willingness to consider providing leftover samples and additional samples for cancer research purposes. Among 784 patients, 683 (87.1%) and 653 (83.3%) were willing to donate leftover tissue and surplus blood after diagnosis, respectively. Six hundred thirty-one (80.5%) were favorably disposed to consider donating both tissue and blood samples for future cancer research. Female patients showed less willingness to donate biospecimens or related clinical data for research. First-hospitalized or older patients were less willing to provide leftover biospecimens or additional blood samples or even clinical data for research. By contrast, patients with a higher education level were more likely to donate leftover tissues after biopsy or surgery for research activities. Most Chinese cancer patients were willing to consider donating blood and tissue samples for cancer research. Several factors, including age, gender, first hospitalization, and education level, can influence their willingness to donate biospecimens. We need to provide proper education to increase understanding of patients in biobanking activities. This study provides novel empirical data on the likelihood of donating surplus and additional biospecimens and clinical health information among Chinese cancer patients.

3D imaging of cells and tissues by focused ion beam/scanning electron microscopy (FIB/SEM).

PubMed

Drobne, Damjana

2013-01-01

Integration of a scanning electron microscope (SEM) and focused ion beam (FIB) technology into a single FIB/SEM system permits use of the FIB as a nano-scalpel to reveal site-specific subsurface microstructures which can be examined in great detail by SEM. The FIB/SEM technology is widely used in the semiconductor industry and material sciences, and recently its use in the life sciences has been initiated. Samples for FIB/SEM investigation can be either embedded in a plastic matrix, the traditional means of preparation of transmission electron microscopy (TEM) specimens, or simply dried as in samples prepared for SEM imaging. Currently, FIB/SEM is used in the life sciences for (a) preparation by the lift-out technique of lamella for TEM analysis, (b) tomography of samples embedded in a matrix, and (c) in situ site-specific FIB milling and SEM imaging using a wide range of magnifications. Site-specific milling and imaging has attracted wide interest as a technique in structural research of single eukaryotic and prokaryotic cells, small animals, and different animal tissue, but it still remains to be explored more thoroughly. In the past, preparation of samples for site-specific milling and imaging by FIB/SEM has typically adopted the embedding techniques used for TEM samples, and which have been very well described in the literature. Sample preparation protocols for the use of dried samples in FIB/SEM have been less well investigated. The aim of this chapter is to encourage application of FIB/SEM on dried biological samples. A detailed description of conventional dried sample preparation and FIB/SEM investigation of dried biological samples is presented. The important steps are described and illustrated, and direct comparison between embedded and dried samples of same tissues is provided. The ability to discover links between gross morphology of the tissue or organ, surface characteristics of any selected region, and intracellular structural details on the nanometer scale is an appealing application of electron microscopy in the life sciences and merits further exploration.

Occurrence of selected contaminants in water, fish tissue, and streambed sediments in central Nebraska, 1992-95

USGS Publications Warehouse

Frenzel, Steven A.

1996-01-01

Surface and ground water in Nebraska may contain contaminants resulting from human activities. For purposes of this publication, a contaminant is any element or compound whose presence may affect the water's suitability for certain uses. For example, herbicide concentrations may exceeed the U.S. Environmental Protection Agency's (USEPA) Health Advisory Levels (HAL) for drinking water or trace-element concentrations may exceed guidelines for the protection of aquatic life. In general, the contaminats discussed in this report enter the aquatic system through nonpoint-source runoff from agricultural lands that dominate the Nebraska landscape. However,because this assessment was conducted as part of a larger, national program, a screening for contaminants with non-agricultural origins was included.The measurement of water quality involves a variety of steps, each contributing unique information while also aggregating to an overall assessment. One aspect of water-quality assesment is to describe the occurrence and distribution of contaminants. Some contaminants may be hundreds or thousands of times more concentrated in the tissues of aquatic organisms or in fine sediments than they are in the water. As a result, fish tissue and streambed sediments are well suited for the detection of certain contaminants. For example, pesticides used in the United States prior to the early 1970's, such as DDT, may have degraded into more stable but still toxic compounds that are highly concentrated in fish tissues. Conversely, other contaminants are not concentrated in sediments or tissues but are readily detected in water samples. Organonitrogen herbicides (such as atrazine), the most commonly used herbicides in Nebraska, are examples of water-soluble contaminants.Several sampling strategies were used to address specific questions. Some sites were sampled repeatedly through time and during all hydrologic conditions, whereas others were sampled only once to determine presence of contaminants. Because a strong relation between concentration and streamflow often exists for contaminants originating from nonpoint sources, streams typically were sampled near gaging stations that monitor streamflow.

Residue depletion of tilmicosin in cattle after subcutaneous administration.

PubMed

Jiang, Haiyang; Ding, Shuangyang; Li, Jiancheng; An, Dianjin; Li, Cun; Shen, Jianzhong

2006-07-12

A study of tissue residue depletion of tilmicosin in cattle was conducted after a single subcutaneous injection at the therapeutic level of 10 mg per kg body weight. Eighteen cross cattle were treated with the tilmicosin oil formulation (30%). Three treated animals (two males and one female) were selected randomly to be scarified at 1, 7, 14, 28, and 35 days withdrawal after injection. Samples of the injection site and of muscle, liver, kidney, and fat were collected. Tilmicosin residue concentrations were determined using a high-performance liquid chromatography (HPLC) method with a UV detector at 290 nm. Using a statistical method recommended by the Committee for Veterinary Medical Products of European Medical Evaluation Agency, the withdrawal time of 34 days was established when all tissue residues except samples in the injection site were below the accepted maximum residue limits.

Selection of internal reference genes for normalization of quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis in the canine brain and other organs.

PubMed

Park, Sang-Je; Huh, Jae-Won; Kim, Young-Hyun; Lee, Sang-Rae; Kim, Sang-Hyun; Kim, Sun-Uk; Kim, Heui-Soo; Kim, Min Kyu; Chang, Kyu-Tae

2013-05-01

Quantitative reverse transcription polymerase chain reaction (qRT-PCR) is a specific and sensitive technique for quantifying gene expression. To analyze qRT-PCR data accurately, suitable reference genes that show consistent expression patterns across different tissues and experimental conditions should be selected. The objective of this study was to obtain the most stable reference genes in dogs, using samples from 13 different brain tissues and 10 other organs. 16 well-known candidate reference genes were analyzed by the geNorm, NormFinder, and BestKeeper programs. Brain tissues were derived from several different anatomical regions, including the forebrain, cerebrum, diencephalon, hindbrain, and metencephalon, and grouped accordingly. Combination of the three different analyses clearly indicated that the ideal reference genes are ribosomal protien S5 (RPS5) in whole brain, RPL8 and RPS5 in whole body tissues, RPS5 and RPS19 in the forebrain and cerebrum, RPL32 and RPS19 in the diencephalon, GAPDH and RPS19 in the hindbrain, and MRPS7 and RPL13A in the metencephalon. These genes were identified as ideal for the normalization of qRT-PCR results in the respective tissues. These findings indicate more suitable and stable reference genes for future studies of canine gene expression.

Segmentation of whole cells and cell nuclei from 3-D optical microscope images using dynamic programming.

PubMed

McCullough, D P; Gudla, P R; Harris, B S; Collins, J A; Meaburn, K J; Nakaya, M A; Yamaguchi, T P; Misteli, T; Lockett, S J

2008-05-01

Communications between cells in large part drive tissue development and function, as well as disease-related processes such as tumorigenesis. Understanding the mechanistic bases of these processes necessitates quantifying specific molecules in adjacent cells or cell nuclei of intact tissue. However, a major restriction on such analyses is the lack of an efficient method that correctly segments each object (cell or nucleus) from 3-D images of an intact tissue specimen. We report a highly reliable and accurate semi-automatic algorithmic method for segmenting fluorescence-labeled cells or nuclei from 3-D tissue images. Segmentation begins with semi-automatic, 2-D object delineation in a user-selected plane, using dynamic programming (DP) to locate the border with an accumulated intensity per unit length greater that any other possible border around the same object. Then the two surfaces of the object in planes above and below the selected plane are found using an algorithm that combines DP and combinatorial searching. Following segmentation, any perceived errors can be interactively corrected. Segmentation accuracy is not significantly affected by intermittent labeling of object surfaces, diffuse surfaces, or spurious signals away from surfaces. The unique strength of the segmentation method was demonstrated on a variety of biological tissue samples where all cells, including irregularly shaped cells, were accurately segmented based on visual inspection.

Effects of selected materials and geometries on the beta dose equivalent rate in a tissue equivalent phantom immersed in infinite clouds of 133Xe

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

Piltingsrud, H.V.; Gels, G.L.

1986-06-01

Most calculations of dose equivalent (D.E.) rates at 70-micron tissue depths in tissue equivalent (T.E.) phantoms from infinite clouds (radius exceeds maximum beta range in air) of /sup 133/Xe do not consider the possible effects of clothing overlays. Consequently, a series of measurements were made using a 1-mm-thick plastic scintillation detector assembly mounted in a tissue equivalent (T.E.) phantom with an overlay of 70 micron of T.E. material. This assembly was placed in an infinite cloud containing a known concentration of /sup 133/Xe. Material samples were placed at selected distances from the detector phantom, both individually and in various combinations.more » Pulse-height spectra resulting from beta radiations were converted to relative D.E. rates at a 70-micron tissue depth. The relative D.E. rates were reduced from values with no clothing cover by as little as 45% when placing a single thin nylon cloth 1 cm from the phantom, to 94% for a T-shirt material plus wool material plus denim placed 1/2, 1 and 3 cm, respectively, from the phantom. The results indicate that even loosely fitting clothing can have an important effect on reducing the D.E. rate. Close-fitting clothing appears to provide better protection.« less

Multi-elemental imaging of paraffin-embedded human samples by laser-induced breakdown spectroscopy

NASA Astrophysics Data System (ADS)

Moncayo, S.; Trichard, F.; Busser, B.; Sabatier-Vincent, M.; Pelascini, F.; Pinel, N.; Templier, I.; Charles, J.; Sancey, L.; Motto-Ros, V.

2017-07-01

Chemical elements play central roles for physiological homeostasis in human cells, and their dysregulation might lead to a certain number of pathologies. Novel imaging techniques that improve the work of pathologists for tissue analysis and diagnostics are continuously sought. We report the use of Laser-Induced Breakdown Spectroscopy (LIBS) to perform multi-elemental images of human paraffin-embedded skin samples on the entire biopsy scale in a complementary and compatible way with microscope histopathological examination. A specific instrumental configuration is proposed in order to detect most of the elements of medical interest (i.e. P, Al, Mg, Na, Zn, Si, Fe, and Cu). As an example of medical application, we selected and analysed skin biopsies, including healthy skin tissue, cutaneous metastasis of melanoma, Merkel-cell carcinoma and squamous cell carcinoma. Clear distinctions in the distribution of chemical elements are observed from the different samples investigated. This study demonstrates the high complementarity of LIBS elemental imaging with conventional histopathology, opening new opportunities for any medical application involving metals.

AutoTag and AutoSnap: Standardized, semi-automatic capture of regions of interest from whole slide images

PubMed Central

Marien, Koen M.; Andries, Luc; De Schepper, Stefanie; Kockx, Mark M.; De Meyer, Guido R.Y.

2015-01-01

Tumor angiogenesis is measured by counting microvessels in tissue sections at high power magnification as a potential prognostic or predictive biomarker. Until now, regions of interest1 (ROIs) were selected by manual operations within a tumor by using a systematic uniform random sampling2 (SURS) approach. Although SURS is the most reliable sampling method, it implies a high workload. However, SURS can be semi-automated and in this way contribute to the development of a validated quantification method for microvessel counting in the clinical setting. Here, we report a method to use semi-automated SURS for microvessel counting: • Whole slide imaging with Pannoramic SCAN (3DHISTECH) • Computer-assisted sampling in Pannoramic Viewer (3DHISTECH) extended by two self-written AutoHotkey applications (AutoTag and AutoSnap) • The use of digital grids in Photoshop® and Bridge® (Adobe Systems) This rapid procedure allows traceability essential for high throughput protein analysis of immunohistochemically stained tissue. PMID:26150998

Study on design and cutting parameters of rotating needles for core biopsy.

PubMed

Giovannini, Marco; Ren, Huaqing; Cao, Jian; Ehmann, Kornel

2018-06-15

Core needle biopsies are widely adopted medical procedures that consist in the removal of biological tissue to better identify a lesion or an abnormality observed through a physical exam or a radiology scan. These procedures can provide significantly more information than most medical tests and they are usually performed on bone lesions, breast masses, lymph nodes and the prostate. The quality of the samples mainly depends on the forces exerted by the needle during the cutting process. The reduction of these forces is critical to extract high-quality tissue samples. The most critical factors that affect the cutting forces are the geometry of the needle tip and its motion while it is penetrating the tissue. However, optimal needle tip configurations and cutting parameters are not well established for rotating insertions. In this paper, the geometry and cutting forces of hollow needles are investigated. The fundamental goal of this study is to provide a series of guidelines for clinicians and surgeons to properly select the optimal tip geometries and speeds. Analytical models related to the cutting angles of several needle tip designs are presented and compared. Several needle tip geometries were manufactured from a 14-gauge cannula, commonly adopted during breast biopsies. The needles were then tested at different speeds and on different phantom tissues. According to these experimental measurements recommendations were formulated for rotating needle insertions. The findings of this study can be applied and extended to several biopsy procedures in which a cannula is used to extract tissue samples. Copyright © 2018 Elsevier Ltd. All rights reserved.

Radioimmunoassay of dermorphin-like peptides in mammalian and non-mammalian tissues.

PubMed

Negri, L; Melchiorri, P; Erspamer, G F; Erspamer, V

1981-01-01

A selective RIA for D-Ala2-Dermorphin (Der), a natural peptide extracted from amphibian skin, has been developed using an antibody raised in rabbits against Der which has been coupled to BSA through its phenolic hydroxyl groups of tyrosine residues with 2,4-Dichloro-6-methoxy-1,3,5-triazine. The cross-reactivity of this antibody with dermorphin analogs, C- and N-terminal fragments of dermorphin molecule, some opioid and gastrointestinal peptides was tested. Der-like immunoreactivity has been identified in tissue extracts of rats, frog and cephalopoda. Der-like peptides were purified by passing methanol extracts of the tissues through a Sephadex G25 column (16 x 100 cm) eluted with 0.1 M acetic acid at 4 degrees C. Der-like immunoreactivity from neural tissue of Dosidicus gigas, Eledone moscata, and rat brain showed a good agreement with an authentic sample of synthetic dermorphin.

Specific detection of rinderpest virus by real-time reverse transcription-PCR in preclincal and clinical samples of experimentally infected cattle

USDA-ARS?s Scientific Manuscript database

A highly sensitive detection test for Rinderpest virus (RPV), based on a real-time reverse transcription-PCR (RT-PR) system, was developed. Five different RPV genomic targets were examined, and one was selected and optimized to detect viral RNA in infected tissue culture fluid with a level of detec...

Clustering behavior in microbial communities from acute endodontic infections.

PubMed

Montagner, Francisco; Jacinto, Rogério C; Signoretti, Fernanda G C; Sanches, Paula F; Gomes, Brenda P F A

2012-02-01

Acute endodontic infections harbor heterogeneous microbial communities in both the root canal (RC) system and apical tissues. Data comparing the microbial structure and diversity in endodontic infections in related ecosystems, such as RC with necrotic pulp and acute apical abscess (AAA), are scarce in the literature. The aim of this study was to examine the presence of selected endodontic pathogens in paired samples from necrotic RC and AAA using polymerase chain reaction (PCR) followed by the construction of cluster profiles. Paired samples of RC and AAA exudates were collected from 20 subjects and analyzed by PCR for the presence of selected strict and facultative anaerobic strains. The frequency of species was compared between the RC and the AAA samples. A stringent neighboring clustering algorithm was applied to investigate the existence of similar high-order groups of samples. A dendrogram was constructed to show the arrangement of the sample groups produced by the hierarchical clustering. All samples harbored bacterial DNA. Porphyromonas endodontalis, Prevotella nigrescens, Filifactor alocis, and Tannerela forsythia were frequently detected in both RC and AAA samples. The selected anaerobic species were distributed in diverse small bacteria consortia. The samples of RC and AAA that presented at least one of the targeted microorganisms were grouped in small clusters. Anaerobic species were frequently detected in acute endodontic infections and heterogeneous microbial communities with low clustering behavior were observed in paired samples of RC and AAA. Copyright © 2012. Published by Elsevier Inc.

Endophytic fungi from selected varieties of soybean (Glycine max L. Merr.) and corn (Zea mays L.) grown in an agricultural area of Argentina.

PubMed

Russo, María L; Pelizza, Sebastián A; Cabello, Marta N; Stenglein, Sebastián A; Vianna, María F; Scorsetti, Ana C

2016-01-01

Endophytic fungi are ubiquitous and live within host plants without causing any noticeable symptoms of disease. Little is known about the diversity and function of fungal endophytes in plants, particularly in economically important species. The aim of this study was to determine the identity and diversity of endophytic fungi in leaves, stems and roots of soybean and corn plants and to determine their infection frequencies. Plants were collected in six areas of the provinces of Buenos Aires and Entre Ríos (Argentina) two areas were selected for sampling corn and four for soybean. Leaf, stem and root samples were surface-sterilized, cut into 1cm(2) pieces using a sterile scalpel and aseptically transferred to plates containing potato dextrose agar plus antibiotics. The species were identified using both morphological and molecular data. Fungal endophyte colonization in soybean plants was influenced by tissue type and varieties whereas in corn plants only by tissue type. A greater number of endophytes were isolated from stem tissues than from leaves and root tissues in both species of plants. The most frequently isolated species in all soybean cultivars was Fusarium graminearum and the least isolated one was Scopulariopsis brevicaulis. Furthermore, the most frequently isolated species in corn plants was Aspergillus terreus whereas the least isolated one was Aspergillus flavus. These results could be relevant in the search for endophytic fungi isolates that could be of interest in the control of agricultural pests. Copyright © 2016 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

Analysis of selected sugars and sugar phosphates in mouse heart tissue by reductive amination and liquid chromatography-electrospray ionization mass spectrometry.

PubMed

Han, Jun; Tschernutter, Vera; Yang, Juncong; Eckle, Tobias; Borchers, Christoph H

2013-06-18

Sensitive and reliable analysis of sugars and sugar phosphates in tissues and cells is essential for many biological and cell engineering studies. However, the successful analysis of these endogenous compounds in biological samples by liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) is often difficult because of their poor chromatographic retention properties in reversed-phase LC, the complex biological matrices, and the ionization suppression in ESI. This situation is further complicated by the existence of their multiple structural isomers in vivo. This work describes the combination of reductive amination using 3-amino-9-ethylcarbazole, with a new LC approach using a pentafluorophenyl core-shell ultrahigh performance (UP) LC column and methylphosphonic acid as an efficient tail-sweeping reagent for improved chromatographic separation. This new method was used for selected detection and accurate quantitation of the major free and phosphorylated reducing sugars in mouse heart tissue. Among the detected compounds, accurate quantitation of glyceraldehyde, ribose, glucose, glycerylaldehyde-3-phosphate, ribose-5-phosphate, glucose-6-phosphate, and mannose-6-phosphate was achieved by UPLC/multiple-reaction monitoring (MRM)-MS, with analytical accuracies ranging from 87.4% to 109.4% and CVs of ≤8.5% (n = 6). To demonstrate isotope-resolved metabolic profiling, we used UPLC/quadrupole time-of-flight (QTOF)-MS to analyze the isotope distribution patterns of C3 to C6 free and phosphorylated reducing sugars in heart tissues from (13)C-labeled wild type and knockout mice. In conclusion, the preanalytical derivatization-LC/ESI-MS method has resulted in selective determination of free and phosphorylated reducing sugars without the interferences from their nonreducing structural isomers in mouse heart tissue, with analytical sensitivities in the femtomole to low picomole range.

Use of an Additional 19-G EBUS-TBNA Needle Increases the Diagnostic Yield of EBUS-TBNA.

PubMed

Garrison, Garth; Leclair, Timothy; Balla, Agnes; Wagner, Sarah; Butnor, Kelly; Anderson, Scott R; Kinsey, C Matthew

2018-06-12

Although endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) has an excellent diagnostic yield, there remain cases where the diagnosis is not obtained. We hypothesized that additional sampling with a 19-G EBUS-TBNA needle may increase diagnostic yield in a subset of cases where additional tissue sampling was required. Indications for use of the 19-G needle following 22-G sampling with rapid on-site cytologic examination were: (1) diagnostic uncertainty of the on-site cytopathologist (eg, nondiagnostic, probable lymphoma, etc.), (2) non-small cell lung cancer with probable need for molecular genetic and/or PD-L1 testing, or (3) need for a larger tissue sample for consideration of inclusion in a research protocol. A 19-G EBUS-TBNA needle was utilized following standard sampling with a 22-G needle in 48 patients (50 sites) during the same procedure. Although the diagnostic yield between the needles was equivalent, the concordance rate was only 83%. The 19-G determined a diagnosis in 4 additional patients (8%) and provided additional histopathologic information in 6 other cases (12%). Conversely, in 3 cases (6%) diagnostic information was provided only by the 22-G needle. Compared with 22-G EBUS-TBNA alone, sampling with both the 22- and 19-G EBUS needles resulted in an increase in diagnostic yield from 92% to 99% (P=0.045) and a number needed to sample of 13 patients to provide one additional diagnosis. There were no significant complications. In select cases where additional tissue may be needed, sampling with a 19-G EBUS needle following standard aspiration with a 22-G needle results in an increase in diagnostic yield.

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.

Concentrations of mercury and other trace elements in walleye, smallmouth bass, and rainbow trout in Franklin D. Roosevelt Lake and the upper Columbia River, Washington, 1994

USGS Publications Warehouse

Munn, M.D.; Cox, S.E.; Dean, C.J.

1995-01-01

Three species of sportfish--walleye, smallmouth bass, and rainbow trout--were collected from Franklin D. Roosevelt Lake and the upstream reach of the Columbia River within the state of Washington, to determine the concentrations of mercury and other selected trace elements in fish tissue. Concentrations of total mercury in walleye fillets ranged from 0.11 to 0.44 milligram per kilogram, with the higher concentrations in the larger fish. Fillets of smallmouth bass and rainbow trout also contained mercury, but generally at lower concentrations. Other selected trace elements were found in fillet samples, but the concentrations were generally low depending on species and the specific trace element. The trace elements cadmium, copper, lead, and zinc were found in liver tissue of these same species with zinc consistently present in the highest concentration.

Expanding the therapeutic use of androgens via selective androgen receptor modulators (SARMs)

PubMed Central

Gao, Wenqing; Dalton, James T.

2007-01-01

Selective androgen receptor modulators (SARMs) are a novel class of androgen receptor (AR) ligands that might change the future of androgen therapy dramatically. With improved pharmacokinetic characteristics and tissue-selective pharmacological activities, SARMs are expected to greatly extend the clinical applications of androgens to osteoporosis, muscle wasting, male contraception and diseases of the prostate. Mechanistic studies with currently available SARMs will help to define the contributions of differential tissue distribution, tissue-specific expression of 5α-reductase, ligand-specific regulation of gene expression and AR interactions with tissue-specific coactivators to their observed tissue selectivity, and lead to even greater expansion of selective anabolic therapies. PMID:17331889

Use of a novel radiometric method to assess the inhibitory effect of donepezil on acetylcholinesterase activity in minimally diluted tissue samples

PubMed Central

Kikuchi, Tatsuya; Okamura, Toshimitsu; Arai, Takuya; Obata, Takayuki; Fukushi, Kiyoshi; Irie, Toshiaki; Shiraishi, Tetsuya

2010-01-01

Background and purpose: Cholinesterase inhibitors have been widely used for the treatment of patients with dementia. Monitoring of the cholinesterase activity in the blood is used as an indicator of the effect of the cholinesterase inhibitors in the brain. The selective measurement of cholinesterase with low tissue dilution is preferred for accurate monitoring; however, the methods have not been established. Here, we investigated the effect of tissue dilution on the action of cholinesterase inhibitors using a novel radiometric method with selective substrates, N-[14C]methylpiperidin-4-yl acetate ([14C]MP4A) and (R)-N-[14C]methylpiperidin-3-yl butyrate ([14C]MP3B_R), for AChE and butyrylcholinesterase (BChE) respectively. Experimental approach: We investigated the kinetics of hydrolysis of [14C]-MP4A and [14C]-MP3B_R by cholinesterases, and evaluated the selectivity of [14C]MP4A and [14C]MP3B_R for human AChE and BChE, respectively, compared with traditional substrates. Then, IC50 values of cholinesterase inhibitors in minimally diluted and highly diluted tissues were measured with [14C]MP4A and [14C]MP3B_R. Key results: AChE and BChE activities were selectively measured as the first-order hydrolysis rates of [14C]-MP4A and [14C]MP3B_R respectively. The AChE selectivity of [14C]MP4A was an order of magnitude higher than traditional substrates used for the AChE assay. The IC50 values of specific AChE and BChE inhibitors, donepezil and ethopropazine, in 1.2-fold diluted human whole blood were much higher than those in 120-fold diluted blood. In addition, the IC50 values of donepezil in monkey brain were dramatically decreased as the tissue was diluted. Conclusions and implications: This method would effectively monitor the activity of cholinesterase inhibitors used for therapeutics, pesticides and chemical warfare agents. PMID:20401964

Single cell gene expression profiling in Alzheimer's disease.

PubMed

Ginsberg, Stephen D; Che, Shaoli; Counts, Scott E; Mufson, Elliott J

2006-07-01

Development and implementation of microarray techniques to quantify expression levels of dozens to hundreds to thousands of transcripts simultaneously within select tissue samples from normal control subjects and neurodegenerative diseased brains has enabled scientists to create molecular fingerprints of vulnerable neuronal populations in Alzheimer's disease (AD) and related disorders. A goal is to sample gene expression from homogeneous cell types within a defined region without potential contamination by expression profiles of adjacent neuronal subpopulations and nonneuronal cells. The precise resolution afforded by single cell and population cell RNA analysis in combination with microarrays and real-time quantitative polymerase chain reaction (qPCR)-based analyses allows for relative gene expression level comparisons across cell types under different experimental conditions and disease progression. The ability to analyze single cells is an important distinction from global and regional assessments of mRNA expression and can be applied to optimally prepared tissues from animal models of neurodegeneration as well as postmortem human brain tissues. Gene expression analysis in postmortem AD brain regions including the hippocampal formation and neocortex reveals selectively vulnerable cell types share putative pathogenetic alterations in common classes of transcripts, for example, markers of glutamatergic neurotransmission, synaptic-related markers, protein phosphatases and kinases, and neurotrophins/neurotrophin receptors. Expression profiles of vulnerable regions and neurons may reveal important clues toward the understanding of the molecular pathogenesis of various neurological diseases and aid in identifying rational targets toward pharmacotherapeutic interventions for progressive, late-onset neurodegenerative disorders such as mild cognitive impairment (MCI) and AD.

Selected water-quality and biological characteristics of streams in some forested basins of North Carolina, 1985-88

USGS Publications Warehouse

Caldwell, W.S.

1992-01-01

Selected physical, chemical and biological components of streams draining undeveloped, forested basins in North Carolina were characterized on the basis of samples collected at nine sites on streams in basins that ranged in size from 0.67 to 11.2 sq mi. Water analysis included specific conductance, dissolved oxygen, water temperature, suspended sediment, pH, major dissolved constituents, nutrients, minor constituents, organochlorine insecticides, and biochemical oxygen demand. Biological characteristics included fish tissue analysis for minor constituents and synthetic organic compounds, fish community structure, and benthic macroinvertebrates. Precipitation is the source of 10 to 40% of the chloride concentration and 20 to 30% of the sulfate concentration in stormflow. Mean total nitrogen concentrations ranged from 0.16 mg/L during low-flow conditions to 1.2 mg/L during stormflow. Organic nitrogen was 60 to 85% of the total nitrogen concentration. Stream water was free of organochlorine insecticides. DDD, DDE, DDT, Lindane, and Mirex were detected in 18 of 60 samples of streambed material. About 35% of fish tissue analyses showed detectable concentrations of copper, lead, mercury and nickel. Synthetic organic chemicals were not detected in fish tissue. Fish community structure data were rated using Karr's Index of Biotic Integrity. Streams rated poor to good because of natural stresses on fish communities. Five streams in the Piedmont and mountains received excellent bioclassification ratings based on benthic macroinvertebrtate data. Two streams in the Coastal Plain rated good to fair because of natural stresses.

Effects of organism preparation in metallothionein and metal analysis in marine invertebrates for biomonitoring marine pollution.

PubMed

Oaten, J F P; Hudson, M D; Jensen, A C; Williams, I D

2015-06-15

Metallothionein (MT) is established as a potentially useful biomarker for monitoring aquatic pollution. This paper addresses widespread inconsistencies in storage conditions, tissue type selection and pre-treatment of samples before MT and metal analysis in biomarker studies. This variation hampers comparability and so the widespread implementation of this monitoring approach. Actively sampled Mytilus edulis in Southampton Water, UK were exposed to different storage temperatures, a variety of tissue types were analysed, and various pre-treatments of transportation on ice, transportation in seawater, depuration, and rapid dissection in the field were examined. Storage temperatures of -20 °C were found to be adequate for periods of at least ten weeks, as MT was not reduced by protein degradation compared with samples kept at -80 °C. Whole tissue and digestive gland concentrations of MT and metals were significantly positively correlated and directly relatable. MT in the digestive gland appeared to be more responsive to metals than in whole tissue, where it may be diluted, masking MT responses. However, longer study periods may suffer the effects of mass changes to the digestive gland, which alters MT concentration, and it may therefore be advisable to measure whole tissue. Depuration and transportation in seawater reduced both MT and metal concentrations in the digestive gland, and few correlations between MT and metals were identified for these treatments. It is therefore recommended that: i) samples are transported to the laboratory on ice and dissected as soon as possible thereafter, ii) depuration should not be used when examining MT response to metal exposure until further research clarifying its utility is reported, iii) either whole tissue or the digestive gland can be used to measure MT, though whole tissue may be preferable on long-term studies, and iv) organisms can be stored at -20 °C before analysis for up to ten weeks. These practices can be applied to future biomonitoring studies and will improve the comparability and repeatability of using MT as a biomarker. Copyright © 2015 Elsevier B.V. All rights reserved.

Optimized optical clearing method for imaging central nervous system

NASA Astrophysics Data System (ADS)

Yu, Tingting; Qi, Yisong; Gong, Hui; Luo, Qingming; Zhu, Dan

2015-03-01

The development of various optical clearing methods provides a great potential for imaging entire central nervous system by combining with multiple-labelling and microscopic imaging techniques. These methods had made certain clearing contributions with respective weaknesses, including tissue deformation, fluorescence quenching, execution complexity and antibody penetration limitation that makes immunostaining of tissue blocks difficult. The passive clarity technique (PACT) bypasses those problems and clears the samples with simple implementation, excellent transparency with fine fluorescence retention, but the passive tissue clearing method needs too long time. In this study, we not only accelerate the clearing speed of brain blocks but also preserve GFP fluorescence well by screening an optimal clearing temperature. The selection of proper temperature will make PACT more applicable, which evidently broaden the application range of this method.

Classification of prostate cancer grade using temporal ultrasound: in vivo feasibility study

NASA Astrophysics Data System (ADS)

Ghavidel, Sahar; Imani, Farhad; Khallaghi, Siavash; Gibson, Eli; Khojaste, Amir; Gaed, Mena; Moussa, Madeleine; Gomez, Jose A.; Siemens, D. Robert; Leveridge, Michael; Chang, Silvia; Fenster, Aaron; Ward, Aaron D.; Abolmaesumi, Purang; Mousavi, Parvin

2016-03-01

Temporal ultrasound has been shown to have high classification accuracy in differentiating cancer from benign tissue. In this paper, we extend the temporal ultrasound method to classify lower grade Prostate Cancer (PCa) from all other grades. We use a group of nine patients with mostly lower grade PCa, where cancerous regions are also limited. A critical challenge is to train a classifier with limited aggressive cancerous tissue compared to low grade cancerous tissue. To resolve the problem of imbalanced data, we use Synthetic Minority Oversampling Technique (SMOTE) to generate synthetic samples for the minority class. We calculate spectral features of temporal ultrasound data and perform feature selection using Random Forests. In leave-one-patient-out cross-validation strategy, an area under receiver operating characteristic curve (AUC) of 0.74 is achieved with overall sensitivity and specificity of 70%. Using an unsupervised learning approach prior to proposed method improves sensitivity and AUC to 80% and 0.79. This work represents promising results to classify lower and higher grade PCa with limited cancerous training samples, using temporal ultrasound.

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.

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.

Multiresidue analysis of sulfonamides, quinolones, and tetracyclines in animal tissues by ultra-high performance liquid chromatography-tandem mass spectrometry.

PubMed

Zhang, Zhiwen; Li, Xiaowei; Ding, Shuangyang; Jiang, Haiyang; Shen, Jianzhong; Xia, Xi

2016-08-01

A multiresidue method for the efficient identification and quantification of 38 compounds from 3 different classes of antibiotics (tetracyclines, sulfonamides, and quinolones) in animal tissues has been developed. The method optimization involved the selection of extraction solutions, comparison of different solid-phase extraction cartridges and different mobile phases. As a result, the samples were extracted with Mcllvaine and phosphate buffers, followed by clean-up step based on solid-phase extraction with Oasis HLB cartridge. All compounds were determined by ultra-high performance liquid chromatography-tandem mass spectrometry, in one single injection with a chromatographic run time of only 9min. The method efficiency was evaluated in 5 tissues including muscle, liver, and kidney, and the mean recoveries ranged from 54% to 102%, with inter-day relative standard deviation lower than 14%. The limits of quantification were between 0.5 and 10μg/kg, which were satisfactory to support future surveillance monitoring. The developed method was applied to the analysis of swine liver and chicken samples from local markets, and sulfamethazine was the most commonly detected compound in the animal samples, with the highest residue level of 998μg/kg. Copyright © 2016 Elsevier Ltd. All rights reserved.

Supercritical carbon dioxide extraction of 2,4-dichlorophenol from food crop tissues.

PubMed

Thomson, C A; Chesney, D J

1992-04-15

Supercritical fluid extraction with carbon dioxide has been found to be effective for the isolation of residue levels (0.1-1 ppm) of 2,4-dichlorophenol from selected plant tissues. The 2,4-dichlorophenol residues were incompletely extracted with supercritical CO2 alone, since a substantial fraction of the 2,4-dichlorophenol was covalently attached to the plant matrix. An acid pretreatment procedure was developed to partially hydrolyze the plant tissue prior to extraction, releasing the bound 2,4-dichlorophenol residues. Steam distillation showed higher residue levels for field-treated straw samples. This is attributed to the greater degree of hydrolysis inherent in the steam distillation procedure. Supercritical CO2 extraction of field-treated seed samples showed higher levels of 2,4-dichlorophenol residues than did steam distillation. The supercritical fluid extractant was able to solvate 2,4-dichlorophenol residues in the interior of the seed and transport them to the surface for collection. The aqueous medium used in steam distillation was unable to penetrate the hydrophobic seed matrix to the same degree. While the actual extraction time experienced in supercritical fluid extraction was far less than that of steam distillation (45 min vs 6 h, respectively), the total sample preparation time was similar in both methods.

Inflammatory cytokine expression following the use of bipolar electrocoagulation, ultracision harmonic scalpel and cold knife biopsy.

PubMed

Litta, Pietro; Saccardi, Carlo; Gizzo, Salvatore; Conte, Lorena; Ambrosi, Giulia; Sissi, Claudia; Palumbo, Manlio

2015-08-01

Electrical surgical devices may determine tissue damage through lateral thermal spread and activation of inflammatory processes. Several tissue effects are associated with the use of different surgical instruments. The aim of the present study was to compare tissue damage following the application of cold knife biopsy, bipolar electrocoagulation and the ultracision harmonic scalpel, through the analysis of inflammatory gene mediator expression. Three fragments of the round ligament (length 0.5 cm) were obtained from 22 females who had undergone total or subtotal laparoscopic hysterectomy using three different modes of resection: Cold knife biopsy, bipolar electrocoagulation and ultracision harmonic scalpel. The tissue fragments were examined by quantitative polymerase chain reaction (qPCR) analysis of selected cytokines. Gene expression analysis demonstrated large standard deviations due to individual variability among patients and indicated variability in the concentrations of cytokines in the three different samples. The quantity of cytokine mRNA in the cold knife biopsy samples was generally greater than those obtained by other techniques. Tumor necrosis factor-α expression was significantly higher in the sample obtained with the ultracision harmonic scalpel and bipolar electrocoagulation (P=0.033) when compared with cold knife biopsy. The inflammatory response was analyzed by the quantification of gene expression through the use of qPCR. The ultracision harmonic scalpel and bipolar electrocoagulation triggered the inflammatory cascade and resulted in an increased production of cytokines compared with cold knife biopsy.

[Distribution of aconitum alkaloids in the corpse died of acute aconite intoxication].

PubMed

Liu, Wei; Shen, Min; Qin, Zhi-Qiang

2009-06-01

To investigate the distribution of aconite alkaloids in biological fluids and tissues in the corpse died of acute aconite intoxication and to provide information for sample selection and result evaluation in forensic identification. The content of aconite alkaloids in biological fluids and tissues were determined by liquid chromatography-tandem mass spectrometry. The content of aconite displayed in decending order of urine, bile, gastric content, heart blood, pancreas, heart, intestine, liver, kidney, stomach, lung, gallbladder and spleen, with no aconite detected in the brain. It was indicated that urine, bile and blood are the best specimens for the determination of aconite in body of the acute aconite intoxication.

Effects of whirling disease on selected hematological parameters in rainbow trout

USGS Publications Warehouse

Densmore, Christine L.; Blazer, V.S.; Waldrop, T.B.; Pooler, P.S.

2001-01-01

Hematological responses to whirling disease in rainbow trout (Oncorhynchus mykiss) were investigated. Two-mo-old fingerling rainbow trout were exposed to cultured triactinomyxon spores of Myxobolus cerebralis at 9,000 spores/fish in December, 1997. Twenty-four wks post-exposure, fish were taken from infected and uninfected groups for peripheral blood and cranial tissue sampling. Histological observations on cranial tissues confirmed M. cerebralis infection in all exposed fish. Differences in hematological parameters between the two groups included significantly lower total leukocyte and small lymphocyte counts for the infected fish. No effects on hematocrit, plasma protein concentration, or other differential leukocyte counts were noted.

Biochemical Fractionation and Stable Isotope Dilution Liquid Chromatography-mass Spectrometry for Targeted and Microdomain-specific Protein Quantification in Human Postmortem Brain Tissue*

PubMed Central

MacDonald, Matthew L.; Ciccimaro, Eugene; Prakash, Amol; Banerjee, Anamika; Seeholzer, Steven H.; Blair, Ian A.; Hahn, Chang-Gyu

2012-01-01

Synaptic architecture and its adaptive changes require numerous molecular events that are both highly ordered and complex. A majority of neuropsychiatric illnesses are complex trait disorders, in which multiple etiologic factors converge at the synapse via many signaling pathways. Investigating the protein composition of synaptic microdomains from human patient brain tissues will yield valuable insights into the interactions of risk genes in many disorders. These types of studies in postmortem tissues have been limited by the lack of proper study paradigms. Thus, it is necessary not only to develop strategies to quantify protein and post-translational modifications at the synapse, but also to rigorously validate them for use in postmortem human brain tissues. In this study we describe the development of a liquid chromatography-selected reaction monitoring method, using a stable isotope-labeled neuronal proteome standard prepared from the brain tissue of a stable isotope-labeled mouse, for the multiplexed quantification of target synaptic proteins in mammalian samples. Additionally, we report the use of this method to validate a biochemical approach for the preparation of synaptic microdomain enrichments from human postmortem prefrontal cortex. Our data demonstrate that a targeted mass spectrometry approach with a true neuronal proteome standard facilitates accurate and precise quantification of over 100 synaptic proteins in mammalian samples, with the potential to quantify over 1000 proteins. Using this method, we found that protein enrichments in subcellular fractions prepared from human postmortem brain tissue were strikingly similar to those prepared from fresh mouse brain tissue. These findings demonstrate that biochemical fractionation methods paired with targeted proteomic strategies can be used in human brain tissues, with important implications for the study of neuropsychiatric disease. PMID:22942359

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

Bulk Genotyping of Biopsies Can Create Spurious Evidence for Hetereogeneity in Mutation Content.

PubMed

Kostadinov, Rumen; Maley, Carlo C; Kuhner, Mary K

2016-04-01

When multiple samples are taken from the neoplastic tissues of a single patient, it is natural to compare their mutation content. This is often done by bulk genotyping of whole biopsies, but the chance that a mutation will be detected in bulk genotyping depends on its local frequency in the sample. When the underlying mutation count per cell is equal, homogenous biopsies will have more high-frequency mutations, and thus more detectable mutations, than heterogeneous ones. Using simulations, we show that bulk genotyping of data simulated under a neutral model of somatic evolution generates strong spurious evidence for non-neutrality, because the pattern of tissue growth systematically generates differences in biopsy heterogeneity. Any experiment which compares mutation content across bulk-genotyped biopsies may therefore suggest mutation rate or selection intensity variation even when these forces are absent. We discuss computational and experimental approaches for resolving this problem.

Discovery of Colorectal Cancer Biomarker Candidates by Membrane Proteomic Analysis and Subsequent Verification using Selected Reaction Monitoring (SRM) and Tissue Microarray (TMA) Analysis*

PubMed Central

Kume, Hideaki; Muraoka, Satoshi; Kuga, Takahisa; Adachi, Jun; Narumi, Ryohei; Watanabe, Shio; Kuwano, Masayoshi; Kodera, Yoshio; Matsushita, Kazuyuki; Fukuoka, Junya; Masuda, Takeshi; Ishihama, Yasushi; Matsubara, Hisahiro; Nomura, Fumio; Tomonaga, Takeshi

2014-01-01

Recent advances in quantitative proteomic technology have enabled the large-scale validation of biomarkers. We here performed a quantitative proteomic analysis of membrane fractions from colorectal cancer tissue to discover biomarker candidates, and then extensively validated the candidate proteins identified. A total of 5566 proteins were identified in six tissue samples, each of which was obtained from polyps and cancer with and without metastasis. GO cellular component analysis predicted that 3087 of these proteins were membrane proteins, whereas TMHMM algorithm predicted that 1567 proteins had a transmembrane domain. Differences were observed in the expression of 159 membrane proteins and 55 extracellular proteins between polyps and cancer without metastasis, while the expression of 32 membrane proteins and 17 extracellular proteins differed between cancer with and without metastasis. A total of 105 of these biomarker candidates were quantitated using selected (or multiple) reaction monitoring (SRM/MRM) with stable synthetic isotope-labeled peptides as an internal control. The results obtained revealed differences in the expression of 69 of these proteins, and this was subsequently verified in an independent set of patient samples (polyps (n = 10), cancer without metastasis (n = 10), cancer with metastasis (n = 10)). Significant differences were observed in the expression of 44 of these proteins, including ITGA5, GPRC5A, PDGFRB, and TFRC, which have already been shown to be overexpressed in colorectal cancer, as well as proteins with unknown function, such as C8orf55. The expression of C8orf55 was also shown to be high not only in colorectal cancer, but also in several cancer tissues using a multicancer tissue microarray, which included 1150 cores from 14 cancer tissues. This is the largest verification study of biomarker candidate membrane proteins to date; our methods for biomarker discovery and subsequent validation using SRM/MRM will contribute to the identification of useful biomarker candidates for various cancers. Data are available via ProteomeXchange with identifier PXD000851. PMID:24687888

Discovery of colorectal cancer biomarker candidates by membrane proteomic analysis and subsequent verification using selected reaction monitoring (SRM) and tissue microarray (TMA) analysis.

PubMed

Kume, Hideaki; Muraoka, Satoshi; Kuga, Takahisa; Adachi, Jun; Narumi, Ryohei; Watanabe, Shio; Kuwano, Masayoshi; Kodera, Yoshio; Matsushita, Kazuyuki; Fukuoka, Junya; Masuda, Takeshi; Ishihama, Yasushi; Matsubara, Hisahiro; Nomura, Fumio; Tomonaga, Takeshi

2014-06-01

Recent advances in quantitative proteomic technology have enabled the large-scale validation of biomarkers. We here performed a quantitative proteomic analysis of membrane fractions from colorectal cancer tissue to discover biomarker candidates, and then extensively validated the candidate proteins identified. A total of 5566 proteins were identified in six tissue samples, each of which was obtained from polyps and cancer with and without metastasis. GO cellular component analysis predicted that 3087 of these proteins were membrane proteins, whereas TMHMM algorithm predicted that 1567 proteins had a transmembrane domain. Differences were observed in the expression of 159 membrane proteins and 55 extracellular proteins between polyps and cancer without metastasis, while the expression of 32 membrane proteins and 17 extracellular proteins differed between cancer with and without metastasis. A total of 105 of these biomarker candidates were quantitated using selected (or multiple) reaction monitoring (SRM/MRM) with stable synthetic isotope-labeled peptides as an internal control. The results obtained revealed differences in the expression of 69 of these proteins, and this was subsequently verified in an independent set of patient samples (polyps (n = 10), cancer without metastasis (n = 10), cancer with metastasis (n = 10)). Significant differences were observed in the expression of 44 of these proteins, including ITGA5, GPRC5A, PDGFRB, and TFRC, which have already been shown to be overexpressed in colorectal cancer, as well as proteins with unknown function, such as C8orf55. The expression of C8orf55 was also shown to be high not only in colorectal cancer, but also in several cancer tissues using a multicancer tissue microarray, which included 1150 cores from 14 cancer tissues. This is the largest verification study of biomarker candidate membrane proteins to date; our methods for biomarker discovery and subsequent validation using SRM/MRM will contribute to the identification of useful biomarker candidates for various cancers. Data are available via ProteomeXchange with identifier PXD000851. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Porphyromonas gingivalis, Treponema denticola and toll-like receptor 2 are associated with hypertensive disorders in placental tissue: a case-control study.

PubMed

Chaparro, A; Blanlot, C; Ramírez, V; Sanz, A; Quintero, A; Inostroza, C; Bittner, M; Navarro, M; Illanes, S E

2013-12-01

To explore the associations between the presence of periodontal pathogens and the expression of toll-like receptors (TLR-2 and TLR-4) in the placental tissue of patients with hypertensive disorders compared to the placentas of healthy normotensive patients. A case-control study was performed. From a cohort composed of 126 pregnant women, 33 normotensive healthy pregnant women were randomly selected, and 25 cases of patients with hypertensive disorders of pregnancy, including gestational hypertension and pre-eclampsia, were selected. Placental biopsy was obtained after aseptic placental collection at the time of delivery. All of the samples were processed and analysed for the detection of Porphyromonas gingivalis, Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum, Treponema denticola and Tannerella forsythia using the polymerase chain reaction (PCR) technique. Determination of the expressions of TLR-2 and TLR-4 was performed in samples of total purified protein isolated from placental tissues and analysed by ELISA. The data were assessed using descriptive statistics. The associations among variables were estimated through multiple logistic regression models and the Mann-Whitney test to evaluate the differences between the two groups. A significant increase was observed in the expression of TLR-2 in the placentas of patients with hypertensive disorders (p = 0.04). Additionally, the multiple logistic regression models demonstrated an association between the presence of T. denticola and P. gingivalis in placental tissues and hypertensive disorders (OR: 9.39, p = 0.001, CI 95% 2.39-36.88 and OR: 7.59, p = 0.019, CI 95% 1.39-41.51, respectively). In the present study, pregnant women with periodontal disease presented an association in the placental tissue between the presence of T. denticola and P. gingivalis and hypertensive disorders. Additionally, increased expression of TLR-2 was observed. However, further studies are required to determine the specific roles of periodontal pathogens and TLRs in the placental tissue of patients with pregnancy-related hypertensive disorders. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Tissue Banking, Bioinformatics, and Electronic Medical Records: The Front-End Requirements for Personalized Medicine

PubMed Central

Suh, K. Stephen; Sarojini, Sreeja; Youssif, Maher; Nalley, Kip; Milinovikj, Natasha; Elloumi, Fathi; Russell, Steven; Pecora, Andrew; Schecter, Elyssa; Goy, Andre

2013-01-01

Personalized medicine promises patient-tailored treatments that enhance patient care and decrease overall treatment costs by focusing on genetics and “-omics” data obtained from patient biospecimens and records to guide therapy choices that generate good clinical outcomes. The approach relies on diagnostic and prognostic use of novel biomarkers discovered through combinations of tissue banking, bioinformatics, and electronic medical records (EMRs). The analytical power of bioinformatic platforms combined with patient clinical data from EMRs can reveal potential biomarkers and clinical phenotypes that allow researchers to develop experimental strategies using selected patient biospecimens stored in tissue banks. For cancer, high-quality biospecimens collected at diagnosis, first relapse, and various treatment stages provide crucial resources for study designs. To enlarge biospecimen collections, patient education regarding the value of specimen donation is vital. One approach for increasing consent is to offer publically available illustrations and game-like engagements demonstrating how wider sample availability facilitates development of novel therapies. The critical value of tissue bank samples, bioinformatics, and EMR in the early stages of the biomarker discovery process for personalized medicine is often overlooked. The data obtained also require cross-disciplinary collaborations to translate experimental results into clinical practice and diagnostic and prognostic use in personalized medicine. PMID:23818899

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

The Structure Of Intact Side Tissue Loss Based On FTIR Spectroscopic Measurements

NASA Astrophysics Data System (ADS)

Hussain, N.; Al-Hadithi, K. O.; Jaafar, M. S.

2009-09-01

Laser applications in dentistry were strongly evolved during the last three decades. Among those applications are laser ablation of dental hard tissue, caries inhibition treatments by localized surface heating, and surface conditioning for bonding. In addition, infra-red lasers are ideally suited for the selective and precise removal of carious dental hard tissue while minimizing the healthy tissue loss. In the present study we applied laser spectroscopy technique FTIR for the study of the structure of intact side tissue of teeth. The aim of the recent work is to study the effect of race and sex (genealogy) on the structure of intact side tissue loss. Our sample consists of twenty Malay females' teeth where the FTIR has been applied. The data show a decrease in the amounts of main substances (like Hydroxyapatite crystals ([Ca5(PO4)3(OH)4], CaF2) than those in healthy teeth. The measured spectra represent the enamel with the characteristic peaks due to the phosphate group in carbonated, hydroxyapatite at 1000 cm-1 and two small peaks near 1500 cm-1 due to the carbonate group. The data explains the effect of the several factors on the intact side tissue loss.

The effect of electronically steering a phased array ultrasound transducer on near-field tissue heating.

PubMed

Payne, Allison; Vyas, Urvi; Todd, Nick; de Bever, Joshua; Christensen, Douglas A; Parker, Dennis L

2011-09-01

This study presents the results obtained from both simulation and experimental techniques that show the effect of mechanically or electronically steering a phased array transducer on proximal tissue heating. The thermal response of a nine-position raster and a 16-mm diameter circle scanning trajectory executed through both electronic and mechanical scanning was evaluated in computer simulations and experimentally in a homogeneous tissue-mimicking phantom. Simulations were performed using power deposition maps obtained from the hybrid angular spectrum (HAS) method and applying a finite-difference approximation of the Pennes' bioheat transfer equation for the experimentally used transducer and also for a fully sampled transducer to demonstrate the effect of acoustic window, ultrasound beam overlap and grating lobe clutter on near-field heating. Both simulation and experimental results show that electronically steering the ultrasound beam for the two trajectories using the 256-element phased array significantly increases the thermal dose deposited in the near-field tissues when compared with the same treatment executed through mechanical steering only. In addition, the individual contributions of both beam overlap and grating lobe clutter to the near-field thermal effects were determined through comparing the simulated ultrasound beam patterns and resulting temperature fields from mechanically and electronically steered trajectories using the 256-randomized element phased array transducer to an electronically steered trajectory using a fully sampled transducer with 40 401 phase-adjusted sample points. Three distinctly different three distinctly different transducers were simulated to analyze the tradeoffs of selected transducer design parameters on near-field heating. Careful consideration of design tradeoffs and accurate patient treatment planning combined with thorough monitoring of the near-field tissue temperature will help to ensure patient safety during an MRgHIFU treatment.

Effect of Carotenoid Supplemented Formula on Carotenoid Bioaccumulation in Tissues of Infant Rhesus Macaques: A Pilot Study Focused on Lutein

PubMed Central

Jeon, Sookyoung; Neuringer, Martha; Johnson, Emily E.; Kuchan, Matthew J.; Pereira, Suzette L.; Johnson, Elizabeth J.; Erdman, John W.

2017-01-01

Lutein is the predominant carotenoid in the developing primate brain and retina, and may have important functional roles. However, its bioaccumulation pattern during early development is not understood. In this pilot study, we investigated whether carotenoid supplementation of infant formula enhanced lutein tissue deposition in infant rhesus macaques. Monkeys were initially breastfed; from 1 to 3 months of age they were fed either a formula supplemented with lutein, zeaxanthin, β-carotene and lycopene, or a control formula with low levels of these carotenoids, for 4 months (n = 2/group). All samples were analyzed by high pressure liquid chromatography (HPLC). Final serum lutein in the supplemented group was 5 times higher than in the unsupplemented group. All brain regions examined showed a selective increase in lutein deposition in the supplemented infants. Lutein differentially accumulated across brain regions, with highest amounts in occipital cortex in both groups. β-carotene accumulated, but zeaxanthin and lycopene were undetectable in any brain region. Supplemented infants had higher lutein concentrations in peripheral retina but not in macular retina. Among adipose sites, abdominal subcutaneous adipose tissue exhibited the highest lutein level and was 3-fold higher in the supplemented infants. The supplemented formula enhanced carotenoid deposition in several other tissues. In rhesus infants, increased intake of carotenoids from formula enhanced their deposition in serum and numerous tissues and selectively increased lutein in multiple brain regions. PMID:28075370

Influence of water content on the laser-induced breakdown spectroscopy analysis of human cell pellet

NASA Astrophysics Data System (ADS)

Moon, Youngmin; Han, Jung Hyun; Lee, Jong Jin; Jeong, Sungho

2015-12-01

The effects of water content change in a biological sample on the emission signal intensity and intensity ratio during LIBS analysis were investigated. To examine the effects of water content only avoiding matrix effects, a homogeneous human cell pellet consisting of cultured human immortalized keratinocyte cell only was used as the sample. LIBS spectra of the human cell pellet sample produced with a Q-switched Nd:YAG laser (λ = 532 nm, τ = 5 ns, top-hat profile) and a six-channel CCD spectrometer (spectral range = 187-1045 nm, spectral resolution = 0.1 nm) revealed that most of the emission lines observed from a tissue sample were also observable from the human cell pellet. The intensity and intensity ratio of the emission lines varied significantly as the water content of the human cell pellet was changed. It was found that a typically selected internal standard in LIBS analysis of biological samples such as carbon could produce inconsistent results, whereas the ratio of properly selected emission lines such as Mg(II) 280.270 nm and Ca(II) 396.847 nm was nearly independent of sample water content.

Report of the clinical donor case workshop of the European Association of Tissue Banks annual meeting 2012.

PubMed

Beele, Hilde; van Wijk, Marja J; Parker, Robert; Sánchez-Ibáňez, Jacinto; Brubaker, Scott A; Wulff, Birgit; Richters, Cornelia D; Cox, Mike; Warwick, Ruth M; Eastlund, Ted

2013-12-01

The European Association of Tissue Banks (EATB) donor case workshop is a forum held within the program of the EATB annual congress. The workshop offers an opportunity to discuss and evaluate approaches taken to challenging situations regarding donor selection, it promotes consensus development in deciding tissue donor acceptability when donor health issues are not addressed in standards and regulations, and serves to strengthen the professional tissue banking networks across Europe and beyond. This report reflects some of the discussion at the workshop during the annual congress in Vienna in 2012. The cases presented dealt with problems encountered by tissue bank facilities concerning idiopathic thrombocytopenia and auto-immune disorders, hemodilution and blood sample identification, premalignant and malignant lesions, and Huntington's disease. The discussions during the workshop demonstrate that the implications on the safety of tissue transplantation of various tissue donor illnesses, physical findings and behaviours, and the preventive measures taken by tissue facilities, may not always be agreed by tissue facility medical directors and other professionals. Moreover, they reveal that operating procedures, regulations and standards cannot comprehensively cover all tissue donor findings, medical histories and circumstances surrounding the cause of death. For many of the issues raised, there is a need for scientific research to provide a better evidence base for future deliberations about the suitability and eligibility of tissue allograft donors.

Provision Of Carbon Nanotube Bucky Paper Cages For Immune Shielding Of Cells, Tissues, and Medical Devices

NASA Technical Reports Server (NTRS)

Loftus, David J. (Inventor)

2006-01-01

System and method for enclosing cells and/or tissue, for purposes of growth, cell differentiation, suppression of cell differentiation, biological processing and/or transplantation of cells and tissues (biological inserts), and for secretion, sensing and monitoring of selected chemical substances and activation of gene expression of biological inserts implanted into a human body. Selected cells and/or tissue are enveloped in a "cage" that is primarily carbon nanotube Bucky paper, with a selected thickness and porosity. Optionally, selected functional groups, proteins and/or peptides are attached to the carbon nanotube cage, or included within the cage, to enhance the growth and/or differentiation of the cells and/or tissue, to select for certain cellular sub-populations, to optimize certain functions of the cells and/or tissue and/or to optimize the passage of chemicals across the cage surface(s). A cage system is also used as an immuns shield and to control operation of a nano-device or macroscopic device, located within the cage, to provide or transform a selected chemical and/or a selected signal.

Selective and self-guided micro-ablation of tissue with plasmonic nanobubbles

PubMed Central

Lukianova-Hleb, Ekaterina Y.; Koneva, Irina I.; Oginsky, Alexander O.; La Francesca, Saverio; Lapotko, Dmitri O.

2010-01-01

Background The accuracy, selectivity and safety of surgical and laser methods for tissue elimination are often limited at microscale. Materials and methods We developed a novel agent, the plasmonic nanobubble (PNB), for optically guided selective elimination of the target tissue with micrometer precision. PNBs were tested in vitro in the two different models of superficial tumors and vascular plaques. Results PNBs were selectively generated around gold nanoparticles (delivered to the target tissues) with short laser pulses. Monolayers of cancerous cells and atherosclerotic plaque tissue were eliminated with PNBs with micrometer accuracy and without thermal and mechanical damage to collateral normal tissues. The effect of the PNB was dynamically controlled through the fluence of laser pulses (532 nm, duration 0.5 and 10 ns) and was guided through the optical scattering by PNB. Conclusions plasmonic nanobubbles were shown to provide precise, tunable, selective and guided ablation of tissue at a microcscopic level and could be employed as a new generation of surgical tools. PMID:21176913

Selective and self-guided micro-ablation of tissue with plasmonic nanobubbles.

PubMed

Lukianova-Hleb, Ekaterina Y; Koneva, Irina I; Oginsky, Alexander O; La Francesca, Saverio; Lapotko, Dmitri O

2011-03-01

The accuracy, selectivity, and safety of surgical and laser methods for tissue elimination are often limited at microscale. We developed a novel agent, the plasmonic nanobubble (PNB), for optically guided selective elimination of the target tissue with micrometer precision. PNBs were tested in vitro in the two different models of superficial tumors and vascular plaques. PNBs were selectively generated around gold nanoparticles (delivered to the target tissues) with short laser pulses. Monolayers of cancerous cells and atherosclerotic plaque tissue were eliminated with PNBs with micrometer accuracy and without thermal and mechanical damage to collateral normal tissues. The effect of the PNB was dynamically controlled through the fluence of laser pulses (532 nm, duration 0.5 and 10 ns) and was guided through the optical scattering by PNB. Plasmonic nanobubbles were shown to provide precise, tunable, selective, and guided ablation of tissue at a microscopic level and could be employed as a new generation of surgical tools. Copyright © 2011 Elsevier Inc. All rights reserved.

Expression Analysis of Previously Verified Fecal and Plasma Dow-regulated MicroRNAs (miR-4478, 1295-3p, 142-3p and 26a-5p), in FFPE Tissue Samples of CRC Patients.

PubMed

Ghanbari, Reza; Rezasoltani, Sama; Hashemi, Javad; Mohamadkhani, Ashraf; Tahmasebifar, Arash; Arefian, Ehsan; Mobarra, Naser; Asadi, Jahanbakhsh; Nazemalhosseini Mojarad, Ehsan; Yazdani, Yaghoub; Knuutila, Sakari; Malekzadeh, Reza

2017-02-01

Colorectal cancer (CRC) is one of the most common causes of cancer-related mortality worldwide. Early diagnosis of this neoplasm is critical and may reduce patients' mortality. MicroRNAs are small non-coding RNA molecules whose expression pattern can be altered in various diseases such as CRC. In this study, we evaluated the expression levels of miR-142-3p, miR-26a-5p (their reduced expression in plasma samples of CRC patients was previously confirmed), miR-4478 and miR-1295-3p (their reduced expression in stool samples of CRC patients was previously confirmed) in tissue samples of CRC patients in comparison to healthy subjects. To achieve this purpose, total RNA including small RNA was extracted from 53 CRC and 35 normal subjects' Formalin-fixed, Paraffin-embedded (FFPE) tissue samples using the miRNeasy FFPE Mini Kit. The expression levels of these four selected miRNAs were measured using quantitative Reverse Transcriptase Polymerase Chain Reaction (qRT-PCR). We found that the expression levels of miR-4478 and miR-1295b-3p (two previously down-regulated fecal miRNAs) were significantly decreased in FFPE samples of CRC patients compared to healthy controls. On the other hand, no significant differences were seen in expression levels of miR-142-3p and miR-26a-5p (two previously down-regulated circulating miRNAs) in FFPE samples between these two groups. Regarding current findings, it may be concluded that to diagnose CRC patients based on the miRNAs approach, stool samples are more likely preferable to plasma samples; nevertheless, additional studies with more samples are needed to confirm the results.

High levels of bioplastic are produced in fertile transplastomic tobacco plants engineered with a synthetic operon for the production of polyhydroxybutyrate.

PubMed

Bohmert-Tatarev, Karen; McAvoy, Susan; Daughtry, Sean; Peoples, Oliver P; Snell, Kristi D

2011-04-01

An optimized genetic construct for plastid transformation of tobacco (Nicotiana tabacum) for the production of the renewable, biodegradable plastic polyhydroxybutyrate (PHB) was designed using an operon extension strategy. Bacterial genes encoding the PHB pathway enzymes were selected for use in this construct based on their similarity to the codon usage and GC content of the tobacco plastome. Regulatory elements with limited homology to the host plastome yet known to yield high levels of plastidial recombinant protein production were used to enhance the expression of the transgenes. A partial transcriptional unit, containing genes of the PHB pathway and a selectable marker gene encoding spectinomycin resistance, was flanked at the 5' end by the host plant's psbA coding sequence and at the 3' end by the host plant's 3' psbA untranslated region. This design allowed insertion of the transgenes into the plastome as an extension of the psbA operon, rendering the addition of a promoter to drive the expression of the transgenes unnecessary. Transformation of the optimized construct into tobacco and subsequent spectinomycin selection of transgenic plants yielded T0 plants that were capable of producing up to 18.8% dry weight PHB in samples of leaf tissue. These plants were fertile and produced viable seed. T1 plants producing up to 17.3% dry weight PHB in samples of leaf tissue and 8.8% dry weight PHB in the total biomass of the plant were also isolated.

Investigation of the "true" extraction recovery of analytes from multiple types of tissues and its impact on tissue bioanalysis using two model compounds.

PubMed

Yuan, Long; Ma, Li; Dillon, Lisa; Fancher, R Marcus; Sun, Huadong; Zhu, Mingshe; Lehman-McKeeman, Lois; Aubry, Anne-Françoise; Ji, Qin C

2016-11-16

LC-MS/MS has been widely applied to the quantitative analysis of tissue samples. However, one key remaining issue is that the extraction recovery of analyte from spiked tissue calibration standard and quality control samples (QCs) may not accurately represent the "true" recovery of analyte from incurred tissue samples. This may affect the accuracy of LC-MS/MS tissue bioanalysis. Here, we investigated whether the recovery determined using tissue QCs by LC-MS/MS can accurately represent the "true" recovery from incurred tissue samples using two model compounds: BMS-986104, a S1P 1 receptor modulator drug candidate, and its phosphate metabolite, BMS-986104-P. We first developed a novel acid and surfactant assisted protein precipitation method for the extraction of BMS-986104 and BMS-986104-P from rat tissues, and determined their recoveries using tissue QCs by LC-MS/MS. We then used radioactive incurred samples from rats dosed with 3 H-labeled BMS-986104 to determine the absolute total radioactivity recovery in six different tissues. The recoveries determined using tissue QCs and incurred samples matched with each other very well. The results demonstrated that, in this assay, tissue QCs accurately represented the incurred tissue samples to determine the "true" recovery, and LC-MS/MS assay was accurate for tissue bioanalysis. Another aspect we investigated is how the tissue QCs should be prepared to better represent the incurred tissue samples. We compared two different QC preparation methods (analyte spiked in tissue homogenates or in intact tissues) and demonstrated that the two methods had no significant difference when a good sample preparation was in place. The developed assay showed excellent accuracy and precision, and was successfully applied to the quantitative determination of BMS-986104 and BMS-986104-P in tissues in a rat toxicology study. Copyright © 2016 Elsevier B.V. All rights reserved.

Mercury concentrations in water and mercury and selenium concentrations in fish from Brownlee Reservoir and selected sites in the Boise and Snake Rivers, Idaho and Oregon, 2013–15

USGS Publications Warehouse

Williams, Marshall L.; MacCoy, Dorene E.

2016-06-30

Mercury (Hg) analyses were conducted on samples of sport fish and water collected from selected sampling sites in Brownlee Reservoir and the Boise and Snake Rivers to meet National Pollution Discharge and Elimination System (NPDES) permit requirements for the City of Boise, Idaho, between 2013 and 2015. City of Boise personnel collected water samples from six sites between October and November 2013 and 2015, with one site sampled in 2014. Total Hg concentrations in unfiltered water samples ranged from 0.48 to 8.8 nanograms per liter (ng/L), with the highest value in Brownlee Reservoir in 2013. All Hg concentrations in water samples were less than the U.S. Environmental Protection Agency (USEPA) Hg chronic aquatic life criterion of 12 ng/L.The USEPA recommended a water-quality criterion of 0.30 milligrams per kilogram (mg/kg) methylmercury (MeHg) expressed as a fish-tissue residue value (wet-weight MeHg in fish tissue). The Idaho Department of Environmental Quality adopted the USEPA’s fish-tissue criterion and established a reasonable potential to exceed (RPTE) threshold 20 percent lower than the criterion or greater than 0.24 mg/kg Hg based on an average concentration of 10 fish from a receiving waterbody. NPDES permitted discharge to waters with fish having Hg concentrations exceeding 0.24 mg/kg are said to have a reasonable potential to exceed the water-quality criterion and thus are subject to additional permit obligations, such as requirements for increased monitoring and the development of a Hg minimization plan. The Idaho Fish Consumption Advisory Program (IFCAP) issues fish advisories to protect general and sensitive populations of fish consumers and has developed an action level of 0.22 mg/kg Hg in fish tissue. Fish consumption advisories are water body- and species-specific and are used to advise allowable fish consumption from specific water bodies. The geometric mean Hg concentration of 10 fish of a single species collected from a single water body (lake or stream) in Idaho is compared to the action level to determine if a fish consumption advisory should be issued.The U.S. Geological Survey collected and analyzed individual fillets of mountain whitefish (Prosopium williamsoni), rainbow trout (Oncorhynchus mykiss), smallmouth bass (Micropterus dolomieu), and channel catfish (Ictalurus punctatus) for Hg. The 2013 average Hg concentration for small mouth bass (0.32 mg/kg) collected at Brownlee Reservoir and for channel catfish (0.33 mg/kg) collected at the Boise River mouth, exceeded the Idaho water quality criterion (>0.3 mg/kg), the Hg RPTE threshold (>0.24 mg/kg), and the IFCAP action level (>0.22 mg/kg). Average Hg concentrations in fish collected in 2014 or 2015 did not exceed evaluation criteria for any of the species assessed.Selenium (Se) analysis was conducted on one composite fish tissue sample per site to assess general concentrations and to provide information for future risk assessments. Composite concentrations of Se in fish tissue collected between 2013 and 2015 ranged from 0.07 and 0.49 mg/kg wet weight with the highest concentration collected from smallmouth bass from the Snake River near Murphy, and the lowest from mountain whitefish from the Boise River at Eckert Road.

Application of neutron capture autoradiography to Boron Delivery seeking techniques for selective accumulation of boron compounds to tumor with intra-arterial administration of boron entrapped water-in-oil-in-water emulsion

NASA Astrophysics Data System (ADS)

Mikado, S.; Yanagie, H.; Yasuda, N.; Higashi, S.; Ikushima, I.; Mizumachi, R.; Murata, Y.; Morishita, Y.; Nishimura, R.; Shinohara, A.; Ogura, K.; Sugiyama, H.; Iikura, H.; Ando, H.; Ishimoto, M.; Takamoto, S.; Eriguchi, M.; Takahashi, H.; Kimura, M.

2009-06-01

It is necessary to accumulate the 10B atoms selectively to the tumor cells for effective Boron Neutron Capture Therapy (BNCT). In order to achieve an accurate measurement of 10B accumulations in the biological samples, we employed a technique of neutron capture autoradiography (NCAR) of sliced samples of tumor tissues using CR-39 plastic track detectors. The CR-39 track detectors attached with the biological samples were exposed to thermal neutrons in the thermal column of the JRR3 of Japan Atomic Energy Agency (JAEA). We obtained quantitative NCAR images of the samples for VX-2 tumor in rabbit liver after injection of 10BSH entrapped water-in-oil-in-water (WOW) emulsion by intra-arterial injection via proper hepatic artery. The 10B accumulations and distributions in VX-2 tumor and normal liver of rabbit were investigated by means of alpha-track density measurements. In this study, we showed the selective accumulation of 10B atoms in the VX-2 tumor by intra-arterial injection of 10B entrapped WOW emulsion until 3 days after injection by using digitized NCAR images (i.e. alpha-track mapping).

International recommendation for a comprehensive neuropathologic workup of epilepsy surgery brain tissue: A consensus Task Force report from the ILAE Commission on Diagnostic Methods.

PubMed

Blümcke, Ingmar; Aronica, Eleonora; Miyata, Hajime; Sarnat, Harvey B; Thom, Maria; Roessler, Karl; Rydenhag, Bertil; Jehi, Lara; Krsek, Pavel; Wiebe, Samuel; Spreafico, Roberto

2016-03-01

Epilepsy surgery is an effective treatment in many patients with drug-resistant focal epilepsies. An early decision for surgical therapy is facilitated by a magnetic resonance imaging (MRI)-visible brain lesion congruent with the electrophysiologically abnormal brain region. Recent advances in the pathologic diagnosis and classification of epileptogenic brain lesions are helpful for clinical correlation, outcome stratification, and patient management. However, application of international consensus classification systems to common epileptic pathologies (e.g., focal cortical dysplasia [FCD] and hippocampal sclerosis [HS]) necessitates standardized protocols for neuropathologic workup of epilepsy surgery specimens. To this end, the Task Force of Neuropathology from the International League Against Epilepsy (ILAE) Commission on Diagnostic Methods developed a consensus standard operational procedure for tissue inspection, distribution, and processing. The aims are to provide a systematic framework for histopathologic workup, meeting minimal standards and maximizing current and future opportunities for morphofunctional correlations and molecular studies for both clinical care and research. Whenever feasible, anatomically intact surgical specimens are desirable to enable systematic analysis in selective hippocampectomies, temporal lobe resections, and lesional or nonlesional neocortical samples. Correct orientation of sample and the sample's relation to neurophysiologically aberrant sites requires good communication between pathology and neurosurgical teams. Systematic tissue sampling of 5-mm slabs along a defined anatomic axis and application of a limited immunohistochemical panel will ensure a reliable differential diagnosis of main pathologies encountered in epilepsy surgery. Wiley Periodicals, Inc. © 2016 International League Against Epilepsy.

Complementary IMAC enrichment methods for HLA-associated phosphopeptide identification by mass spectrometry

PubMed Central

Abelin, Jennifer G; Trantham, Paisley D; Penny, Sarah A; Patterson, Andrea M; Ward, Stephen T; Hildebrand, William H; Cobbold, Mark; Bai, Dina L; Shabanowitz, Jeffrey; Hunt, Donald F

2015-01-01

Phosphorylation events within cancer cells often become dysregulated, leading to oncogenic signaling and abnormal cell growth. Phosphopeptides derived from aberrantly phosphorylated proteins that are presented on tumors and not on normal tissues by human leukocyte antigen (HLA) class I molecules are promising candidates for future cancer immunotherapies, because they are tumor specific and have been shown to elicit cytotoxic T cell responses. Robust phosphopeptide enrichments that are suitable for low input amounts must be developed to characterize HLA-associated phosphopeptides from clinical samples that are limited by material availability. We present two complementary mass spectrometry–compatible, iron(III)-immobilized metal affinity chromatography (IMAC) methods that use either nitrilotriacetic acid (NTA) or iminodiacetic acid (IDA) in-house-fabricated columns. We developed these protocols to enrich for subfemtomole-level phosphopeptides from cell line and human tissue samples containing picograms of starting material, which is an order of magnitude less material than what is commonly used. In addition, we added a peptide esterification step to increase phosphopeptide specificity from these low-input samples. To date, hundreds of phosphopeptides displayed on melanoma, ovarian cancer, leukemia and colorectal cancer have been identified using these highly selective phosphopeptide enrichment protocols in combination with a program called ‘CAD Neutral Loss Finder’ that identifies all spectra containing the characteristic neutral loss of phosphoric acid from phosphorylated serine and threonine residues. This methodology enables the identification of HLA-associated phosphopeptides presented by human tissue samples containing as little as nanograms of peptide material in 2 d. PMID:26247297

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

Mineral lung burden of an urban population

NASA Astrophysics Data System (ADS)

Paoletti, L.; Falchi, M.; Batisti, D.; Carrieri, M. P.; Petrelli, M. G.; Ciallella, C.; Donelli, G.

A study was carried out on mineral lung burden in 85 autopsy cases who died accidentally. Subjects of both sexes aged from 15 to 70 years were selected from all the autopsies performed at the Institute of Forensic Medicine in Rome. These subjects were living in an urban area and were not affected by neoplasm diseases. All selected subjects were residing in Rome at the time of their death. Information on years of legal residence in urban areas, smoking habits and occupational history were obtained by interviews with relatives. Lung parenchyma samples were obtained from the right upper lobe. The mineral particulate matter present in the tissue samples was studied by means of analytical transmission electron microscopy (ATEM) techniques: 16 mineral varieties and 22 metallic elements were identified. Smoke, age and residence seem to have influence on the lung burden.

Facile synthesis of titanium (IV) ion immobilized adenosine triphosphate functionalized silica nanoparticles for highly specific enrichment and analysis of intact phosphoproteins.

PubMed

Wang, Hao; Tian, Zhixin

2018-06-06

Analysis of phosphoproteins always faces the challenge of low stoichiometry, which demands highly selective and efficient enrichment in the initial sample preparation. Here we report our synthesis of the novel titanium (IV) ion immobilized adenosine triphosphate functionalized silica nanoparticles (Ti 4+ -ATP-NPs) for efficient enrichment of intact phosphoproteins. The average diameter of Ti 4+ -ATP-NPs was about 128 nm with good dispersibility and the saturated adsorption capacity for β-casein was 1046.5 mg/g. In addition, Ti 4+ -ATP-NPs exhibited high specificity and selectivity in enriching phosphoproteins from both standard protein mixtures and complex biological samples (non-fat milk, chicken egg white and mouse heart tissue extract) as demonstrated by SDS-PAGE. Copyright © 2018 Elsevier B.V. All rights reserved.

Assessment of real-time PCR method for detection of EGFR mutation using both supernatant and cell pellet of malignant pleural effusion samples from non-small-cell lung cancer patients.

PubMed

Shin, Saeam; Kim, Juwon; Kim, Yoonjung; Cho, Sun-Mi; Lee, Kyung-A

2017-10-26

EGFR mutation is an emerging biomarker for treatment selection in non-small-cell lung cancer (NSCLC) patients. However, optimal mutation detection is hindered by complications associated with the biopsy procedure, tumor heterogeneity and limited sensitivity of test methodology. In this study, we evaluated the diagnostic utility of real-time PCR using malignant pleural effusion samples. A total of 77 pleural fluid samples from 77 NSCLC patients were tested using the cobas EGFR mutation test (Roche Molecular Systems). Pleural fluid was centrifuged, and separated cell pellets and supernatants were tested in parallel. Results were compared with Sanger sequencing and/or peptide nucleic acid (PNA)-mediated PCR clamping of matched tumor tissue or pleural fluid samples. All samples showed valid real-time PCR results in one or more DNA samples extracted from cell pellets and supernatants. Compared with other molecular methods, the sensitivity of real-time PCR method was 100%. Concordance rate of real-time PCR and Sanger sequencing plus PNA-mediated PCR clamping was 98.7%. We have confirmed that real-time PCR using pleural fluid had a high concordance rate compared to conventional methods, with no failed samples. Our data demonstrated that the parallel real-time PCR testing using supernatant and cell pellet could offer reliable and robust surrogate strategy when tissue is not available.

High Resolution X-ray Phase Contrast Imaging with Acoustic Tissue-Selective Contrast Enhancement

DTIC Science & Technology

2008-06-01

Imaging with Acoustic Tissue-Selective Contrast Enhancement PRINCIPAL INVESTIGATOR: Gerald J. Diebold, Ph.D. CONTRACTING... Contrast Imaging with Acoustic Tissue-Selective Contrast Enhancement 5b. GRANT NUMBER W81XWH-04-1-0481 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S...additional phase contrast features are visible at the interfaces of soft tissues as slight contrast enhancements . The image sequence in Fig. 2 shows an image

A novel device to stretch multiple tissue samples with variable patterns: application for mRNA regulation in tissue-engineered constructs.

PubMed

Imsirovic, Jasmin; Derricks, Kelsey; Buczek-Thomas, Jo Ann; Rich, Celeste B; Nugent, Matthew A; Suki, Béla

2013-01-01

A broad range of cells are subjected to irregular time varying mechanical stimuli within the body, particularly in the respiratory and circulatory systems. Mechanical stretch is an important factor in determining cell function; however, the effects of variable stretch remain unexplored. In order to investigate the effects of variable stretch, we designed, built and tested a uniaxial stretching device that can stretch three-dimensional tissue constructs while varying the strain amplitude from cycle to cycle. The device is the first to apply variable stretching signals to cells in tissues or three dimensional tissue constructs. Following device validation, we applied 20% uniaxial strain to Gelfoam samples seeded with neonatal rat lung fibroblasts with different levels of variability (0%, 25%, 50% and 75%). RT-PCR was then performed to measure the effects of variable stretch on key molecules involved in cell-matrix interactions including: collagen 1α, lysyl oxidase, α-actin, β1 integrin, β3 integrin, syndecan-4, and vascular endothelial growth factor-A. Adding variability to the stretching signal upregulated, downregulated or had no effect on mRNA production depending on the molecule and the amount of variability. In particular, syndecan-4 showed a statistically significant peak at 25% variability, suggesting that an optimal variability of strain may exist for production of this molecule. We conclude that cycle-by-cycle variability in strain influences the expression of molecules related to cell-matrix interactions and hence may be used to selectively tune the composition of tissue constructs.

Adipose Tissue Responses to Breaking Sitting in Men and Women with Central Adiposity.

PubMed

Chen, Yung-Chih; Betts, James A; Walhin, Jean-Philippe; Thompson, Dylan

2018-04-27

Breaking prolonged sitting reduces postprandial glucose and insulin concentrations and influences skeletal muscle molecular signalling pathways but it is unknown whether breaking sitting also affects adipose tissue. Eleven central overweight participants (7 men and 4 post-menopausal women) aged 50 ± 5 years (means ± SD) completed two mixed-meal feeding trials (PROLONGED SITTING versus BREAKING SITTING) in a randomised, counterbalanced design. The BREAKING SITTING intervention comprised walking for 2 min every 20 min over 5.5 h. Blood samples were taken at regular intervals to examine metabolic biomarkers and adipokine concentrations. Adipose tissue samples were taken at baseline and at 5.5 h to examine changes in mRNA expression and secretion of selected adipokines ex-vivo. Postprandial glycaemia and insulinaemia were attenuated by approximately 50% and 40% in BREAKING SITTING compared to PROLONGED SITTING (iAUC: 359 ± 117 versus 697 ± 218 mmol·330 min·L, p = 0.001 and 202 ± 71 versus 346 ± 150 nmol·330 min·L, p = 0.001, respectively). Despite these pronounced and sustained differences in postprandial glucose and insulin concentrations, adipose tissue mRNA expression for various genes (IL-6, leptin, adiponectin, PDK4, IRS1/2, PI3K and Akt1, etc.) and ex-vivo adipose tissue secretion of IL-6, leptin and adiponectin were not different between trials. This study demonstrates that breaking sitting with short bouts of physical activity has very pronounced effects on systemic postprandial glucose and insulin concentrations but this does not translate into corresponding effects within adipose tissue.

Comparison of gene expression profiles between dental pulp and periodontal ligament tissues in humans

PubMed Central

Gong, Ai-Xiu; Zhang, Jing-Han; Li, Jing; Wu, Jun; Wang, Lin; Miao, Deng-Shun

2017-01-01

There are anatomical and functional differences between human dental pulp (DP) and periodontal ligament (PDL). However, the molecular biological differences and function of these tissues are poorly understood. In the present study, we employed a cDNA microarray array to screen for differentially expressed genes (DEGs) between human DP and PDL tissues, and used the online software WebGestalt to perform the functional analysis of the DEGs. In addition, the STRING database and KEGG pathway analysis were applied for interaction network and pathway analysis of the DEGs. DP and PDL samples were obtained from permanent premolars (n=16) extracted for orthodontic purposes. The results of the microarray assay were confirmed by RT-qPCR. The DEGs were found to be significantly associated with the extracellular matrix and focal adhesion. A total of 10 genes were selected to confirm the results. The mRNA levels of integrin alpha 4 (ITGA4), integrin alpha 8 (ITGA8), neurexin 1 (NRXN1) and contactin 1 (CNTN1) were significantly higher in the DP than in the PDL tissues. However, the levels of collagen type XI alpha 1 (COL11A1), aggrecan (ACAN), collagen type VI alpha 1 (COL6A1), chondroadherin (CHAD), laminin gamma 2 (LAMC2) and laminin alpha 3 (LAMA3) were higher in the PDL than in the DP samples. The gene expression profiles provide novel insight into the characterization of DP and PDL tissues, and contribute to our understanding of the potential molecular mechanisms of dental tissue mineralization and regeneration. PMID:28713908

Tamoxifen Downregulates Ets-oncogene Family Members ETV4 and ETV5 in Benign Breast Tissue: Implications for Durable Risk Reduction

PubMed Central

Euhus, David; Bu, Dawei; Xie, Xian-Jin; Sarode, Venetia; Ashfaq, Raheela; Hunt, Kelly; Xia, Weiya; O’Shaughnessy, Joyce; Grant, Michael; Arun, Banu; Dooley, William; Miller, Alexander; Flockhart, David; Lewis, Cheryl

2011-01-01

Background Five years of tamoxifen reduces breast cancer risk by nearly 50% but is associated with significant side-effects and toxicities. A better understanding of the direct and indirect effects of tamoxifen in benign breast tissue could elucidate new mechanisms of breast carcinogenesis, suggest novel chemoprevention targets, and provide relevant early response biomarkers for Phase II prevention trials. Methods Seventy-three women at increased risk for breast cancer were randomized to tamoxifen (20 mg daily) or placebo for three months. Blood and breast tissue samples were collected at baseline and post-treatment. Sixty-nine women completed all study activities (37 tamoxifen and 32 placebo). The selected biomarkers focused on estradiol and IGFs in the blood, DNA methylation and cytology in random periareolar fine needle aspirates, and tissue morphometry, proliferation, apoptosis, and gene expression (microarray and RT-PCR) in the tissue core samples. Results Tamoxifen downregulated ets-oncogene transcription factor family members ETV4 and ETV5 and reduced breast epithelial cell proliferation independent of CYP2D6 genotypes or effects on estradiol, ESR1 or IGFs. Reduction in proliferation was correlated with downregulation of ETV4 and DNAJC12. Tamoxifen reduced the expression of ETV4- and ETV5-regulated genes implicated in epithelial-stromal interaction and tissue remodeling. Three months of tamoxifen did not affect breast tissue composition, cytological atypia, preneoplasia or apoptosis. Conclusions A plausible mechanism for the chemopreventive effects of tamoxifen is restriction of lobular expansion into stroma through downregulation of ETV4 and ETV5. Multipotential progenitor cap cells of terminal end buds may be the primary target. PMID:21778330

Clinical usefulness of fine needle aspiration cytology in patients less than 20 years old: a 10-year experience at a single institution.

PubMed

Kim, Sunzoo; Jang, Eun Jeong; Jeong, Ji Yun; Park, Ji Young

2013-01-01

The purpose of this study was to identify the spectrum of cytological diagnoses and evaluate the diagnostic effectiveness of fine needle aspiration cytology (FNA) in patients less than 20 years old. The subjects were selected by retrospectively reviewing records from 1999 to 2009. Selected patients less than 20 years old underwent FNA. Cytological and histological slides of samples from the subjects were reviewed. Our study included a total of 909 subjects with a mean age of 14.6 years. The majority of the FNA samples were taken from lymph nodes (n = 448, 49.3%), with the remaining aspirates obtained from the thyroid gland (n = 247, 27.2%), soft tissues of head and neck masses (n = 106, 11.7%), salivary glands (n = 75, 8.3%), breasts (n = 18, 1.9%), skins (n = 9, 1.0%) and soft tissues of extremity (n = 6, 0.7%). The majority (87.6%, n = 796) of the FNA samples were categorized as 'benign', with the remaining designated as 'atypical lesion' (n = 18, 2.0%), 'malignant' (n = 24, 2.6%), or 'inadequate specimen' (n = 71, 7.8%). FNA accuracy was 92% for diagnosing cancer. Specificity and sensitivity were 99% and 63%, respectively. Our study first revealed that FNA has a high specificity for diagnosing cancer in various anatomical locations in young patients and can be confidently used as an effective tool for diagnosing malignancies in young individuals with a clinically suspicious lesion.

Non-specific filtering of beta-distributed data.

PubMed

Wang, Xinhui; Laird, Peter W; Hinoue, Toshinori; Groshen, Susan; Siegmund, Kimberly D

2014-06-19

Non-specific feature selection is a dimension reduction procedure performed prior to cluster analysis of high dimensional molecular data. Not all measured features are expected to show biological variation, so only the most varying are selected for analysis. In DNA methylation studies, DNA methylation is measured as a proportion, bounded between 0 and 1, with variance a function of the mean. Filtering on standard deviation biases the selection of probes to those with mean values near 0.5. We explore the effect this has on clustering, and develop alternate filter methods that utilize a variance stabilizing transformation for Beta distributed data and do not share this bias. We compared results for 11 different non-specific filters on eight Infinium HumanMethylation data sets, selected to span a variety of biological conditions. We found that for data sets having a small fraction of samples showing abnormal methylation of a subset of normally unmethylated CpGs, a characteristic of the CpG island methylator phenotype in cancer, a novel filter statistic that utilized a variance-stabilizing transformation for Beta distributed data outperformed the common filter of using standard deviation of the DNA methylation proportion, or its log-transformed M-value, in its ability to detect the cancer subtype in a cluster analysis. However, the standard deviation filter always performed among the best for distinguishing subgroups of normal tissue. The novel filter and standard deviation filter tended to favour features in different genome contexts; for the same data set, the novel filter always selected more features from CpG island promoters and the standard deviation filter always selected more features from non-CpG island intergenic regions. Interestingly, despite selecting largely non-overlapping sets of features, the two filters did find sample subsets that overlapped for some real data sets. We found two different filter statistics that tended to prioritize features with different characteristics, each performed well for identifying clusters of cancer and non-cancer tissue, and identifying a cancer CpG island hypermethylation phenotype. Since cluster analysis is for discovery, we would suggest trying both filters on any new data sets, evaluating the overlap of features selected and clusters discovered.

Organic Compounds and Trace Elements in Fish Tissue and Bed Sediment in the Delaware River Basin, New Jersey, Pennsylvania, New York, and Delaware, 1998-2000

USGS Publications Warehouse

Romanok, Kristin M.; Fischer, Jeffrey M.; Riva-Murray, Karen; Brightbill, Robin; Bilger, Michael

2006-01-01

As part of the National Water-Quality Assessment (NAWQA) program activities in the Delaware River Basin (DELR), samples of fish tissue from 21 sites and samples of bed sediment from 35 sites were analyzed for a suite of organic compounds and trace elements. The sampling sites, within subbasins ranging in size from 11 to 600 square miles, were selected to represent 5 main land-use categories in the DELR -forest, low-agricultural, agricultural, urban, and mixed use. Samples of both fish tissue and bed sediment were also collected from 4 'large-river' sites that represented drainage areas ranging from 1,300 to 6,800 square miles, areas in which the land is used for a variety of purposes. One or more of the organochlorine compounds-DDT and chlordane metabolites, polychlorinated biphenyls (total PCBs), and dieldrin- were detected frequently in samples collected over a wide geographic area. One or more of these compounds were detected in fish-tissue samples from 92 percent of the sites and in bed-sediment samples from 82 percent of the sites. Concentrations of total DDT, total chlordanes, total PCBs, and dieldrin in whole white suckers and in bed sediment were significantly related to urban/industrial basin characteristics, such as percentage of urban land use and population density. Semi-volatile organic compounds (SVOCs)-total polycyclic aromatic hydrocarbons (PAHs), total phthalates, and phenols- were detected frequently in bed-sediment samples. All three types of SVOCs were detected in samples from at least one site in each land-use category. The highest detection rates and concentrations typically were in samples from sites in the urban and mixed land-use categories, as well as from the large-river sites. Concentrations of total PAHs and total phthalates in bed-sediment samples were found to be statistically related to percentages of urban land use and to population density in the drainage areas represented by the sampling sites. The samples of fish tissue and bed sediment collected throughout the DELR were analyzed for a large suite of trace elements, but results of the analyses for eight elements-arsenic, cadmium, chromium, copper, lead, nickel, mercury, and zinc- that are considered contaminants of concern are described in this report. One or more of the eight trace elements were detected in samples from every fish tissue and bed-sediment sampling site, and all of the trace elements were detected in samples from 97 percent of the bed-sediment sites. The concentrations of organic compounds and trace elements in the DELR samples were compared to applicable guidelines for the protection of wildlife and other biological organisms. Concentrations of total DDT, total chlordanes, total PCBs, and dieldrin in fish-tissue samples from 14 sites exceeded one or more of the Wildlife Protective Guidelines established by the New York State Department of Environmental Conservation. Concentrations of one or more organic compounds in samples from 16 bed-sediment sites exceeded the Threshold Effects Concentrations (TEC) of the Canadian Sediment Quality Guidelines, and concentrations of one or more of the eight trace elements in samples from 38 bed-sediment sites exceeded the TEC. (The TEC is the concentration below which adverse biological effects in freshwater ecosystems are expected to be rare.) Concentrations of organic compounds in samples from some bed-sediment sites exceeded the Canadian Probable Effects Concentrations (PEC), and concentrations of trace elements in samples from 18 sites exceeded the PEC. (The PEC is the concentration above which adverse effects to biological organisms are expected to occur frequently). Concentrations of organic compounds and trace elements in samples from the DELR were compared to similar data from other NAWQA study units in the northeastern United States and also data from the Mobile River (Alabama) Basin and the Northern Rockies Intermontane Basin study units. Median concentrations of to

A Review of Mid-Infrared and Near-Infrared Imaging: Principles, Concepts and Applications in Plant Tissue Analysis.

PubMed

Türker-Kaya, Sevgi; Huck, Christian W

2017-01-20

Plant cells, tissues and organs are composed of various biomolecules arranged as structurally diverse units, which represent heterogeneity at microscopic levels. Molecular knowledge about those constituents with their localization in such complexity is very crucial for both basic and applied plant sciences. In this context, infrared imaging techniques have advantages over conventional methods to investigate heterogeneous plant structures in providing quantitative and qualitative analyses with spatial distribution of the components. Thus, particularly, with the use of proper analytical approaches and sampling methods, these technologies offer significant information for the studies on plant classification, physiology, ecology, genetics, pathology and other related disciplines. This review aims to present a general perspective about near-infrared and mid-infrared imaging/microspectroscopy in plant research. It is addressed to compare potentialities of these methodologies with their advantages and limitations. With regard to the organization of the document, the first section will introduce the respective underlying principles followed by instrumentation, sampling techniques, sample preparations, measurement, and an overview of spectral pre-processing and multivariate analysis. The last section will review selected applications in the literature.

Monitoring Dopamine ex Vivo during Electrical Stimulation Using Liquid-Microjunction Surface Sampling.

PubMed

Gill, Emily L; Marks, Megan; Yost, Richard A; Vedam-Mai, Vinata; Garrett, Timothy J

2017-12-19

Liquid-microjunction surface sampling (LMJ-SS) is an ambient ionization technique based on the continuous flow of solvent using an in situ microextraction device in which solvent moves through the probe, drawing in the analytes in preparation for ionization using an electrospray ionization source. However, unlike traditional mass spectrometry (MS) techniques, it operates under ambient pressure and requires no sample preparation, thereby making it ideal for rapid sampling of thicker tissue sections for electrophysiological and other neuroscientific research studies. Studies interrogating neural synapses, or a specific neural circuit, typically employ thick, ex vivo tissue sections maintained under near-physiological conditions to preserve tissue viability and maintain the neural networks. Deep brain stimulation (DBS) is a surgical procedure used to treat the neurological symptoms that are associated with certain neurodegenerative and neuropsychiatric diseases. Parkinson's disease (PD) is a neurological disorder which is commonly treated with DBS therapy. PD is characterized by the degeneration of dopaminergic neurons in the substantia nigra pars compacta portion of the brain. Here, we demonstrate that the LMJ-SS methodology can provide a platform for ex vivo analysis of the brain during electrical stimulation, such as DBS. We employ LMJ-SS in the ex vivo analysis of mouse brain tissue for monitoring dopamine during electrical stimulation of the striatum region. The mouse brain tissue was sectioned fresh post sacrifice and maintained in artificial cerebrospinal fluid to create near-physiological conditions before direct sampling using LMJ-SS. A selection of metabolites, including time-sensitive metabolites involved in energy regulation in the brain, were identified using standards, and the mass spectral database mzCloud was used to assess the feasibility of the methodology. Thereafter, the intensity of m/z 154 corresponding to protonated dopamine was monitored before and after electrical stimulation of the striatum region, showing an increase in signal directly following a stimulation event. Dopamine is the key neurotransmitter implicated in PD, and although electrochemical detectors have shown such increases in dopamine post-DBS, this is the first study to do so using MS methodologies.

Selection and Validation of Appropriate Reference Genes for qRT-PCR Analysis in Isatis indigotica Fort.

PubMed Central

Li, Tao; Wang, Jing; Lu, Miao; Zhang, Tianyi; Qu, Xinyun; Wang, Zhezhi

2017-01-01

Due to its sensitivity and specificity, real-time quantitative PCR (qRT-PCR) is a popular technique for investigating gene expression levels in plants. Based on the Minimum Information for Publication of Real-Time Quantitative PCR Experiments (MIQE) guidelines, it is necessary to select and validate putative appropriate reference genes for qRT-PCR normalization. In the current study, three algorithms, geNorm, NormFinder, and BestKeeper, were applied to assess the expression stability of 10 candidate reference genes across five different tissues and three different abiotic stresses in Isatis indigotica Fort. Additionally, the IiYUC6 gene associated with IAA biosynthesis was applied to validate the candidate reference genes. The analysis results of the geNorm, NormFinder, and BestKeeper algorithms indicated certain differences for the different sample sets and different experiment conditions. Considering all of the algorithms, PP2A-4 and TUB4 were recommended as the most stable reference genes for total and different tissue samples, respectively. Moreover, RPL15 and PP2A-4 were considered to be the most suitable reference genes for abiotic stress treatments. The obtained experimental results might contribute to improved accuracy and credibility for the expression levels of target genes by qRT-PCR normalization in I. indigotica. PMID:28702046

A sensitive and selective immunoaffinity column clean up coupled to UPLC-MS/MS for determination of trace methyl-3-quinoxaline-2-carboxylic acid in animal tissues.

PubMed

Li, Peipei; Zhang, Xiaojun; Zhang, Juanmei; Yan, Zhongyong; Zhang, Shuai; Chen, Si; Fang, Yi

2018-02-01

This paper described a reliable and simple method for the selective determination of MQCA in animal tissues using ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). A highly targeted immunoaffinity column was used for sample purification after enzymatic hydrolysis. The purified extracts were analyzed by reversed-phase HPLC-MS/MS in positive ESI and multiple reaction monitoring mode. The calibration curves showed good linearity with correlation coefficient (r 2 ) larger than 0.995. The average recoveries at the spiked levels of 0.5, 2.0 and 20μgkg -1 were 90.2% to 103.5% with intra-day and inter-day relatives standard deviations (RSD, n=6) ranging from 1.8% to 6.7% and 3.5% to 7.6% respectively. The limit of quantification (LOQ) was 0.5μgkg -1 , which can fulfil the maximum residue level (MRL) of 4.0μgkg -1 stipulated by the Agricultural Minister of China and the requirement of the confirmatory criteria according to the European Commission Decision 2002/657/EC. The method is sensitive, accurate, convenient and rapid, and has been successfully applied in real samples. Copyright © 2018 Elsevier B.V. All rights reserved.

Efficacy of Exclusive Lingual Nerve Block versus Conventional Inferior Alveolar Nerve Block in Achieving Lingual Soft-tissue Anesthesia.

PubMed

Balasubramanian, Sasikala; Paneerselvam, Elavenil; Guruprasad, T; Pathumai, M; Abraham, Simin; Krishnakumar Raja, V B

2017-01-01

The aim of this randomized clinical trial was to assess the efficacy of exclusive lingual nerve block (LNB) in achieving selective lingual soft-tissue anesthesia in comparison with conventional inferior alveolar nerve block (IANB). A total of 200 patients indicated for the extraction of lower premolars were recruited for the study. The samples were allocated by randomization into control and study groups. Lingual soft-tissue anesthesia was achieved by IANB and exclusive LNB in the control and study group, respectively. The primary outcome variable studied was anesthesia of ipsilateral lingual mucoperiosteum, floor of mouth and tongue. The secondary variables assessed were (1) taste sensation immediately following administration of local anesthesia and (2) mouth opening and lingual nerve paresthesia on the first postoperative day. Data analysis for descriptive and inferential statistics was performed using SPSS (IBM SPSS Statistics for Windows, Version 22.0, Armonk, NY: IBM Corp. Released 2013) and a P < 0.05 was considered statistically significant. In comparison with the control group, the study group (LNB) showed statistically significant anesthesia of the lingual gingiva of incisors, molars, anterior floor of the mouth, and anterior tongue. Exclusive LNB is superior to IAN nerve block in achieving selective anesthesia of lingual soft tissues. It is technically simple and associated with minimal complications as compared to IAN block.

Efficacy of Exclusive Lingual Nerve Block versus Conventional Inferior Alveolar Nerve Block in Achieving Lingual Soft-tissue Anesthesia

PubMed Central

Balasubramanian, Sasikala; Paneerselvam, Elavenil; Guruprasad, T; Pathumai, M; Abraham, Simin; Krishnakumar Raja, V. B.

2017-01-01

Objective: The aim of this randomized clinical trial was to assess the efficacy of exclusive lingual nerve block (LNB) in achieving selective lingual soft-tissue anesthesia in comparison with conventional inferior alveolar nerve block (IANB). Materials and Methods: A total of 200 patients indicated for the extraction of lower premolars were recruited for the study. The samples were allocated by randomization into control and study groups. Lingual soft-tissue anesthesia was achieved by IANB and exclusive LNB in the control and study group, respectively. The primary outcome variable studied was anesthesia of ipsilateral lingual mucoperiosteum, floor of mouth and tongue. The secondary variables assessed were (1) taste sensation immediately following administration of local anesthesia and (2) mouth opening and lingual nerve paresthesia on the first postoperative day. Results: Data analysis for descriptive and inferential statistics was performed using SPSS (IBM SPSS Statistics for Windows, Version 22.0, Armonk, NY: IBM Corp. Released 2013) and a P < 0.05 was considered statistically significant. In comparison with the control group, the study group (LNB) showed statistically significant anesthesia of the lingual gingiva of incisors, molars, anterior floor of the mouth, and anterior tongue. Conclusion: Exclusive LNB is superior to IAN nerve block in achieving selective anesthesia of lingual soft tissues. It is technically simple and associated with minimal complications as compared to IAN block. PMID:29264294

Crystal violet stain as a selective stain for the assessment of mitotic figures in oral epithelial dysplasia and oral squamous cell carcinoma.

PubMed

Jadhav, Kiran B; Ahmed Mujib, B R; Gupta, Nidhi

2012-01-01

Assessment of mitotic figures (MFs) is routinely practiced as prognostic indicator in oral epithelial dysplasia (OED) and oral squamous cell carcinoma (OSCC), but identification of MFs poses a problem in terms of staining characteristics. To evaluate effectiveness of crystal violet stain for staining of MFs and its comparison with hematoxylin and eosin (H and E) stain. Study sample includes archival tissues embedded in paraffin blocks diagnosed as OED (n = 30) and OSCC (n = 30). The control group comprised of tissue specimen from oral mucosa of healthy volunteers (n = 30). Two serial sections of each tissue specimen were stained separately with H and E stain and 1% crystal violet stain. The stained sections were observed under microscope for identification and counting of MFs. Data obtained was statistically analyzed by using the Man-Whitney U test. A significant increase in number of MFs was observed in OED and OSCC in comparison with normal oral mucosa. There was a highly significant increase in number of MFs in crystal violet stained tissue sections when compared with H and E stain. Metaphase is the most commonly observed phase of mitosis in crystal violet stain when compared with H and E stain for all three groups. Crystal violet stain can be considered as selective stain for mitotic figures.

Serum Autoantibodies in Chronic Prostate Inflammation in Prostate Cancer Patients.

PubMed

Schlick, Bettina; Massoner, Petra; Lueking, Angelika; Charoentong, Pornpimol; Blattner, Mirjam; Schaefer, Georg; Marquart, Klaus; Theek, Carmen; Amersdorfer, Peter; Zielinski, Dirk; Kirchner, Matthias; Trajanoski, Zlatko; Rubin, Mark A; Müllner, Stefan; Schulz-Knappe, Peter; Klocker, Helmut

2016-01-01

Chronic inflammation is frequently observed on histological analysis of malignant and non-malignant prostate specimens. It is a suspected supporting factor for prostate diseases and their progression and a main cause of false positive PSA tests in cancer screening. We hypothesized that inflammation induces autoantibodies, which may be useful biomarkers. We aimed to identify and validate prostate inflammation associated serum autoantibodies in prostate cancer patients and evaluate the expression of corresponding autoantigens. Radical prostatectomy specimens of prostate cancer patients (N = 70) were classified into high and low inflammation groups according to the amount of tissue infiltrating lymphocytes. The corresponding pre-surgery blood serum samples were scrutinized for autoantibodies using a low-density protein array. Selected autoantigens were identified in prostate tissue and their expression pattern analyzed by immunohistochemistry and qPCR. The identified autoantibody profile was cross-checked in an independent sample set (N = 63) using the Luminex-bead protein array technology. Protein array screening identified 165 autoantibodies differentially abundant in the serum of high compared to low inflammation patients. The expression pattern of three corresponding antigens were established in benign and cancer tissue by immunohistochemistry and qPCR: SPAST (Spastin), STX18 (Syntaxin 18) and SPOP (speckle-type POZ protein). Of these, SPAST was significantly increased in prostate tissue with high inflammation. All three autoantigens were differentially expressed in primary and/or castration resistant prostate tumors when analyzed in an inflammation-independent tissue microarray. Cross-validation of the inflammation autoantibody profile on an independent sample set using a Luminex-bead protein array, retrieved 51 of the significantly discriminating autoantibodies. Three autoantibodies were significantly upregulated in both screens, MUT, RAB11B and CSRP2 (p>0.05), two, SPOP and ZNF671, close to statistical significance (p = 0.051 and 0.076). We provide evidence of an inflammation-specific autoantibody profile and confirm the expression of corresponding autoantigens in prostate tissue. This supports evaluation of autoantibodies as non-invasive markers for prostate inflammation.

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.

Pharmacokinetic, bioavailability and tissue distribution study of MP3950, a new gastroprokinetic candidate compound, in rat using UPLC-MS/MS.

PubMed

Zhao, Yu; Zhao, Min; Jiang, Qi; Qin, Feng; Wang, Chengying; Xiong, Zhili; Wang, Shaojie; He, Zhonggui; Guo, Xingjie; Zhao, Longshan

2018-06-02

MP3950 is being developed as a gastroprokinetic candidate compound. To illustrate the pharmacokinetic profiles, absolute bioavailability after intravenous administration and oral administration with MP3950 as well as tissue distribution in vivo, an UPLC-MS/MS approach which was rapid and selective was developed to determine MP3950 in plasma and tissue of rat. Sample pre-treatment of plasma sample was one-step protein precipitation. 0.1% formic acid containing 5 mmol/L ammonium acetate-methanol(55/45,v/v) was used for isocratic elution on a Waters ACQUITY UPLC® BEH C18 (50 mm × 2.1 mm, 1.7 μm) to achieve the separation. The analysis was performed in MRM mode via positive ESI mode. LLOQ of the method was 10 ng/mL, and the linearity up to 10,000 ng/mL. The intra-day precision (relative standard deviation, RSD) was 4.0-9.0% and the inter-day precision was 4.2-10.6%. The accuracy (relative error, RE) was -1.2-2.4%. Tissue samples were collected from the brain, heart, liver, spleen, lung, kidney, stomach, duodenum, small intestine, large intestine, appendix and skeletal muscle. The same liquid chromatographic and mass spectrometric conditions were used, and it's proven that this method was feasible to analyze the MP3950 in tissues with good precision and accuracy over the range from 10 to 5000 ng·mL -1 . It was found that the concentration of MP3950 is higher in digestive system. The tissue distribution, pharmacokinetic and bioavailability of MP3950 in rats were carried out by the method for the first time, which can provide enough information for the further development and investigation of MP3950. Copyright © 2018. Published by Elsevier B.V.

Serum Autoantibodies in Chronic Prostate Inflammation in Prostate Cancer Patients

PubMed Central

Schlick, Bettina; Massoner, Petra; Lueking, Angelika; Charoentong, Pornpimol; Blattner, Mirjam; Schaefer, Georg; Marquart, Klaus; Theek, Carmen; Amersdorfer, Peter; Zielinski, Dirk; Kirchner, Matthias; Trajanoski, Zlatko; Rubin, Mark A.; Müllner, Stefan; Schulz-Knappe, Peter; Klocker, Helmut

2016-01-01

Background Chronic inflammation is frequently observed on histological analysis of malignant and non-malignant prostate specimens. It is a suspected supporting factor for prostate diseases and their progression and a main cause of false positive PSA tests in cancer screening. We hypothesized that inflammation induces autoantibodies, which may be useful biomarkers. We aimed to identify and validate prostate inflammation associated serum autoantibodies in prostate cancer patients and evaluate the expression of corresponding autoantigens. Methods Radical prostatectomy specimens of prostate cancer patients (N = 70) were classified into high and low inflammation groups according to the amount of tissue infiltrating lymphocytes. The corresponding pre-surgery blood serum samples were scrutinized for autoantibodies using a low-density protein array. Selected autoantigens were identified in prostate tissue and their expression pattern analyzed by immunohistochemistry and qPCR. The identified autoantibody profile was cross-checked in an independent sample set (N = 63) using the Luminex-bead protein array technology. Results Protein array screening identified 165 autoantibodies differentially abundant in the serum of high compared to low inflammation patients. The expression pattern of three corresponding antigens were established in benign and cancer tissue by immunohistochemistry and qPCR: SPAST (Spastin), STX18 (Syntaxin 18) and SPOP (speckle-type POZ protein). Of these, SPAST was significantly increased in prostate tissue with high inflammation. All three autoantigens were differentially expressed in primary and/or castration resistant prostate tumors when analyzed in an inflammation-independent tissue microarray. Cross-validation of the inflammation autoantibody profile on an independent sample set using a Luminex-bead protein array, retrieved 51 of the significantly discriminating autoantibodies. Three autoantibodies were significantly upregulated in both screens, MUT, RAB11B and CSRP2 (p>0.05), two, SPOP and ZNF671, close to statistical significance (p = 0.051 and 0.076). Conclusions We provide evidence of an inflammation-specific autoantibody profile and confirm the expression of corresponding autoantigens in prostate tissue. This supports evaluation of autoantibodies as non-invasive markers for prostate inflammation. PMID:26863016

Grouped fuzzy SVM with EM-based partition of sample space for clustered microcalcification detection.

PubMed

Wang, Huiya; Feng, Jun; Wang, Hongyu

2017-07-20

Detection of clustered microcalcification (MC) from mammograms plays essential roles in computer-aided diagnosis for early stage breast cancer. To tackle problems associated with the diversity of data structures of MC lesions and the variability of normal breast tissues, multi-pattern sample space learning is required. In this paper, a novel grouped fuzzy Support Vector Machine (SVM) algorithm with sample space partition based on Expectation-Maximization (EM) (called G-FSVM) is proposed for clustered MC detection. The diversified pattern of training data is partitioned into several groups based on EM algorithm. Then a series of fuzzy SVM are integrated for classification with each group of samples from the MC lesions and normal breast tissues. From DDSM database, a total of 1,064 suspicious regions are selected from 239 mammography, and the measurement of Accuracy, True Positive Rate (TPR), False Positive Rate (FPR) and EVL = TPR* 1-FPR are 0.82, 0.78, 0.14 and 0.72, respectively. The proposed method incorporates the merits of fuzzy SVM and multi-pattern sample space learning, decomposing the MC detection problem into serial simple two-class classification. Experimental results from synthetic data and DDSM database demonstrate that our integrated classification framework reduces the false positive rate significantly while maintaining the true positive rate.

TOFSIMS-P: a web-based platform for analysis of large-scale TOF-SIMS data.

PubMed

Yun, So Jeong; Park, Ji-Won; Choi, Il Ju; Kang, Byeongsoo; Kim, Hark Kyun; Moon, Dae Won; Lee, Tae Geol; Hwang, Daehee

2011-12-15

Time-of-flight secondary ion mass spectrometry (TOF-SIMS) has been a useful tool to profile secondary ions from the near surface region of specimens with its high molecular specificity and submicrometer spatial resolution. However, the TOF-SIMS analysis of even a moderately large size of samples has been hampered due to the lack of tools for automatically analyzing the huge amount of TOF-SIMS data. Here, we present a computational platform to automatically identify and align peaks, find discriminatory ions, build a classifier, and construct networks describing differential metabolic pathways. To demonstrate the utility of the platform, we analyzed 43 data sets generated from seven gastric cancer and eight normal tissues using TOF-SIMS. A total of 87 138 ions were detected from the 43 data sets by TOF-SIMS. We selected and then aligned 1286 ions. Among them, we found the 66 ions discriminating gastric cancer tissues from normal ones. Using these 66 ions, we then built a partial least square-discriminant analysis (PLS-DA) model resulting in a misclassification error rate of 0.024. Finally, network analysis of the 66 ions showed disregulation of amino acid metabolism in the gastric cancer tissues. The results show that the proposed framework was effective in analyzing TOF-SIMS data from a moderately large size of samples, resulting in discrimination of gastric cancer tissues from normal tissues and identification of biomarker candidates associated with the amino acid metabolism.

Near-infrared image-guided laser ablation of artificial caries lesions.

PubMed

Tao, You-Chen; Fan, Kenneth; Fried, Daniel

2007-01-01

Laser removal of dental hard tissue can be combined with optical, spectral or acoustic feedback systems to selectively ablate dental caries and restorative materials. Near-infrared (NIR) imaging has considerable potential for the optical discrimination of sound and demineralized tissue. The objective of this study was to test the hypothesis that two-dimensional NIR images of demineralized tooth surfaces can be used to guide CO(2) laser ablation for the selective removal of artificial caries lesions. Highly patterned artificial lesions were produced by submerging 5 × 5 mm(2) bovine enamel samples in demineralized solution for a 9-day period while sound areas were protected with acid resistant varnish. NIR imaging and polarization sensitive optical coherence tomography (PS-OCT) were used to acquire depth-resolved images at a wavelength of 1310-nm. An imaging processing module was developed to analyze the NIR images and to generate optical maps. The optical maps were used to control a CO(2) laser for the selective removal of the lesions at a uniform depth. This experiment showed that the patterned artificial lesions were removed selectively using the optical maps with minimal damage to sound enamel areas. Post-ablation NIR and PS-OCT imaging confirmed that demineralized areas were removed while sound enamel was conserved. This study successfully demonstrated that near-IR imaging can be integrated with a CO(2) laser ablation system for the selective removal of dental caries.

Near-infrared image-guided laser ablation of artificial caries lesions

PubMed Central

Tao, You-Chen; Fan, Kenneth; Fried, Daniel

2012-01-01

Laser removal of dental hard tissue can be combined with optical, spectral or acoustic feedback systems to selectively ablate dental caries and restorative materials. Near-infrared (NIR) imaging has considerable potential for the optical discrimination of sound and demineralized tissue. The objective of this study was to test the hypothesis that two–dimensional NIR images of demineralized tooth surfaces can be used to guide CO2 laser ablation for the selective removal of artificial caries lesions. Highly patterned artificial lesions were produced by submerging 5 × 5 mm2 bovine enamel samples in demineralized solution for a 9-day period while sound areas were protected with acid resistant varnish. NIR imaging and polarization sensitive optical coherence tomography (PS-OCT) were used to acquire depth-resolved images at a wavelength of 1310-nm. An imaging processing module was developed to analyze the NIR images and to generate optical maps. The optical maps were used to control a CO2 laser for the selective removal of the lesions at a uniform depth. This experiment showed that the patterned artificial lesions were removed selectively using the optical maps with minimal damage to sound enamel areas. Post-ablation NIR and PS-OCT imaging confirmed that demineralized areas were removed while sound enamel was conserved. This study successfully demonstrated that near-IR imaging can be integrated with a CO2 laser ablation system for the selective removal of dental caries. PMID:22866210

Near-infrared image-guided laser ablation of artificial caries lesions

NASA Astrophysics Data System (ADS)

Tao, You-Chen; Fan, Kenneth; Fried, Daniel

2007-02-01

Laser removal of dental hard tissue can be combined with optical, spectral or acoustic feedback systems to selectively ablate dental caries and restorative materials. Near-infrared (NIR) imaging has considerable potential for the optical discrimination of sound and demineralized tissue. The objective of this study was to test the hypothesis that two-dimensional NIR images of demineralized tooth surfaces can be used to guide CO II laser ablation for the selective removal of artificial caries lesions. Highly patterned artificial lesions were produced by submerging 5 x 5 mm2 bovine enamel samples in demineralized solution for a 9-day period while sound areas were protected with acid resistant varnish. NIR imaging and polarization sensitive optical coherence tomography (PS-OCT) were used to acquire depth-resolved images at a wavelength of 1310-nm. An imaging processing module was developed to analyze the NIR images and to generate optical maps. The optical maps were used to control a CO II laser for the selective removal of the lesions at a uniform depth. This experiment showed that the patterned artificial lesions were removed selectively using the optical maps with minimal damage to sound enamel areas. Post-ablation NIR and PS-OCT imaging confirmed that demineralized areas were removed while sound enamel was conserved. This study successfully demonstrated that near-IR imaging can be integrated with a CO II laser ablation system for the selective removal of dental caries.

Histopathology Image Analysis in Two Long-Term Animal Experiments with Helical Flow Total Artificial Heart.

PubMed

Wotke, Jiri; Homolka, Pavel; Vasku, Jaromír; Dobsak, Petr; Palanova, Petra; Mrkvicova, Veronika; Konecny, Petr; Soska, Vladimir; Pohanka, Michal; Novakova, Marie; Yurimoto, Terumi; Saito, Itsuro; Inoue, Yusuke; Isoyama, Takashi; Abe, Yusuke

2016-12-01

Histopathological analysis can provide important information in long-term experiments with total artificial heart (TAH). Recently, a new type of blood pump, the helical flow total artificial heart (HF-TAH) was developed. This study aimed to investigate the changes in selected vital organs in animal experiments with implanted HF-TAH. Samples from lung, liver, and kidneys from two female goats (No. 1301 and No. 1304) with implanted HF-TAH were analyzed. Tissue samples were fixed in 10% formaldehyde and 4 µm thick transverse sections were stained with hematoxylin-eosin (HE). Additional staining was done for detection of connective tissue (Masson-Goldner stain) and for detection of iron (hemosiderin) deposits (Perls stain). Sections were scanned at 100× and 500× magnification with a light microscope. Experiment no. 1301 survived 100 days (cause of termination was heavy damage of the right pump); experimental goat no.1304 survived 68 days and was sacrificed due to severe right hydrodynamic bearing malfunction. Histopathological analysis of liver samples proved signs of chronic venostasis with limited focal necrotic zones. Dilated tubules, proteinaceous material in tubular lumen, and hemosiderin deposits were detected in kidney samples. Contamination of the organs by embolized micro-particles was suspected at the autopsy after discovery of visible damage (scratches) of the pump impeller surface (made from titanium alloy) in both experiments. Sporadic deposits of foreign micro-particles (presumably titanium) were observed in most of the analyzed parenchymal organs. However, the described deposits were not in direct connection with inflammatory reactions in the analyzed tissues. Histopathological analysis showed the presence of minimal contamination of the lung, kidney, and liver tissue samples by foreign material (titanium very likely). The analysis showed only limited pathological changes, especially in liver and kidneys, which might be attributed to the influence of artificial perfusion often observed in chronic TAH experiments. © 2016 Wiley Periodicals, Inc. and International Center for Artificial Organs and Transplantation.

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.

Impact of Ischemia and Procurement Conditions on Gene Expression in Renal Cell Carcinoma

PubMed Central

Liu, Nick W.; Sanford, Thomas; Srinivasan, Ramaprasad; Liu, Jack L.; Khurana, Kiranpreet; Aprelikova, Olga; Valero, Vladimir; Bechert, Charles; Worrell, Robert; Pinto, Peter A.; Yang, Youfeng; Merino, Maria; Linehan, W. Marston; Bratslavsky, Gennady

2013-01-01

Purpose Previous studies have shown that ischemia alters gene expression in normal and malignant tissues. There are no studies that evaluated effects of ischemia in renal tumors. This study examines the impact of ischemia and tissue procurement conditions on RNA integrity and gene expression in renal cell carcinoma. Experimental Design Ten renal tumors were resected without renal hilar clamping from 10 patients with renal clear cell carcinoma. Immediately after tumor resection, a piece of tumor was snap frozen. Remaining tumor samples were stored at 4C, 22C and 37C and frozen at 5, 30, 60, 120, and 240 minutes. Histopathologic evaluation was performed on all tissue samples, and only those with greater than 80% tumor were selected for further analysis. RNA integrity was confirmed by electropherograms and quantitated using RIN index. Altered gene expression was assessed by paired, two-sample t-test between the zero time point and aliquots from various conditions obtained from the same tumor. Results One hundred and forty microarrays were performed. Some RNA degradation was observed 240 mins after resection at 37C. The expression of over 4,000 genes was significantly altered by ischemia times or storage conditions. The greatest gene expression changes were observed with longer ischemia time and warmer tissue procurement conditions. Conclusion RNA from kidney cancer remains intact for up to 4 hours post surgical resection regardless of storage conditions. Despite excellent RNA preservation, time after resection and procurement conditions significantly influence gene expression profiles. Meticulous attention to pre-acquisition variables is of paramount importance for accurate tumor profiling. PMID:23136194

Development of an improved sample preparation platform for acidic endogenous hormones in plant tissues using electromembrane extraction.

PubMed

Suh, Joon Hyuk; Han, Sang Beom; Wang, Yu

2018-02-02

Despite their importance in pivotal signaling pathways due to trace quantities and complex matrices, the analysis of plant hormones is a challenge. Here, to improve this issue, we present an electromembrane extraction technology combined with liquid chromatography-tandem mass spectrometry for determination of acidic plant hormones including jasmonic acid, abscisic acid, salicylic acid, benzoic acid, gibberellic acid and gibberellin A 4 in plant tissues. Factors influencing extraction efficiency, such as voltage, extraction time and stirring rate were optimized using a design of experiments. Analytical performance was evaluated in terms of specificity, linearity, limit of quantification, precision, accuracy, recovery and repeatability. The results showed good linearity (r 2  > 0.995), precision and acceptable accuracy. The limit of quantification ranged from 0.1 to 10 ng mL -1 , and the recoveries were 34.6-50.3%. The developed method was applied in citrus leaf samples, showing better clean-up efficiency, as well as higher sensitivity compared to a previous method using liquid-liquid extraction. Organic solvent consumption was minimized during the process, making it an appealing method. More noteworthy, electromembrane extraction has been scarcely applied to plant tissues, and this is the first time that major plant hormones were extracted using this technology, with high sensitivity and selectivity. Taken together, this work gives not only a novel sample preparation platform using an electric field for plant hormones, but also a good example of extracting complex plant tissues in a simple and effective way. Copyright © 2017 Elsevier B.V. All rights reserved.

Select tissue mineral concentrations and chronic wasting disease status in mule deer from North-central Colorado.

PubMed

Wolfe, Lisa L; Conner, Mary M; Bedwell, Cathy L; Lukacs, Paul M; Miller, Michael W

2010-07-01

Trace mineral imbalances have been suggested as having a causative or contributory role in chronic wasting disease (CWD), a prion disease of several North American cervid species. To begin exploring relationships between tissue mineral concentrations and CWD in natural systems, we measured liver tissue concentrations of copper, manganese, and molybdenum in samples from 447 apparently healthy, adult (> or = 2 yr old) mule deer (Odocoileus hemionus) culled or vehicle killed from free-ranging populations in north-central Colorado, United States, where CWD occurs naturally; we also measured copper concentrations in brain-stem (medulla oblongata at the obex) tissue from 181 of these deer. Analyses revealed a wide range of concentrations of all three minerals among sampled deer (copper: 5.6-331 ppm in liver, 1.5-31.9 ppm in obex; manganese: 0.1-21.4 ppm in liver; molybdenum: 0.5-4.0 ppm in liver). Bayesian multiple regression analysis revealed a negative association between obex copper (-0.097; 95% credible interval -0.192 to -0.006) and the probability of sampled deer also being infected with CWD, as well as a positive association between liver manganese (0.158; 95% credible interval 0.066 to 0.253) and probability of infection. We could not discern whether the tendencies toward lower brain-stem copper concentrations or higher systemic manganese concentrations in infected deer preceded prion infection or rather were the result of infection and its subsequent effects, although the distribution of trace mineral concentrations in infected deer seemed more suggestive of the latter.

Full-genome dengue virus sequencing in mosquito saliva shows lack of convergent positive selection during transmission by Aedes aegypti

PubMed Central

Cao-Lormeau, Van-Mai; Lambrechts, Louis

2017-01-01

Abstract Like other pathogens with high mutation and replication rates, within-host dengue virus (DENV) populations evolve during infection of their main mosquito vector, Aedes aegypti. Within-host DENV evolution during transmission provides opportunities for adaptation and emergence of novel virus variants. Recent studies of DENV genetic diversity failed to detect convergent evolution of adaptive mutations in mosquito tissues such as midgut and salivary glands, suggesting that convergent positive selection is not a major driver of within-host DENV evolution in the vector. However, it is unknown whether this conclusion extends to the transmitted viral subpopulation because it is technically difficult to sequence DENV genomes in mosquito saliva. Here, we achieved DENV full-genome sequencing by pooling saliva samples collected non-sacrificially from 49 to 163 individual Ae. aegypti mosquitoes previously infected with one of two DENV-1 genotypes. We compared the transmitted viral subpopulations found in the pooled saliva samples collected in time series with the input viral population present in the infectious blood meal. In all pooled saliva samples examined, the full-genome consensus sequence of the input viral population was unchanged. Although the pooling strategy prevents analysis of individual saliva samples, our results demonstrate the lack of strong convergent positive selection during a single round of DENV transmission by Ae. aegypti. This finding reinforces the idea that genetic drift and purifying selection are the dominant evolutionary forces shaping within-host DENV genetic diversity during transmission by mosquitoes. PMID:29497564

Heterogeneous histopathology of cortical microbleeds in cerebral amyloid angiopathy.

PubMed

van Veluw, Susanne J; Biessels, Geert Jan; Klijn, Catharina J M; Rozemuller, Annemieke J M

2016-03-01

To investigate the histopathologic substrate of microbleeds detected on 7T postmortem MRI in autopsy cases with severe cerebral amyloid angiopathy (CAA) and Alzheimer pathology. Five decedents (mean age at death 79.6 ± 5.7 years) with documented severe CAA and Alzheimer pathology on standard neuropathologic examination were selected from a local database. Formalin-fixed coronal brain slices were scanned at 7T MRI, including high-resolution T2- and T2*-weighted sequences. Representative microbleeds from each case were sampled for histopathologic analysis, including the presence of blood, blood breakdown products, and markers of ischemic tissue injury. On MRI, we identified >300 cortical and 4 subcortical microbleeds. Two out of 15 sampled cortical microbleeds corresponded histologically to erythrocytes (suggestive of recent hemorrhages), 4 to vasculopathies (fibrinoid necrosis in 3 and a cavernoma) without substantial parenchymal tissue injury, and 9 to accumulations of iron-positive siderophages without erythrocytes (suggestive of old hemorrhages) combined with mild to moderate degrees of chronic ischemic tissue injury. This study provides evidence for heterogeneous pathologic substrates and possibly different pathophysiologic mechanisms underlying MRI-observed cortical microbleeds in the context of advanced CAA and Alzheimer disease. © 2016 American Academy of Neurology.

Protocol for Biomarker Ratio Imaging Microscopy with Specific Application to Ductal Carcinoma In situ of the Breast

PubMed Central

Clark, Andrea J.; Petty, Howard R.

2016-01-01

This protocol describes the methods and steps involved in performing biomarker ratio imaging microscopy (BRIM) using formalin fixed paraffin-embedded (FFPE) samples of human breast tissue. The technique is based on the acquisition of two fluorescence images of the same microscopic field using two biomarkers and immunohistochemical tools. The biomarkers are selected such that one biomarker correlates with breast cancer aggressiveness while the second biomarker anti-correlates with aggressiveness. When the former image is divided by the latter image, a computed ratio image is formed that reflects the aggressiveness of tumor cells while increasing contrast and eliminating path-length and other artifacts from the image. For example, the aggressiveness of epithelial cells may be assessed by computing ratio images of N-cadherin and E-cadherin images or CD44 and CD24 images, which specifically reflect the mesenchymal or stem cell nature of the constituent cells, respectively. This methodology is illustrated for tissue samples of ductal carcinoma in situ (DCIS) and invasive breast cancer. This tool should be useful in tissue studies of experimental cancer as well as the management of cancer patients. PMID:27857940

Gene expression analysis of colorectal cancer by bioinformatics strategy.

PubMed

Cui, Meng; Yuan, Junhua; Li, Jun; Sun, Bing; Li, Tao; Li, Yuantao; Wu, Guoliang

2014-10-01

We used bioinformatics technology to analyze gene expression profiles involved in colorectal cancer tissue samples and healthy controls. In this paper, we downloaded the gene expression profile GSE4107 from Gene Expression Omnibus (GEO) database, in which a total of 22 chips were available, including normal colonic mucosa tissue from normal healthy donors (n=10), colorectal cancer tissue samples from colorectal patients (n=33). To further understand the biological functions of the screened DGEs, the KEGG pathway enrichment analysis were conducted. Then we built a transcriptome network to study differentially co-expressed links. A total of 3151 DEGs of CRC were selected. Besides, total 164 DCGs (Differentially Coexpressed Gene, DCG) and 29279 DCLs (Differentially Co-expressed Link, DCL) were obtained. Furthermore, the significantly enriched KEGG pathways were Endocytosis, Calcium signaling pathway, Vascular smooth muscle contraction, Linoleic acid metabolism, Arginine and proline metabolism, Inositol phosphate metabolism and MAPK signaling pathway. Our results show that the generation of CRC involves multiple genes, TFs and pathways. Several signal and immune pathways are linked to CRC and give us more clues in the process of CRC. Hence, our work would pave ways for novel diagnosis of CRC, and provided theoretical guidance into cancer therapy.

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

Liu, Xing; Xu, Yanli; Meng, Qian

Colorectal cancer (CRC) is one of the most common types of malignant tumor worldwide. Currently, although many researchers have been devoting themselves in CRC studies, the process of locating biomarkers for CRC early diagnosis and prognostic is still very slow. Using a centrifugal proteomic reactor-based proteomic analysis of minute amount of colonic biopsies by enteroscopy sampling, 2620 protein groups were quantified between cancer mucosa and adjacent normal colorectal mucosa. Of which, 403 protein groups were differentially expressed with statistic significance between cancer and normal tissues, including 195 up-regulated and 208 down-regulated proteins in cancer tissues. Three proteins (SOD3, PRELP andmore » NGAL) were selected for further Western blot validation. And the resulting Western blot experimental results were consistent with the quantitative proteomic data. SOD3 and PRELP are down-regulated in CRC mucosa comparing to adjacent normal tissue, while NGAL is up-regulated in CRC mucosa. In conclusion, the centrifugal proteomic reactor-based label-free quantitative proteomic approach provides a highly sensitive and powerful tool for analyzing minute protein sample from tiny colorectal biopsies, which may facilitate CRC biomarkers discovery for diagnoses and prognoses. -- Highlights: •Minute amount of colonic biopsies by endoscopy is suitable for proteomic analysis. •Centrifugal proteomic reactor can be used for processing tiny clinic biopsy sample. •SOD3 and PRELP are down-regulated in CRC, while NGAL is up-regulated in CRC.« less

Detection, quantitation and identification of enteroviruses from surface waters and sponge tissue from the Florida Keys using real-time RT-PCR

USGS Publications Warehouse

Donaldson, K.A.; Griffin, Dale W.; Paul, J.H.

2002-01-01

A method was developed for the quantitative detection of pathogenic human enteroviruses from surface waters in the Florida Keys using Taqman (R) one-step Reverse transcription (RT)-PCR with the Model 7700 ABI Prism (R) Sequence Detection System. Viruses were directly extracted from unconcentrated grab samples of seawater, from seawater concentrated by vortex flow filtration using a 100kD filter and from sponge tissue. Total RNA was extracted from the samples, purified and concentrated using spin-column chromatography. A 192-196 base pair portion of the 5??? untranscribed region was amplified from these extracts. Enterovirus concentrations were estimated using real-time RT-PCR technology. Nine of 15 sample sites or 60% were positive for the presence of pathogenic human enteroviruses. Considering only near-shore sites, 69% were positive with viral concentrations ranging from 9.3viruses/ml to 83viruses/g of sponge tissue (uncorrected for extraction efficiency). Certain amplicons were selected for cloning and sequencing for identification. Three strains of waterborne enteroviruses were identified as Coxsackievirus A9, Coxsackievirus A16, and Poliovirus Sabin type 1. Time and cost efficiency of this one-step real-time RT-PCR methodology makes this an ideal technique to detect, quantitate and identify pathogenic enteroviruses in recreational waters. Copyright ?? 2002 Elsevier Science Ltd.

Simultaneous analysis of D-alanine, D-aspartic acid, and D-serine using chiral high-performance liquid chromatography-tandem mass spectrometry and its application to the rat plasma and tissues.

PubMed

Karakawa, Sachise; Shimbo, Kazutaka; Yamada, Naoyuki; Mizukoshi, Toshimi; Miyano, Hiroshi; Mita, Masashi; Lindner, Wolfgang; Hamase, Kenji

2015-11-10

A highly sensitive and selective chiral LC-MS/MS method for D-alanine, D-aspartic acid and D-serine has been developed using the precolumn derivatization reagents, 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AccQ-Tag) or p-N,N,N-trimethylammonioanilyl N'-hydroxysuccinimidyl carbamate iodide (TAHS). The thus N-tagged enantiomers of the derivatized amino acids were nicely separated within 20min using the cinchona alkaloid-based zwittterionic ion-exchange type enantioselective column, Chiralpak ZWIX(+). The selected reaction monitoring was applied for detecting the target d-amino acids in biological matrices. By using the present chiral LC-MS/MS method, the three d-amino acids and their l-forms could be simultaneously determined in the range of 0.1-500nmol/mL. Finally, the technique was successfully applied to rat plasma and tissue samples. Copyright © 2015 Elsevier B.V. All rights reserved.

VAT=TAAT-SAAT: innovative anthropometric model to predict visceral adipose tissue without resort to CT-Scan or DXA.

PubMed

Samouda, Hanen; Dutour, Anne; Chaumoitre, Kathia; Panuel, Michel; Dutour, Olivier; Dadoun, Frédéric

2013-01-01

To investigate whether a combination of a selected but limited number of anthropometric measurements predicts visceral adipose tissue (VAT) better than other anthropometric measurements, without resort to medical imaging. Abdominal anthropometric measurements are total abdominal adipose tissue indicators and global measures of VAT and SAAT (subcutaneous abdominal adipose tissue). Therefore, subtracting the anthropometric measurement the more correlated possible with SAAT while being the least correlated possible with VAT, from the most correlated abdominal anthropometric measurement with VAT while being highly correlated with TAAT, may better predict VAT. BMI participants' range was from 16.3 to 52.9 kg m(-2) . Anthropometric and abdominal adipose tissues data by computed tomography (CT-Scan) were available in 253 patients (18-78 years) (CHU Nord, Marseille) and used to develop the anthropometric VAT prediction models. Subtraction of proximal thigh circumference from waist circumference, adjusted to age and/or BMI, predicts better VAT (Women: VAT = 2.15 × Waist C - 3.63 × Proximal Thigh C + 1.46 × Age + 6.22 × BMI - 92.713; R(2) = 0.836. Men: VAT = 6 × Waist C - 4.41 × proximal thigh C + 1.19 × Age - 213.65; R(2) = 0.803) than the best single anthropometric measurement or the association of two anthropometric measurements highly correlated with VAT. Both multivariate models showed no collinearity problem. Selected models demonstrate high sensitivity (97.7% in women, 100% in men). Similar predictive abilities were observed in the validation sample (Women: R(2) = 76%; Men: R(2) = 70%). Bland and Altman method showed no systematic estimation error of VAT. Validated in a large range of age and BMI, our results suggest the usefulness of the anthropometric selected models to predict VAT in Europides (South of France). Copyright © 2013 The Obesity Society.

VAT=TAAT-SAAT: Innovative Anthropometric Model to Predict Visceral Adipose Tissue Without Resort to CT-Scan or DXA

PubMed Central

Samouda, Hanen; Dutour, Anne; Chaumoitre, Kathia; Panuel, Michel; Dutour, Olivier; Dadoun, Frédéric

2013-01-01

Objective To investigate whether a combination of a selected but limited number of anthropometric measurements predicts visceral adipose tissue (VAT) better than other anthropometric measurements, without resort to medical imaging. Hypothesis Abdominal anthropometric measurements are total abdominal adipose tissue indicators and global measures of VAT and SAAT (subcutaneous abdominal adipose tissue). Therefore, subtracting the anthropometric measurement the more correlated possible with SAAT while being the least correlated possible with VAT, from the most correlated abdominal anthropometric measurement with VAT while being highly correlated with TAAT, may better predict VAT. Design and Methods BMI participants' range was from 16.3 to 52.9 kg m−2. Anthropometric and abdominal adipose tissues data by computed tomography (CT-Scan) were available in 253 patients (18-78 years) (CHU Nord, Marseille) and used to develop the anthropometric VAT prediction models. Results Subtraction of proximal thigh circumference from waist circumference, adjusted to age and/or BMI, predicts better VAT (Women: VAT = 2.15 × Waist C − 3.63 × Proximal Thigh C + 1.46 × Age + 6.22 × BMI − 92.713; R2 = 0.836. Men: VAT = 6 × Waist C − 4.41 × proximal thigh C + 1.19 × Age − 213.65; R2 = 0.803) than the best single anthropometric measurement or the association of two anthropometric measurements highly correlated with VAT. Both multivariate models showed no collinearity problem. Selected models demonstrate high sensitivity (97.7% in women, 100% in men). Similar predictive abilities were observed in the validation sample (Women: R2 = 76%; Men: R2 = 70%). Bland and Altman method showed no systematic estimation error of VAT. Conclusion Validated in a large range of age and BMI, our results suggest the usefulness of the anthropometric selected models to predict VAT in Europides (South of France). PMID:23404678

A Prospective Study of Chronic Inflammation in Benign Prostate Tissue and Risk of Prostate Cancer: Linked PCPT and SELECT Cohorts

PubMed Central

Platz, Elizabeth A.; Kulac, Ibrahim; Barber, John R.; Drake, Charles G.; Joshu, Corinne E.; Nelson, William G.; Lucia, M. Scott; Klein, Eric A.; Lippman, Scott M.; Parnes, Howard L.; Thompson, Ian M.; Goodman, Phyllis J.; Tangen, Catherine M.; De Marzo, Angelo M.

2017-01-01

Background We leveraged two trials to test the hypothesis of an inflammation-prostate cancer link prospectively in men without indication for biopsy. Methods Prostate Cancer Prevention Trial (PCPT) participants who had an end-of-study biopsy performed per protocol that was negative for cancer and who subsequently enrolled in the Selenium and Vitamin E Cancer Prevention Trial (SELECT) were eligible. We selected all 100 cases and sampled 200 frequency-matched controls and used PCPT end-of-study biopsies as “baseline”. Five men with PSA >4 ng/mL at end-of-study biopsy were excluded. Tissue was located for 92 cases and 193 controls. We visually assessed inflammation in benign tissue. We estimated odds ratios (ORs) and 95% confidence intervals (CIs) using logistic regression adjusting for age and race. Results Mean time between biopsy and diagnosis was 5.9 years. In men previously in the PCPT placebo arm, 78.1% of cases (N=41) and 68.2% of controls (N=85) had at least one baseline biopsy core (~5 evaluated per man) with inflammation. The odds of prostate cancer (N=41 cases) appeared to increase with increasing mean percentage of tissue area with inflammation, a trend that was statistically significant for Gleason sum <4+3 disease (N=31 cases; versus 0%, >0–<1.8% OR=1.70, 1.8%–<5.0% OR=2.39, ≥5% OR=3.31, p-trend=0.047). In men previously in the finasteride arm, prevalence of inflammation did not differ between cases (76.5%; N=51) and controls (75.0%; N=108). Conclusions Benign tissue inflammation was positively associated with prostate cancer. Impact This first prospective study of men without biopsy indication supports the hypothesis that inflammation influences prostate cancer development. PMID:28754796

Genome-wide prediction and analysis of human tissue-selective genes using microarray expression data

PubMed Central

2013-01-01

Background Understanding how genes are expressed specifically in particular tissues is a fundamental question in developmental biology. Many tissue-specific genes are involved in the pathogenesis of complex human diseases. However, experimental identification of tissue-specific genes is time consuming and difficult. The accurate predictions of tissue-specific gene targets could provide useful information for biomarker development and drug target identification. Results In this study, we have developed a machine learning approach for predicting the human tissue-specific genes using microarray expression data. The lists of known tissue-specific genes for different tissues were collected from UniProt database, and the expression data retrieved from the previously compiled dataset according to the lists were used for input vector encoding. Random Forests (RFs) and Support Vector Machines (SVMs) were used to construct accurate classifiers. The RF classifiers were found to outperform SVM models for tissue-specific gene prediction. The results suggest that the candidate genes for brain or liver specific expression can provide valuable information for further experimental studies. Our approach was also applied for identifying tissue-selective gene targets for different types of tissues. Conclusions A machine learning approach has been developed for accurately identifying the candidate genes for tissue specific/selective expression. The approach provides an efficient way to select some interesting genes for developing new biomedical markers and improve our knowledge of tissue-specific expression. PMID:23369200

Evaluation of several two-dimensional gel electrophoresis techniques in cardiac proteomics.

PubMed

Li, Zhao Bo; Flint, Paul W; Boluyt, Marvin O

2005-09-01

Two-dimensional gel electrophoresis (2-DE) is currently the best method for separating complex mixtures of proteins, and its use is gradually becoming more common in cardiac proteome analysis. A number of variations in basic 2-DE have emerged, but their usefulness in analyzing cardiac tissue has not been evaluated. The purpose of the present study was to systematically evaluate the capabilities and limitations of several 2-DE techniques for separating proteins from rat heart tissue. Immobilized pH gradient strips of various pH ranges, parameters of protein loading and staining, subcellular fractionation, and detection of phosphorylated proteins were studied. The results provide guidance for proteome analysis of cardiac and other tissues in terms of selection of the isoelectric point separating window for cardiac proteins, accurate quantitation of cardiac protein abundance, stabilization of technical variation, reduction of sample complexity, enrichment of low-abundant proteins, and detection of phosphorylated proteins.

Water-quality, bed-sediment, and biological data (October 2010 through September 2011) and statistical summaries of data for streams in the Clark Fork basin, Montana

USGS Publications Warehouse

Dodge, Kent A.; Hornberger, Michelle I.; Dyke, Jessica

2013-01-01

Water, bed sediment, and biota were sampled in streams from Butte to near Missoula, Montana, as part of a monitoring program in the upper Clark Fork basin of western Montana; additional water samples were collected from near Galen to near Missoula at select sites as part of a supplemental sampling program. The sampling program was conducted by the U.S. Geological Survey in cooperation with the U.S. Environmental Protection Agency to characterize aquatic resources in the Clark Fork basin, with emphasis on trace elements associated with historic mining and smelting activities. Sampling sites were located on the Clark Fork and selected tributaries. Water samples were collected periodically at 20 sites from October 2010 through September 2011. Bed-sediment and biota samples were collected once at 14 sites during August 2011. This report presents the analytical results and quality-assurance data for water-quality, bed-sediment, and biota samples collected at sites from October 2010 through September 2011. Water-quality data include concentrations of selected major ions, trace elements, and suspended sediment. Turbidity was analyzed for water samples collected at the four sites where seasonal daily values of turbidity were being determined. Daily values of suspended-sediment concentration and suspended-sediment discharge were determined for four sites. Bed-sediment data include trace-element concentrations in the fine-grained fraction. Biological data include trace-element concentrations in whole-body tissue of aquatic benthic insects. Statistical summaries of water-quality, bed-sediment, and biological data for sites in the upper Clark Fork basin are provided for the period of record since 1985.

Water-quality, bed-sediment, and biological data (October 2015 through September 2016) and statistical summaries of data for streams in the Clark Fork Basin, Montana

USGS Publications Warehouse

Dodge, Kent A.; Hornberger, Michelle I.; Turner, Matthew A.

2018-03-30

Water, bed sediment, and biota were sampled in selected streams from Butte to near Missoula, Montana, as part of a monitoring program in the upper Clark Fork Basin of western Montana. The sampling program was led by the U.S. Geological Survey, in cooperation with the U.S. Environmental Protection Agency, to characterize aquatic resources in the Clark Fork Basin, with emphasis on trace elements associated with historic mining and smelting activities. Sampling sites were on the Clark Fork and selected tributaries. Water samples were collected periodically at 20 sites from October 2015 through September 2016. Bed-sediment and biota samples were collected once at 13 sites during August 2016.This report presents the analytical results and quality-assurance data for water-quality, bed-sediment, and biota samples collected at sites from October 2015 through September 2016. Water-quality data include concentrations of selected major ions, trace elements, and suspended sediment. Samples for analysis of turbidity were collected at 13 sites, whereas samples for analysis of dissolved organic carbon were collected at 10 sites. In addition, samples for analysis of nitrogen (nitrate plus nitrite) were collected at two sites. Daily values of mean suspended-sediment concentration and suspended-sediment discharge were determined for three sites. Seasonal daily values of turbidity were determined for five sites. Bed-sediment data include trace-element concentrations in the fine-grained (less than 0.063 millimeter) fraction. Biological data include trace-element concentrations in whole-body tissue of aquatic benthic insects. Statistical summaries of water-quality, bed-sediment, and biological data for sites in the upper Clark Fork Basin are provided for the period of record.

An exploratory study of contrast agents for soft tissue visualization by means of high resolution X-ray computed tomography imaging.

PubMed

Pauwels, E; Van Loo, D; Cornillie, P; Brabant, L; Van Hoorebeke, L

2013-04-01

High resolution X-ray computed tomography (CT), or microCT, is a promising and already widely used technique in various scientific fields. Also for histological purposes it has great potential. Although microCT has proven to be a valuable technique for the imaging of bone structures, the visualization of soft tissue structures is still an important challenge due to their low inherent X-ray contrast. One way to achieve contrast enhancement is to make use of contrast agents. However, contrary to light and electron microscopy, knowledge about contrast agents and staining procedures is limited for X-ray CT. The purpose of this paper is to identify useful X-ray contrast agents for soft tissue visualization, which can be applied in a simple way and are also suited for samples larger than (1 cm)(3) . And 28 chemical substances have been investigated. All chemicals were applied in the form of concentrated aqueous solutions in which the samples were immersed. First, strips of green Bacon were stained to evaluate contrast enhancement between muscle and adipose tissue. Furthermore it was also tested whether the contrast agents remained fixed in the tissue after staining by re-immersing them in water. Based on the results, 12 contrast agents were selected for further testing on postmortem mice hind legs, containing a variety of different tissues, including muscle, fat, bone, cartilage and tendons. It was evaluated whether the contrast agents allowed a clearer distinction between the different soft tissue structures present. Finally also penetration depth was measured. And 26 chemicals resulted in contrast enhancement between muscle and adipose tissue in the Bacon strips. Mercury(II)chloride (HgCl2 ), phosphotungstic acid (PTA), phosphomolybdic acid (PMA) and ammonium orthomolybdate ((NH4 )2 MoO4 ) remained fixed after re-immersion in water. The penetration tests showed that potassium iodide (KI) and sodium tungstate can be most efficiently used for large samples of the order of several tens of cm(3) . PMA, PTA, HgCl2 and also to a lesser extent Na2 WO4 and (NH4 )2 MoO4 allowed a clearer distinction between the different soft tissue structures present. © 2013 The Authors Journal of Microscopy © 2013 Royal Microscopical Society.

Tissue-specific concentrations and patterns of perfluoroalkyl carboxylates and sulfonates in East Greenland polar bears.

PubMed

Greaves, Alana K; Letcher, Robert J; Sonne, Christian; Dietz, Rune; Born, Erik W

2012-11-06

Several perfluoroalkyl carboxylates (PFCAs) and perfluoroalkyl sulfonates (PFSAs) of varying chain length are bioaccumulative in biota. However, wildlife reports have focused on liver and with very little examination of other tissues, and thus there is a limited understanding of their distribution and potential effects in the mammalian body. In the present study, the comparative accumulation of C(6) to C(15) PFCAs, C(4), C(6), C(8) and C(10) PFSAs, and select precursors were examined in the liver, blood, muscle, adipose, and brain of 20 polar bears (Ursus maritimus) from Scoresby Sound, Central East Greenland. Overall, PFSA and PFCA concentrations were highest in liver followed by blood > brain > muscle ≈ adipose. Liver and blood samples contained proportionally more of the shorter/medium chain length (C(6) to C(11)) PFCAs, whereas adipose and brain samples were dominated by longer chain (C(13) to C(15)) PFCAs. PFCAs with lower lipophilicities accumulated more in the liver, whereas the brain accumulated PFCAs with higher lipophilicities. The concentration ratios (±SE) between perfluorooctane sulfonate and its precursor perfluorooctane sulfonamide varied among tissues from 9 (±1):1 (muscle) to 36 (±7):1 (liver). PFCA and PFSA patterns in polar bears indicate that the pharmacokinetics of these compounds are to some extent tissue-specific, and are the result of several factors that may include differing protein interactions throughout the body.

A new classification method for MALDI imaging mass spectrometry data acquired on formalin-fixed paraffin-embedded tissue samples.

PubMed

Boskamp, Tobias; Lachmund, Delf; Oetjen, Janina; Cordero Hernandez, Yovany; Trede, Dennis; Maass, Peter; Casadonte, Rita; Kriegsmann, Jörg; Warth, Arne; Dienemann, Hendrik; Weichert, Wilko; Kriegsmann, Mark

2017-07-01

Matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) shows a high potential for applications in histopathological diagnosis, and in particular for supporting tumor typing and subtyping. The development of such applications requires the extraction of spectral fingerprints that are relevant for the given tissue and the identification of biomarkers associated with these spectral patterns. We propose a novel data analysis method based on the extraction of characteristic spectral patterns (CSPs) that allow automated generation of classification models for spectral data. Formalin-fixed paraffin embedded (FFPE) tissue samples from N=445 patients assembled on 12 tissue microarrays were analyzed. The method was applied to discriminate primary lung and pancreatic cancer, as well as adenocarcinoma and squamous cell carcinoma of the lung. A classification accuracy of 100% and 82.8%, resp., could be achieved on core level, assessed by cross-validation. The method outperformed the more conventional classification method based on the extraction of individual m/z values in the first application, while achieving a comparable accuracy in the second. LC-MS/MS peptide identification demonstrated that the spectral features present in selected CSPs correspond to peptides relevant for the respective classification. This article is part of a Special Issue entitled: MALDI Imaging, edited by Dr. Corinna Henkel and Prof. Peter Hoffmann. Copyright © 2016 Elsevier B.V. All rights reserved.

Actinides in deer tissues at the rocky flats environmental technology site.

PubMed

Todd, Andrew S; Sattelberg, R Mark

2005-11-01

Limited hunting of deer at the future Rocky Flats National Wildlife Refuge has been proposed in U.S. Fish and Wildlife planning documents as a compatible wildlife-dependent public use. Historically, Rocky Flats site activities resulted in the contamination of surface environmental media with actinides, including isotopes of americium, plutonium, and uranium. In this study, measurements of actinides [Americium-241 (241Am); Plutonium-238 (238Pu); Plutonium-239,240 (239,240Pu); uranium-233,244 (233,234U); uranium-235,236 (235,236U); and uranium-238 (238U)] were completed on select liver, muscle, lung, bone, and kidney tissue samples harvested from resident Rocky Flats deer (N = 26) and control deer (N = 1). In total, only 17 of the more than 450 individual isotopic analyses conducted on Rocky Flats deer tissue samples measured actinide concentrations above method detection limits. Of these 17 detects, only 2 analyses, with analytical uncertainty values added, exceeded threshold values calculated around a 1 x 10(-6) risk level (isotopic americium, 0.01 pCi/g; isotopic plutonium, 0.02 pCi/g; isotopic uranium, 0.2 pCi/g). Subsequent, conservative risk calculations suggest minimal human risk associated with ingestion of these edible deer tissues. The maximum calculated risk level in this study (4.73 x 10(-6)) is at the low end of the U.S. Environmental Protection Agency's acceptable risk range.

Ultrasound Imaging Techniques for Spatiotemporal Characterization of Composition, Microstructure, and Mechanical Properties in Tissue Engineering.

PubMed

Deng, Cheri X; Hong, Xiaowei; Stegemann, Jan P

2016-08-01

Ultrasound techniques are increasingly being used to quantitatively characterize both native and engineered tissues. This review provides an overview and selected examples of the main techniques used in these applications. Grayscale imaging has been used to characterize extracellular matrix deposition, and quantitative ultrasound imaging based on the integrated backscatter coefficient has been applied to estimating cell concentrations and matrix morphology in tissue engineering. Spectral analysis has been employed to characterize the concentration and spatial distribution of mineral particles in a construct, as well as to monitor mineral deposition by cells over time. Ultrasound techniques have also been used to measure the mechanical properties of native and engineered tissues. Conventional ultrasound elasticity imaging and acoustic radiation force imaging have been applied to detect regions of altered stiffness within tissues. Sonorheometry and monitoring of steady-state excitation and recovery have been used to characterize viscoelastic properties of tissue using a single transducer to both deform and image the sample. Dual-mode ultrasound elastography uses separate ultrasound transducers to produce a more potent deformation force to microscale characterization of viscoelasticity of hydrogel constructs. These ultrasound-based techniques have high potential to impact the field of tissue engineering as they are further developed and their range of applications expands.

Assessment of status of three water bodies in Serbia based on tissue metal and metalloid concentration (ICP-OES) and genotoxicity (comet assay).

PubMed

Sunjog, Karolina; Kolarević, Stoimir; Kračun-Kolarević, Margareta; Višnjić-Jeftić, Željka; Skorić, Stefan; Gačić, Zoran; Lenhardt, Mirjana; Vasić, Nebojša; Vuković-Gačić, Branka

2016-06-01

Metals and metalloids are natural components of the biosphere, which are not produced per se by human beings, but whose form and distribution can be affected by human activities. Like all substances, they are a contaminant if present in excess compared to background levels and/or in a form that would not normally occur in the environment. Samples of liver, gills, gonads and muscle from European chub, Squalius cephalus, were analyzed for Al, As, B, Ba, Cr, Cu, Fe, Hg, Mn, Mo, Sr and Zn using inductively coupled plasma optical emission spectrometry (ICP-OES) to highlight the importance of tissue selection in monitoring research. The comet assay or single cell gel electrophoresis (SCGE) was selected as an in vivo genotoxicity assay, a rapid and sensitive method for measuring genotoxic effects in blood, liver and gills of the European chub. Microscopic images of comets were scored using Comet IV Computer Software (Perceptive Instruments, UK). The objective of our study was to investigate two reservoirs, Zlatar and Garasi, and one river, Pestan by: (i) determining and comparing metal and metalloid concentrations in sediment, water and tissues of European chub: liver, gills, muscle and gonads (ii) comparing these findings with genotoxicity of water expressed through DNA damage of fish tissues. A clear link between the level of metals in water, sediment and tissues and between metal and genotoxicity levels at examined sites was not found. This suggests that other xenobiotics (possibly the organic compounds), contribute to DNA damage. Copyright © 2016 Elsevier Ltd. All rights reserved.

High Resolution X-Ray Phase Contrast Imaging with Acoustic Tissue-Selective Contrast Enhancement

DTIC Science & Technology

2005-06-01

Ultrasonics Symp 1319 (1999). 17. Sarvazyan, A. P. Shear Wave Elasticity Imaging: A New Ultrasonic Technology of Medical Diagnostics. Ultrasound in...samples using acoustically modulated X-ray phase contrast imaging. 15. SUBJECT TERMS x-ray, ultrasound, phase contrast, imaging, elastography 16...x-rays, phase contrast imaging is based on phase changes as x-rays traverse a body resulting in wave interference that result in intensity changes in

FTA Cards for Preservation of Nucleic Acids for Molecular Assays: A Review on the Use of Cytologic/Tissue Samples.

PubMed

da Cunha Santos, Gilda

2018-03-01

- Traditional methods for storing histologic and cytologic specimens for future use in molecular assays have consisted of either snap-freezing with cryopreservation or formalin-fixing, paraffin-embedding the samples. Although snap-freezing with cryopreservation is recommended for better preservation of nucleic acids, the infrastructure and space required for archiving impose challenges for high-volume pathology laboratories. Cost-effective, long-term storage at room temperature; relatively easy shipment; and standardized handling can be achieved with formalin-fixed, paraffin-embedded samples, but formalin fixation induces fragmentation and chemical modification of nucleic acids. Advances in next-generation sequencing platforms, coupled with an increase in diagnostic, prognostic, and predictive molecular biomarkers have created a demand for high-quality nucleic acids. To address issues of the quality of nucleic acid and logistics in sample acquisition, alternatives for specimen preservation and long-term storage have been described and include novel universal tissue fixatives, stabilizers, and technologies. - To collect, retrieve, and review information from studies describing the use of nucleic acids recovered from cytologic/tissue specimens stored on Flinders Technology Associates (FTA, GE Whatman, Maidstone, Kent, United Kingdom) cards for downstream molecular applications. - An electronic literature search in the PubMed (National Center for Biotechnology Information, Bethesda, Maryland) database allowed the selection of manuscripts addressing the use of FTA cards for storage of cytologic samples for molecular analysis. Only articles published in English were retrieved. - The use of FTA cards is a versatile method for fostering multicenter, international collaborations and clinical trials that require centralized testing, long-distance shipment, and high-quality nucleic acids for molecular techniques. Studies with controlled temperature are required to test the quality of recovered RNA after long-term storage.

Tissue distribution, metabolism, and residue depletion study in Atlantic salmon following oral administration of [3H]emamectin benzoate.

PubMed

Kim-Kang, Heasook; Bova, Alice; Crouch, Louis S; Wislocki, Peter G; Robinson, Robert A; Wu, Jinn

2004-04-07

Atlantic salmon (approximately 1.3 kg) maintained in tanks of seawater at 5 +/- 1 degrees C were dosed with [3H]emamectin B1 benzoate in feed at a nominal rate of 50 microg of emamectin benzoate/kg/day for 7 consecutive days. Tissues, blood, and bile were collected from 10 fish each at 3 and 12 h and at 1, 3, 7, 15, 30, 45, 60, and 90 days post final dose. Feces were collected daily from the tanks beginning just prior to dosing to 90 days post final dose. The total radioactive residues (TRR) of the daily feces samples during dosing were 0.25 ppm maximal, and >97% of the TRR in pooled feces covering the dosing period was emamectin B1a. Feces TRR then rapidly declined to approximately 0.05 ppm by 1 day post final dose. The ranges of mean TRR for tissues over the 90 days post dose period were as follows: kidney, 1.4-3 ppm; liver, 1.0-2.3 ppm; skin, 0.04-0.09 ppm; muscle, 0.02-0.06 ppm; and bone, <0.01 ppm. The residue components of liver, kidney, muscle, and skin samples pooled by post dose interval were emamectin B1a (81-100% TRR) and desmethylemamectin B1a (0-17% TRR) with N-formylemamectin B1a seen in trace amounts (<2%) in some muscle samples. The marker residue selected for regulatory surveillance of emamectin residues was emamectin B1a. The emamectin B1a level was quantified in individual samples of skin and muscle using HPLC-fluorometry and was below 85 ppb in all samples analyzed (3 h to 30 days post dose).

In vitro and in vivo evaluation of fluoroquinolone resistance associated with DNA gyrase mutations in Francisella tularensis, including in tularaemia patients with treatment failure.

PubMed

Sutera, V; Hoarau, G; Renesto, P; Caspar, Y; Maurin, M

2017-09-01

Fluoroquinolones (FQs) are highly effective for treating tularaemia, a zoonosis caused by Francisella tularensis, but failures and relapses remain common in patients with treatment delay or immunocompromised status. FQ-resistant strains of F. tularensis harboring mutations in the quinolone-resistance determining region (QRDR) of gyrA and gyrB, the genes encoding subunits A and B of DNA gyrase, have been selected in vitro. Such mutants have never been isolated from humans as this microorganism is difficult to culture. In this study, the presence of FQ-resistant mutants of F. tularensis was assessed in tularaemia patients using combined culture- and PCR-based approaches. We analyzed 42 F. tularensis strains and 82 tissue samples collected from 104 tularaemia cases, including 32 (30.7%) with FQ treatment failure or relapse. Forty F. tularensis strains and 55 clinical samples were obtained before any FQ treatment, while 2 strains and 15 tissue samples were collected after treatment. FQ resistance was evaluated by the minimum inhibitory concentration (MIC) for the bacterial strains, and by newly developed PCR-based methods targeting the gyrA and gyrB QRDRs for both the bacterial strains and the clinical samples. None of the F. tularensis strains displayed an increased MIC compared with FQ-susceptible controls. Neither gyrA nor gyrB QRDR mutation was found in bacterial strains and tissue samples tested, including those from patients with FQ treatment failure or relapse. Further phenotypic and genetic resistance traits should be explored to explain the poor clinical response to FQ treatment in such tularaemia patients. Copyright © 2017 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

Determination of volatile bases in seafood using the ammonia ion selective electrode: collaborative study.

PubMed

Ellis, P C; Pivarnik, L F; Thiam, M; Ellis, P C; Pivarnik, L F; Thiam, M

2000-01-01

Nine collaborating laboratories tested a combination of 23 seafood samples for volatile bases using an ammonia ion selective electrode. Results were reported as mg NH3/100 g fish, but the method reflected levels of both ammonia and trimethylamine, which permeated the ammonia membrane. The 23 samples were broken down into 8 blind duplicate pairs, 2 Youden matched pairs, and 3 single samples covering fresh to spoiled product ranging from 8 to 82 mg NH3/100 g. Seven species were evaluated: Atlantic cod, squid, Atlantic halibut, gray sole, monkfish, dogfish, and Atlantic mackerel. The ammonia electrode assay was performed on an aqueous homogenate consisting of 95 mL distilled water and 5.0 g sample tissue. Alkaline ion strength adjusting solution (2 mL) was added to the homogenate to liberate ammonia that was sensed by the ion specific electrode and measured on a precalibrated portable meter. Repeatability standard deviations (RSDr) ranged from 4.2 to 17%; reproducibility standard deviations (RSDR) ranged from 8.8 to 21%. A standard ammonium chloride solution was provided to all laboratories to spike 3 different samples at 10 mg NH3/100 g. Recoveries of added ammonia as ammonium chloride for fresh, borderline, and spoiled samples were 88.6, 107, and 128%, respectively.

High Levels of Bioplastic Are Produced in Fertile Transplastomic Tobacco Plants Engineered with a Synthetic Operon for the Production of Polyhydroxybutyrate1[C][OA

PubMed Central

Bohmert-Tatarev, Karen; McAvoy, Susan; Daughtry, Sean; Peoples, Oliver P.; Snell, Kristi D.

2011-01-01

An optimized genetic construct for plastid transformation of tobacco (Nicotiana tabacum) for the production of the renewable, biodegradable plastic polyhydroxybutyrate (PHB) was designed using an operon extension strategy. Bacterial genes encoding the PHB pathway enzymes were selected for use in this construct based on their similarity to the codon usage and GC content of the tobacco plastome. Regulatory elements with limited homology to the host plastome yet known to yield high levels of plastidial recombinant protein production were used to enhance the expression of the transgenes. A partial transcriptional unit, containing genes of the PHB pathway and a selectable marker gene encoding spectinomycin resistance, was flanked at the 5′ end by the host plant’s psbA coding sequence and at the 3′ end by the host plant’s 3′ psbA untranslated region. This design allowed insertion of the transgenes into the plastome as an extension of the psbA operon, rendering the addition of a promoter to drive the expression of the transgenes unnecessary. Transformation of the optimized construct into tobacco and subsequent spectinomycin selection of transgenic plants yielded T0 plants that were capable of producing up to 18.8% dry weight PHB in samples of leaf tissue. These plants were fertile and produced viable seed. T1 plants producing up to 17.3% dry weight PHB in samples of leaf tissue and 8.8% dry weight PHB in the total biomass of the plant were also isolated. PMID:21325565

Raman spectral feature selection using ant colony optimization for breast cancer diagnosis.

PubMed

Fallahzadeh, Omid; Dehghani-Bidgoli, Zohreh; Assarian, Mohammad

2018-06-04

Pathology as a common diagnostic test of cancer is an invasive, time-consuming, and partially subjective method. Therefore, optical techniques, especially Raman spectroscopy, have attracted the attention of cancer diagnosis researchers. However, as Raman spectra contain numerous peaks involved in molecular bounds of the sample, finding the best features related to cancerous changes can improve the accuracy of diagnosis in this method. The present research attempted to improve the power of Raman-based cancer diagnosis by finding the best Raman features using the ACO algorithm. In the present research, 49 spectra were measured from normal, benign, and cancerous breast tissue samples using a 785-nm micro-Raman system. After preprocessing for removal of noise and background fluorescence, the intensity of 12 important Raman bands of the biological samples was extracted as features of each spectrum. Then, the ACO algorithm was applied to find the optimum features for diagnosis. As the results demonstrated, by selecting five features, the classification accuracy of the normal, benign, and cancerous groups increased by 14% and reached 87.7%. ACO feature selection can improve the diagnostic accuracy of Raman-based diagnostic models. In the present study, features corresponding to ν(C-C) αhelix proline, valine (910-940), νs(C-C) skeletal lipids (1110-1130), and δ(CH2)/δ(CH3) proteins (1445-1460) were selected as the best features in cancer diagnosis.

Genetic parameters for sensory traits in longissimus muscle and their associations with tenderness, marbling score, and intramuscular fat in Angus cattle.

PubMed

Mateescu, R G; Garrick, D J; Garmyn, A J; VanOverbeke, D L; Mafi, G G; Reecy, J M

2015-01-01

The objective of this study was to estimate heritabilities for sensory traits and genetic correlations among sensory traits and with marbling score (MS), Warner-Bratzler shear force (WBSF), and intramuscular fat content (IMFC). Samples of LM from 2,285 Angus cattle were obtained and fabricated into steaks for laboratory analysis and 1,720 steaks were analyzed by a trained sensory panel. Restricted maximum likelihood procedures were used to obtain estimates of variance and covariance components under a multitrait animal model. Estimates of heritability for MS, IMFC, WBSF, tenderness, juiciness, and connective tissue traits were 0.67, 0.38, 0.19, 0.18, 0.06, and 0.25, respectively. The genetic correlations of MS with tenderness, juiciness, and connective tissue were estimated to be 0.57 ± 0.14, 1.00 ± 0.17, and 0.49 ± 0.13, all positive and strong. Estimated genetic correlations of IMFC with tenderness, juiciness, and connective tissue were 0.56 ± 0.16, 1.00 ± 0.21, and 0.50 ± 0.15, respectively. The genetic correlations of WBSF with tenderness, juiciness, and connective tissue were all favorable and estimated to be -0.99 ± 0.08, -0.33 ± 0.30 and -0.99 ± 0.07, respectively. Strong and positive genetic correlations were estimated between tenderness and juiciness (0.54 ± 0.28) and between connective tissue and juiciness (0.58 ± 0.26). In general, genetic correlations were large and favorable, which indicated that strong relationships exist and similar gene and gene networks may control MS, IMFC, and juiciness or WBSF, panel tenderness, and connective tissue. The results from this study confirm that MS currently used in selection breeding programs has positive genetic correlations with tenderness and juiciness and, therefore, is an effective indicator trait for the improvement of tenderness and juiciness in beef. This study also indicated that a more objective measure, particularly WBSF, a trait not easy to improve through phenotypic selection, is an excellent candidate trait for genomic selection aimed at improving eating satisfaction.

Microscopic fluorescence spectral analysis of basal cell carcinomas

NASA Astrophysics Data System (ADS)

He, Qingli; Lui, Harvey; Zloty, David; Cowan, Bryce; Warshawski, Larry; McLean, David I.; Zeng, Haishan

2007-05-01

Background and Objectives. Laser-induced autofluorescence (LIAF) is a promising tool for cancer diagnosis. This method is based on the differences in autofluorescence spectra between normal and cancerous tissues, but the underlined mechanisms are not well understood. The objective of this research is to study the microscopic origins and intrinsic fluorescence properties of basal cell carcinoma (BCC) for better understanding of the mechanism of in vivo fluorescence detection and margin delineation of BCCs on skin patients. A home-made micro- spectrophotometer (MSP) system was used to image the fluorophore distribution and to measure the fluorescence spectra of various microscopic structures and regions on frozen tissue sections. Materials and Methods. BCC tissue samples were obtained from 14 patients undergoing surgical resections. After surgical removal, each tissue sample was immediately embedded in OCT medium and snap-frozen in liquid nitrogen. The frozen tissue block was then cut into 16-μm thickness sections using a cryostat microtome and placed on microscopic glass slides. The sections for fluorescence study were kept unstained and unfixed, and then analyzed by the MSP system. The adjacent tissue sections were H&E stained for histopathological examination and also served to help identify various microstructures on the adjacent unstained sections. The MSP system has all the functions of a conventional microscope, plus the ability of performing spectral analysis on selected micro-areas of a microscopic sample. For tissue fluorescence analysis, 442nm He-Cd laser light is used to illuminate and excite the unstained tissue sections. A 473-nm long pass filter was inserted behind the microscope objective to block the transmitted laser light while passing longer wavelength fluorescence signal. The fluorescence image of the sample can be viewed through the eyepieces and also recorded by a CCD camera. An optical fiber is mounted onto the image plane of the photograph port of the microscope to collect light from a specific micro area of the sample. The collected light is transmitted via the fiber to a disperserve type CCD spectrometer for spectral analysis. Results. The measurement results showed significant spectral differences between normal and cancerous tissues. For normal tissue regions, the spectral results agreed with our previous findings on autofluorescence of normal skin sections. For the cancerous regions, the epidermis showed very weak fluorescence signal, while the stratum corneum exhibited fluorescence emissions peaking at about 510 nm. In the dermis, the basal cell island and a band of surrounding areas showed very weak fluorescence signal, while distal dermis above and below the basal cell island showed greater fluorescence signal but with different spectral shapes. The very weak autofluorescence from the basal cell island and its surrounding area may be attributed to their degenerative properties that limited the production of collagens. Conclusions. The obtained microscopic results very well explain the in vivo fluorescence properties of BCC lesions in that they have decreased fluorescence intensity compared to the surrounding normal skin. The intrinsic spectra of various microstructures and the microscopic fluorescence images (corresponding fluorophore distribution in tissue) obtained in this study will be used for further theoretical modeling of in vivo fluorescence spectroscopy and imaging of skin cancers.

Alteration of organic matter during infaunal polychaete gut passage and links to sediment organic geochemistry. Part I: Amino acids

NASA Astrophysics Data System (ADS)

Woulds, Clare; Middelburg, Jack J.; Cowie, Greg L.

2012-01-01

Of the factors which control the quantity and composition of organic matter (OM) buried in marine sediments, the links between infaunal ingestion and gut passage and sediment geochemistry have received relatively little attention. This study aimed to use feeding experiments and novel isotope tracing techniques to quantify amino acid net accumulation and loss during polychaete gut passage, and to link this to patterns of selective preservation and decay in sediments. Microcosms containing either Arenicolamarina or Hediste (formerly Nereis) diversicolor were constructed from defaunated sediment and filtered estuarine water, and maintained under natural temperature and light conditions. They were fed with 13C-labelled diatoms daily for 8 days, and animals were transferred into fresh, un-labelled sediment after ∼20 days. Samples of fauna, microcosm sediment and faecal matter were collected after 8, ∼20 and ∼40 days, and analysed for their bulk isotopic signatures and 13C-labelled amino acid compositions. Bulk isotopic data showed that, consistent with their feeding modes, Hediste assimilated added 13C more quickly, and attained a higher labelling level than Arenicola. Both species retained the added 13C in their biomass even after removal from the food. A principal component analysis of 13C-labelled amino acid mole percentages showed clear differences in composition between the algae, faunal tissues, and sediment plus faecal matter. Further, the two species of polychaete showed different compositions in their tissues. The amino acids phenylalanine, valine, leucine, iso-leucine, threonine and proline showed net accumulation in polychaete tissues. Serine, methionine, lysine, aspartic and glutamic acids and tyrosine were rapidly lost through metabolism, consistent with their presence in easily digestible cell components (as opposed to cell walls which offer physical protection). All sample types (polychaete tissues, sediments and faecal matter) were enriched in labelled glycine. Possible mechanisms for this enrichment include accumulation through inclusion in tissues with long residence times, preferential preservation (i.e. selection against) during metabolism, production from other labelled amino acids during varied metabolic processes, and accumulation in refractory by-products of secondary bacterial production. Overall, similarities were observed between amino-acid decay patterns in faunated microcosms, afaunal controls, and those previously reported in marine sediments. Thus, while polychaete gut passage did produce compound-selective accumulation and losses of certain amino acids in polychaete tissues and faecal matter, the impact of polychaete gut passage on sediment organic geochemistry was difficult to deconvolve from microbial decay. Despite processing large volumes of organic matter, polychaetes may not have distinctive influence on sediment compositions, possibly because metabolic processes concerning amino acids may be broadly similar across a wide range of organisms.

Detection of Acetaminophen-Protein Adducts in Decedents with Suspected Opioid-Acetaminophen Combination Product Overdose.

PubMed

Thomas, Karen C; Wilkins, Diana G; Curry, Steven C; Grey, Todd C; Andrenyak, David M; McGill, Lawrence D; Rollins, Douglas E

2016-09-01

Acetaminophen overdose is a leading cause of drug-induced liver failure in the United States. Acetaminophen-protein adducts have been suggested as a biomarker of hepatotoxicity. The purpose of this study was to determine whether protein-derived acetaminophen-protein adducts are quantifiable in postmortem samples. Heart blood, femoral blood, and liver tissue were collected at autopsy from 22 decedents suspected of opioid-acetaminophen overdose. Samples were assayed for protein-derived acetaminophen-protein adducts, acetaminophen, and selected opioids found in combination products containing acetaminophen. Protein-derived APAP-CYS was detected in 17 of 22 decedents and was measurable in blood that was not degraded or hemolyzed. Heart blood concentrations ranged from 11 ng/mL (0.1 μM) to 7817 ng/mL (28.9 μM). Protein-derived acetaminophen-protein adducts were detectable in liver tissue for 20 of 22 decedents. Liver histology was also performed for all decedents, and no evidence of centrilobular hepatic necrosis was observed. © 2016 American Academy of Forensic Sciences.

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.

Assessment of biological effects of environmental pollution in Mersin Bay (Turkey, northeastern Mediterranean Sea) using Mullus barbatus and Liza ramada as target organisms.

PubMed

Yılmaz, Doruk; Kalay, Mustafa; Dönmez, Erdem; Yılmaz, Nejat

2016-01-01

The increasing emphasis on the assessment and monitoring of marine ecosystems has revealed the need to use appropriate biological indicators for these areas. Enzyme activities and histopathology are increasingly being used as indicators of environmental stress since they provide a definite biological end-point of pollutant exposure. As part of an ecotoxicological assessment of Mersin Bay, EROD enzyme activity and histopathological response in selected organs and tissues of two species of fish, Mullus barbatus (red mullet) and Liza ramada (thinlip grey mullet), captured from area were examined. Pollutant (Organochlorines (OC), alkylphenols (APs) and BPA) levels and biomarker responses in tissue samples were evaluated together for their potential to alter the metabolism and cellular aspects in liver and gonad. Elevated induction of EROD activity and histopathological alterations in contaminated samples from Mersin Bay was observed compared to reference site indicating the exposure to potential pollutants. Copyright © 2015 Elsevier Ltd. All rights reserved.

Sperm Competition Selects for Sperm Quantity and Quality in the Australian Maluridae

PubMed Central

Rowe, Melissah; Pruett-Jones, Stephen

2011-01-01

When ejaculates from rival males compete for fertilization, there is strong selection for sperm traits that enhance fertilization success. Sperm quantity is one such trait, and numerous studies have demonstrated a positive association between sperm competition and both testes size and the number of sperm available for copulations. Sperm competition is also thought to favor increases in sperm quality and changes in testicular morphology that lead to increased sperm production. However, in contrast to sperm quantity, these hypotheses have received considerably less empirical support and remain somewhat controversial. In a comparative study using the Australian Maluridae (fairy-wrens, emu-wrens, grasswrens), we tested whether increasing levels of sperm competition were associated with increases in both sperm quantity and quality, as well as an increase in the relative amount of seminiferous tubule tissue contained within the testes. After controlling for phylogeny, we found positive associations between sperm competition and sperm numbers, both in sperm reserves and in ejaculate samples. Additionally, as sperm competition level increased, the proportion of testicular spermatogenic tissue also increased, suggesting that sperm competition selects for greater sperm production per unit of testicular tissue. Finally, we also found that sperm competition level was positively associated with multiple sperm quality traits, including the proportion of motile sperm in ejaculates and the proportion of both viable and morphologically normal sperm in sperm reserves. These results suggest multiple ejaculate traits, as well as aspects of testicular morphology, have evolved in response to sperm competition in the Australian Maluridae. Furthermore, our findings emphasize the importance of post-copulatory sexual selection as an evolutionary force shaping macroevolutionary differences in sperm phenotype. PMID:21283577

Sperm competition selects for sperm quantity and quality in the Australian Maluridae.

PubMed

Rowe, Melissah; Pruett-Jones, Stephen

2011-01-25

When ejaculates from rival males compete for fertilization, there is strong selection for sperm traits that enhance fertilization success. Sperm quantity is one such trait, and numerous studies have demonstrated a positive association between sperm competition and both testes size and the number of sperm available for copulations. Sperm competition is also thought to favor increases in sperm quality and changes in testicular morphology that lead to increased sperm production. However, in contrast to sperm quantity, these hypotheses have received considerably less empirical support and remain somewhat controversial. In a comparative study using the Australian Maluridae (fairy-wrens, emu-wrens, grasswrens), we tested whether increasing levels of sperm competition were associated with increases in both sperm quantity and quality, as well as an increase in the relative amount of seminiferous tubule tissue contained within the testes. After controlling for phylogeny, we found positive associations between sperm competition and sperm numbers, both in sperm reserves and in ejaculate samples. Additionally, as sperm competition level increased, the proportion of testicular spermatogenic tissue also increased, suggesting that sperm competition selects for greater sperm production per unit of testicular tissue. Finally, we also found that sperm competition level was positively associated with multiple sperm quality traits, including the proportion of motile sperm in ejaculates and the proportion of both viable and morphologically normal sperm in sperm reserves. These results suggest multiple ejaculate traits, as well as aspects of testicular morphology, have evolved in response to sperm competition in the Australian Maluridae. Furthermore, our findings emphasize the importance of post-copulatory sexual selection as an evolutionary force shaping macroevolutionary differences in sperm phenotype.

Data-driven sampling method for building 3D anatomical models from serial histology

NASA Astrophysics Data System (ADS)

Salunke, Snehal Ulhas; Ablove, Tova; Danforth, Theresa; Tomaszewski, John; Doyle, Scott

2017-03-01

In this work, we investigate the effect of slice sampling on 3D models of tissue architecture using serial histopathology. We present a method for using a single fully-sectioned tissue block as pilot data, whereby we build a fully-realized 3D model and then determine the optimal set of slices needed to reconstruct the salient features of the model objects under biological investigation. In our work, we are interested in the 3D reconstruction of microvessel architecture in the trigone region between the vagina and the bladder. This region serves as a potential avenue for drug delivery to treat bladder infection. We collect and co-register 23 serial sections of CD31-stained tissue images (6 μm thick sections), from which four microvessels are selected for analysis. To build each model, we perform semi-automatic segmentation of the microvessels. Subsampled meshes are then created by removing slices from the stack, interpolating the missing data, and re-constructing the mesh. We calculate the Hausdorff distance between the full and subsampled meshes to determine the optimal sampling rate for the modeled structures. In our application, we found that a sampling rate of 50% (corresponding to just 12 slices) was sufficient to recreate the structure of the microvessels without significant deviation from the fullyrendered mesh. This pipeline effectively minimizes the number of histopathology slides required for 3D model reconstruction, and can be utilized to either (1) reduce the overall costs of a project, or (2) enable additional analysis on the intermediate slides.

DNA degrades during storage in formalin-fixed and paraffin-embedded tissue blocks.

PubMed

Guyard, Alice; Boyez, Alice; Pujals, Anaïs; Robe, Cyrielle; Tran Van Nhieu, Jeanne; Allory, Yves; Moroch, Julien; Georges, Odette; Fournet, Jean-Christophe; Zafrani, Elie-Serge; Leroy, Karen

2017-10-01

Formalin-fixed paraffin-embedded (FFPE) tissue blocks are widely used to identify clinically actionable molecular alterations or perform retrospective molecular studies. Our goal was to quantify degradation of DNA occurring during mid to long-term storage of samples in usual conditions. We selected 46 FFPE samples of surgically resected carcinomas of lung, colon, and urothelial tract, of which DNA had been previously extracted. We performed a second DNA extraction on the same blocks under identical conditions after a median period of storage of 5.5 years. Quantitation of DNA by fluorimetry showed a 53% decrease in DNA quantity after storage. Quantitative PCR (qPCR) targeting KRAS exon 2 showed delayed amplification of DNA extracted after storage in all samples but one. The qPCR/fluorimetry quantification ratio decreased from 56 to 15% after storage (p < 0.001). Overall, remaining proportion of DNA analyzable by qPCR represented only 11% of the amount obtained at first extraction. Maximal length of amplifiable DNA fragments assessed with a multiplex PCR was reduced in DNA extracted from stored tissue, indicating that DNA fragmentation had increased in the paraffin blocks during storage. Next-generation sequencing was performed on 12 samples and showed a mean 3.3-fold decrease in library yield and a mean 4.5-fold increase in the number of single-nucleotide variants detected after storage. In conclusion, we observed significant degradation of DNA extracted from the same FFPE block after 4 to 6 years of storage. Better preservation strategies should be considered for storage of FFPE biopsy specimens.

Water-quality, bed-sediment, and biological data (October 2014 through September 2015) and statistical summaries of data for streams in the Clark Fork Basin, Montana

USGS Publications Warehouse

Dodge, Kent A.; Hornberger, Michelle I.; Turner, Matthew A.

2017-01-19

Water, bed sediment, and biota were sampled in selected streams from Butte to near Missoula, Montana, as part of a monitoring program in the upper Clark Fork Basin of western Montana. The sampling program was led by the U.S. Geological Survey, in cooperation with the U.S. Environmental Protection Agency, to characterize aquatic resources in the Clark Fork Basin, with emphasis on trace elements associated with historic mining and smelting activities. Sampling sites were located on the Clark Fork and selected tributaries. Water samples were collected periodically at 20 sites from October 2014 through September 2015. Bed-sediment and biota samples were collected once at 13 sites during August 2015.This report presents the analytical results and quality-assurance data for water-quality, bed-sediment, and biota samples collected at sites from October 2014 through September 2015. Water-quality data include concentrations of selected major ions, trace elements, and suspended sediment. At 12 sites, samples for analysis of dissolved organic carbon and turbidity were collected. In addition, samples for analysis of nitrogen (nitrate plus nitrite) were collected at two sites. Daily values of mean suspended-sediment concentration and suspended-sediment discharge were determined for three sites. Seasonal daily values of turbidity were determined for four sites. Bed-sediment data include trace-element concentrations in the fine-grained fraction. Biological data include trace-element concentrations in whole-body tissue of aquatic benthic insects. Statistical summaries of water-quality, bed-sediment, and biological data for sites in the upper Clark Fork Basin are provided for the period of record.

Volumetric bioimaging based on light field microscopy with temporal focusing illumination

NASA Astrophysics Data System (ADS)

Hsu, Feng-Chun; Sie, Yong Da; Lai, Feng-Jie; Chen, Shean-Jen

2018-02-01

Light field technique at a single shot can get the whole volume image of observed sample. Therefore, the original frame rate of the optical system can be taken as the volumetric image rate. For dynamically imaging whole micron-scale biosample, a light field microscope with temporal focusing illumination has been developed. In the light field microscope, the f-number of the microlens array (MLA) is adopted to match that of the objective; hence, the subimages via adjacent lenslets do not overlay each other. A three-dimensional (3D) deconvolution algorithm is utilized to deblur the out-of-focusing part. Conventional light field microscopy (LFM) illuminates whole volume sample even noninteresting parts; nevertheless, whole volume excitation causes even more damage on bio-sample and also increase the background noise from the out of range. Therefore, temporal focusing is integrated into the light field microscope for selecting the illumination volume. Herein, a slit on the back focal plane of the objective is utilized to control the axial excitation confinement for selecting the illumination volume. As a result, the developed light field microscope with the temporal focusing multiphoton illumination (TFMPI) can reconstruct 3D images within the selected volume, and the lateral resolution approaches to the theoretical value. Furthermore, the 3D Brownian motion of two-micron fluorescent beads is observed as the criterion of dynamic sample. With superior signal-to-noise ratio and less damage to tissue, the microscope is potential to provide volumetric imaging for vivo sample.

Biomonitoring of Environmental Status and Trends (BEST) Program: Environmental contaminants and their effects on fish in the Columbia River Basin

USGS Publications Warehouse

Hinck, Jo Ellen; Schmitt, Christopher J.; Bartish, Timothy M.; Denslow, Nancy D.; Blazer, Vicki; Anderson, Patrick J.; Coyle, James J.; Dethloff, Gail M.; Tillitt, Donald E.

2004-01-01

Common carp (Cyprinus carpio), black basses (Micropterus sp.), and largescale sucker (Catostomus macrocheilus) together accounted for 80% of the fish sampled during the study. Fish were weighed and measured then field-examined for external and internal lesions, and liver, spleen, and gonads were weighed to compute somatic indices. Selected tissues and fluids were obtained and preserved for analysis of fish health and reproductive biomarkers. Composite samples of whole fish from each station were grouped by species and gender and analyzed for persistent organic and inorganic contaminants and for dioxin-like activity using H4IIE rat hepatoma cell bioassay.

AGS16F Is a Novel Antibody Drug Conjugate Directed against ENPP3 for the Treatment of Renal Cell Carcinoma.

PubMed

Doñate, Fernando; Raitano, Arthur; Morrison, Kendall; An, Zili; Capo, Linnette; Aviña, Hector; Karki, Sher; Morrison, Karen; Yang, Peng; Ou, Jimmy; Moriya, Ryuichi; Shostak, Yuriy; Malik, Faisal; Nadell, Rossana; Liu, Wendy; Satpayev, Daulet; Atkinson, John; Joseph, Ingrid B J; Pereira, Daniel S; Challita-Eid, Pia M; Stover, David R

2016-04-15

New cancer-specific antigens are required for the design of novel antibody-drug conjugates (ADC) that deliver tumor-specific and highly potent cytotoxic therapy. Suppression subtractive hybridization identified ectonucleotide pyrophosphatase/phosphodiesterase 3 (ENPP3 or CD203c) as a potential human cancer-specific antigen. Antibodies targeting the extracellular domain of human ENPP3 were produced and selected for specific binding to ENPP3. Expression of ENPP3 in normal and cancer tissue specimens was evaluated by immunohistochemistry (IHC). ADCs comprising anti-ENPP3 Ab conjugated with maleimidocaproyl monomethyl auristatin F via a noncleavable linker (mcMMAF) were selected for therapeutic potential using binding and internalization assays, cytotoxicity assays, and tumor growth inhibition in mouse xenograft models. Pharmacodynamic markers were evaluated by IHC in tissues and ELISA in blood. ENPP3 was highly expressed in clear cell renal cell carcinoma: 92.3% of samples were positive and 83.9% showed high expression. By contrast, expression was negligible in normal tissues examined, with the exception of the kidney. High expression was less frequent in papillary renal cell carcinoma and hepatocellular carcinoma samples. AGS16F, an anti-ENPP3 antibody-mcMMAF conjugate, inhibited tumor growth in three different renal cell carcinoma (RCC) xenograft models. AGS16F localized to tumors, formed the active metabolite Cys-mcMMAF, induced cell-cycle arrest and apoptosis, and increased blood levels of caspase-cleaved cytokeratin-18, a marker of epithelial cell death. AGS16F is a promising new therapeutic option for patients with RCC and is currently being evaluated in a phase I clinical trial. ©2015 American Association for Cancer Research.

Chemokine expression in rheumatoid arthritis (RA): evidence of RANTES and macrophage inflammatory protein (MIP)-1 beta production by synovial T cells.

PubMed Central

Robinson, E; Keystone, E C; Schall, T J; Gillett, N; Fish, E N

1995-01-01

Earlier studies from this laboratory provided evidence for restricted cytokine expression in the T cell population in RA tissues. Specifically, IL-2, IL-4, IL-6 and interferon-gamma (IFN-gamma) gene expression levels were low. The selective chemoattractant and activation effects of chemokines on leucocytes identify them as potentially ideal candidates in mediating selective inflammatory processes in RA. Accordingly, we undertook studies to examine constitutive chemokine gene expression in RA tissues. RANTES, monocyte chemotactic protein-1 (MCP-1) and MIP-1 beta gene expression was examined in both the T and non-T cell populations in RA peripheral blood (PB), synovial fluid (SF) and synovial tissues (ST). Our results identified elevated levels of both RANTES and MIP-1 beta gene expression in circulating RA PB and SF T cells. By contrast, MCP-1 expression was virtually absent in RA PB, yet elevated MCP-1 mRNA levels were detected primarily in the non-T cell populations of the SF and ST samples. Histological examination of affected rheumatoid joints revealed extensive RANTES and MIP-1 beta expression in sites of lymphocyte infiltration and cell proliferation, namely the synovial lining and sublining layers. Fractionation or RA ST patient samples revealed that RANTES expression was restricted to the T cells, whereas MIP-1 beta expression was detected in both T and non-T fractions. These data suggest that MCP-1, MIP-1 beta and RANTES may have a central role in the trafficking of reactive molecules involved in immunoregulation and in the inflammatory processes in RA. Images Fig. 4 PMID:7545093

Selected organic compounds and trace elements in streambed sediments and fish tissues, Cook Inlet Basin, Alaska

USGS Publications Warehouse

Frenzel, Steven A.

2000-01-01

Organochlorines, semivolatile organic compounds (SVOCs), and trace elements were investigated in streambed sediments and fish tissues at selected sites in the Cook Inlet Basin, Alaska, during 1998. At most sites, SVOCs and organochlorine compounds were either not detected or detected at very low concentrations. Chester Creek at Arctic Boulevard at Anchorage, which was the only site sampled with a significant degree of development in the watershed, had elevated levels of many SVOCs in streambed sediment. Coring of sediments from two ponds on Chester Creek confirmed the presence of elevated concentrations of a variety of organic compounds. Moose Creek, a stream with extensive coal deposits in its watershed, had low concentrations of numerous SVOCs in streambed sediment. Three sites located in national parks or in a national wildlife refuge had no detectable concentrations of SVOCs. Trace elements were analyzed in both streambed sediments and tissues of slimy sculpin. The two media provided similar evidence for elevated concentrations of cadmium, lead, and zinc at Chester Creek. In this study, 'probable effect levels '(PELs) were determined from sediments finer than 0.063 millimeters, where concentrations tend to be greatest. Arsenic and chromium concentrations exceeded the PEL at eight and six sites respectively. Zinc exceeded the PEL at one site. Cadmium and copper concentrations were smaller than the PEL at all sites. Mercury concentrations in streambed sediments from the Deshka River were near the PEL, and selenium concentrations at that site also appear to be elevated above background levels. At half the sites where slimy sculpin were sampled, selenium concentrations were at levels that may cause adverse effects in some species.

Occurrence of specific environmental risk factors in brain tissues of sudden infant death and sudden intrauterine unexpected death victims assessed with gas chromatography-tandem mass spectrometry.

PubMed

Termopoli, Veronica; Famiglini, Giorgio; Palma, Pierangela; Magrini, Laura; Cappiello, Achille

2015-03-01

Sudden infant death syndrome (SIDS) and sudden intrauterine unexpected death syndrome (SIUDS) are an unresolved teaser in the social-medical and health setting of modern medicine and are the result of multifactorial interactions. Recently, prenatal exposure to environmental contaminants has been associated with negative pregnancy outcomes, and verification of their presence in fetal and newborn tissues is of crucial importance. A gas chromatography-tandem mass spectrometry (MS/MS) method, using a triple quadrupole analyzer, is proposed to assess the presence of 20 organochlorine pesticides, two organophosphate pesticides, one carbamate (boscalid), and a phenol (bisphenol A) in human brain tissues. Samples were collected during autopsies of infants and fetuses that died suddenly without any evident cause. The method involves a liquid-solid extraction using n-hexane as the extraction solvent. The extracts were purified with Florisil cartridges prior to the final determination. Recovery experiments using lamb brain spiked at three different concentrations in the range of 1-50 ng g(-1) were performed, with recoveries ranging from 79 to 106%. Intraday and interday repeatability were evaluated, and relative standard deviations lower than 10% and 18%, respectively, were obtained. The selectivity and sensitivity achieved in multiple reaction monitoring mode allowed us to achieve quantification and confirmation in a real matrix at levels as low as 0.2-0.6 ng g(-1). Two MS/MS transitions were acquired for each analyte, using the Q/q ratio as the confirmatory parameter. This method was applied to the analysis of 14 cerebral cortex samples (ten SIUDS and four SIDS cases), and confirmed the presence of several selected compounds.

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.

Position specific morphological characteristics of top-level male handball players.

PubMed

Srhoj, V; Marinović, M; Rogulj, N

2002-06-01

With the aim of determining the common morphological features of senior male handball players and of analyzing their specificities i.e. the probable variation between the four playing positions (back court players, wings, pivots, goalkeepers), 25 anthropometric measures were applied to the sample of 49 players, members of three top-quality handball teams. The basic descriptive parameters were calculated and then an analysis of variance and a cluster analysis were performed. A mesomorphic, athletic body- type was obtained with a marked longitudinal skeleton dimensionality, but also with a balanced ratio between the skeletal system and muscle mass and with low values of subcutaneous fat tissue. Back court players and goalkeepers are superior in terms of outstanding skeletal dimensions and circumferences. Line players, i.e. wings and pivots, have somewhat lower longitudinal measures, whereas pronounced voluminosity and a slightly higher fat tissue value differentiate pivots from players in other positions. Cluster analysis revealed a greater homogeneity in the morphological profiles of the samples of line players (wings and pivots) than in the profiles of backs or goalkeepers. The obtained position-related differences in morphological characteristics of players suggest that the experienced players from the sample were earlier successfully subjected to the selection process and oriented to a particular playing position because of the observed correspondence between their body-type and specific kinesiological demands of the position in question. Therefore, in top-quality team-handball it would be recommendable to select players whose morphological profiles are as compatible as possible with positional specificities in the demand for the game.

Singular structure of Mueller matrices images of biological crystal networks for diagnostic human tissues pathological changes

NASA Astrophysics Data System (ADS)

Sakhnovskiy, M. Y.; Ushenko, V. A.

2013-09-01

The process of converting of laser radiation by optically anisotropic crystals of biological networks are singular in the sense of total (simultaneous) of mechanisms of orientation and phase (birefringence) anisotropy the formation of polarization-inhomogeneous field of scattered radiation. This work is aimed at developing a method of polarization selection mechanisms of blood plasma polycrystalline networks anisotropy. The relationship between statistics, correlation and fractal parameters of polarization-inhomogeneous images of blood plasma and by linear dichroism and linear birefringence of polycrystalline networks albumin and globulin was found. The criteria of differentiation and diagnostic images of polarization-inhomogeneous plasma samples of the control group (donor) and a group of patients with malignant changes of breast tissue was identified.

Caries-removal effectiveness of a papain-based chemo-mechanical agent: A quantitative micro-CT study.

PubMed

Neves, Aline A; Lourenço, Roseane A; Alves, Haimon D; Lopes, Ricardo T; Primo, Laura G

2015-01-01

The aim of this study was to access the effectiveness and specificity of a papain-based chemo-mechanical caries-removal agent in providing minimum residual caries after cavity preparation. In order to do it, extracted carious molars were selected and scanned in a micro-CT before and after caries-removal procedures with the papain-based gel. Similar parameters for acquisition and reconstruction of the image stacks were used between the scans. After classification of the dentin substrate based on mineral density intervals and establishment of a carious tissue threshold, volumetric parameters related to effectiveness (mineral density of removed dentin volume and residual dentin tissue) and specificity (relation between carious dentin in removed volume and initial caries) of this caries-removal agent were obtained. In general, removed dentin volume was similar or higher than the initial carious volume, indicating that the method was able to effectively remove dentin tissue. Samples with an almost perfect accuracy in carious dentin removal also showed an increased removal of caries-affected tissue. On the contrary, less or no affected dentin was removed in samples where some carious tissue was left in residual dentin. Mineral density values in residual dentin were always higher or similar to the threshold for mineral density values in carious dentin. In conclusion, the papain-based gel was effective in removing carious dentin up to a conservative in vitro threshold. Lesion characteristics, such as activity and morphology of enamel lesion, may also influence caries-removal properties of the method. © Wiley Periodicals, Inc.

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.

Physiological comparisons of plasma and tissue metrics of selected inland and coastal steelhead kelts.

USGS Publications Warehouse

Penney, Zachary L.; Moffitt, Christine M.; Jones, Bryan; Marston, Brian

2016-01-01

The physiological status of migrating steelhead kelts (Oncorhynchus mykiss) from the Situk River, Alaska, and two tributaries of the Clearwater River, Idaho, was evaluated to explore potential differences in post-spawning survival related to energy reserves. Blood plasma samples were analyzed for metrics related to nutritional and osmotic status, and samples of white muscle tissue collected from recent mortalities at weirs were analyzed for proximate constituents. Female kelts from the Situk River had significantly higher plasma cholesterol, triglycerides, glucose and calcium concentrations, all of which suggested higher lipid and energy stores. Additional support for energy limitation in kelts was provided by evaluating the presence of detectable proteins in the plasma. Most all kelts sampled from the Situk River populations had detectable plasma proteins, in contrast to kelts sampled from the Clearwater River tributary populations where 27 % of kelts from one tributary, and 68 % of the second tributary were below the limits of detection. We found proximate constituents of kelt mortalities were similar between the Situk and Clearwater River populations, and the lipid fraction of white muscle averaged 0.1 and 0.2 %. Our findings lend support to the hypothesis that energetic limitations likely affect post-spawn survival in the Clearwater River kelts.

A Stratified Transcriptomics Analysis of Polygenic Fat and Lean Mouse Adipose Tissues Identifies Novel Candidate Obesity Genes

PubMed Central

Morton, Nicholas M.; Nelson, Yvonne B.; Michailidou, Zoi; Di Rollo, Emma M.; Ramage, Lynne; Hadoke, Patrick W. F.; Seckl, Jonathan R.; Bunger, Lutz; Horvat, Simon; Kenyon, Christopher J.; Dunbar, Donald R.

2011-01-01

Background Obesity and metabolic syndrome results from a complex interaction between genetic and environmental factors. In addition to brain-regulated processes, recent genome wide association studies have indicated that genes highly expressed in adipose tissue affect the distribution and function of fat and thus contribute to obesity. Using a stratified transcriptome gene enrichment approach we attempted to identify adipose tissue-specific obesity genes in the unique polygenic Fat (F) mouse strain generated by selective breeding over 60 generations for divergent adiposity from a comparator Lean (L) strain. Results To enrich for adipose tissue obesity genes a ‘snap-shot’ pooled-sample transcriptome comparison of key fat depots and non adipose tissues (muscle, liver, kidney) was performed. Known obesity quantitative trait loci (QTL) information for the model allowed us to further filter genes for increased likelihood of being causal or secondary for obesity. This successfully identified several genes previously linked to obesity (C1qr1, and Np3r) as positional QTL candidate genes elevated specifically in F line adipose tissue. A number of novel obesity candidate genes were also identified (Thbs1, Ppp1r3d, Tmepai, Trp53inp2, Ttc7b, Tuba1a, Fgf13, Fmr) that have inferred roles in fat cell function. Quantitative microarray analysis was then applied to the most phenotypically divergent adipose depot after exaggerating F and L strain differences with chronic high fat feeding which revealed a distinct gene expression profile of line, fat depot and diet-responsive inflammatory, angiogenic and metabolic pathways. Selected candidate genes Npr3 and Thbs1, as well as Gys2, a non-QTL gene that otherwise passed our enrichment criteria were characterised, revealing novel functional effects consistent with a contribution to obesity. Conclusions A focussed candidate gene enrichment strategy in the unique F and L model has identified novel adipose tissue-enriched genes contributing to obesity. PMID:21915269

Associations between cruciferous vegetable intake and selected biomarkers among women scheduled for breast biopsies.

PubMed

Zhang, Zhenzhen; Atwell, Lauren L; Farris, Paige E; Ho, Emily; Shannon, Jackilen

2016-05-01

To examine the relationship between dietary cruciferous vegetable intake and selected tumour biomarkers for histone acetylation (H3K9ac, H3K18ac, HDAC3 and HDAC6), proliferation (Ki-67) and cell-cycle regulation (p21) from breast tissue. The study used baseline data of women recruited to participate in a clinical trial of sulforaphane supplement. Dietary cruciferous vegetable intake was collected through a validated Arizona Cruciferous Vegetable Intake Questionnaire. Breast tissue was obtained from biopsy samples. Spearman correlations were calculated between intake of specific cruciferous vegetables and biomarkers. Tissue biomarkers were log2-transformed to obtain approximate normality. Linear regression analyses were conducted to examine associations between cruciferous vegetable intake and biomarkers adjusting for age and use of non-steroidal anti-inflammatory drugs. False discovery rate (FDR) was used to account for multiple comparisons. Clinical trial baseline. Fifty-four women who had abnormal mammogram findings and were scheduled for breast biopsy. Mean intake of total cruciferous vegetables from all food sources was 81·7 (sd 57·3) g/d. Mean urinary total sulforaphane metabolites was 0·08 (sd 0·07) µm/mm creatinine. Total cruciferous vegetable intake was inversely associated with Ki-67 protein expression in breast ductal carcinoma in situ (DCIS) tissue (β=-0·004; se=0·001; FDR q value=0·03), but not in benign or invasive ductal carcinoma (IDC) tissue. No association was found for other biomarkers measured (HDAC3, HDAC6, H3K9, H3K18 and p21) in all tissues examined (benign, DCIS and IDC). The present study sought to provide additional evidence for the potential role of sulforaphane in histone acetylation and cell proliferation. Here, we report that total cruciferous vegetable intake is associated with decreased cell proliferation in breast DCIS tissue.

First plasma and tissue pharmacokinetic study of the YSNSG cyclopeptide, a new integrin antagonist, using microdialysis.

PubMed

Slimano, Florian; Djerada, Zoubir; Bouchene, Salim; Van Gulick, Laurence; Brassart-Pasco, Sylvie; Dukic, Sylvain

2017-07-15

The YSNSG peptide is a synthetic peptide targeting α v β 3 integrin. This peptide exhibits promising activity in vitro and in vivo against melanoma. To determine pharmacokinetic parameters and predictive active doses in the central nervous system (CNS) and subcutaneous tissue (SC), we conducted microdialysis coupled with pharmacokinetic modeling and Monte Carlo simulation. After a recovery period of surgical procedures, a microdialysis probe was inserted in the caudate and in subcutaneous tissue. Plasma samples and dialysates collected 5h after YSNSG intravenous administration (10mg/kg) were analyzed by UPLC-MS/MS. A nonlinear mixed-effect modeling approach implemented in Monolix® 2016R1 was performed. Model selection and evaluation were based on the usual diagnostic plot, precision and information criteria. The primary plasma and tissue pharmacokinetic parameters were comparable with those of other integrin antagonists, such as cilengitide or ATN-161. Tissue/plasma and brain/plasma area under the curve (AUC) ratio were 66.2±21.6% and 3.6±4.7%, respectively. Two models of 2-compartments with an additional microdialysis compartment, parameterized as rate constants (k for elimination, k12/k21 and k13/k31 for distribution) and volumes (central V1 and peripheral microdialysis compartment V3) with zero-order input were selected to describe the dialysate concentrations in CNS and SC. The inter-individual variability (IIV) was described by exponential terms, and residual variability was described by a combined additive and proportional error model. Individual AUC (plasma and tissues) values were derived for each animal using the Empirical-Bayes-Estimates of the individual parameters. The regimens needed to achieve an in vitro predetermined target concentration in tissues were studied by Monte Carlo simulations using Monolix® 2016R1. YSNSG pharmacokinetic parameters show promising results in terms of subcutaneous disposition. Further investigations into such processes as encapsulation and intratumoral disposition are currently being conducted. Copyright © 2017 Elsevier B.V. All rights reserved.

A stratified transcriptomics analysis of polygenic fat and lean mouse adipose tissues identifies novel candidate obesity genes.

PubMed

Morton, Nicholas M; Nelson, Yvonne B; Michailidou, Zoi; Di Rollo, Emma M; Ramage, Lynne; Hadoke, Patrick W F; Seckl, Jonathan R; Bunger, Lutz; Horvat, Simon; Kenyon, Christopher J; Dunbar, Donald R

2011-01-01

Obesity and metabolic syndrome results from a complex interaction between genetic and environmental factors. In addition to brain-regulated processes, recent genome wide association studies have indicated that genes highly expressed in adipose tissue affect the distribution and function of fat and thus contribute to obesity. Using a stratified transcriptome gene enrichment approach we attempted to identify adipose tissue-specific obesity genes in the unique polygenic Fat (F) mouse strain generated by selective breeding over 60 generations for divergent adiposity from a comparator Lean (L) strain. To enrich for adipose tissue obesity genes a 'snap-shot' pooled-sample transcriptome comparison of key fat depots and non adipose tissues (muscle, liver, kidney) was performed. Known obesity quantitative trait loci (QTL) information for the model allowed us to further filter genes for increased likelihood of being causal or secondary for obesity. This successfully identified several genes previously linked to obesity (C1qr1, and Np3r) as positional QTL candidate genes elevated specifically in F line adipose tissue. A number of novel obesity candidate genes were also identified (Thbs1, Ppp1r3d, Tmepai, Trp53inp2, Ttc7b, Tuba1a, Fgf13, Fmr) that have inferred roles in fat cell function. Quantitative microarray analysis was then applied to the most phenotypically divergent adipose depot after exaggerating F and L strain differences with chronic high fat feeding which revealed a distinct gene expression profile of line, fat depot and diet-responsive inflammatory, angiogenic and metabolic pathways. Selected candidate genes Npr3 and Thbs1, as well as Gys2, a non-QTL gene that otherwise passed our enrichment criteria were characterised, revealing novel functional effects consistent with a contribution to obesity. A focussed candidate gene enrichment strategy in the unique F and L model has identified novel adipose tissue-enriched genes contributing to obesity.

Molecular detection of Coxiella burnetii from the formalin-fixed tissues of Q fever patients with acute hepatitis.

PubMed

Jang, Young-Rock; Shin, Yong; Jin, Choong Eun; Koo, Bonhan; Park, Se Yoon; Kim, Min-Chul; Kim, Taeeun; Chong, Yong Pil; Lee, Sang-Oh; Choi, Sang-Ho; Kim, Yang Soo; Woo, Jun Hee; Kim, Sung-Han; Yu, Eunsil

2017-01-01

Serologic diagnosis is one of the most widely used diagnostic methods for Q fever, but the window period in antibody response of 2 to 3 weeks after symptom onset results in significant diagnostic delay. We investigated the diagnostic utility of Q fever PCR from formalin-fixed liver tissues in Q fever patients with acute hepatitis. We reviewed the clinical and laboratory data in patients with Q fever hepatitis who underwent liver biopsy during a 17-year period, and whose biopsied tissues were available. We also selected patients who revealed granuloma in liver biopsy and with no Q fever diagnosis within the last 3 years as control. Acute Q fever hepatitis was diagnosed if two or more of the following clinical, serologic, or histopathologic criteria were met: (1) an infectious hepatitis-like clinical feature such as fever (≥ 38°C) with elevated hepatic transaminase levels; (2) exhibition of a phase II immunoglobulin G (IgG) antibodies titer by IFA of ≥ 1:128 in single determination, or a four-fold or greater rise between two separate samples obtained two or more weeks apart; (3) histologic finding of biopsy tissue showing characteristic fibrin ring granuloma. A total of 11 patients with acute Q fever hepatitis were selected and analyzed. Of the 11 patients, 3 (27%) had exposure to zoonotic risk factors and 7 (63%) met the serologic criteria. Granulomas with either circumferential or radiating fibrin deposition were observed in 10 cases on liver biopsy and in 1 case on bone marrow biopsy. 8 (73%) revealed positive Coxiella burnetii PCR from their formalin-fixed liver tissues. In contrast, none of 10 patients with alternative diagnosis who had hepatic granuloma revealed positive C. burnetii PCR from their formalin-fixed liver tissues. Q fever PCR from formalin-fixed liver tissues appears to be a useful adjunct for diagnosing Q fever hepatitis.

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.

CD13 as target for tissue factor induced tumor vascular infarction in small cell lung cancer.

PubMed

Schmidt, Lars Henning; Stucke-Ring, Janine; Brand, Caroline; Schliemann, Christoph; Harrach, Saliha; Muley, Thomas; Herpel, Esther; Kessler, Torsten; Mohr, Michael; Görlich, Dennis; Kreuter, Michael; Lenz, Georg; Wardelmann, Eva; Thomas, Michael; Berdel, Wolfgang E; Schwöppe, Christian; Hartmann, Wolfgang

2017-11-01

Zinc-binding protease aminopeptidase N (CD13) is expressed on tumor vascular cells and tumor cells. It represents a potential candidate for molecular targeted therapy, e.g. employing truncated tissue factor (tTF)-NGR, which can bind CD13 and thereby induce tumor vascular infarction. We performed a comprehensive analysis of CD13 expression in a clinically well characterized cohort of patients with small cell lung cancer (SCLC) to evaluate its potential use for targeted therapies in this disease. CD13 expression was analyzed immunohistochemically in 27 SCLC patients and correlated with clinical course and outcome. In CD-1 nude mice bearing human HTB119 SCLC xenotransplants, the systemic effects of the CD13-targeting fusion protein tTF-NGR on tumor growth were tested. In 52% of the investigated SCLC tissue samples, CD13 was expressed in tumor stroma cells, while the tumor cells were negative for CD13. No prognostic effect was found in the investigated SCLC study collective with regard to overall survival (p>0.05). In CD-1 nude mice, xenografts of CD13 negative HTB119 SCLC cells showed CD13 expression in the intratumoral vascular and perivascular cells, and the systemic application of CD13-targeted tissue factor tTF-NGR led to a significant reduction of tumor growth. We here present first data on the expression of CD13 in SCLC tumor samples. Our results strongly recommend the further investigation of tTF-NGR and other molecules targeted by NGR-peptides in SCLC patients. Considering the differential expression of CD13 in SCLC samples pre-therapeutic CD13 analysis is proposed for testing as investigational predictive biomarker for patient selection. Copyright © 2017 Elsevier B.V. All rights reserved.

Development of suspect and non-target screening methods for detection of organic contaminants in highway runoff and fish tissue with high-resolution time-of-flight mass spectrometry.

PubMed

Du, Bowen; Lofton, Jonathan M; Peter, Katherine T; Gipe, Alexander D; James, C Andrew; McIntyre, Jenifer K; Scholz, Nathaniel L; Baker, Joel E; Kolodziej, Edward P

2017-09-20

Untreated urban stormwater runoff contributes to poor water quality in receiving waters. The ability to identify toxicants and other bioactive molecules responsible for observed adverse effects in a complex mixture of contaminants is critical to effective protection of ecosystem and human health, yet this is a challenging analytical task. The objective of this study was to develop analytical methods using liquid chromatography coupled to high-resolution quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) to detect organic contaminants in highway runoff and in runoff-exposed fish (adult coho salmon, Oncorhynchus kisutch). Processing of paired water and tissue samples facilitated contaminant prioritization and aided investigation of chemical bioavailability and uptake processes. Simple, minimal processing effort solid phase extraction (SPE) and elution procedures were optimized for water samples, and selective pressurized liquid extraction (SPLE) procedures were optimized for fish tissues. Extraction methods were compared by detection of non-target features and target compounds (e.g., quantity and peak area), while minimizing matrix interferences. Suspect screening techniques utilized in-house and commercial databases to prioritize high-risk detections for subsequent MS/MS characterization and identification efforts. Presumptive annotations were also screened with an in-house linear regression (log K ow vs. retention time) to exclude isobaric compounds. Examples of confirmed identifications (via reference standard comparison) in highway runoff include ethoprophos, prometon, DEET, caffeine, cotinine, 4(or 5)-methyl-1H-methylbenzotriazole, and acetanilide. Acetanilide was also detected in runoff-exposed fish gill and liver samples. Further characterization of highway runoff and fish tissues (14 and 19 compounds, respectively with tentative identification by MS/MS data) suggests that many novel or poorly characterized organic contaminants exist in urban stormwater runoff and exposed biota.

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.

Effect of soil and water environment on typeability of PowerPlex Y (Promega) in selected tissue samples.

PubMed

Niemcunowicz-Janica, Anna; Pepinski, Witold; Janica, Jacek Robert; Skawronska, Malgorzata; Janica, Jerzy; Koc-Zorawska, Ewa; Stolyszewski, Ireneusz

2007-01-01

In cases of decomposed bodies Y chromosomal STR markers may be useful in identification of a male relative. The authors assessed typeability PowerPlex Y (Promega) loci in tissue material stored in water and soil environment. Tissue material was collected during autopsies of five persons aged 20-30 years with time of death determined within the limit of 14 hours. Heart muscle, liver and lung specimens were stored in pond water, sea water, sand and peat soil. DNA was extracted by organic method from tissue samples collected in 7-day intervals. Liver specimens were typeable in all PowerPlex Y loci within 100 days of storage in pond water with gradual decline at DYS392 in sea water. Heart muscle specimens stored in pond water exhibited allelic loss at DYS19, DYS385, DYS389II and DYS392, while all loci were typeable in sea water stored samples. For lung specimens allelic loss was noted throughout the profile. Storage of liver specimens in peat soil for more than 14 days resulted in allelic drop-out, and after 21 days no profiles were typeable. Heart muscle specimens were typeable in all PowerPlex Y systems after 35-day storage in sand, while allelic drop-out and subsequent lack of profiles were noted after 14 and 35 days respectively. Lung specimens stored in garden soil exhibited allelic drop-out and subsequent lack of profiles after 7 and 21 days, respectively. All PowerPlex Y loci were typeable in the latter material in sand up to day 35 with gradual decline of longer amplicons (DYS19, DYS385, DYS389II and DYS392).

Concentrations of selected trace elements in fish tissue and streambed sediment in the Clark Fork-Pend Oreille and Spokane River basins, Washington, Idaho, and Montana, 1998

USGS Publications Warehouse

Maret, Terry R.; Skinner, K.D.

2000-01-01

Fish tissue and bed sediment samples were collected from 16 stream sites in the Northern Rockies Intermontane Basins study area in 1998 as part of the U.S. Geological Survey National Water-Quality Assessment Program. Bed sediment samples were analyzed for 45 trace elements, and fish livers and sportfish fillets were analyzed for 22 elements to characterize the occurrence and distribution of these elements in relation to stream characteristics and land use activities. Nine trace elements of environmental concern—arsenic, cadmium, chromium, copper, lead, mercury, nickel, selenium, and zinc—were detected in bed sediment, but not all of these elements were detected in fish tissue. Trace-element concentrations were highest in bed sediment samples collected at sites downstream from significant natural mineral deposits and (or) mining activities. Arsenic, cadmium, copper, lead, mercury, and zinc in bed sediment at some sites were elevated relative to national median concentrations, and some concentrations were at levels that can adversely affect aquatic biota. Although trace-element concentrations in bed sediment exceeded various guidelines, no concentrations in sportfish fillets exceeded U.S. Environmental Protection Agency screening values for the protection of human health. Correlations between most trace-element concentrations in bed sediment and fish tissue (liver and fillet) were not significant (r0.05). Concentrations of arsenic, cadmium, copper, lead, mercury, nickel, selenium, and zinc in bed sediment were significantly correlated (r=0.53 to 0.88, p2=0.95 and 0.99, p<0.001) that corresponded to trace-element enrichment categories. These strong relations warrant further study using mine density as an explanatory variable to predict trace-element concentrations in bed sediment.

Evaluation of two main RNA-seq approaches for gene quantification in clinical RNA sequencing: polyA+ selection versus rRNA depletion.

PubMed

Zhao, Shanrong; Zhang, Ying; Gamini, Ramya; Zhang, Baohong; von Schack, David

2018-03-19

To allow efficient transcript/gene detection, highly abundant ribosomal RNAs (rRNA) are generally removed from total RNA either by positive polyA+ selection or by rRNA depletion (negative selection) before sequencing. Comparisons between the two methods have been carried out by various groups, but the assessments have relied largely on non-clinical samples. In this study, we evaluated these two RNA sequencing approaches using human blood and colon tissue samples. Our analyses showed that rRNA depletion captured more unique transcriptome features, whereas polyA+ selection outperformed rRNA depletion with higher exonic coverage and better accuracy of gene quantification. For blood- and colon-derived RNAs, we found that 220% and 50% more reads, respectively, would have to be sequenced to achieve the same level of exonic coverage in the rRNA depletion method compared with the polyA+ selection method. Therefore, in most cases we strongly recommend polyA+ selection over rRNA depletion for gene quantification in clinical RNA sequencing. Our evaluation revealed that a small number of lncRNAs and small RNAs made up a large fraction of the reads in the rRNA depletion RNA sequencing data. Thus, we recommend that these RNAs are specifically depleted to improve the sequencing depth of the remaining RNAs.

Proteomic Profiling of Paraffin-Embedded Samples Identifies Metaplasia-Specific and Early-Stage Gastric Cancer Biomarkers

PubMed Central

Sousa, Josane F.; Ham, Amy-Joan L.; Whitwell, Corbin; Nam, Ki Taek; Lee, Hyuk-Joon; Yang, Han-Kwang; Kim, Woo Ho; Zhang, Bing; Li, Ming; LaFleur, Bonnie; Liebler, Daniel C.; Goldenring, James R.

2013-01-01

Early diagnosis and curative resection are the predominant factors associated with increased survival in patients with gastric cancer. However, most gastric cancer cases are still diagnosed at later stages. Since most pathologic specimens are archived as FFPE samples, the ability to use them to generate expression profiles can greatly improve cancer biomarker discovery. We sought to uncover new biomarkers for stomach preneoplastic metaplasias and neoplastic lesions by generating proteome profiles using FFPE samples. We combined peptide isoelectric focusing and liquid chromatography–tandem mass spectrometry analysis to generate proteomic profiles from FFPE samples of intestinal-type gastric cancer, metaplasia, and normal mucosa. The expression patterns of selected proteins were analyzed by immunostaining first in single tissue sections from normal stomach, metaplasia, and gastric cancer and later in larger tissue array cohorts. We detected 60 proteins up-regulated and 87 proteins down-regulated during the progression from normal mucosa to metaplasia to gastric cancer. Two of the up-regulated proteins, LTF and DMBT1, were validated as specific markers for spasmolytic polypeptide–expressing metaplasia and intestinal metaplasia, respectively. In cancers, significantly lower levels of DMBT1 or LTF correlated with more advanced disease and worse prognosis. Thus, proteomic profiling using FFPE samples has led to the identification of two novel markers for stomach metaplasias and gastric cancer prognosis. PMID:22944598

Identification, Characterization, and Utilization of Adult Meniscal Progenitor Cells

DTIC Science & Technology

2017-11-01

approach including row scaling and Ward’s minimum variance method was chosen. This analysis revealed two groups of four samples each. For the selected...articular cartilage in an ovine model. Am J Sports Med. 2008;36(5):841-50. 7. Deshpande BR, Katz JN, Solomon DH, Yelin EH, Hunter DJ, Messier SP, et al...Miosge1,* 1Tissue Regeneration Work Group , Department of Prosthodontics, Medical Faculty, Georg-August-University, 37075 Goettingen, Germany 2Institute of

Biomarkers for Early Detection of Clinically Relevant Prostate Cancer. A Multi-Institutional Validation Trial

DTIC Science & Technology

2016-10-01

Study (PASS). We are in the process of evaluating these three biomarker panels in tissue, blood, and urine samples with well annotated clinical and...impacting both the initial choice of therapy and decision-making during AS. The objective of the study is to utilize analytically validated assays that...predict reclassification from Gleason 6 cancer to Gleason 7 or greater. The analysis plan was determined before specimens were selected for the study

Biomarkers for Early Detection of Clinically Relevant Prostate Cancer. A Multi-Institutional Validation Trial

DTIC Science & Technology

2016-10-01

Study (PASS). We are in the process of evaluating these three biomarker panels in tissue, blood, and urine samples with well annotated clinical and...choice of therapy and decision-making during AS. The objective of the study is to utilize analytically validated assays that take into account tumor...Gleason 6 cancer to Gleason 7 or greater. The analysis plan was determined before specimens were selected for the study , and included 7 breaking

OpenMSI Arrayed Analysis Tools v2.0

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

BOWEN, BENJAMIN; RUEBEL, OLIVER; DE ROND, TRISTAN

2017-02-07

Mass spectrometry imaging (MSI) enables high-resolution spatial mapping of biomolecules in samples and is a valuable tool for the analysis of tissues from plants and animals, microbial interactions, high-throughput screening, drug metabolism, and a host of other applications. This is accomplished by desorbing molecules from the surface on spatially defined locations, using a laser or ion beam. These ions are analyzed by a mass spectrometry and collected into a MSI 'image', a dataset containing unique mass spectra from the sampled spatial locations. MSI is used in a diverse and increasing number of biological applications. The OpenMSI Arrayed Analysis Tool (OMAAT)more » is a new software method that addresses the challenges of analyzing spatially defined samples in large MSI datasets, by providing support for automatic sample position optimization and ion selection.« less

Optical diffusion property of chicken tissue

NASA Astrophysics Data System (ADS)

Schneider, Patricia S.; Flamholz, Alex; Wong, Peter K.; Lieberman, David H.; Cheung, Tak D.; Itoka, Harriet; Minott, Troy; Quizhpi, Janie; Rodriguez, Jacquelin

2004-11-01

Chicken tissue acts as a turbid medium in optical wavelength. Optical characterization data of fresh chicken dark and white meat were studied using the theory of light diffusion. The gaussian-like transmission profile was used to determine the transport mean free path and absorption. The refractive index, a fundamental parameter, was extracted via transmission correlation function analysis without using index-matching fluid. The variation in refractive index also produced various small shifts in the oscillatory feature of the intensity spatial correlation function at distance shorter than the transport mean free path. The optical system was calibrated with porous silicate slabs containing different water contents and also with a solid alumina slab. The result suggested that the selective scattering/absorption of myoglobin and mitochondria in the dark tissues is consistent with the transmission data. The refractive index was similar for dark and white tissues at the He-Ne wavelength and suggested that the index could serve as a marker for quality control. Application to chicken lunchmeat samples revealed that higher protein and lower carbohydrate would shift the correlation toward smaller distance. The pure fat refractive index was different from that of the meat tissue. Application of refractive index as a fat marker is also discussed

Genome-wide DNA methylation sequencing reveals miR-663a is a novel epimutation candidate in CIMP-high endometrial cancer

PubMed Central

Yanokura, Megumi; Banno, Kouji; Adachi, Masataka; Aoki, Daisuke; Abe, Kuniya

2017-01-01

Aberrant DNA methylation is widely observed in many cancers. Concurrent DNA methylation of multiple genes occurs in endometrial cancer and is referred to as the CpG island methylator phenotype (CIMP). However, the features and causes of CIMP-positive endometrial cancer are not well understood. To investigate DNA methylation features characteristic to CIMP-positive endometrial cancer, we first classified samples from 25 patients with endometrial cancer based on the methylation status of three genes, i.e. MLH1, CDH1 (E-cadherin) and APC: CIMP-high (CIMP-H, 2/25, 8.0%), CIMP-low (CIMP-L, 7/25, 28.0%) and CIMP-negative (CIMP(-), 16/25, 64.0%). We then selected two samples each from CIMP-H and CIMP(-) classes, and analyzed DNA methylation status of both normal (peripheral blood cells: PBCs) and cancer tissues by genome-wide, targeted bisulfite sequencing. Genomes of the CIMP-H cancer tissues were significantly hypermethylated compared to those of the CIMP(-). Surprisingly, in normal tissues of the CIMP-H patients, promoter region of the miR-663a locus is hypermethylated relative to CIMP(-) samples. Consistent with this finding, miR-663a expression was lower in the CIMP-H PBCs than in the CIMP(-) PBCs. The same region of the miR663a locus is found to be highly methylated in cancer tissues of both CIMP-H and CIMP(-) cases. This is the first report showing that aberrant DNA methylation of the miR-663a promoter can occur in normal tissue of the cancer patients, suggesting a possible link between this epigenetic abnormality and endometrial cancer. This raises the possibility that the hypermethylation of the miR-663a promoter represents an epimutation associated with the CIMP-H endometrial cancers. Based on these findings, relationship of the aberrant DNA methylation and CIMP-H phenotype is discussed. PMID:28440489

Genome-wide DNA methylation sequencing reveals miR-663a is a novel epimutation candidate in CIMP-high endometrial cancer.

PubMed

Yanokura, Megumi; Banno, Kouji; Adachi, Masataka; Aoki, Daisuke; Abe, Kuniya

2017-06-01

Aberrant DNA methylation is widely observed in many cancers. Concurrent DNA methylation of multiple genes occurs in endometrial cancer and is referred to as the CpG island methylator phenotype (CIMP). However, the features and causes of CIMP-positive endometrial cancer are not well understood. To investigate DNA methylation features characteristic to CIMP-positive endometrial cancer, we first classified samples from 25 patients with endometrial cancer based on the methylation status of three genes, i.e. MLH1, CDH1 (E-cadherin) and APC: CIMP-high (CIMP-H, 2/25, 8.0%), CIMP-low (CIMP-L, 7/25, 28.0%) and CIMP-negative (CIMP(-), 16/25, 64.0%). We then selected two samples each from CIMP-H and CIMP(-) classes, and analyzed DNA methylation status of both normal (peripheral blood cells: PBCs) and cancer tissues by genome-wide, targeted bisulfite sequencing. Genomes of the CIMP-H cancer tissues were significantly hypermethylated compared to those of the CIMP(-). Surprisingly, in normal tissues of the CIMP-H patients, promoter region of the miR-663a locus is hypermethylated relative to CIMP(-) samples. Consistent with this finding, miR-663a expression was lower in the CIMP-H PBCs than in the CIMP(-) PBCs. The same region of the miR663a locus is found to be highly methylated in cancer tissues of both CIMP-H and CIMP(-) cases. This is the first report showing that aberrant DNA methylation of the miR-663a promoter can occur in normal tissue of the cancer patients, suggesting a possible link between this epigenetic abnormality and endometrial cancer. This raises the possibility that the hypermethylation of the miR-663a promoter represents an epimutation associated with the CIMP-H endometrial cancers. Based on these findings, relationship of the aberrant DNA methylation and CIMP-H phenotype is discussed.

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.

Comparative anti-microbial efficacy of Azadirachta indica irrigant with standard endodontic irrigants: A preliminary study

PubMed Central

Dutta, Arindam; Kundabala, Mala

2014-01-01

Objective: The anti-microbial efficacy of 2.5% sodium hypochlorite (SHC) and 0.2% chlorhexidine gluconate were compared with an experimental irrigant formulated from the Neem tree, Azadirachta indica A. Juss. Materials and Methods: A sample of 36 single rooted anterior teeth with periapical radiolucency and absence of response to vitality tests that required root canal treatment were selected for this study. The test irrigants and their combinations were assigned to five different groups and saline served as the control. Access cavities were prepared using an aseptic technique and samples collected for both anaerobic culture and Gram stained smears, followed by irrigation and sample collection again. The number of organisms were expressed in colony forming units/ml after 72 h of incubation; the smears were analyzed for their microbial loads and tissue clearance and assessed as per defined criteria. Results: Our results found the maximum reduction in microbial loads, when analyzed by culture method, with a combination of SHC and the experimental neem irrigant. Maximum tissue clearance on the Gram Stained smears was also found with the same combination. Conclusion: Neem irrigant has anti-microbial efficacy and can be considered for endodontic use. PMID:24778508

Automated palpation for breast tissue discrimination based on viscoelastic biomechanical properties.

PubMed

Tsukune, Mariko; Kobayashi, Yo; Miyashita, Tomoyuki; Fujie, G Masakatsu

2015-05-01

Accurate, noninvasive methods are sought for breast tumor detection and diagnosis. In particular, a need for noninvasive techniques that measure both the nonlinear elastic and viscoelastic properties of breast tissue has been identified. For diagnostic purposes, it is important to select a nonlinear viscoelastic model with a small number of parameters that highly correlate with histological structure. However, the combination of conventional viscoelastic models with nonlinear elastic models requires a large number of parameters. A nonlinear viscoelastic model of breast tissue based on a simple equation with few parameters was developed and tested. The nonlinear viscoelastic properties of soft tissues in porcine breast were measured experimentally using fresh ex vivo samples. Robotic palpation was used for measurements employed in a finite element model. These measurements were used to calculate nonlinear viscoelastic parameters for fat, fibroglandular breast parenchyma and muscle. The ability of these parameters to distinguish the tissue types was evaluated in a two-step statistical analysis that included Holm's pairwise [Formula: see text] test. The discrimination error rate of a set of parameters was evaluated by the Mahalanobis distance. Ex vivo testing in porcine breast revealed significant differences in the nonlinear viscoelastic parameters among combinations of three tissue types. The discrimination error rate was low among all tested combinations of three tissue types. Although tissue discrimination was not achieved using only a single nonlinear viscoelastic parameter, a set of four nonlinear viscoelastic parameters were able to reliably and accurately discriminate fat, breast fibroglandular tissue and muscle.

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.

Solvent Separating Secondary Metabolites Directly from Biosynthetic Tissue for Surface-Assisted Laser Desorption Ionisation Mass Spectrometry

PubMed Central

Rudd, David; Benkendorff, Kirsten; Voelcker, Nicolas H.

2015-01-01

Marine bioactive metabolites are often heterogeneously expressed in tissues both spatially and over time. Therefore, traditional solvent extraction methods benefit from an understanding of the in situ sites of biosynthesis and storage to deal with heterogeneity and maximize yield. Recently, surface-assisted mass spectrometry (MS) methods namely nanostructure-assisted laser desorption ionisation (NALDI) and desorption ionisation on porous silicon (DIOS) surfaces have been developed to enable the direct detection of low molecular weight metabolites. Since direct tissue NALDI-MS or DIOS-MS produce complex spectra due to the wide variety of other metabolites and fragments present in the low mass range, we report here the use of “on surface” solvent separation directly from mollusc tissue onto nanostructured surfaces for MS analysis, as a mechanism for simplifying data annotation and detecting possible artefacts from compound delocalization during the preparative steps. Water, ethanol, chloroform and hexane selectively extracted a range of choline esters, brominated indoles and lipids from Dicathais orbita hypobranchial tissue imprints. These compounds could be quantified on the nanostructured surfaces by comparison to standard curves generated from the pure compounds. Surface-assisted MS could have broad utility for detecting a broad range of secondary metabolites in complex marine tissue samples. PMID:25786067

Classification model based on Raman spectra of selected morphological and biochemical tissue constituents for identification of atherosclerosis in human coronary arteries.

PubMed

Peres, Marines Bertolo; Silveira, Landulfo; Zângaro, Renato Amaro; Pacheco, Marcos Tadeu Tavares; Pasqualucci, Carlos Augusto

2011-09-01

This study presents the results of Raman spectroscopy applied to the classification of arterial tissue based on a simplified model using basal morphological and biochemical information extracted from the Raman spectra of arteries. The Raman spectrograph uses an 830-nm diode laser, imaging spectrograph, and a CCD camera. A total of 111 Raman spectra from arterial fragments were used to develop the model, and those spectra were compared to the spectra of collagen, fat cells, smooth muscle cells, calcification, and cholesterol in a linear fit model. Non-atherosclerotic (NA), fatty and fibrous-fatty atherosclerotic plaques (A) and calcified (C) arteries exhibited different spectral signatures related to different morphological structures presented in each tissue type. Discriminant analysis based on Mahalanobis distance was employed to classify the tissue type with respect to the relative intensity of each compound. This model was subsequently tested prospectively in a set of 55 spectra. The simplified diagnostic model showed that cholesterol, collagen, and adipocytes were the tissue constituents that gave the best classification capability and that those changes were correlated to histopathology. The simplified model, using spectra obtained from a few tissue morphological and biochemical constituents, showed feasibility by using a small amount of variables, easily extracted from gross samples.

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

Quantitative Evaluation of Atherosclerotic Plaque Using Ultrasound Tissue Characterization.

NASA Astrophysics Data System (ADS)

Yigiter, Ersin

Evaluation of therapeutic methods directed toward interrupting and/or delaying atherogenesis is impeded by the lack of a reliable, non-invasive means for monitoring progression or regression of disease. The ability to characterize the predominant component of plaque may be very valuable in the study of this disease's natural history. The earlier the lesion, the more likely is lipid to be the predominant component. Progression of plaque is usually by way of overgrowth of fibrous tissues around the fatty pool. Calcification is usually a feature of the older or complicated lesion. To explore the feasibility of using ultrasound to characterize plaque we have conducted measurements of the acoustical properties of various atherosclerotic lesions found in freshly excised samples of human abdominal aorta. Our objective has been to determine whether or not the acoustical properties of plaque correlate with the type and/or chemical composition of plaque and, if so, to define a measurement scheme which could be done in-vivo and non-invasively. Our current data base consists of individual tissue samples from some 200 different aortas. Since each aorta yields between 10 to 30 tissue samples for study, we have data on some 4,468 different lesions or samples. Measurements of the acoustical properties of plaque were found to correlate well with the chemical composition of plaque. In short, measurements of impedance and attenuation seem sufficient to classify plaque as to type and to composition. Based on the in-vitro studies, the parameter of attenuation was selected as a means of classifying the plaque. For these measurements, an intravascular ultrasound scanner was modified according to our specifications. Signal processing algorithms were developed which would analyze the complex ultrasound waveforms and estimate tissue properties such as attenuation. Various methods were tried to estimate the attenuation from the pulse-echo backscattered signal. Best results were obtained by comparing averaged power spectra in small time windows at different depths for a series of A-lines. Comparisons between consequent averaged spectra at different depths provided the magnitude and frequency dependence of attenuation. Non-invasive characterization of the physical state of the tissue with quantitative ultrasound holds great promise for the extension of the diagnostic power of conventional B-mode imaging.

Selection of reference genes for quantitative gene expression normalization in flax (Linum usitatissimum L.).

PubMed

Huis, Rudy; Hawkins, Simon; Neutelings, Godfrey

2010-04-19

Quantitative real-time PCR (qRT-PCR) is currently the most accurate method for detecting differential gene expression. Such an approach depends on the identification of uniformly expressed 'housekeeping genes' (HKGs). Extensive transcriptomic data mining and experimental validation in different model plants have shown that the reliability of these endogenous controls can be influenced by the plant species, growth conditions and organs/tissues examined. It is therefore important to identify the best reference genes to use in each biological system before using qRT-PCR to investigate differential gene expression. In this paper we evaluate different candidate HKGs for developmental transcriptomic studies in the economically-important flax fiber- and oil-crop (Linum usitatissimum L). Specific primers were designed in order to quantify the expression levels of 20 different potential housekeeping genes in flax roots, internal- and external-stem tissues, leaves and flowers at different developmental stages. After calculations of PCR efficiencies, 13 HKGs were retained and their expression stabilities evaluated by the computer algorithms geNorm and NormFinder. According to geNorm, 2 Transcriptional Elongation Factors (TEFs) and 1 Ubiquitin gene are necessary for normalizing gene expression when all studied samples are considered. However, only 2 TEFs are required for normalizing expression in stem tissues. In contrast, NormFinder identified glyceraldehyde-3-phosphate dehydrogenase (GADPH) as the most stably expressed gene when all samples were grouped together, as well as when samples were classed into different sub-groups.qRT-PCR was then used to investigate the relative expression levels of two splice variants of the flax LuMYB1 gene (homologue of AtMYB59). LuMYB1-1 and LuMYB1-2 were highly expressed in the internal stem tissues as compared to outer stem tissues and other samples. This result was confirmed with both geNorm-designated- and NormFinder-designated-reference genes. The use of 2 different statistical algorithms results in the identification of different combinations of flax HKGs for expression data normalization. Despite such differences, the use of geNorm-designated- and NormFinder-designated-reference genes enabled us to accurately compare the expression levels of a flax MYB gene in different organs and tissues. Our identification and validation of suitable flax HKGs will facilitate future developmental transcriptomic studies in this economically-important plant.

Clonal evolution and resistance to EGFR blockade in the blood of colorectal cancer patients.

PubMed

Siravegna, Giulia; Mussolin, Benedetta; Buscarino, Michela; Corti, Giorgio; Cassingena, Andrea; Crisafulli, Giovanni; Ponzetti, Agostino; Cremolini, Chiara; Amatu, Alessio; Lauricella, Calogero; Lamba, Simona; Hobor, Sebastijan; Avallone, Antonio; Valtorta, Emanuele; Rospo, Giuseppe; Medico, Enzo; Motta, Valentina; Antoniotti, Carlotta; Tatangelo, Fabiana; Bellosillo, Beatriz; Veronese, Silvio; Budillon, Alfredo; Montagut, Clara; Racca, Patrizia; Marsoni, Silvia; Falcone, Alfredo; Corcoran, Ryan B; Di Nicolantonio, Federica; Loupakis, Fotios; Siena, Salvatore; Sartore-Bianchi, Andrea; Bardelli, Alberto

2015-07-01

Colorectal cancers (CRCs) evolve by a reiterative process of genetic diversification and clonal evolution. The molecular profile of CRC is routinely assessed in surgical or bioptic samples. Genotyping of CRC tissue has inherent limitations; a tissue sample represents a single snapshot in time, and it is subjected to spatial selection bias owing to tumor heterogeneity. Repeated tissue samples are difficult to obtain and cannot be used for dynamic monitoring of disease progression and response to therapy. We exploited circulating tumor DNA (ctDNA) to genotype colorectal tumors and track clonal evolution during treatment with the epidermal growth factor receptor (EGFR)-specific antibodies cetuximab or panitumumab. We identified alterations in ctDNA of patients with primary or acquired resistance to EGFR blockade in the following genes: KRAS, NRAS, MET, ERBB2, FLT3, EGFR and MAP2K1. Mutated KRAS clones, which emerge in blood during EGFR blockade, decline upon withdrawal of EGFR-specific antibodies, indicating that clonal evolution continues beyond clinical progression. Pharmacogenomic analysis of CRC cells that had acquired resistance to cetuximab reveals that upon antibody withdrawal KRAS clones decay, whereas the population regains drug sensitivity. ctDNA profiles of individuals who benefit from multiple challenges with anti-EGFR antibodies exhibit pulsatile levels of mutant KRAS. These results indicate that the CRC genome adapts dynamically to intermittent drug schedules and provide a molecular explanation for the efficacy of rechallenge therapies based on EGFR blockade.

Identification and typing of Brucella spp. in stranded harbour porpoises (Phocoena phocoena) on the Dutch coast.

PubMed

Maio, Elisa; Begeman, Lineke; Bisselink, Yvette; van Tulden, Peter; Wiersma, Lidewij; Hiemstra, Sjoukje; Ruuls, Robin; Gröne, Andrea; Roest, Hendrik-Ido-Jan; Willemsen, Peter; van der Giessen, Joke

2014-09-17

The presence of Brucella (B.) spp. in harbour porpoises stranded between 2008 and 2011 along the Dutch coast was studied. A selection of 265 tissue samples from 112 animals was analysed using conventional and molecular methods. In total, 4.5% (5/112) of the animals corresponding with 2.3% (6/265) Brucella positive tissue samples were Brucella positive by culture and these were all confirmed by real-time polymerase chain reaction (real-time PCR) based on the insertion element 711 (IS711). In addition, two more Brucella-positive tissue samples from two animals collected in 2011 were identified using real-time PCR resulting in an overall Brucella prevalence of 6.3% (7/112 animals). Brucella spp. were obtained from lungs (n=3), pulmonary lymph node (n=3) and lungworms (n=2). Multi Locus Variable Number of Tandem Repeats (VNTR) Analysis (MLVA) typing based on the MLVA-16 showed that the Brucella isolates were B. ceti. Additional in silico Multi Locus Sequence typing (MLST) after whole genome sequencing of the 6 Brucella isolates confirmed B. ceti ST 23. According to the Brucella 2010 MLVA database, the isolated Brucella strains encountered were of five genotypes, in two distinct subclusters divided in two different time periods of harbour porpoises collection. This study is the first population based analyses for Brucella spp. infections in cetaceans stranded along the Dutch coast. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

Nanomanipulation-coupled nanospray mass spectrometry as an approach for single cell analysis

NASA Astrophysics Data System (ADS)

Phelps, Mandy; Hamilton, Jason; Verbeck, Guido F.

2014-12-01

Electrospray mass spectrometry is now a widely used technique for observing cell content of various biological tissues. However, electrospray techniques (liquid chromatography and direct infusion) often involve lysing a group of cells and extracting the biomolecules of interest, rather than a sensitive, individual cell method to observe local chemistry. Presented here is an approach of combining a nanomanipulator workstation with nanospray mass spectrometry, which allows for extraction of a single cell, followed by rapid mass analysis that can provide a detailed metabolic profile. Triacylglycerol content was profiled with this tool coupled to mass spectrometry to investigate heterogeneity between healthy and tumorous tissues as well as lipid droplet containing adipocytes in vitro as proof of concept. This selective approach provides cellular resolution and complements existing bioanalytical techniques with minimal invasion to samples. In addition, the coupling of nanomanipulation and mass spectrometry holds the potential to be used in a great number of applications for individual organelles, diseased tissues, and in vitro cell cultures for observing heterogeneity even amongst cells and organelles of the same tissue.

Fluorescence of Picrosirius Red Multiplexed With Immunohistochemistry for the Quantitative Assessment of Collagen in Tissue Sections.

PubMed

Wegner, Kyle A; Keikhosravi, Adib; Eliceiri, Kevin W; Vezina, Chad M

2017-08-01

The low cost and simplicity of picrosirius red (PSR) staining have driven its popularity for collagen detection in tissue sections. We extended the versatility of this method by using fluorescent imaging to detect the PSR signal and applying automated quantification tools. We also developed the first PSR protocol that is fully compatible with multiplex immunostaining, making it possible to test whether collagen structure differs across immunohistochemically labeled regions of the tissue landscape. We compared our imaging method with two gold standards in collagen imaging, linear polarized light microscopy and second harmonic generation imaging, and found that it is at least as sensitive and robust to changes in sample orientation. As proof of principle, we used a genetic approach to overexpress beta catenin in a patchy subset of mouse prostate epithelial cells distinguished only by immunolabeling. We showed that collagen fiber length is significantly greater near beta catenin overexpressing cells than near control cells. Our fluorescent PSR imaging method is sensitive, reproducible, and offers a new way to guide region of interest selection for quantifying collagen in tissue sections.

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.

Evaluation of a gas chromatography method for azelaic acid determination in selected biological samples

PubMed Central

Garelnabi, Mahdi; Litvinov, Dmitry; Parthasarathy, Sampath

2010-01-01

Background: Azelaic acid (AzA) is the best known dicarboxilic acid to have pharmaceutical benefits and clinical applications and also to be associated with some diseases pathophysiology. Materials and Methods: We extracted and methylesterified AzA and determined its concentration in human plasma obtained from healthy individuals and also in mice fed AzA containing diet for three months. Results: AzA was detected in Gas Chromatography (GC) and confirmed by Liquid chromatography mass spectrometry (LCMS), and gas chromatography mass spectrometry (GCMC). Our results have shown that AzA can be determined efficiently in selected biological samples by GC method with 1nM limit of detection (LoD) and the limit of quantification (LoQ); was established at 50nM. Analytical Sensitivity as assayed by hexane demonstrated an analytical sensitivity at 0.050nM. The method has demonstrated 8-10% CV batch repeatability across the sample types and 13-18.9% CV for the Within-Lab Precision analysis. The method has shown that AzA can efficiently be recovered from various sample preparation including liver tissue homogenate (95%) and human plasma (97%). Conclusions: Because of its simplicity and lower limit of quantification, the present method provides a useful tool for determining AzA in various biological sample preparations. PMID:22558586

Evaluation of a gas chromatography method for azelaic acid determination in selected biological samples.

PubMed

Garelnabi, Mahdi; Litvinov, Dmitry; Parthasarathy, Sampath

2010-09-01

Azelaic acid (AzA) is the best known dicarboxilic acid to have pharmaceutical benefits and clinical applications and also to be associated with some diseases pathophysiology. We extracted and methylesterified AzA and determined its concentration in human plasma obtained from healthy individuals and also in mice fed AzA containing diet for three months. AzA was detected in Gas Chromatography (GC) and confirmed by Liquid chromatography mass spectrometry (LCMS), and gas chromatography mass spectrometry (GCMC). Our results have shown that AzA can be determined efficiently in selected biological samples by GC method with 1nM limit of detection (LoD) and the limit of quantification (LoQ); was established at 50nM. Analytical Sensitivity as assayed by hexane demonstrated an analytical sensitivity at 0.050nM. The method has demonstrated 8-10% CV batch repeatability across the sample types and 13-18.9% CV for the Within-Lab Precision analysis. The method has shown that AzA can efficiently be recovered from various sample preparation including liver tissue homogenate (95%) and human plasma (97%). Because of its simplicity and lower limit of quantification, the present method provides a useful tool for determining AzA in various biological sample preparations.

Immediate versus Delayed Treatment in the Anterior Maxilla Using Single Implants with a Laser-Microtextured Collar: 3-Year Results of a Case Series on Hard- and Soft-Tissue Response and Esthetics.

PubMed

Guarnieri, Renzo; Belleggia, Fabrizio; Grande, Maurizio

2016-02-01

To compare peri-implant marginal bone loss, soft tissue response, and esthetics following single immediate implant treatment (IIT) and delayed implant treatment (DIT) in the esthetic zone of the maxilla in well-selected patients. Adequate bone volume and ideal soft tissue level/contour were considered requirements for implant therapy, with additional prerequisites for IIT of residual alveolar bone wall integrity and a thick gingival biotype. IIT included immediate placement and provisionalization, while DIT included extraction socket preservation followed by implant placement and provisionalization 4 months later. Cortical bone levels and peri-implant mucosal conditions were evaluated at regular intervals. The esthetic outcome was objectively rated after 3 years using the pink esthetic score (PES) and white esthetic score (WES). Twelve patients received an immediate Laser-Lok® implant, and 13 patients received a delayed Laser-Lok® implant. No significant differences were found between the study groups regarding survival rate (100%). The mean bone level from the implant/abutment interface was 0.35 ± 0.18 mm for IIT and 0.42 ± 0.21 mm for DIT after 3 years (p > 0.05). Mesial and distal papillae remained stable over time in DIT. A tendency for regrowth of mesial and distal papillae was found following IIT (p < 0.05). Midfacial soft tissues remained stable over time following DIT and IIT. Within the limitations of this study (e.g., small sample size, short follow-up duration), the results suggest that regarding success rate, hard/soft tissue responses, and esthetics, DIT and IIT with single Laser-Lok® implants in the anterior maxilla are comparable and predictable options for well-selected patients. © 2015 by the American College of Prosthodontists.

Bioaccumulation of selected heavy metals and histopathological and hematobiochemical alterations in backyard chickens reared in an industrial area, India.

PubMed

Kar, Indrajit; Mukhopadhayay, Sunit Kumar; Patra, Amlan Kumar; Pradhan, Saktipada

2018-02-01

This study was carried out to determine the concentrations of four heavy metals, cadmium (Cd), lead (Pb), copper (Cu), and cobalt (Co), and histopathological lesions in tissues of chickens reared in an industrial area of West Bengal, India. In particular, Mejhia Block was selected as a heavy metal-exposed area and Vatar Block (120 km away from industrially polluted areas) as a reference site. Samples were collected from the backyard chickens in these areas. Concentrations of heavy metals in soil, water, feedstuffs, tissues (liver, kidney, spleen, lung, and muscle), and droppings were greater (p < 0.05) in the polluted site than in the reference site. The greatest tissue concentrations of Cd and Pb were noted in the kidney, followed by the liver, lung, spleen, and muscle, whereas Cu and Co concentrations accumulated in these tissues resulted in the following: liver > kidney > lung > spleen > muscle. Heavy metal concentrations were greater in older chickens (> 1 year) than in young chickens (< 1 year). Packed cell volume, total erythrocyte counts, hemoglobin, total protein, and total leucocyte counts were significantly (p < 0.01) lower in the blood picture of chickens in the exposed area. Different serum enzymes and biochemical profiles were altered due to heavy metal exposure. Histological changes revealed necrotic lesions and tubulitis in the kidney, degeneration and necrosis in liver parenchyma, and periarteriolitis, peribronchiolitis, and presence of hemosiderin pigment in the lung of chickens in the exposed site. Results indicated that backyard chickens in heavy metal-exposed site may show pathological lesions in different tissues due to accumulation of heavy metals, and thus, the consumption of chicken meat from the industrially exposed site may pose a potential health risk to local residents of polluted sites.

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.

Molecular discrimination of Echinococcus granulosus and Echinococcus multilocularis by sequencing and a new PCR-RFLP method with the potential use for other Echinococcus species.

PubMed

Şakalar, Çağrı; Kuk, Salih; Erensoy, Ahmet; Dağli, Adile Ferda; Özercan, İbrahim Hanifi; Çetınkaya, Ülfet; Yazar, Süleyman

2014-01-01

To develop a novel polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) protocol using a new genomic marker sequence and a novel set of restriction enzymes in order to detect and discriminate 2 Echinococcus species, E. granulosus and E. multilocularis, found in formalin-fixed paraffin-embedded (FFPE) human tissues. DNA was isolated from 11 FFPE human tissue samples positive for cystic echinococcosis or alveolar echinococcosis. A mitochondrial genomic marker region was amplified and sequenced using a novel primer pair and a new PCR-RFLP protocol was developed for the detection and discrimination of E. granulosus and E. multilocularis using a set of restriction enzymes including AccI, MboI, MboII, and TsoI. The selected marker region was amplified using DNA isolated from FFPE human tissue samples positive for cystic echinococcosis or alveolar echinococcosis and the discrimination of E. granulosus and E. multilocularis was accomplished by use of the novel PCR-RFLP method. In this PCR-RFLP protocol, use of any single restriction enzyme is enough for the discrimination of E. granulosus and E. multilocularis. The PCR-RFLP protocol can be potentially used for the discrimination of 5 other Echinococcus species: E. oligarthus, E. shiquicus, E. ortleppi, E. canadensis, and E. vogeli.

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.

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.

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.

Measurements of optical parameters of phantom solution and bulk animal tissues ex vivo at 650 nm

NASA Astrophysics Data System (ADS)

Sun, Ping; Wang, Yu; Liu, Jian

2008-12-01

Optical parameters of biological tissues, including absorption coefficient (μa), reduced scattering coefficient (μs') or scattering coefficient (μs), anisotropy factor (g) and refractive index (n) are investigated extensively and systemically at wavelength of 650 nm. Intralipid solution was selected to be the tissue phantom in order to test the validity of measurements. Considering the factors of fiber orientation and haemoglobin content, we chose some fresh bulk animal tissues in vitro which were bovine adipose, bovine muscle, porcine adipose, porcine muscle, porcine kidney, porcine liver, mutton and chicken breast. The basic assumption is that in vitro samples are a reasonable representation of the in vivo situation. We have gained numbers of experimental data of Intralipid and some tissues. Particularly, we have set up the close relationships among six optical parameters involving μa, μs', μs, g, n and μt. The experimental results show that for animal tissues, μa, μs' or μs and n rely deeply on muscle fiber orientations. Both of μs and μt range from 10mm-1 to 20mm-1. μa ranges from 10-2 mm-1 to 10-3 mm-1 and g from 0.95 to 0.99. The results of this study will be helpful in further understanding of optical properties of tissues.

Organochlorine compounds and trace elements in fish tissue and bed sediments in the lower Snake River basin, Idaho and Oregon

USGS Publications Warehouse

Clark, Gregory M.; Maret, Terry R.

1998-01-01

Fish-tissue and bed-sediment samples were collected to determine the occurrence and distribution of organochlorine compounds and trace elements in the lower Snake River Basin. Whole-body composite samples of suckers and carp from seven sites were analyzed for organochlorine compounds; liver samples were analyzed for trace elements. Fillets from selected sportfish were analyzed for organochlorine compounds and trace elements. Bed-sediment samples from three sites were analyzed for organochlorine compounds and trace elements. Twelve different organochlorine compounds were detected in 14 fish-tissue samples. All fish-tissue samples contained DDT or its metabolites. Concentrations of total DDT ranged from 11 micrograms per kilogram wet weight in fillets of yellow perch from C.J. Strike Reservoir to 3,633 micrograms per kilogram wet weight in a whole-body sample of carp from Brownlee Reservoir at Burnt River. Total DDT concentrations in whole-body samples of sucker and carp from the Snake River at C.J. Strike Reservoir, Snake River at Swan Falls, Snake River at Nyssa, and Brownlee Reservoir at Burnt River exceeded criteria established for the protection of fish-eating wildlife. Total PCB concentrations in a whole-body sample of carp from Brownlee Reservoir at Burnt River also exceeded fish-eating wildlife criteria. Concentrations of organochlorine compounds in whole-body samples, in general, were larger than concentrations in sportfish fillets. However, concentrations of dieldrin and total DDT in fillets of channel catfish from the Snake River at Nyssa and Brownlee Reservoir at Burnt River, and concentrations of total DDT in fillets of smallmouth bass and white crappie from Brownlee Reservoir at Burnt River exceeded a cancer risk screening value of 10-6 established by the U.S. Environmental Protection Agency. Concentrations of organochlorine compounds in bed sediment were smaller than concentrations in fish tissue. Concentrations of p,p'DDE, the only compound detected in all three bed-sediment samples, ranged from 1.1 micrograms per kilogram dry weight in C.J. Strike Reservoir to 11 micrograms per kilogram dry weight in Brownlee Reservoir at Burnt River. Data from this study, compared with data collected in the upper Snake River Basin from 1992 to 1994, indicates that, in general, organochlorine concentrations in fish tissue and bed sediment increased from the headwaters of the Snake River in Wyoming downstream to Brownlee Reservoir. The largest trace-element concentrations in fish tissue were in liver samples from carp from Brownlee Reservoir at Burnt River and suckers from the Boise River near Twin Springs. Concentrations of most trace elements were larger in livers than in the sport- fish fillets. However, mercury concentrations were generally larger in the sportfish fillets; they ranged from 0.08 microgram per gram wet weight in yellow perch from C.J. Strike Reservoir to 0.32 microgram per gram wet weight in channel catfish from Brownlee Reservoir at Burnt River. None of the trace-element concentrations in fillets exceeded median international standards or U.S. Food and Drug Administration action levels. Large trace-element concentrations in the upper Snake River Basin were reported in liver samples from suckers from headwater streams, probably a result of historical mining and weathering of metal-rich rocks. Concentrations of most trace elements in the bed-sediment samples were largest in Brownlee Reservoir at Mountain Man Lodge. Concentrations of arsenic, cadmium, chromium, copper, nickel, and zinc in bed sediment from the Mountain Man Lodge site exceeded either the threshold effect level or probable effect level established by the Canadian Government for the protection of benthic life. Arsenic, chromium, copper, and nickel concentrations in bed sediment from Brownlee Reservoir at Burnt River and chromium, copper, and nickel in bed sediment from C.J. Strike Reservoir also exceeded the threshold effect level.

[Analysis of effects of salt stress on absorption and accumulation of mineral elements in Elymus spp. using atomic absorption spectrophotometer].

PubMed

Jia, Ya-xiong; Sun, Lei; He, Feng; Wan, Li-qiang; Yuan, Qing-hua; Li, Xiang-lin

2008-12-01

Salinization contributes significantly to soil degradation and the growth and survival of plants. A high level of salts imposes both ionic and osmotic stresses on plants, resulting in an excessive accumulation of sodium (Na) in plant tissues. Na toxicity disrupts the uptake of soil nutrients. Plant uptake and absorption of macro-elements under salt stress have been studied in plants, but there is little literature addressing the effect of salt stress on plant accumulation and absorption of micro-elements. Species in Elymus genus are among the most important forage plants on high-salinity soils in China An experiment was conducted to study the effect of salt stress on accumulation and absorption of both macro- and micro-elements by wild plants of Elymus genus. Plant samples taken from two populations with different salt tolerance were tested and the level of 4 macro-elements, namely Na, K, Ca and Mg, and 4 micro-elements, namely Cu, Fe, Mn, Zn was determined using atomic absorption spectrophotometer. The relationship between the selection of elements in the process of absorption and accumulation and salt tolerance was also analyzed. The results showed that the level of Na in root and leaf tissues increased with increasing salt stress. The level of Na in leaf tissue of plants with high salt tolerance (HS) was significantly higher than that in plants with low salt tolerance (P<0.05). The level of K and Ca decreased in response to increasing salt stress, while that in HS was higher than in LS. The level of Fe and Zn in the tissues of both roots and leaves increased. No significant difference was detected between HS and LS samples in the level of Cu in root tissues, while that of Cu in leaf tissue of both samples increased. The level of Mn decreased with increasing salt stress, but was higher in HS than in LS. Fe and Zn in roots and leaves of HS were lower than in those of LS.

Socio-demographic, lifestyle, and dietary determinants of essential and possibly-essential trace element levels in adipose tissue from an adult cohort.

PubMed

Rodríguez-Pérez, Celia; Vrhovnik, Petra; González-Alzaga, Beatriz; Fernández, Mariana F; Martin-Olmedo, Piedad; Olea, Nicolás; Fiket, Željka; Kniewald, Goran; Arrebola, Juan P

2018-05-01

There is increasing evidence linking levels of trace elements (TEs) in adipose tissue with certain chronic conditions (e.g., diabetes or obesity). The objectives of this study were to assess concentrations of a selection of nine essential and possibly-essential TEs in adipose tissue samples from an adult cohort and to explore their socio-demographic, dietary, and lifestyle determinants. Adipose tissue samples were intraoperatively collected from 226 volunteers recruited in two public hospitals from Granada province. Trace elements (Co, Cr, Cu, Fe, Mn, Mo, Se, V, and Zn) were analyzed in adipose tissue by high-resolution inductively coupled plasma mass spectrometry (HR-ICP-MS). Data were collected on socio-demographic characteristics, lifestyle, diet, and health status by face-to-face interview. Predictors of TE concentrations were assessed by using multivariable linear and logistic regression. All TEs were detected in all samples with the exception of Se (53.50%). Iron, zinc, and copper showed the highest concentrations (42.60 mg/kg, 9.80 mg/kg, and 0.68 mg/kg, respectively). Diet was the main predictor of Cr, Fe, Mo, and Se concentrations. Body mass index was negatively associated with all TEs (β coefficients = -0.018 to -0.593, p = 0.001-0.090) except for Mn and V. Age showed a borderline-significant positive correlation with Cu (β = 0.004, p = 0.089). Residence in a rural or semi-rural area was associated with increased Co, Cr, Fe, Mo, Mn, V and Zn concentrations and with β coefficients ranging from 0.196 to 0.544 (p < 0.05). Furthermore, individuals with higher educational level showed increased Cr, Co, Fe and V concentrations (β coefficients = 0.276-0.368, p = 0.022-0.071). This is the first report on the distribution of these TEs in adipose tissue and on their determinants in a human cohort and might serve as an initial step in the elucidation of their clinical relevance. Copyright © 2017 Elsevier Ltd. All rights reserved.

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.

Identification of Specific miRNA Signature in Paired Sera and Tissue Samples of Indian Women with Triple Negative Breast Cancer

PubMed Central

Thakur, Seema; Grover, Rajesh K.; Gupta, Sanjay; Yadav, Ajay K.; Das, Bhudev C.

2016-01-01

Of several subtypes of breast cancer, triple negative breast cancer (TNBC) is a highly aggressive tumor that lacks expression of hormone receptors for estrogen, progesterone and human epidermal growth factor receptor 2 and shows a worst prognosis. The small noncoding RNAs (miRNAs) considered as master regulator of gene expression play a key role in cancer initiation, progression and drug resistance and have emerged as attractive molecular biomarkers for diagnosis, prognosis and treatment targets in cancer. We have done expression profiling of selected miRNAs in paired serum and tissue samples of TNBC patients and corresponding cell lines and compared with that of other subtypes, in order to identify novel serum miRNA biomarkers for early detection and progression of TNBC. A total of 85 paired tumor tissues and sera with an equal number of adjacent normal tissue margins and normal sera from age matched healthy women including tissue and sera samples from 15 benign fibroadenomas were employed for the study. We report for the first time an extremely high prevalence (73.9%) of TNBC in premenopausal women below 35 years of age and a significant altered expression of a panel of three specific oncogenic miRNAs- miR-21, miR-221, miR-210, and three tumor suppressor miRNAs- miR-195, miR-145 and Let-7a in both tissues and corresponding sera of TNBC patients when compared with triple positive breast cancer (TPBC) patients. While miR-21, miR-221 and miR-210 showed significant over-expression, miR-195 and miR-145 were downregulated and well correlated with various clinicopathological and demographic risk factors, tumor grade, clinical stage and hormone receptor status. Interestingly, despite being a known tumor suppressor, Let-7a showed a significant overexpression in TNBCs. It is suggested that this panel of six miRNA signature may serve as a minimally invasive biomarker for an early detection of TNBC patients. PMID:27404381

Multi-spectral brain tissue segmentation using automatically trained k-Nearest-Neighbor classification.

PubMed

Vrooman, Henri A; Cocosco, Chris A; van der Lijn, Fedde; Stokking, Rik; Ikram, M Arfan; Vernooij, Meike W; Breteler, Monique M B; Niessen, Wiro J

2007-08-01

Conventional k-Nearest-Neighbor (kNN) classification, which has been successfully applied to classify brain tissue in MR data, requires training on manually labeled subjects. This manual labeling is a laborious and time-consuming procedure. In this work, a new fully automated brain tissue classification procedure is presented, in which kNN training is automated. This is achieved by non-rigidly registering the MR data with a tissue probability atlas to automatically select training samples, followed by a post-processing step to keep the most reliable samples. The accuracy of the new method was compared to rigid registration-based training and to conventional kNN-based segmentation using training on manually labeled subjects for segmenting gray matter (GM), white matter (WM) and cerebrospinal fluid (CSF) in 12 data sets. Furthermore, for all classification methods, the performance was assessed when varying the free parameters. Finally, the robustness of the fully automated procedure was evaluated on 59 subjects. The automated training method using non-rigid registration with a tissue probability atlas was significantly more accurate than rigid registration. For both automated training using non-rigid registration and for the manually trained kNN classifier, the difference with the manual labeling by observers was not significantly larger than inter-observer variability for all tissue types. From the robustness study, it was clear that, given an appropriate brain atlas and optimal parameters, our new fully automated, non-rigid registration-based method gives accurate and robust segmentation results. A similarity index was used for comparison with manually trained kNN. The similarity indices were 0.93, 0.92 and 0.92, for CSF, GM and WM, respectively. It can be concluded that our fully automated method using non-rigid registration may replace manual segmentation, and thus that automated brain tissue segmentation without laborious manual training is feasible.

Seabird tissue archival and monitoring project: Protocol for collecting and banking seabird eggs

USGS Publications Warehouse

Weston-York, Geoff; Porter, Barbara J.; Pugh, Rebecca S.; Roseneau, David G.; Simac, Kristin S.; Becker, Paul R.; Thorsteinson, Lyman K.; Wise, Stephen A.

2001-01-01

Archiving biological and environmental samples for retrospective analysis is a major component of systematic environmental monitoring. The long-term storage of carefully selected, representative samples in an environmental specimen bank is an important complement to the real-time monitoring of the environment. These archived samples permit:The use of subsequently developed innovative analytical technology that was not available at the time the samples were archived, for clear state-of-art identification an~ quantification of analytes of interest,The identification and quantification of analytes that are of subsequent interest but that were not of interest at the time the samples were archived, andThe comparison of present and past analytical techniques and values, providing continued credibility of past analytical values, and allowing flexibility in environmental monitoring programs.Seabirds, including albatrosses, pelicans, cormorants, terns, kittiwakes, murres, guillemots, and puffins spend most of their lives at sea and have special adaptations for feeding in the marine environment, including the ability to excrete the excess salt obtained from ingesting seawater. Many species nest in dense groups (colonies) on steep, precipitous sea-cliffs and headlands.Seabirds are long-lived and slow to mature. They occupy high positions in the marine food web and are considered sensitive indicators for the marine environment (prey includes krill, small fish, and squid). Breeding success, timing of nesting, diets, and survival rates may provide early indications of changing environmental conditions (e.g., see Hatch et aI., 1993). Chemical analysis of seabird tissues, including egg contents, can be particularly useful in determining whether contaminants (and potential biological effects) associated with human industrial activities, such as offshore petroleum and mineral exploration and development, are accumulating in marine environments. The collection and archival of seabird tissues over a period of several years will be a resource for future analyses, providing samples that can be used to determine historical baseline contaminant levels.

[Values and limits in fine needle aspiration in the diagnosis of Warthin tumour of the parotid gland].

PubMed

Righi, A; Foschini, M P

2006-12-01

Recent paper state that cytological diagnosis of Warthin Tumour of the parotid gland is poorly reproducible and that aspiration procedures damage the tumoral tissue as to make difficult the histological diagnosis. Aim of the present study is to review cytological sampling performed with fine needle sampling without aspiration and to compare them with the relative surgical specimens in order to determine the value of fine needle aspiration in the diagnosis of Warthin Tumour and to quantify' the damage in histological tissue. All cases with cytological and/or histological diagnosis of Warthin Tumour during the period 1/1/98 to 30/6/04 were retrieved. In all cases both cytological and histological slides were reviewed and compared. In all cases fine needle aspiration (FNA) was performed according to the technique described by Zajdela et al. in 1987. A cytologic sample was considered diagnostic when there were in the same sample mucoid substance, lymphocytes scattered with a range of maturation and oncocytes organized in flat sheets of cells. 38 cases were selected corresponding to 37 patients (1 case of Warthin Tumour bilateral): all tumors were localized in the parotid gland. Fine needle sampling were 43. Nine cytological sample (20.9%) were inadequate. Sensitivity and specificity, in the preent series, had the same value, being 97.1%. A spectrum of histologic alterations were observed in 30/38 cases (79%). These alterations were squamous metaplasia (7 cases), acute and chronic hemorrhage and inflammation with multinucleated giant cells (26 cases) and granulation tissue with subsequent fibrosis (24 cases). In all cases the histologic alterations from fine needle were focal, limited to less than 20% of the tumoral area (first category of Batsakis 'classification), and did not prevent the histologic diagnosis of Warthin Tumour. These data suggest that fine needle aspiration without aspiration is a valuable tool for the per-operative diagnosis of Warthin Tumour. In addition it creates only minor histological changes, that always allow the histological diagnosis.

Mid-IR Spectral Investigation of Normal and Malignant Breast and Cervical Tissue Samples Using a Quantum Cascade Laser-Based Microscope

NASA Astrophysics Data System (ADS)

Haugen, Paul

Mid-infrared (MIR) spectroscopy has been a tool used to identify specific features of normal and malignant tissue samples by utilizing MIR characteristics, specifically in the "fingerprint" region. The fingerprint region is a biologically significant spectral region typically identified between 1500 and 500 cm-1. MIR spectroscopy can be used to study molecular changes and variations occurring in samples, which can then be used to fingerprint specific spectral characteristics and biomarkers in order to categorize the specimens. The most common instruments currently used in this analysis are Fourier transform infrared (FTIR) spectrometers, although properties inherent in these instruments, such as slow data collection time and an inability to specify sample location for the spectral data collection, have placed a ceiling on the clinical practicality of their use for specimen classification and identification. In this thesis, we use a prototype of an infrared hyperspectral imaging microscopy platform based around tunable quantum cascade laser (QCL) technology that has a spectral coverage from 1800-900 cm-1. The quantum cascade lasers are coupled with a series of MIR refractive objectives and an uncooled microbolometer camera. The speed of spectral imaging improves to 30 frames per second, and the high magnification objective has a 1.34 microm pixel resolution with a 0.70 numerical aperture and 4.3 microm spatial resolution. We are able to specify data collection at specific discrete wavelengths as opposed to the full spectrum, which improves the data collection time and de-clutters the data for analysis expediency. Finally, we perform spectral imaging real-time, which aides in selecting precise regions of interest on the target sample. This thesis demonstrates the advantages of exploiting the capabilities of the QCL microscope to advance MIR spectroscopy in the identification of distinguishing traits of normal and malignant breast and cervical tissue samples.

Raman spectroscopy differentiates squamous cell carcinoma (SCC) from normal skin following treatment with a high-powered CO2 laser.

PubMed

Fox, Sara A; Shanblatt, Ashley A; Beckman, Hugh; Strasswimmer, John; Terentis, Andrew C

2014-12-01

The number of cases of non-melanoma skin cancer (NMSC), which include squamous cell carcinoma (SCC) and basal cell carcinoma (BCC), continues to rise as the aging population grows. Mohs micrographic surgery has become the treatment of choice in many cases but is not always necessary or feasible. Ablation with a high-powered CO2 laser offers the advantage of highly precise, hemostatic tissue removal. However, confirmation of complete cancer removal following ablation is difficult. In this study we tested for the first time the feasibility of using Raman spectroscopy as an in situ diagnostic method to differentiate NMSC from normal tissue following partial ablation with a high-powered CO2 laser. Twenty-five tissue samples were obtained from eleven patients undergoing Mohs micrographic surgery to remove NMSC tumors. Laser treatment was performed with a SmartXide DOT Fractional CO2 Laser (DEKA Laser Technologies, Inc.) emitting a wavelength of 10.6 μm. Treatment levels ranged from 20 mJ to 1200 mJ total energy delivered per laser treatment spot (350 μm spot size). Raman spectra were collected from both untreated and CO2 laser-treated samples using a 785 nm diode laser. Principal Component Analysis (PCA) and Binary Logistic Regression (LR) were used to classify spectra as originating from either normal or NMSC tissue, and from treated or untreated tissue. Partial laser ablation did not adversely affect the ability of Raman spectroscopy to differentiate normal from cancerous residual tissue, with the spectral classification model correctly identifying SCC tissue with 95% sensitivity and 100% specificity following partial laser ablation, compared with 92% sensitivity and 60% selectivity for untreated NMSC tissue. The main biochemical difference identified between normal and NMSC tissue was high levels of collagen in the normal tissue, which was lacking in the NMSC tissue. The feasibility of a combined high-powered CO2 laser ablation, Raman diagnostic procedure for the treatment of NMSC is demonstrated since CO2 laser treatment does not hinder the ability of Raman spectroscopy to differentiate normal from diseased tissue. This combined approach could be employed clinically to greatly enhance the speed and effectiveness of NMSC treatment in many cases. © 2014 Wiley Periodicals, Inc.

Methods for the identification, characterization and banking of human DPSCs: current strategies and perspectives.

PubMed

Tirino, Virginia; Paino, Francesca; d'Aquino, Riccardo; Desiderio, Vincenzo; De Rosa, Alfredo; Papaccio, Gianpaolo

2011-09-01

Dental pulp stem cells (DPSCs), originating from neural crests, can be found within dental pulp. Up to now, it has been demonstrated that these cells are capable of producing bone tissue, both in vitro and in vivo and differentiate into adipocytes, endotheliocytes, melanocytes, neurons, glial cells, and can be easily cryopreserved and stored. Moreover, recent attention has been focused on tissue engineering and on the properties of these cells. In addition, adult bone tissue with good vascularisation has been obtained in grafts. The latest use in clinical trials for bone repair enforces the notion that DPSCs can be used successfully in patients. Therefore, their isolation, selection, differentiation and banking is of great importance. The isolation and detection techniques used in most laboratories are based on the use of antibodies revealed by flow-cytometers with cell sorter termed FACS (fluorescent activated cell sorter). In this report, we focus our attention on the main procedures used in the selection of DPSCs by flow cytometry, cell culture, freezing/thawing, cell cycle evaluation, histochemistry/immunofluorescence and differentiation of DPSCs. In addition, new methods/protocols to select and isolate stem cells without staining by fluorescent markers for implementation in biomedical/clinical laboratories are discuss. We emphasize that the new methods must address simplicity and short times of preparation and use of samples, complete sterility of cells, the potential disposable, low cost and complete maintenance of the viability and integrity of the cells with real-time response for subsequent applications in the biomedical/clinical/surgical fields.

Extraction of nucleic acids from yeast cells and plant tissues using ethanol as medium for sample preservation and cell disruption.

PubMed

Linke, Bettina; Schröder, Kersten; Arter, Juliane; Gasperazzo, Tatiana; Woehlecke, Holger; Ehwald, Rudolf

2010-09-01

Here we report that dehydrated ethanol is an excellent medium for both in situ preservation of nucleic acids and cell disruption of plant and yeast cells. Cell disruption was strongly facilitated by prior dehydration of the ethanol using dehydrated zeolite. Following removal of ethanol, nucleic acids were extracted from the homogenate pellet using denaturing buffers. The method provided DNA and RNA of high yield and integrity. Whereas cell wall disruption was essential for extraction of DNA and large RNA molecules, smaller molecules such as tRNAs could be selectively extracted from undisrupted, ethanol-treated yeast cells. Our results demonstrate the utility of absolute ethanol for sample fixation, cell membrane and cell wall disruption, as well as preservation of nucleic acids during sample storage.

Blind Biobanking of the Prostatectomy Specimen: Critical Evaluation of the Existing Techniques and Development of the New 4-Level Tissue Extraction Model With High Sampling Efficacy.

PubMed

Tolkach, Yuri; Eminaga, Okyaz; Wötzel, Fabian; Huss, Sebastian; Bettendorf, Olaf; Eltze, Elke; Abbas, Mahmoud; Imkamp, Florian; Semjonow, Axel

2017-03-01

Fresh tissue is mandatory to perform high-quality translation studies. Several models for tissue extraction from prostatectomy specimens without guidance by frozen sections are already introduced. However, little is known about the sampling efficacy of these models, which should provide representative tissue in adequate volumes, account for multifocality and heterogeneity of tumor, not violate the routine final pathological examination, and perform quickly without frozen section-based histological control. The aim of the study was to evaluate the sampling efficacy of the existing tissue extraction models without guidance by frozen sections ("blind") and to develop an optimized model for tissue extraction. Five hundred thirty-three electronic maps of the tumor distribution in prostates from a single-center cohort of the patients subjected to radical prostatectomy were used for analysis. Six available models were evaluated in silico for their sampling efficacy. Additionally, a novel model achieving the best sampling efficacy was developed. The available models showed high efficacies for sampling "any part" from the tumor (up to 100%), but were uniformly low in efficacy to sample all tumor foci from the specimens (with the best technique sampling only 51.6% of the all tumor foci). The novel 4-level extraction model achieved a sampling efficacy of 93.1% for all tumor foci. The existing "blind" tissue extraction models from prostatectomy specimens without frozen sections control are suitable to target tumor tissues but these tissues do not represent the whole tumor. The novel 4-level model provides the highest sampling efficacy and a promising potential for integration into routine. Prostate 77: 396-405, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Efficacy of Alkaline Hydrolysis as an Alternative Method for Treatment and Disposal of Infectious Animal Waste.

PubMed

Kaye, Gordon; Weber, Peter; Evans, Ann; Venezia, Richard

1998-05-01

The efficacy of alkaline hydrolysis as an alternative for incineration or autoclaving during treatment and disposal of infectious waste was evaluated by testing for the destruction of samples of pure cultures of selected infectious microorganisms during digestion of 114 to 136-kg loads of animal carcasses in an animal tissue digestor at the Albany Medical College. Ten milliliter samples of pure cultures of each microorganism were divided among 3 dialysis bags made from narrow diameter dialysis tubing, and each of these bags was placed inside another dialysis bag made from larger diameter dialysis tubing. Each double-bagged sample was suspended from the cover of the carcass basket of the tissue digestor so that it was completely covered by hot alkaline digestion solution during the carcass digestion process. The following organisms were required by the New York State Department of Health as representative pathogens for testing sterilization capabilities of the procedure: Staphylococcus aureus, Mycobacterium fortuitum, Candida albicans, Bacillus subtilis, Pseudomonas aeruginosa, Aspergillus fumigatus, Mycobacterium bovis BCG, MS-2 bacteriophage, and Giardia muris. Animal carcasses included pigs, sheep, rabbits, dogs, rats, mice, and guinea pigs. The tissue digestor was operated at 110 to 120 C and approximately 15 lb/in2 (gauge) for 18 h before the system was allowed to cool to 50 C and dialysis bags were retrieved and submitted for microbial culture. None of the samples obtained from the dialysis bags after the digestion process yielded indicator bacteria or yeast. Giardia cysts were completely destroyed; only small fragments of what appeared to be cyst wall could be recognized with light microscopic examination. No plaque-forming units were detected with MS-2 bacteriophage after digestion. Samples of the hydrolyzate also did not yield growth on culture media. Animal carcasses were completely solubilized and digested, with only the inorganic components of the bones and teeth remaining after draining and rinsing of the digestion vessel. Alkaline hydrolysis, as performed in this tissue digestor, completely destroyed all representative classes of potentially infectious agents as well as disposing of animal carcasses by solubilization and digestion.

Viscoelastic Properties of Human Tracheal Tissues.

PubMed

Safshekan, Farzaneh; Tafazzoli-Shadpour, Mohammad; Abdouss, Majid; Shadmehr, Mohammad B

2017-01-01

The physiological performance of trachea is highly dependent on its mechanical behavior, and therefore, the mechanical properties of its components. Mechanical characterization of trachea is key to succeed in new treatments such as tissue engineering, which requires the utilization of scaffolds which are mechanically compatible with the native human trachea. In this study, after isolating human trachea samples from brain-dead cases and proper storage, we assessed the viscoelastic properties of tracheal cartilage, smooth muscle, and connective tissue based on stress relaxation tests (at 5% and 10% strains for cartilage and 20%, 30%, and 40% for smooth muscle and connective tissue). After investigation of viscoelastic linearity, constitutive models including Prony series for linear viscoelasticity and quasi-linear viscoelastic, modified superposition, and Schapery models for nonlinear viscoelasticity were fitted to the experimental data to find the best model for each tissue. We also investigated the effect of age on the viscoelastic behavior of tracheal tissues. Based on the results, all three tissues exhibited a (nonsignificant) decrease in relaxation rate with increasing the strain, indicating viscoelastic nonlinearity which was most evident for cartilage and with the least effect for connective tissue. The three-term Prony model was selected for describing the linear viscoelasticity. Among different models, the modified superposition model was best able to capture the relaxation behavior of the three tracheal components. We observed a general (but not significant) stiffening of tracheal cartilage and connective tissue with aging. No change in the stress relaxation percentage with aging was observed. The results of this study may be useful in the design and fabrication of tracheal tissue engineering scaffolds.

Rapid electrochemical assessment of tumor suppressor gene methylations in raw human serum, and tumor cells and tissues using immuno-magnetic beads and selective DNA hybridization.

PubMed

Povedano, Eloy; Valverde, Alejandro; Ruiz-Valdepeñas Montiel, Víctor; Pedrero, María; Yáñez-Sedeño, Paloma; Barderas, Rodrigo; San Segundo-Acosta, Pablo; Peláez-García, Alberto; Mendiola, Marta; Hardisson, David; Campuzano, Susana; Pingarron, José Manuel

2018-05-09

We report a rapid and sensitive electrochemical strategy for the detection of gene-specific 5-methylcytosine DNA methylation. Magnetic beads (MBs) modified with an antibody specific for 5-methylcytosines (5-mC) are employed for the selective capture of any 5-mC methylated single-stranded (ss)DNA sequence. A flanking region next to the 5-mCs of the captured methylated ssDNA is recognized by selective hybridization with a synthetic biotinylated DNA sequence, further labeled with an HRP streptavidin conjugate. Amperometric transduction at disposable screen-printed carbon electrodes (SPCEs) is employed. The developed biosensor exhibits a dynamic range from 3.9 to 500 pM and a detection limit of 1.2 pM for the methylated synthetic sequence of the tumor suppressor gene O-6-methylguanine-DNA methyltransferase (MGMT) promoter region. The applicability of this strategy is demonstrated through the 45 min-analysis of specific methylation in the MGMT promoter region directly in raw spiked human serum samples and in genomic DNA extracted from U-87 glioblastoma cells and paraffin-embedded brain tumor tissues without any amplification and pretreatment step. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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.

Fatty Acid Composition of Muscle, Adipose Tissue and Liver from Muskoxen (Ovibos moschatus) Living in West Greenland

PubMed Central

Alves, Susana P.; Raundrup, Katrine; Cabo, Ângelo; Bessa, Rui J. B.; Almeida, André M.

2015-01-01

Information about lipid content and fatty acid (FA) composition of muskoxen (Ovibos moschatos) edible tissues is very limited in comparison to other meat sources. Thus, this work aims to present the first in-depth characterization of the FA profile of meat, subcutaneous adipose tissue and liver of muskoxen living in West Greenland. Furthermore, we aim to evaluate the effect of sex in the FA composition of these edible tissues. Samples from muscle (Longissimus dorsi), subcutaneous adipose tissue and liver were collected from female and male muskoxen, which were delivered at the butchery in Kangerlussuaq (West Greenland) during the winter hunting season. The lipid content of muscle, adipose tissue and liver averaged 284, 846 and 173 mg/g of dry tissue, respectively. This large lipid contents confirms that in late winter, when forage availability is scarce, muskoxen from West Greenland still have high fat reserves, demonstrating that they are well adapted to seasonal feed restriction. A detailed characterization of FA and dimethylacetal composition of muskoxen muscle, subcutaneous adipose tissue and liver showed that there are little differences on FA composition between sexes. Nevertheless, the 18:1cis-9 was the most abundant FA in muscle and adipose tissue, reaching 43% of total FA in muscle. The high content of 18:1cis-9 suggests that it can be selectively stored in muskoxen tissues. Regarding the nutritional composition of muskoxen edible tissues, they are not a good source of polyunsaturated FA; however, they may contribute to a higher fat intake. Information about the FA composition of muskoxen meat and liver is scarce, so this work can contribute to the characterization of the nutritional fat properties of muskoxen edible tissues and can be also useful to update food composition databases. PMID:26678792

Standardization and optimization of fluorescence in situ hybridization (FISH) for HER-2 assessment in breast cancer: A single center experience.

PubMed

Bogdanovska-Todorovska, Magdalena; Petrushevska, Gordana; Janevska, Vesna; Spasevska, Liljana; Kostadinova-Kunovska, Slavica

2018-05-20

Accurate assessment of human epidermal growth factor receptor 2 (HER-2) is crucial in selecting patients for targeted therapy. Commonly used methods for HER-2 testing are immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). Here we presented the implementation, optimization and standardization of two FISH protocols using breast cancer samples and assessed the impact of pre-analytical and analytical factors on HER-2 testing. Formalin fixed paraffin embedded (FFPE) tissue samples from 70 breast cancer patients were tested for HER-2 using PathVysion™ HER-2 DNA Probe Kit and two different paraffin pretreatment kits, Vysis/Abbott Paraffin Pretreatment Reagent Kit (40 samples) and DAKO Histology FISH Accessory Kit (30 samples). The concordance between FISH and IHC results was determined. Pre-analytical and analytical factors (i.e., fixation, baking, digestion, and post-hybridization washing) affected the efficiency and quality of hybridization. The overall hybridization success in our study was 98.6% (69/70); the failure rate was 1.4%. The DAKO pretreatment kit was more time-efficient and resulted in more uniform signals that were easier to interpret, compared to the Vysis/Abbott kit. The overall concordance between IHC and FISH was 84.06%, kappa coefficient 0.5976 (p < 0.0001). The greatest discordance (82%) between IHC and FISH was observed in IHC 2+ group. A standardized FISH protocol for HER-2 assessment, with high hybridization efficiency, is necessary due to variability in tissue processing and individual tissue characteristics. Differences in the pre-analytical and analytical steps can affect the hybridization quality and efficiency. The use of DAKO pretreatment kit is time-saving and cost-effective.

Hyperspectral interventional imaging for enhanced tissue visualization and discrimination combining band selection methods.

PubMed

Nouri, Dorra; Lucas, Yves; Treuillet, Sylvie

2016-12-01

Hyperspectral imaging is an emerging technology recently introduced in medical applications inasmuch as it provides a powerful tool for noninvasive tissue characterization. In this context, a new system was designed to be easily integrated in the operating room in order to detect anatomical tissues hardly noticed by the surgeon's naked eye. Our LCTF-based spectral imaging system is operative over visible, near- and middle-infrared spectral ranges (400-1700 nm). It is dedicated to enhance critical biological tissues such as the ureter and the facial nerve. We aim to find the best three relevant bands to create a RGB image to display during the intervention with maximal contrast between the target tissue and its surroundings. A comparative study is carried out between band selection methods and band transformation methods. Combined band selection methods are proposed. All methods are compared using different evaluation criteria. Experimental results show that the proposed combined band selection methods provide the best performance with rich information, high tissue separability and short computational time. These methods yield a significant discrimination between biological tissues. We developed a hyperspectral imaging system in order to enhance some biological tissue visualization. The proposed methods provided an acceptable trade-off between the evaluation criteria especially in SWIR spectral band that outperforms the naked eye's capacities.

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

Persistence of highly pathogenic avian influenza virus (H7N1) in infected chickens: feather as a suitable sample for diagnosis.

PubMed

Busquets, Núria; Abad, F Xavier; Alba, Anna; Dolz, Roser; Allepuz, Alberto; Rivas, Raquel; Ramis, Antonio; Darji, Ayub; Majó, Natàlia

2010-09-01

Selection of an ideal sample is a vital element in early detection of influenza infection. Rapid identification of infectious individuals or animals is crucial not only for avian influenza virus (AIV) surveillance programmes, but also for treatment and containment strategies. This study used a combination of quantitative real-time RT-PCR with an internal positive control and a cell-titration system to examine the presence of virus in different samples during active experimental AIV infection and its persistence in the infected carcasses. Oropharyngeal/cloacal swabs as well as feather pulp and blood samples were collected from 15-day-old chicks infected with H7N1 highly pathogenic AIV (HPAIV) and the kinetics of virus shedding during active infection were evaluated. Additionally, several samples (muscle, skin, brain, feather pulp and oropharyngeal and cloacal swabs) were examined to assess the persistence of virus in the HPAIV-infected carcasses. Based on the results, feather pulp was found to be the best sample to detect and isolate HPAIV from infected chicks from 24 h after inoculation onwards. Kinetic studies on the persistence of virus in infected carcasses revealed that tissues such as muscle could potentially transmit infectious virus for 3 days post-mortem (p.m.), whilst other tissues such as skin, feather pulp and brain retained their infectivity for as long as 5-6 days p.m. at environmental temperature (22-23 degrees C). These results strongly favour feather as a useful sample for HPAIV diagnosis in infected chickens as well as in carcasses.

Dual-view plane illumination microscopy for rapid and spatially isotropic imaging

PubMed Central

Kumar, Abhishek; Wu, Yicong; Christensen, Ryan; Chandris, Panagiotis; Gandler, William; McCreedy, Evan; Bokinsky, Alexandra; Colón-Ramos, Daniel A; Bao, Zhirong; McAuliffe, Matthew; Rondeau, Gary; Shroff, Hari

2015-01-01

We describe the construction and use of a compact dual-view inverted selective plane illumination microscope (diSPIM) for time-lapse volumetric (4D) imaging of living samples at subcellular resolution. Our protocol enables a biologist with some prior microscopy experience to assemble a diSPIM from commercially available parts, to align optics and test system performance, to prepare samples, and to control hardware and data processing with our software. Unlike existing light sheet microscopy protocols, our method does not require the sample to be embedded in agarose; instead, samples are prepared conventionally on glass coverslips. Tissue culture cells and Caenorhabditis elegans embryos are used as examples in this protocol; successful implementation of the protocol results in isotropic resolution and acquisition speeds up to several volumes per s on these samples. Assembling and verifying diSPIM performance takes ~6 d, sample preparation and data acquisition take up to 5 d and postprocessing takes 3–8 h, depending on the size of the data. PMID:25299154

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.

A highly efficient protocol for micropropagation of Begonia tuberous.

PubMed

Duong, Tan Nhut; Nguyen, Thanh Hai; Mai, Xuan Phan

2010-01-01

A protocol for micropropagation of begonia was established utilizing a thin cell layer (TCL) system. This system has been employed to produce several thousand shoots per sample. Explant size and position, and plant growth regulators (PGRs) contribute to the tissue morphogenesis. By optimizing the size of the tissue and applying an improved selection procedure, shoots were elongated in 8 weeks of culture, with an average number of 210 +/- 9.7 shoots per segment. This system has facilitated a number of studies using TCL as a model for micropropagation and will enable the large-scale production of begonia. On an average, the best treatment would allow production of about 10,000 plantlets by the micropropagation of the axillary buds of one plant with five petioles, within a period of 8 months.

Organic conjugated small molecule materials based optical probe for rapid, colorimetric and UV-vis spectral detection of phosphorylated protein in placental tissue

NASA Astrophysics Data System (ADS)

Wang, Yanfang; Yang, Na; Liu, Yi

2018-04-01

A novel organic small molecule with D-Pi-A structure was prepared, which was found to be a promising colorimetric and ratiometric UV-vis spetral probe for detection of phosphorylated proteins with the help of tetravalent zirconium ion. Such optical probe based on chromophore WYF-1 shows a rapid response (within 10 s) and high selectivity and sensitivity for phosphorylated proteins, giving distinct colorimetric and ratiometric UV-vis changes at 720 and 560 nm. The detection limit for phosphorylated proteins was estimated to be 100 nM. In addition, detection of phosphorylated proteins in placental tissue samples with this probe was successfully applied, which indicates that this probe holds great potential for phosphorylated proteins detection.

Increased Liposome Extravasation in Selected Tissues: Effect of Substance P

NASA Astrophysics Data System (ADS)

Rosenecker, Joseph; Zhang, Weiming; Hong, Keelung; Lausier, James; Geppetti, Pierangelo; Yoshihara, Shigemi; Papahadjopoulos, Demetrios; Nadel, Jay A.

1996-07-01

We have used a pharmacologic mediator to open intercellular connections in selected vessels to allow liposomes to escape from the blood stream and to extravasate into tissues that have appropriate receptors. We have examined the effects of substance P (SP), a peptide known to increase vascular permeability in selected tissues, such as trachea, esophagus, and urinary bladder in rats. We used quantitative fluorescence analysis of tissues to measure two fluorescent markers, one attached to the lipid (rhodamine-phosphatidylethanolamine) and another, doxorubicin (an antitumor drug), encapsulated within the aqueous interior. We have also examined the deposition of liposomes microscopically by the use of encapsulated colloidal gold and silver enhancement. Analysis of the biochemical and morphological observations indicate the following: (i) Injection of SP produces a striking increase in both liposome labels, but only in tissues that possess receptors for SP in postcapillary venules; (ii) liposome material in these tissues has extravasated and is found extracellularly near a variety of cells beyond the endothelial layer over the first few hours; (iii) 24 h following injection of liposomes and SP, liposome material is found in these tissues, localized intracellularly in both endothelial cells and macrophages. We propose that appropriate application of tissue-specific mediators can result in liposome extravasation deep within tissues that normally do not take up significant amounts of liposomes from the blood. Such liposomes are able to carry a variety of pharmacological agents that can be released locally within selected target tissues for therapeutic purposes.

Validation of reference genes for RT-qPCR studies of gene expression in banana fruit under different experimental conditions.

PubMed

Chen, Lei; Zhong, Hai-ying; Kuang, Jian-fei; Li, Jian-guo; Lu, Wang-jin; Chen, Jian-ye

2011-08-01

Reverse transcription quantitative real-time PCR (RT-qPCR) is a sensitive technique for quantifying gene expression, but its success depends on the stability of the reference gene(s) used for data normalization. Only a few studies on validation of reference genes have been conducted in fruit trees and none in banana yet. In the present work, 20 candidate reference genes were selected, and their expression stability in 144 banana samples were evaluated and analyzed using two algorithms, geNorm and NormFinder. The samples consisted of eight sample sets collected under different experimental conditions, including various tissues, developmental stages, postharvest ripening, stresses (chilling, high temperature, and pathogen), and hormone treatments. Our results showed that different suitable reference gene(s) or combination of reference genes for normalization should be selected depending on the experimental conditions. The RPS2 and UBQ2 genes were validated as the most suitable reference genes across all tested samples. More importantly, our data further showed that the widely used reference genes, ACT and GAPDH, were not the most suitable reference genes in many banana sample sets. In addition, the expression of MaEBF1, a gene of interest that plays an important role in regulating fruit ripening, under different experimental conditions was used to further confirm the validated reference genes. Taken together, our results provide guidelines for reference gene(s) selection under different experimental conditions and a foundation for more accurate and widespread use of RT-qPCR in banana.

The protective effect of different airway humidification liquids to lung after tracheotomy in traumatic brain injury: The role of pulmonary surfactant protein-A (SP-A).

PubMed

Su, Xinyang; Li, Zefu; Wang, Meilin; Li, Zhenzhu; Wang, Qingbo; Lu, Wenxian; Li, Xiaoli; Zhou, Youfei; Xu, Hongmei

2016-02-10

The purpose of this study was to establish a rat model of a brain injury with tracheotomy and compared the wetting effects of different airway humidification liquids, afterward, the best airway humidification liquid was selected for the clinical trial, thus providing a theoretical basis for selecting a proper airway humidification liquid in a clinical setting. Rats were divided into a sham group, group A (0.9% NaCl), group B (0.45% NaCl), group C (0.9% NaCl+ambroxol) and group D (0.9% NaCl+Pulmicort). An established rat model of traumatic brain injury with tracheotomy was used. Brain tissue samples were taken to determine water content, while lung tissue samples were taken to determine wet/dry weight ratio (W/D), histological changes and expression levels of SP-A mRNA and SP-A protein. 30 patients with brain injury and tracheotomy were selected and divided into two groups based on the airway humidification liquid instilled in the trachea tube, 0.45% NaCl and 0.9% NaCl+ambroxol. Blood was then extracted from the patients to measure the levels of SP-A, interleukin-6 (IL-6), interleukin-8 (IL-8) and tumour necrosis factor-α (TNF-α). The difference between group C and other groups in lung W/D and expression levels of SP-A mRNA and SP-A protein was significant (P<0.05). In comparison, the histological changes showed that the lung tissue damage was smallest in group C compared to the three other groups. Aspect of patients, 0.45% NaCl group and 0.9% NaCl+ambroxol group were significantly different in the levels of SP-A, IL-6, IL-8 and TNF-α (P<0.01). In the present study, 0.9% NaCl+ambroxol promote the synthesis and secretion of pulmonary surfactant, and has anti-inflammatory and antioxidant effects, which inhibit the release of inflammatory factors and cytokines, making it an ideal airway humidification liquid. Copyright © 2015 Elsevier B.V. All rights reserved.

Alteration of organic matter during infaunal polychaete gut passage and links to sediment organic geochemistry. Part II: Fatty acids and aldoses

NASA Astrophysics Data System (ADS)

Woulds, Clare; Middelburg, Jack J.; Cowie, Greg L.

2014-07-01

The activities of sediment-dwelling fauna are known to influence the rates of and pathways through which organic matter is cycled in marine sediments, and thus to influence eventual organic carbon burial or decay. However, due to methodological constraints, the role of faunal gut passage in determining the subsequent composition and thus degradability of organic matter is relatively little studied. Previous studies of organic matter digestion by benthic fauna have been unable to detect uptake and retention of specific biochemicals in faunal tissues, and have been of durations too short to fit digestion into the context of longer-term sedimentary degradation processes. Therefore this study aimed to investigate the aldose and fatty acid compositional alterations occurring to organic matter during gut passage by the abundant and ubiquitous polychaetes Hediste diversicolor and Arenicola marina, and to link these to longer-term changes typically observed during organic matter decay. This aim was approached through microcosm experiments in which selected polychaetes were fed with 13C-labelled algal detritus, and organisms, sediments, and faecal pellets were sampled at three timepoints over ∼6 weeks. Samples were analysed for their 13C-labelled aldose and fatty acid contents using GC-MS and GC-IRMS. Compound-selective net accumulation of biochemicals in polychaete tissues was observed for both aldoses and fatty acids, and the patterns of this were taxon-specific. The dominant patterns included an overall loss of glucose and polyunsaturated fatty acids; and preferential preservation or production of arabinose, microbial compounds (rhamnose, fucose and microbial fatty acids), and animal-synthesised fatty acids. These patterns may have been driven by fatty acid essentiality, preferential metabolism of glucose, and A. marina grazing on bacteria. Fatty acid suites in sediments from faunated microcosms showed greater proportions of saturated fatty acids and bacterial markers than those from afaunal controls. Aldose suite alterations were similar in faunated microcosms and afaunal controls, however the impact of faunal gut passage on sedimentary aldose compositions may be observable over longer timescales. Therefore this study provides direct evidence that polychaete gut passage influences OM composition both through taxon-specific selective assimilation and retention in polychaete tissues, and also through interactions with the microbial community. Polychaete gut passage will result in selective loss, preservation, and retention in polychaete tissues of specific aldoses and fatty acids. The pattern of selectivity will be taxon specific. Changes observed during gut passage will align with those commonly observed during OM decay, thus indicating that macrofaunal gut passage is one of the factors controlling sedimentary OM composition. Together with a previous publication reporting amino acid data from the same experiments (Woulds et al., 2012), this study represents the most complete description of OM alteration during gut passage that is available to date.

Tissue distribution of pretomanid in rat brain via mass spectrometry imaging.

PubMed

Shobo, Adeola; Bratkowska, Dominika; Baijnath, Sooraj; Naiker, Suhashni; Somboro, Anou M; Bester, Linda A; Singh, Sanil D; Naicker, Tricia; Kruger, Hendrik G; Govender, Thavendran

2016-01-01

1. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) combines the sensitivity and selectivity of mass spectrometry with spatial analysis to provide a new dimension for histological analyses of the distribution of drugs in tissue. Pretomanid is a pro-drug belonging to a class of antibiotics known as nitroimidizoles, which have been proven to be active under hypoxic conditions and to the best of our knowledge there have been no studies investigating the distribution and localisation of this class of compounds in the brain using MALDI MSI. 2. Herein, we report on the distribution of pretomanid in the healthy rat brain after intraperitoneal administration (20 mg/kg) using MALDI MSI. Our findings showed that the drug localises in specific compartments of the rat brain viz. the corpus callosum, a dense network of neurons connecting left and right cerebral hemispheres. 3. This study proves that MALDI MSI technique has great potential for mapping the pretomanid distribution in uninfected tissue samples, without the need for molecular labelling.

An Efficient Method for Genomic DNA Extraction from Different Molluscs Species

PubMed Central

Pereira, Jorge C.; Chaves, Raquel; Bastos, Estela; Leitão, Alexandra; Guedes-Pinto, Henrique

2011-01-01

The selection of a DNA extraction method is a critical step when subsequent analysis depends on the DNA quality and quantity. Unlike mammals, for which several capable DNA extraction methods have been developed, for molluscs the availability of optimized genomic DNA extraction protocols is clearly insufficient. Several aspects such as animal physiology, the type (e.g., adductor muscle or gills) or quantity of tissue, can explain the lack of efficiency (quality and yield) in molluscs genomic DNA extraction procedure. In an attempt to overcome these aspects, this work describes an efficient method for molluscs genomic DNA extraction that was tested in several species from different orders: Veneridae, Ostreidae, Anomiidae, Cardiidae (Bivalvia) and Muricidae (Gastropoda), with different weight sample tissues. The isolated DNA was of high molecular weight with high yield and purity, even with reduced quantities of tissue. Moreover, the genomic DNA isolated, demonstrated to be suitable for several downstream molecular techniques, such as PCR sequencing among others. PMID:22174651

Distribution of opiate alkaloids in brain tissue of experimental animals

PubMed Central

Pilija, Vladimir; Mimica-Dukic, Neda; Budakov, Branislav; Cvjeticanin, Stanko

2012-01-01

The present study examined regional distribution of opiate alkaloids from seized heroin in brain regions of experimental animals in order to select parts with the highest content of opiates. Their analysis should contribute to resolve causes of death due to heroin intake. The tests were performed at different time periods (5, 15, 45 and 120 min) after male and female Wistar rats were treated with seized heroin. Opiate alkaloids (codeine, morphine, acetylcodeine, 6-acetylmorphine and 3,6-diacetylmorphine) were quantitatively determined in brain regions known for their high concentration of µ-opiate receptors: cortex, brainstem, amygdala and basal ganglia, by using gas chromatography–mass spectrometry (GC–MS). The highest content of opiate alkaloids in the brain tissue of female animals was found 15 min and in male animals 45 min after treatment. The highest content of opiates was determined in the basal ganglia of the animals of both genders, indicating that this part of brain tissue presents a reliable sample for identifying and assessing contents of opiates after heroin intake. PMID:23554560

Fish assemblages

USGS Publications Warehouse

McGarvey, Daniel J.; Falke, Jeffrey A.; Li, Hiram W.; Li, Judith; Hauer, F. Richard; Lamberti, G.A.

2017-01-01

Methods to sample fishes in stream ecosystems and to analyze the raw data, focusing primarily on assemblage-level (all fish species combined) analyses, are presented in this chapter. We begin with guidance on sample site selection, permitting for fish collection, and information-gathering steps to be completed prior to conducting fieldwork. Basic sampling methods (visual surveying, electrofishing, and seining) are presented with specific instructions for estimating population sizes via visual, capture-recapture, and depletion surveys, in addition to new guidance on environmental DNA (eDNA) methods. Steps to process fish specimens in the field including the use of anesthesia and preservation of whole specimens or tissue samples (for genetic or stable isotope analysis) are also presented. Data analysis methods include characterization of size-structure within populations, estimation of species richness and diversity, and application of fish functional traits. We conclude with three advanced topics in assemblage-level analysis: multidimensional scaling (MDS), ecological networks, and loop analysis.

Dynamic landscape and regulation of RNA editing in mammals

PubMed Central

Tan, Meng How; Li, Qin; Shanmugam, Raghuvaran; Piskol, Robert; Kohler, Jennefer; Young, Amy N.; Liu, Kaiwen Ivy; Zhang, Rui; Ramaswami, Gokul; Ariyoshi, Kentaro; Gupte, Ankita; Keegan, Liam P.; George, Cyril X.; Ramu, Avinash; Huang, Ni; Pollina, Elizabeth A.; Leeman, Dena S.; Rustighi, Alessandra; Sharon Goh, Y. P.; Chawla, Ajay; Del Sal, Giannino; Peltz, Gary; Brunet, Anne; Conrad, Donald F.; Samuel, Charles E.; O’Connell, Mary A.; Walkley, Carl R.; Nishikura, Kazuko; Li, Jin Billy

2017-01-01

Adenosine-to-inosine (A-to-I) RNA editing is a conserved post-transcriptional mechanism mediated by ADAR enzymes that diversifies the transcriptome by altering selected nucleotides in RNA molecules1. Although many editing sites have recently been discovered2–7, the extent to which most sites are edited and how the editing is regulated in different biological contexts are not fully understood8–10. Here we report dynamic spatiotemporal patterns and new regulators of RNA editing, discovered through an extensive profiling of A-to-I RNA editing in 8,551 human samples (representing 53 body sites from 552 individuals) from the Genotype-Tissue Expression (GTEx) project and in hundreds of other primate and mouse samples. We show that editing levels in non-repetitive coding regions vary more between tissues than editing levels in repetitive regions. Globally, ADAR1 is the primary editor of repetitive sites and ADAR2 is the primary editor of non-repetitive coding sites, whereas the catalytically inactive ADAR3 predominantly acts as an inhibitor of editing. Cross-species analysis of RNA editing in several tissues revealed that species, rather than tissue type, is the primary determinant of editing levels, suggesting stronger cis-directed regulation of RNA editing for most sites, although the small set of conserved coding sites is under stronger trans-regulation. In addition, we curated an extensive set of ADAR1 and ADAR2 targets and showed that many editing sites display distinct tissue-specific regulation by the ADAR enzymes in vivo. Further analysis of the GTEx data revealed several potential regulators of editing, such as AIMP2, which reduces editing in muscles by enhancing the degradation of the ADAR proteins. Collectively, our work provides insights into the complex cis- and trans-regulation of A-to-I editing. PMID:29022589

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

Measuring temperature induced phase change kinetics in subcutaneous fatty tissues using near infrared spectroscopy, magnetic resonance imaging and optical coherence tomography (Conference Presentation)

NASA Astrophysics Data System (ADS)

Sajjadi, Amir Y.; Carp, Stefan A.; Manstein, Dieter

2017-02-01

Monitoring phase transition in adipose tissue and formation of lipid crystals is important in Cryo-procedures such as cryosurgery or Selective Cryolipolysis (SC). In this work, we exploited a Near-Infrared Spectroscopy (NIRS) method to monitor the onset of fat freezing/melting. Concurrent measurements using frequency domain NIRS and MR Spectroscopy during cooling/heating were performed on an in vitro porcine skin sample with a thick subcutaneous fat layer in a human MR scanner. The NIRS probe was placed on the skin measuring the average optical scattering of the fatty layer. Two fiber optic temperature probes were inserted in the area of the MRS and NIRS measurements. To further investigate the microscopic features of the phase-transition, an identical cooling/heating procedure was replicated on the same fat tissue while being imaged by Optical Coherence Tomography. The temperature relationships of optical scattering, MRS peak characteristics and OCT reflection intensity were analyzed to find signatures related to the onset of phase transition. The optical scattering in the fatty tissues decreases during the heating and increases by cooling. However, there is an inflexion in the rate of change of the scattering while the phase transition happens in the fatty layer. The methylene fat peaks on the MR Spectrum are also shown to be broadened during the cooling. OCT intensity displays a sharp increase at the transition temperature. The results from multiple samples show two transition points around 5-10 ˚C (cooling) and 15-20 ˚C (heating) through all three methods, demonstrating that adipose tissue phase change can be monitored non-invasively.

Dynamic landscape and regulation of RNA editing in mammals.

PubMed

Tan, Meng How; Li, Qin; Shanmugam, Raghuvaran; Piskol, Robert; Kohler, Jennefer; Young, Amy N; Liu, Kaiwen Ivy; Zhang, Rui; Ramaswami, Gokul; Ariyoshi, Kentaro; Gupte, Ankita; Keegan, Liam P; George, Cyril X; Ramu, Avinash; Huang, Ni; Pollina, Elizabeth A; Leeman, Dena S; Rustighi, Alessandra; Goh, Y P Sharon; Chawla, Ajay; Del Sal, Giannino; Peltz, Gary; Brunet, Anne; Conrad, Donald F; Samuel, Charles E; O'Connell, Mary A; Walkley, Carl R; Nishikura, Kazuko; Li, Jin Billy

2017-10-11

Adenosine-to-inosine (A-to-I) RNA editing is a conserved post-transcriptional mechanism mediated by ADAR enzymes that diversifies the transcriptome by altering selected nucleotides in RNA molecules. Although many editing sites have recently been discovered, the extent to which most sites are edited and how the editing is regulated in different biological contexts are not fully understood. Here we report dynamic spatiotemporal patterns and new regulators of RNA editing, discovered through an extensive profiling of A-to-I RNA editing in 8,551 human samples (representing 53 body sites from 552 individuals) from the Genotype-Tissue Expression (GTEx) project and in hundreds of other primate and mouse samples. We show that editing levels in non-repetitive coding regions vary more between tissues than editing levels in repetitive regions. Globally, ADAR1 is the primary editor of repetitive sites and ADAR2 is the primary editor of non-repetitive coding sites, whereas the catalytically inactive ADAR3 predominantly acts as an inhibitor of editing. Cross-species analysis of RNA editing in several tissues revealed that species, rather than tissue type, is the primary determinant of editing levels, suggesting stronger cis-directed regulation of RNA editing for most sites, although the small set of conserved coding sites is under stronger trans-regulation. In addition, we curated an extensive set of ADAR1 and ADAR2 targets and showed that many editing sites display distinct tissue-specific regulation by the ADAR enzymes in vivo. Further analysis of the GTEx data revealed several potential regulators of editing, such as AIMP2, which reduces editing in muscles by enhancing the degradation of the ADAR proteins. Collectively, our work provides insights into the complex cis- and trans-regulation of A-to-I editing.

New methods to improve the safety assessment of cryopreserved ovarian tissue for fertility preservation in breast cancer patients.

PubMed

Rodríguez-Iglesias, Beatriz; Novella-Maestre, Edurne; Herraiz, Sonia; Díaz-García, César; Pellicer, Nuria; Pellicer, Antonio

2015-12-01

To develop a novel molecular panel of markers to detect breast cancer (BC) disseminated malignant cells in ovarian tissue, and to improve the safety of ovarian tissue transplantation. Experimental study. University hospital. Ten ovarian biopsies from healthy patients, 13 biopsies with diagnosed BC metastasis, and 4 biopsies from primary BC tumor for designing a diagnostic panel of BC cell contamination; 60 ovarian biopsies from BC patients undergoing fertility preservation for validating the panel. Female nude mice. A novel panel for BC malignant cell detection by reverse-transcription polymerase chain reaction (RT-PCR), inmmunohistochemical analysis, in vitro invasion assay and xenotransplantation assayed in ovarian tissue from BC patients. Expression of GCDFP15, MGB1, SBEM, MUC1, WT-1, and NY-BR-01, selected as markers, assessed by quantitative RT-PCR in samples with confirmed BC metastasis. The most sensitive markers were confirmed by immunohistochemistry, and tested in vitro and in vivo. GCDFP15, MGB1, and SBEM were the most sensitive and specific markers to detect BC metastatic cells when at least one was expressed by quantitative RT-PCR. The panel was validated in 60 patients and confirmed in an in vitro invasion assay, where no invasive cells were observed. Samples negative for BC cells cannot develop disease when xenografted. GCDFP15, MGB1, and SBEM were the most sensitive molecules to create a diagnostic panel for BC malignant cell contamination, which may make ovarian tissue cryopreservation and transplantation a safe technique for fertility preservation in BC patients. Copyright © 2015 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Determination of diagnostic standards on saturated soil extracts for cut roses grown in greenhouses.

PubMed

Franco-Hermida, John Jairo; Quintero, María Fernanda; Cabrera, Raúl Iskander; Guzman, José Miguel

2017-01-01

This work comprises the theoretical determination and validation of diagnostic standards for the analysis of saturated soil extracts for cut rose flower crops (Rosa spp.) growing in the Bogota Plateau, Colombia. The data included 684 plant tissue analyses and 684 corresponding analyses of saturated soil extracts, all collected between January 2009 and June 2013. The tissue and soil samples were selected from 13 rose farms, and from cultivars grafted on the 'Natal Briar' rootstock. These concurrent samples of soil and plant tissues represented 251 production units (locations) of approximately 10,000 m2 distributed across the study area. The standards were conceived as a tool to improve the nutritional balance in the leaf tissue of rose plants and thereby define the norms for expressing optimum productive potential relative to nutritional conditions in the soil. To this end, previously determined diagnostic standard for rose leaf tissues were employed to obtain rates of foliar nutritional balance at each analyzed location and as criteria for determining the diagnostic norms for saturated soil extracts. Implementing this methodology to foliar analysis, showed a higher significant correlation for diagnostic indices. A similar behavior was observed in saturated soil extracts analysis, becoming a powerful tool for integrated nutritional diagnosis. Leaf analyses determine the most limiting nutrients for high yield and analyses of saturated soil extracts facilitate the possibility of correcting the fertigation formulations applied to soils or substrates. Recommendations are proposed to improve the balance in soil-plant system with which the possibility of yield increase becomes more probable. The main recommendations to increase and improve rose crop flower yields would be: continuously check pH values of SSE, reduce the amounts of P, Fe, Zn and Cu in fertigation solutions and carefully analyze the situation of Mn in the soil-plant system.

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.

Three Minute Method for Amino Acid Analysis by UHPLC and high resolution quadrupole orbitrap mass spectrometry

PubMed Central

Nemkov, Travis; D'Alessandro, Angelo; Hansen, Kirk C.

2015-01-01

Amino acid analysis is a powerful bioanalytical technique for many biomedical research endeavors, including cancer, emergency medicine, nutrition and neuroscience research. In the present study, we present a three minute analytical method for underivatized amino acid analysis that employs ultra-high performance liquid chromatography and high resolution quadrupole orbitrap mass spectrometry. This method has demonstrated linearity (mM to nM range), reproducibility (intra-day<5%, inter-day<20%), sensitivity (low fmol) and selectivity. Here, we illustrate the rapidity and accuracy of the method through comparison with conventional liquid chromatography-mass spectrometry methods. We further demonstrate the robustness and sensitivity of this method on a diverse range of biological matrices. Using this method we were able to selectively discriminate murine pancreatic cancer cells with and without knocked down expression of Hypoxia Inducible Factor 1α; plasma, lymph and bronchioalveolar lavage fluid samples from control versus hemorrhaged rats; and muscle tissue samples harvested from rats subjected to both low fat and high fat diets. Furthermore, we were able to exploit the sensitivity of the method to detect and quantify the release of glutamate from sparsely isolated murine taste buds. Spiked in light or heavy standards (13C6-arginine, 13C6-lysine, 13C515N2-glutamine) or xenometabolites were used to determine coefficient of variations, confirm linearity of relative quantitation in four different matrices, and overcome matrix effects for absolute quantitation. The presented method enables high-throughput analysis of low abundance samples requiring only one percent of the material extracted from 100,000 cells, 10 μl of biological fluid, or 2 mg of muscle tissue. PMID:26058356

Boron delivery with liposomes for boron neutron capture therapy (BNCT): biodistribution studies in an experimental model of oral cancer demonstrating therapeutic potential.

PubMed

Heber, Elisa M; Kueffer, Peter J; Lee, Mark W; Hawthorne, M Frederick; Garabalino, Marcela A; Molinari, Ana J; Nigg, David W; Bauer, William; Hughes, Andrea Monti; Pozzi, Emiliano C C; Trivillin, Verónica A; Schwint, Amanda E

2012-05-01

Boron neutron capture therapy (BNCT) combines selective accumulation of (10)B carriers in tumor tissue with subsequent neutron irradiation. We previously demonstrated the therapeutic efficacy of BNCT in the hamster cheek pouch oral cancer model. Optimization of BNCT depends largely on improving boron targeting to tumor cells. Seeking to maximize the potential of BNCT for the treatment for head and neck cancer, the aim of the present study was to perform boron biodistribution studies in the oral cancer model employing two different liposome formulations that were previously tested for a different pathology, i.e., in experimental mammary carcinoma in BALB/c mice: (1) MAC: liposomes incorporating K[nido-7-CH(3)(CH(2))(15)-7,8-C(2)B(9)H(11)] in the bilayer membrane and encapsulating a hypertonic buffer, administered intravenously at 6 mg B per kg body weight, and (2) MAC-TAC: liposomes incorporating K[nido-7-CH(3)(CH(2))(15)-7,8-C(2)B(9)H(11)] in the bilayer membrane and encapsulating a concentrated aqueous solution of the hydrophilic species Na(3) [ae-B(20)H(17)NH(3)], administered intravenously at 18 mg B per kg body weight. Samples of tumor, precancerous and normal pouch tissue, spleen, liver, kidney, and blood were taken at different times post-administration and processed to measure boron content by inductively coupled plasma mass spectrometry. No ostensible clinical toxic effects were observed with the selected formulations. Both MAC and MAC-TAC delivered boron selectively to tumor tissue. Absolute tumor values for MAC-TAC peaked to 66.6 ± 16.1 ppm at 48 h and to 43.9 ± 17.6 ppm at 54 h with very favorable ratios of tumor boron relative to precancerous and normal tissue, making these protocols particularly worthy of radiobiological assessment. Boron concentration values obtained would result in therapeutic BNCT doses in tumor without exceeding radiotolerance in precancerous/normal tissue at the thermal neutron facility at RA-3.

Boron delivery with liposomes for boron neutron capture therapy (BNCT): biodistribution studies in an experimental model of oral cancer demonstrating therapeutic potential

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

David W. Nigg

2012-05-01

Boron neutron capture therapy (BNCT) combines selective accumulation of 10B carriers in tumor tissue with subsequent neutron irradiation. We previously demonstrated the therapeutic efficacy of BNCT in the hamster cheek pouch oral cancer model. Optimization of BNCT depends largely on improving boron targeting to tumor cells. Seeking to maximize the potential of BNCT for the treatment for head and neck cancer, the aim of the present study was to perform boron biodistribution studies in the oral cancer model employing two different liposome formulations that were previously tested for a different pathology, i.e., in experimental mammary carcinoma in BALB/c mice: (1)more » MAC: liposomes incorporating K[nido-7-CH3(CH2)15-7,8-C2B9H11] in the bilayer membrane and encapsulating a hypertonic buffer, administered intravenously at 6 mg B per kg body weight, and (2) MAC-TAC: liposomes incorporating K[nido-7-CH3(CH2)15-7,8-C2B9H11] in the bilayer membrane and encapsulating a concentrated aqueous solution of the hydrophilic species Na3 [ae-B20H17NH3], administered intravenously at 18 mg B per kg body weight. Samples of tumor, precancerous and normal pouch tissue, spleen, liver, kidney, and blood were taken at different times post-administration and processed to measure boron content by inductively coupled plasma mass spectrometry. No ostensible clinical toxic effects were observed with the selected formulations. Both MAC and MAC-TAC delivered boron selectively to tumor tissue. Absolute tumor values for MAC-TAC peaked to 66.6 {+-} 16.1 ppm at 48 h and to 43.9 {+-} 17.6 ppm at 54 h with very favorable ratios of tumor boron relative to precancerous and normal tissue, making these protocols particularly worthy of radiobiological assessment. Boron concentration values obtained would result in therapeutic BNCT doses in tumor without exceeding radiotolerance in precancerous/normal tissue at the thermal neutron facility at RA-3.« less

A workflow to preserve genome-quality tissue samples from plants in botanical gardens and arboreta1

PubMed Central

Gostel, Morgan R.; Kelloff, Carol; Wallick, Kyle; Funk, Vicki A.

2016-01-01

Premise of the study: 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. Methods and Results: 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. Conclusions: 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. PMID:27672517

A Hybrid Hierarchical Approach for Brain Tissue Segmentation by Combining Brain Atlas and Least Square Support Vector Machine

PubMed Central

Kasiri, Keyvan; Kazemi, Kamran; Dehghani, Mohammad Javad; Helfroush, Mohammad Sadegh

2013-01-01

In this paper, we present a new semi-automatic brain tissue segmentation method based on a hybrid hierarchical approach that combines a brain atlas as a priori information and a least-square support vector machine (LS-SVM). The method consists of three steps. In the first two steps, the skull is removed and the cerebrospinal fluid (CSF) is extracted. These two steps are performed using the toolbox FMRIB's automated segmentation tool integrated in the FSL software (FSL-FAST) developed in Oxford Centre for functional MRI of the brain (FMRIB). Then, in the third step, the LS-SVM is used to segment grey matter (GM) and white matter (WM). The training samples for LS-SVM are selected from the registered brain atlas. The voxel intensities and spatial positions are selected as the two feature groups for training and test. SVM as a powerful discriminator is able to handle nonlinear classification problems; however, it cannot provide posterior probability. Thus, we use a sigmoid function to map the SVM output into probabilities. The proposed method is used to segment CSF, GM and WM from the simulated magnetic resonance imaging (MRI) using Brainweb MRI simulator and real data provided by Internet Brain Segmentation Repository. The semi-automatically segmented brain tissues were evaluated by comparing to the corresponding ground truth. The Dice and Jaccard similarity coefficients, sensitivity and specificity were calculated for the quantitative validation of the results. The quantitative results show that the proposed method segments brain tissues accurately with respect to corresponding ground truth. PMID:24696800

Target and Tissue Selectivity Prediction by Integrated Mechanistic Pharmacokinetic-Target Binding and Quantitative Structure Activity Modeling.

PubMed

Vlot, Anna H C; de Witte, Wilhelmus E A; Danhof, Meindert; van der Graaf, Piet H; van Westen, Gerard J P; de Lange, Elizabeth C M

2017-12-04

Selectivity is an important attribute of effective and safe drugs, and prediction of in vivo target and tissue selectivity would likely improve drug development success rates. However, a lack of understanding of the underlying (pharmacological) mechanisms and availability of directly applicable predictive methods complicates the prediction of selectivity. We explore the value of combining physiologically based pharmacokinetic (PBPK) modeling with quantitative structure-activity relationship (QSAR) modeling to predict the influence of the target dissociation constant (K D ) and the target dissociation rate constant on target and tissue selectivity. The K D values of CB1 ligands in the ChEMBL database are predicted by QSAR random forest (RF) modeling for the CB1 receptor and known off-targets (TRPV1, mGlu5, 5-HT1a). Of these CB1 ligands, rimonabant, CP-55940, and Δ 8 -tetrahydrocanabinol, one of the active ingredients of cannabis, were selected for simulations of target occupancy for CB1, TRPV1, mGlu5, and 5-HT1a in three brain regions, to illustrate the principles of the combined PBPK-QSAR modeling. Our combined PBPK and target binding modeling demonstrated that the optimal values of the K D and k off for target and tissue selectivity were dependent on target concentration and tissue distribution kinetics. Interestingly, if the target concentration is high and the perfusion of the target site is low, the optimal K D value is often not the lowest K D value, suggesting that optimization towards high drug-target affinity can decrease the benefit-risk ratio. The presented integrative structure-pharmacokinetic-pharmacodynamic modeling provides an improved understanding of tissue and target selectivity.

Identification of tissue-specific targeting peptide

NASA Astrophysics Data System (ADS)

Jung, Eunkyoung; Lee, Nam Kyung; Kang, Sang-Kee; Choi, Seung-Hoon; Kim, Daejin; Park, Kisoo; Choi, Kihang; Choi, Yun-Jaie; Jung, Dong Hyun

2012-11-01

Using phage display technique, we identified tissue-targeting peptide sets that recognize specific tissues (bone-marrow dendritic cell, kidney, liver, lung, spleen and visceral adipose tissue). In order to rapidly evaluate tissue-specific targeting peptides, we performed machine learning studies for predicting the tissue-specific targeting activity of peptides on the basis of peptide sequence information using four machine learning models and isolated the groups of peptides capable of mediating selective targeting to specific tissues. As a representative liver-specific targeting sequence, the peptide "DKNLQLH" was selected by the sequence similarity analysis. This peptide has a high degree of homology with protein ligands which can interact with corresponding membrane counterparts. We anticipate that our models will be applicable to the prediction of tissue-specific targeting peptides which can recognize the endothelial markers of target tissues.

Chlordane, DDT, PCB's, and other selected organic compounds in Asiatic clams and yellow bullhead in the Potomac River Basin, 1992

USGS Publications Warehouse

Zappia, Humbert

1996-01-01

Chlordane, DDT (dichlor-diphenyl-trichloroethane), and PCB's (polychlorinated biphenyls) were the most widespread organic contaminants detected during a 1992 survey of aquatic biological tissues in the Potomac River Basin. On the basis of existing U.S. Food and Drug Administration criteria, no new threats to human health were discovered, although chlordane concentrations may pose a threat to fish-eating wildlife. Chlordane exceeded the National Academy of Science and National Academy of Engineering recommended maximum concentration for the protection of fish-eating wildlife at two sites. The survey, conducted by the U.S. Geological Survey's National Water-Quality Assessment Program, sampled Asiatic clams (Corbicula fluminea) and yellow bullhead (Ameiurus natalis) at 16 sites to determine the occurrence and distribution of 29 hydrophobic organic compounds. Thirteen of these organic compounds were detected in the survey. Sites with the greatest number of compounds detected include the Potomac River near Alexandria, Va., with 6 compounds detected in Asiatic clam tissue, and Accotink Creek near Annandale, Va., with 11 compounds in yellow bullhead tissue. Chlordane was detected at six sites, with maximum concentrations of 31.1 ?g/kg (micrograms per kilograms) in Asiatic clam tissue and 127 ?g/kg in yellow bullhead whole-fish tissue. DDT was detected at five sites, with maximum concentrations of 12.9 ?g/kg in Asiatic clam tissue and 7.6 ?g/kg in yellow bullhead whole-fish tissue. PCB's were detected at nine sites, with maximum concentrations of 162 ?g/kg in Asiatic clam tissue and 146 mg/ kg in yellow bullhead whole-fish tissue.

The system spatial-frequency filtering of birefringence images of human blood layers

NASA Astrophysics Data System (ADS)

Ushenko, A. G.; Boychuk, T. M.; Mincer, O. P.; Angelsky, P. O.; Bodnar, N. B.; Oleinichenko, B. P.; Bizer, L. I.

2013-09-01

Among various opticophysical methods [1 - 3] of diagnosing the structure and properties of the optical anisotropic component of various biological objects a specific trend has been singled out - multidimensional laser polarimetry of microscopic images of the biological tissues with the following statistic, correlative and fractal analysis of the coordinate distributions of the azimuths and ellipticity of polarization in approximating of linear birefringence polycrystalline protein networks [4 - 10]. At the same time, in most cases, experimental obtaining of tissue sample is a traumatic biopsy operation. In addition, the mechanisms of transformation of the state of polarization of laser radiation by means of the opticoanisotropic biological structures are more varied (optical dichroism, circular birefringence). Hereat, real polycrystalline networks can be formed by different types, both in size and optical properties of biological crystals. Finally, much more accessible for an experimental investigation are biological fluids such as blood, bile, urine, and others. Thus, further progress of laser polarimetry can be associated with the development of new methods of analysis and processing (selection) of polarization- heterogeneous images of biological tissues and fluids, taking into account a wider set of mechanisms anisotropic mechanisms. Our research is aimed at developing experimental method of the Fourier polarimetry and a spatialfrequency selection for distributions of the azimuth and the ellipticity polarization of blood plasma laser images with a view of diagnosing prostate cancer.

Monitoring of trace metals in tissues of Wallago attu (lanchi) from the Indus River as an indicator of environmental pollution

PubMed Central

Al-Ghanim, K.A.; Mahboob, Shahid; Seemab, Sadia; Sultana, S.; Sultana, T.; Al-Misned, Fahad; Ahmed, Z.

2015-01-01

We aimed to assess the bioaccumulation of selected four trace metals (Cd, Ni, Zn and Co) in four tissues (muscles, skin, gills and liver) of a freshwater fish Wallago attu (lanchi) from three different sites (upstream, middle stream and downstream) of the Indus River in Mianwali district of Pakistan. Heavy metal contents in water samples and from different selected tissues of fish were examined by using flame atomic absorption spectrometry. The data were statistically compared to study the effects of the site and fish organs and their interaction on the bioaccumulation pattern of these metals at P < 0.05. In W. attu the level of cadmium ranged from 0.004 to 0.24; nickel 0.003–0.708; cobalt 0.002–0.768 and zinc 47.4–1147.5 μg/g wet weight. The magnitude of metal bioaccumulation in different organs of fish species had the following order gills > liver > skin > muscle. The order of bioaccumulation of these metals was Ni < Zn < Co < Cd. Heavy metal concentrations were increased during the dry season as compared to the wet season. The results of this study indicate that freshwater fish produced and marketed in Mianwali have concentrations below the standards of FEPA/WHO for these toxic metals. PMID:26858541

Monitoring of trace metals in tissues of Wallago attu (lanchi) from the Indus River as an indicator of environmental pollution.

PubMed

Al-Ghanim, K A; Mahboob, Shahid; Seemab, Sadia; Sultana, S; Sultana, T; Al-Misned, Fahad; Ahmed, Z

2016-01-01

We aimed to assess the bioaccumulation of selected four trace metals (Cd, Ni, Zn and Co) in four tissues (muscles, skin, gills and liver) of a freshwater fish Wallago attu (lanchi) from three different sites (upstream, middle stream and downstream) of the Indus River in Mianwali district of Pakistan. Heavy metal contents in water samples and from different selected tissues of fish were examined by using flame atomic absorption spectrometry. The data were statistically compared to study the effects of the site and fish organs and their interaction on the bioaccumulation pattern of these metals at P < 0.05. In W. attu the level of cadmium ranged from 0.004 to 0.24; nickel 0.003-0.708; cobalt 0.002-0.768 and zinc 47.4-1147.5 μg/g wet weight. The magnitude of metal bioaccumulation in different organs of fish species had the following order gills > liver > skin > muscle. The order of bioaccumulation of these metals was Ni < Zn < Co < Cd. Heavy metal concentrations were increased during the dry season as compared to the wet season. The results of this study indicate that freshwater fish produced and marketed in Mianwali have concentrations below the standards of FEPA/WHO for these toxic metals.

Effects of pulsed CO2 laser in caries selective ablation

NASA Astrophysics Data System (ADS)

Colojoara, Carmen; David, Ion; Marinovici, Mariana

1995-03-01

We have evaluated the effect of pulsed carbon dioxide laser in the treatment for deep carious decay. The so called `caries profonda' is still a problem for conservative dentistry. A `Valvfivre' Master 20S carbon dioxide laser was pulsed to determine the effects on dentine and for testing the properties of softened dentine in selective ablation. Laser treatment parameters were from 1 to 2 W, 50 to 150 ms, 200 to 320 Hz. Fifteen human teeth samples were exposed to irradiation: extracted third molar were exposed to CO2 pulsed laser to determine in vitro the effects on pulp morphology. The tissue samples were analyzed histologically and by means of scanning electron microscopy for evidence of thermal damage. Next, we have evaluated the morphologic changes in vivo on 10 cases in patients with deep carious decay. Pulsed infrared lasers are capable of inducing physical and chemical changes in dentine structure. The results showed an artificially sclerosing and micro-hardness on the remaining dentine. CO2 laser can vaporized carious dentine.

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

The Nature of the Dietary Protein Impacts the Tissue-to-Diet 15N Discrimination Factors in Laboratory Rats

PubMed Central

Poupin, Nathalie; Bos, Cécile; Mariotti, François; Huneau, Jean-François; Tomé, Daniel; Fouillet, Hélène

2011-01-01

Due to the existence of isotope effects on some metabolic pathways of amino acid and protein metabolism, animal tissues are 15N-enriched relative to their dietary nitrogen sources and this 15N enrichment varies among different tissues and metabolic pools. The magnitude of the tissue-to-diet discrimination (Δ15N) has also been shown to depend on dietary factors. Since dietary protein sources affect amino acid and protein metabolism, we hypothesized that they would impact this discrimination factor, with selective effects at the tissue level. To test this hypothesis, we investigated in rats the influence of a milk or soy protein-based diet on Δ15N in various nitrogen fractions (urea, protein and non-protein fractions) of blood and tissues, focusing on visceral tissues. Regardless of the diet, the different protein fractions of blood and tissues were generally 15N-enriched relative to their non-protein fraction and to the diet (Δ15N>0), with large variations in the Δ15N between tissue proteins. Δ15N values were markedly lower in tissue proteins of rats fed milk proteins compared to those fed soy proteins, in all sampled tissues except in the intestine, and the amplitude of Δ15N differences between diets differed between tissues. Both between-tissue and between-diet Δ15N differences are probably related to modulations of the relative orientation of dietary and endogenous amino acids in the different metabolic pathways. More specifically, the smaller Δ15N values observed in tissue proteins with milk than soy dietary protein may be due to a slightly more direct channeling of dietary amino acids for tissue protein renewal and to a lower recycling of amino acids through fractionating pathways. In conclusion, the present data indicate that natural Δ15N of tissue are sensitive markers of the specific subtle regional modifications of the protein and amino acid metabolism induced by the protein dietary source. PMID:22132207

Veterinary Medicine and Multi-Omics Research for Future Nutrition Targets: Metabolomics and Transcriptomics of the Common Degenerative Mitral Valve Disease in Dogs.

PubMed

Li, Qinghong; Freeman, Lisa M; Rush, John E; Huggins, Gordon S; Kennedy, Adam D; Labuda, Jeffrey A; Laflamme, Dorothy P; Hannah, Steven S

2015-08-01

Canine degenerative mitral valve disease (DMVD) is the most common form of heart disease in dogs. The objective of this study was to identify cellular and metabolic pathways that play a role in DMVD by performing metabolomics and transcriptomics analyses on serum and tissue (mitral valve and left ventricle) samples previously collected from dogs with DMVD or healthy hearts. Gas or liquid chromatography followed by mass spectrophotometry were used to identify metabolites in serum. Transcriptomics analysis of tissue samples was completed using RNA-seq, and selected targets were confirmed by RT-qPCR. Random Forest analysis was used to classify the metabolites that best predicted the presence of DMVD. Results identified 41 known and 13 unknown serum metabolites that were significantly different between healthy and DMVD dogs, representing alterations in fat and glucose energy metabolism, oxidative stress, and other pathways. The three metabolites with the greatest single effect in the Random Forest analysis were γ-glutamylmethionine, oxidized glutathione, and asymmetric dimethylarginine. Transcriptomics analysis identified 812 differentially expressed transcripts in left ventricle samples and 263 in mitral valve samples, representing changes in energy metabolism, antioxidant function, nitric oxide signaling, and extracellular matrix homeostasis pathways. Many of the identified alterations may benefit from nutritional or medical management. Our study provides evidence of the growing importance of integrative approaches in multi-omics research in veterinary and nutritional sciences.

Development and Testing of a Method for Validating Chemical Inactivation of Ebola Virus.

PubMed

Alfson, Kendra J; Griffiths, Anthony

2018-03-13

Complete inactivation of infectious Ebola virus (EBOV) is required before a sample may be removed from a Biosafety Level 4 laboratory. The United States Federal Select Agent Program regulations require that procedures used to demonstrate chemical inactivation must be validated in-house to confirm complete inactivation. The objective of this study was to develop a method for validating chemical inactivation of EBOV and then demonstrate the effectiveness of several commonly-used inactivation methods. Samples containing infectious EBOV ( Zaire ebolavirus ) in different matrices were treated, and the sample was diluted to limit the cytopathic effect of the inactivant. The presence of infectious virus was determined by assessing the cytopathic effect in Vero E6 cells. Crucially, this method did not result in a loss of infectivity in control samples, and we were able to detect less than five infectious units of EBOV ( Zaire ebolavirus ). We found that TRIzol LS reagent and RNA-Bee inactivated EBOV in serum; TRIzol LS reagent inactivated EBOV in clarified cell culture media; TRIzol reagent inactivated EBOV in tissue and infected Vero E6 cells; 10% neutral buffered formalin inactivated EBOV in tissue; and osmium tetroxide vapors inactivated EBOV on transmission electron microscopy grids. The methods described herein are easily performed and can be adapted to validate inactivation of viruses in various matrices and by various chemical methods.

Wavelength selection-based nonlinear calibration for transcutaneous blood glucose sensing using Raman spectroscopy

PubMed Central

Dingari, Narahara Chari; Barman, Ishan; Kang, Jeon Woong; Kong, Chae-Ryon; Dasari, Ramachandra R.; Feld, Michael S.

2011-01-01

While Raman spectroscopy provides a powerful tool for noninvasive and real time diagnostics of biological samples, its translation to the clinical setting has been impeded by the lack of robustness of spectroscopic calibration models and the size and cumbersome nature of conventional laboratory Raman systems. Linear multivariate calibration models employing full spectrum analysis are often misled by spurious correlations, such as system drift and covariations among constituents. In addition, such calibration schemes are prone to overfitting, especially in the presence of external interferences that may create nonlinearities in the spectra-concentration relationship. To address both of these issues we incorporate residue error plot-based wavelength selection and nonlinear support vector regression (SVR). Wavelength selection is used to eliminate uninformative regions of the spectrum, while SVR is used to model the curved effects such as those created by tissue turbidity and temperature fluctuations. Using glucose detection in tissue phantoms as a representative example, we show that even a substantial reduction in the number of wavelengths analyzed using SVR lead to calibration models of equivalent prediction accuracy as linear full spectrum analysis. Further, with clinical datasets obtained from human subject studies, we also demonstrate the prospective applicability of the selected wavelength subsets without sacrificing prediction accuracy, which has extensive implications for calibration maintenance and transfer. Additionally, such wavelength selection could substantially reduce the collection time of serial Raman acquisition systems. Given the reduced footprint of serial Raman systems in relation to conventional dispersive Raman spectrometers, we anticipate that the incorporation of wavelength selection in such hardware designs will enhance the possibility of miniaturized clinical systems for disease diagnosis in the near future. PMID:21895336

Methods to measure target site penetration of antibiotics in critically ill patients.

PubMed

Schwameis, Richard; Zeitlinger, Markus

2013-02-01

While several tools are necessary to repair a car, the engineer knows exactly which instrument he has to utilize at different parts of the broken machine. Likewise, depending on the information we are interested in, we have to choose different tools to investigate and consecutively understand the multiple aspects that are involved in pharmacokinetics of antimicrobial agents in critically ill patients. Some techniques, like blood sampling, microdialysis or positrons emission tomography (PET) will allow for obtaining continues concentration time profiles while others like bronchoalveolar lavage (BAL), biopsy or surgical tissue samples can only be used a limited number of times per subject. PET and methods based on tissue homogenization will deliver an average of the actual concentrations in intra - and extracellular compartments while investigations in isolated blood cells or microdialysis allow for more distinguished allocation of a concentration to a defined compartment. The present review aims at discussing the advantages and disadvantages of the various methods used for assessing pharmacokinetics in critically ill patients with regard to specific aspects of pharmacokinetic research and further reviews data of selected antibiotics as examples for applications of the individual techniques.

The value of intraoperative ultrasonography during the resection of relapsed irradiated malignant gliomas in the brain.

PubMed

Mursch, Kay; Scholz, Martin; Brück, Wolfgang; Behnke-Mursch, Julianne

2017-01-01

The aim of this study was to investigate whether intraoperative ultrasonography (IOUS) helped the surgeon navigate towards the tumor as seen in preoperative magnetic resonance imaging and whether IOUS was able to distinguish between tumor margins and the surrounding tissue. Twenty-five patients suffering from high-grade gliomas who were previously treated by surgery and radiotherapy were included. Intraoperatively, two histopathologic samples were obtained a sample of unequivocal tumor tissue (according to anatomical landmarks and the surgeon's visual and tactile impressions) and a small tissue sample obtained using a navigated needle when the surgeon decided to stop the resection. This specimen was considered to be a boundary specimen, where no tumor tissue was apparent. The decision to take the second sample was not influenced by IOUS. The effect of IOUS was analyzed semi-quantitatively. All 25 samples of unequivocal tumor tissue were histopathologically classified as tumor tissue and were hyperechoic on IOUS. Of the boundary specimens, eight were hypoechoic. Only one harbored tumor tissue (P=0.150). Seventeen boundaries were moderately hyperechoic, and these samples contained all possible histological results (i.e., tumor, infiltration, or no tumor). During surgery performed on relapsed, irradiated, high-grade gliomas, IOUS provided a reliable method of navigating towards the core of the tumor. At borders, it did not reliably distinguish between remnants or tumor-free tissue, but hypoechoic areas seldom contained tumor tissue.

Genetic identification of Iberian rodent species using both mitochondrial and nuclear loci: application to noninvasive sampling.

PubMed

Barbosa, S; Pauperio, J; Searle, J B; Alves, P C

2013-01-01

Species identification through noninvasive sampling is increasingly used in animal conservation genetics, given that it obviates the need to handle free-living individuals. Noninvasive sampling is particularly valuable for elusive and small species such as rodents. Although rodents are not usually assumed to be the most obvious target for conservation, of the 21 species or near-species present in Iberia, three are considered endangered and declining, while several others are poorly studied. Here, we develop a genetic tool for identifying all rodent species in Iberia by noninvasive genetic sampling. To achieve this purpose, we selected one mitochondrial gene [cytochrome b (cyt-b)] and one nuclear gene [interphotoreceptor retinoid-binding protein (IRBP)], which we first sequenced using tissue samples. Both genes allow for the phylogenetic distinction of all species except the sibling species Microtus lusitanicus and Microtus duodecimcostatus. Overall, cyt-b showed higher resolution than IRBP, revealing a clear barcoding gap. To allow these markers to be applied to noninvasive samples, we selected a short highly diagnostic fragment from each gene, which we used to obtain sequences from faeces and bones from owl pellets. Amplification success for the cyt-b and IRBP fragment was 85% and 43% in faecal and 88% and 64% in owl-pellet DNA extractions, respectively. The method allows the unambiguous identification of the great majority of Iberian rodent species from noninvasive samples, with application in studies of distribution, spatial ecology and population dynamics, and for conservation. © 2012 Blackwell Publishing Ltd.

5-ALA-induced fluorescence as a marker for diagnostic tissue in stereotactic biopsies of intracranial lymphomas: experience in 41 patients.

PubMed

Kiesel, Barbara; Millesi, Matthias; Woehrer, Adelheid; Furtner, Julia; Bavand, Anahita; Roetzer, Thomas; Mischkulnig, Mario; Wolfsberger, Stefan; Preusser, Matthias; Knosp, Engelbert; Widhalm, Georg

2018-06-01

OBJECTIVE Stereotactic needle biopsies are usually performed for histopathological confirmation of intracranial lymphomas to guide adequate treatment. During biopsy, intraoperative histopathology is an effective tool to avoid acquisition of nondiagnostic samples. In the last years, 5-aminolevulinic acid (5-ALA)-induced fluorescence has been increasingly used for visualization of diagnostic brain tumor tissue during stereotactic biopsies. Recently, visible fluorescence was reported in the first cases of intracranial lymphomas as well. The aim of this study is thus to investigate the technical and clinical utility of 5-ALA-induced fluorescence in a large series of stereotactic biopsies for intracranial lymphoma. METHODS This prospective study recruited adult patients who underwent frameless stereotactic needle biopsy for a radiologically suspected intracranial lymphoma after oral 5-ALA administration. During biopsy, samples from the tumor region were collected for histopathological analysis, and presence of fluorescence (strong, vague, or no fluorescence) was assessed with a modified neurosurgical microscope. In tumors with available biopsy samples from at least 2 different regions the intratumoral fluorescence homogeneity was additionally investigated. Furthermore, the influence of potential preoperative corticosteroid treatment or immunosuppression on fluorescence was analyzed. Histopathological tumor diagnosis was established and all collected biopsy samples were screened for diagnostic lymphoma tissue. RESULTS The final study cohort included 41 patients with intracranial lymphoma. Stereotactic biopsies with assistance of 5-ALA were technically feasible in all cases. Strong fluorescence was found as maximum level in 30 patients (75%), vague fluorescence in 2 patients (4%), and no visible fluorescence in 9 patients (21%). In 28 cases, samples were obtained from at least 2 different tumor regions; homogenous intratumoral fluorescence was found in 16 of those cases (57%) and inhomogeneous intratumoral fluorescence in 12 (43%). According to histopathological analysis, all samples with strong or vague fluorescence contained diagnostic lymphoma tissue, resulting in a positive predictive value of 100%. Analysis showed no influence of preoperative corticosteroids or immunosuppression on fluorescence. CONCLUSIONS The data obtained in this study demonstrate the technical and clinical utility of 5-ALA-induced fluorescence in stereotactic biopsies of intracranial lymphomas. Thus, 5-ALA can serve as a useful tool to select patients not requiring intraoperative histopathology, and its application should markedly reduce operation time and related costs in the future.

A probable risk factor of female breast cancer: study on benign and malignant breast tissue samples.

PubMed

Rehman, Sohaila; Husnain, Syed M

2014-01-01

The study reports enhanced Fe, Cu, and Zn contents in breast tissues, a probable risk factor of breast cancer in females. Forty-one formalin-fixed breast tissues were analyzed using atomic absorption spectrophotometry. Twenty malignant, six adjacent to malignant and 15 benign tissues samples were investigated. The malignant tissues samples were of grade 11 and type invasive ductal carcinoma. The quantitative comparison between the elemental levels measured in the two types of specimen (benign and malignant) tissues (removed after surgery) suggests significant elevation of these metals (Fe, Cu, and Zn) in the malignant tissue. The specimens were collected just after mastectomy of women aged 19 to 59 years from the hospitals of Islamabad and Rawalpindi, Pakistan. Most of the patients belong to urban areas of Pakistan. Findings of study depict that these elements have a promising role in the initiation and development of carcinoma as consistent pattern of elevation for Fe, Cu, and Zn was observed. The results showed the excessive accumulation of Fe (229 ± 121 mg/L) in malignant breast tissue samples of patients (p < 0.05) to that in benign tissues samples (49.1 ± 11.4 mg/L). Findings indicated that excess accumulation of iron in malignant tissues can be a risk factor of breast cancer. In order to validate our method of analysis, certified reference material muscle tissue lyophilized (IAEA) MA-M-2/TM was analyzed for metal studied. Determined concentrations were quite in good agreement with certified levels. Asymmetric concentration distribution for Fe, Cu, and Zn was observed in both malignant and benign tissue samples.

A pre-clinical pharmacokinetic study in rats of three naturally occurring iridoid glycosides, Picroside-I, II and III, using a validated simultaneous HPLC-MS/MS assay.

PubMed

Zhu, Jianwei; Xue, Bingyang; Ma, Bo; Zhang, Qi; Liu, Ming; Liu, Lei; Yao, Di; Qi, Huanhuan; Wang, Yonglu; Ying, Hanjie; Wu, Zimei

2015-07-01

A selective and sensitive high-performance liquid chromatography-electro-spray ionization tandem mass spectrometry (LC-ESI-MS/MS) method was developed for the simultaneous quantitative determination of Picroside-I, II, and III in rat plasma and tissue homogenate to aid the pre-clinical studies. The chromatographic separation was performed on a Hypersil GOLD AQ C18 column using a gradient elution program with a mobile phase consisting of 2mM ammonium acetate and acetonitrile. The detection was achieved using a triple quadrupole tandem MS in negative ionization multiple reaction monitoring (MRM) mode. One-step protein precipitation was selected for plasma and tissue sample preparation while liquid-liquid extraction failed to achieve satisfactory recoveries. The calibration curves of all three analytes in either plasma or tissue homogenate showed good linearity over the concentration range of 0.5-500ng/mL with a limit of quantitation at 0.5ng/mL. Both the intra- and inter-day accuracy and precision were within ±10%. The extraction recoveries were >70%, and the relative matrix effect ranged from 80.4% to 107.4% in all the biological samples. All the analytes were stable in matrices for at least 24h at room temperature, or 21 days in frozen. Three freeze/thaw cycles did not cause degradation. The method was successfully applied for quantification of the three iridoid glycosides in the collected plasma and various tissues following intravenous administration in rats. Picroside-I, II, and III were all eliminated rapidly with large volume of distribution. Among the three glycosides, Picroside-II showed the highest liver uptake, and only Picroside-I and II were found to get across the blood brain barrier (BBB). These results were consistent with their hepatoprotective or neuroprotective effects reported clinically. With the aid of the efficient and reliable simultaneous LC-ESI-MS/MS assay this pharmacokinetic study provided insights into their therapeutic targets of these three iridoid glycosides as well as valuable experimental basis for an expansion of their clinical indications. Copyright © 2015 Elsevier B.V. All rights reserved.

The development of cat testicular sperm cryopreservation protocols: Effects of tissue fragments or sperm cell suspension.

PubMed

Chatdarong, Kaywalee; Thuwanut, Paweena; Morrell, Jane M

2016-01-15

In endangered animals that have been found dead or sterilized for medical reasons, testis is the ultimate source of haploid DNA or sperm. Thus, preservation of testicular sperm may be performed to rescue their genetics. The aim of this study was to evaluate protocols for testicular sperm freezing: as tissue fragments or cell suspension in domestic cats as a model. A pair of testes from each cat (n = 9) were cut into eight equal pieces. Four randomly selected pieces were cryopreserved as: (1) tissue pieces using two-step freezing; (2) tissue pieces using a slow passive cooling device (CoolCell); (3) sperm suspension after single-layer centrifugation (SLC) through colloids; and (4) sperm suspension without being processed through SLC. A testicular piece from each cat served as fresh control. Testicular sperm membrane and DNA integrity were evaluated before, and after, the cryopreservation process. In addition, spermatogenic cell types (testicular sperm, spermatogonia, spermatocyte, and spermatid) present in the suspension samples were counted before and after SLC. The results found that testicular sperm membrane integrity in the suspension after SLC process was higher than that in the fragment form neither using the two-step nor CoolCell freezing, both before and after freezing (before freezing: 92.3 ± 3.4 vs. 81 ± 4.5 and 80.0 ± 7.0; after freezing: 84.5 ± 4.6 vs. 71.2 ± 12 and 76.2 ± 4.6; P ≤ 0.05). Testicular sperm DNA integrity was, however, not different among groups. Furthermore, the samples processed through the SLC had higher ration of sperm cells: other spermatogenic cells than those were not processed through the SLC (88.9 ± 3.8 vs. 30 ± 7.9; P ≤ 0.05). In summary, testicular sperm cryopreserved as a minced suspension is considered suitable in terms of preventing sperm membrane integrity, and SLC is considered a selection tool for enriching haploid sperm cells from castrated or postmortem cats. Copyright © 2016 Elsevier Inc. All rights reserved.

Serum, tissue and body fluid concentrations of tigecycline after a single 100 mg dose.

PubMed

Rodvold, Keith A; Gotfried, Mark H; Cwik, Michael; Korth-Bradley, Joan M; Dukart, Gary; Ellis-Grosse, Evelyn J

2006-12-01

The purpose of this study was to determine the tissue and corresponding serum concentration of tigecycline at selected time points in gall bladder, bile, colon, bone, synovial fluid (SF), lung and CSF in subjects undergoing surgical or medical procedures. One hundred and four adult subjects (aged 24-83 years; 64 women, 40 men) received a single intravenous (i.v.) dose of tigecycline (100 mg infused over 30 min). Subjects were randomly assigned to one of four collection times at 4, 8, 12 and 24 h after the start of the infusion. For CSF, samples were collected at approximately 1.5 and 24 h after the start of the infusion. All subjects had serum samples collected before the administration of tigecycline, at the end of the infusion and at the time corresponding to tissue or body fluid collection. Drug concentrations in serum, tissues and body fluids were determined by LC/MS/MS. The area under the mean concentration-time curve from 0 to 24 h (AUC(0-24)) was determined for the comparison of systemic exposure between tissue or body fluid to serum. The mean serum concentrations of tigecycline were similar to those previously published. Tissue penetration, expressed as the ratio of AUC(0-24) in tissue or body fluid to serum, was 537 for bile, 23 for gall bladder, 2.6 for colon, 2.0 for lung, 0.41 for bone, 0.31 for SF and 0.11 for CSF. A single 100 mg dose of intravenous tigecycline produced considerably higher tissue/fluid concentrations in bile, gall bladder, colon and lung compared with simultaneous serum concentrations. On average, the systemic exposure of tigecycline in bone, SF and CSF ranged from 11% to 41% of serum concentrations. The results in bone are inconsistent with previous radiolabelled studies in animals and it is unclear if tight binding to bone (versus low bone uptake) or poor extraction of tigecycline for LC/MS/MS detection or both may have contributed to the differences we observed in humans.

Imaging biological tissues with electrical conductivity contrast below 1 S m−1 by means of magnetoacoustic tomography with magnetic induction

PubMed Central

Hu, Gang; Li, Xu; He, Bin

2010-01-01

Magnetoacoustic tomography with magnetic induction (MAT-MI) is a recently introduced imaging modality for noninvasive electrical impedance imaging, with ultrasound imaging resolution and a contrast reflecting the electrical conductivity properties of tissues. However, previous MAT-MI systems can only image samples that are much more conductive than real human or animal tissues. To image real biological tissue samples, a large-current-carrying coil that can give stronger magnetic stimulations and stronger MAT-MI acoustic signals is employed in this study. The conductivity values of all the tissue samples employed in this study are also directly measured using a well calibrated four-electrode system. The experimental results demonstrated the feasibility to image biological tissues with electrical conductivity contrast below 1.0 S∕m using the MAT-MI technique with safe level of electromagnetic energy applied to tissue samples. PMID:20938494

Imaging biological tissues with electrical conductivity contrast below 1 S m-1 by means of magnetoacoustic tomography with magnetic induction

NASA Astrophysics Data System (ADS)

Hu, Gang; Li, Xu; He, Bin

2010-09-01

Magnetoacoustic tomography with magnetic induction (MAT-MI) is a recently introduced imaging modality for noninvasive electrical impedance imaging, with ultrasound imaging resolution and a contrast reflecting the electrical conductivity properties of tissues. However, previous MAT-MI systems can only image samples that are much more conductive than real human or animal tissues. To image real biological tissue samples, a large-current-carrying coil that can give stronger magnetic stimulations and stronger MAT-MI acoustic signals is employed in this study. The conductivity values of all the tissue samples employed in this study are also directly measured using a well calibrated four-electrode system. The experimental results demonstrated the feasibility to image biological tissues with electrical conductivity contrast below 1.0 S/m using the MAT-MI technique with safe level of electromagnetic energy applied to tissue samples.

The efficacy of selective calculus ablation at 400 nm: comparison to conventional calculus removal methods

NASA Astrophysics Data System (ADS)

Schoenly, Joshua E.; Seka, Wolf; Romanos, Georgios; Rechmann, Peter

A desired outcome of scaling and root planing is the complete removal of calculus and infected root tissue and preservation of healthy cementum for rapid healing of periodontal tissues. Conventional periodontal treatments for calculus removal, such as hand instrument scaling and ultrasonic scaling, often deeply scrape the surface of the underlying hard tissue and may leave behind a smear layer. Pulsed lasers emitting at violet wavelengths (specifically, 380 to 400 nm) are a potential alternative treatment since they can selectively ablate dental calculus without ablating pristine hard tissue (i.e., enamel, cementum, and dentin). In this study, light and scanning electron microscopy are used to compare and contrast the efficacy of in vitro calculus removal for several conventional periodontal treatments (hand instruments, ultrasonic scaler, and Er:YAG laser) to calculus removal with a frequency-doubled Ti:sapphire (λ = 400 nm). After calculus removal, enamel and cementum surfaces are investigated for calculus debris and damage to the underlying hard tissue surface. Compared to the smear layer, grooves, and unintentional hard tissue removal typically found using these conventional treatments, calculus removal using the 400-nm laser is complete and selective without any removal of pristine dental hard tissue. Based on these results, selective ablation from the 400-nm laser appears to produce a root surface that would be more suitable for successful healing of periodontal tissues.

Long non-coding RNA expression profile in cervical cancer tissues

PubMed Central

Zhu, Hua; Chen, Xiangjian; Hu, Yan; Shi, Zhengzheng; Zhou, Qing; Zheng, Jingjie; Wang, Yifeng

2017-01-01

Cervical cancer (CC), one of the most common types of cancer of the female population, presents an enormous challenge in diagnosis and treatment. Long non-coding (lnc)RNAs, non-coding (nc)RNAs with length >200 nucleotides, have been identified to be associated with multiple types of cancer, including CC. This class of nc transcripts serves an important role in tumor suppression and oncogenic signaling pathways. In the present study, the microarray method was used to obtain the expression profile of lncRNAs and protein-coding mRNAs and to compare the expression of lncRNAs between CC tissues and corresponding adjacent non-cancerous tissues in order to screen potential lncRNAs for associations with CC. Overall, 3356 lncRNAs with significantly different expression pattern in CC tissues compared with adjacent non-cancerous tissues were identified, while 1,857 of them were upregulated. These differentially expressed lncRNAs were additionally classified into 5 subgroups. Reverse transcription quantitative polymerase chain reactions were performed to validate the expression pattern of 5 random selected lncRNAs, and 2lncRNAs were identified to have significantly different expression in CC samples compared with adjacent non-cancerous tissues. This finding suggests that those lncRNAs with different expression may serve important roles in the development of CC, and the expression data may provide information for additional study on the involvement of lncRNAs in CC. PMID:28789353

Efficient Cancer Detection Using Multiple Neural Networks.

PubMed

Shell, John; Gregory, William D

2017-01-01

The inspection of live excised tissue specimens to ascertain malignancy is a challenging task in dermatopathology and generally in histopathology. We introduce a portable desktop prototype device that provides highly accurate neural network classification of malignant and benign tissue. The handheld device collects 47 impedance data samples from 1 Hz to 32 MHz via tetrapolar blackened platinum electrodes. The data analysis was implemented with six different backpropagation neural networks (BNN). A data set consisting of 180 malignant and 180 benign breast tissue data files in an approved IRB study at the Aurora Medical Center, Milwaukee, WI, USA, were utilized as a neural network input. The BNN structure consisted of a multi-tiered consensus approach autonomously selecting four of six neural networks to determine a malignant or benign classification. The BNN analysis was then compared with the histology results with consistent sensitivity of 100% and a specificity of 100%. This implementation successfully relied solely on statistical variation between the benign and malignant impedance data and intricate neural network configuration. This device and BNN implementation provides a novel approach that could be a valuable tool to augment current medical practice assessment of the health of breast, squamous, and basal cell carcinoma and other excised tissue without requisite tissue specimen expertise. It has the potential to provide clinical management personnel with a fast non-invasive accurate assessment of biopsied or sectioned excised tissue in various clinical settings.

Efficient Cancer Detection Using Multiple Neural Networks

PubMed Central

Gregory, William D.

2017-01-01

The inspection of live excised tissue specimens to ascertain malignancy is a challenging task in dermatopathology and generally in histopathology. We introduce a portable desktop prototype device that provides highly accurate neural network classification of malignant and benign tissue. The handheld device collects 47 impedance data samples from 1 Hz to 32 MHz via tetrapolar blackened platinum electrodes. The data analysis was implemented with six different backpropagation neural networks (BNN). A data set consisting of 180 malignant and 180 benign breast tissue data files in an approved IRB study at the Aurora Medical Center, Milwaukee, WI, USA, were utilized as a neural network input. The BNN structure consisted of a multi-tiered consensus approach autonomously selecting four of six neural networks to determine a malignant or benign classification. The BNN analysis was then compared with the histology results with consistent sensitivity of 100% and a specificity of 100%. This implementation successfully relied solely on statistical variation between the benign and malignant impedance data and intricate neural network configuration. This device and BNN implementation provides a novel approach that could be a valuable tool to augment current medical practice assessment of the health of breast, squamous, and basal cell carcinoma and other excised tissue without requisite tissue specimen expertise. It has the potential to provide clinical management personnel with a fast non-invasive accurate assessment of biopsied or sectioned excised tissue in various clinical settings. PMID:29282435

In situ hybridisation for the detection of Leishmania species in paraffin wax-embedded canine tissues using a digoxigenin-labelled oligonucleotide probe

PubMed Central

Dinhopl, N.; Mostegl, M. M.; Richter, B.; Nedorost, N.; Maderner, A.; Fragner, K.; Weissenböck, H.

2011-01-01

The diagnosis of canine leishmaniosis (CanL) is currently predominantly achieved by cytological or histological identification of amastigotes in biopsy samples, demonstration of specific anti-Leishmania antibodies and PCR-based approaches. All these methods have the advantage of being sensitive and more or less specific; nevertheless, most of them also have disadvantages. A chromogenic in situ hybridisation (ISH) procedure with a digoxigenin-labelled probe, targeting a fragment of the 5.8S rRNA was developed for the detection of all species of Leishmania parasites in routinely paraffin wax-embedded canine tissues. This method was validated in comparison with traditional techniques (histology, PCR), on various tissues from three dogs with histological changes consistent with a florid leishmaniosis. Amastigote forms of Leishmania gave clear signals and were easily identified using ISH. Various tissues from 10 additional dogs with clinical suspicion or/and a positive serological test but without histological presence of amastigotes did not show any ISH signals. Potential cross-reactivity of the probe was ruled out by negative outcome of the ISH against selected protozoa (including the related Trypanosoma cruzi) and fungi. Thus, ISH proved to be a powerful tool for unambiguous detection of Leishmania parasites in paraffin wax-embedded tissues. PMID:21921059

76 FR 71315 - Endangered and Threatened Species; Take of Anadromous Fish

Federal Register 2010, 2011, 2012, 2013, 2014

2011-11-17

... the fish and would then sample them for biological information (fin tissue and scale samples). They..., measured, weighed, tissue-sampled, and checked for external marks and coded-wire tags depending on the.... Then the researchers would remove and preserve fish body tissues, otoliths, and coded wire tags (from...

Age estimation based on aspartic acid racemization in human sclera.

PubMed

Klumb, Karolin; Matzenauer, Christian; Reckert, Alexandra; Lehmann, Klaus; Ritz-Timme, Stefanie

2016-01-01

Age estimation based on racemization of aspartic acid residues (AAR) in permanent proteins has been established in forensic medicine for years. While dentine is the tissue of choice for this molecular method of age estimation, teeth are not always available which leads to the need to identify other suitable tissues. We examined the suitability of total tissue samples of human sclera for the estimation of age at death. Sixty-five samples of scleral tissue were analyzed. The samples were hydrolyzed and after derivatization, the extent of aspartic acid racemization was determined by gas chromatography. The degree of AAR increased with age. In samples from younger individuals, the correlation of age and D-aspartic acid content was closer than in samples from older individuals. The age-dependent racemization in total tissue samples proves that permanent or at least long-living proteins are present in scleral tissue. The correlation of AAR in human sclera and age at death is close enough to serve as basis for age estimation. However, the precision of age estimation by this method is lower than that of age estimation based on the analysis of dentine which is due to molecular inhomogeneities of total tissue samples of sclera. Nevertheless, the approach may serve as a valuable alternative or addition in exceptional cases.

Tissue culture specificity of the tobacco ASA2 promoter driving hpt as a selectable marker for soybean transformation selection.

PubMed

Zernova, Olga; Zhong, Wei; Zhang, Xing-Hai; Widholm, Jack

2008-11-01

This study was carried out to determine if the tobacco anthranilate synthase ASA2 2.3 kb promoter drives tissue culture specific expression and if it is strong enough to drive hpt (hygromycin phosphotransferase) gene expression at a level sufficient to allow selection of transformed soybean embryogenic culture lines. A number of transformed cell lines were selected showing that the promoter was strong enough. Northern blot analysis of plant tissues did not detect hpt mRNA in the untransformed control or in the ASA2-hpt plants except in developing seeds while hpt mRNA was detected in all tissues of the CaMV35S-hpt positive control line plants. However, when the more sensitive RT-PCR assay was used all tissues of the ASA2-hpt plants except roots and mature seeds were found to contain detectable hpt mRNA. Embryogenic tissue cultures initiated from the ASA2-hpt plants contained hpt mRNA detectable by both northern and RT-PCR analysis and the cultures were hygromycin resistant. Friable callus initiated from leaves of ASA2-hpt plants did in some cases contain hpt mRNA that was only barely detectable by northern hybridization even though the callus was very hygromycin resistant. Thus the ASA2 promoter is strong enough to drive sufficient hpt expression in soybean embryogenic cultures for hygromycin selection and only very low levels of expression were found in most plant tissues with none in mature seeds.

Extreme Hypoxic Conditions Induce Selective Molecular Responses and Metabolic Reset in Detached Apple Fruit

PubMed Central

Cukrov, Dubravka; Zermiani, Monica; Brizzolara, Stefano; Cestaro, Alessandro; Licausi, Francesco; Luchinat, Claudio; Santucci, Claudio; Tenori, Leonardo; Van Veen, Hans; Zuccolo, Andrea; Ruperti, Benedetto; Tonutti, Pietro

2016-01-01

The ripening physiology of detached fruit is altered by low oxygen conditions with profound effects on quality parameters. To study hypoxia-related processes and regulatory mechanisms, apple (Malus domestica, cv Granny Smith) fruit, harvested at commercial ripening, were kept at 1°C under normoxic (control) and hypoxic (0.4 and 0.8 kPa oxygen) conditions for up to 60 days. NMR analyses of cortex tissue identified eight metabolites showing significantly different accumulations between samples, with ethanol and alanine displaying the most pronounced difference between hypoxic and normoxic treatments. A rapid up-regulation of alcohol dehydrogenase and pyruvate-related metabolism (lactate dehydrogenase, pyruvate decarboxylase, alanine aminotransferase) gene expression was detected under both hypoxic conditions with a more pronounced effect induced by the lowest (0.4 kPa) oxygen concentration. Both hypoxic conditions negatively affected ACC synthase and ACC oxidase transcript accumulation. Analysis of RNA-seq data of samples collected after 24 days of hypoxic treatment identified more than 1000 genes differentially expressed when comparing 0.4 vs. 0.8 kPa oxygen concentration samples. Genes involved in cell-wall, minor and major CHO, amino acid and secondary metabolisms, fermentation and glycolysis as well as genes involved in transport, defense responses, and oxidation-reduction appeared to be selectively affected by treatments. The lowest oxygen concentration induced a higher expression of transcription factors belonging to AUX/IAA, WRKY, HB, Zinc-finger families, while MADS box family genes were more expressed when apples were kept under 0.8 kPa oxygen. Out of the eight group VII ERF members present in apple genome, two genes showed a rapid up-regulation under hypoxia, and western blot analysis showed that apple MdRAP2.12 proteins were differentially accumulated in normoxic and hypoxic samples, with the highest level reached under 0.4 kPa oxygen. These data suggest that ripe apple tissues finely and specifically modulate sensing and regulatory mechanisms in response to different hypoxic stress conditions. PMID:26909091

Multi-mycotoxin analysis of animal feed and animal-derived food using LC-MS/MS system with timed and highly selective reaction monitoring.

PubMed

Zhao, Zhiyong; Liu, Na; Yang, Lingchen; Deng, Yifeng; Wang, Jianhua; Song, Suquan; Lin, Shanhai; Wu, Aibo; Zhou, Zhenlei; Hou, Jiafa

2015-09-01

Mycotoxins have the potential to enter the human food chain through carry-over of contaminants from feed into animal-derived products. The objective of the study was to develop a reliable and sensitive method for the analysis of 30 mycotoxins in animal feed and animal-derived food (meat, edible animal tissues, and milk) using liquid chromatography-tandem mass spectrometry (LC-MS/MS). In the study, three extraction procedures, as well as various cleanup procedures, were evaluated to select the most suitable sample preparation procedure for different sample matrices. In addition, timed and highly selective reaction monitoring on LC-MS/MS was used to filter out isobaric matrix interferences. The performance characteristics (linearity, sensitivity, recovery, precision, and specificity) of the method were determined according to Commission Decision 2002/657/EC and 401/2006/EC. The established method was successfully applied to screening of mycotoxins in animal feed and animal-derived food. The results indicated that mycotoxin contamination in feed directly influenced the presence of mycotoxin in animal-derived food. Graphical abstract Multi-mycotoxin analysis of animal feed and animal-derived food using LC-MS/MS.