Increased numbers of total nucleated and CD34+ cells in blood group O cord blood: an analysis of neonatal innate factors in the Korean population.

PubMed

Lee, Hye Ryun; Park, Jeong Su; Shin, Sue; Roh, Eun Youn; Yoon, Jong Hyun; Han, Kyou Sup; Kim, Byung Jae; Storms, Robert W; Chao, Nelson J

2012-01-01

We analyzed neonatal factors that could affect hematopoietic variables of cord blood (CB) donated from Korean neonates. The numbers of total nucleated cells (TNCs), CD34+ cells, and CD34+ cells/TNCs of CB in neonates were compared according to sex, gestational age, birth weight, birth weight centile for gestational age, and ABO blood group. With 11,098 CB units analyzed, blood group O CB showed an increased number of TNCs, CD34+ cells, and CD34+ cells/TNCs compared with other blood groups. Although TNC counts were lower in males, no difference in the number of CD34+ cells was demonstrated because the number of CD34+ cells/TNCs was higher in males. An increase in the gestational age resulted in an increase in the number of TNCs and decreases in the number of CD34+ cells and CD34+ cells/TNCs. The numbers of TNCs, CD34+ cells, and CD34+ cells/TNCs increased according to increased birth weight centile as well as birth weight. CB with blood group O has unique hematologic variables in this large-scale analysis of Korean neonates, although the impact on the storage policies of CB banks or the clinical outcome of transplantation remains to be determined. © 2011 American Association of Blood Banks.

Radix Ilicis Pubescentis total flavonoids combined with mobilization of bone marrow stem cells to protect against cerebral ischemia/reperfusion injury

PubMed Central

Miao, Ming-san; Guo, Lin; Li, Rui-qi; Ma, Xiao

2016-01-01

Previous studies have shown that Radix Ilicis Pubescentis total flavonoids have a neuroprotective effect, but it remains unclear whether Radix Ilicis Pubescentis total flavonoids have a synergistic effect with the recombinant human granulocyte colony stimulating factor-mobilized bone marrow stem cell transplantation on cerebral ischemia/reperfusion injury. Rat ischemia models were administered 0.3, 0.15 and 0.075 g/kg Radix Ilicis Pubescentis total flavonoids from 3 days before modeling to 2 days after injury. Results showed that Radix Ilicis Pubescentis total flavonoids could reduce pathological injury in rats with cerebral ischemia/reperfusion injury. The number of Nissl bodies increased, Bax protein expression decreased, Bcl-2 protein expression increased and the number of CD34-positive cells increased. Therefore, Radix Ilicis Pubescentis total flavonoids can improve the bone marrow stem cell mobilization effect, enhance the anti-apoptotic ability of nerve cells, and have a neuroprotective effect on cerebral ischemia/reperfusion injury in rats. PMID:27073381

Effect of cryopreservation and in vitro culture of bovine fibroblasts on histone acetylation levels and in vitro development of hand-made cloned embryos

USGS Publications Warehouse

Chacon, L.; Gomez, M.C.; Jenkins, J.A.; Leibo, S.P.; Wirtu, G.; Dresser, B.L.; Pope, C.E.

2011-01-01

In this study, the relative acetylation levels of histone 3 in lysine 9 (H3K9ac) in cultured and cryopreserved bovine fibroblasts was measured and we determined the influence of the epigenetic status of three cultured (C1, C2 and C3) donor cell lines on the in vitro development of reconstructed bovine embryos. Results showed that cryopreservation did not alter the overall acetylation levels of H3K9 in bovine fibroblasts analysed immediately after thawing (frozen/thawed) compared with fibroblasts cultured for a period of time after thawing. However, reduced cleavage rates were noted in embryos reconstructed with fibroblasts used immediately after thawing. Cell passage affects the levels of H3K9ac in bovine fibroblasts, decreasing after P1 and donor cells with lower H3K9ac produced a greater frequency of embryo development to the blastocyst stage. Cryopreservation did not influence the total cell and ICM numbers, or the ICM/TPD ratios of reconstructed embryos. However, the genetic source of donor cells did influence the total number of cells and the trophectoderm cell numbers, and the cell passage influenced the total ICM cell numbers. ?? Copyright Cambridge University Press 2010.

Total numbers of neurons and glial cells in cortex and basal ganglia of aged brains with Down syndrome--a stereological study.

PubMed

Karlsen, Anna Schou; Pakkenberg, Bente

2011-11-01

The total numbers of neurons and glial cells in the neocortex and basal ganglia in adults with Down syndrome (DS) were estimated with design-based stereological methods, providing quantitative data on brains affected by delayed development and accelerated aging. Cell numbers, volume of regions, and densities of neurons and glial cell subtypes were estimated in brains from 4 female DS subjects (mean age 66 years) and 6 female controls (mean age 70 years). The DS subjects were estimated to have about 40% fewer neocortical neurons in total (11.1 × 10(9) vs. 17.8 × 10(9), 2p ≤ 0.001) and almost 30% fewer neocortical glial cells with no overlap to controls (12.8 × 10(9) vs. 18.2 × 10(9), 2p = 0.004). In contrast, the total number of neurons in the basal ganglia was the same in the 2 groups, whereas the number of oligodendrocytes in the basal ganglia was reduced by almost 50% in DS (405 × 10(6) vs. 816 × 10(6), 2p = 0.01). We conclude that trisomy 21 affects cortical structures more than central gray matter emphasizing the differential impairment of brain development. Despite concomitant Alzheimer-like pathology, the neurodegenerative outcome in a DS brain deviates from common Alzheimer disease.

Regional differences in cell loss associated with binge-like alcohol exposure during the first two trimesters equivalent in the rat.

PubMed

Maier, S E; West, J R

2001-01-01

Women who abuse alcohol during pregnancy may deliver offspring who could be diagnosed with fetal alcohol syndrome (FAS) or a less severe deficit involving cognitive and behavioral disorders. The severity of the deficits may involve the interaction of several known risk factors, such as alcohol consumption pattern or duration, the timing of alcohol consumption relative to critical windows of vulnerability, or the inherent differential vulnerability among the various brain regions to alcohol-induced brain injury. In this study, we explore the vulnerability of the different brain regions by making cell counts from multiple brain regions. Specifically, we used stereological cell-counting techniques to estimate the total cell numbers in the cerebellum (Purkinje and granule cells), olfactory bulb (mitral and granule cells), hippocampus (CA1 and CA3 cells), and dentate gyrus (granule cells). Groups of timed-pregnant Sprague-Dawley rats were assigned to one of five treatments: alcohol by intragastric intubation (2.25, 4.5, or 6.5 g/kg/day), nutritional control [pairfed and intubated=Pairfed) and intubated], and normal control (Chow). Treatments began on embryonic day 1 (E1) and continued through E20. On E33 (usually postnatal day 10), all offspring were perfused intracardially with saline followed by fixatives. Representative forebrains, cerebella, and olfactory bulb from each group were processed for cell counting. The optical dissector was used to obtain cell densities, while Cavalieri's principle was used to calculate the reference volume. The product of density and volume gave unbiased estimates of the total neuronal number within each brain region. Overall peak BACs (regardless of sampling day) for the three alcohol groups averaged 136, 290, and 422 mg/dl for the 2.25-, 4.5-, and 6.5-g/kg groups, respectively. The total number of cerebellar Purkinje cells was reduced in the 6.5-g/kg group relative to controls, while the total number of olfactory bulb mitral cells and hippocampal CA1 and CA3 pyramidal cells from all alcohol-treated groups was not different from controls. Total numbers of granule neurons were reduced in the cerebellum and olfactory bulb of offspring exposed to 4.5 or 6.5 g/kg/day, but granule cell numbers in the dentate gyrus were not affected by the prenatal alcohol treatment. Taken together with previous findings, these data demonstrate that prenatal alcohol exposure results in regional vulnerability of various brain structures and underscores the variability of deleterious effects of alcohol on brain development.

Total extract of Korean red ginseng facilitates human bone marrow hematopoietic colony formation in vitro

PubMed Central

Kim, Sang-Gyung; Bae, Sung Hwa; Kim, Seong-Mo; Lee, Ji-Hye; Kim, Min Ji; Jang, Hae-Bong

2014-01-01

Background The number of CD34+ cells in a peripheral blood stem cell collection is the key factor in predicting successful treatment of hematologic malignancies. Korean Red Ginseng (KRG) (Panax ginseng C.A. Meyer) is the most popular medicinal herb in Korea. The objective of this study was to determine the effect of KRG on hematopoietic colony formation. Methods Bone marrow (BM) samples were obtained from 8 human donors after acquiring informed consent. BM mononuclear cells (MNCs) were isolated, and CD34+ cells were sorted using magnetic beads. The sorted CD34+ cells were incubated with or without total extract of KRG (50 µg/mL, 100 µg/mL) or Ginsenoside Rg1 (100 µg/mL), and the hematopoietic colony assay was performed using methylcellulose semisolid medium. The CD34+ cell counts were measured by a single platform assay using flow cytometry. Results The numbers of human BM-MNCs and CD34+ cells obtained after purification were variable among donors (5.6×107 and 1.3-48×107 and 8.9×104 and 1.8-80×104, respectively). The cells expanded 1,944 times after incubation for 12 d. Total extract of KRG added to the hematopoietic stem cell (HSC)-specific medium increased CD34+ cell counts 3.6 times compared to 2.6 times when using HSC medium alone. Total numbers of hematopoietic colonies in KRG medium were more than those observed in conventional medium, especially that of erythroid colonies such as burst forming unit-erythroid. Conclusion Total extract of KRG facilitated CD34+ cell expansion and hematopoietic colony formation, especially of the erythroid lineage. PMID:25325037

Ursodeoxycholic acid inhibits the proliferation of colon cancer cells by regulating oxidative stress and cancer stem-like cell growth.

PubMed

Kim, Eun-Kyung; Cho, Jae Hee; Kim, EuiJoo; Kim, Yoon Jae

2017-01-01

The regulation of reactive oxygen species (ROS) exists as a therapeutic target for cancer treatments. Previous studies have shown that ursodeoxycholic acid (UDCA) suppresses the proliferation of colon cancer cells. The aim of this study was to evaluate the effect of UDCA upon the proliferation of colon cancer cells as a direct result of the regulation of ROS. Colon cancer cell lines (HT29 and HCT116) were treated with UDCA. The total number of cells and the number of dead cells were determined using cell counters. A fluorescein isothiocyanate-bromodeoxyuridine flow kit was used to analyze cell cycle variations. Upon exposure to UDCA, the protein levels of p27, p21, CDK2, CDK4 and CDK6 were determined using western blotting, and qRT-PCR was used to determine levels of mRNA. We preformed dichlorofluorescindiacetate (DCF-DA) staining to detect alteration of intracellular ROS using fluorescence activated cell sorting (FACS). Colon cancer stem-like cell lines were generated by tumorsphere culture and treated with UDCA for seven days. The total number of tumorspheres was determined using microscopy. We found that UDCA reduced the total number of colon cancer cells, but did not increase the number of dead cells. UDCA inhibited the G1/S and G2/M transition phases in colon cancer cells. UDCA induced expression of cell cycle inhibitors such as p27 and p21. However, it was determined that UDCA suppressed levels of CDK2, CDK4, and CDK6. UDCA regulated intracellular ROS generation in colon cancer cells, and induced activation of Erk1/2. Finally, UDCA inhibited formation of colon cancer stem-like cells. Our results indicate that UDCA suppresses proliferation through regulation of oxidative stress in colon cancer cells, as well as colon cancer stem-like cells.

Ursodeoxycholic acid inhibits the proliferation of colon cancer cells by regulating oxidative stress and cancer stem-like cell growth

PubMed Central

Kim, EuiJoo

2017-01-01

Introduction The regulation of reactive oxygen species (ROS) exists as a therapeutic target for cancer treatments. Previous studies have shown that ursodeoxycholic acid (UDCA) suppresses the proliferation of colon cancer cells. The aim of this study was to evaluate the effect of UDCA upon the proliferation of colon cancer cells as a direct result of the regulation of ROS. Method Colon cancer cell lines (HT29 and HCT116) were treated with UDCA. The total number of cells and the number of dead cells were determined using cell counters. A fluorescein isothiocyanate-bromodeoxyuridine flow kit was used to analyze cell cycle variations. Upon exposure to UDCA, the protein levels of p27, p21, CDK2, CDK4 and CDK6 were determined using western blotting, and qRT-PCR was used to determine levels of mRNA. We preformed dichlorofluorescindiacetate (DCF-DA) staining to detect alteration of intracellular ROS using fluorescence activated cell sorting (FACS). Colon cancer stem-like cell lines were generated by tumorsphere culture and treated with UDCA for seven days. The total number of tumorspheres was determined using microscopy. Results We found that UDCA reduced the total number of colon cancer cells, but did not increase the number of dead cells. UDCA inhibited the G1/S and G2/M transition phases in colon cancer cells. UDCA induced expression of cell cycle inhibitors such as p27 and p21. However, it was determined that UDCA suppressed levels of CDK2, CDK4, and CDK6. UDCA regulated intracellular ROS generation in colon cancer cells, and induced activation of Erk1/2. Finally, UDCA inhibited formation of colon cancer stem-like cells. Conclusion Our results indicate that UDCA suppresses proliferation through regulation of oxidative stress in colon cancer cells, as well as colon cancer stem-like cells. PMID:28708871

Basolateral amygdala volume and cell numbers in major depressive disorder: a postmortem stereological study.

PubMed

Rubinow, Marisa J; Mahajan, Gouri; May, Warren; Overholser, James C; Jurjus, George J; Dieter, Lesa; Herbst, Nicole; Steffens, David C; Miguel-Hidalgo, Jose J; Rajkowska, Grazyna; Stockmeier, Craig A

2016-01-01

Functional imaging studies consistently report abnormal amygdala activity in major depressive disorder (MDD). Neuroanatomical correlates are less clear: imaging studies have produced mixed results on amygdala volume, and postmortem neuroanatomic studies have only examined cell densities in portions of the amygdala or its subregions in MDD. Here, we present a stereological analysis of the volume of, and the total number of, neurons, glia, and neurovascular (pericyte and endothelial) cells in the basolateral amygdala in MDD. Postmortem tissues from 13 subjects with MDD and 10 controls were examined. Sections (~15/subject) taken throughout the rostral-caudal extent of the basolateral amygdala (BLA) were stained for Nissl substance and utilized for stereological estimation of volume and cell numbers. Results indicate that depressed subjects had a larger lateral nucleus than controls and a greater number of total BLA neurovascular cells than controls. There were no differences in the number or density of neurons or glia between depressed and control subjects. These findings present a more detailed picture of BLA cellular anatomy in depression than has previously been available. Further studies are needed to determine whether the greater number of neurovascular cells in depressed subjects may be related to increased amygdala activity in depression.

Mobilization of hematopoietic progenitor cells with granulocyte colony stimulating factors for autologous transplant in hematologic malignancies: a single center experience

PubMed Central

Gabús, Raul; Borelli, Gabriel; Ferrando, Martín; Bódega, Enrique; Citrín, Estela; Jiménez, Constanza Olivera; Álvarez, Ramón

2011-01-01

Background In 2006 the Hematology Service of Hospital Maciel published its experience with peripheral blood progenitor cell harvesting for autologous stem cell transplantation using Filgen JP (Clausen Filgrastim). After mobilization with a mean filgrastim dose of 78 mcg/Kg, 4.7 x 106 CD34+ cells/Kg were obtained by apheresis. Age above 50, multiple myeloma as underlying disease and a malignancy that was not in remission were identified as frequent characteristics among patients showing complex mobilization. Objective The aim of this study was to compare stem cell mobilization using different brands of filgrastim. Methods One hundred and fifty-seven mobilizations performed between 1997 and 2006 were analyzed. This retrospective analysis comparative two groups of patients: those mobilized with different brands of filgrastim (Group A) and those who received Filgen JP (Clausen Filgrastim) as mobilizing agent (Group B). A cluster analysis technique was used to identify four clusters of individuals with different behaviors differentiated by age, total dose of filgrastim required, number of apheresis and harvested CD34+ cells. Results The mean total dose of filgrastim administered was 105 mcg/Kg, the median number of apheresis was 2 procedures and the mean number of harvested stem cells was 4.98 x 106 CD34+ cells/Kg. No significant differences were observed between Groups A and B regarding the number of apheresis, harvested CD34+ cells and number of mobilization failures, however the total dose of filgrastim was significantly lower in Group B. Conclusions Among other factors, the origin of the cytokine used as mobilizing agent is an element to be considered when evaluating CD34+ cell mobilization results. PMID:23049356

Mobilization of hematopoietic progenitor cells with granulocyte colony stimulating factors for autologous transplant in hematologic malignancies: a single center experience.

PubMed

Gabús, Raul; Borelli, Gabriel; Ferrando, Martín; Bódega, Enrique; Citrín, Estela; Jiménez, Constanza Olivera; Alvarez, Ramón

2011-01-01

In 2006 the Hematology Service of Hospital Maciel published its experience with peripheral blood progenitor cell harvesting for autologous stem cell transplantation using Filgen JP (Clausen Filgrastim). After mobilization with a mean filgrastim dose of 78 mcg/Kg, 4.7 x 10(6) CD34(+) cells/Kg were obtained by apheresis. Age above 50, multiple myeloma as underlying disease and a malignancy that was not in remission were identified as frequent characteristics among patients showing complex mobilization. The aim of this study was to compare stem cell mobilization using different brands of filgrastim. One hundred and fifty-seven mobilizations performed between 1997 and 2006 were analyzed. This retrospective analysis comparative two groups of patients: those mobilized with different brands of filgrastim (Group A) and those who received Filgen JP (Clausen Filgrastim) as mobilizing agent (Group B). A cluster analysis technique was used to identify four clusters of individuals with different behaviors differentiated by age, total dose of filgrastim required, number of apheresis and harvested CD34(+) cells. The mean total dose of filgrastim administered was 105 mcg/Kg, the median number of apheresis was 2 procedures and the mean number of harvested stem cells was 4.98 x 10(6) CD34(+) cells/Kg. No significant differences were observed between Groups A and B regarding the number of apheresis, harvested CD34(+) cells and number of mobilization failures, however the total dose of filgrastim was significantly lower in Group B. Among other factors, the origin of the cytokine used as mobilizing agent is an element to be considered when evaluating CD34(+) cell mobilization results.

Determining the number of fingers in the lifting Hele-Shaw problem

NASA Astrophysics Data System (ADS)

Miranda, Jose; Dias, Eduardo

2013-11-01

The lifting Hele-Shaw cell flow is a variation of the celebrated radial viscous fingering problem for which the upper cell plate is lifted uniformly at a specified rate. This procedure causes the formation of intricate interfacial patterns. Most theoretical studies determine the total number of emerging fingers by maximizing the linear growth rate, but this generates discrepancies between theory and experiments. In this work, we tackle the number of fingers selection problem in the lifting Hele-Shaw cell by employing the recently proposed maximum-amplitude criterion. Our linear stability analysis accounts for the action of capillary, viscous normal stresses, and wetting effects, as well as the cell confinement. The comparison of our results with very precise laboratory measurements for the total number of fingers shows a significantly improved agreement between theoretical predictions and experimental data. We thank CNPq (Brazilian Sponsor) for financial support.

Peripheral vestibular pathology in Mondini dysplasia.

PubMed

Kaya, Serdar; Hızlı, Ömer; Kaya, Fatıma Kübra; Monsanto, Rafael DaCosta; Paparella, Michael M; Cureoglu, Sebahattin

2017-01-01

In this study, our objective was to histopathologically analyze the peripheral vestibular system in patients with Mondini dysplasia. Comparative human temporal bone study. We assessed the sensory epithelium of the human vestibular system with a focus on the number of type I and type II hair cells, as well as the total number of hair cells. We compared those numbers in our Mondini dysplasia group versus our control group. The loss of type I and type II hair cells in the cristae of the superior, lateral, and posterior semicircular canals, as well as in the saccular and utricular macula, was significantly higher in our Mondini dysplasia group than in our control group. The total number of hair cells significantly decreased in the cristae of the superior, lateral, and posterior semicircular canals, as well as in the saccular and utricular macula, in our Mondini dysplasia group. Loss of vestibular hair cells can lead to vestibular dysfunction in patients with Mondini dysplasia. NA Laryngoscope, 127:206-209, 2017. © 2016 The American Laryngological, Rhinological and Otological Society, Inc.

Image analysis for the automated estimation of clonal growth and its application to the growth of smooth muscle cells.

PubMed

Gavino, V C; Milo, G E; Cornwell, D G

1982-03-01

Image analysis was used for the automated measurement of colony frequency (f) and colony diameter (d) in cultures of smooth muscle cells, Initial studies with the inverted microscope showed that number of cells (N) in a colony varied directly with d: log N = 1.98 log d - 3.469 Image analysis generated the complement of a cumulative distribution for f as a function of d. The number of cells in each segment of the distribution function was calculated by multiplying f and the average N for the segment. These data were displayed as a cumulative distribution function. The total number of colonies (fT) and the total number of cells (NT) were used to calculate the average colony size (NA). Population doublings (PD) were then expressed as log2 NA. Image analysis confirmed previous studies in which colonies were sized and counted with an inverted microscope. Thus, image analysis is a rapid and automated technique for the measurement of clonal growth.

Evaluation of the percentage of ganglion cells in the ganglion cell layer of the rodent retina

PubMed Central

Schlamp, Cassandra L.; Montgomery, Angela D.; Mac Nair, Caitlin E.; Schuart, Claudia; Willmer, Daniel J.

2013-01-01

Purpose Retinal ganglion cells comprise a percentage of the neurons actually residing in the ganglion cell layer (GCL) of the rodent retina. This estimate is useful to extrapolate ganglion cell loss in models of optic nerve disease, but the values reported in the literature are highly variable depending on the methods used to obtain them. Methods We tested three retrograde labeling methods and two immunostaining methods to calculate ganglion cell number in the mouse retina (C57BL/6). Additionally, a double-stain retrograde staining method was used to label rats (Long-Evans). The number of total neurons was estimated using a nuclear stain and selecting for nuclei that met specific criteria. Cholinergic amacrine cells were identified using transgenic mice expressing Tomato fluorescent protein. Total neurons and total ganglion cell numbers were measured in microscopic fields of 104 µm2 to determine the percentage of neurons comprising ganglion cells in each field. Results Historical estimates of the percentage of ganglion cells in the mouse GCL range from 36.1% to 67.5% depending on the method used. Experimentally, retrograde labeling methods yielded a combined estimate of 50.3% in mice. A retrograde method also yielded a value of 50.21% for rat retinas. Immunolabeling estimates were higher at 64.8%. Immunolabeling may introduce overestimates, however, with non-specific labeling effects, or ectopic expression of antigens in neurons other than ganglion cells. Conclusions Since immunolabeling methods may overestimate ganglion cell numbers, we conclude that 50%, which is consistently derived from retrograde labeling methods, is a reliable estimate of the ganglion cells in the neuronal population of the GCL. PMID:23825918

High dosage of monosodium glutamate causes deficits of the motor coordination and the number of cerebellar Purkinje cells of rats.

PubMed

Prastiwi, D; Djunaidi, A; Partadiredja, G

2015-11-01

Monosodium glutamate (MSG) has been widely used throughout the world as a flavoring agent of food. However, MSG at certain dosages is also thought to cause damage to many organs, including cerebellum. This study aimed at investigating the effects of different doses of MSG on the motor coordination and the number of Purkinje cells of the cerebellum of Wistar rats. A total of 24 male rats aged 4 to 5 weeks were divided into four groups, namely, control (C), T2.5, T3, and T3.5 groups, which received intraperitoneal injection of 0.9% sodium chloride solution, 2.5 mg/g body weight (bw) of MSG, 3.0 mg/g bw of MSG, and 3.5 mg/g bw of MSG, respectively, for 10 consecutive days. The motor coordination of the rats was examined prior and subsequent to the treatment. The number of cerebellar Purkinje cells was estimated using physical fractionator method. It has been found that the administration of MSG at a dosage of 3.5 mg/g bw, but not at lower dosages, caused a significant decrease of motor coordination and the estimated total number of Purkinje cells of rats. There was also a significant correlation between motor coordination and the total number of Purkinje cells. © The Author(s) 2015.

Hypoxia-Induced Expression of VEGF Splice Variants and Protein in Four Retinal Cell Types

PubMed Central

Watkins, William M.; McCollum, Gary W.; Savage, Sara R.; Capozzi, Megan E.; Penn, John S.; Morrison, David G.

2014-01-01

The purpose of this study was to investigate the hypoxia-induced Vegf120, Vegf164 and Vegf188 mRNA expression profiles in rat Müller cells (MC), astrocytes, retinal pigmented epithelial cells (RPE) and retinal microvascular endothelial cells (RMEC) and correlate these findings to VEGF secreted protein. Cultured cells were exposed to normoxia or hypoxia. Total RNA was isolated from cell lysates and Vegf splice variant mRNA copy numbers were assayed by a validated qRT-PCR external calibration curve method. mRNA copy numbers were normalized to input total RNA. Conditioned medium was collected from cells and assayed for total VEGF protein by ELISA. Hypoxia increased total Vegf mRNA and secreted protein in all the retinal cell types, with the highest levels observed in MC and astrocytes ranking second. Total Vegf mRNA levels in hypoxic RPE and RMEC were comparable; however, the greatest hypoxic induction of each Vegf splice variant mRNA was observed in RMEC. RPE and RMEC ranked 3rd and 4th respectively, in terms of secreted total VEGF protein in hypoxia. The Vegf120, Vegf164 and Vegf188 mRNA splice variants were all increased in hypoxic cells compared to normoxic controls. In normoxia, the relative Vegf splice variant mRNA levels ranked from highest to lowest for each cell type were Vegf164>Vegf120>Vegf188. Hypoxic induction did not alter this ranking, although it did favor an increased stoichiometry of Vegf164 mRNA over the other two splice variants. MC and astrocytes are likely to be the major sources of total Vegf, and Vegf164 splice variant mRNAs, and VEGF protein in retinal hypoxia. PMID:24076411

[A comparative study on inducing non-homologous mesenchymal stem cells to differentiate into neural stem cells using non-homologous cerebrospinal fluid].

PubMed

Ren, Chao; Liu, Xiaoyun; Wan, Meirong; Geng, Deqin; Ge, Wei; Li, Jinmei; Zhang, Weiwei

2013-12-01

In order to set up a base for stem cells to be widely used in clinical medicine, we tried to optimize, in this study, the technique that induces human mesenchymal stem cells (hMSCs) to differentiate into neural stem cells by using cerebrospinal fluid (CSF) from the different groups. After the induction, presence of neural stem cells was confirmed with microscope observation, flow cytometry analysis, immunohistochemistry and fluorescent immunohistochemistry. At the same time, we also compared and analysed the data of the number of stem cells when it totally met the requirements for clinical treatment and the days required. At last, we confirmed that hMSCs could be induced to differentiate into neural stem cells, and that the number of cells totally met the requirements for clinical treatment. But there were some differences both in the number of cells and the days required. Among the groups, the group that marrow mesenchymal stem cells from patients own induced by CSF from healthy volunteers used the shortest time and the quantity of the cells was significantly higher than those of the others.

B-Cell Depletion Promotes Aortic Infiltration of Immunosuppressive Cells and Is Protective of Experimental Aortic Aneurysm.

PubMed

Schaheen, Basil; Downs, Emily A; Serbulea, Vlad; Almenara, Camila C P; Spinosa, Michael; Su, Gang; Zhao, Yunge; Srikakulapu, Prasad; Butts, Cherié; McNamara, Coleen A; Leitinger, Norbert; Upchurch, Gilbert R; Meher, Akshaya K; Ailawadi, Gorav

2016-11-01

B-cell depletion therapy is widely used for treatment of cancers and autoimmune diseases. B cells are abundant in abdominal aortic aneurysms (AAA); however, it is unknown whether B-cell depletion therapy affects AAA growth. Using experimental models of murine AAA, we aim to examine the effect of B-cell depletion on AAA formation. Wild-type or apolipoprotein E-knockout mice were treated with mouse monoclonal anti-CD20 or control antibodies and subjected to an elastase perfusion or angiotensin II infusion model to induce AAA, respectively. Anti-CD20 antibody treatment significantly depleted B1 and B2 cells, and strikingly suppressed AAA growth in both models. B-cell depletion resulted in lower circulating IgM levels, but did not affect the levels of IgG or cytokine/chemokine levels. Although the total number of leukocyte remained unchanged in elastase-perfused aortas after anti-CD20 antibody treatment, the number of B-cell subtypes was significantly lower. Interestingly, plasmacytoid dendritic cells expressing the immunomodulatory enzyme indole 2,3-dioxygenase were detected in the aortas of B-cell-depleted mice. In accordance with an increase in indole 2,3-dioxygenase+ plasmacytoid dendritic cells, the number of regulatory T cells was higher, whereas the expression of proinflammatory genes was lower in aortas of B-cell-depleted mice. In a coculture model, the presence of B cells significantly lowered the number of indole 2,3-dioxygenase+ plasmacytoid dendritic cells without affecting total plasmacytoid dendritic cell number. The present results demonstrate that B-cell depletion protects mice from experimental AAA formation and promotes emergence of an immunosuppressive environment in aorta. © 2016 American Heart Association, Inc.

Effects of hypergravity on the development of cell number and asymmetry in fish brain nuclei

NASA Astrophysics Data System (ADS)

Anken, R. H.; Werner, K.; Rahmann, H.

Larval cichlid fish ( Oreochromis mossambicus) siblings were subjected to 3g hypergravity (hg) and total darkness for 21 days during development and subsequently processed for conventional histology. Further siblings reared at 1g and alternating light/dark (12h:12h) conditions served as contros. Cell number counts of the visual Nucleus isthmi (Ni) versus the vestibular Nucleus magnocellularis (Nm) revealed that in experimental animals total cell number was decreased in the Ni, possibly due to retarded growth as a result of the lack of visual input whereas no effect was observed in the Nm. Calculating the percentual asymmetry in cell number (i.e., right vs. the left side of the brain), no effects of hg/darkness were seen in the Ni, whereas asymmetry was slightly increased in the Nm. Since the asymmetry of inner ear otoliths is decreased under hg, this finding may indicate efferent vestibular action of the CNS on the level of the Nm by means of a feedback mechanism.

Fetal hemoglobin in sickle cell anemia: a glass half full?

PubMed

Steinberg, Martin H; Chui, David H K; Dover, George J; Sebastiani, Paola; Alsultan, Abdulrahman

2014-01-23

Fetal hemoglobin (HbF) modulates the phenotype of sickle cell anemia by inhibiting deoxy sickle hemoglobin (HbS) polymerization. The blood concentration of HbF, or the number of cells with detectable HbF (F-cells), does not measure the amount of HbF/F-cell. Even patients with high HbF can have severe disease because HbF is unevenly distributed among F-cells, and some cells might have insufficient concentrations to inhibit HbS polymerization. With mean HbF levels of 5%, 10%, 20%, and 30%, the distribution of HbF/F-cell can greatly vary, even if the mean is constant. For example, with 20% HbF, as few as 1% and as many as 24% of cells can have polymer-inhibiting, or protective, levels of HbF of ∼10 pg; with lower HbF, few or no protected cells can be present. Only when the total HbF concentration is near 30% is it possible for the number of protected cells to approach 70%. Rather than the total number of F-cells or the concentration of HbF in the hemolysate, HbF/F-cell and the proportion of F-cells that have enough HbF to thwart HbS polymerization is the most critical predictor of the likelihood of severe sickle cell disease.

Ratio of inner cell mass and trophoblastic cells in demi- and intact pig embryos.

PubMed

Tao, T; Reichelt, B; Niemann, H

1995-07-01

Pig morulae, early blastocysts and blastocysts were microsurgically bisected to produce zona-free demi-embryos or remained nonbisected with or without zona pellucida, and the presence of inner cell mass cells was determined using a differential fluorochrome staining technique. After 24 h of in vitro culture, all demi-embryos were classified into three categories, based on morphological criteria: 1, excellent; 2, fair; and 3, degenerated. The average number of total cells and inner cell mass cells in intact embryos cultured without zona pellucida for 24 h was higher (P < 0.05) than that for those with zona pellucida in morulae and early blastocysts. The percentage of demi-embryos without inner cell mass cells in these different morphological categories was 18.7%, 22.2% and 29.8% for morulae, respectively; 3.8%, 16.7% and 30.8% for early blastocysts, respectively; and 3.7%, 32.0% and 36.4% for blastocysts, respectively. The percentage of demi-embryos without inner cell mass cells was lower (P < 0.01) in demi-embryos classified in category 1 compared with category 3 in early blastocysts and in category 1 compared with categories 2 and 3 in blastocysts. Significant differences in the total number of cells and the number of inner cell mass cells were apparent among the three morphological categories of demi-embryos derived from morulae, early blastocysts and blastocysts. The ratio of total cells to inner cell mass cells was similar among intact pig embryos and the different morphological categories of demi-embryos derived from morulae, early blastocysts and blastocysts, with the exception of that between demi-blastocysts of category 1 and the other groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Improving the quality of miniature pig somatic cell nuclear transfer blastocysts: aggregation of SCNT embryos at the four-cell stage.

PubMed

Terashita, Y; Sugimura, S; Kudo, Y; Amano, R; Hiradate, Y; Sato, E

2011-04-01

Miniature pigs share many similar characteristics such as anatomy, physiology and body size with humans and are expected to become important animal models for therapeutic cloning using embryonic stem cells (ESCs) derived by somatic cell nuclear transfer (SCNT). In the present study, we observed that miniature pig SCNT blastocysts possessed a lower total number of nuclei and a lower percentage of POU5F1-positive cells than those possessed by in vitro fertilized (IVF) blastocysts. To overcome these problems, we evaluated the applicability of aggregating miniature pig SCNT embryos at the four-cell stage. We showed that (i) aggregation of two or three miniature pig SCNT embryos at the four-cell stage improves the total number of nuclei and the percentage of POU5F1-positive cells in blastocysts, and (ii) IVF blastocysts with low cell numbers induced by the removal of two blastomeres at the four-cell stage did not exhibit a decrease in the percentage of POU5F1-positive cells. These results suggest that the aggregation of miniature pig SCNT embryos at the four-cell stage can be a useful technique for improving the quality of miniature pig SCNT blastocysts and indicating that improvement in the percentage of POU5F1-positive cells in aggregated SCNT embryos is not simply the consequence of increased cell numbers. © 2010 Blackwell Verlag GmbH.

Effect of Early Neonatal Exposure to Deltamethrin on the Purkinje Cell Number in Rat Cerebellum

PubMed Central

Asari, Mohd Asnizam; Abdullah, Mohammad Shukri; Abdullah, Suryati

2008-01-01

Deltamethrin is a widely used insecticide belonging to the class of pyrethroid. Although the neurotoxicity of pyrethroids including deltamethrin is well established, it is still unclear whether exposure to deltamethrin during neonatal period has any deleterious effect on the survival of the Purkinje cells in the cerebellum. In the study, we investigated the total number of Purkinje cells in experimental rats exposed to deltamethrin using a stereological method, the fractionator. Deltamethrin in a dose of 1 mg/kg/day (corresponds to 20% of LD50 ) was administered through oral gavage to male pups from 2nd to 5th postnatal day (PND). At PND 21 the animals were sacrificed and their cerebelli were removed. The cerebelli were systematically sampled using the fractionator method and stained with cresyl fast violet. The number of the Purkinje cells was counted for each cerebellum. The results showed that there was no significant difference in the total number of Purkinje cells in the deltamethrin-treated group as compared to the control animals. This suggests that deltamethrin exposure at the current dosage during the neonatal period do not have any significant effect on the survival of the Purkinje cells in the cerebellum. PMID:22570585

Microbes Characteristics in Groundwater Flow System in Mountainous Area

NASA Astrophysics Data System (ADS)

Yamamoto, Chisato; Tsujimura, Maki; Kato, Kenji; Sakakibara, Koichi; Ogawa, Mahiro; Sugiyama, Ayumi; Nagaosa, Kazuyo

2017-04-01

We focus on a possibility of microbes as a tracer for groundwater flow investigation. Some previous papers showed that the total number of prokaryotes in groundwater has correlation with depth and geology (Parkes et al., 1994; Griebler et al., 2009; Kato et al., 2012). However, there are few studies investigating both microbe characteristics and groundwater flow system. Therefore, we investigated a relationship between the total number of prokaryotes and age of spring water and groundwater. Intensive field survey was conducted at four mountainous areas, namely Mt. Fuji (volcano), a headwater at Mt. Setohachi, a headwater at River Oi and a headwater at River Nagano underlain by volcanic lava at Mt. Fuji, granite at Mt. Setohachi and sedimentary rock at River Oi and River Nagano. We collected totally 40 spring water/ groundwater samples in these mountainous areas in October 2015, August, October and November 2016 and analyzed concentration of inorganic ions, the stable isotopes of oxygen - 18, deuterium, CFCs and SF6. Also, we counted prokaryotic cells under the epifluorescence microscopy after fixation and filteration. The total number of prokaryotes in the spring water/ groundwater ranged from 1.0×102 to 7.0×103cells mL-1 at the Mt. Fuji, 1.3×104 to 2.7×105cells mL-1 at Mt. Setohachi, 3.1×104cells mL-1 at River Oi and 1.8×105 to 3.2×106cells mL-1 at River Nagano. The SF6 age of the spring water/ groundwater ranged from 8 to 64 years at Mt. Fuji, 2 to 32.5 years at Mt. Setohachi, 2.5 years at River Oi and 15 to 16 years at River Nagano. The total number of prokaryotes showed a clear negative correlation with residence time of spring water/ groundwater in all regions. Especially the prokaryotes number increased in the order of 102 cells mL-1 with decreasing of residence time in approximately 10 years in the groundwater and spring water with the age less than 15 years.

Long-term Erythrocytapheresis Is Associated With Reduced Liver Iron Concentration in Sickle Cell Disease.

PubMed

Myers, Scott N; Eid, Ryan; Myers, John; Bertolone, Salvatore; Panigrahi, Arun; Mullinax, Jennifer; Raj, Ashok B

2016-01-01

Erythrocytapheresis procedures are increasingly used in sickle cell disease. Serum ferritin and noninvasive magnetic resonance imaging measurements of liver iron concentration (LIC) are frequently used to monitor iron overload secondary to hypertransfusion. There is a paucity of data describing the impact of long-term erythrocytapheresis (LTE) on LIC. We measured magnetic resonance imaging liver and cardiac iron on LTE subjects and stratified them into 2 groups: higher LIC (>3 mg/g) and lower LIC (<3 mg/g). χ(2) and t test were used to test for differences between the 2 groups. Logistic regression and generalized linear mixed-effects models were used to test what impacted LIC. None of 29 sickle cell disease subjects maintained on LTE had high cardiac iron concentration. LIC was associated with serum ferritin (r=0.697, P<0.001) but was not associated with the total number of LTE procedures (r=-0.088, P=0.656) or total number of simple transfusions (r=0.316, P=0.108). The total number of LTE procedures was not associated with serum ferritin (r=0.040, P=0.838), the total number of simple transfusions (r=-0.258, P=0.184), or LIC group (r=-0.111, P=0.566). There was no significant correlation between duration of LTE maintenance and LIC.

Influence of laser and LED irradiation on mast cells of cutaneous wounds of rats with iron deficiency anemia

NASA Astrophysics Data System (ADS)

Becher Rosa, Cristiane; Oliveira Sampaio, Susana C. P.; Monteiro, Juliana S. C.; Ferreira, Maria F. L.; Zanini, Fátima A. A.; Santos, Jean N.; Cangussú, Maria Cristina T.; Pinheiro, Antonio L. B.

2011-03-01

This work aimed to study histologically the effect of Laser or LED phototherapy on mast cells on cutaneous wounds of rats with iron deficiency. 18 rats were used and fed with special peleted iron-free diet. An excisional wound was created on the dorsum of each animal which were divided into: Group I - Control with anemia + no treatment; Group II - Anemia + Laser; Group III - Anemia + LED; Group IV - Healthy + no treatment; Group V - Healthy + Laser; Group VI - Healthy + LED. Irradiation was performed using a diode Laser (λ660nm, 40mW, CW, total dose of 10J/cm2, 4X2.5J/cm2) or a RED-LED ( λ700nm, 15mW, CW, total dose of 10J/cm2). Histological specimens were routinely processed, cut and stained with toluidine blue and mast cell counts performed. No significant statistic difference was found between groups as to the number of degranulated, non-degradulated or total mast cells. Greater mean values were found for degranulated mast cells in the Anemia + LED. LED irradiation on healthy specimens resulted in a smaller number of degranulated mast cells. Our results leads to conclude that there are no significant differences in the number of mast cells seven days after irradiation following Laser or LED phototherapy.

Three-Dimensional Analysis of Peeled Internal Limiting Membrane Using Focused Ion Beam/Scanning Electron Microscopy

PubMed Central

Murata, Kazuhisa; Hayashi, Ken; Nakamura, Kei-ichiro

2018-01-01

Purpose To reevaluate the effect of internal limiting membrane peeling during vitrectomy on the Müller cell damage, we examined the ultrastructure of the internal limiting membrane by using focused ion beam/scanning electron microscopy (FIB/SEM). Methods A total of 12 internal limiting membranes obtained during surgery in both the macular hole and the idiopathic epiretinal membrane groups were processed for observation by FIB/SEM. Three-dimensional structures of the internal limiting membrane were analyzed. Results The number of cell fragments in the macular hole group was 5.07 ± 1.03 per unit area of internal limiting membrane (100 μm2). The total volume of cell fragments was 3.54 ± 1.24 μm3/100 μm2. In contrast, the number of cell fragments in the epiretinal membrane group was 12.85 ± 3.45/100 μm2, and the total volume of cell fragments was 10.45 ± 2.77 μm3/100 μm2. Data for both values were significantly higher than those observed in the macular hole group (P = 0.0024 and P = 0.0022, respectively, Mann-Whitney U test). No statistical difference was found for the mean volume of the cell fragment between the two groups. Conclusions All of the internal limiting membrane examined in this study showed cell fragments on the retinal surface of the internal limiting membrane. As compared with macular hole, epiretinal membrane exhibited a higher number and total volume of cell fragments, indicating that internal limiting membrane peeling for epiretinal membrane might have a higher risk of causing inner retinal damage. Translational Relevance FIB/SEM was a useful tool for three-dimensional quantitative analysis of the internal limiting membrane. PMID:29423341

A Low Power Linear Phase Programmable Long Delay Circuit.

PubMed

Rodriguez-Villegas, Esther; Logesparan, Lojini; Casson, Alexander J

2014-06-01

A novel linear phase programmable delay is being proposed and implemented in a 0.35 μm CMOS process. The delay line consists of N cascaded cells, each of which delays the input signal by Td/N, where Td is the total line delay. The delay generated by each cell is programmable by changing a clock frequency and is also fully independent of the frequency of the input signal. The total delay hence depends only on the chosen clock frequency and the total number of cascaded cells. The minimum clock frequency is limited by the maximum time a voltage signal can effectively be held by an individual cell. The maximum number of cascaded cells will be limited by the effects of accumulated offset due to transistor mismatch, which eventually will affect the operating mode of the individual transistors in a cell. This latter limitation has however been dealt with in the topology by having an offset compensation mechanism that makes possible having a large number of cascaded cells and hence a long resulting delay. The delay line has been designed for scalp-based neural activity analysis that is predominantly in the sub-100 Hz frequency range. For these signals, the delay generated by a 31-cell cascade has been demonstrated to be programmable from 30 ms to 3 s. Measurement results demonstrate a 31 stage, 50 Hz bandwidth, 0.3 s delay that operates from a 1.1 V supply with power consumption of 270 nW.

CELL RESPIRATION STUDIES

PubMed Central

Daland, Geneva A.; Isaacs, Raphael

1927-01-01

1. The oxygen consumption of blood of normal individuals, when the hemoglobin is saturated with oxygen, is practically zero within the limits of experimental error of the microspirometer used. 2. The oxygen consumed in a microspirometer by the blood of patients with chronic myelogenous leucemia with a high white blood cell count, and of one with leucocytosis from sepsis, was proportional to the number of adult polymorphonuclear neutrophils in the blood. 3. No correlation could be made between the rate of oxygen absorption and the total number of white blood cells in the blood, or the total number of immature cells, or the number of red blood cells, or the amount of oxyhemoglobin. 4. The blood of patients with chronic myelogenous leucemia continued to use oxygen in the microspirometer longer than that of normal individuals, and the hemoglobin, in the leucemic bloods, became desaturated even though exposed to air. 5. In blood in which the bulk. of the cells were immature and the mature cells few, the oxygen consumption was lower than in blood in which the mature cells predominated. The rate of oxygen consumption of the immature cells was relatively low as compared to the mature. 6. The slower rate of oxygen absorption by the immature leucocytes in chronic myelogenous leucemia as compared to the mature cells, places them, in accord with Warburg's reports, in the class of the malignant tissues in this respect rather than in the group of young or embryonic cells. PMID:19869329

Effects of donor fibroblast cell type and transferred cloned embryo number on the efficiency of pig cloning.

PubMed

Li, Zicong; Shi, Junsong; Liu, Dewu; Zhou, Rong; Zeng, Haiyu; Zhou, Xiu; Mai, Ranbiao; Zeng, Shaofen; Luo, Lvhua; Yu, Wanxian; Zhang, Shouquan; Wu, Zhenfang

2013-02-01

Currently, cloning efficiency in pigs is very low. Donor cell type and number of cloned embryos transferred to an individual surrogate are two major factors that affect the successful rate of somatic cell nuclear transfer (SCNT) in pigs. This study aimed to compare the influence of different donor fibroblast cell types and different transferred embryo numbers on recipients' pregnancy rate and delivery rate, the average number of total clones born, clones born alive and clones born healthy per litter, and the birth rate of healthy clones (=total number of healthy cloned piglets born /total number of transferred cloned embryos). Three types of donor fibroblasts were tested in large-scale production of cloned pigs, including fetal fibroblasts (FFBs) from four genetically similar Western swine breeds of Pietrain (P), Duroc (D), Landrace (L), and Yorkshire (Y), which are referred to as P,D,LY-FFBs, adult fibroblasts (AFBs) from the same four breeds, which are designated P,D,L,Y-AFBs, and AFBs from a Chinese pig breed of Laiwu (LW), which is referred to as LW-AFBs. Within each donor fibroblast cell type group, five transferred cloned embryo number groups were tested. In each embryo number group, 150-199, 200-249, 250-299, 300-349, or 350-450 cloned embryos were transferred to each individual recipient sow. For the entire experiment, 92,005 cloned embryos were generated from nearly 115,000 matured oocytes and transferred to 328 recipients; in total, 488 cloned piglets were produced. The results showed that the mean clones born healthy per litter resulted from transfer of embryos cloned from LW-AFBs (2.53 ± 0.34) was similar with that associated with P,D,L,Y-FFBs (2.72 ± 0.29), but was significantly higher than that resulted from P,D,L,Y-AFBs (1.47 ± 0.18). Use of LW-AFBs as donor cells for SCNT resulted in a significantly higher pregnancy rate (72.00% vs. 59.30% and 48.11%) and delivery rate (60.00% vs. 45.93% and 35.85%) for cloned embryo recipients, and a significantly higher birth rate of healthy clones (0.5009% vs. 0.3362% and 0.2433%) than that resulting from P,D,L,Y-AFBs and P,D,L,Y-FFBs. This suggests that using LW-AFBs as donor cells results in a higher cloning efficiency in pigs, compared with the other two donor fibroblast cell types. The birth rate of healthy clones was significantly improved when the number of transferred cloned embryos was increased from 150-199 to 200-450 per recipient. However, increase of the number of transferred embryos from 200-249 to 250-450 per surrogate did not change the birth rate of healthy clones. This suggests that transfer of excessive (250-450) cloned embryos to an individual surrogate is not necessary for increasing the cloning efficiency in pigs, and the relatively optimal number of reconstructed embryos transferred to individual recipient is 200-249. Furthermore, our results indicated that the numbers of total born clones, clones born alive, and clones born healthy per litter have a significantly high positive correlation with each other. The present study provides useful information for improving SCNT efficiency in pigs.

Effects of Donor Fibroblast Cell Type and Transferred Cloned Embryo Number on the Efficiency of Pig Cloning

PubMed Central

Li, Zicong; Shi, Junsong; Liu, Dewu; Zhou, Rong; Zeng, Haiyu; Zhou, Xiu; Mai, Ranbiao; Zeng, Shaofen; Luo, Lvhua; Yu, Wanxian; Zhang, Shouquan

2013-01-01

Abstract Currently, cloning efficiency in pigs is very low. Donor cell type and number of cloned embryos transferred to an individual surrogate are two major factors that affect the successful rate of somatic cell nuclear transfer (SCNT) in pigs. This study aimed to compare the influence of different donor fibroblast cell types and different transferred embryo numbers on recipients' pregnancy rate and delivery rate, the average number of total clones born, clones born alive and clones born healthy per litter, and the birth rate of healthy clones (=total number of healthy cloned piglets born /total number of transferred cloned embryos). Three types of donor fibroblasts were tested in large-scale production of cloned pigs, including fetal fibroblasts (FFBs) from four genetically similar Western swine breeds of Pietrain (P), Duroc (D), Landrace (L), and Yorkshire (Y), which are referred to as P,D,LY-FFBs, adult fibroblasts (AFBs) from the same four breeds, which are designated P,D,L,Y-AFBs, and AFBs from a Chinese pig breed of Laiwu (LW), which is referred to as LW-AFBs. Within each donor fibroblast cell type group, five transferred cloned embryo number groups were tested. In each embryo number group, 150–199, 200–249, 250–299, 300–349, or 350–450 cloned embryos were transferred to each individual recipient sow. For the entire experiment, 92,005 cloned embryos were generated from nearly 115,000 matured oocytes and transferred to 328 recipients; in total, 488 cloned piglets were produced. The results showed that the mean clones born healthy per litter resulted from transfer of embryos cloned from LW-AFBs (2.53±0.34) was similar with that associated with P,D,L,Y-FFBs (2.72±0.29), but was significantly higher than that resulted from P,D,L,Y-AFBs (1.47±0.18). Use of LW-AFBs as donor cells for SCNT resulted in a significantly higher pregnancy rate (72.00% vs. 59.30% and 48.11%) and delivery rate (60.00% vs. 45.93% and 35.85%) for cloned embryo recipients, and a significantly higher birth rate of healthy clones (0.5009% vs. 0.3362% and 0.2433%) than that resulting from P,D,L,Y-AFBs and P,D,L,Y-FFBs. This suggests that using LW-AFBs as donor cells results in a higher cloning efficiency in pigs, compared with the other two donor fibroblast cell types. The birth rate of healthy clones was significantly improved when the number of transferred cloned embryos was increased from 150–199 to 200–450 per recipient. However, increase of the number of transferred embryos from 200–249 to 250–450 per surrogate did not change the birth rate of healthy clones. This suggests that transfer of excessive (250–450) cloned embryos to an individual surrogate is not necessary for increasing the cloning efficiency in pigs, and the relatively optimal number of reconstructed embryos transferred to individual recipient is 200–249. Furthermore, our results indicated that the numbers of total born clones, clones born alive, and clones born healthy per litter have a significantly high positive correlation with each other. The present study provides useful information for improving SCNT efficiency in pigs. PMID:23256540

Decrease in Numbers of Naive and Resting B Cells in HIV-Infected Kenyan Adults Leads to a Proportional Increase in Total and Plasmodium falciparum-Specific Atypical Memory B Cells.

PubMed

Frosch, Anne E; Odumade, Oludare A; Taylor, Justin J; Ireland, Kathleen; Ayodo, George; Ondigo, Bartholomew; Narum, David L; Vulule, John; John, Chandy C

2017-06-15

Human immunodeficiency virus type 1 (HIV-1) infection is associated with B cell activation and exhaustion, and hypergammaglobulinemia. How these changes influence B cell responses to coinfections such as malaria is poorly understood. To address this, we compared B cell phenotypes and Abs specific for the Plasmodium falciparum vaccine candidate apical membrane Ag-1 (AMA1) in HIV-infected and uninfected adults living in Kenya. Surprisingly, HIV-1 infection was not associated with a difference in serum AMA1-specific Ab levels. HIV-infected individuals had a higher proportion of total atypical and total activated memory B cells (MBCs). Using an AMA1 tetramer to detect AMA1-specific B cells, HIV-infected individuals were also shown to have a higher proportion of AMA1-specific atypical MBCs. However, this proportional increase resulted in large part from a loss in the number of naive and resting MBCs rather than an increase in the number of atypical and activated cells. The loss of resting MBCs and naive B cells was mirrored in a population of cells specific for an Ag to which these individuals were unlikely to have been chronically exposed. Together, the data show that changes in P. falciparum Ag-specific B cell subsets in HIV-infected individuals mirror those in the overall B cell population, and suggest that the increased proportion of atypical MBC phenotypes found in HIV-1-infected individuals results from the loss of naive and resting MBCs. Copyright © 2017 by The American Association of Immunologists, Inc.

Forskolin improves the cryosurvival of in vivo-derived porcine embryos at very early stages using two vitrification methods.

PubMed

Gomis, J; Cuello, C; Sanchez-Osorio, J; Gil, M A; Parrilla, I; Angel, M A; Vazquez, J M; Roca, J; Martinez, E A

2013-04-01

This study was aimed to determine the effect of forskolin on the viability of in vivo-derived porcine embryos vitrified by the superfine open pulled straw (SOPS) or solid surface vitrification (SSV) methods at the 2-cell, 4-cell, and blastocyst stages. Zygotes, 2- to 4-cell embryos, and morulae were obtained from superovulated sows. After collection, embryos were cultured for 24h with 0 or 10 μM forskolin and then vitrified using the SOPS and SSV method, or not vitrified (fresh controls). Fresh and vitrified-warmed 2-cells, 4-cells, and blastocysts were cultured for additional 96 h, 72 h and 24 h, respectively. At the end of the culture, embryos were evaluated for progression to the blastocyst stage and total cell number. The vitrification method did not affect any of the parameters evaluated for any embryo stage. Forskolin increased (P<0.01) the blastocyst formation and the final developmental stage of vitrified 2- and 4-cell embryos. However, these embryos exhibited lower (P<0.003) blastocyst formation rates than their fresh counterparts. The total cell number and hatching rate were similar in both groups (vitrified and fresh) of 2- and 4-cell embryos. Vitrified blastocysts exhibited viabilities, final developmental stages, hatching rates, and total cell numbers that were similar to those of their fresh counterparts, regardless of the addition of forskolin. In conclusion, the SOPS and SSV methods are suitable for the cryopreservation of in vivo-derived 2- to 4-cell porcine embryos. Pre-treatment with forskolin for 24h before vitrification improves the cryotolerance of 2- and 4-cell porcine embryos. Copyright © 2013 Elsevier Inc. All rights reserved.

[Response of HeLa cells to mitomycine C. III. The analysis of nucleoli of mother and daughter cells].

PubMed

Petrov, Iu P; Neguliaev, Iu A; Tsupkina, N V

2014-01-01

The comparative analysis of the number of nucleoli in cells of the established HeLa-M line was carried out before and after exposure to mitomycin C in a concentration of 10 μg/ml for 2 h. Using time-lapse microscopy, nucleoli in mother and their respective daughter cells were computed. It has been shown that the average number of nucleoli per cell is generally higher in daughter cells than in mother cells, and a standard deviation, on the contrary, decreases. An average number of nucleoli in daughter cells, whose mother cells had been treated with mitomycin C, was higher than in corresponding cells of control group. The separate analysis has been performed for the cells having from 1 to 4 nucleoli. Nonrandom complete coincidence of the number of nucleoli in mather and daughter cells has been typicaly shown for about 1/7 of the total cell population. Mitomycin C reduces this value of about 1.5 times.

SIDR: simultaneous isolation and parallel sequencing of genomic DNA and total RNA from single cells.

PubMed

Han, Kyung Yeon; Kim, Kyu-Tae; Joung, Je-Gun; Son, Dae-Soon; Kim, Yeon Jeong; Jo, Areum; Jeon, Hyo-Jeong; Moon, Hui-Sung; Yoo, Chang Eun; Chung, Woosung; Eum, Hye Hyeon; Kim, Sangmin; Kim, Hong Kwan; Lee, Jeong Eon; Ahn, Myung-Ju; Lee, Hae-Ock; Park, Donghyun; Park, Woong-Yang

2018-01-01

Simultaneous sequencing of the genome and transcriptome at the single-cell level is a powerful tool for characterizing genomic and transcriptomic variation and revealing correlative relationships. However, it remains technically challenging to analyze both the genome and transcriptome in the same cell. Here, we report a novel method for simultaneous isolation of genomic DNA and total RNA (SIDR) from single cells, achieving high recovery rates with minimal cross-contamination, as is crucial for accurate description and integration of the single-cell genome and transcriptome. For reliable and efficient separation of genomic DNA and total RNA from single cells, the method uses hypotonic lysis to preserve nuclear lamina integrity and subsequently captures the cell lysate using antibody-conjugated magnetic microbeads. Evaluating the performance of this method using real-time PCR demonstrated that it efficiently recovered genomic DNA and total RNA. Thorough data quality assessments showed that DNA and RNA simultaneously fractionated by the SIDR method were suitable for genome and transcriptome sequencing analysis at the single-cell level. The integration of single-cell genome and transcriptome sequencing by SIDR (SIDR-seq) showed that genetic alterations, such as copy-number and single-nucleotide variations, were more accurately captured by single-cell SIDR-seq compared with conventional single-cell RNA-seq, although copy-number variations positively correlated with the corresponding gene expression levels. These results suggest that SIDR-seq is potentially a powerful tool to reveal genetic heterogeneity and phenotypic information inferred from gene expression patterns at the single-cell level. © 2018 Han et al.; Published by Cold Spring Harbor Laboratory Press.

SIDR: simultaneous isolation and parallel sequencing of genomic DNA and total RNA from single cells

PubMed Central

Han, Kyung Yeon; Kim, Kyu-Tae; Joung, Je-Gun; Son, Dae-Soon; Kim, Yeon Jeong; Jo, Areum; Jeon, Hyo-Jeong; Moon, Hui-Sung; Yoo, Chang Eun; Chung, Woosung; Eum, Hye Hyeon; Kim, Sangmin; Kim, Hong Kwan; Lee, Jeong Eon; Ahn, Myung-Ju; Lee, Hae-Ock; Park, Donghyun; Park, Woong-Yang

2018-01-01

Simultaneous sequencing of the genome and transcriptome at the single-cell level is a powerful tool for characterizing genomic and transcriptomic variation and revealing correlative relationships. However, it remains technically challenging to analyze both the genome and transcriptome in the same cell. Here, we report a novel method for simultaneous isolation of genomic DNA and total RNA (SIDR) from single cells, achieving high recovery rates with minimal cross-contamination, as is crucial for accurate description and integration of the single-cell genome and transcriptome. For reliable and efficient separation of genomic DNA and total RNA from single cells, the method uses hypotonic lysis to preserve nuclear lamina integrity and subsequently captures the cell lysate using antibody-conjugated magnetic microbeads. Evaluating the performance of this method using real-time PCR demonstrated that it efficiently recovered genomic DNA and total RNA. Thorough data quality assessments showed that DNA and RNA simultaneously fractionated by the SIDR method were suitable for genome and transcriptome sequencing analysis at the single-cell level. The integration of single-cell genome and transcriptome sequencing by SIDR (SIDR-seq) showed that genetic alterations, such as copy-number and single-nucleotide variations, were more accurately captured by single-cell SIDR-seq compared with conventional single-cell RNA-seq, although copy-number variations positively correlated with the corresponding gene expression levels. These results suggest that SIDR-seq is potentially a powerful tool to reveal genetic heterogeneity and phenotypic information inferred from gene expression patterns at the single-cell level. PMID:29208629

The effect of interferon on the receptor sites to rabies virus on mouse neuroblastoma cells

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

Briggs, D.J.

1989-01-01

The binding of rabies virus to mouse neuroblastoma cells (MNA) primed with alpha interferon (IFN-{alpha}), beta interferon (IFN-{beta}), or alpha bungarotoxin (BTX) was examined. A saturable number of receptor sites to rabies virus was calculated by increasing the amount of {sup 3}H-CVS added to a constant number of untreated MNA cells. MNA cells were then exposed to 20 I.U. of IFN-{alpha}, IFN-{beta}, or 1 {mu}g of BTX and assayed to determine if these treatments had an effect on the number of receptor sites to rabies virus. Total amount of {sup 3}H-CVS bound to MNA cells was determined during a threemore » hour incubation period. Cold competition assays using 1,000 fold excess unlabeled CVS were used to determine non-specific binding for each treatment. Specific binding was then calculated by subtracting non-specific binding from the total amount of CVS bound to MNA cells. A similar amount of total viral protein bound to untreated and IFN-{beta}, and BTX treated cells after 180 minutes of incubation. The bound protein varied by only 0.07 {mu}g. However, the amount of specific and non-specific binding varied a great deal between treatments. BTX caused an increase in non-specific and a decrease in specific binding of rabies virus. IFN-{beta} produced variable results in non-specific and specific binding while IFN-{alpha} caused mainly specific binding to occur. The most significant change brought about by IFN-{alpha} was an increase in the rate of viral attachment. At 30 minutes post-infection, IFN-{alpha} treated cells had bound 90% of the total amount of virus bound to untreated cells after 180 minutes. The increased binding rate did not cause a productive infection of rabies virus. No viral production was evident after an incubation period of 48 hours in either IFN-{alpha} or IFN-{beta} treated cells.« less

White Nail Radio Transmitter: Billion Dollar Savings through Energy Efficiency

DTIC Science & Technology

2011-05-10

increase efficiency and reduce overall energy consumption ashore by 50 percent CNO, Navy Energy Vision, P 10 White Nail Vision Your Cell Phone Cell...Estimated Total Number of transmitters 3,000,000 Estimated total power saved Watt 1,250,000,000 Cell Phone Transmitter Efficiency 1.25 Gigawatts saved...Greenhouse Gas Power 4 1 Energy Navy Use 7.3 Billion kWh White Nail Cell Phone Savings 11 Billion kWh One and a half times!!! Saves the output of four of

The humoral immune response of mice exposed to manual metal arc stainless steel-welding fumes.

PubMed

Anderson, Stacey E; Meade, B Jean; Butterworth, Leon F; Munson, Albert E

2007-01-01

Arc welding is one of the most common forms of welding and includes the use of stainless steel electrodes that emit fumes containing chromium and nickel. Epidemological studies suggest a correlation between arc welding and adverse respiratory health effects. Studies evaluating the immunotoxic effects of welding fumes are limited due to the large number of variables associated with welding. This work investigates the immunotoxic effects of welding fumes by analyzing the in vivo and in vitro IgM response to a T-dependent antigen after welding fume exposure. Significant decreases in the total IgM activity/10(6) viable cells and total IgM activity/well were observed in splenocytes exposed to 5 mu g/ml of either total or soluble welding fumes. A significant reduction in the specific IgM activity in lung associated lymph node cells was also observed following four pharyngeal aspirations of 10 mg/kg total or soluble welding fumes to mice. Significant elevations in the absolute lymph node cell numbers for both B- and T-cells including the CD4(+) and CD8(+) subsets were observed. These results demonstrate that exposure to manual metal-stainless steel welding fumes is immunosuppressive in the presence of increased lymphoctye numbers in mice and raises concerns regarding the potential for adverse immunological effects to impact respiratory health in humans.

Zinc supplements for treating thalassaemia and sickle cell disease.

PubMed

Swe, Kye Mon Min; Abas, Adinegara B L; Bhardwaj, Amit; Barua, Ankur; Nair, N S

2013-06-28

Haemoglobinopathies, inherited disorders of haemoglobin synthesis (thalassaemia) or structure (sickle cell disease), are responsible for significant morbidity and mortality throughout the world. The WHO estimates that, globally, 5% of adults are carriers of a haemoglobin condition, 2.9% are carriers of thalassaemia and 2.3% are carriers of sickle cell disease. Carriers are found worldwide as a result of migration of various ethnic groups to different regions of the world. Zinc is an easily available supplement and intervention programs have been carried out to prevent deficiency in people with thalassaemia or sickle cell anaemia. It is important to evaluate the role of zinc supplementation in the treatment of thalassaemia and sickle cell anaemia to reduce deaths due to complications. To assess the effect of zinc supplementation in the treatment of thalassaemia and sickle cell disease. We searched the Cochrane Cystic Fibrosis and Genetic Disorders Group's Haemoglobinopathies Trials Register comprising references identified from comprehensive electronic database searches and handsearches of relevant journals and abstract books of conference proceedings.Date of most recent search: 01 February 2013. Randomised, placebo-controlled trials of zinc supplements for treating thalassaemia or sickle cell disease administered at least once a week for at least a month. Two review authors assessed the eligibility and risk of bias of the included trials, extracted and analysed data and wrote the review. We summarised results using risk ratios or rate ratios for dichotomous data and mean differences for continuous data. We combined trial results where appropriate. We identified nine trials for inclusion with all nine contributing outcome data. Two trials reported on people with thalassaemia (n = 152) and seven on sickle cell anaemia (n = 307).In people with thalassaemia, in one trial, the serum zinc level value showed no difference between the zinc supplemented group and the control group, mean difference 47.40 (95% confidence interval -12.95 to 107.99). Regarding anthropometry, in one trial, height velocity was significantly increased in patients who received zinc supplementation for one to seven years duration, mean difference 3.37 (95% confidence interval 2.36 to 4.38) (total number of participants = 26). In one trial, however, there was no difference in body mass index between treatment groups.Zinc acetate supplementation for three months (in one trial) and one year (in two trials) (total number of participants = 71) was noted to increase the serum zinc level significantly in patients with sickle cell anaemia, mean difference 14.90 (95% confidence interval 6.94 to 22.86) and 20.25 (95% confidence interval 11.73 to 28.77) respectively. There was no significant difference in haemoglobin level between intervention and control groups, at either three months (one trial) or one year (one trial), mean difference 0.06 (95% confidence interval -0.84 to 0.96) and mean difference -0.07 (95% confidence interval -1.40 to 1.26) respectively. Regarding anthropometry, one trial showed no significant changes in body mass index or weight after one year of zinc acetate supplementation. In patients with sickle cell disease, the total number of sickle cell crises at one year were significantly decreased in the zinc sulphate supplemented group as compared to controls, mean difference -2.83 (95% confidence interval -3.51 to -2.15) (total participants 130), but not in zinc acetate group, mean difference 1.54 (95% confidence interval -2.01 to 5.09) (total participants 22). In one trial at three months and another at one year, the total number of clinical infections were significantly decreased in the zinc supplemented group as compared to controls, mean difference 0.05 (95% confidence interval 0.01 - 0.43) (total number of participants = 36), and mean difference -7.64 (95% confidence interval -10.89 to -4.39) (total number of participants = 21) respectively. According to the results, there is no evidence from randomised controlled trials to indicate any benefit of zinc supplementation with regards to serum zinc level in patients with thalassaemia. However, height velocity was noted to increase among those who received this intervention.There is mixed evidence on the benefit of using zinc supplementation in people with sickle cell disease. For instance, there is evidence that zinc supplementation for one year increased the serum zinc levels in patients with sickle cell disease. However, though serum zinc level was raised in patients receiving zinc supplementation, haemoglobin level and anthropometry measurements were not significantly different between groups. Evidence of benefit is seen with the reduction in the number of sickle cell crises among sickle cell patients who received one year of zinc sulphate supplementation and with the reduction in the total number of clinical infections among sickle cell patients who received zinc supplementation for both three months and for one year.The conclusion is based on the data from a small group of trials,which were generally of good quality, with a low risk of bias. The authors recommend that more trials on zinc supplementation in thalassaemia and sickle cell disease be conducted given that the literature has shown the benefits of zinc in these types of diseases.

CELL RESPIRATION STUDIES : II. A COMPARATIVE STUDY OF THE OXYGEN CONSUMPTION OF BLOOD FROM NORMAL INDIVIDUALS AND PATIENTS WITH INCREASED LEUCOCYTE COUNTS (SEPSIS; CHRONIC MYELOGENOUS LEUCEMIA).

PubMed

Daland, G A; Isaacs, R

1927-06-30

1. The oxygen consumption of blood of normal individuals, when the hemoglobin is saturated with oxygen, is practically zero within the limits of experimental error of the microspirometer used. 2. The oxygen consumed in a microspirometer by the blood of patients with chronic myelogenous leucemia with a high white blood cell count, and of one with leucocytosis from sepsis, was proportional to the number of adult polymorphonuclear neutrophils in the blood. 3. No correlation could be made between the rate of oxygen absorption and the total number of white blood cells in the blood, or the total number of immature cells, or the number of red blood cells, or the amount of oxyhemoglobin. 4. The blood of patients with chronic myelogenous leucemia continued to use oxygen in the microspirometer longer than that of normal individuals, and the hemoglobin, in the leucemic bloods, became desaturated even though exposed to air. 5. In blood in which the bulk. of the cells were immature and the mature cells few, the oxygen consumption was lower than in blood in which the mature cells predominated. The rate of oxygen consumption of the immature cells was relatively low as compared to the mature. 6. The slower rate of oxygen absorption by the immature leucocytes in chronic myelogenous leucemia as compared to the mature cells, places them, in accord with Warburg's reports, in the class of the malignant tissues in this respect rather than in the group of young or embryonic cells.

The effect of protein-coated contact lenses on the adhesion and viability of gram negative bacteria.

PubMed

Williams, Timothy J; Schneider, Rene P; Willcox, Mark D P

2003-10-01

Gram negative bacterial adhesion to contact lenses can cause adverse responses. During contact lens wear, components of the tear film adsorb to the contact lens. This study aimed to investigate the effect of this conditioning film on the viability of bacteria. Bacteria adhered to contact lenses which were either unworn, worn for daily-, extended- or overnight-wear or coated with lactoferrin or lysozyme. Numbers of viable and total cells were estimated. The number of viable attached cells was found to be significantly lower than the total number of cells on worn (50% for strain Paer1 on daily-wear lenses) or lactoferrin-coated lenses (56% for strain Paer1). Lysozyme-coated lenses no statistically significant effect on adhesion. The conditioning film gained through wear may not inhibit bacterial adhesion, but may act adversely upon those bacteria that succeed in attaching.

ACEA (a highly selective cannabinoid CB1 receptor agonist) stimulates hippocampal neurogenesis in mice treated with antiepileptic drugs.

PubMed

Andres-Mach, Marta; Haratym-Maj, Agnieszka; Zagaja, Miroslaw; Rola, Radoslaw; Maj, Maciej; Chrościńska-Krawczyk, Magdalena; Luszczki, Jarogniew J

2015-10-22

Hippocampal neurogenesis plays a very important role in learning and memory functions. In a search for best neurological drugs that protect neuronal cells and stimulate neurogenesis with no side effects, cannabinoids proved to be a strong group of substances having many beneficial properties. The aim of this study was to evaluate the impact of ACEA (arachidonyl-2'-chloroethylamide--a highly selective cannabinoid CB1 receptor agonist) combined with a classical antiepileptic drug sodium valproate (VPA) on neural precursor cells' proliferation and differentiation in the mouse brain. All experiments were performed on adolescent CB57/BL male mice injected i.p. with VPA (10mg/kg), ACEA (10mg/kg) and PMSF (30 mg/kg) (phenylmethylsulfonyl fluoride--a substance protecting ACEA against degradation by the fatty-acid amidohydrolase) for 10 days. Next an acute response of proliferating neural precursor cells to ACEA and VPA administration was evaluated with Ki-67 staining (Time point 1). Next, in order to determine whether acute changes translated into long-term alterations in neurogenesis, proliferating cells were labeled with 5-bromo-2deoxyuridine (BrdU) followed by confocal microscopy used to determine the percentage of BrdU-labeled cells that showed mature cell phenotypes (Time point 2). Results indicate that ACEA with PMSF significantly increase the total number of Ki-67-positive cells when compared to the control group. Moreover, ACEA in combination with VPA increased the number of Ki-67-positive cells, whereas VPA administered alone had no impact on proliferating cells' population. Accordingly, neurogenesis study results indicate that the combination of ACEA+PMSF administered alone and in combination with VPA considerably increases the total number of BrdU-positive cells in comparison to the control group while ACEA+PMSF alone and in combination with VPA increased total numbers of BrdU-positive cells, newly born neurons and astrocytes as compared to VPA group but not to the control group. VPA administered alone decreased the number of newly born neurons with no significant impact on neurogenesis. These data provide substantial evidence that VPA administered chronically slightly decreases the proliferation and differentiation of newly born cells while combination of VPA+ACEA significantly increases the level of newborn neurons in the dentate subgranular zone. Copyright © 2015 Elsevier B.V. All rights reserved.

Receptor activator NFkappaB-ligand and osteoprotegerin protein expression in human periapical cysts and granulomas.

PubMed

Menezes, Renato; Bramante, Clóvis Monteiro; da Silva Paiva, Katiúcia Batista; Letra, Ariadne; Carneiro, Everdan; Fernando Zambuzzi, Willian; Granjeiro, José Mauro

2006-09-01

The purpose of this study was to determine the expression of receptor activator of NFkappaB ligand (RANKL) and osteoprotegerin (OPG) associated with bone destruction in periapical cysts and granulomas. Forty human dental chronic periapical lesions were collected after periapical surgery. The lesions collected were fixed in 10% buffered formalin and histologically processed. At least 2 sections of each specimen were stained with hematoxylin and eosin for microscopic diagnosis. After that, 10 human periapical granulomas and 10 cysts were selected for immunohistochemical analysis for RANKL, OPG, and CD68+. Polymorphonuclear neutrophils, macrophages, endothelial cells, and lymphocytes were stained for RANKL and OPG in both lesions. Epithelial cells were also stained for RANKL and OPG in periapical cysts. Quantitative analysis was conducted and the results were expressed as a ratio of the number of immunostained cells over the total number of cells in the field (n = 100). The ratio of RANKL+/total cells was higher than OPG+/total cells in periapical granulomas (0.553 +/- 0.153 and 0.483 +/- 0.189, respectively; P < .0012; paired t test) and in cysts (0.519 +/- 0.09 and 0.339 +/- 0.117, respectively; P < .0001; paired t test). The ratios of OPG+/total cells (P < .0001; paired t test) and RANKL+/total cells (P < .0322; paired t test) were greater in granulomas than in cysts. However, the ratio RANKL+/OPG+ in granulomas (1.336 +/- 0.723) and cysts (1.404 +/- 0.385) was not significantly different. The ratio of CD68+/total cells was significantly higher in granulomas (0.381 +/- 0.040) than in cysts (0.307 +/- 0.068) (P < .0001; unpaired t test with Welch correction). Taking into account the limitations of the experimental approach employed, our findings indicate the presence of RANKL and OPG in cysts and granulomas, strongly suggesting the involvement of these gene products in the development of periapical lesions.

Rapid staining and enumeration of small numbers of total bacteria in water by solid-phase laser cytometry

NASA Technical Reports Server (NTRS)

Broadaway, Susan C.; Barton, Stephanie A.; Pyle, Barry H.

2003-01-01

The nucleic acid stain SYBR Green I was evaluated for use with solid-phase laser cytometry to obtain total bacterial cell counts from several water sources with small bacterial numbers. Results were obtained within 30 min and exceeded or equaled counts on R2A agar plates incubated for 14 days at room temperature.

Estimation of neuronal numbers in rat hippocampus following neonatal amphetamine exposure: a stereology study.

PubMed

Smith, Andrew M; Pappalardo, Dana; Chen, Wei-Jung A

2008-01-01

In this study, the effects of amphetamine exposure during a portion of the brain growth spurt on the total number of hippocampal pyramidal cells (CA1/CA3 subregions) and the granule cells (dentate gyrus) were examined in both neonatal and adult rats. Intragastric intubation was used to administer 5, 15 or 25 mg/kg/day of amphetamine to Sprague-Dawley rat pups from PDs 4-9. Unbiased stereology was used to estimate the total number of cells present within each hippocampal subregion at both PD 9 and PD 68. The results indicated that neonatal amphetamine exposure did not alter the cell number, the reference volume or the density in any of the hippocampal subregions assessed, regardless of age. However, amphetamine significantly altered the rate of neuronal incorporation in both the hippocampal CA3 subregion and the dentate gyrus, and this effect appeared to be dose-related with the most robust effect observed in the highest amphetamine dose. While these findings did not demonstrate significant injurious effects of neonatal amphetamine treatment on the number of hippocampal neurons, these data suggest that amphetamine may interfere with proper hippocampal development. Future studies employing more sensitive measurements or exposing amphetamine during an alternate period of development may provide more information regarding amphetamine-mediated developmental neurotoxicity.

Application of Real-Time PCR for Quantification of Microcystin Genotypes in a Population of the Toxic Cyanobacterium Microcystis sp.

PubMed Central

Kurmayer, Rainer; Kutzenberger, Thomas

2003-01-01

The cyanobacterium Microcystis sp. frequently develops water blooms consisting of organisms with different genotypes that either produce or lack the hepatotoxin microcystin. In order to monitor the development of microcystin (mcy) genotypes during the seasonal cycle of the total population, mcy genotypes were quantified by means of real-time PCR in Lake Wannsee (Berlin, Germany) from June 1999 to October 2000. Standard curves were established by relating cell concentrations to the threshold cycle (the PCR cycle number at which the fluorescence passes a set threshold level) determined by the Taq nuclease assay (TNA) for two gene regions, the intergenic spacer region within the phycocyanin (PC) operon to quantify the total population and the mcyB gene, which is indicative of microcystin synthesis. In laboratory batch cultures, the cell numbers inferred from the standard curve by TNA correlated significantly with the microscopically determined cell numbers on a logarithmic scale. The TNA analysis of 10 strains revealed identical amplification efficiencies for both genes. In the field, the proportion of mcy genotypes made up the smaller part of the PC genotypes, ranging from 1 to 38%. The number of mcyB genotypes was one-to-one related to the number of PC genotypes, and parallel relationships between cell numbers estimated via the inverted microscope technique and TNA were found for both genes. It is concluded that the mean proportion of microcystin genotypes is stable from winter to summer and that Microcystis cell numbers could be used to infer the mean proportion of mcy genotypes in Lake Wannsee. PMID:14602633

Total knee replacement induces peripheral blood lymphocytes apoptosis and it is not prevented by regional anesthesia - a randomized study.

PubMed

Kosel, Juliusz; Rusak, Małgorzata; Gołembiewski, Łukasz; Dąbrowska, Milena; Siemiątkowski, Andrzej

2016-01-01

Among the many changes caused by a surgical insult one of the least studied is postoperative immunosuppression. This phenomenon is an important cause of infectious complications of surgery such as surgical site infection or hospital acquired pneumonia. One of the mechanisms leading to postoperative immunosuppression is the apoptosis of immunological cells. Anesthesia during surgery is intended to minimize harmful changes and maintain perioperative homeostasis. The aim of the study was evaluation of the effect of the anesthetic technique used for total knee replacement on postoperative peripheral blood lymphocyte apoptosis. 34 patients undergoing primary total knee replacement were randomly assigned to two regional anesthetic protocols: spinal anesthesia and combined spinal-epidural anesthesia. 11 patients undergoing total knee replacement under general anesthesia served as control group. Before surgery, immediately after surgery, during first postoperative day and seven days after the surgery venous blood samples were taken and the immunological status of the patient was assessed with the use of flow cytometry, along with lymphocyte apoptosis using fluorescent microscopy. Peripheral blood lymphocyte apoptosis was seen immediately in the postoperative period and was accompanied by a decrease of the number of T cells and B cells. There were no significant differences in the number of apoptotic lymphocytes according to the anesthetic protocol. Changes in the number of T CD3/8 cells and the number of apoptotic lymphocytes were seen on the seventh day after surgery. Peripheral blood lymphocyte apoptosis is an early event in the postoperative period that lasts up to seven days and is not affected by the choice of the anesthetic technique. Copyright © 2014 Sociedade Brasileira de Anestesiologia. Published by Elsevier Editora Ltda. All rights reserved.

[Morpho-functional parameters of nucleoli in polyploid mucous and albumen cells of salivary gland in the snail Succinea lauta].

PubMed

Anisimova, A A; Anisimov, A P

2005-01-01

Variation of some characteristics of nucleoli of polyploid mucous and albumen cells was examined in salivary glands of the snail Succinea lauta. The number, total area and Ag-protein content of nucleoli, and DNA content in each nucleus were estimated on squashed preparations incubated with AgNO3, decolorized and then Feulgen stained. The ultrastructure of nucleoli was studied by electron microscopy. Differentiated mucous cells had 4c-8c-16c-32c nuclei; albumen cells had 8c-16c-32c-64c-128c nuclei. The ultrastructure of nucleoli of the two cell types was essentially the same. Normally, a large fibrous to granular zone was observed in the nucleoli, without a clear distinction between fibrous and granular components. At the same time, aggregations of granular matter could be discerned at the periphery of nucleoli. No fibrous centers were observed. Occassionally, nucleolonema-like structures occurred. Normally each nucleolus contacted several chromosomes. On squashed preparations, the least size of nucleoli was 2-3 microm, and the largest size amounted to 14 microm in mucous cells, and to 50-80 microm in albumen cells. The number of nucleoli rose from 1-2 in tetraploid nuclei to 2-3 in 32c-nuclei, and to 5-7 in 128c-nuclei. The disparity between the ploidy levels of nuclei and the numbers of nucleoli may be due, presumably, to aggregation of chromosome NORs. The Ag-protein content in the nucleoli, and the total nucleolar area displayed a strong mutual correlation. Both parameters differed significantly by 1.5-2.2 times in mucous and albumen cells of the same ploidy level. Thus, in albumen and mucous cells the total Ag-protein content in octaploid nuclei was 3.3 and 2.2 relative units (r. u.), respectively. In 16c- and 32c-nuclei of albumen cells, it was 7.6 and 15.1 r. u.; and in the same nuclei of mucous cells--3.8 and 6.8 r. u., respectively. On the whole, in albumen cells, in the course of 4 endocycles (4c-128c), the total Ag-protein content increased by 17 times. Therefore, the mean multiplication factor for this parameter was found to be 2.05 per endocycle. In mucous cells, in the course of 3 endocycles (4c-32c), the total Ag-protein content increased by 5.2 times against 8 times expected, with the mean multiplication factor equal to 1.75 per endocycle. Thus, in the course of polyploidization of albumen and mucous cell nuclei, the gene dosage effect was fully pronounced in the former, and only partly in the latter. This differtence is due obviously to peculiarities of differentiation of the two cell types, in particular, to differences in the number of activated ribosomal genes.

Quantitative Detection of Viable Bifidobacterium bifidum BF-1 Cells in Human Feces by Using Propidium Monoazide and Strain-Specific Primers

PubMed Central

Fujimoto, Junji

2013-01-01

We developed a PCR-based method to detect and quantify viable Bifidobacterium bifidum BF-1 cells in human feces. This method (PMA-qPCR) uses propidium monoazide (PMA) to distinguish viable from dead cells and quantitative PCR using a BF-1-specific primer set designed from the results of randomly amplified polymorphic DNA analysis. During long-term culture (10 days), the number of viable BF-1 cells detected by counting the number of CFU on modified MRS agar, by measuring the ATP contents converted to CFU, and by using PMA-qPCR decreased from about 1010 to 106 cells/ml; in contrast, the total number of (viable and dead) BF-1 cells detected by counting 4′,6-diamidino-2-phenylindolee (DAPI)-stained cells and by using qPCR without PMA and reverse transcription-qPCR remained constant. The number of viable BF-1 cells in fecal samples detected by using PMA-qPCR was highly and significantly correlated with the number of viable BF-1 cells added to the fecal samples, within the range of 105.3 to 1010.3 cells/g feces (wet weight) (r > 0.99, P < 0.001). After 12 healthy subjects ingested 1010.3 to 1011.0 CFU of BF-1 in a fermented milk product daily for 28 days, 104.5 ± 1.5 (mean ± standard deviation [SD]) BF-1 CFU/g was detected in fecal samples by using strain-specific selective agar; in contrast, 106.2 ± 0.4 viable BF-1 cells/g were detected by using PMA-qPCR, and a total of 107.6 ± 0.7 BF-1 cells/g were detected by using qPCR without PMA. Thus, the number of viable BF-1 cells detected by PMA-qPCR was about 50 times higher (P < 0.01) than that detected by the culture-dependent method. We conclude that strain-specific PMA-qPCR can be used to quickly and accurately evaluate viable BF-1 in feces. PMID:23354719

Multifactorial analysis of human blood cell responses to clinical total body irradiation

NASA Technical Reports Server (NTRS)

Yuhas, J. M.; Stokes, T. R.; Lushbaugh, C. C.

1972-01-01

Multiple regression analysis techniques are used to study the effects of therapeutic radiation exposure, number of fractions, and time on such quantal responses as tumor control and skin injury. The potential of these methods for the analysis of human blood cell responses is demonstrated and estimates are given of the effects of total amount of exposure and time of protraction in determining the minimum white blood cell concentration observed after exposure of patients from four disease groups.

Tier-2 studies on monocrotaline immunotoxicity in C57BL/6 mice.

PubMed

Deyo, J A; Kerkvliet, N I

1991-01-01

Monocrotaline (MCT) is a member of a class of naturally occurring phytotoxins known as pyrrolizidine alkaloids, and is a toxicological concern to both man and his livestock. The purpose of these studies was to evaluate the effect of a 14-day oral MCT (0-100 mg/kg per day) exposure on the functional integrity of various immunocyte effector systems in C57BL/6 mice, as well as to investigate potential mechanisms for its immunotoxicity. Decreases in lymphoid organ weights and cellularity, and resident peritoneal exudate cell (PEC) number were only observed after exposure to the highest dose of 100 mg/kg MCT. This dose also inhibited NK cell cytotoxicity, while the total number of NK lytic units per spleen was decreased (-53%) after exposure to 50 mg/kg MCT. Following i.p. injection of SRBC, the percentage of PEC macrophages containing engulfed SRBC was significantly increased in MCT-exposed mice, while the percentage of large vacuolated (activated) macrophages was decreased in a dose-dependent manner. Exposure to MCT significantly decreased the total number of Ig+ cells without altering the number of CD4+ and CD8+ cells. The antibody responses (PFC/10(6) spleen cells) to two T cell-independent antigens, TNP-LPS and DNP-Ficoll, were significantly decreased at all MCT doses, and the degree of suppression of both responses was identical at coincident doses. MCT exposure (25 mg/kg) significantly suppressed the blastogenic response to the T cell mitogen concanavalin A (-38%), and to the B cell mitogen lipopolysaccharide (-58%). These results indicate that exposure to MCT can alter the functional integrity of various immune effector responses in a dose-dependent manner, and suggest that the B cell may be a relatively more sensitive target of MCT immunotoxicity compared to T cells, macrophages and NK cells.

Effects of several immunostimulants on phenoloxidase and hemocytes of the crab Charybdis japonica

NASA Astrophysics Data System (ADS)

Fan, Tingjun; Yu, Miaomiao; Yang, Lingling; Shi, Zhenping; Sun, Wenjie; Cong, Rishan; Yang, Xiuxia; Jiang, Guojian

2009-09-01

To investigate the stimulating effects of immunostimulants on the autogenous immunocompetence of crabs and the possible mechanisms involved, the immunostimulating effects of β-1,3-glucan, lipopolysaccharide (LPS), inactivated Vibrio harveyi and Vibrio anguillarum on phenoloxidase (PO) and hemocytes of Charybdis japonica were investigated in this study. It was found that the yields and the enzymatic activities of purified PO in C. japonica increased significantly after the crabs were treated with immunostimulants, while the unit enzymatic activities remained almost the same. After treatment with β-1,3-glucan and LPS, the amount of rough endoplasmic reticulum (RER) and the number of mitochondria in both semigranular cells and granular cells increased greatly, and the number of cytoplasmic granules decreased but with enlarged volume. However, the corresponding characteristics of hyaline cells remained almost the same. On the other hand, the number of granules in semigranular cells decreased greatly, and the number of mitochondria of hyaline cells increased greatly, after treatment with inactivated vibrios. It may be concluded that the effect of polysaccharide immunostimulants on the innate immune system of C. japonica is different from that of inactivated vibrio immunostimulants. The immunity-enhancing mechanism of polysaccharides in crab autogenous immunocompetence is probably accomplished by the increased yields of PO and total PO activities, while that of inactivated vibrios is probably accomplished by the partially increased yields of PO and total PO activities as well as the significantly improved phagocytotic abilities of semigranular cells and hyaline cells.

Effect of repeated arthrocentesis on cytologic analysis of synovial fluid in dogs.

PubMed

Berg, R I M; Sykes, J E; Kass, P H; Vernau, W

2009-01-01

Serial arthrocentesis and synovial fluid examination can be used to monitor treatment efficacy in immune-mediated polyarthritis (IMPA), but whether this procedure induces inflammation that interferes with test result interpretation is unknown. The aim of this study was to determine the effect of repeated arthrocentesis on synovial fluid cytology in healthy dogs. Nine healthy client-owned dogs. Prospective study. Arthrocentesis was performed under sedation on 4 joints (both carpi, 1 tarsus, 1 stifle) on each dog every 3 weeks, a total of 4 times. Automated cell counts were done on stifle fluid, smears were made, and differential cell counts done on smears from all joints. Slides were evaluated microscopically for erythrocyte numbers, total nucleated cell count, differential cell count, and cell morphology. Data were analyzed by 2-way analysis of variance. A total of 144 synovial fluid samples were examined. Repeated arthrocentesis was not associated with increases in synovial fluid neutrophil numbers. Mild mononuclear inflammation was detected in 13 samples from 6 dogs. Serial arthrocentesis at 3-week intervals can rarely be associated with mild mononuclear joint inflammation, but does not appear to induce neutrophilic inflammation, at least in healthy dogs, and can be useful to monitor treatment response in canine IMPA.

Chronic Binge Alcohol Administration Increases Intestinal T-Cell Proliferation and Turnover in Rhesus Macaques.

PubMed

Veazey, Ronald S; Amedee, Angela; Wang, Xiaolei; Bernice Kaack, M; Porretta, Constance; Dufour, Jason; Welsh, David; Happel, Kyle; Pahar, Bapi; Molina, Patricia E; Nelson, Steve; Bagby, Gregory J

2015-08-01

Alcohol use results in changes in intestinal epithelial cell turnover and microbial translocation, yet less is known about the consequences on intestinal lymphocytes in the gut. Here, we compared T-cell subsets in the intestine of macaques before and after 3 months of chronic alcohol administration to examine the effects of alcohol on intestinal T-cell subsets. Rhesus macaques received either alcohol or isocaloric sucrose as a control treatment daily over a 3-month period via indwelling gastric catheters. Intestinal lymphocyte subsets were identified in biopsy samples by flow cytometry. Twenty-four hours prior to sampling, animals were inoculated with bromo-deoxyuridine (BrdU) to assess lymphocyte proliferation. Immunohistochemistry was performed on tissue samples to quantitate CD3+ cells. Animals receiving alcohol had increased rates of intestinal T-cell turnover of both CD4+ and CD8+ T cells as reflected by increased BrdU incorporation. However, absolute numbers of T cells were decreased in intestinal tissues as evidenced by immunohistochemistry for total CD3 expression per mm(2) intestinal lamina propria in tissue sections. Combining immunohistochemistry and flow cytometry data showed that the absolute numbers of CD8+ T cells were significantly decreased, whereas absolute numbers of total CD4+ T cells were minimally decreased. Collectively, these data indicate that alcohol exposure to the small intestine results in marked loss of CD3+ T cells, accompanied by marked increases in CD4+ and CD8+ T-cell proliferation and turnover, which we speculate is an attempt to maintain stable numbers of T cells in tissues. This suggests that alcohol results in accelerated T-cell turnover in the gut, which may contribute to premature T-cell senescence. Further, these data indicate that chronic alcohol administration results in increased levels of HIV target cells (proliferating CD4+ T cells) that may support higher levels of HIV replication in intestinal tissues. Copyright © 2015 by the Research Society on Alcoholism.

Magnetic beads (Dynabead) toxicity to endothelial cells at high bead concentration: implication for tissue engineering of vascular prosthesis.

PubMed

Tiwari, A; Punshon, G; Kidane, A; Hamilton, G; Seifalian, A M

2003-10-01

Magnetic beads (Dynabeads) have been used for the purification of endothelial cells. One application for this procedure may be for single-stage seeding of bypass grafts. The number of endothelial cells (EC) isolated is crucial and therefore to increase the number of cells extracted, a higher number of Dynabeads per cell may need to be used. The effect of large numbers of CD31 Dynabeads on cell proliferation/metabolism is unknown. We undertook this study using CD31-coated Dynabeads and EC from human umbilical vein. EC were coated at concentrations of 4, 10, or 50 beads per cell. The cells were cultured for 6 days with control being normal EC. Cellular proliferation was assessed by trypsinization of cells and metabolism assessed with an Alamar blue viability assay. In a further experiment a compliant polyurethane graft was single-stage seeded with both coated Dynabeads and normal EC. The results showed that using a higher number of beads per cell resulted in a reduction in cell proliferation and a reduction in cell metabolism. The total number of Dynabeads-coated cells in culture compared to controls (%) by day 6 were 30.7 +/- 2.56, 41.3 +/- 9.8 and 59.2 +/- 7.3 for 50, 10, and 4 beads per cell, respectively. The corresponding results for Alamar blue were 43.7 +/- 1.2, 61.8 +/- 1.4, and 72.1 +/- 4.3. The seeded grafts showed reduced metabolism with the Dynabeads-coated EC. In conclusion, high numbers of beads per cell have a late detrimental effect on cell proliferation and metabolism. Therefore for single-stage seeding lower numbers of Dynabeads will need to be used with resultant reduction in the number of available EC.

Effect of electromagnetic microwave radiation on the growth of Ehrlich ascites carcinoma.

PubMed

Kryukova, O V; Pyankov, V F; Kopylov, A F; Khlebopros, R G

2016-09-01

Daily exposure of mouse recipients of Ehrlich ascites carcinoma to electromagnetic radiation of the microwave range leads to a change in the dynamics of tumor growth by decreasing the total number of cells. The number of tumor cells with blebbing morphological signs after microwave radiation increases gradually with tumor growth. The maximum content of tumor cells in the state of blebbing is observed during active proliferation in tumor-recipient mice of the control group (without irradiation).

Study of physical and biological factors involved in the disruption of E. coli by hydrodynamic cavitation.

PubMed

Balasundaram, B; Harrison, S T L

2006-01-01

Hydrodynamic cavitation results in flow restriction in a flow system causing rapid pressure fluctuations and significant fluid forces. These can be harnessed to mediate microbial cell damage. Hydrodynamic cavitation was studied for the partial disruption of E. coli and selective release of specific proteins relative to the total soluble protein. The effects of the cavitation number, the number of passes, and the specific growth rate of E. coli on the release of periplasmic and cytoplasmic proteins were studied. At the optimum cavitation number of 0.17 for this experimental configuration, 48% of the total soluble protein, 88% of acid phosphatase, and 67% of beta-galactosidase were released by hydrodynamic cavitation in comparison with the maximum release attained using multiple passes through the French Press. The higher release of the acid phosphatase over the total soluble protein suggested preferred release of periplasmic compounds. This was supported by SDS-PAGE analysis. The absence of micronization of cell material resulting in the potential for ease of solid-liquid separation downstream of the cell disruption operation was confirmed by TEM microscopy. E. coli cells cultivated at a higher specific growth rate (0.36 h(-1)) were more easily disrupted than slower grown cells (0.11 h(-1)). The specific activity of the enzyme of interest released by hydrodynamic cavitation, defined as the units of enzyme in solution per milligram of total soluble protein, was greater than that obtained on release by the French Press, high-pressure homogenization, osmotic shock, and EDTA treatment. The selectivity offered indicates the potential of enzyme release by hydrodynamic cavitation to ease the purification in the subsequent downstream processing.

Exposure to fine and ultrafine particulate matter during gestation alters postnatal oligodendrocyte maturation, proliferation capacity, and myelination.

PubMed

Klocke, Carolyn; Allen, Joshua L; Sobolewski, Marissa; Blum, Jason L; Zelikoff, Judith T; Cory-Slechta, Deborah A

2018-03-01

Accumulating studies indicate that the brain is a direct target of air pollution exposure during the fetal period. We have previously demonstrated that exposure to concentrated ambient particles (CAPs) during gestation produces ventriculomegaly, periventricular hypermyelination, and enlargement of the corpus callosum (CC) during postnatal development in mice. This study aimed to further characterize the cellular basis of the observed hypermyelination and determine if this outcome, among other effects, persisted as the brain matured. Analysis of CC-1 + mature oligodendrocytes in the CC at postnatal days (PNDs) 11-15 suggest a premature maturational shift in number and proportion of total cells in prenatally CAPs-exposed males and females, with no overall change in total CC cellularity. The overall number of Olig2 + lineage cells in the CC was not affected in either sex at the same postnatal timepoint. Assessment of myelin status at early brain maturity (PNDs 57-61) revealed persistent hypermyelination in CAPs-exposed animals of both sexes. In addition, ventriculomegaly was persistent in CAPs-treated females, with possible amelioration of ventriculomegaly in CAPs-exposed males. When oligodendrocyte precursor cell (OPC) pool status was analyzed at PNDs 57-61, there were significant CAPs-induced alterations in cycling Ki67 + /Olig2 + cell number and proportion of total cells in the female CC. Total CC cellularity was slightly elevated in CAPs-exposed males at PNDs 57-61. Overall, these data support a growing body of evidence that demonstrate the vulnerability of the developing brain to environmental insults such as ambient particulate matter. The sensitivity of oligodendrocytes and myelin, in particular, to such an insult warrants further investigation into the mechanistic underpinnings of OPC and myelin disruption by constituent air pollutants. Copyright © 2017 Elsevier B.V. All rights reserved.

Comparison of fluorescence microscopy and solid-phase cytometry methods for counting bacteria in water

USGS Publications Warehouse

Lisle, John T.; Hamilton, Martin A.; Willse, Alan R.; McFeters, Gordon A.

2004-01-01

Total direct counts of bacterial abundance are central in assessing the biomass and bacteriological quality of water in ecological and industrial applications. Several factors have been identified that contribute to the variability in bacterial abundance counts when using fluorescent microscopy, the most significant of which is retaining an adequate number of cells per filter to ensure an acceptable level of statistical confidence in the resulting data. Previous studies that have assessed the components of total-direct-count methods that contribute to this variance have attempted to maintain a bacterial cell abundance value per filter of approximately 106 cells filter-1. In this study we have established the lower limit for the number of bacterial cells per filter at which the statistical reliability of the abundance estimate is no longer acceptable. Our results indicate that when the numbers of bacterial cells per filter were progressively reduced below 105, the microscopic methods increasingly overestimated the true bacterial abundance (range, 15.0 to 99.3%). The solid-phase cytometer only slightly overestimated the true bacterial abundances and was more consistent over the same range of bacterial abundances per filter (range, 8.9 to 12.5%). The solid-phase cytometer method for conducting total direct counts of bacteria was less biased and performed significantly better than any of the microscope methods. It was also found that microscopic count data from counting 5 fields on three separate filters were statistically equivalent to data from counting 20 fields on a single filter.

Data on the number and frequency of scientific literature citations for established medulloblastoma cell lines.

PubMed

Ivanov, D P; Walker, D A; Coyle, B; Grabowska, A M

2016-12-01

This article collates information about the number of scientific articles mentioning each of the established medulloblastoma cell lines, derived through a systematic search of Web of Science, Scopus and Google Scholar in 2016. The data for each cell line have been presented as raw number of citations, percentage share of the total citations for each search engine and as an average percentage between the three search engines. In order to correct for the time since each cell line has been in use, the raw citation data have also been divided by the number of years since the derivation of each cell line. This is a supporting article for a review of in vitro models of medulloblastoma published in "in vitro models of medulloblastoma: choosing the right tool for the job" (D.P. Ivanov, D.A. Walker, B. Coyle, A.M. Grabowska, 2016) [1].

Characterization of winemaking yeast by cell number-size distribution analysis through flow field-flow fractionation with multi-wavelength turbidimetric detection.

PubMed

Zattoni, Andrea; Melucci, Dora; Reschiglian, Pierluigi; Sanz, Ramsés; Puignou, Lluís; Galceran, Maria Teresa

2004-10-29

Yeasts are widely used in several areas of food industry, e.g. baking, beer brewing, and wine production. Interest in new analytical methods for quality control and characterization of yeast cells is thus increasing. The biophysical properties of yeast cells, among which cell size, are related to yeast cell capabilities to produce primary and secondary metabolites during the fermentation process. Biophysical properties of winemaking yeast strains can be screened by field-flow fractionation (FFF). In this work we present the use of flow FFF (FlFFF) with turbidimetric multi-wavelength detection for the number-size distribution analysis of different commercial winemaking yeast varieties. The use of a diode-array detector allows to apply to dispersed samples like yeast cells the recently developed method for number-size (or mass-size) analysis in flow-assisted separation techniques. Results for six commercial winemaking yeast strains are compared with data obtained by a standard method for cell sizing (Coulter counter). The method here proposed gives, at short analysis time, accurate information on the number of cells of a given size, and information on the total number of cells.

Studies on the Biochemistry and Fine Structure of Silica Shell Formation in Diatoms. Chemical Composition of Navicula pelliculosa during Silicon-Starvation Synchrony 1

PubMed Central

Coombs, J.; Darley, W. M.; Holm-Hansen, O.; Volcani, B. E.

1967-01-01

Changes are reported in total cellular organic carbon, nucleic acids, proteins, carbohydrates, lipids and chlorophylls during the course of silicon-starvation synchrony of Navicula pelliculosa. All constituents increased at the same rate, relative to cell number, for 30 hours of exponential growth during which silicon was depleted from the medium. Increase in cell number then stopped, but net synthesis of most components continued for a further 5 to 7 hours before ceasing. Deoxyribonucleic acids and lipids accumulated throughout the 14 hour silicon-starvation period. When silicon was resupplied, lipid synthesis ceased and organic carbon and carbohydrates decreased slightly. Net synthesis remained low during the 4 hour silicon uptake period but was resumed at higher rates as cell number began to rise. In cultures transferred to the dark 1 hour prior to readdition of silicon, total carbon, carbohydrates, and lipids decreased markedly during silicon uptake and cell separation. This was due in part to conversion of protein which maintained the protein level of the dark cells close to that of cells kept in the light. Mechanisms by which silicon starvation and reintroduction of silicon might affect rates of cellular synthesis are discussed. PMID:6080872

Classification of Mouse Retinal Bipolar Cells: Type-Specific Connectivity with Special Reference to Rod-Driven AII Amacrine Pathways

PubMed Central

Tsukamoto, Yoshihiko; Omi, Naoko

2017-01-01

We confirmed the classification of 15 morphological types of mouse bipolar cells by serial section transmission electron microscopy and characterized each type by identifying chemical synapses and gap junctions at axon terminals. Although whether the previous type 5 cells consist of two or three types was uncertain, they are here clustered into three types based on the vertical distribution of axonal ribbons. Next, while two groups of rod bipolar (RB) cells, RB1, and RB2, were previously proposed, we clarify that a half of RB1 cells have the intermediate characteristics, suggesting that these two groups comprise a single RB type. After validation of bipolar cell types, we examined their relationship with amacrine cells then particularly with AII amacrine cells. We found a strong correlation between the number of amacrine cell synaptic contacts and the number of bipolar cell axonal ribbons. Formation of bipolar cell output at each ribbon synapse may be effectively regulated by a few nearby inhibitory inputs of amacrine cells which are chosen from among many amacrine cell types. We also found that almost all types of ON cone bipolar cells frequently have a minor group of midway ribbons along the axon passing through the OFF sublamina as well as a major group of terminal ribbons in the ON sublamina. AII amacrine cells are connected to five of six OFF bipolar cell types via conventional chemical synapses and seven of eight ON (cone) bipolar cell types via electrical synapses (gap junctions). However, the number of synapses is dependent on bipolar cell types. Type 2 cells have 69% of the total number of OFF bipolar chemical synaptic contacts with AII amacrine cells and type 6 cells have 46% of the total area of ON bipolar gap junctions with AII amacrine cells. Both type 2 and 6 cells gain the greatest access to AII amacrine cell signals also share those signals with other types of bipolar cells via networked gap junctions. These findings imply that the most sensitive scotopic signal may be conveyed to the center by ganglion cells that have the most numerous synapses with type 2 and 6 cells. PMID:29114208

Cytoprotection: Immune and Matrix Modulation of Tissue Repair

DTIC Science & Technology

2012-04-01

bronchoalveolar lavage (BAL) fluid for cytokines, cell counts and eosinophilia, and histological analysis of airway hyper-responsiveness and remodeling...OVA administered intra-nasally on Days 21–25 with or without 0.1% XHA. B) Total leukocyte and eosinophil counts in bronchoalveolar lavage (BAL) fluid...and different doses of HA peptide. The absolute number of Tmr+ and FOXP3+ populations was determined using total cell counts and flow cytometry for Tmr

The increase in the number of astrocytes in the total cerebral ischemia model in rats

NASA Astrophysics Data System (ADS)

Kudabayeva, M.; Kisel, A.; Chernysheva, G.; Smol'yakova, V.; Plotnikov, M.; Khodanovich, M.

2017-08-01

Astrocytes are the most abundant cell class in the CNS. Astrocytic therapies have a huge potential for neuronal repair after stroke. The majority of brain stroke studies address the damage to neurons. Modern studies turn to the usage of morphological and functional changes in astroglial cells after stroke in regenerative medicine. Our study is focused on the changes in the number of astrocytes in the hippocampus (where new glia cells divide) after brain ischemia. Ischemia was modeled by occlusion of tr. brachiocephalicus, a. subclavia sin., a. carotis communis sin. Astrocytes were determined using immunohistochemical labeling with anti GFAP antibody. We found out that the number of astrocytes increased on the 10th and 30th days after stroke in the CA1, CA2 fields, the granular layer of dentate gyrus (GrDG) and hilus. The morphology of astrocytes became reactive in these regions. Therefore, our results revealed long-term reactive astrogliosis in the hippocampus region after total ischemia in rats.

Measuring mRNA copy-number in individual Escherichia coli cells using single-molecule fluorescent in situ hybridization (smFISH)

PubMed Central

Skinner, Samuel O.; Sepúlveda, Leonardo A.; Xu, Heng; Golding, Ido

2014-01-01

We present a method for measuring the absolute number of mRNA molecules from a gene of interest in individual, chemically fixed Escherichia coli cells. A set of fluorescently-labeled oligonucleotide probes are hybridized to the target mRNA, so that each mRNA molecule is decorated by a known number of fluorescent dyes. Cells are then imaged using fluorescence microscopy. The number of target mRNA is estimated from the total intensity of fluorescent foci in the cell, rather than from counting discrete “spots” as in other currently available protocols. Image analysis is performed using an automated algorithm. The measured mRNA copy-number distribution obtained from many individual cells can be used to extract the parameters of stochastic gene activity, namely the frequency and size of transcription bursts from the gene of interest. The experimental procedure takes 2 days, with another 2-3 days typically required for image and data analysis. PMID:23680982

Health investigations of depleted-uranium clean-up workers.

PubMed

Milacic, Snezana

2008-01-01

The soil contaminated by depleted uranium (238U) ammunition during the NATO bombing of Serbia and Montenegro was cleaned-up for four months in 2002. A team of 11 clean-up workers (expert members) were medically examined three times: before decontamination as a preliminary medical check-up, immediately after decontamination, and four years after cleaning up contaminated ground. This short report presents investigations and health risk assessments of clean-up workers in radioactive decontamination operations and an assessment of the environmental health perspectives for citizens living in surrounding areas. The method of initial health disorders was used, analyzing the most sensitive biological materials, such as blood cells or chromosome damage, DNA strand breaks, radio-toxicological examination of urine. The total number of blood cells did not change, but variations of the relative number (percentage) of cells in the leukocyte formula were observed. The total number of DNA alterations was higher immediately after decontamination than before decontamination. Four years after decontamination the number of DNA alterations had decreased. However, the number of damaged cells (lymphocytes containing chromosome lesions) was higher in both medical examinations, immediately after and four years after decontamination. Disease or tumours due to 238U did not develop in the group of depleted uranium clean-up workers during the investigation period of four years. Further monitoring of haematological and chromosomal effects and the health condition of workers is necessary.

Effect of topical rebamipide on goblet cells in the lid wiper of human conjunctiva

PubMed Central

Kase, Satoru; Shinohara, Toshiya; Kase, Manabu; Ishida, Susumu

2017-01-01

It has been demonstrated that topical administration of rebamipide, which is an antiulcer agent, increases the mucin level of the tear film and ameliorates ocular surface conditions such as lid wiper epitheliopathy. The aim of the present study was to analyze the changes in goblet cell number, cell proliferation, and epidermal growth factor receptor (EGFR) induced by topical rebamipide addition to the lid wiper of humans. A total of 30 eyelid tissue samples were obtained during involutional entropion surgeries, fixed in paraformaldehyde, embedded in paraffin and divided into two groups: Rebamipide or non-rebamipide. The tissues in the rebamipide group were obtained from patients who had a medical history of topical rebamipide use prior to surgery. The number of goblet cells was counted under light microscopy. A total of 22 eyelid tissue samples were further examined using immunohistochemistry with anti-Ki-67 and anti-EGFR antibodies to evaluate cell proliferation and EGFR expression, respectively. Histologically, the lid wiper and palpebral conjunctiva were clearly identified in the tissues. The number of goblet cells was significantly higher in the rebamipide group compared with the non-rebamipide group (P=0.0367). There was no significant difference in lid wiper cell proliferation between the rebamipide and non-rebamipide groups. Immunohistochemistry revealed that EGFR levels in the lid wiper epithelial cells were significantly higher in the rebamipide group compared with the non-rebamipide group (P=0.0237). These results suggest that topical rebamipide application increases the number of goblet cells in the lid wiper, which in turn upregulates the expression of EGFR. These findings may be clinically relevant and provide a therapeutic basis for the treatment of ocular disease such as dry eye and lid wiper epitheliopathy. PMID:28587435

Effect of topical rebamipide on goblet cells in the lid wiper of human conjunctiva.

PubMed

Kase, Satoru; Shinohara, Toshiya; Kase, Manabu; Ishida, Susumu

2017-06-01

It has been demonstrated that topical administration of rebamipide, which is an antiulcer agent, increases the mucin level of the tear film and ameliorates ocular surface conditions such as lid wiper epitheliopathy. The aim of the present study was to analyze the changes in goblet cell number, cell proliferation, and epidermal growth factor receptor (EGFR) induced by topical rebamipide addition to the lid wiper of humans. A total of 30 eyelid tissue samples were obtained during involutional entropion surgeries, fixed in paraformaldehyde, embedded in paraffin and divided into two groups: Rebamipide or non-rebamipide. The tissues in the rebamipide group were obtained from patients who had a medical history of topical rebamipide use prior to surgery. The number of goblet cells was counted under light microscopy. A total of 22 eyelid tissue samples were further examined using immunohistochemistry with anti-Ki-67 and anti-EGFR antibodies to evaluate cell proliferation and EGFR expression, respectively. Histologically, the lid wiper and palpebral conjunctiva were clearly identified in the tissues. The number of goblet cells was significantly higher in the rebamipide group compared with the non-rebamipide group (P=0.0367). There was no significant difference in lid wiper cell proliferation between the rebamipide and non-rebamipide groups. Immunohistochemistry revealed that EGFR levels in the lid wiper epithelial cells were significantly higher in the rebamipide group compared with the non-rebamipide group (P=0.0237). These results suggest that topical rebamipide application increases the number of goblet cells in the lid wiper, which in turn upregulates the expression of EGFR. These findings may be clinically relevant and provide a therapeutic basis for the treatment of ocular disease such as dry eye and lid wiper epitheliopathy.

Critical telomerase activity for uncontrolled cell growth

NASA Astrophysics Data System (ADS)

Wesch, Neil L.; Burlock, Laura J.; Gooding, Robert J.

2016-08-01

The lengths of the telomere regions of chromosomes in a population of cells are modelled using a chemical master equation formalism, from which the evolution of the average number of cells of each telomere length is extracted. In particular, the role of the telomere-elongating enzyme telomerase on these dynamics is investigated. We show that for biologically relevant rates of cell birth and death, one finds a critical rate, R crit, of telomerase activity such that the total number of cells diverges. Further, R crit is similar in magnitude to the rates of mitosis and cell death. The possible relationship of this result to replicative immortality and its associated hallmark of cancer is discussed.

Intercellular Variability in Protein Levels from Stochastic Expression and Noisy Cell Cycle Processes

PubMed Central

Soltani, Mohammad; Vargas-Garcia, Cesar A.; Antunes, Duarte; Singh, Abhyudai

2016-01-01

Inside individual cells, expression of genes is inherently stochastic and manifests as cell-to-cell variability or noise in protein copy numbers. Since proteins half-lives can be comparable to the cell-cycle length, randomness in cell-division times generates additional intercellular variability in protein levels. Moreover, as many mRNA/protein species are expressed at low-copy numbers, errors incurred in partitioning of molecules between two daughter cells are significant. We derive analytical formulas for the total noise in protein levels when the cell-cycle duration follows a general class of probability distributions. Using a novel hybrid approach the total noise is decomposed into components arising from i) stochastic expression; ii) partitioning errors at the time of cell division and iii) random cell-division events. These formulas reveal that random cell-division times not only generate additional extrinsic noise, but also critically affect the mean protein copy numbers and intrinsic noise components. Counter intuitively, in some parameter regimes, noise in protein levels can decrease as cell-division times become more stochastic. Computations are extended to consider genome duplication, where transcription rate is increased at a random point in the cell cycle. We systematically investigate how the timing of genome duplication influences different protein noise components. Intriguingly, results show that noise contribution from stochastic expression is minimized at an optimal genome-duplication time. Our theoretical results motivate new experimental methods for decomposing protein noise levels from synchronized and asynchronized single-cell expression data. Characterizing the contributions of individual noise mechanisms will lead to precise estimates of gene expression parameters and techniques for altering stochasticity to change phenotype of individual cells. PMID:27536771

30 CFR 7.44 - Technical requirements.

Code of Federal Regulations, 2010 CFR

2010-07-01

... the top of the wall. (i) Cell terminals shall be burned on, except that bolted connectors using two or... direct access to cell terminals and other uninsulated current carrying parts. The total minimum... following formula: EC15NO91.011 N = Number of cells in battery box. R = Rated 6 hour battery capacity in...

Comparison of Mast Cells and Inflammatory Cells within Periapical Lesions and Comparison of Degranulated Mast Cells Between Fibrous and Inflamed Area in Radicular Cysts: An Immunohistochemical Study.

PubMed

Shiromany, Aseem; Sood, Rahul; Akifuddin, Syed; Sidhu, Gagandeep Kaur; Khan, Nadia; Singla, Kapil

2014-12-01

The role of mast cells as the key effector of allergic inflammation, anaphylactic inflammatory reactions and in the pathogenesis of chronic inflammation, is well-known. The present study is adopted to compare mast cells and inflammatory cells within periapical granuloma and cysts and localize the mast cells and quantify their number in the periapical cysts so as to propose a role of mast cells in the pathogenesis of this lesion. Biopsy specimens of 30 periapical lesions were stained with hematoxylin-eosin, and immunohistochemical Mast Cell Tryptase from Bio SB (IHC detection system kit) antibody. The tryptase positive mast cells and mononuclear inflammatory cells were counted in 10 consecutive high power fields (100X) using the binocular microscope from Motic attached to a computer with Motic Advanced Images 3.2 software. Comparative microscopic analysis indicated that periapical cyst shows more percentage of mast cells and less percentage of inflammatory cell than periapical granuloma (comparison of mean and standard deviation of total number of mast cells and inflammatory cells, mast cells 3.15±1.39 in the granuloma group and 4.43±1.91in the cyst group, inflammatory cells, 67.11±1.2 in the granuloma group and 52.66±0.8 in the cyst group). Numerous degranulated mast cells were observed in the fibrous wall than the inflammatory infiltrate of the periapical cysts. The mean and standard deviation of degranulated mast cells between the inflammatory and fibrous zone within the cyst group, being 0.95±1.10 and1.68±1.34 respectively. The values varied significantly between the two zones. The number of inflammatory cells in the cyst group is less than periapical granuloma and total number of mast cells in the cyst group is more as compared to periapical granuloma. The degranulated cells were quantified and they were higher in the fibrous area of the cysts than the inflammatory zone. This study could support the fact that the various mediators released on degranulation play a role in the connective tissue remodeling, chronicity and expansion of the periapical lesion.

FORSPAN Model Users Guide

USGS Publications Warehouse

Klett, T.R.; Charpentier, Ronald R.

2003-01-01

The USGS FORSPAN model is designed for the assessment of continuous accumulations of crude oil, natural gas, and natural gas liquids (collectively called petroleum). Continuous (also called ?unconventional?) accumulations have large spatial dimensions and lack well defined down-dip petroleum/water contacts. Oil and natural gas therefore are not localized by buoyancy in water in these accumulations. Continuous accumulations include ?tight gas reservoirs,? coalbed gas, oil and gas in shale, oil and gas in chalk, and shallow biogenic gas. The FORSPAN model treats a continuous accumulation as a collection of petroleumcontaining cells for assessment purposes. Each cell is capable of producing oil or gas, but the cells may vary significantly from one another in their production (and thus economic) characteristics. The potential additions to reserves from continuous petroleum resources are calculated by statistically combining probability distributions of the estimated number of untested cells having the potential for additions to reserves with the estimated volume of oil and natural gas that each of the untested cells may potentially produce (total recovery). One such statistical method for combination of number of cells with total recovery, used by the USGS, is called ACCESS.

Satellite cell response to concurrent resistance exercise and high-intensity interval training in sedentary, overweight/obese, middle-aged individuals.

PubMed

Pugh, Jamie K; Faulkner, Steve H; Turner, Mark C; Nimmo, Myra A

2018-02-01

Sarcopenia can begin from the 4-5th decade of life and is exacerbated by obesity and inactivity. A combination of resistance exercise (RE) and endurance exercise is recommended to combat rising obesity and inactivity levels. However, work continues to elucidate whether interference in adaptive outcomes occur when RE and endurance exercise are performed concurrently. This study examined whether a single bout of concurrent RE and high-intensity interval training (HIIT) alters the satellite cell response following exercise compared to RE alone. Eight sedentary, overweight/obese, middle-aged individuals performed RE only (8 × 8 leg extensions at 70% 1RM), or RE + HIIT (10 × 1 min at 90% HR max on a cycle ergometer). Muscle biopsies were collected from the vastus lateralis before and 96 h after the RE component to determine muscle fiber type-specific total (Pax7 + cells) and active (MyoD + cells) satellite cell number using immunofluorescence microscopy. Type-I-specific Pax7 + (P = 0.001) cell number increased after both exercise trials. Type-I-specific MyoD + (P = 0.001) cell number increased after RE only. However, an elevated baseline value in RE + HIIT compared to RE (P = 0.046) was observed, with no differences between exercise trials at 96 h (P = 0.21). Type-II-specific Pax7 + and MyoD + cell number remained unchanged after both exercise trials (all P ≥ 0.13). Combining a HIIT session after a single bout of RE does not interfere with the increase in type-I-specific total, and possibly active, satellite cell number, compared to RE only. Concurrent RE + HIIT may offer a time-efficient way to maximise the physiological benefits from a single bout of exercise in sedentary, overweight/obese, middle-aged individuals.

Immature germ cells in semen - correlation with total sperm count and sperm motility.

PubMed

Patil, Priya S; Humbarwadi, Rajendra S; Patil, Ashalata D; Gune, Anita R

2013-07-01

Current data regarding infertility suggests that male factor contributes up to 30% of the total cases of infertility. Semen analysis reveals the presence of spermatozoa as well as a number of non-sperm cells, presently being mentioned in routine semen report as "round cells" without further differentiating them into leucocytes or immature germ cells. The aim of this work was to study a simple, cost-effective, and convenient method for differentiating the round cells in semen into immature germ cells and leucocytes and correlating them with total sperm counts and motility. Semen samples from 120 males, who had come for investigation for infertility, were collected, semen parameters recorded, and stained smears studied for different round cells. Statistical analysis of the data was done to correlate total sperm counts and sperm motility with the occurrence of immature germ cells and leucocytes. The average shedding of immature germ cells in different groups with normal and low sperm counts was compared. The clinical significance of "round cells" in semen and their differentiation into leucocytes and immature germ cells are discussed. Round cells in semen can be differentiated into immature germ cells and leucocytes using simple staining methods. The differential counts mentioned in a semen report give valuable and clinically relevant information. In this study, we observed a negative correlation between total count and immature germ cells, as well as sperm motility and shedding of immature germ cells. The latter was statistically significant with a P value 0.000.

Engineering cord blood to improve engraftment after cord blood transplant

PubMed Central

Dave, Hema; Bollard, Catherine M.; Shpall, Elizabeth J.

2017-01-01

Umbilical cord blood transplant (CBT) has traditionally been associated with slower engraftment of neutrophils, delayed immune reconstitution and consequently higher risk of infections as compared with peripheral blood progenitor cell (PBPC) or bone marrow (BM) transplants. This is primarily due to low numbers of total nucleated cells (TNCs) and the naive nature of CB immune cells. The use of double unit CB transplant (DCBT) increases the total cell dose in the graft, but it still does not produce as rapid engraftment as seen with PBPC or even BM transplants. Herein, we discuss strategies to improve engraftment after CBT. We describe methods of (I) expansion of CB graft ex vivo to increase the total cell dose; and (II) enhancement of BM homing capability of CB progenitor cells; (III) ex vivo expansion of CB derived T cells for improving T cell function against viruses, tumors and protection from graft versus host disease (GVHD). With these novel approaches, engraftment after CBT is now reaching levels comparable to that of other graft types. PMID:28607915

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

Wagemaker, G.; Visser, T.P.; van Bekkum, D.W.

alpha-Thalassemic heterozygous (Hbath/+) mice were used to investigate the possible selective advantage of transplanted normal (+/+) hemopoietic cells. Without conditioning by total-body irradiation (TBI), infusion of large numbers of normal bone marrow cells failed to correct the thalassemic peripheral blood phenotype. Since the recipients' stem cells are normal with respect to number and differentiation capacity, it was thought that the transplanted stem cells were not able to lodge, or that they were not stimulated to proliferate. Therefore, a nonlethal dose of TBI was given to temporarily reduce endogenous stem cell numbers and hemopoiesis. TBI doses of 2 or 3 Gymore » followed by infusion of normal bone marrow cells proved to be effective in replacing the thalassemic red cells by normal red cells, whereas a dose of 1 Gy was ineffective. It is concluded that cure of thalassemia by bone marrow transplantation does not necessarily require eradication of thalassemic stem cells. Consequently, the objectives of conditioning regimens for bone marrow transplantation of thalassemic patients (and possibly other nonmalignant hemopoietic disorders) should be reconsidered.« less

Systemic immune cell response in rats after pulmonary exposure to manganese-containing particles collected from welding aerosols.

PubMed

Antonini, James M; Zeidler-Erdely, Patti C; Young, Shih-Houng; Roberts, Jenny R; Erdely, Aaron

2012-01-01

Welding fume inhalation affects the immune system of exposed workers. Manganese (Mn) in welding fume may induce immunosuppressive effects. The goal was to determine if Mn in welding fume alters immunity by reducing the number of circulating total leukocytes and specific leukocyte sub-populations. Sprague-Dawley rats were treated by intratracheal instillation (ITI) with either a single dose (2.00 mg/rat) or repeated doses (0.125 or 2.00 mg/rat for 7 weeks) with welding fumes that contained different levels of Mn. Additional rats were treated by ITI once a week for 7 weeks with the two doses of manganese chloride (MnCl₂). Bronchoalveolar lavage was performed to assess lung inflammation. Also, whole blood was recovered, and the number of circulating total leukocytes, as well as specific lymphocyte subsets, was determined by flow cytometry. The welding fume highest in Mn content significantly increased lung inflammation, injury, and production of inflammatory cytokines and chemokines compared to all other treatment groups. In addition, the same group expressed significant decreases in the number of circulating CD4⁺ and CD8⁺ T-lymphocytes after a single exposure, and significant reductions in the number of circulating total lymphocytes, primarily CD4⁺ and CD8⁺ T-lymphocytes, after repeated exposures (compared to control values). Repeated MnCl₂ exposure led to a trend of a reduction (but not statistically significant) in circulating total lymphocytes, attributable to the changes in the CD4⁺ T-lymphocyte population levels. The welding fume with the lower concentration of Mn had no significant effect on the numbers of blood lymphocytes and lymphocyte subsets compared to control values. Evidence from this study indicates that pulmonary exposure to certain welding fumes cause decrements in systemic immune cell populations, specifically circulating T-lymphocytes, and these alterations in immune cell number are not dependent exclusively on Mn, but likely a combination of other metals present in welding fume.

Retinal specializations and visual ecology in an animal with an extremely elaborate pupil shape: The Little skate Leucoraja (Raja) erinacea Mitchell, 1825.

PubMed

Jinson, S Terrell; Liebich, Jan; Senft, Stephen L; Mäthger, Lydia M

2018-05-14

Investigating retinal specializations offers insights into eye functionality. Using retinal wholemount techniques, we investigated the distribution of retinal ganglion cells in the Little skate Leucoraja erinacea by (1) dye-backfilling into the optic nerve prior to retinal wholemounting; (2) Nissl-staining of retinal wholemounts. Retinas were examined for regional specializations (higher numbers) of ganglion cells that would indicate higher visual acuity in those areas. Total ganglion cell number were low compared to other elasmobranchs (backfilled: average 49,713 total ganglion cells, average peak cell density 1,315 ganglion cells mm -2 ; Nissl-stained: average 47,791 total ganglion cells, average peak cell density 1,319 ganglion cells mm -2 ). Ganglion cells fit into three size categories: small (5-20µm); medium (20-30µm); large: (≥ 30µm), and they were not homogeneously distributed across the retina. There was a dorsally located horizontal visual streak with increased ganglion cell density; additionally, there were approximately 3 local maxima in ganglion cell distribution (potential areae centrales) within this streak in which densities were highest. Using computerized tomography (CT) and micro-CT, geometrical dimensions of the eye were obtained. Combined with ganglion cell distributions, spatial resolving power was determined to be between 1.21 to 1.37 cycles per degree. Additionally, photoreceptor sizes across different retinal areas varied; photoreceptors were longest within the horizontal visual streak. Variations in the locations of retinal specializations appear to be related to the animal's anatomy: shape of the head and eyes, position of eyes, location of tapetum, and shape of pupil, as well as the visual demands associated with lifestyle and habitat type. This article is protected by copyright. All rights reserved. © 2018 Wiley Periodicals, Inc.

Serum protein profile and blood cell counts in adult toads Bufo arenarum(Amphibia: Anura: Bufonidae): effects of sublethal lead acetate.

PubMed

Chiesa, María E; Rosenberg, Carolina E; Fink, Nilda E; Salibián, Alfredo

2006-04-01

Lead is a multiple-source pollutant, well known for its toxicity, of great risk both for the environment and human health. The main target organs of lead are the hematopoietic, nervous, and renal systems; there are also reports in support of its impairment effects on the reproductive and immune systems. It is well known that most of the metal is accumulated in the blood cells and that many of the deleterious effects are related to its circulating concentrations. These adverse effects have been described not only in humans but also in a number of other vertebrates such as fish and birds. The purpose of the present work was to evaluate the effects of weekly administration of sublethal Pb (as acetate, 50 mg x kg(-1)) during 6 weeks on the profile of the serum proteins and blood cell counts of the adult South American toad, Bufo arenarum (Anura: Bufonidae). The electrophoretic patterns of serum proteins pointed out the presence of four fractions; the metal provoked a significant decrease in both total proteins and albumin fraction; among the globulin fractions, the G3 resulted augmented. These findings may be related to the impact of lead on the toads' hepatic cells and immune system. The number of total red blood cells (RBC) showed a tendency to decrease after the injections of the metal, whereas the number of white blood cells (WBC) increased significantly; the differential leukocyte counts showed a statistically significant increase in the absolute number and in the relative percentage of blast-like cells. The decrease in RBC was attributed to the negative impact of the metals on the hemoglobin synthesis. The increasing of the WBC counts may be interpreted as a consequence of the induction of proliferation of pluripotential hematopoietic cells.

Correlation of quantitative histopathological morphology and quantitative radiological analysis during aseptic loosening of hip endoprostheses.

PubMed

Bertz, S; Kriegsmann, J; Eckardt, A; Delank, K-S; Drees, P; Hansen, T; Otto, M

2006-01-01

Aseptic hip prosthesis loosening is the most important long-term complication in total hip arthroplasty. Polyethylene (PE) wear is the dominant etiologic factor in aseptic loosening, which together with other factors induces mechanisms resulting in bone loss, and finally in implant loosening. The single-shot radiograph analysis (EBRA, abbreviation for the German term "Einzel-Bild-Röntgenanalyse") is a computerized method for early radiological prediction of aseptic loosening. In this study, EBRA parameters were correlated with histomorphological parameters of the periprosthetic membrane. Periprosthetic membranes obtained from 19 patients during revision surgery of loosened ABG I-type total hip pros-theses were analyzed histologically and morphometrically. The pre-existing EBRA parameters, the thickness of the PE debris lay-er and the dimension of inclination and anteversion, were compared with the density of macrophages and giant cells. Addi-tionally, the semiquantitatively determined density of lymphocytes, plasma cells, giant cells and the size of the necrotic areas were correlated with the EBRA results. All periprosthetic membranes were classified as debris-induced type membranes. We found a positive correlation between the number of giant cells and the thickness of the PE debris layer. There was no significant correlation between the number of macrophages or all semiquantitative parameters and EBRA parameters. The number of giant cells decreased with implant duration. The morphometrically measured number of foreign body giant cells more closely reflects the results of the EBRA. The semiquantitative estimation of giant cell density could not substitute for the morphometrical analysis. The density of macrophages, lymphocytes, plasma cells and the size of necrotic areas did not correlate with the EBRA parameters, indicating that there is no correlation with aseptic loosening.

Flow cytometry analysis of inflammatory cells isolated from the sciatic nerve and DRG after chronic constriction injury in mice.

PubMed

Liu, Liping; Yin, Yan; Li, Fei; Malhotra, Charvi; Cheng, Jianguo

2017-06-01

Cellular responses to nerve injury play a central role in the pathogenesis of neuropathic pain. However, the analysis of site specific cellular responses to nerve injury and neuropathic pain is limited to immunohistochemistry staining with numerous limitations. We proposed to apply flow cytometry to overcome some of the limitations and developed two protocols for isolation of cells from small specimens of the sciatic nerve and dorsal root ganglion (DRG) in mice. RESULTS AND COMPARASION WITH EXISTING: methods We found that both the non-enzymatic and enzymatic approaches were highly effective in harvesting a sufficient number of cells for flow cytometry analysis in normal and pathological conditions. The total number of cells in the injury site of the sciatic and its DRGs increased significantly 14days after chronic constriction injury (CCI) of the sciatic nerve, compared to sham surgery control or the contralateral control. The enzymatic approach yielded a significantly higher total number of cells and CD45 negative cells, suggesting that this approach allows for harvest of more resident cells, compared to the non-enzymatic method. The percentage of CD45 + /CD11b + cells was significantly increased in the sciatic nerve but not in the DRG. These results were consistent with both protocols. We thus offer two simple and effective protocols that allow for application of flow cytometry to the investigation of cellular and molecular mechanisms of neuropathic pain. Copyright © 2017 Elsevier B.V. All rights reserved.

Control of the red tide dinoflagellate Cochlodinium polykrikoides by ozone in seawater.

PubMed

Shin, Minjung; Lee, Hye-Jin; Kim, Min Sik; Park, Noh-Back; Lee, Changha

2017-02-01

The inactivation of C. polykrikoides, a red tide dinoflagellate, by ozonation was investigated in seawater by monitoring numbers of viable and total cells. Parameters affecting the inactivation efficacy of C. polykrikoides such as the ozone dose, initial cell concentration, pH, and temperature were examined. The viable cell number rapidly decreased in the initial stage of the reaction (mostly in 1-2 min), whereas the decrease in total cell number was relatively slow and steady. Increasing ozone dose and decreasing initial cell concentration increased the inactivation efficacy of C. polykrikoides, while increasing pH and temperature decreased the cell inactivation efficacy. The addition of humic acid (a promoter for the ozone decomposition) inhibited the inactivation of C. polykrikoides, whereas bicarbonate ion (an inhibitor for the ozone decomposition) accelerated the C. polykrikoides inactivation. Observations regarding the effects of pH, temperature, humic acid, and bicarbonate ion collectively indicate that the inactivation of C. polykrikoides by ozonation is mainly attributed to oxidative cell damages by molecular ozone, rather than by hydroxyl radical, produced during the ozone decomposition. At high ozone dose (e.g., 5 mg/L), hypobromous acid formed by the reaction of bromide with ozone may partially contribute to cell inactivation. The use of ozone of less than 1 mg/L produced 0.75-2.03 μg/L bromate. Copyright © 2016 Elsevier Ltd. All rights reserved.

[A Paired Case Controlled Study Comparing the Short-term Outcomes of Da Vinci RATS and VATS Approach for Non-small Cell Lung Cancer].

PubMed

Dai, Feng; Xu, Shiguang; Xu, Wei; Ding, Renquan; Liu, Bo; Meng, Hao; Kang, Yunteng; Meng, Xiangrui; Lin, Jie; Wang, Shumin

2018-03-20

Da Vinci Surgical System is one of the greatest inventions of the 20th century, which represents the development direction of the precise minimally invasive surgical techniques, the aim of this study was to comparing the short-term outcomes between da Vinci robot-assisted lobectomy and video-assisted thoracic surgery (VATS) lobectomy for non-small cell lung cancer. 45 pairs of non-small cell lung cancer patients underwent pulmonary lobectomy with da Vinci Robotic assisted thoracoscopic (RATS) and VATS approach during the same period from January 2014 to January 2017. The operative time, estimated blood loss (EBL), total number and total groups of dissected lymph nodes, postoperative duration of drainage, the first day volume of drainage, total volume of drainage were compared. No perioperative death and convertion to thoracotomy occured in both groups. There were significant difference between RATS group and VATS group in EBL [(50.30±32.33) mL vs (208.60±132.63) mL], the first day volume of drainage [(275.00±145.42) mL vs (347.60±125.80) mL], the dissected total number [(22.67±9.67) vs (15.51±5.41)] and total team [(6.31±1.43) vs (4.91±1.04)] of lymph node. There were no significant difference in other outcomes. RATS is safe and effective and took better short-outcomes than VATS in non-small cell lung cancer.

[Analysis of characteristics of mononuclear cells remaining in the leukoreduction system chamber of Trima Accel and their differentiation into dendritic cells].

PubMed

Lee, Yangsoon; Kim, Sinyoung; Lee, Seung-Tae; Kim, Han-Soo; Baek, Eun-Jung; Kim, Hyung Jin; Lee, MeeKyung; Kim, Hyun Ok

2009-08-01

We investigated the characteristics of the mononuclear cells remaining in the leukoreduction system (LRS) chambers of Trima Accel in comparison with those of standard buffy coat cells, and evaluated their potential for differentiation into dendritic cells. Twenty-six LRS chambers of Trima Accel were collected after platelet pheresis from healthy adults. Flow cytometric analysis for T, B, NK, and CD14+ cells was performed and the number of CD34+ cells was counted. Differentiation and maturation into dendritic cells were induced using CD14+ cells seperated via Magnetic cell sorting (MACS) Seperation (Miltenyi Biotec Inc., USA). Total white blood cell (WBC) count in LRS chambers was 10.8 x 10(8) (range 7.7-18.0 x 10(8)). The median values (range) of proportions of each cells were CD4+ T cell 29.6% (18.7-37.6), CD8+ T cell 27.7% (19.2-40.0), B cell 5.5% (2.2-12.1), NK cell 15.7% (13.7-19.9), and CD14+ cells 12.4% (8.6-32.3) respectively. Although total WBC count was significantly higher in the buffy coat (whole blood of 400 mL) than the LRS chambers, the numbers of lymphocytes and monocytes were not statistically different. The numbers of B cells and CD4+ cells were significantly higher in the buffy coat than the LRS chambers (P<0.05). The median value (range) of CD34+ cells obtained from the LRS chambers was 0.9 x 10(6) (0.2-2.6 x 10(6)). After 7 days of cytokine-supplemented culture, the CD14+ cells were successfully differentiated into dendritic cells. The mononuclear cells in LRS chambers of Trima Accel are an excellent alternative source of viable and functional human blood cells, which can be used for research purposes.

Ratio of metastatic lymph nodes to total number of nodes resected is prognostic for survival in esophageal carcinoma.

PubMed

Kelty, Clive J; Kennedy, Catherine W; Falk, Gregory L

2010-09-01

The role of the number of metastatic nodes in esophageal cancer surgery is of interest. We assess predictors of survival after oesophagectomy for esophageal and gastroesophageal junction malignancy. Prospective data of consecutive patients undergoing oesophagectomy and systematic lymphadenectomy between 1991 and 2007. Of 224 patients, 148 patients (66%) had adenocarcinoma, 70 (31%) squamous cell carcinoma, and 6 (2.6%) were other tumor types. Five-year survival was 43% with hospital mortality of 3.5%. Locoregional recurrence occurred in 14%. The total number of affected nodes significantly reduced survival (four or more metastatic nodes). Further analysis of the ratio of nodes affected to the total number resected showed a significant decrease in survival as the percentage of positive nodes increased (p < 0.001). Patients undergoing surgery for esophageal cancer should be staged according to a minimum total number of metastatic lymph nodes and ratios because this more accurately predicts survival than current staging systems.

Stacked microbial desalination cells to enhance water desalination efficiency.

PubMed

Chen, Xi; Xia, Xue; Liang, Peng; Cao, Xiaoxin; Sun, Haotian; Huang, Xia

2011-03-15

Microbial desalination cell (MDC) is a new method to obtain clean water from brackish water using electricity generated from organic matters by exoelectrogenic bacteria. Anions and cations, derived from salt solution filled in the desalination chamber between the anode and cathode, move to the anode and cathode chambers under the force of electrical field, respectively. On the basis of the primitive single-desalination-chambered MDC, stacked microbial desalination cells (SMDCs) were developed in order to promote the desalination rate in the present study. The effects of desalination chamber number and external resistance were investigated. Results showed that a remarkable increase in the total desalination rate (TDR) could be obtained by means of increasing the desalination cell number and reducing the external resistance, which caused the charge transfer efficiency increased since the SMDCs enabled more pairs of ions separated while one electron passed through the external circuit. The maximum TDR of 0.0252 g/h was obtained using a two-desalination-chambered SMDC with an external resistance of 10 Ω, which was 1.4 times that of single-desalination-chambered MDC. SMDCs proved to be an effective approach to increase the total water desalination rate if provided a proper desalination chamber number and external resistance.

CD23 surface density on B cells is associated with IgE levels and determines IgE-facilitated allergen uptake, as well as activation of allergen-specific T cells.

PubMed

Selb, Regina; Eckl-Dorna, Julia; Neunkirchner, Alina; Schmetterer, Klaus; Marth, Katharina; Gamper, Jutta; Jahn-Schmid, Beatrice; Pickl, Winfried F; Valenta, Rudolf; Niederberger, Verena

2017-01-01

Increasing evidence suggests that the low-affinity receptor for IgE, CD23, plays an important role in controlling the activity of allergen-specific T cells through IgE-facilitated allergen presentation. We sought to determine the number of CD23 molecules on immune cells in allergic patients and to investigate whether the number of CD23 molecules on antigen-presenting cells is associated with IgE levels and influences allergen uptake and allergen-specific T-cell activation. Numbers of CD23 molecules on immune cells of allergic patients were quantified by using flow cytometry with QuantiBRITE beads and compared with total and allergen-specific IgE levels, as well as with allergen-induced immediate skin reactivity. Allergen uptake and allergen-specific T-cell activation in relation to CD23 surface density were determined by using flow cytometry in combination with confocal microscopy and T cells transfected with the T-cell receptor specific for the birch pollen allergen Bet v 1, respectively. Defined IgE-allergen immune complexes were formed with human monoclonal allergen-specific IgE and Bet v 1. In allergic patients the vast majority of CD23 molecules were expressed on naive IgD + B cells. The density of CD23 molecules on B cells but not the number of CD23 + cells correlated with total IgE levels (R S  = 0.53, P = .03) and allergen-induced skin reactions (R S  = 0.63, P = .008). Uptake of allergen-IgE complexes into B cells and activation of allergen-specific T cells depended on IgE binding to CD23 and were associated with CD23 surface density. Addition of monoclonal IgE to cultured PBMCs significantly (P = .04) increased CD23 expression on B cells. CD23 surface density on B cells of allergic patients is correlated with allergen-specific IgE levels and determines allergen uptake and subsequent activation of T cells. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Optimal spatial sampling techniques for ground truth data in microwave remote sensing of soil moisture

NASA Technical Reports Server (NTRS)

Rao, R. G. S.; Ulaby, F. T.

1977-01-01

The paper examines optimal sampling techniques for obtaining accurate spatial averages of soil moisture, at various depths and for cell sizes in the range 2.5-40 acres, with a minimum number of samples. Both simple random sampling and stratified sampling procedures are used to reach a set of recommended sample sizes for each depth and for each cell size. Major conclusions from statistical sampling test results are that (1) the number of samples required decreases with increasing depth; (2) when the total number of samples cannot be prespecified or the moisture in only one single layer is of interest, then a simple random sample procedure should be used which is based on the observed mean and SD for data from a single field; (3) when the total number of samples can be prespecified and the objective is to measure the soil moisture profile with depth, then stratified random sampling based on optimal allocation should be used; and (4) decreasing the sensor resolution cell size leads to fairly large decreases in samples sizes with stratified sampling procedures, whereas only a moderate decrease is obtained in simple random sampling procedures.

Protective effect of total flavonoids of seabuckthorn (Hippophae rhamnoides) in simulated high-altitude polycythemia in rats.

PubMed

Zhou, Ji-Yin; Zhou, Shi-Wen; Du, Xiao-Huang; Zeng, Sheng-Ya

2012-09-28

Seabuckthorn (Hippophae rhamnoides L.) has been used to treat high altitude diseases. The effects of five-week treatment with total flavonoids of seabuckthorn (35, 70, 140 mg/kg, ig) on cobalt chloride (5.5 mg/kg, ip)- and hypobaric chamber (simulating 5,000 m)-induced high-altitude polycythemia in rats were measured. Total flavonoids decreased red blood cell number, hemoglobin, hematocrit, mean corpuscular hemoglobin levels, span of red blood cell electrophoretic mobility, aggregation index of red blood cell, plasma viscosity, whole blood viscosity, and increased deformation index of red blood cell, erythropoietin level in serum. Total flavonoids increased pH, pO₂, Sp(O₂), pCO₂ levels in arterial blood, and increased Na⁺, HCO₃⁻, Cl⁻, but decreased K⁺ concentrations. Total flavonoids increased mean arterial pressure, left ventricular systolic pressure, end-diastolic pressure, maximal rate of rise and decrease, decreased heart rate and protected right ventricle morphology. Changes in hemodynamic, hematologic parameters, and erythropoietin content suggest that administration of total flavonoids from seabuckthorn may be useful in the prevention of high altitude polycythaemia in rats.

Cancer cell uptake behavior of Au nanoring and its localized surface plasmon resonance induced cell inactivation

NASA Astrophysics Data System (ADS)

Chu, Che-Kuan; Tu, Yi-Chou; Chang, Yu-Wei; Chu, Chih-Ken; Chen, Shih-Yang; Chi, Ting-Ta; Kiang, Yean-Woei; Yang, Chih-Chung

2015-02-01

Au nanorings (NRIs), which have the localized surface plasmon resonance (LSPR) wavelength around 1058 nm, either with or without linked antibodies, are applied to SAS oral cancer cells for cell inactivation through the LSPR-induced photothermal effect when they are illuminated by a laser of 1065 nm in wavelength. Different incubation times of cells with Au NRIs are considered for observing the variations of cell uptake efficiency of Au NRI and the threshold laser intensity for cell inactivation. In each case of incubation time, the cell sample is washed for evaluating the total Au NRI number per cell adsorbed and internalized by the cells based on inductively coupled plasma mass spectrometry measurement. Also, the Au NRIs remaining on cell membrane are etched with KI/I2 solution to evaluate the internalized Au NRI number per cell. The threshold laser intensities for cell inactivation before washout, after washout, and after KI/I2 etching are calibrated from the circular area sizes of inactivated cells around the illuminated laser spot center with various laser power levels. By using Au NRIs with antibodies, the internalized Au NRI number per cell increases monotonically with incubation time up to 24 h. However, the number of Au NRI remaining on cell membrane reaches a maximum at 12 h in incubation time. The cell uptake behavior of an Au NRI without antibodies is similar to that with antibodies except that the uptake NRI number is significantly smaller and the incubation time for the maximum NRI number remaining on cell membrane is delayed to 20 h. By comparing the threshold laser intensities before and after KI/I2 etching, it is found that the Au NRIs remaining on cell membrane cause more effective cancer cell inactivation, when compared with the internalized Au NRIs.

MORPHOLOGICAL AND CULTURAL COMPARISON OF MICROORGANISMS IN SURFACE SOIL AND SUBSURFACE SEDIMENTS AT A PRISTINE STUDY SITE IN OKLAHOMA (JOURNAL VERSION)

EPA Science Inventory

Surface-soil and subsurface microfloras at the site of a shallow aquifer in Oklahoma were examined and compared with respect to (1) total and viable cell numbers, (2) colony and cell types that grew on various plating media, (3) cell morphologies seen in flotation films stripped ...

[Changes and clinical significance of peripheral blood natural killer cells in neonates with bacterial pneumonia].

PubMed

Li, Qiuling; Weng, Kaizhi; Zhu, Ling; Mei, Xuqiao; Xu, Liping; Lin, Jiehua

2014-10-01

To detect the percentage of total natural killer (NK) cells and its different populations in the peripheral blood from neonates with bacterial pneumonia and discuss the clinical significance of NK cells in the pathogenesis of bacterial pneumonia. Flow cytometry was performed to detect the percentages of NK cells and its subsets in peripheral blood lymphocytes from 38 cases of neonatal bacterial pneumonias and 18 cases of normal neonates. Patients recruited were divided into two groups according to hospitalization days and numbers of peripheral leukocytes: hospitalization days within 10 days (including 10 days) as group A, and more than 10 days as group B; the number of peripheral blood leukocytes <5.0×10(9)/L or >20.0×10(9)/L as severe infection group, and 5.0×10(9)/L< number of peripheral blood leukocytes <20.0×10(9)/L as mild infection group. The percentages of peripheral blood NK cells and CD3(-)CD56(neg)CD16(bright) subset in the neonates with bacterial pneumonia were significantly lower than those of the normal newborns (P<0.01), but there were no statistically significant differences in CD3(-)CD56(bright)CD16(neg/dim) and CD3(-)CD56(dim)CD16(bright) subsets. The percentage of CD3(-)CD56(neg)CD16(bright) subset in group A was significantly lower than that of the normal newborns (P<0.01), while the percentages of the total NK cells and other subsets had no statistical significance. The neonates with bacterial pneumonia had significantly lower percentages of the total NK cells and CD3(-)CD56(neg)CD16(bright) subset in group B as compared with the normal neonates (P<0.01). And the percentages of the total NK cells and its subsets in group B were also lower than those in group A (P<0.05). The percentages of NK cells and each subset in severe infection group were significantly lower than those in mild infection group (P<0.05). To the neonates who suffer from bacterial pneumonia, the more serious and the longer hospital stay, the lower the percentages of NK cells and its subsets are.

HIV dynamics with multiple infections of target cells.

PubMed

Dixit, Narendra M; Perelson, Alan S

2005-06-07

The high incidence of multiple infections of cells by HIV sets the stage for rapid HIV evolution by means of recombination. Yet how HIV dynamics proceeds with multiple infections remains poorly understood. Here, we present a mathematical model that describes the dynamics of viral, target cell, and multiply infected cell subpopulations during HIV infection. Model calculations reproduce several experimental observations and provide key insights into the influence of multiple infections on HIV dynamics. We find that the experimentally observed scaling law, that the number of cells coinfected with two distinctly labeled viruses is proportional to the square of the total number of infected cells, can be generalized so that the number of triply infected cells is proportional to the cube of the number of infected cells, etc. Despite the expectation from Poisson statistics, we find that this scaling relationship only holds under certain conditions, which we predict. We also find that multiple infections do not influence viral dynamics when the rate of viral production from infected cells is independent of the number of times the cells are infected, a regime expected when viral production is limited by cellular rather than viral factors. This result may explain why extant models, which ignore multiple infections, successfully describe viral dynamics in HIV patients. Inhibiting CD4 down-modulation increases the average number of infections per cell. Consequently, altering CD4 down-modulation may allow for an experimental determination of whether viral or cellular factors limit viral production.

HIV dynamics with multiple infections of target cells

PubMed Central

Dixit, Narendra M.; Perelson, Alan S.

2005-01-01

The high incidence of multiple infections of cells by HIV sets the stage for rapid HIV evolution by means of recombination. Yet how HIV dynamics proceeds with multiple infections remains poorly understood. Here, we present a mathematical model that describes the dynamics of viral, target cell, and multiply infected cell subpopulations during HIV infection. Model calculations reproduce several experimental observations and provide key insights into the influence of multiple infections on HIV dynamics. We find that the experimentally observed scaling law, that the number of cells coinfected with two distinctly labeled viruses is proportional to the square of the total number of infected cells, can be generalized so that the number of triply infected cells is proportional to the cube of the number of infected cells, etc. Despite the expectation from Poisson statistics, we find that this scaling relationship only holds under certain conditions, which we predict. We also find that multiple infections do not influence viral dynamics when the rate of viral production from infected cells is independent of the number of times the cells are infected, a regime expected when viral production is limited by cellular rather than viral factors. This result may explain why extant models, which ignore multiple infections, successfully describe viral dynamics in HIV patients. Inhibiting CD4 down-modulation increases the average number of infections per cell. Consequently, altering CD4 down-modulation may allow for an experimental determination of whether viral or cellular factors limit viral production. PMID:15928092

Mechanisms for capacity fading in the NiH2 cell and its effects on cycle life

NASA Technical Reports Server (NTRS)

Zimmerman, Albert H.

1993-01-01

During recent years there have been a number of instances where the capacity of nickel hydrogen battery cells has proven to be unstable during storage. The capacity losses seen after periods of cell or battery storage have typically varied from only a small amount of fading, up to about 30 percent of the total cell capacity. Detailed studies into the root causes for such fading have been carried out in a number of instances. This report provides an overview of the different mechanisms that have been found to be responsible for such capacity fading in nickel hydrogen cells, and summarizes the presently available data on how each responsible mechanism affects ultimate cell cycle life.

The effect of short G-CSF administration on the numbers and clonogenic efficiency of hematopoietic progenitor cells in bone marrow and peripheral blood of normal donors.

PubMed

Zaucha, J M; Knopińska-Posłuszny, W; Bieniaszewska, M; Myśliwski, A; Hellmann, A

2000-01-01

We have analysed the cellularity, the number of clonogenic cells and their clonogenic efficiency (the number of clonogenic cells/2 x 10(5) MNC) in peripheral blood (PB) and bone marrow (BM) during and after filgrastim (rhG-CSF) mobilization of CD34+ cells in 12 healthy donors for allogeneic stem cell donation. G-CSF was administrated subcutaneously for 5 consecutive days at a dose of 10 micrograms/kg/day. WBC, MNC, CD34+ cell counts, CFU-GM and BFU-E assays in PB were performed at baseline and then daily 12 hours after each G-CSF dose. BM was assayed before start (day 1) and after the last dose (day 6) of G-CSF. Results are given as medians, with ranges in parentheses. In PB the total WBC and MNC increased 7.4-fold (6.0-12.3) and 3.3-fold (1.5-9.4), respectively, reaching a peak of 49.4 x 10(9)/l (32.5-66.6) on day 6 for WBC and 6.28 x 10(9)/l (4.7-13.3) for MNC on day 5. CD34+ cell number reached a peak value of 48.0 x 10(6)/l (45.6-285) on day 6 whereas CFU-GM and BFU-E reached their peaks on day 5, 0.95 x 10(4)/ml (0.05-6.08) and 1.04 x 10(4)/ml, respectively. CFU-MIX, not detectable at baseline, reached a peak of 0.95 x 10(4)/ml (0.006-0.51) on day 5 as well. This was accompanied by an increase in CFU-GM, BFU-E and CFU-MIX clonogenic efficiency: 23-fold (3-150), 9.75-fold (2.2-27.8) and 20-fold (2.5-210), respectively. In BM the total WBC number increased 2.5-fold (1.3-4.9) from the baseline value of 52.6 x 10(9)/l (7.9-137.0) whereas the MNC count increased 2.0-fold (0.81-3.7) from a baseline of 13.6 x 10(9)/l (3.5-54.8). This was, however, not significant. The number of CD34+ cells increased significantly 2.9-fold (0.8-8.3). In 8 donors CFU-MIX were detectable before but not after G-CSF treatment. A similar decrease in CFU-GM and BFU-E clonogenic efficiency occurred but was not significant. CFU-GM and BFU-E numbers did not change. We conclude that the total body numbers of lineage committed progenitors increased during G-CSF administration, which indicate their proliferation in addition to mobilization. The effect of G-CSF on the number of more primitive progenitors in BM is less clear and needs further investigation.

Peripheral natural killer cytotoxicity and CD56(pos)CD16(pos) cells increase during early pregnancy in women with a history of recurrent spontaneous abortion.

PubMed

Emmer, P M; Nelen, W L; Steegers, E A; Hendriks, J C; Veerhoek, M; Joosten, I

2000-05-01

For diagnostic purposes we assessed peripheral natural killer (NK) cell cytotoxicity and NK and T cell numbers to assess their putative predictive value in recurrent spontaneous abortion (RSA). A total of 43 women with subsequent pregnancy, 37 healthy controls and 39 women successfully partaking in an in-vitro fertilization (IVF) procedure, were included in the study. We show that before pregnancy, levels of NK cytotoxicity and numbers of both single CD56(pos) and double CD56(pos)CD16(pos) cells were similar between RSA women and controls. But notably, within the RSA group, NK cell numbers of <12% were strongly associated with a subsequent pregnancy carried to term. Supplementation of folic acid led to an increase of single CD56(pos) cells, but cytotoxic function appeared unaffected. The expression pattern of killer inhibitory receptors on CD56(pos) cells was not different between patients and controls. A longitudinal study revealed that, compared with controls, in RSA women higher numbers of double CD56(pos)CD16(pos) cells were present during early pregnancy, paralleled by an increase in cytotoxic NK cell reactivity. The single CD56(pos) population decreased in number. In conclusion, the analysis of peripheral NK cell characteristics appears a suitable diagnostic tool in RSA. Immunomodulation aimed at NK cell function appears a promising therapeutic measure.

Poly(ADP-ribose) polymerase-1 (Parp-1)-deficient mice demonstrate abnormal antibody responses

PubMed Central

Ambrose, Helen E; Willimott, Shaun; Beswick, Richard W; Dantzer, Françoise; de Murcia, Josiane Ménissier; Yelamos, José; Wagner, Simon D

2009-01-01

Poly(ADP-ribosylation) of acceptor proteins is an epigenetic modification involved in DNA strand break repair, recombination and transcription. Here we provide evidence for the involvement of poly(ADP-ribose) polymerase-1 (Parp-1) in antibody responses. Parp-1−/− mice had increased numbers of T cells and normal numbers of total B cells. Marginal zone B cells were mildly reduced in number, and numbers of follicular B cells were preserved. There were abnormal levels of basal immunoglobulins, with reduced levels of immunoglobulin G2a (IgG2a) and increased levels of IgA and IgG2b. Analysis of specific antibody responses showed that T cell-independent responses were normal but T cell-dependent responses were markedly reduced. Germinal centres were normal in size and number. In vitro purified B cells from Parp-1−/− mice proliferated normally and showed normal IgM secretion, decreased switching to IgG2a but increased IgA secretion. Collectively our results demonstrate that Parp-1 has essential roles in normal T cell-dependent antibody responses and the regulation of isotype expression. We speculate that Parp-1 forms a component of the protein complex involved in resolving the DNA double-strand breaks that occur during class switch recombination. PMID:18778284

Effects of first-line chemotherapy on natural killer cells in adult T-cell leukemia-lymphoma and peripheral T-cell lymphoma.

PubMed

Ogura, Michinori; Ishida, Takashi; Tsukasaki, Kunihiro; Takahashi, Takeshi; Utsunomiya, Atae

2016-07-01

Natural killer (NK) cells are well known to be the most important effector cells mediating antibody-dependent cellular cytotoxicity (ADCC) which is an important mechanism of action of antibody drugs. We evaluated the effects of chemotherapy on the cell number and activity of NK cells from patients who received the vincristine-cyclophosphamide-doxorubicin-prednisone (VCAP), doxorubicin-ranimustine-prednisone (AMP), and vindesine-etoposide-carboplatin-prednisone (VECP) (mLSG15) or mLSG15-like (-L) regimen, which is one of the standard of cares for newly diagnosed adult T-cell leukemia-lymphoma (ATL), or the cyclophosphamide-doxorubicin-vincristine-prednisone (CHOP) or CHOP-L regimen which is another standard of care for ATL and peripheral T-cell lymphoma (PTCL). The number of lymphocytes and NK cells, and NK cell activity, were assessed using flow cytometry and a (51)Cr release assay, respectively. A total of 26 patients with untreated ATL or PTCL were enrolled, and blood samples from 25 patients were evaluable. NK cell number in ATL decreased after mLSG15/-L treatment, and the degree of decrease in the NK cell number was more prominent just before VECP therapy (Day 15-17 of each cycle) than just before VCAP therapy (Day 1 of each cycle). The NK cell number in ATL after CHOP/-L treatment also decreased. Interestingly, the NK cell activity showed a tendency to increase after the treatment. NK cell number in PTCL did not decrease by CHOP/-L regimen, but the activity was slightly decreased after the treatment. These results indicate that the effects of chemotherapeutic agents on NK cells vary according to the disease type and intensity of chemotherapy.

Integrated micro-optofluidic platform for real-time detection of airborne microorganisms

NASA Astrophysics Data System (ADS)

Choi, Jeongan; Kang, Miran; Jung, Jae Hee

2015-11-01

We demonstrate an integrated micro-optofluidic platform for real-time, continuous detection and quantification of airborne microorganisms. Measurements of the fluorescence and light scattering from single particles in a microfluidic channel are used to determine the total particle number concentration and the microorganism number concentration in real-time. The system performance is examined by evaluating standard particle measurements with various sample flow rates and the ratios of fluorescent to non-fluorescent particles. To apply this method to real-time detection of airborne microorganisms, airborne Escherichia coli, Bacillus subtilis, and Staphylococcus epidermidis cells were introduced into the micro-optofluidic platform via bioaerosol generation, and a liquid-type particle collection setup was used. We demonstrate successful discrimination of SYTO82-dyed fluorescent bacterial cells from other residue particles in a continuous and real-time manner. In comparison with traditional microscopy cell counting and colony culture methods, this micro-optofluidic platform is not only more accurate in terms of the detection efficiency for airborne microorganisms but it also provides additional information on the total particle number concentration.

Chloroplast Growth and Replication in Germinating Spinach Cotyledons following Massive γ-Irradiation of the Seed

PubMed Central

Rose, Ray; Possingham, John

1976-01-01

Spinach seeds (Spinacia oleracea L.) given massive doses of γ-irradiation (500 krad) germinate and form a seedling with two green cotyledons and a radicle, but develop no further. Irradiated cotyledons show no increase in cell number or total DNA over a 7-day period in the light, while in control cotyledons there is a small increase in cell number and large increases in total DNA and chloroplast number. The chloroplasts of irradiated cotyledons are delayed in their division, become greatly enlarged and contain large amounts of starch. The whole population of chloroplasts subsequently undergoes a wave of division. The daughter chloroplasts show normal thylakoid development, but have some abnormal structural features caused by the radiation stress. Information on the effect of X-irradiation, ultraviolet irradiation, and 5-fluorodeoxyuridine on chloroplast replication and on chloroplast and nuclear DNA synthesis was obtained from cultured spinach leaf discs. It appears that chloroplast replication is more resistant to ionizing radiation than cell division and can proceed in the absence of nuclear DNA synthesis and greatly reduced chloroplast DNA synthesis. Images PMID:16659421

Integrated micro-optofluidic platform for real-time detection of airborne microorganisms

PubMed Central

Choi, Jeongan; Kang, Miran; Jung, Jae Hee

2015-01-01

We demonstrate an integrated micro-optofluidic platform for real-time, continuous detection and quantification of airborne microorganisms. Measurements of the fluorescence and light scattering from single particles in a microfluidic channel are used to determine the total particle number concentration and the microorganism number concentration in real-time. The system performance is examined by evaluating standard particle measurements with various sample flow rates and the ratios of fluorescent to non-fluorescent particles. To apply this method to real-time detection of airborne microorganisms, airborne Escherichia coli, Bacillus subtilis, and Staphylococcus epidermidis cells were introduced into the micro-optofluidic platform via bioaerosol generation, and a liquid-type particle collection setup was used. We demonstrate successful discrimination of SYTO82-dyed fluorescent bacterial cells from other residue particles in a continuous and real-time manner. In comparison with traditional microscopy cell counting and colony culture methods, this micro-optofluidic platform is not only more accurate in terms of the detection efficiency for airborne microorganisms but it also provides additional information on the total particle number concentration. PMID:26522006

The organization of human epidermis: functional epidermal units and phi proportionality.

PubMed

Hoath, Steven B; Leahy, D G

2003-12-01

The concept that mammalian epidermis is structurally organized into functional epidermal units has been proposed on the basis of stratum corneum (SC) architecture, proliferation kinetics, melanocyte:keratinocyte ratios (1:36), and, more recently, Langerhans cell: epidermal cell ratios (1:53). This article examines the concept of functional epidermal units in human skin in which the maintenance of phi (1.618034) proportionality provides a central organizing principle. The following empirical measurements were used: 75,346 nucleated epidermal cells per mm2, 1394 Langerhans cells per mm2, 1999 melanocytes per mm2, 16 (SC) layers, 900-microm2 corneocyte surface area, 17,778 corneocytes per mm2, 14-d (SC) turnover time, and 93,124 per mm2 total epidermal cells. Given these empirical data: (1) the number of corneocytes is a mean proportional between the sum of the Langerhans cell + melanocyte populations and the number of epidermal cells, 3393/17,778-17,778/93,124; (2) the ratio of nucleated epidermal cells over corneocytes is phi proportional, 75,346/17,778 approximately phi3; (3) assuming similar 14-d turnover times for the (SC) and Malpighian epidermis, the number of corneocytes results from subtraction of a cellular fraction equal to approximately 2/phi2 x the number of living cells, 75,436 - (2/phi2 x 75,346) approximately 17,778; and (4) if total epidermal turnover time equals (SC) turnover time x the ratio of living/dead cells, then compartmental turnover times are unequal (14 d for (SC) to 45.3 d for nucleated epidermis approximately 1/2phi) and cellular replacement rates are 52.9 corneocytes/69.3 keratinocytes per mm2 per h approximately 2/phi2. These empirically derived equivalences provide logicomathematical support for the presence of functional epidermal units in human skin. Validation of a phi proportional unit architecture in human epidermis will be important for tissue engineering of skin and the design of instruments for skin measurement.

Hippocampal neurogenesis and cortical cellular plasticity in Wahlberg's epauletted fruit bat: a qualitative and quantitative study.

PubMed

Gatome, Catherine W; Mwangi, Deter K; Lipp, Hans-Peter; Amrein, Irmgard

2010-01-01

Species-specific characteristics of neuronal plasticity emerging from comparative studies can address the functional relevance of hippocampal or cortical plasticity in the light of ecological adaptation and evolutionary history of a given species. Here, we present a quantitative and qualitative analysis of neurogenesis in young and adult free-living Wahlberg's epauletted fruit bats. Using the markers for proliferating cell nuclear antigen (PCNA), bromodeoxyuridine (BrdU), doublecortin (DCX) and polysialic acid neural cell adhesion molecule (PSA-NCAM), our findings in the hippocampus, olfactory bulb and cortical regions are described and compared to reports in other mammals. Expressed as a percentage of the total number of granule cells, PCNA- and BrdU-positive cells accounted for 0.04 in young to 0.01% in adult animals; DCX-positive cells for 0.05 (young) to 0.01% (adult); PSA-NCAM-positive cells for 0.1 (young) to 0.02% (adult), and pyknotic cells for 0.007 (young) to 0.005% (adult). The numbers were comparable to other long-lived, late-maturing mammals such as primates. A significant increase in the total granule cell number from young to adult animals demonstrated the successful formation and integration of new cells. In adulthood, granule cell number appeared stable and was surprisingly low in comparison to other species. Observations in the olfactory bulb and rostral migratory stream were qualitatively similar to descriptions in other species. In the ventral horn of the lateral ventricle, we noted prominent expression of DCX and PSA-NCAM forming a temporal migratory stream targeting the piriform cortex, possibly reflecting the importance of olfaction to these species. Low, but persistent hippocampal neurogenesis in non-echolocating fruit bats contrasted the findings in echolocating microbats, in which hippocampal neurogenesis was largely absent. Together with the observed intense cortical plasticity in the olfactory system of fruit bats we suggest a differential influence of sensory modalities on hippocampal and cortical plasticity in this mammalian order. Copyright © 2010 S. Karger AG, Basel.

Cell apoptosis and lipid content of in vitro-produced, vitrified bovine embryos treated with forskolin.

PubMed

Paschoal, Daniela Martins; Sudano, Mateus José; Schwarz, Kátia Regina Lancellotti; Maziero, Rosiára Rosário Dias; Guastali, Midyan Daroz; Crocomo, Letícia Ferrari; Magalhães, Luis Carlos Oña; Martins, Alício; Leal, Claudia Lima Verde; Landim-Alvarenga, Fernanda da Cruz

2017-01-01

The presence of fetal calf serum in culture medium influences embryo quality, causing a reduction in postcryopreservation survival. Forskolin has been used to induce lipolysis and increase cryotolerance, functioning as an activator of adenylate cyclase and elevating cAMP levels. In the present experiment, bovine zygotes were cultured in synthetic oviduct fluid with amino acid plus 2.5% fetal calf serum for 6 days, when forskolin was added in three concentrations: 2.5, 5, and 10 μM. Treatment with forskolin lasted for 24 hours. Blastocyst formation rate, quantification of lipid granules, total cell numbers, and apoptosis rate were evaluated. In a second assessment, embryos were vitrified, and warming, re-expansion rate, total cell numbers, and apoptosis rate were also evaluated. There was no difference due to forskolin in blastocyst formation or re-expansion rates after vitrification. However, lipid measurements were lower (control: 136.8 and F 2.5 μM: 128.5; P < 0.05), and number of cells per embryo higher (control: 140.1 and F 2.5 μM: 173.5; P < 0.05) than controls for 2.5 μM forskolin but not for higher forskolin concentrations. The number of intact cells per embryo was higher, and the rate of apoptosis was lower in fresh than in vitrified embryos (number of cells of warmed embryos, control: 104.1, F 2.5 μM: 101.3, F 5 μM: 115.4, F 10 μM: 95.1; apoptotic of fresh cells, control: 12.1%, F 2.5 μM: 16.7%, F 5 μM: 11.1%, F 10 μM: 14.2%; and apoptotic warmed embryos, control: 22.3%, F 2.5 μM: 37.3%, F 5 μM: 33.2%, F 10 μM: 30.3%; P < 0.05). It was concluded that forskolin is an effective lipolytic agent even at low concentrations, leading to formation of blastocysts with a comparatively larger number of cells. Copyright © 2016 Elsevier Inc. All rights reserved.

The number of satellite cells in slow and fast fibres from human vastus lateralis muscle.

PubMed

Kadi, Fawzi; Charifi, Nadia; Henriksson, Jan

2006-07-01

The aim of this investigation was to study the distribution of satellite cells in slow (type I fibres) and fast (type II fibres) fibres from human vastus lateralis muscle. This muscle is characterised by a mixed fibre type composition and is considered as the site of choice for biopsies in research work and for clinical diagnosis. Biopsy samples were obtained from five healthy young volunteers and a total of 1,747 type I fibres and 1,760 type II fibres were assessed. Satellite cells and fibre type composition were studied on serial muscle cross-sections stained with specific monoclonal antibodies. From a total of 218 satellite cells, 116 satellite cells were found in contact with type I fibres (53.6+/-8% of the satellite cells associated to type I fibres) and 102 satellite cells in contact with type II fibres (46.4+/-8% of the satellite cells associated to type II fibres). There was no significant difference (P=0.4) between the percentages of satellite cells in contact with type I and with type II fibres. Additionally, there was no relationship between the mean number of satellite cells per fibre and the mean cross-sectional area of muscle fibres. In conclusion, our results show that there is no fibre type-specific distribution of satellite cells in a human skeletal muscle with mixed fibre type composition.

Positioning of the NOR-bearing chromosomes in relation to nucleoli in daughter cells after mitosis.

PubMed

Kalmárová, M; Smirnov, E; Kovácik, L; Popov, A; Raska, I

2008-01-01

It is known that chromosomes occupy non-random positions in the cell nucleus. However, it is not clear to what extent their nuclear positions, together with their neighborhood, are conserved in daughter cells. To address specific aspects of this problem, we used the model of the chromosomes carrying ribosomal genes that are organized in clusters termed Nucleolus Organizer Regions (NORs). We compared the association of chosen NOR-bearing chromosomes (NOR-chromosomes) with nucleoli, as well as the numbers of nucleoli, in the pairs of daughter cells, and established how frequently the daughter cells had equal numbers of the homologs of certain NOR-chromosomes associated with individual nucleoli. The daughter cells typically had different numbers of nucleoli. At the same time, using immuno-FISH with probes for chromosomes 14 and 15 in HeLa cells, we found that the cell pairs with identical combinations appeared significantly more frequently than predicted by the random model. Thus, although the total number of chromosomes associated with nucleoli is variable, our data indicate that the position of the NOR-bearing chromosomes in relation to nucleoli is partly conserved through mitosis.

Positioning of the NOR-Bearing Chromosomes in Relation to Nucleoli in Daughter Cells after Mitosis

PubMed Central

KALMÁROVÁ, M.; SMIRNOV, E.; KOVÁČIK, L.; POPOV, A.; RAŠKA, I.

2008-01-01

Summary It is known that chromosomes occupy non-random positions in the cell nucleus. However, it is not clear to what extent their nuclear positions, together with their neighborhood, are conserved in daughter cells. To address specific aspects of this problem, we used the model of the chromosomes carrying ribosomal genes that are organized in clusters termed Nucleolus Organizer Regions (NORs). We compared the association of chosen NOR-bearing chromosomes (NOR-chromosomes) with nucleoli, as well as the numbers of nucleoli, in the pairs of daughter cells, and established how frequently the daughter cells had equal numbers of the homologs of certain NOR-chromosomes associated with individual nucleoli. The daughter cells typically had different numbers of nucleoli. At the same time, using immuno-FISH with probes for chromosomes 14 and 15 in HeLa cells, we found that the cell pairs with identical combinations appeared significantly more frequently than predicted by the random model. Thus, although the total number of chromosomes associated with nucleoli is variable, our data indicate that the position of the NOR-bearing chromosomes in relation to nucleoli is partly conserved through mitosis. PMID:18597585

Are periprosthetic tissue reactions observed after revision of total disc replacement comparable to the reactions observed after total hip or knee revision surgery?

PubMed Central

Punt, Ilona M.; Austen, Shennah; Cleutjens, Jack P.M.; Kurtz, Steven M.; ten Broeke, René H.M.; van Rhijn, Lodewijk W.; Willems, Paul C.; van Ooij, André

2011-01-01

Study design Comparative study. Objective To compare periprosthetic tissue reactions observed after total disc replacement (TDR), total hip arthroplasty (THA) and total knee arthroplasty (TKA) revision surgery. Summary of background data Prosthetic wear debris leading to particle disease, followed by osteolysis, is often observed after THA and TKA. Although the presence of polyethylene (PE) particles and periprosthetic inflammation after TDR has been proven recently, osteolysis is rarely observed. The clinical relevance of PE wear debris in the spine remains poorly understood. Methods Number, size and shape of PE particles, as well as quantity and type of inflammatory cells in periprosthetic tissue retrieved during Charité TDR (n=22), THA (n=10) and TKA (n=4) revision surgery were compared. Tissue samples were stained with hematoxylin/eosin and examined by using light microscopy with bright field and polarized light. Results After THA, large numbers of PE particles <6 µm were observed, which were mainly phagocytosed by macrophages. The TKA group had a broad size range with many larger PE particles and more giant cells. In TDR, the size range was similar to that observed in TKA. However, the smallest particles were the most prevalent with 75% of the particles being <6 µm, as seen in revision THA. In TDR, both macrophages and giant cells were present with a higher number of macrophages. Conclusions Both small and large PE particles are present after TDR revision surgery compatible with both THA and TKA wear patterns. The similarities between periprosthetic tissue reactions in the different groups may give more insight in the clinical relevance of PE particles and inflammatory cells in the lumbar spine. The current findings may help to improve TDR design as applied from technologies previously developed in THA and TKA with the goal of a longer survival of TDR. PMID:21336235

Effect of bevacizumab combined with boron neutron capture therapy on local tumor response and lung metastasis

PubMed Central

MASUNAGA, SHIN-ICHIRO; SAKURAI, YOSHINORI; TANO, KEIZO; TANAKA, HIROKI; SUZUKI, MINORU; KONDO, NATSUKO; NARABAYASHI, MASARU; WATANABE, TSUBASA; NAKAGAWA, YOSUKE; MARUHASHI, AKIRA; ONO, KOJI

2014-01-01

The aim of the present study was to evaluate the effect of bevacizumab on local tumor response and lung metastatic potential during boron neutron capture therapy (BNCT) and in particular, the response of intratumor quiescent (Q) cells. B16-BL6 melanoma tumor-bearing C57BL/6 mice were continuously administered bromodeoxyuridine (BrdU) to label all proliferating (P) tumor cells. The tumors were irradiated with thermal neutron beams following the administration of a 10B-carrier [L-para-boronophenylalanine-10B (BPA) or sodium mercaptoundecahydrododecaborate-10B (BSH)], with or without the administration of bevacizumab. This was further combined with an acute hypoxia-releasing agent (nicotinamide) or mild temperature hyperthermia (MTH, 40°C for 60 min). Immediately following the irradiation, cells from certain tumors were isolated and incubated with a cytokinesis blocker. The responses of the Q cells and the total (P+Q) cell populations were assessed based on the frequency of micronuclei using immunofluorescence staining for BrdU. In other tumor-bearing mice, 17 days following irradiation, lung metastases were enumerated. Three days following bevacizumab administration, the sensitivity of the total tumor cell population following BPA-BNCT had increased more than that following BSH-BNCT. The combination with MTH, but not with nicotinamide, further enhanced total tumor cell population sensitivity. Regardless of the presence of a 10B-carrier, MTH enhanced the sensitivity of the Q cell population. Regardless of irradiation, the administration of bevacizumab, as well as nicotinamide treatment, demonstrated certain potential in reducing the number of lung metastases especially in BPA-BNCT compared with BSH-BNCT. Thus, the current study revealed that BNCT combined with bevacizumab has the potential to sensitize total tumor cells and cause a reduction in the number of lung metastases to a similar level as nicotinamide. PMID:24944637

Antibody production studied by means of the LHG assay*

PubMed Central

Wortis, H. H.; Taylor, R. B.; Dresser, D. W.

1966-01-01

By means of a modified plaque assay the numbers of cells producing 19S antibody (direct PFC) and 7S antibody (developed PFC) have been studied separately. Significant and somewhat surprising differences in the responses of the two kinds of antibody-producing cell have been noted. It is clear that the route of injection of the antigen is of importance in determining the response in the spleen. The response of both types of PFC was found to be biphasic. The total increase in spleen cell number after immunization was not readily accounted for by the increase in specific PFC. PMID:5954775

Effects of Aging on Hippocampal Neurogenesis After Irradiation

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

Cheng, Zoey; Institute of Medical Science, University of Toronto, Toronto, Ontario; Li, Yu-Qing

Purpose: To assess the influence of aging on hippocampal neuronal development after irradiation (IR). Methods and Materials: Male mice, 2, 4, 6, 12, and 18 months of age, were given a single dose of 0 or 5 Gy of IR. A bromodeoxyuridine (BrdU) incorporation study was used to label newborn cells. Neural progenitors, newborn neurons, and microglia in dentate gyrus (DG) were identified by phenotypic markers, and their numbers were quantified by nonbiased stereology 9 weeks after IR. Results: BrdU-positive or newborn cells in DG decreased with aging and after IR. The number of neuroblasts and newborn neurons decreased with aging, and amore » further significant reduction was observed after IR. Total type 1 cells (the putative neural stem cells), and newborn type 1 cells decreased with aging, and further reduction in total type 1 cells was observed after IR. Aging-associated activation of microglia in hippocampus was enhanced after IR. Conclusions: The aging-associated decline in hippocampal neurogenesis was further inhibited after IR. Ablation of neural progenitors and activation of microglia may contribute to the inhibition of neuronal development after IR across all ages.« less

Chronic binge alcohol administration increases intestinal T cell proliferation and turnover in rhesus macaques

PubMed Central

Veazey, Ronald S.; Amedee, Angela; Wang, Xiaolei; Kaack, M. Bernice; Porretta, Constance; Dufour, Jason; Welsh, David; Happel, Kyle; Pahar, Bapi; Molina, Patricia E.; Nelson, Steve; Bagby, Gregory J.

2015-01-01

Background Alcohol use results in changes in intestinal epithelial cell turnover and microbial translocation, yet less in known about the consequences on intestinal lymphocytes in the gut. Here we compared T cell subsets in the intestine of macaques before and after 3 months of chronic alcohol administration to examine the effects of alcohol on intestinal T cell subsets. Methods Rhesus macaques received either alcohol or isocaloric sucrose as a control treatment daily over a 3 month period via indwelling gastric catheters. Intestinal lymphocytes subsets were identified in biopsy samples by flow cytometry. Twenty-four hours prior to sampling, animals were inoculated with BrdU to assess lymphocyte proliferation. Immunohistochemistry was performed on tissue samples to quantitate CD3+ cells. Results Animals receiving alcohol had increased rates of intestinal T cell turnover of both CD4+ and CD8+ T cells as reflected by increased BrdU incorporation. However, absolute numbers of T cells were decreased in intestinal tissues as evidenced by immunohistochemistry for total CD3 expression per mm2 intestinal lamina propria in tissue sections. Combining immunohistochemistry and flow cytometry data showed that the absolute numbers of CD8+ T cells were significantly decreased, whereas total of CD4+ T cells were minimally decreased. Conclusions Collectively, these data indicate alcohol exposure to the small intestine results in marked loss of CD3+ T cells, accompanied by marked increases in CD4+ and CD8+ T cell proliferation and turnover, which we speculate is an attempt to maintain stable numbers of T cells in tissues. This suggests alcohol results in accelerated T cell turnover in the gut, which may contribute to premature T cell senescence. Further these data indicate that chronic alcohol administration results in increased levels of HIV target cells (proliferating CD4+ T cells) that may support higher levels of HIV replication in intestinal tissues. PMID:26146859

Total synthesis of panicein A2

PubMed Central

Yeung, Lili; Pilkington, Lisa I; Cadelis, Melissa M; Copp, Brent R

2015-01-01

Summary The first total synthesis of the unusual aromatic sesquiterpene panicein A2 is reported and the structure of the natural product has been confirmed. When tested by the NCI against a range of human cancer cell lines, it was found that panicein A2 exhibits very little antiproliferative activity at 10 μM – an observation that is at odds with the earlier report that stated panicein A2 exhibits in vitro cytotoxicity against a number of tumour cell lines. PMID:26664619

Prognostic Factors for Survival in Patients Treated With Stereotactic Radiosurgery for Recurrent Brain Metastases After Prior Whole Brain Radiotherapy

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

Caballero, Jorge A.; Sneed, Penny K., E-mail: psneed@radonc.ucsf.edu; Lamborn, Kathleen R.

2012-05-01

Purpose: To evaluate prognostic factors for survival after stereotactic radiosurgery (SRS) for new, progressive, or recurrent brain metastases (BM) after prior whole brain radiotherapy (WBRT). Methods and Materials: Patients treated between 1991 and 2007 with Gamma Knife SRS for BM after prior WBRT were retrospectively reviewed. Potential prognostic factors were analyzed overall and by primary site using univariate and stepwise multivariate analyses and recursive partitioning analysis, including age, Karnofsky performance status (KPS), primary tumor control, extracranial metastases, number of BM treated, total SRS target volume, and interval from WBRT to SRS. Results: A total of 310 patients were analyzed, includingmore » 90 breast, 113 non-small-cell lung, 31 small-cell lung, 42 melanoma, and 34 miscellaneous patients. The median age was 56, KPS 80, number of BM treated 3, and interval from WBRT to SRS 8.1 months; 76% had controlled primary tumor and 60% had extracranial metastases. The median survival was 8.4 months overall and 12.0 vs. 7.9 months for single vs. multiple BM treated (p = 0.001). There was no relationship between number of BM and survival after excluding single-BM patients. On multivariate analysis, favorable prognostic factors included age <50, smaller total target volume, and longer interval from WBRT to SRS in breast cancer patients; smaller number of BM, KPS >60, and controlled primary in non-small-cell lung cancer patients; and smaller total target volume in melanoma patients. Conclusions: Among patients treated with salvage SRS for BM after prior WBRT, prognostic factors appeared to vary by primary site. Although survival time was significantly longer for patients with a single BM, the median survival time of 7.9 months for patients with multiple BM seems sufficiently long for salvage SRS to appear to be worthwhile, and no evidence was found to support the use of a cutoff for number of BM appropriate for salvage SRS.« less

Basal cell carcinoma of the nipple. Report of two cases.

PubMed

Cain, R J; Sau, P; Benson, P M

1990-02-01

Two cases of basal cell carcinoma of the nipple are presented, bringing the total number of reported cases to 15. The majority, including our two patients, are elderly men. This finding suggests a causal role of exposure to ultraviolet radiation. In our cases excision was curative.

Comparison of Mast Cells and Inflammatory Cells within Periapical Lesions and Comparison of Degranulated Mast Cells Between Fibrous and Inflamed Area in Radicular Cysts: An Immunohistochemical Study

PubMed Central

Sood, Rahul; Akifuddin, Syed; Sidhu, Gagandeep Kaur; Khan, Nadia; Singla, Kapil

2014-01-01

Objective: The role of mast cells as the key effector of allergic inflammation, anaphylactic inflammatory reactions and in the pathogenesis of chronic inflammation, is well-known. The present study is adopted to compare mast cells and inflammatory cells within periapical granuloma and cysts and localize the mast cells and quantify their number in the periapical cysts so as to propose a role of mast cells in the pathogenesis of this lesion. Materials and Methods: Biopsy specimens of 30 periapical lesions were stained with hematoxylin–eosin, and immunohistochemical Mast Cell Tryptase from Bio SB (IHC detection system kit) antibody. The tryptase positive mast cells and mononuclear inflammatory cells were counted in 10 consecutive high power fields (100X) using the binocular microscope from Motic attached to a computer with Motic Advanced Images 3.2 software. Results: Comparative microscopic analysis indicated that periapical cyst shows more percentage of mast cells and less percentage of inflammatory cell than periapical granuloma (comparison of mean and standard deviation of total number of mast cells and inflammatory cells, mast cells 3.15±1.39 in the granuloma group and 4.43±1.91in the cyst group, inflammatory cells, 67.11±1.2 in the granuloma group and 52.66±0.8 in the cyst group). Numerous degranulated mast cells were observed in the fibrous wall than the inflammatory infiltrate of the periapical cysts. The mean and standard deviation of degranulated mast cells between the inflammatory and fibrous zone within the cyst group, being 0.95±1.10 and1.68±1.34 respectively. The values varied significantly between the two zones. Conclusion: The number of inflammatory cells in the cyst group is less than periapical granuloma and total number of mast cells in the cyst group is more as compared to periapical granuloma. The degranulated cells were quantified and they were higher in the fibrous area of the cysts than the inflammatory zone. This study could support the fact that the various mediators released on degranulation play a role in the connective tissue remodeling, chronicity and expansion of the periapical lesion. PMID:25654034

The Epstein-Barr virus DNA load in the peripheral blood of transplant recipients does not accurately reflect the burden of infected cells.

PubMed

Fink, Susanne; Tsai, Ming-Han; Schnitzler, Paul; Zeier, Martin; Dreger, Peter; Wuchter, Patrick; Bulut, Olcay C; Behrends, Uta; Delecluse, Henri-Jacques

2017-01-01

Transplant recipients frequently exhibit an increased Epstein-Barr virus (EBV) load in the peripheral blood. Here, we quantitated the EBV-infected cells in the peripheral blood of these patients and defined the mode of viral infection, latent or lytic. These data indicated that there is no strong correlation between the number of infected cells and the EBV load (EBVL). This can be explained by a highly variable number of EBV copies per infected cell and by lytic replication in some cells. The plasma of these patients did not contain any free infectious viruses, but contained nevertheless EBV DNA, sometimes in large amounts, that probably originates from cell debris and contributed to the total EBVL. Some of the investigated samples carried a highly variable number of infected cells in active latency, characterized by an expression of the Epstein-Barr nuclear antigens (EBNA2) protein. However, a third of the samples expressed neither EBNA2 nor lytic proteins. Patients with an increased EBVL represent a heterogeneous group of patients whose infection cannot be characterized by this method alone. Precise characterization of the origin of an increased EBVL, in particular, in terms of the number of EBV-infected cells, requires additional investigations including the number of EBV-encoded small RNA-positive cells. © 2016 Steunstichting ESOT.

Towards a Minimal Architecture for a Printable, Modular, and Robust Sensing Skin

DTIC Science & Technology

2014-04-27

hard copies of all or part of this work for personal or classroom use is granted without fee provided that copies are not made or distributed for...the total logic complexity and reduce sensor throughput. The final selection can be made to balance these effects given a specific application. Sensor...Company (TSMC)’s 65-nm GPLUSTC CMOS standard cells. Table II shows the number of gates (standard cells) and flip -flops generated for the given number of

Effects of estradiol on incorporation of new cells in the developing zebra finch song system: potential relationship to expression of ribosomal proteins L17 and L37.

PubMed

Tang, Yu Ping; Wade, Juli

2009-06-01

Mechanisms regulating masculinization of the zebra finch song system are unclear; both estradiol and sex-specific genes may be important. This study was designed to investigate relationships between estrogen and ribosomal proteins (RPL17 and RPL37; sex-linked genes) that exhibit greater expression in song control nuclei in juvenile males than females. Four studies on zebra finches were conducted using bromodeoxyuridine (BrdU) injections on posthatching days 6-10 with immunohistochemistry for the ribosomal proteins and the neuronal marker HuC/D at day 25. Volumes of brain regions were also assessed in Nissl-stained tissue. Most BrdU+ cells expressed RPL17 and RPL37. The density and percentage of cells co-expressing BrdU and HuC/D was greatest in Area X. The density of BrdU+ cells in Area X (or its equivalent) and the percentage of these cells that were neurons were greater in males than females. In RA and HVC, total BrdU+ cells were increased in males. A variety of effects of estradiol were also detected, including inducing an Area X in females with a masculine total number of BrdU+ cells, and increasing the volume and percentage of new neurons in the HVC of females. The same manipulation in males decreased the density of BrdU+ cells in Area X, total number of BrdU+ cells in RA, and density of new neurons in HVC and RA. These data are consistent with the idea that RPL17, RPL37, and estradiol might all influence sexual differentiation, perhaps with the hormone and proteins interacting, such that an appropriate balance is required for normal development.

Effects of Estradiol on Incorporation of New Cells in the Developing Zebra Finch Song System: Potential Relationship to Expression of Ribosomal Proteins L17 and L37

PubMed Central

Tang, Yu Ping; Wade, Juli

2009-01-01

Mechanisms regulating masculinization of the zebra finch song system are unclear; both estradiol and sex-specific genes may be important. This study was designed to investigate relationships between estrogen and ribosomal proteins (RPL17 and RPL37; sex-linked genes) that exhibit greater expression in song control nuclei in juvenile males than females. Four studies on zebra finches were conducted using bromodeoxyuridine (BrdU) injections on posthatching days 6-10 with immunohistochemistry for the ribosomal proteins and the neuronal marker HuC/D at day 25. Volumes of brain regions were also assessed in Nissl-stained tissue. Most BrdU+ cells expressed RPL17 and RPL37. The density and percentage of cells co-expressing BrdU and HuC/D was greatest in Area X. The density of BrdU+ cells in Area X (or its equivalent) and the percentage of these cells that were neurons were greater in males than females. In RA and HVC, total BrdU+ cells were increased in males. A variety of effects of estradiol were also detected, including inducing an Area X in females with a masculine total number of BrdU+ cells, and increasing the volume and percentage of new neurons in the HVC of females. The same manipulation in males decreased the density of BrdU+ cells in Area X, total number of BrdU+ cells in RA, and density of new neurons in HVC and RA. These data are consistent with the ideas that RPL17, RPL37, and estradiol might all influence sexual differentiation, perhaps with the hormone and proteins interacting, such that an appropriate balance is required for normal development. PMID:19373862

In vitro and ex vivo angiogenic effects of roxarsone on rat endothelial cells.

PubMed

Zhu, Jiaqiao; Cui, Weibo; Liu, Xue; Ying, Jun; Hu, Chengyun; Zhang, Yumei

2013-11-25

Roxarsone, a feed additive, is being used worldwide to promote animal growth. However, the potential effect of roxarsone on angiogenesis has not been extensively characterized. We examined the ability of roxarsone to promote angiogenesis of rat endothelial cells in vitro and from rat aorta rings ex vivo. Endothelial cells from rats were exposed to 0.01-10.00μM roxarsone, 5ng/mL vascular endothelial growth factor (VEGF) as a positive control or phosphate buffer saline (PBS) as a negative control. Cell proliferation was measured by MTT assay, and the content of VEGF in supernatants was measured by enzyme-linked immunosorbent assay and Western blotting. A Matrigel-induced tube formation assay was used to evaluate the effects of roxarsone on endothelial cells. Additionally, the total number and length of microvessels sprouted from rat aortic rings were measured for ex vivo investigation of angiogenesis. Results showed that the cell viability and total number and length of capillary-like tube formations after roxarsone treatment was significantly higher than that of negative (P<0.05), with a maximum effect at 1.00μM exposure. Furthermore, the number of microvessels sprouted from aortic rings treated for 4h with 0.1-10.0μM roxarsone was significantly higher than that of PBS treatment, with a peak value of 1.0μM. These results further demonstrate the potential of roxarsone to promote angiogenesis in vitro and ex vivo. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Maximal exercise increases mucosal associated invariant T cell frequency and number in healthy young men.

PubMed

Hanson, Erik D; Danson, Eli; Nguyen-Robertson, Catriona V; Fyfe, Jackson J; Stepto, Nigel K; Bartlett, David B; Sakkal, Samy

2017-11-01

Mucosal associated invariant T (MAIT) cells have properties of the innate and acquired immune systems. While the response to vigorous exercise has been established for most leukocytes, MAIT cells have not been investigated. Therefore, the purpose was to determine if MAIT cell lymphocytosis occurs with acute maximal aerobic exercise and if this response is influenced by exercise duration, cardiovascular fitness, or body composition. Twenty healthy young males with moderate fitness levels performed an extended graded exercise test until volitional fatigue. Peripheral blood mononuclear cells were isolated from venous blood obtained prior and immediately after exercise and were labeled to identify specific T cell populations using flow cytometry. The percentage of MAIT cells relative to total T cells significantly increased from 3.0 to 3.8% and absolute MAIT cell counts increased by 2.2-fold following maximal exercise. MAIT cell subpopulation proportions were unchanged with exercise. Within cytotoxic T lymphocytes (CTL), MAIT cells consisted of 8% of these cells and this remained constant after exercise. MAIT cell counts and changes with exercise were not affected by body composition, VO 2peak , or exercise duration. Maximal exercise doubled MAIT cell numbers and showed preferential mobilization within total T cells but the response was not influenced by fitness levels, exercise duration, or body composition. These results suggest that acute exercise could be used to offset MAIT cell deficiencies observed with certain pathologies. MAIT cells also make up a substantial proportion of CTLs, which may have implications for cytotoxicity assays using these cells.

The impact of temperature on microbial diversity and AOA activity in the Tengchong Geothermal Field, China

NASA Astrophysics Data System (ADS)

Li, Haizhou; Yang, Qunhui; Li, Jian; Gao, Hang; Li, Ping; Zhou, Huaiyang

2015-11-01

Using a culture-independent method that combines CARD-FISH, qPCR and 16S rDNA, we investigated the abundance, community structure and diversity of microbes along a steep thermal gradient (50-90 °C) in the Tengchong Geothermal Field. We found that Bacteria and Archaea abundance changed markedly with temperature changes and that the number of cells was lowest at high temperatures (90.8 °C). Under low-temperature conditions (52.3-74.6 °C), the microbial communities were dominated by Bacteria, which accounted for 60-80% of the total number of cells. At 74.6 °C, Archaea were dominant, and at 90.8 °C, they accounted for more than 90% of the total number of cells. Additionally, the microbial communities at high temperatures (74.6-90.8 °C) were substantially simpler than those at the low-temperature sites. Only a few genera (e.g., bacterial Caldisericum, Thermotoga and Thermoanaerobacter, archaeal Vulcanisaeta and Hyperthermus) often dominated in high-temperature environments. Additionally, a positive correlation between Ammonia-Oxidizing Archaea (AOA) activity and temperature was detected. AOA activity increased from 17 to 52 pmol of NO2- per cell d-1 with a temperature change from 50 to 70 °C.

The impact of temperature on microbial diversity and AOA activity in the Tengchong Geothermal Field, China.

PubMed

Li, Haizhou; Yang, Qunhui; Li, Jian; Gao, Hang; Li, Ping; Zhou, Huaiyang

2015-11-26

Using a culture-independent method that combines CARD-FISH, qPCR and 16S rDNA, we investigated the abundance, community structure and diversity of microbes along a steep thermal gradient (50-90 °C) in the Tengchong Geothermal Field. We found that Bacteria and Archaea abundance changed markedly with temperature changes and that the number of cells was lowest at high temperatures (90.8 °C). Under low-temperature conditions (52.3-74.6 °C), the microbial communities were dominated by Bacteria, which accounted for 60-80% of the total number of cells. At 74.6 °C, Archaea were dominant, and at 90.8 °C, they accounted for more than 90% of the total number of cells. Additionally, the microbial communities at high temperatures (74.6-90.8 °C) were substantially simpler than those at the low-temperature sites. Only a few genera (e.g., bacterial Caldisericum, Thermotoga and Thermoanaerobacter, archaeal Vulcanisaeta and Hyperthermus) often dominated in high-temperature environments. Additionally, a positive correlation between Ammonia-Oxidizing Archaea (AOA) activity and temperature was detected. AOA activity increased from 17 to 52 pmol of NO2(-) per cell d(-1) with a temperature change from 50 to 70 °C.

Effect of chronic antipsychotic exposure on astrocyte and oligodendrocyte numbers in macaque monkeys

PubMed Central

Konopaske, Glenn T.; Dorph-Petersen, Karl-Anton; Sweet, Robert A.; Pierri, Joseph N.; Zhang, Wei; Sampson, Allan R.; Lewis, David A.

2008-01-01

Background Both in vivo and post-mortem studies suggest that oligodendrocyte and myelination alterations are present in individuals with schizophrenia. However, it is unclear whether prolonged treatment with antipsychotic medications contributes to these disturbances. We recently reported that chronic exposure of macaque monkeys to haloperidol or olanzapine was associated with a 10−18% lower glial cell number in the parietal grey matter. Consequently, in this study we sought to determine whether the lower glial cell number was due to fewer oligodendrocytes as opposed to lower numbers of astrocytes. Methods Using fluorescent immunocytochemical techniques, we optimized the visualization of each cell type throughout the entire thickness of tissue sections, while minimizing final tissue shrinkage. As a result, we were able to obtain robust stereological estimates of total oligodendrocyte and astrocyte numbers in the parietal grey matter using the optical fractionator method. Results We found a significant 20.5% lower astrocyte number with a non-significant 12.9% lower oligodendrocyte number in the antipsychotic-exposed monkeys. Similar effects were seen in both the haloperidol and olanzapine groups. Conclusion These findings suggest that studies investigating glial cell alterations in schizophrenia must take into account the effect of antipsychotic treatment. PMID:17945195

Chemical Toxicity on HeLa Cells.

PubMed

Verma, Rajeshwar P; Hansch, Corwin

2006-01-01

HeLa cells were named for Henrietta Lacks, who died in 1952 from an infection of a special type of cancer. Margaret Gey, her physician, started working with these cancer cells that are still used for medical research. In the present review, an attempt has been made to collect the data for the effects of different chemicals on HeLa cells and to discuss them by the formulation of a total number of 22 QSAR.

Comparison of cell counting methods in rodent pulmonary toxicity studies: automated and manual protocols and considerations for experimental design

PubMed Central

Zeidler-Erdely, Patti C.; Antonini, James M.; Meighan, Terence G.; Young, Shih-Houng; Eye, Tracy J.; Hammer, Mary Ann; Erdely, Aaron

2016-01-01

Pulmonary toxicity studies often use bronchoalveolar lavage (BAL) to investigate potential adverse lung responses to a particulate exposure. The BAL cellular fraction is counted, using automated (i.e. Coulter Counter®), flow cytometry or manual (i.e. hemocytometer) methods, to determine inflammatory cell influx. The goal of the study was to compare the different counting methods to determine which is optimal for examining BAL cell influx after exposure by inhalation or intratracheal instillation (ITI) to different particles with varying inherent pulmonary toxicities in both rat and mouse models. General findings indicate that total BAL cell counts using the automated and manual methods tended to agree after inhalation or ITI exposure to particle samples that are relatively nontoxic or at later time points after exposure to a pneumotoxic particle when the response resolves. However, when the initial lung inflammation and cytotoxicity was high after exposure to a pneumotoxic particle, significant differences were observed when comparing cell counts from the automated, flow cytometry and manual methods. When using total BAL cell count for differential calculations from the automated method, depending on the cell diameter size range cutoff, the data suggest that the number of lung polymorphonuclear leukocytes (PMN) varies. Importantly, the automated counts, regardless of the size cutoff, still indicated a greater number of total lung PMN when compared with the manual method, which agreed more closely with flow cytometry. The results suggest that either the manual method or flow cytometry would be better suited for BAL studies where cytotoxicity is an unknown variable. PMID:27251196

Comparison of cell counting methods in rodent pulmonary toxicity studies: automated and manual protocols and considerations for experimental design.

PubMed

Zeidler-Erdely, Patti C; Antonini, James M; Meighan, Terence G; Young, Shih-Houng; Eye, Tracy J; Hammer, Mary Ann; Erdely, Aaron

2016-08-01

Pulmonary toxicity studies often use bronchoalveolar lavage (BAL) to investigate potential adverse lung responses to a particulate exposure. The BAL cellular fraction is counted, using automated (i.e. Coulter Counter®), flow cytometry or manual (i.e. hemocytometer) methods, to determine inflammatory cell influx. The goal of the study was to compare the different counting methods to determine which is optimal for examining BAL cell influx after exposure by inhalation or intratracheal instillation (ITI) to different particles with varying inherent pulmonary toxicities in both rat and mouse models. General findings indicate that total BAL cell counts using the automated and manual methods tended to agree after inhalation or ITI exposure to particle samples that are relatively nontoxic or at later time points after exposure to a pneumotoxic particle when the response resolves. However, when the initial lung inflammation and cytotoxicity was high after exposure to a pneumotoxic particle, significant differences were observed when comparing cell counts from the automated, flow cytometry and manual methods. When using total BAL cell count for differential calculations from the automated method, depending on the cell diameter size range cutoff, the data suggest that the number of lung polymorphonuclear leukocytes (PMN) varies. Importantly, the automated counts, regardless of the size cutoff, still indicated a greater number of total lung PMN when compared with the manual method, which agreed more closely with flow cytometry. The results suggest that either the manual method or flow cytometry would be better suited for BAL studies where cytotoxicity is an unknown variable.

Evolution of TUNEL-labeling in the rat lens after in vivo exposure to just above threshold dose UVB.

PubMed

Kronschläger, Martin; Yu, Zhaohua; Talebizadeh, Nooshin; Meyer, Linda M; Hallböök, Finn; Söderberg, Per G

2013-08-01

To quantitatively analyse the evolution of TUNEL-labeling, after in vivo exposure to UVB. Altogether, 16 Sprague Dawley rats were unilaterally exposed in vivo for 15 min to close to threshold dose, 5 kJ/m(2), of ultraviolet radiation in the 300 nm wavelength region. Animals were sacrificed in groups of 4 at 1, 5, 24 and 120 h after exposure. For each animal, both eye globes were removed and frozen. The frozen eye was cryo-sectioned in 10 µm thick midsagittal sections. From each globe, three midsagittal sections with at least five sections interval in between were mounted on a microscope slide. Sections were TUNEL-labeled and counter stained with DAPI. For quantification of apoptosis, a fluorescence microscope was used. In sections with a continuous epithelial cell surface, the number of lens epithelial cell nuclei and the number of TUNEL-positive epithelial cell nuclei was counted. The total number of TUNEL-positive epithelial cell nuclei for all three sections of one lens in relation to the total number of epithelial cell nuclei for all three sections of the same lens was compared between exposed and contralateral not exposed lens for each animal. The relative difference of the fraction of TUNEL-positive nuclei between exposed and contralateral not exposed lens increased gradually, peaked in the time interval 5-120 h after exposure, and then declined. Close to threshold dose of UVB induces TUNEL-labeling that peaks in the time window 5-120 h after exposure to UVB.

Single-Cell RNA-Sequencing Reveals a Continuous Spectrum of Differentiation in Hematopoietic Cells.

PubMed

Macaulay, Iain C; Svensson, Valentine; Labalette, Charlotte; Ferreira, Lauren; Hamey, Fiona; Voet, Thierry; Teichmann, Sarah A; Cvejic, Ana

2016-02-02

The transcriptional programs that govern hematopoiesis have been investigated primarily by population-level analysis of hematopoietic stem and progenitor cells, which cannot reveal the continuous nature of the differentiation process. Here we applied single-cell RNA-sequencing to a population of hematopoietic cells in zebrafish as they undergo thrombocyte lineage commitment. By reconstructing their developmental chronology computationally, we were able to place each cell along a continuum from stem cell to mature cell, refining the traditional lineage tree. The progression of cells along this continuum is characterized by a highly coordinated transcriptional program, displaying simultaneous suppression of genes involved in cell proliferation and ribosomal biogenesis as the expression of lineage specific genes increases. Within this program, there is substantial heterogeneity in the expression of the key lineage regulators. Overall, the total number of genes expressed, as well as the total mRNA content of the cell, decreases as the cells undergo lineage commitment. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Progenitor cells are mobilized by acute psychological stress but not beta-adrenergic receptor agonist infusion

PubMed Central

Riddell, Natalie E.; Burns, Victoria E.; Wallace, Graham R.; Edwards, Kate M.; Drayson, Mark; Redwine, Laura S.; Hong, Suzi; Bui, Jack D.; Fischer, Johannes C.; Mills, Paul J.; Bosch, Jos A.

2015-01-01

Objectives Stimuli that activate the sympathetic nervous system, such as acute psychological stress, rapidly invoke a robust mobilization of lymphocytes into the circulation. Experimental animal studies suggest that bone marrow-derived progenitor cells (PCs) also mobilize in response to sympathetic stimulation. Here we tested the effects of acute psychological stress and brief pharmacological β-adrenergic (βAR) stimulation on peripheral PC numbers in humans. Methods In two studies, we investigated PC mobilization in response to an acute speech task (n=26) and βAR-agonist (isoproterenol) infusion (n=20). A subset of 8 participants also underwent the infusion protocol with concomitant administration of the βAR-antagonist propranolol. Flow cytometry was used to enumerate lymphocyte subsets, total progenitor cells, total haematopoietic stem cells (HSC), early HSC (multi-lineage potential), late HSC (lineage committed), and endothelial PCs (EPCs). Results Both psychological stress and βAR-agonist infusion caused the expected mobilization of total monocytes and lymphocytes and CD8+ T lymphocytes. Psychological stress also induced a modest, but significant, increase in total PCs, HSCs, and EPC numbers in peripheral blood. However, infusion of a βAR-agonist did not result in a significant change in circulating PCs. Conclusion PCs are rapidly mobilized by psychological stress via mechanisms independent of βAR-stimulation, although the findings do not exclude βAR-stimulation as a possible cofactor. Considering the clinical and physiological relevance, further research into the mechanisms involved in stress-induced PC mobilization seems warranted. PMID:25747743

Progenitor cells are mobilized by acute psychological stress but not beta-adrenergic receptor agonist infusion.

PubMed

Riddell, Natalie E; Burns, Victoria E; Wallace, Graham R; Edwards, Kate M; Drayson, Mark; Redwine, Laura S; Hong, Suzi; Bui, Jack C; Fischer, Johannes C; Mills, Paul J; Bosch, Jos A

2015-10-01

Stimuli that activate the sympathetic nervous system, such as acute psychological stress, rapidly invoke a robust mobilization of lymphocytes into the circulation. Experimental animal studies suggest that bone marrow-derived progenitor cells (PCs) also mobilize in response to sympathetic stimulation. Here we tested the effects of acute psychological stress and brief pharmacological β-adrenergic (βAR) stimulation on peripheral PC numbers in humans. In two studies, we investigated PC mobilization in response to an acute speech task (n=26) and βAR-agonist (isoproterenol) infusion (n=20). A subset of 8 participants also underwent the infusion protocol with concomitant administration of the βAR-antagonist propranolol. Flow cytometry was used to enumerate lymphocyte subsets, total progenitor cells, total haematopoietic stem cells (HSC), early HSC (multi-lineage potential), late HSC (lineage committed), and endothelial PCs (EPCs). Both psychological stress and βAR-agonist infusion caused the expected mobilization of total monocytes and lymphocytes and CD8(+) T lymphocytes. Psychological stress also induced a modest, but significant, increase in total PCs, HSCs, and EPC numbers in peripheral blood. However, infusion of a βAR-agonist did not result in a significant change in circulating PCs. PCs are rapidly mobilized by psychological stress via mechanisms independent of βAR-stimulation, although the findings do not exclude βAR-stimulation as a possible cofactor. Considering the clinical and physiological relevance, further research into the mechanisms involved in stress-induced PC mobilization seems warranted. Copyright © 2015 Elsevier Inc. All rights reserved.

Communities of ammonia-oxidizing bacteria in activated sludge of various sewage treatment plants in Tokyo.

PubMed

Limpiyakorn, Tawan; Shinohara, Yuko; Kurisu, Futoshi; Yagi, Osami

2005-10-01

We investigated ammonia-oxidizing bacteria in activated sludge collected from 12 sewage treatment systems, whose ammonia removal and treatment processes differed, during three different seasons. We used real-time PCR quantification to reveal total bacterial numbers and total ammonia oxidizer numbers, and used specific PCR followed by denaturing gel gradient electrophoresis, cloning, and sequencing of 16S rRNA genes to analyze ammonia-oxidizing bacterial communities. Total bacterial numbers and total ammonia oxidizer numbers were in the range of 1.6 x 10(12) - 2.4 x 10(13) and 1.0 x 10(9) - 9.2 x 10(10)cellsl(-1), respectively. Seasonal variation was observed in the total ammonia oxidizer numbers, but not in the ammonia-oxidizing bacterial communities. Members of the Nitrosomonas oligotropha cluster were found in all samples, and most sequences within this cluster grouped within two of the four sequence types identified. Members of the clusters of Nitrosomonas europaea-Nitrosococcus mobilis, Nitrosomonas cryotolerans, and unknown Nitrosomonas, occurred solely in one anaerobic/anoxic/aerobic (A2O) system. Members of the Nitrosomonas communis cluster occurred almost exclusively in association with A2O and anaerobic/aerobic systems. Solid residence time mainly influenced the total numbers of ammonia-oxidizing bacteria, whereas dissolved oxygen concentration primarily affected the ammonia-oxidizing activity per ammonia oxidizer cell.

Involvement of the major histocompatibility complex region in the genetic regulation of circulating CD8 T-cell numbers in humans.

PubMed

Cruz, E; Vieira, J; Gonçalves, R; Alves, H; Almeida, S; Rodrigues, P; Lacerda, R; Porto, G

2004-07-01

Variability in T-lymphocyte numbers is partially explained by a genetic regulation. From studies in animal models, it is known that the Major Histocompatibility Complex (MHC) is involved in this regulation. In humans, this has not been shown yet. The objective of the present study was to test the hypothesis that genes in the MHC region influence the regulation of T-lymphocyte numbers. Two approaches were used. Association studies between T-cell counts (CD4(+) and CD8(+)) or total lymphocyte counts and HLA class I alleles (A and B) or mutations in the HFE (C282Y and H63D), the hemochromatosis gene, in an unrelated population (n = 264). A second approach was a sibpair correlation analysis of the same T-cell counts in relation to HLA-HFE haplotypes in subjects belonging to 48 hemochromatosis families (n = 456 sibpairs). In the normal population, results showed a strong statistically significant association of the HLA-A*01 with high numbers of CD8(+) T cells and a less powerful association with the HLA-A*24 with low numbers of CD8(+) T cells. Sibpair correlations revealed the most significant correlation for CD8(+) T-cell numbers for sibpairs with HLA-HFE-identical haplotypes. This was not observed for CD4(+) T cells. These results show that the MHC region is involved in the genetic regulation of CD8(+) T-cell numbers in humans. Identification of genes responsible for this control may have important biological and clinical implications.

Increased Skin Inflammation and Blood Vessel Density in Human and Experimental Diabetes

PubMed Central

Tellechea, Ana; Kafanas, Antonios; Leal, Ermelindo C; Tecilazich, Francesco; Kuchibhotla, Sarada; Auster, Michael E; Kontoes, Iraklis; Paolino, Jacqueline; Carvalho, Eugenia; Nabzdyk, Leena Pradhan; Veves, Aristidis

2013-01-01

Systemic inflammation is associated with impaired wound healing in diabetic patients. Using immunohistochemistry techniques, the authors investigated changes in skin inflammation and skin blood vessels in human and experimental diabetes. Comparing to the non-DM human subjects, the total number of inflammatory cells per biopsy and the number of inflammatory cells around blood vessels, a strong indication of inflammation, were higher in DM subjects irrespective of their risk for developing diabetic foot ulcer. Inflammatory cell infiltration was robustly increased in all diabetic animal models compared to their non-diabetic controls. The number and density of blood vessels and CD31 positive proliferating endothelial cells around pre-existing skin vessels was also higher in the DM patients. However, there were no differences in the skin blood flow between the non-DM and DM subjects. The number of skin blood vessels was also increased in the DM animals; however, these differences were less obvious than the ones observed for inflammatory cells. We conclude that skin inflammation and skin blood vessel density is increased in diabetic human subjects and in rodent and rabbit models of diabetes. PMID:23446362

Effects of spaceflight on rat peripheral blood leukocytes and bone marrow progenitor cells

NASA Technical Reports Server (NTRS)

Ichiki, A. T.; Gibson, L. A.; Jago, T. L.; Strickland, K. M.; Johnson, D. L.; Lange, R. D.; Allebban, Z.

1996-01-01

The white blood cell (WBC) elements and the bone marrow myeloid progenitor cell populations were analyzed to ascertain adaptation to micro-gravity and subsequent readaptation to 1 G in rats flown on the 14-day Spacelab Life Sciences-2 (SLS-2) mission. Bone marrow cells were harvested from one group of rats killed inflight (FD13) and blood was drawn from three other groups at various times. The WBC level was normal on FD14 with the exception of neutrophilia. On FD13, numbers of colony-forming units-granulocyte (CFU-G), CFU-GM, and CFU-M from flight animals were decreased compared with ground controls when incubated with recombinant rat interleukin-3 (rrIL-3) alone or in combination with recombinant human erythropoietin (rhEpo). On recovery (R + 0), flight rats had decreased numbers of total leukocytes and absolute numbers of lymphocytes and monocytes with elevated neutrophils compared with control rats. They had lower numbers of CD4, CD8, CD2, CD3, and B cells in the peripheral blood but no differences in spleen lymphocytes.

L-carnosine enhanced reproductive potential of the Saccharomyces cerevisiae yeast growing on medium containing glucose as a source of carbon.

PubMed

Kwolek-Mirek, Magdalena; Molon, Mateusz; Kaszycki, Pawel; Zadrag-Tecza, Renata

2016-08-01

Carnosine is an endogenous dipeptide composed of β-alanine and L-histidine, which occurs in vertebrates, including humans. It has a number of favorable properties including buffering, chelating, antioxidant, anti-glycation and anti-aging activities. In our study we used the Saccharomyces cerevisiae yeast as a model organism to examine the impact of L-carnosine on the cell lifespan. We demonstrated that L-carnosine slowed down the growth and decreased the metabolic activity of cells as well as prolonged their generation time. On the other hand, it allowed for enhancement of the yeast reproductive potential and extended its reproductive lifespan. These changes may be a result of the reduced mitochondrial membrane potential and decreased ATP content in the yeast cells. However, due to reduction of the post-reproductive lifespan, L-carnosine did not have an influence on the total lifespan of yeast. In conclusion, L-carnosine does not extend the total lifespan of S. cerevisiae but rather it increases the yeast's reproductive capacity by increasing the number of daughter cells produced.

Quantitation of blood lymphocytes secreting antibodies to pneumococcal polysaccharides after in vivo antigenic stimulation.

PubMed

Heilmann, C; Pedersen, F K

1986-02-01

An indirect plaque-forming cell assay detecting B cells secreting IgM, IgG and IgA antibodies against pneumococcal polysaccharides (PPS) is described. The numbers of anti-PPS-secreting cells (SC) and Ig-SC in the blood of normal persons immunized with a polyvalent PPS vaccine were quantitated. Anti-PPS-SC were recorded from the fourth to the twelfth post-vaccination day, and the maximum number was found between days 6 and 9. Quantitatively IgA anti-PPS-SC outnumbered the IgM and IgG anti-PPS-SC. Concomitant with the increase in the numbers of antibody-SC an increase in polyclonally activated IgM-, IgG- and IgA-SC was recorded. The specific anti-PPS-antibody-SC constituted 20-80% of the total numbers of Ig-SC from the sixth to the ninth post-vaccination day.

Long live the liver: immunohistochemical and stereological study of hepatocytes, liver sinusoidal endothelial cells, Kupffer cells and hepatic stellate cells of male and female rats throughout ageing.

PubMed

Marcos, Ricardo; Correia-Gomes, Carla

2016-12-01

Male/female differences in enzyme activity and gene expression in the liver are known to be attenuated with ageing. Nevertheless, the effect of ageing on liver structure and quantitative cell morphology remains unknown. Male and female Wistar rats aged 2, 6, 12 and 18 months were examined by means of stereological techniques and immunohistochemical tagging of hepatocytes (HEP), liver sinusoidal endothelial cells (LSEC), Kupffer cells (KC) and hepatic stellate cells (HSC) in order to assess the total number and number per gram of these cells throughout life. The mean cell volume of HEP and HSC, the lobular position and the collagen content of the liver were also evaluated with stereological techniques. The number per gram of HSC was similar for both genders and was maintained throughout ageing. The mean volume of HSC was also conserved but differences in the cell body and lobular location were observed. Statistically significant gender differences in HEP were noted in young rats (females had smaller and more binucleated HEP) but were attenuated with ageing. The same occurred for KC and LSEC, since the higher number per gram in young females disappeared in older animals. Liver collagen increased with ageing but only in males. Thus, the numbers of these four cell types are related throughout ageing, with well-defined cell ratios. The shape and lobular position of HSC change with ageing in both males and females. Gender dimorphism in HEP, KC and LSEC of young rat liver disappears with ageing.

Shaping the shoot: the relative contribution of cell number and cell shape to variations in internode length between parent and hybrid apple trees.

PubMed

Ripetti, V; Escoute, J; Verdeil, J L; Costes, E

2008-01-01

Genetic control of plant size and shape is a promising perspective, particularly in fruit trees, in order to select desirable genotypes. A recent study on architectural traits in an apple progeny showed that internode length was a highly heritable character. However, few studies have been devoted to internode cellular patterning in dicotyledonous stems, and the interplay between the two elementary cell processes that contribute to their length, i.e. cell division and elongation, is not fully understood. The present study aimed at unravelling their contributions in the genetic variation of internode length in a selection of F(1) and parent genotypes of apple tree, by exploring the number of cells and cell shape within mature internodes belonging to the main axes. The results highlighted that both the variables were homogeneous in samples collected either along a sagital line or along the pith width, and suggest that cell lengthening was homogeneous during internode development. They allowed the total number of cells to be estimated on the internode scale and opened up new perspectives for simplifying tissue sampling procedures for further investigations. Differences in internode length were observed between the genotypes, in particular between the parents, and partly resulted from a compensation between cell number and cell length. However, genetic variations in internode length primarily involved the number of cells, while cell length was more secondary. These results argue for an interplay between cellular and organismal control of internode shape that may involve the rib meristem.

Effects of employing a 10B-carrier and manipulating intratumour hypoxia on local tumour response and lung metastatic potential in boron neutron capture therapy

PubMed Central

Masunaga, S; Sakurai, Y; Tanaka, H; Suzuki, M; Liu, Y; Kondo, N; Maruhashi, A; Kinashi, Y; Ono, K

2012-01-01

Objectives To evaluate the effects of employing a 10B-carrier and manipulating intratumour hypoxia on local tumour response and lung metastatic potential in boron neutron capture therapy (BNCT) by measuring the response of intratumour quiescent (Q) cells. Methods B16-BL6 melanoma tumour-bearing C57BL/6 mice were continuously given 5-bromo-2′-deoxyuridine (BrdU) to label all proliferating (P) cells. The tumours received reactor thermal neutron beam irradiation following the administration of a 10B-carrier [L-para-boronophenylalanine-10B (BPA) or sodium mercaptoundecahydrododecaborate-10B (BSH)] in combination with an acute hypoxia-releasing agent (nicotinamide) or mild temperature hyperthermia (MTH). Immediately after the irradiation, cells from some tumours were isolated and incubated with a cytokinesis blocker. The responses of the Q and total (P+Q) cell populations were assessed based on the frequency of micronuclei using immunofluorescence staining for BrdU. In other tumour-bearing mice, macroscopic lung metastases were enumerated 17 days after irradiation. Results BPA-BNCT increased the sensitivity of the total tumour cell population more than BSH-BNCT. However, the sensitivity of Q cells treated with BPA was lower than that of BSH-treated Q cells. With or without a 10B–carrier, MTH enhanced the sensitivity of the Q cell population. Without irradiation, nicotinamide treatment decreased the number of lung metastases. With irradiation, BPA-BNCT, especially in combination with nicotinamide treatment, showed the potential to reduce the number of metastases more than BSH-BNCT. Conclusion BSH-BNCT in combination with MTH improves local tumour control, while BPA-BNCT in combination with nicotinamide may reduce the number of lung metastases. PMID:22391496

Temporal Changes in Microscale Colonization of the Phylloplane by Aureobasidium pullulans

PubMed Central

McGrath, Molly J.; Andrews, John H.

2006-01-01

Colonization of apple leaves by the yeastlike fungus Aureobasidium pullulans was followed quantitatively and spatially at a microscale level throughout two growing seasons. Ten field leaves were sampled on 11 dates in 2003 and 15 dates in 2004. Using an A. pullulans-specific fluorescence in situ hybridization probe and epifluorescence microscopy, we enumerated total cells, swollen-cells and chlamydospores (SCC), and blastospores/mm2 on leaf features, including the midvein, other (smaller) veins, and the interveinal regions. By 7 July 2003 and 7 June 2004, the total numbers of A. pullulans cells/mm2 were significantly higher (P < 0.05) on the midvein and other veins than in the interveinal regions. This pattern remained consistent thereafter. The primary colonizing morphotype in all regions at all dates was the SCC form, although blastospores always occurred in low numbers. Occupancy was quantified based on the percentage of microscope fields of a particular leaf feature containing ≥1 A. pullulans cell. In general, as seasons progressed, the percent occupancy of features increased and, for most midvein and veinal features, approximated 100% at the end of both growing seasons. Except for early collections, when A. pullulans cell numbers were low, the percent occupancy of interveinal regions was lower than that of the midvein or other veinal regions. A. pullulans was distributed primarily as single cells throughout the seasons in interveinal regions. On the midvein and other veins, colonies of ≥4 cells developed over time, and more cells occurred in colonies than as singletons by August. Our results demonstrate that A. pullulans primarily colonizes veins, where populations appear to increase by growth in situ. This pattern is established early in the growing season and persists. PMID:16957250

Karyotyping of Transformed Human Epithelial Cells from Exposures of Heavy Ions

NASA Technical Reports Server (NTRS)

Yeshitla, Samrawit

2013-01-01

It is most likely that the untreated transformed single clone (clone #2) cell undergoes unequal segregation of chromosome in two daughter cell that result in 94 chromosome during mitosis, particularly in anaphase stage. Chromosome aberration observed. I. Breakage of part of chromosome 7. II. One additional number of chromosome 8 instead of the total chromosome can only be explained by early abnormal cell division. III. Complete lost of chromosome and translocation and fusion of chromosome 3 and X-chromosome. IV. Our result for translocation and fusion of chromosome 3 and X- Chromosome is conformed by mBAND pattern. There is no different between the transformed parental cell and the single cloned transformed cell. Both harbor the chromosome 5 and 16 translocation and both harbor has the trisomy chromosome 20. Transformed cells may have the number of chromosomes greater or less than 46. Doubling of chromosome numbers is a signature of tumor. Chromosomal aberration was observed on HBEC-3kt non-irradiated-soft agar (Clone #2) sample, and indication of chromosome instability in the tumor development process.

Myostatin inhibitors as therapies for muscle wasting associated with cancer and other disorders

PubMed Central

Smith, Rosamund C.; Lin, Boris K.

2013-01-01

Purpose of review This review summarizes recent progress in the development of myostatin inhibitors for the treatment of muscle wasting disorders. It also focuses on findings in myostatin biology that may have implications for the development of antimyostatin therapies. Recent findings There has been progress in evaluating antimyostatin therapies in animal models of muscle wasting disorders. Some programs have progressed into clinical development with initial results showing positive impact on muscle volume. In normal mice myostatin deficiency results in enlarged muscles with increased total force but decreased specific force (total force/total mass). An increase in myofibrillar protein synthesis without concomitant satellite cell proliferation and fusion leads to muscle hypertrophy with unchanged myonuclear number. A specific force reduction is not observed when atrophied muscle, the predominant therapeutic target of myostatin inhibitor therapy, is made myostatindeficient. Myostatin has been shown to be expressed by a number of tumor cell lines in mice and man. Summary Myostatin inhibition remains a promising therapeutic strategy for a range of muscle wasting disorders. PMID:24157714

Myostatin inhibitors as therapies for muscle wasting associated with cancer and other disorders.

PubMed

Smith, Rosamund C; Lin, Boris K

2013-12-01

This review summarizes recent progress in the development of myostatin inhibitors for the treatment of muscle wasting disorders. It also focuses on findings in myostatin biology that may have implications for the development of antimyostatin therapies. There has been progress in evaluating antimyostatin therapies in animal models of muscle wasting disorders. Some programs have progressed into clinical development with initial results showing positive impact on muscle volume.In normal mice myostatin deficiency results in enlarged muscles with increased total force but decreased specific force (total force/total mass). An increase in myofibrillar protein synthesis without concomitant satellite cell proliferation and fusion leads to muscle hypertrophy with unchanged myonuclear number. A specific force reduction is not observed when atrophied muscle, the predominant therapeutic target of myostatin inhibitor therapy, is made myostatindeficient.Myostatin has been shown to be expressed by a number of tumor cell lines in mice and man. Myostatin inhibition remains a promising therapeutic strategy for a range of muscle wasting disorders.

Human Embryonic Stem Cell-Derived Neurons Are Highly Permissive for Varicella-Zoster Virus Lytic Infection.

PubMed

Sadaoka, Tomohiko; Schwartz, Cindi L; Rajbhandari, Labchan; Venkatesan, Arun; Cohen, Jeffrey I

2018-01-01

Varicella-zoster virus (VZV) is highly cell associated when grown in culture and has a much higher (4,000- to 20,000-fold increased) particle-to-PFU ratio in vitro than herpes simplex virus (HSV). In contrast, VZV is highly infectious in vivo by airborne transmission. Neurons are major targets for VZV in vivo ; in neurons, the virus can establish latency and reactivate to produce infectious virus. Using neurons derived from human embryonic stem cells (hESC) and cell-free wild-type (WT) VZV, we demonstrated that neurons are nearly 100 times more permissive for WT VZV infection than very-early-passage human embryonic lung cells or MRC-5 diploid human fibroblasts, the cells used for vaccine production or virus isolation. The peak titers achieved after infection were ∼10-fold higher in human neurons than in MRC-5 cells, and the viral genome copy number-to-PFU ratio for VZV in human neurons was 500, compared with 50,000 for MRC-5 cells. Thus, VZV may not necessarily have a higher particle-to-PFU ratio than other herpesviruses; instead, the cells previously used to propagate virus in vitro may have been suboptimal. Furthermore, based on electron microscopy, neurons infected with VZV produced fewer defective or incomplete viral particles than MRC-5 cells. Our data suggest that neurons derived from hESC may have advantages compared to other cells for studies of VZV pathogenesis, for obtaining stocks of virus with high titers, and for isolating VZV from clinical specimens. IMPORTANCE Varicella-zoster virus (VZV) causes chickenpox and shingles. Cell-free VZV has been difficult to obtain, both for in vitro studies and for vaccine production. While numerous cells lines have been tested for their ability to produce high titers of VZV, the number of total virus particles relative to the number of viral particles that can form plaques in culture has been reported to be extremely high relative to that in other viruses. We show that VZV grows to much higher titers in human neurons than in other cell types in vitro and that the number of total virus genomes relative to the number of viral particles that can form plaques in culture is much lower in human neurons than other cultured cells. These findings indicate that human neurons may be useful for studying VZV in vitro , for growing preparations of virus with high titers, and for isolating the virus from human samples. Copyright © 2017 American Society for Microbiology.

Ecto-5′-Nucleotidase Activity in Human T Cell Subsets. DECREASED NUMBERS OF ECTO-5′-NUCLEOTIDASE POSITIVE CELLS FROM BOTH OKT4+ AND OKT8+ CELLS IN PATIENTS WITH HYPOGAMMAGLOBULINEMIA

PubMed Central

Thompson, Linda F.; Saxon, Andrew; O'Connor, Richard D.; Fox, Robert I.

1983-01-01

T lymphocytes from control subjects were separated into subsets using monoclonal antibodies of the OKT series and complement lysis and analyzed for ecto-5′-nucleotidase activity both by quantitative radiochemical assay and a histochemical stain. T cells from 15 control subjects contained 54±4% OKT4+ (helper/inducer) cells and 32±3% OKT8+ (cytotoxic/suppressor) cells. Total T cell ecto-5′-nucleotidase activity was 10.9±2.1 nmol/h per 106 cells with 25±7% positive by histochemical stain. Ecto-5′-nucleotidase activity in OKT4-enriched populations was 5.43±1.8 nmol/h per 106 cells with 14±2% positive by histochemical stain; that in OKT8-enriched populations was 17.1±5.9 nmol/h per 106 cells with 35±8% positive by histochemical stain. Two of four patients with congenital agammaglobulinemia and four of seven patients with common variable immunodeficiency had decreased proportions of OKT4+ T cells with corresponding increases in the proportions of OKT8+ T cells (OKT4/OKT8 = 0.60 to 1.0 as compared with 1.7±0.2 for control subjects). All four patients with congenital agammaglobulinemia, and three of seven patients with common variable immunodeficiency also had low T cell ecto-5′-nucleotidase activity (<5.5 nmol/h per 106 cells). Ecto-5′-nucleotidase activity in OKT4- enriched populations isolated from four patients with low total T cell activity was 2.85±0.90 nmol/h per 106 cells with 10±4% positive by histochemical stain; that in OKT8-enriched populations was 6.82±1.7 nmol/h per 106 cells with 7.5±3% positive by histochemical stain. Thus, the number of ecto-5′-nucleotidase positive cells is decreased, especially in the OKT8+ subpopulation, and the low total T cell ecto-5′-nucleotidase activity seen in these patients is due to fewer positive cells rather than to substantially less activity per cell. Our data indicate that ecto-5′-nucleotidase activity defines two subpopulations of T lymphocytes (ecto-5′-nucleotidase positive and negative), the proportions of which are markedly altered in many patients with hypogammaglobulinemia. In preliminary studies with seven patients, increased numbers of ecto-5′-nucleotidase negative T cells appeared to correlate with increased suppressor T cell activity toward in vitro immunoglobulin synthesis. Therefore, ecto-5′-nucleotidase may be a useful cell surface marker in the study of imbalances of regulatory T cell subsets in patients with antibody synthesis disorders. PMID:6300192

Secretory vesicles of immune cells contain only a limited number of interleukin 6 molecules.

PubMed

Verboogen, Daniëlle R J; Ter Beest, Martin; Honigmann, Alf; van den Bogaart, Geert

2018-05-01

Immune cells communicate by releasing large quantities of cytokines. Although the mechanisms of cytokine secretion are increasingly understood, quantitative knowledge of the number of cytokines per vesicle is still lacking. Here, we measured with quantitative microscopy the release rate of vesicles potentially carrying interleukin-6 (IL-6) in human dendritic cells. By comparing this to the total secreted IL-6, we estimate that secretory vesicles contain about 0.5-3 IL-6 molecules, but with a large spread among cells/donors. Moreover, IL-6 did not accumulate within most cells, indicating that synthesis and not trafficking is the bottleneck for IL-6 production. IL-6 accumulated in the Golgi apparatus only in ~ 10% of the cells. Understanding how immune cells produce cytokines is important for designing new immunomodulatory drugs. © 2018 The Authors. FEBS Letters published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.

The Distinctive Sensitivity to Microgravity of Immune Cell Subpopulations

NASA Astrophysics Data System (ADS)

Chen, Hui; Luo, Haiying; Liu, Jing; Wang, Peng; Dong, Dandan; Shang, Peng; Zhao, Yong

2015-11-01

Immune dysfunction in astronauts is well documented after spaceflights. Microgravity is one of the key factors directly suppressing the function of immune system. However, it is unclear which subpopulations of immune cells including innate and adaptive immune cells are more sensitive to microgravity We herein investigated the direct effects of modeled microgravity (MMg) on different immune cells in vitro. Mouse splenocytes, thymocytes and bone marrow cells were exposed to MMg for 16 hrs. The survival and the phenotypes of different subsets of immune cells including CD4+T cells, CD8+T cells, CD4+Foxp3+ regulatory T cells (Treg), B cells, monocytes/macrophages, dendritic cells (DCs), natural killer cells (NK) were determined by flow cytometry. After splenocytes were cultured under MMg for 16h, the cell frequency and total numbers of monocytes, macrophages and CD4+Foxp3+T cells were significantly decreased more than 70 %. MMg significantly decreased the cell numbers of CD8+ T cells, B cells and neutrophils in splenocytes. The cell numbers of CD4+T cells and NK cells were unchanged significantly when splenocytes were cultured under MMg compared with controls. However, MMg significantly increased the ratio of mature neutrophils to immature neutrophils in bone marrow and the cell number of DCs in splenocytes. Based on the cell survival ability, monocytes, macrophages and CD4+Foxp3+Treg cells are most sensitive to microgravity; CD4+T cells and NK cells are resistant to microgravity; CD8+T cells and neutrophils are impacted by short term microgravity exposure. Microgravity promoted the maturation of neutrophils and development of DCs in vitro. The present studies offered new insights on the direct effects of MMg on the survival and homeostasis of immune cell subsets.

Molecular Determinants of Radioresponse in Prostate Cancer

DTIC Science & Technology

2002-08-01

SECURITY CLASSIFICATION OF ABSTRACT Unclassified 15. NUMBER OF PAGES 72 16. PRICE CODE 20. LIMITATION OF ABSTRACT Unlimited Standard Form...irradiated LNCaP cells; right panel, irradiated RAT-1 HO 15.19 positive control cells (magnification lOOOx); (b) Time course of IR-induced apoptosis... right ) at 5 days post-irradiation following doses of OGy (control, mock- irradiated), 2Gy and lOGy. Shown is a representative experiment with total cell

Exploring the Role of Microbiota in the Limiting of B1 and MZ B-Cell Numbers by Naturally Secreted Immunoglobulins.

PubMed

Mohr, Elodie; Lino, Andreia C

2017-01-01

Immunoglobulins (Igs)-or antibodies (Ab)-are important to combat foreign pathogens but also to the immune system homeostasis. We developed the AID -/- μS -/- mouse model devoid of total soluble Igs and suitable to monitor the role of Igs on immune homeostasis. We used this experimental system to uncover a negative feedback control of marginal zone (MZ) and B1 B cells numbers by naturally secreted Igs. We raised AID -/- μS -/- mice in germ-free conditions demonstrating that this effect of natural secreted Igs is independent of the microbiota. Herein, we provide a comprehensive description of the protocols to establish and use the AID -/- μS -/- mice to study the role of total secreted Igs or of different Ig classes. This study involves Igs injections to AID -/- μS -/- mice or establishment of AID -/- μS -/- mixed bone marrow chimeras that provide a powerful system to study AID -/- μS -/- B cells in the presence of stable concentrations of different Ig classes. While we describe flow cytometric and histological methods to analyze MZ and B1 B cell subsets, AID -/- μS -/- mice can be used to study the effects of natural Igs on other B cell subsets or immune cells.

Analysis of B Cell Repertoire Dynamics Following Hepatitis B Vaccination in Humans, and Enrichment of Vaccine-specific Antibody Sequences.

PubMed

Galson, Jacob D; Trück, Johannes; Fowler, Anna; Clutterbuck, Elizabeth A; Münz, Márton; Cerundolo, Vincenzo; Reinhard, Claudia; van der Most, Robbert; Pollard, Andrew J; Lunter, Gerton; Kelly, Dominic F

2015-12-01

Generating a diverse B cell immunoglobulin repertoire is essential for protection against infection. The repertoire in humans can now be comprehensively measured by high-throughput sequencing. Using hepatitis B vaccination as a model, we determined how the total immunoglobulin sequence repertoire changes following antigen exposure in humans, and compared this to sequences from vaccine-specific sorted cells. Clonal sequence expansions were seen 7 days after vaccination, which correlated with vaccine-specific plasma cell numbers. These expansions caused an increase in mutation, and a decrease in diversity and complementarity-determining region 3 sequence length in the repertoire. We also saw an increase in sequence convergence between participants 14 and 21 days after vaccination, coinciding with an increase of vaccine-specific memory cells. These features allowed development of a model for in silico enrichment of vaccine-specific sequences from the total repertoire. Identifying antigen-specific sequences from total repertoire data could aid our understanding B cell driven immunity, and be used for disease diagnostics and vaccine evaluation.

Antigen-specific tolerance inhibits autoimmune uveitis in pre-sensitized animals by deletion and CD4+CD25+ T-regulatory cells.

PubMed

Matta, Bharati; Jha, Purushottam; Bora, Puran S; Bora, Nalini S

2010-02-01

The objective of this study was to inhibit experimental autoimmune anterior uveitis (EAAU) by establishing antigen-specific immune tolerance in animals pre-sensitized with melanin-associated antigen (MAA). Intravenous administration of MAA on days 6, 7, 8 and 9 post-immunization induced tolerance and inhibited EAAU in all Lewis rats. The number of cells (total T cells, CD4(+) T cells and CD8(+) T cells) undergoing apoptosis dramatically increased in the popliteal lymph nodes (LNs) of the tolerized animals compared with non-tolerized animals. In addition, Fas ligand (FasL), TNF receptor 1 (TNFR1) and caspase-8 were upregulated in tolerized rats. Proliferation of total lymphocytes, CD4(+)T cells and CD8(+) T cells (harvested from the popliteal LNs) in response to antigenic stimulation was drastically reduced in the state of tolerance compared with the cells from non-tolerized animals. The level of interferon (IFN)-gamma and IL-2 decreased, whereas TGF-beta2 was elevated in the state of tolerance. Furthermore, the number of CD4(+)CD25(+)FoxP3(+) regulatory T cells (Tregs) increased in the popliteal LNs of tolerized animals compared with non-tolerized animals. In conclusion, our results suggest that deletion of antigen-specific T cells by apoptosis and active suppression mediated by Tregs has an important role in the induction of antigen specific immune tolerance in animals with an established immune response against MAA.

Acute aerocystitis in Nile tilapia bred in net cages and supplemented with chromium carbochelate and Saccharomyces cerevisiae.

PubMed

Castro, Marcello P; Claudiano, Gustavo S; Petrillo, Thalita R; Shimada, Marina Tie; Belo, Marco A A; Marzocchi-Machado, Cleni M; Moraes, Julieta R E; Manrique, G Wilson; Moraes, Flávio R

2014-01-01

Oreochromis niloticus bred in net cages were supplemented with cell wall of Saccharomyces cerevisiae (Sc) (0.3%) or chromium carbochelate (Cr) (18 mg/kg of feed) or in association (Sc + Cr), for 90 days. After this period, acute inflammation was induced in the swim bladder by inoculation of 3 × 10(8) CFU of inactivated Streptococcus agalactiae, and another group received 0.65% saline solution (control). Twelve, 24, and 48 h after stimulation, the inflammation was evaluated through total and differential counting of accumulated cells, and through leukocyte respiratory burst in the blood, cortisolemia, glycemia and serum lysozyme concentration. The results showed that there were greater total numbers of cells in the exudate of fish inoculated with inactivated bacterium than in those injected with saline solution, with predominance of lymphocytes, thrombocytes, macrophages and granulocytes. Tilapia supplemented with Cr presented increased total numbers of cells with significant accumulation of lymphocytes and reductions in cortisolemia and glycemia, but the different treatments did not have any influence on leukocyte respiratory burst or serum lysozyme concentration. Tilapia supplemented with Sc and the Cr + Sc association did not present significant changes to the variables evaluated, despite higher accumulation of lymphocytes in the inflammatory exudate from fish treated with Sc. The results indicate that tilapia bred in net cages and supplemented with Cr presented higher total accumulation of cells at the inflammatory focus, thus indicating an increase in the inflammatory response induced by the bacterium, probably due to the reduction in cortisolemia and higher glucose consumption. Thus, supplementation with Cr had beneficial action, which facilitated development of acute inflammation induced by the bacterium, but did not affect neither leukocyte respiratory burst in the blood nor serum lysozyme concentration. Copyright © 2013 Elsevier Ltd. All rights reserved.

Reducing intratumour acute hypoxia through bevacizumab treatment, referring to the response of quiescent tumour cells and metastatic potential

PubMed Central

Masunaga, S; Liu, Y; Tanaka, H; Sakurai, Y; Suzuki, M; Kondo, N; Maruhashi, A; Ono, K

2011-01-01

Objectives The aim was to evaluate the influence of bevacizumab on intratumour oxygenation status and lung metastasis following radiotherapy, with specific reference to the response of quiescent (Q) cell populations within irradiated tumours. Methods B16-BL6 melanoma tumour-bearing C57BL/6 mice were continuously given 5-bromo-2-deoxyuridine (BrdU) to label all proliferating (P) cells. They received γ-ray irradiation following treatment with the acute hypoxia-releasing agent nicotinamide or local mild temperature hyperthermia (MTH) with or without the administration of bevacizumab under aerobic conditions or totally hypoxic conditions, achieved by clamping the proximal end of the tumours. Immediately after the irradiation, cells from some tumours were isolated and incubated with a cytokinesis blocker. The responses of the Q and total (P + Q) cell populations were assessed based on the frequency of micronuclei using immunofluorescence staining for BrdU. In the other tumour-bearing mice, macroscopic lung metastases were enumerated 17 days after irradiation. Results 3 days after bevacizumab administration, acute hypoxia-rich total cell population in the tumour showed a remarkably enhanced radiosensitivity to γ-rays, and the hypoxic fraction (HF) was reduced, even after MTH treatment. However, the hypoxic fraction was not reduced after nicotinamide treatment. With or without γ-ray irradiation, bevacizumab administration showed some potential to reduce the number of lung metastases as well as nicotinamide treatment. Conclusion Bevacizumab has the potential to reduce perfusion-limited acute hypoxia and some potential to cause a decrease in the number of lung metastases as well as nicotinamide. PMID:21586505

Recycled Cell Phones - A Treasure Trove of Valuable Metals

USGS Publications Warehouse

Sullivan, Daniel E.

2006-01-01

This U.S. Geological Survey (USGS) Fact Sheet examines the potential value of recycling the metals found in obsolete cell phones. Cell phones seem ubiquitous in the United States and commonplace throughout most of the world. There were approximately 1 billion cell phones in use worldwide in 2002. In the United States, the number of cell phone subscribers increased from 340,000 in 1985 to 180 million in 2004. Worldwide, cell phone sales have increased from slightly more than 100 million units per year in 1997 to an estimated 779 million units per year in 2005. Cell phone sales are projected to exceed 1 billion units per year in 2009, with an estimated 2.6 billion cell phones in use by the end of that year. The U.S. Environmental Protection Agency estimated that, by 2005, as many as 130 million cell phones would be retired annually in the United States. The nonprofit organization INFORM, Inc., anticipated that, by 2005, a total of 500 million obsolete cell phones would have accumulated in consumers' desk drawers, store rooms, or other storage, awaiting disposal. Typically, cell phones are used for only 1 1/2 years before being replaced. Less than 1 percent of the millions of cell phones retired and discarded annually are recycled. When large numbers of cell phones become obsolete, large quantities of valuable metals end up either in storage or in landfills. The amount of metals potentially recoverable would make a significant addition to total metals recovered from recycling in the United States and would supplement virgin metals derived from mining.

Necrotic cell death by hydrogen peroxide in immortal DF-1 chicken embryo fibroblast cells expressing deregulated MnSOD and catalase.

PubMed

Kim, H; You, S; Kong, B W; Foster, L K; Farris, J; Foster, D N

2001-08-22

The reactive oxygen species are known as endogenous toxic oxidant damaging factors in a variety of cell types, and in response, the antioxidant genes have been implicated in cell proliferation, senescence, immortalization, and tumorigenesis. The expression of manganese superoxide dismutase mRNA was shown to increase in most of the immortal chicken embryo fibroblast (CEF) cells tested, while expression of catalase mRNA appeared to be dramatically decreased in all immortal CEF cells compared to their primary counterparts. The expression of copper-zinc superoxide dismutase mRNA was shown to increase slightly in some immortal CEF cells. The glutathione peroxidase expressed relatively similar levels in both primary and immortal CEF cells. As primary and immortal DF-1 CEF cells were treated with 10-100 microM of hydrogen peroxide (concentrations known to be sublethal in human diploid fibroblasts), immortal DF-1 CEF cells were shown to be more sensitive to hydrogen peroxide, and total cell numbers were dramatically reduced when compared with primary cell counterparts. This increased sensitivity to hydrogen peroxide in immortal DF-1 cells occurred without evident changes in either antioxidant gene expression, mitochondrial membrane potential, cell cycle distribution or chromatin condensation. However, the total number of dead cells without chromatin condensation was dramatically elevated in immortal DF-1 CEFs treated with hydrogen peroxide, indicating that the inhibition of immortal DF-1 cell growth by low concentrations of hydrogen peroxide is due to increased necrotic cell death, but not apoptosis. Taken together, our observation suggests that the balanced antioxidant function might be important for cell proliferation in response to toxic oxidative damage by hydrogen peroxide.

Neuroprotective effects of diazoxide and its antagonism by glibenclamide in pyramidal neurons of rat hippocampus subjected to ischemia-reperfusion-induced injury.

PubMed

Zarch, Anoushiravan Vakili; Toroudi, Hamidreza Pazoki; Soleimani, Mansooreh; Bakhtiarian, Azam; Katebi, Majid; Djahanguiri, Bijan

2009-01-01

Mitochondrial ATP-sensitive potassium channel opener, diazoxide, is shown to have protective effect on the heart and brain following ischemia-reperfusion-induced injury (IR/II). However, the detailed effect of diazoxide and its antagonist on neuronal death, mitochondrial changes, and apoptosis in cerebral IR/II has not fully studied. IR/II was induced in rats by the 4-vessel occlusion model. Neuronal cell death and mitochondrial changes in CA1-CA4 pyramidal cells of the hippocampus were studied by light and electron microscopy, respectively. Apoptosis was assessed by measuring the amount of protein expressed by Bax and Bcl-2 genes. In light microscopy studies, the number of total and normal cells were increased only following 18 mg/kg of diazoxide. Lower doses (2 and 6 mg/kg) failed to change the cell numbers. All three doses of glibenclamide (1, 5, and 25 mg/kg) decreased the number of total and normal cell populations. In electron microscopy studies, different doses of diazoxide and glibenclamide prevented and aggravated the IR-induced morphological changes, respectively. Western blot analysis showed that diazoxide and glibenclamide inhibited and enhanced Bax protein expression respectively. Regarding Bcl-2 expression, only diazoxide showed a significant enhancement of gene expression. In conclusion, the results show that diazoxide can exhibit neuroprotective effects against IR/II in hippocampal regions, possibly through the opening of mitochondrial ATP-sensitive K(+) channels.

Global transcriptome analysis of the C57BL/6J mouse testis by SAGE: evidence for nonrandom gene order.

PubMed

Divina, Petr; Vlcek, Cestmír; Strnad, Petr; Paces, Václav; Forejt, Jirí

2005-03-05

We generated the gene expression profile of the total testis from the adult C57BL/6J male mice using serial analysis of gene expression (SAGE). Two high-quality SAGE libraries containing a total of 76 854 tags were constructed. An extensive bioinformatic analysis and comparison of SAGE transcriptomes of the total testis, testicular somatic cells and other mouse tissues was performed and the theory of male-biased gene accumulation on the X chromosome was tested. We sorted out 829 genes predominantly expressed from the germinal part and 944 genes from the somatic part of the testis. The genes preferentially and specifically expressed in total testis and testicular somatic cells were identified by comparing the testis SAGE transcriptomes to the available transcriptomes of seven non-testis tissues. We uncovered chromosomal clusters of adjacent genes with preferential expression in total testis and testicular somatic cells by a genome-wide search and found that the clusters encompassed a significantly higher number of genes than expected by chance. We observed a significant 3.2-fold enrichment of the proportion of X-linked genes specific for testicular somatic cells, while the proportions of X-linked genes specific for total testis and for other tissues were comparable. In contrast to the tissue-specific genes, an under-representation of X-linked genes in the total testis transcriptome but not in the transcriptomes of testicular somatic cells and other tissues was detected. Our results provide new evidence in favor of the theory of male-biased genes accumulation on the X chromosome in testicular somatic cells and indicate the opposite action of the meiotic X-inactivation in testicular germ cells.

Global transcriptome analysis of the C57BL/6J mouse testis by SAGE: evidence for nonrandom gene order

PubMed Central

Divina, Petr; Vlček, Čestmír; Strnad, Petr; Pačes, Václav; Forejt, Jiří

2005-01-01

Background We generated the gene expression profile of the total testis from the adult C57BL/6J male mice using serial analysis of gene expression (SAGE). Two high-quality SAGE libraries containing a total of 76 854 tags were constructed. An extensive bioinformatic analysis and comparison of SAGE transcriptomes of the total testis, testicular somatic cells and other mouse tissues was performed and the theory of male-biased gene accumulation on the X chromosome was tested. Results We sorted out 829 genes predominantly expressed from the germinal part and 944 genes from the somatic part of the testis. The genes preferentially and specifically expressed in total testis and testicular somatic cells were identified by comparing the testis SAGE transcriptomes to the available transcriptomes of seven non-testis tissues. We uncovered chromosomal clusters of adjacent genes with preferential expression in total testis and testicular somatic cells by a genome-wide search and found that the clusters encompassed a significantly higher number of genes than expected by chance. We observed a significant 3.2-fold enrichment of the proportion of X-linked genes specific for testicular somatic cells, while the proportions of X-linked genes specific for total testis and for other tissues were comparable. In contrast to the tissue-specific genes, an under-representation of X-linked genes in the total testis transcriptome but not in the transcriptomes of testicular somatic cells and other tissues was detected. Conclusion Our results provide new evidence in favor of the theory of male-biased genes accumulation on the X chromosome in testicular somatic cells and indicate the opposite action of the meiotic X-inactivation in testicular germ cells. PMID:15748293

"I Have Sickle Cell Disease, But Sickle Cell Doesn't Have Me" | NIH MedlinePlus the Magazine

MedlinePlus

... May 2009, and it has really changed my life. I haven't had a hospital admission in 13 months, and my blood numbers are wonderful. My doctor is totally pleased with it. I haven't had any negative side effects from the medicine. On staying healthy: It's much ...

Blocking variant surface glycoprotein synthesis alters endoplasmic reticulum exit sites/Golgi homeostasis in Trypanosoma brucei.

PubMed

Ooi, Cher-Pheng; Smith, Terry K; Gluenz, Eva; Wand, Nadina Vasileva; Vaughan, Sue; Rudenko, Gloria

2018-06-01

The predominant secretory cargo of bloodstream form Trypanosoma brucei is variant surface glycoprotein (VSG), comprising ~10% total protein and forming a dense protective layer. Blocking VSG translation using Morpholino oligonucleotides triggered a precise pre-cytokinesis arrest. We investigated the effect of blocking VSG synthesis on the secretory pathway. The number of Golgi decreased, particularly in post-mitotic cells, from 3.5 ± 0.6 to 2.0 ± 0.04 per cell. Similarly, the number of endoplasmic reticulum exit sites (ERES) in post-mitotic cells dropped from 3.9 ± 0.6 to 2.7 ± 0.1 eight hours after blocking VSG synthesis. The secretory pathway was still functional in these stalled cells, as monitored using Cathepsin L. Rates of phospholipid and glycosylphosphatidylinositol-anchor biosynthesis remained relatively unaffected, except for the level of sphingomyelin which increased. However, both endoplasmic reticulum and Golgi morphology became distorted, with the Golgi cisternae becoming significantly dilated, particularly at the trans-face. Membrane accumulation in these structures is possibly caused by reduced budding of nascent vesicles due to the drastic reduction in the total amount of secretory cargo, that is, VSG. These data argue that the total flux of secretory cargo impacts upon the biogenesis and maintenance of secretory structures and organelles in T. brucei, including the ERES and Golgi. © 2018 The Authors. Traffic published by John Wiley & Sons Ltd.

Stereological studies of the effects of sodium benzoate or ascorbic acid on rats` cerebellum.

PubMed

Noorafshan, Ali; Erfanizadeh, Mahboobeh; Karbalay-Doust, Saied

2014-12-01

To evaluate the cerebellar structure in sodium benzoate (NaB) or ascorbic acid (AA) treated rats. This experimental study was conducted between May and September 2013 in the Laboratory Animal Center of Shiraz University of Medical Sciences, Shiraz, Iran. The rats received distilled either water, NaB (200mg/kg/day), AA (100mg/kg/day), or NaB+AA. The hemispheres were removed after 28 days and underwent quantitative study. The total volume of the cerebellar hemisphere, its cortex, intracerebellar nuclei; the total number of the Purkinje, Bergman, granule, neurons, and glial cells of the molecular layer; and neurons and glial cells of the intracerebellar nuclei reduced by 21-52% in the NaB-treated rats compared with the distilled water group (p=0.004). The total number of the Purkinje, Bergman, Golgi, and granule cells was 29-45% higher in the AA-treated rats compared with the distilled water group (p=0.05). However, these measures reduced by 17-50% in the NaB+AA-treated rats compared with the distilled water group (p=0.004). The NaB+AA group did not induce any significant structural changes in comparison with the NaB group (p>0.05). The NaB exposure with or without AA treatment could alter the cerebellum. Yet, AA could prevent the loss of some cells in the cerebellum. 

Distribution of culturable microorganisms in Fennoscandian Shield groundwater.

PubMed

Haveman, Shelley A; Pedersen, Karsten

2002-02-01

Microbial populations in 16 groundwater samples from six Fennoscandian Shield sites in Finland and Sweden were investigated. The average total cell number was 3.7x10(5) cells ml(-1), and there was no change in the mean of the total cell numbers to a depth of 1390 m. Culture media were designed based on the chemical composition of each groundwater sample and used successfully to culture anaerobic microorganisms from all samples between 65 and 1350 m depth. Between 0.0084 and 14.8% of total cells were cultured from groundwater samples. Sulfate-reducing bacteria, iron-reducing bacteria and heterotrophic acetogenic bacteria were cultured from groundwater sampled at 65-686 m depth in geographically distant sites. Different microbial populations were cultured from deeper, older and more saline groundwater from 863 to 1350 m depth. Principal component analysis of groundwater chemistry data showed that sulfate- and iron-reducing bacteria were not detected in the most saline groundwater. Iron-reducing bacteria and acetogens were cultured from deep groundwater that contained 0.35-3.5 mM sulfate, while methanogens and acetogens were cultured from deep sulfate-depleted groundwater. In one borehole from which autotrophic methanogens were cultured, dissolved inorganic carbon was enriched in (13)C compared to other Fennoscandian Shield groundwater samples, suggesting that autotrophs were active. It can be concluded that a diverse microbial community is present from the surface to over 1300 m depth in the Fennoscandian Shield.

Multiple intravenous injections of allogeneic equine mesenchymal stem cells do not induce a systemic inflammatory response but do alter lymphocyte subsets in healthy horses.

PubMed

Kol, Amir; Wood, Joshua A; Carrade Holt, Danielle D; Gillette, Jessica A; Bohannon-Worsley, Laurie K; Puchalski, Sarah M; Walker, Naomi J; Clark, Kaitlin C; Watson, Johanna L; Borjesson, Dori L

2015-04-15

Intravenous (IV) injection of mesenchymal stem cells (MSCs) is used to treat systemic human diseases and disorders but is not routinely used in equine therapy. In horses, MSCs are isolated primarily from adipose tissue (AT) or bone marrow (BM) and used for treatment of orthopedic injuries through one or more local injections. The objective of this study was to determine the safety and lymphocyte response to multiple allogeneic IV injections of either AT-derived MSCs (AT-MSCs) or BM-derived MSCs (BM-MSCs) to healthy horses. We injected three doses of 25 × 10(6) allogeneic MSCs from either AT or BM (a total of 75 × 10(6) MSCs per horse) into five and five, respectively, healthy horses. Horses were followed up for 35 days after the first MSC infusion. We evaluated host inflammatory and immune response, including total leukocyte numbers, serum cytokine concentration, and splenic lymphocyte subsets. Repeated injection of allogeneic AT-MSCs or BM-MSCs did not elicit any clinical adverse effects. Repeated BM-MSC injection resulted in increased blood CD8(+) T-cell numbers. Multiple BM-MSC injections also increased splenic regulatory T cell numbers compared with AT-MSC-injected horses but not controls. These data demonstrate that multiple IV injections of allogeneic MSCs are well tolerated by healthy horses. No clinical signs or clinico-pathologic measurements of organ toxicity or systemic inflammatory response were recorded. Increased numbers of circulating CD8(+) T cells after multiple IV injections of allogeneic BM-MSCs may indicate a mild allo-antigen-directed cytotoxic response. Safety and efficacy of allogeneic MSC IV infusions in sick horses remain to be determined.

Structural analysis, electronic properties, and band gaps of a graphene nanoribbon: A new 2D materials

NASA Astrophysics Data System (ADS)

Dass, Devi

2018-03-01

Graphene nanoribbon (GNR), a new 2D carbon nanomaterial, has some unique features and special properties that offer a great potential for interconnect, nanoelectronic devices, optoelectronics, and nanophotonics. This paper reports the structural analysis, electronic properties, and band gaps of a GNR considering different chirality combinations obtained using the pz orbital tight binding model. In structural analysis, the analytical expressions for GNRs have been developed and verified using the simulation for the first time. It has been found that the total number of unit cells and carbon atoms within an overall unit cell and molecular structure of a GNR have been changed with the change in their chirality values which are similar to the values calculated using the developed analytical expressions thus validating both the simulation as well as analytical results. Further, the electronic band structures at different chirality values have been shown for the identification of metallic and semiconductor properties of a GNR. It has been concluded that all zigzag edge GNRs are metallic with very small band gaps range whereas all armchair GNRs show both the metallic and semiconductor nature with very small and high band gaps range. Again, the total number of subbands in each electronic band structure is equal to the total number of carbon atoms present in overall unit cell of the corresponding GNR. The semiconductors GNRs can be used as a channel material in field effect transistor suitable for advanced CMOS technology whereas the metallic GNRs could be used for interconnect.

Cell Size Influences the Reproductive Potential and Total Lifespan of the Saccharomyces cerevisiae Yeast as Revealed by the Analysis of Polyploid Strains.

PubMed

Zadrag-Tecza, Renata; Kwolek-Mirek, Magdalena; Alabrudzińska, Małgorzata; Skoneczna, Adrianna

2018-01-01

The total lifespan of the yeast Saccharomyces cerevisiae may be divided into two phases: the reproductive phase, during which the cell undergoes mitosis cycles to produce successive buds, and the postreproductive phase, which extends from the last division to cell death. These phases may be regulated by a common mechanism or by distinct ones. In this paper, we proposed a more comprehensive approach to reveal the mechanisms that regulate both reproductive potential and total lifespan in cell size context. Our study was based on yeast cells, whose size was determined by increased genome copy number, ranging from haploid to tetraploid. Such experiments enabled us to test the hypertrophy hypothesis, which postulates that excessive size achieved by the cell-the hypertrophy state-is the reason preventing the cell from further proliferation. This hypothesis defines the reproductive potential value as the difference between the maximal size that a cell can reach and the threshold value, which allows a cell to undergo its first cell cycle and the rate of the cell size to increase per generation. Here, we showed that cell size has an important impact on not only the reproductive potential but also the total lifespan of this cell. Moreover, the maximal cell size value, which limits its reproduction capacity, can be regulated by different factors and differs depending on the strain ploidy. The achievement of excessive size by the cell (hypertrophic state) may lead to two distinct phenomena: the cessation of reproduction without "mother" cell death and the cessation of reproduction with cell death by bursting, which has not been shown before.

Radiation protocols determine acute graft-versus-host disease incidence after allogeneic bone marrow transplantation in murine models.

PubMed

Schwarte, Sebastian; Bremer, Michael; Fruehauf, Joerg; Sorge, Yanina; Skubich, Susanne; Hoffmann, Matthias W

2007-09-01

Effects of radiation sources used for total body irradiation (TBI) on Graft-versus-Host Disease (GvHD) induction were examined. In a T cell receptor (TCR) transgenic mouse model, single fraction TBI was performed with different radiation devices ((60)Cobalt; (137)Cesium; 6 MV linear accelerator), dose rates (0.85; 1.5; 2.9; 5 Gy/min) and total doses before allogeneic bone marrow transplantation (BMT). Recipients were observed for 120 days. Different tissues were examined histologically. Acute GvHD was induced by a dose rate of 0.85 Gy/min ((60)Cobalt) and a total dose of 9 Gy and injection of 5 x 10(5) lymph node cells plus 5 x 10(6) bone marrow cells. Similar results were obtained using 6 MV linear accelerator- (linac-) photons with a dose rate of 1.5 Gy/min and 0.85 Gy/min, a total dose of 9.5 Gy and injection of same cell numbers. TBI with (137)Cesium (dose rate: 2.5 Gy/min) did not lead reproducibly to lethal acute GvHD. Experimental TBI in murine models may induce different immunological responses, depending on total energy, total single dose and dose rate. GvHD might also be induced by TBI with low dose rates.

Assessments of Total and Viable Escherichia coli O157:H7 on Field and Laboratory Grown Lettuce

PubMed Central

Moyne, Anne-Laure; Harris, Linda J.; Marco, Maria L.

2013-01-01

Leafy green produce has been associated with numerous outbreaks of foodborne illness caused by strains of Escherichia coli O157:H7. While the amounts of culturable E. coli O157:H7 rapidly decline after introduction onto lettuce in the field, it remains to be determined whether the reduction in cell numbers is due to losses in cell viability, cell injury and a subsequent inability to be detected by standard laboratory culturing methods, or a lack of adherence and hence rapid removal of the organism from the plants during application. To assess which of these options is most relevant for E. coli O157:H7 on leafy green produce, we developed and applied a propidium monoazide (PMA) real-time PCR assay to quantify viable (with PMA) and total (without PMA) E. coli O157:H7 cells on growth chamber and field-grown lettuce. E. coli O157:H7, suspended in 0.1% peptone, was inoculated onto 4-week-old lettuce plants at a level of approximately 106 CFU/plant. In the growth chamber at low relative humidity (30%), culturable amounts of the nontoxigenic E. coli O157:H7 strain ATCC 700728 and the virulent strain EC4045 declined 100 to 1000-fold in 24 h. Fewer E. coli O157:H7 cells survived when applied onto plants in droplets with a pipette compared with a fine spray inoculation. Total cells for both strains were equivalent to inoculum levels for 7 days after application, and viable cell quantities determined by PMA real-time PCR were approximately 104 greater than found by colony enumeration. Within 2 h after application onto plants in the field, the number of culturable E. coli ATCC 700728 was reduced by up to 1000-fold, whereas PCR-based assessments showed that total cell amounts were equivalent to inoculum levels. These findings show that shortly after inoculation onto plants, the majority of E. coli O157:H7 cells either die or are no longer culturable. PMID:23936235

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

Effect of Native and Minimally Modified Low-density Lipoprotein on the Activation of Monocyte Subsets.

PubMed

Blanco-Favela, Francisco; Espinosa-Luna, José Esteban; Chávez-Rueda, Adriana Karina; Madrid-Miller, Alejandra; Chávez-Sánchez, Luis

2017-07-01

In atherosclerosis, monocytes are essential and secrete pro-inflammatory cytokines in response to modified low-density lipoprotein (LDL). Human CD14 ++ CD16 - , CD14 ++ CD16 + and CD14 + CD16 ++ monocytes produce different cytokines. The objective of this research was to determine the number of monocyte subsets positives to cytokines in response to native (nLDL) and minimally modified LDL (mmLDL). Human monocytes from healthy individuals were purified by negative selection and were stimulated with nLDL, mmLDL or LPS. Subsequently, human total monocytes were incubated with monoclonal antibodies specific for CD14 or both CD14 and CD16 to characterize total monocytes and monocyte subsets and with antibodies specific to anti-tumor necrosis factor (TNF)-α, anti-interleukin (IL)-6 and anti-IL-10. The number of cells positive for cytokines was determined and cells cultured with nLDL, mmLDL and LPS were compared with cells cultured only with culture medium. We found that nLDL does not induce in the total monocyte population or in the three monocyte subsets positives to cytokines. MmLDL induced in total monocytes positives to TNF-α and IL-6 as well as in both CD14 ++ CD16 + and CD14 + CD16 ++ and in CD14 ++ CD16 + monocytes, respectively. Moreover, total monocytes and the three monocyte subsets expressed few amounts of cells positives to IL-10 in response to mmLDL. Our study demonstrated that nLDL did not induce cells positives to cytokines and that the CD14 ++ CD16 + and CD14 + CD16 ++ monocyte subsets could be the main sources of TNF-α and IL-6, respectively, in response to mmLDL, which promotes the development and progression of atherosclerotic plaque. Copyright © 2017 IMSS. Published by Elsevier Inc. All rights reserved.

[Resveratrol derived from rhizoma et radix polygoni cuspidati and its liposomal form protect nigral cells of Parkinsonian rats].

PubMed

Wang, Yanchun; Xu, Hanlin; Fu, Qin; Ma, Rong; Xiang, Jizhou

2011-04-01

Oxidative stress is a hallmark in the pathogenesis of Parkinson disease (PD), which involves the selective loss of nigral dopaminergic neurons in PD. Resveratrol (3,5,4'-trihydroxy-trans-stilbene) is well known for its powerful antioxidant property and a wide range of other biological effects. In this study, we investigated the protective effect of resveratrol derived from Rhizoma Et Radix Polygoni Cuspidati and its liposomal form on the nigral cells of PD rats induced by unilateral microinjection of 6-hydroxy dopamine in the striatum. The results showed that after 14 days gavage of resveratrol and resveratrol liposome respectively (20 mg x kg(-1) WB per day), the abnormal rotational behavior of PD rats were deceased evidently, the numbers of total nigral cells, total nigral neurons and TH immuno-positive neurons were more than that of PD rats without given resveratrol or resveratrol liposome, simultaneously, the number of apoptotic nigral cells were decreased obviously. The results also showed that resveratrol and resveratrol liposome could decrease the total ROS activity, increase the total antioxidant capability of the nigral tissues. All the data indicated that resveratrol liposome performed stronger effects than resveratrol except for behavioral improvement. Our study confirmed that resveratrol derived from Rhizoma Et Radix Polygoni Cuspidati and its liposomal form could inhibit the loss of dopaminergic neurons of PD rats, the underlying mechanism may be attributed to their radical scavenging effect and antioxidant property. Due to presumably increased bioavailability, resveratrol liposome possesses the stronger therapeutic effect and may become a better clinical agent for the treatment of PD than free resveratrol.

The impact of temperature on microbial diversity and AOA activity in the Tengchong Geothermal Field, China

PubMed Central

Li, Haizhou; Yang, Qunhui; Li, Jian; Gao, Hang; Li, Ping; Zhou, Huaiyang

2015-01-01

Using a culture-independent method that combines CARD-FISH, qPCR and 16S rDNA, we investigated the abundance, community structure and diversity of microbes along a steep thermal gradient (50–90 °C) in the Tengchong Geothermal Field. We found that Bacteria and Archaea abundance changed markedly with temperature changes and that the number of cells was lowest at high temperatures (90.8 °C). Under low-temperature conditions (52.3–74.6 °C), the microbial communities were dominated by Bacteria, which accounted for 60–80% of the total number of cells. At 74.6 °C, Archaea were dominant, and at 90.8 °C, they accounted for more than 90% of the total number of cells. Additionally, the microbial communities at high temperatures (74.6–90.8 °C) were substantially simpler than those at the low-temperature sites. Only a few genera (e.g., bacterial Caldisericum, Thermotoga and Thermoanaerobacter, archaeal Vulcanisaeta and Hyperthermus) often dominated in high-temperature environments. Additionally, a positive correlation between Ammonia-Oxidizing Archaea (AOA) activity and temperature was detected. AOA activity increased from 17 to 52 pmol of NO2− per cell d−1 with a temperature change from 50 to 70 °C. PMID:26608685

Measurement uncertainty for the Uniform Engine Testing Program conducted at NASA Lewis Research Center

NASA Technical Reports Server (NTRS)

Abdelwahab, Mahmood; Biesiadny, Thomas J.; Silver, Dean

1987-01-01

An uncertainty analysis was conducted to determine the bias and precision errors and total uncertainty of measured turbojet engine performance parameters. The engine tests were conducted as part of the Uniform Engine Test Program which was sponsored by the Advisory Group for Aerospace Research and Development (AGARD). With the same engines, support hardware, and instrumentation, performance parameters were measured twice, once during tests conducted in test cell number 3 and again during tests conducted in test cell number 4 of the NASA Lewis Propulsion Systems Laboratory. The analysis covers 15 engine parameters, including engine inlet airflow, engine net thrust, and engine specific fuel consumption measured at high rotor speed of 8875 rpm. Measurements were taken at three flight conditions defined by the following engine inlet pressure, engine inlet total temperature, and engine ram ratio: (1) 82.7 kPa, 288 K, 1.0, (2) 82.7 kPa, 288 K, 1.3, and (3) 20.7 kPa, 288 K, 1.3. In terms of bias, precision, and uncertainty magnitudes, there were no differences between most measurements made in test cells number 3 and 4. The magnitude of the errors increased for both test cells as engine pressure level decreased. Also, the level of the bias error was two to three times larger than that of the precision error.

Significance of manipulating tumour hypoxia and radiation dose rate in terms of local tumour response and lung metastatic potential, referring to the response of quiescent cell populations

PubMed Central

Masunaga, S; Matsumoto, Y; Kashino, G; Hirayama, R; Liu, Y; Tanaka, H; Sakurai, Y; Suzuki, M; Kinashi, Y; Maruhashi, A; Ono, K

2010-01-01

The purpose of this study was to evaluate the influence of manipulating intratumour oxygenation status and radiation dose rate on local tumour response and lung metastases following radiotherapy, referring to the response of quiescent cell populations within irradiated tumours. B16-BL6 melanoma tumour-bearing C57BL/6 mice were continuously given 5-bromo-2′-deoxyuridine (BrdU) to label all proliferating (P) cells. They received γ-ray irradiation at high dose rate (HDR) or reduced dose rate (RDR) following treatment with the acute hypoxia-releasing agent nicotinamide or local hyperthermia at mild temperatures (MTH). Immediately after the irradiation, cells from some tumours were isolated and incubated with a cytokinesis blocker. The responses of the quiescent (Q) and total (proliferating + Q) cell populations were assessed based on the frequency of micronuclei using immunofluorescence staining for BrdU. In other tumour-bearing mice, 17 days after irradiation, macroscopic lung metastases were enumerated. Following HDR irradiation, nicotinamide and MTH enhanced the sensitivity of the total and Q-cell populations, respectively. The decrease in sensitivity at RDR irradiation compared with HDR irradiation was slightly inhibited by MTH, especially in Q cells. Without γ-ray irradiation, nicotinamide treatment tended to reduce the number of lung metastases. With γ-rays, in combination with nicotinamide or MTH, especially the former, HDR irradiation decreased the number of metastases more remarkably than RDR irradiation. Manipulating both tumour hypoxia and irradiation dose rate have the potential to influence lung metastasis. The combination with the acute hypoxia-releasing agent nicotinamide may be more promising in HDR than RDR irradiation in terms of reducing the number of lung metastases. PMID:20739345

Quantitative analysis of taste bud cell numbers in fungiform and soft palate taste buds of mice.

PubMed

Ohtubo, Yoshitaka; Yoshii, Kiyonori

2011-01-07

Mammalian taste bud cells (TBCs) consist of several cell types equipped with different taste receptor molecules, and hence the ratio of cell types in a taste bud constitutes the taste responses of the taste bud. Here we show that the population of immunohistochemically identified cell types per taste bud is proportional to the number of total TBCs in the taste bud or the area of the taste bud in fungiform papillae, and that the proportions differ among cell types. This result is applicable to soft palate taste buds. However, the density of almost all cell types, the population of cell types divided by the area of the respective taste buds, is significantly higher in soft palates. These results suggest that the turnover of TBCs is regulated to keep the ratio of each cell type constant, and that taste responsiveness is different between fungiform and soft palate taste buds. Copyright © 2010 Elsevier B.V. All rights reserved.

Naïve and memory CD8 T cell pool homeostasis in advanced aging: impact of age and of antigen-specific responses to cytomegalovirus.

PubMed

Vescovini, Rosanna; Fagnoni, Francesco Fausto; Telera, Anna Rita; Bucci, Laura; Pedrazzoni, Mario; Magalini, Francesca; Stella, Adriano; Pasin, Federico; Medici, Maria Cristina; Calderaro, Adriana; Volpi, Riccardo; Monti, Daniela; Franceschi, Claudio; Nikolich-Žugich, Janko; Sansoni, Paolo

2014-04-01

Alterations in the circulating CD8+ T cell pool, with a loss of naïve and accumulation of effector/effector memory cells, are pronounced in older adults. However, homeostatic forces that dictate such changes remain incompletely understood. This observational cross-sectional study explored the basis for variability of CD8+ T cell number and composition of its main subsets: naïve, central memory and effector memory T cells, in 131 cytomegalovirus (CMV) seropositive subjects aged over 60 years. We found great heterogeneity of CD8+ T cell numbers, which was mainly due to variability of the CD8 + CD28- T cell subset regardless of age. Analysis, by multiple regression, of distinct factors revealed that age was a predictor for the loss in absolute number of naïve T cells, but was not associated with changes in central or effector memory CD8+ T cell subsets. By contrast, the size of CD8+ T cells specific to pp65 and IE-1 antigens of CMV, predicted CD28 - CD8+ T cell, antigen-experienced CD8+ T cell, and even total CD8+ T cell numbers, but not naïve CD8+ T cell loss. These results indicate a clear dichotomy between the homeostasis of naïve and antigen-experienced subsets of CD8+ T cells which are independently affected, in human later life, by age and antigen-specific responses to CMV, respectively.

Low Copy Numbers of DC-SIGN in Cell Membrane Microdomains: Implications for Structure and Function

PubMed Central

Liu, Ping; Wang, Xiang; Itano, Michelle S.; Neumann, Aaron K.; de Silva, Aravinda M.; Jacobson, Ken; Thompson, Nancy L.

2014-01-01

Presently, there are few estimates of the number of molecules occupying membrane domains. Using a total internal reflection fluorescence microscopy (TIRFM) imaging approach, based on comparing the intensities of fluorescently labeled microdomains with those of single fluorophores, we measured the occupancy of DC-SIGN, a C-type lectin, in membrane microdomains. DC-SIGN or its mutants were labeled with primary monoclonal antibodies (mAbs) in either dendritic cells (DCs) or NIH3T3 cells, or expressed as GFP fusions in NIH3T3 cells. The number of DC-SIGN molecules per microdomain ranges from only a few to over 20, while microdomain dimensions range from the diffraction limit to > 1μm. The largest fraction of microdomains, appearing at the diffraction limit, in either immature DCs or 3T3 cells contains only 4-8 molecules of DC-SIGN, consistent with our preliminary super-resolution Blink microscopy estimates. We further show that these small assemblies are sufficient to bind and efficiently internalize a small (~50nm) pathogen, dengue virus, leading to infection of host cells. PMID:24313910

Number of negative lymph nodes as a prognostic factor in esophageal squamous cell carcinoma.

PubMed

Ma, Mingquan; Tang, Peng; Jiang, Hongjing; Gong, Lei; Duan, Xiaofeng; Shang, Xiaobin; Yu, Zhentao

2017-10-01

The aim of this study is to investigate the number of negative lymph nodes (NLNs) as a prognostic factor for survival in patients with resected esophageal squamous cell carcinoma. A total of 381 esophageal squamous cell carcinoma patients who had underwent surgical resection as the primary treatment was enrolled into this retrospective study. The impact of number of NLNs on patient's overall survival was assessed and compared with the factors among the current tumor-nodes-metastasis (TNM) staging system. The number of NLNs was closely related to the overall survival, and the 5-year survival rate was 45.4% for number of NLNs of >20 (142 cases) and 26.4% for NLNs ≤ 20 (239 cases) (P = 0.001). In multivariate survival analysis, the number of NLNs remained an independent prognostic factor (P = 0.002) as did the other current TNM factors. For subgroup analysis, the predictive value of number of NLNs was significant in patients with T3 or T4 disease (P = 0.001) and patients with N1 and N2-3 disease (P = 0.025, 0.043), but not in patients with T1 or T2 disease or patients with N0 disease. The number of NLNs, which represents the extent of lymphadenectomy for esophageal squamous cell carcinoma, could impact the overall survival of patients with resected esophageal squamous cell carcinoma, especially among those with nodal-positive disease and advanced T-stage tumor. © 2016 John Wiley & Sons Australia, Ltd.

Early effect of boron neutron capture therapy mediated by boronophenylalanine (BPA-BNCT) on mast cells in premalignant tissue and tumors of the hamster cheek pouch.

PubMed

Aromando, Romina F; Trivillin, Verónica A; Heber, Elisa M; Pozzi, Emiliano; Schwint, Amanda E; Itoiz, María E

2010-05-01

Mast cell (MC) activation in the hamster cheek pouch cancerization model is associated with the increase in tumor cell proliferation, mediated in turn by tryptase, a protease released from mast cell granules after activation. Tryptase induces tumor cell proliferation through the activation of PAR-2 (protease activated receptor-2) on the plasma membrane of carcinoma cells. The therapeutic success of boron neutron capture therapy mediated by boronophenylalanine (BPA-BNCT) in tumor control in the hamster cheek pouch oral cancer model has been previously reported by our laboratory. Early effects of BPA-BNCT on tumors of the hamster cheek pouch include a reduction in DNA-synthesis with the concomitant decrease in the proliferation of malignant cells. The aim of the present study was to investigate the early histological changes in mast cells after BPA-BNCT in tumors and premalignant tissue of the hamster cheek pouch. Tumor-bearing pouches were treated with BPA-BNCT or beam only (neutron irradiation without prior administration of the boron compound) and sacrificed 1day after treatment. The samples were fixed in Carnoy fixative and stained with alcian blue-safranin to identify all the populations of mast cells. Total, active and inactive mast cells (MC) were counted in the connective tissue and the adventitious tissue underlying the pouch wall and at the base of the tumors in pouches treated with BPA-BNCT, in keeping with a previously described technique. BPA-BNCT induced a marked reduction in the total number of mast cells in the pouch (p<0.05). This reduction in the total number of mast cells was due to a reduction in mast cells at the base of the tumor (p<0.005) and it occurred at the expense of the active mast cells (p<0.05). A slight reduction that did not reach statistical significance also occurred in the amount of mast cells in the pouch wall (that corresponds to the premalignant tissue in tumor-bearing pouches), and in the adventitious tissue. In this case the reduction was seen in the inactive population. Both BPA-BNCT and beam only elicited a qualitative change in the secretion modality of the granule content. Although further studies are needed to evaluate the subcellular effect of BNCT on mast cell granule secretion, the reduction in cell proliferation induced by BPA-BNCT would be partially due to the decrease in total mast cells in the hamster check pouch. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

Cell Size Influences the Reproductive Potential and Total Lifespan of the Saccharomyces cerevisiae Yeast as Revealed by the Analysis of Polyploid Strains

PubMed Central

Kwolek-Mirek, Magdalena; Alabrudzińska, Małgorzata

2018-01-01

The total lifespan of the yeast Saccharomyces cerevisiae may be divided into two phases: the reproductive phase, during which the cell undergoes mitosis cycles to produce successive buds, and the postreproductive phase, which extends from the last division to cell death. These phases may be regulated by a common mechanism or by distinct ones. In this paper, we proposed a more comprehensive approach to reveal the mechanisms that regulate both reproductive potential and total lifespan in cell size context. Our study was based on yeast cells, whose size was determined by increased genome copy number, ranging from haploid to tetraploid. Such experiments enabled us to test the hypertrophy hypothesis, which postulates that excessive size achieved by the cell—the hypertrophy state—is the reason preventing the cell from further proliferation. This hypothesis defines the reproductive potential value as the difference between the maximal size that a cell can reach and the threshold value, which allows a cell to undergo its first cell cycle and the rate of the cell size to increase per generation. Here, we showed that cell size has an important impact on not only the reproductive potential but also the total lifespan of this cell. Moreover, the maximal cell size value, which limits its reproduction capacity, can be regulated by different factors and differs depending on the strain ploidy. The achievement of excessive size by the cell (hypertrophic state) may lead to two distinct phenomena: the cessation of reproduction without “mother” cell death and the cessation of reproduction with cell death by bursting, which has not been shown before. PMID:29743970

The toxicity, in vitro, of silicon carbide whiskers.

PubMed

Vaughan, G L; Jordan, J; Karr, S

1991-10-01

To mouse cells in culture, SiC whiskers (SiCW) and asbestos are similarly cytotoxic, disrupting cell membranes and killing cells. Both shorten cell generation time, increase the rate of DNA synthesis, increase total cell DNA content, and cause a loss in growth control often associated with malignant cellular transformation. Within the narrow size range of materials examined, the amount of damage appeared to be more a function of the number of whiskers present than of their size. Silicon carbide whiskers, if mishandled, may pose a serious health hazard to humans.

The significance of gallstones in children with sickle cell anemia.

PubMed Central

Alexander-Reindorf, C.; Nwaneri, R. U.; Worrell, R. G.; Ogbonna, A.; Uzoma, C.

1990-01-01

Infection is the most common cause of high morbidity, hospitalization, and mortality in children with sickle cell anemia. In this study of pediatric sickle cell anemia patients, aged 1 to 19, we explore the hypothesis that gallstones (usually pigment stones) create a nidus of infection, predisposing the affected patients to high morbidity. Our study involved 86 children with sickle cell anemia at the Howard University Center for Sickle Cell Disease, who had been followed at the clinic for a total of 602 patient years. Review of their records revealed that patients with gallstones had a mean number of 10.24 hospitalizations and 25.35 ambulatory visits; those without gallstones had a mean number of only 4.26 hospitalizations and 13.41 ambulatory visits. In children with sickle cell anemia and gallstones, elective cholecystectomy (or, in the future, cholelithotripsy) could reduce the high morbidity caused by infection. PMID:2213913

Morphometry of the midgut of Melipona quadrifasciata anthidioides (Lepeletier) (Hymenoptera: Apidae) during metamorphosis.

PubMed

Cruz, L C; Araújo, V A; Dolder, H; Araújo, A P A; Serrão, J E; Neves, C A

2011-01-01

In Hymenoptera, midgut changes begin in the last instar. At this stage, the larval epithelial digestive cells degenerate, leaving only the basal membrane and the regenerative cells which will develop into a new epithelium during the pupal stage and in the adult. Epithelium renewal is followed by changes in volume and shape of the midgut. Morphometric analysis of digestive cells and total midgut volume of Melipona quadrifasciata anthidioides (Lepeletier) were conducted to verify whether cell volume increase are sufficient to account for the total midgut volume increase that occurs during metamorphosis. An increase in midgut volume was verified in spite of the scarcity of cell proliferation found during metamorphosis. At the end of metamorphosis, the increase in cell volume was not sufficient to explain the increase in volume of the midgut, indicating that an increase in the number of digestive cells is apparently necessary. Nevertheless, the mechanism by which regenerative cells reconstitute the epithelium during metamorphosis remains unknown.

Impact of Aging, Cytomegalovirus Infection, and Long-Term Treatment for Human Immunodeficiency Virus on CD8+ T-Cell Subsets

PubMed Central

Veel, Ellen; Westera, Liset; van Gent, Rogier; Bont, Louis; Otto, Sigrid; Ruijsink, Bram; Rabouw, Huib H.; Mudrikova, Tania; Wensing, Annemarie; Hoepelman, Andy I. M.; Borghans, José A. M.; Tesselaar, Kiki

2018-01-01

Both healthy aging and human immunodeficiency virus (HIV) infection lead to a progressive decline in naive CD8+ T-cell numbers and expansion of the CD8+ T-cell memory and effector compartments. HIV infection is therefore often considered a condition of premature aging. Total CD8+ T-cell numbers of HIV-infected individuals typically stay increased even after long-term (LT) combination antiretroviral treatment (cART), which is associated with an increased risk of non-AIDS morbidity and mortality. The causes of these persistent changes in the CD8+ T-cell pool remain debated. Here, we studied the impact of age, CMV infection, and LT successful cART on absolute cell numbers in different CD8+ T-cell subsets. While naïve CD8+ T-cell numbers in cART-treated individuals (N = 38) increased to healthy levels, central memory (CM), effector memory (EM), and effector CD8+ T-cell numbers remained higher than in (unselected) age-matched healthy controls (N = 107). Longitudinal analysis in a subset of patients showed that cART did result in a loss of memory CD8+ T-cells, mainly during the first year of cART, after which memory cell numbers remained relatively stable. As CMV infection is known to increase CD8+ T-cell numbers in healthy individuals, we studied whether any of the persistent changes in the CD8+ T-cell pools of cART-treated patients could be a direct reflection of the high CMV prevalence among HIV-infected individuals. We found that EM and effector CD8+ T-cell numbers in CMV+ healthy individuals (N = 87) were significantly higher than in CMV− (N = 170) healthy individuals. As a result, EM and effector CD8+ T-cell numbers in successfully cART-treated HIV-infected individuals did not deviate significantly from those of age-matched CMV+ healthy controls (N = 39). By contrast, CM T-cell numbers were quite similar in CMV+ and CMV− healthy individuals across all ages. The LT expansion of the CM CD8+ T-cell pool in cART-treated individuals could thus not be attributed directly to CMV and was also not related to residual HIV RNA or to the presence of HIV-specific CM T-cells. It remains to be investigated why the CM CD8+ T-cell subset shows seemingly irreversible changes despite years of effective treatment. PMID:29619031

Hydrogen-Rich Water Ameliorates Total Body Irradiation-Induced Hematopoietic Stem Cell Injury by Reducing Hydroxyl Radical

PubMed Central

Xue, Xiaolei; Han, Xiaodan; Li, Yuan; Lu, Lu; Li, Deguan

2017-01-01

We examined whether consumption of hydrogen-rich water (HW) could ameliorate hematopoietic stem cell (HSC) injury in mice with total body irradiation (TBI). The results indicated that HW alleviated TBI-induced HSC injury with respect to cell number alteration and to the self-renewal and differentiation of HSCs. HW specifically decreased hydroxyl radical (∙OH) levels in the c-kit+ cells of 4 Gy irradiated mice. Proliferative bone marrow cells (BMCs) increased and apoptotic c-kit+ cells decreased in irradiated mice uptaken with HW. In addition, the mean fluorescence intensity (MFI) of γ-H2AX and percentage of 8-oxoguanine positive cells significantly decreased in HW-treated c-kit+ cells, indicating that HW can alleviate TBI-induced DNA damage and oxidative DNA damage in c-kit+ cells. Finally, the cell cycle (P21), cell apoptosis (BCL-XL and BAK), and oxidative stress (NRF2, HO-1, NQO1, SOD, and GPX1) proteins were significantly altered by HW in irradiated mouse c-kit+ cells. Collectively, the present results suggest that HW protects against TBI-induced HSC injury. PMID:28243358

Hydrogen-Rich Water Ameliorates Total Body Irradiation-Induced Hematopoietic Stem Cell Injury by Reducing Hydroxyl Radical.

PubMed

Zhang, Junling; Xue, Xiaolei; Han, Xiaodan; Li, Yuan; Lu, Lu; Li, Deguan; Fan, Saijun

2017-01-01

We examined whether consumption of hydrogen-rich water (HW) could ameliorate hematopoietic stem cell (HSC) injury in mice with total body irradiation (TBI). The results indicated that HW alleviated TBI-induced HSC injury with respect to cell number alteration and to the self-renewal and differentiation of HSCs. HW specifically decreased hydroxyl radical ( ∙ OH) levels in the c-kit + cells of 4 Gy irradiated mice. Proliferative bone marrow cells (BMCs) increased and apoptotic c-kit + cells decreased in irradiated mice uptaken with HW. In addition, the mean fluorescence intensity (MFI) of γ -H2AX and percentage of 8-oxoguanine positive cells significantly decreased in HW-treated c-kit + cells, indicating that HW can alleviate TBI-induced DNA damage and oxidative DNA damage in c-kit + cells. Finally, the cell cycle (P21), cell apoptosis (BCL-XL and BAK), and oxidative stress (NRF2, HO-1, NQO1, SOD, and GPX1) proteins were significantly altered by HW in irradiated mouse c-kit + cells. Collectively, the present results suggest that HW protects against TBI-induced HSC injury.

Usefulness of Daily Fractionated Administration of Wortmannin Combined With γ-Ray Irradiation in Terms of Local Tumor Response and Lung Metastasis

PubMed Central

Masunaga, Shin-ichiro; Sakurai, Yoshinori; Tanaka, Hiroki; Suzuki, Minoru; Kondo, Natsuko; Narabayashi, Masaru; Tano, Keizo; Maruhashi, Akira; Ono, Koji

2013-01-01

Background To evaluate the usefulness of fractionated administration of wortmannin combined with γ-ray irradiation in terms of local tumor response and lung metastatic potential, referring to the response of intratumor quiescent (Q) cells. Methods B16-BL6 melanoma tumor-bearing C57BL/6 mice were continuously given 5-bromo-2’-deoxyuridine (BrdU) to label all proliferating (P) cells. The tumor-bearing mice then received γ-ray irradiation after wortmannin treatment through a single or 4 consecutive daily intraperitoneal administrations up to a total dose of 4 mg/kg in combination with an acute hypoxia-releasing agent (nicotinamide) or mild temperature hyperthermia (MTH). Immediately after the irradiation, cells from some tumors were isolated and incubated with a cytokinesis blocker. The responses of the Q and total (= P + Q) cell populations were assessed based on the frequency of micronuclei using immunofluorescence staining for BrdU. In other tumor-bearing mice, 17 days after irradiation, macroscopic lung metastases were enumerated. Results Wortmannin raised the sensitivity of Q cells more remarkably than the total cell population in both single and daily administrations. Daily administration of wortmannin elevated the sensitivity of both the total and Q cell populations, but especially the total cell population, compared with single administration. Daily administration, especially combined with MTH, decreased the number of lung metastases. Conclusion Daily fractionated administration of wortmannin in combination with γ-ray irradiation was thought to be more promising than single administration because of its potential to enhance local tumor response and repress lung metastatic potential. PMID:29147327

Dynamic Adhesion of Umbilical Cord Blood Endothelial Progenitor Cells under Laminar Shear Stress

PubMed Central

Angelos, Mathew G.; Brown, Melissa A.; Satterwhite, Lisa L.; Levering, Vrad W.; Shaked, Natan T.; Truskey, George A.

2010-01-01

Late outgrowth endothelial progenitor cells (EPCs) represent a promising cell source for rapid reendothelialization of damaged vasculature after expansion ex vivo and injection into the bloodstream. We characterized the dynamic adhesion of umbilical-cord-blood-derived EPCs (CB-EPCs) to surfaces coated with fibronectin. CB-EPC solution density affected the number of adherent cells and larger cells preferentially adhered at lower cell densities. The number of adherent cells varied with shear stress, with the maximum number of adherent cells and the shear stress at maximum adhesion depending upon fluid viscosity. CB-EPCs underwent limited rolling, transiently tethering for short distances before firm arrest. Immediately before arrest, the instantaneous velocity decreased independent of shear stress. A dimensional analysis indicated that adhesion was a function of the net force on the cells, the ratio of cell diffusion to sliding speed, and molecular diffusivity. Adhesion was not limited by the settling rate and was highly specific to α5β1 integrin. Total internal reflection fluorescence microscopy showed that CB-EPCs produced multiple contacts of α5β1 with the surface and the contact area grew during the first 20 min of attachment. These results demonstrate that CB-EPC adhesion from blood can occur under physiological levels of shear stress. PMID:21112278

Number and size of nucleoli in the spermatocytes of chicken and Japanese quail.

PubMed

Andraszek, Katarzyna; Gryzińska, Magdalena; Knaga, Sebastian; Wójcik, Ewa; Smalec, Elzbieta

2012-01-01

Nucleoli are the product of nucleolus organizing region activity (NOR) of specific chromosomes. Their basic function is to synthetise ribosomal RNA precursors and promote the maturation and assemblage of preribosomal RNP molecules. Information on rRNA-coding gene activity can be provided by the analysis of the number and size of nucleoli in the prophase of the first meiotic division. The morphology and ultrastructure of a nucleolus depends, among others, on the species and cell growth cycle as well as the physiological and pathological state of an organism. The purpose of this research was to determine the number and size of nucleoli in the spermatocytes of the domestic chicken and the Japanese quail. Diverse numbers and sizes of nucleoli in the cells of the analysed birds were observed. 1-4 nucleoli were identified in chicken cells (1.91 +/- 0.63 on average) and 1-2 in quail cells (1.13 +/- 0.33 on average). For the total of 957 nucleoli observed in Gallus cells, 329 were classified as large and 628 as small. In Coturnix cells, 563 nucleoli were identified (66 large and 497 small ones). An analysis of the numbers and sizes of nucleoli can be performed at the cytogenetic level and serve as an alternative source of information on rRNA encoding gene and nucleolus organising region (NOR) activities.

Application of advanced cytometric and molecular technologies to minimal residual disease monitoring

NASA Astrophysics Data System (ADS)

Leary, James F.; He, Feng; Reece, Lisa M.

2000-04-01

Minimal residual disease monitoring presents a number of theoretical and practical challenges. Recently it has been possible to meet some of these challenges by combining a number of new advanced biotechnologies. To monitor the number of residual tumor cells requires complex cocktails of molecular probes that collectively provide sensitivities of detection on the order of one residual tumor cell per million total cells. Ultra-high-speed, multi parameter flow cytometry is capable of analyzing cells at rates in excess of 100,000 cells/sec. Residual tumor selection marker cocktails can be optimized by use of receiver operating characteristic analysis. New data minimizing techniques when combined with multi variate statistical or neural network classifications of tumor cells can more accurately predict residual tumor cell frequencies. The combination of these techniques can, under at least some circumstances, detect frequencies of tumor cells as low as one cell in a million with an accuracy of over 98 percent correct classification. Detection of mutations in tumor suppressor genes requires insolation of these rare tumor cells and single-cell DNA sequencing. Rare residual tumor cells can be isolated at single cell level by high-resolution single-cell cell sorting. Molecular characterization of tumor suppressor gene mutations can be accomplished using a combination of single- cell polymerase chain reaction amplification of specific gene sequences followed by TA cloning techniques and DNA sequencing. Mutations as small as a single base pair in a tumor suppressor gene of a single sorted tumor cell have been detected using these methods. Using new amplification procedures and DNA micro arrays it should be possible to extend the capabilities shown in this paper to screening of multiple DNA mutations in tumor suppressor and other genes on small numbers of sorted metastatic tumor cells.

Molecular diagnosis of bacterial vaginosis: Does adjustment for total bacterial load or human cellular content improve diagnostic performance?

PubMed

Plummer, E L; Garland, S M; Bradshaw, C S; Law, M G; Vodstrcil, L A; Hocking, J S; Fairley, C K; Tabrizi, S N

2017-02-01

We investigated the utility of quantitative PCR assays for diagnosis of bacterial vaginosis and found that while the best model utilized bacterial copy number adjusted for total bacterial load (sensitivity=98%, specificity=93%, AUC=0.95[95%CI=0.93,0.97]), adjusting for total bacterial or human cell load did not consistently increase the diagnostic performance of the assays. Copyright © 2017 Elsevier B.V. All rights reserved.

The Search for True Numbers of Neurons and Glial Cells in the Human Brain: A Review of 150 Years of Cell Counting

PubMed Central

von Bartheld, Christopher S.; Bahney, Jami; Herculano-Houzel, Suzana

2016-01-01

For half a century, the human brain was believed to contain about 100 billion neurons and one trillion glial cells, with a glia:neuron ratio of 10:1. A new counting method, the isotropic fractionator, has challenged the notion that glia outnumber neurons and revived a question that was widely thought to have been resolved. The recently validated isotropic fractionator demonstrates a glia:neuron ratio of less than 1:1 and a total number of less than 100 billion glial cells in the human brain. A survey of original evidence shows that histological data always supported a 1:1 ratio of glia to neurons in the entire human brain, and a range of 40–130 billion glial cells. We review how the claim of one trillion glial cells originated, was perpetuated, and eventually refuted. We compile how numbers of neurons and glial cells in the adult human brain were reported and we examine the reasons for an erroneous consensus about the relative abundance of glial cells in human brains that persisted for half a century. Our review includes a brief history of cell counting in human brains, types of counting methods that were and are employed, ranges of previous estimates, and the current status of knowledge about the number of cells. We also discuss implications and consequences of the new insights into true numbers of glial cells in the human brain, and the promise and potential impact of the newly validated isotropic fractionator for reliable quantification of glia and neurons in neurological and psychiatric diseases. PMID:27187682

The search for true numbers of neurons and glial cells in the human brain: A review of 150 years of cell counting.

PubMed

von Bartheld, Christopher S; Bahney, Jami; Herculano-Houzel, Suzana

2016-12-15

For half a century, the human brain was believed to contain about 100 billion neurons and one trillion glial cells, with a glia:neuron ratio of 10:1. A new counting method, the isotropic fractionator, has challenged the notion that glia outnumber neurons and revived a question that was widely thought to have been resolved. The recently validated isotropic fractionator demonstrates a glia:neuron ratio of less than 1:1 and a total number of less than 100 billion glial cells in the human brain. A survey of original evidence shows that histological data always supported a 1:1 ratio of glia to neurons in the entire human brain, and a range of 40-130 billion glial cells. We review how the claim of one trillion glial cells originated, was perpetuated, and eventually refuted. We compile how numbers of neurons and glial cells in the adult human brain were reported and we examine the reasons for an erroneous consensus about the relative abundance of glial cells in human brains that persisted for half a century. Our review includes a brief history of cell counting in human brains, types of counting methods that were and are employed, ranges of previous estimates, and the current status of knowledge about the number of cells. We also discuss implications and consequences of the new insights into true numbers of glial cells in the human brain, and the promise and potential impact of the newly validated isotropic fractionator for reliable quantification of glia and neurons in neurological and psychiatric diseases. J. Comp. Neurol. 524:3865-3895, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Time-dependent cell disintegration kinetics in lung tumors after irradiation

NASA Astrophysics Data System (ADS)

Chvetsov, Alexei V.; Palta, Jatinder J.; Nagata, Yasushi

2008-05-01

We study the time-dependent disintegration kinetics of tumor cells that did not survive radiotherapy treatment. To evaluate the cell disintegration rate after irradiation, we studied the volume changes of solitary lung tumors after stereotactic radiotherapy. The analysis is performed using two approximations: (1) tumor volume is a linear function of the total cell number in the tumor and (2) the cell disintegration rate is governed by the exponential decay with constant risk, which is defined by the initial cell number and a half-life T1/2. The half-life T1/2 is determined using the least-squares fit to the clinical data on lung tumor size variation with time after stereotactic radiotherapy. We show that the tumor volume variation after stereotactic radiotherapy of solitary lung tumors can be approximated by an exponential function. A small constant component in the volume variation does not change with time; however, this component may be the residual irregular density due to radiation fibrosis and was, therefore, subtracted from the total volume variation in our computations. Using computerized fitting of the exponent function to the clinical data for selected patients, we have determined that the average half-life T1/2 of cell disintegration is 28.2 days for squamous cell carcinoma and 72.4 days for adenocarcinoma. This model is needed for simulating the tumor volume variation during radiotherapy, which may be important for time-dependent treatment planning of proton therapy that is sensitive to density variations.

Niche recycling through division-independent egress of hematopoietic stem cells

PubMed Central

Czechowicz, Agnieszka; Ooi, A.G. Lisa; Rossi, Derrick J.; Bryder, David; Weissman, Irving L.

2009-01-01

Hematopoietic stem cells (HSCs) are thought to reside in discrete niches through stable adhesion, yet previous studies have suggested that host HSCs can be replaced by transplanted donor HSCs, even in the absence of cytoreductive conditioning. To explain this apparent paradox, we calculated, through cell surface phenotyping and transplantation of unfractionated blood, that ∼1–5% of the total pool of HSCs enters into the circulation each day. Bromodeoxyuridine (BrdU) feeding experiments demonstrated that HSCs in the peripheral blood incorporate BrdU at the same rate as do HSCs in the bone marrow, suggesting that egress from the bone marrow to the blood can occur without cell division and can leave behind vacant HSC niches. Consistent with this, repetitive daily transplantations of small numbers of HSCs administered as new niches became available over the course of 7 d led to significantly higher levels of engraftment than did large, single-bolus transplantations of the same total number of HSCs. These data provide insight as to how HSC replacement can occur despite the residence of endogenous HSCs in niches, and suggest therapeutic interventions that capitalize upon physiological HSC egress. PMID:19887396

SU-D-BRB-06: Treating Glioblastoma Multiforme (GBM) as a Chronic Disease: Implication of Temporal-Spatial Dose Fractionation Optimization Including Cancer Stem Cell Dynamics

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

Yu, V; Nguyen, D; Pajonk, F

Purpose: To explore the feasibility of improving GBM treatment outcome with temporal-spatial dose optimization of an ordinary differential equation (ODE) that models the differentiation and distinct radiosensitivity between cancer stem cells (CSC) and differentiated cancer cells (DCC). Methods: The ODE was formulated into a non-convex optimization problem with the objective to minimize remaining total cancer cells 500 days from the onset of radiotherapy when the total cancer cell number was 3.5×10{sup 7}, while maintaining normal tissue biological effective dose (BED) of 100Gy resulted from standard prescription of 2Gyx30. Assuming spatially separated CSC and DCC, optimization was also performed to exploremore » the potential benefit from dose-painting the two compartments. Dose escalation to a sub-cell-population in the GTV was also examined assuming that a 2 cm margin around the GTV allows sufficient dose drop-off to 100Gy BED. The recurrence time was determined as the time at which the total cancer cell number regrows to 10{sup 9} cells. Results: The recurrence time with variable fractional doses administered once per week, bi-week and month for one year were found to be 615, 593 and 570 days, superior to the standard-prescription recurrence time of 418 days. The optimal dose-fraction size progression for both uniform and dose-painting to the tumor is low and relatively constant in the beginning and gradually increases to more aggressive fractions at end of the treatment course. Dose escalation to BED of 200Gy to the whole tumor alongside with protracted weekly treatment was found to further delay recurrence to 733 days. Dose-painting of 200 and 500Gy BED to CSC on a year-long weekly schedule further extended recurrence to 736 and 1076 days, respectively. Conclusion: GBM treatment outcome can possibly be improved with a chronic treatment approach. Further dose escalation to the entire tumor or CSC targeted killing is needed to achieve total tumor control. This work is supported by the NSF Graduate Research Fellowship (DGE-1144087)« less

Distinct abnormalities in the innate immune system of children with Down syndrome.

PubMed

Bloemers, Beatrijs L P; van Bleek, Grada M; Kimpen, Jan L L; Bont, Louis

2010-05-01

To analyze the frequency and phenotype of cells of the innate immune system in the peripheral blood of children with Down syndrome (DS). Flow cytometric analysis of expression of cell surface markers was performed in children with DS (n = 41) and healthy age-matched controls (n = 41). Compared with controls, children with DS had significantly lower absolute total leukocyte counts, lymphocytes, monocytes, and granulocytes, but 1.5-times higher absolute numbers of CD14(dim)CD16(+) monocytes (147 x 10(6)/L vs 93 x 10(6)/L; P = .02). This difference is fully explained by a higher percentage of CD14(dim)CD16(+) monocytes within the monocyte compartment (28.7% vs 13.4%; P <.001). The absolute numbers of myeloid dendritic cells were lower in DS (13.8 x 10(6)/L vs 22.7 x 10(6)/L; P <.001). The numbers of plasmacytoid dendritic cells and natural killer cells were normal. Absolute numbers of invariant natural killer T cells were very low overall, but significantly lower in children with DS than in controls (1.2 x 10(6)/L vs 3.7 x 10(6)/L; P = .01). Children with DS exhibited distinct abnormalities in cells of the innate immune system. Most strikingly, they had a high number of proinflammatory CD14(dim)CD16(+) monocytes. This elevated level of CD14(dim)CD16(+) monocytes may play an important role in the onset and maintenance of chronic inflammatory disease in DS.

A novel method to generate and culture human mast cells: Peripheral CD34+ stem cell-derived mast cells (PSCMCs).

PubMed

Schmetzer, Oliver; Valentin, Patricia; Smorodchenko, Anna; Domenis, Rossana; Gri, Giorgia; Siebenhaar, Frank; Metz, Martin; Maurer, Marcus

2014-11-01

The identification and characterization of human mast cell (MC) functions are hindered by the shortage of MC populations suitable for investigation. Here, we present a novel technique for generating large numbers of well differentiated and functional human MCs from peripheral stem cells (=peripheral stem cell-derived MCs, PSCMCs). Innovative and key features of this technique include 1) the use of stem cell concentrates, which are routinely discarded by blood banks, as the source of CD34+ stem cells, 2) cell culture in serum-free medium and 3) the addition of LDL as well as selected cytokines. In contrast to established and published protocols that use CD34+ or CD133+ progenitor cells from full blood, we used a pre-enriched cell population obtained from stem cell concentrates, which yielded up to 10(8) differentiated human MCs per batch after only three weeks of culture starting with 10(6) total CD34+ cells. The total purity on MCs (CD117+, FcεR1+) generated by this method varied between 55 and 90%, of which 4-20% were mature MCs that contain tryptase and chymase and show expression of FcεRI and CD117 in immunohistochemistry. PSCMCs showed robust histamine release in response to stimulation with anti-FcεR1 or IgE/anti-IgE, and increased proliferation and differentiation in response to IL-1β or IFN-γ. Taken together, this new protocol of the generation of large numbers of human MCs provides for an innovative and suitable option to investigate the biology of human MCs. Copyright © 2014 Elsevier B.V. All rights reserved.

Discrete virus infection model of hepatitis B virus.

PubMed

Zhang, Pengfei; Min, Lequan; Pian, Jianwei

2015-01-01

In 1996 Nowak and his colleagues proposed a differential equation virus infection model, which has been widely applied in the study for the dynamics of hepatitis B virus (HBV) infection. Biological dynamics may be described more practically by discrete events rather than continuous ones. Using discrete systems to describe biological dynamics should be reasonable. Based on one revised Nowak et al's virus infection model, this study introduces a discrete virus infection model (DVIM). Two equilibriums of this model, E1 and E2, represents infection free and infection persistent, respectively. Similar to the case of the basic virus infection model, this study deduces a basic virus reproductive number R0 independing on the number of total cells of an infected target organ. A proposed theorem proves that if the basic virus reproductive number R0<1 then the virus free equilibrium E1 is locally stable. The DVIM is more reasonable than an abstract discrete susceptible-infected-recovered model (SIRS) whose basic virus reproductive number R0 is relevant to the number of total cells of the infected target organ. As an application, this study models the clinic HBV DNA data of a patient who was accepted via anti-HBV infection therapy with drug lamivudine. The results show that the numerical simulation is good in agreement with the clinic data.

Longitudinal analysis of Prototheca zopfii-specific immune responses: correlation with disease progression and carriage in dairy cows.

PubMed

Roesler, Uwe; Hensel, Andreas

2003-03-01

In order to characterize the humoral and cellular immune responses to bovine mammary protothecosis, serum and whey samples obtained from 72 dairy cows assigned to four different clinical stages of infection were examined for specific antibodies by indirect enzyme-linked immunosorbent assay techniques. Milk samples were analyzed for the total numbers of excreted algal cells and somatic cells. After characterization of the course of immune induction in bovine protothecal mastitis, a long-term sentinel study was performed in an affected herd in order to investigate disease progression. A total of 61 dairy cows with protothecal mastitis were examined for shedding of algae cells and for local immune responses three times in 6-month intervals. During acute and chronic stages of protothecosis, significantly elevated specific antibody activities in sera were detected. A strong correlation of whey immunoglobulin A (IgA) and whey IgG1 antibody activity with the total counts of somatic cells in milk was observed, whereas only a weak correlation of whey IgA and whey IgG1 concentrations to the number of algal cells excreted with the milk was seen. Our results from the sentinel long-term study of infected cows revealed that 70.5% of the persistently infected animals were continuously shedding the pathogen. About 4.9% of the animals showed an intermittent shedding, whereas 18% of the cows were tested culturally negative throughout the study. It can be assumed that Prototheca zopfii mastitis in dairy cows is maintained on the herd level by subclinically infected alga-shedding cows.

Longitudinal Analysis of Prototheca zopfii-Specific Immune Responses: Correlation with Disease Progression and Carriage in Dairy Cows

PubMed Central

Roesler, Uwe; Hensel, Andreas

2003-01-01

In order to characterize the humoral and cellular immune responses to bovine mammary protothecosis, serum and whey samples obtained from 72 dairy cows assigned to four different clinical stages of infection were examined for specific antibodies by indirect enzyme-linked immunosorbent assay techniques. Milk samples were analyzed for the total numbers of excreted algal cells and somatic cells. After characterization of the course of immune induction in bovine protothecal mastitis, a long-term sentinel study was performed in an affected herd in order to investigate disease progression. A total of 61 dairy cows with protothecal mastitis were examined for shedding of algae cells and for local immune responses three times in 6-month intervals. During acute and chronic stages of protothecosis, significantly elevated specific antibody activities in sera were detected. A strong correlation of whey immunoglobulin A (IgA) and whey IgG1 antibody activity with the total counts of somatic cells in milk was observed, whereas only a weak correlation of whey IgA and whey IgG1 concentrations to the number of algal cells excreted with the milk was seen. Our results from the sentinel long-term study of infected cows revealed that 70.5% of the persistently infected animals were continuously shedding the pathogen. About 4.9% of the animals showed an intermittent shedding, whereas 18% of the cows were tested culturally negative throughout the study. It can be assumed that Prototheca zopfii mastitis in dairy cows is maintained on the herd level by subclinically infected alga-shedding cows. PMID:12624049

Vaccine draining lymph nodes are a source of antigen-specific B cells.

PubMed

Pero, Stephanie C; Sun, Yu-Jing; Shukla, Girja S; Carman, Chelsea L; Krag, Christopher C; Teuscher, Cory; Krementsov, Dimitry N; Krag, David N

2017-03-01

Our research is focused on using vaccine draining lymph nodes as a source of immune cells to better understand the immune response and to attempt to generate new anti-cancer reagents. Following a vaccine, harvesting the lymph node can only be done once. We endeavored to determine the range of times that B cells secreting anti-KLH antibodies were present in the node of KLH-vaccinated mice. Following vaccination the total number of mononuclear cells (MNCs) increased in the vaccine-draining lymph node (VDN). The percentage of MNCs that were B cells nearly doubled. B cells recovered from the node that secreted anti-KLH antibodies were evident by day 7. The number continued to increase and then slowly decreased over the observed time range to 28days after vaccination. The VDN, compared to the spleen, the bone marrow and the nonVDN, contained a higher percentage of B cells that secreted anti-KLH antibodies. After a vaccine, there is a multi-week window of time when an increasing number of B cells are present in a VDN that secrete anti-KLH antibodies. These results support using the VDN as a source for B cells that secrete anti-vaccine antibodies. Copyright © 2017 Elsevier Ltd. All rights reserved.

Decreased frequency and function of circulating plasmocytoid dendritic cells (pDC) in hepatitis B virus infected humans.

PubMed

Duan, Xue-Zhang; Wang, Min; Li, Han-Wei; Zhuang, Hui; Xu, Dongping; Wang, Fu-Sheng

2004-11-01

The Type 2 precursor plasmacytoid dendritic cells (pDC) represent the most important cell type in antiviral innate immunity. To understand the function of pDC during hepatitis B virus infection, the frequency and function of circulating pDC were analyzed by flow cytometric analysis, and IFN-alpha secretion of total PBMCs was determined by ELISA assay in 25 healthy subjects and 116 patients at various stages of chronic hepatitis B virus infection (CHB). The number of circulating pDC was found to be significantly lower in patients with CHB and associated liver cirrhosis (LC). The ability of PBMCs to secrete IFN-alpha also decreased significantly. There was a corresponding decrease of circulating NK cells and CD8+ T cells. We observed that lamuvidine antiviral therapy restored the number of circulating pDC and there was a reversal of pDC frequency with the control of HBV replication in chronic HBV patients, indicating these subjects are unlikely to be totally immunocompromised. The decrease of pDC was found to be related to nosocomial infections in LC patients. Our results suggest that CHB patients probably have a quantitative and qualitative impairment of circulating pDC or NK cells, which may be associated with HBV persistent infection as well as the nosocomial infections that arise in LC patients.

Effects of perfluorinated chemicals on adipocyte development ...

EPA Pesticide Factsheets

Obesity is a growing concern in the US population. Current interest is high in the role played by environmental factors called obesogens that may contribute to obesity through developmental exposure. One class of potential obesogens is the family of perfluorinated chemicals used as surfactants in a variety of industrial applications. Given the importance of understanding the role these compounds play in lipid homeostasis we used pre-adipocyte 3T3-L1 mouse fibroblast cells (Zen-Bio, RTP NC) to study their effects on adipogenesis and lipid accumulation. These cells differentiate into adipocytes accumulating large lipid droplets. Cultures were treated with perfluorooctanoic acid (PFOA) (1-200uM), perfluorononanoic acid (PFNA) (5-lOOuM), perfluorooctane sulfonate (PFOS) (5O-300uM), and perfluorohexane sulfonate (PFHxS) (40- 250uM). Cell size number, and lipid content were assessed using morphomeiric analysis. All four compounds decreased cell size compared to control, and PFNA was most potent, in terms of lowest observed effect concentration (LOEC), whereas PFOA was least potent. Cell number increased for all perfluorinated chemicals tested, most potently for PFNA and least for PFOS. Interestingly, average lipid area per cell for all four chemicals decreased compared to control, but PFOS and PFHxS had increased total lipid area. Additionally, significant increases in total triglyceride were noted for all compounds compared to controls. PFOA and PFNA increased trigly

Hallmarks in prostate cancer imaging with Ga68-PSMA-11-PET/CT with reference to detection limits and quantitative properties.

PubMed

Sanchez-Crespo, Alejandro; Jussing, Emma; Björklund, Ann-Charlotte; Pokrovskaja Tamm, Katja

2018-04-04

Gallium-68-labeled prostate-specific antigen positron emission tomography/computed tomography imaging (Ga68-PSMA-11-PET/CT) has emerged as a potential gold standard for prostate cancer (PCa) diagnosis. However, the imaging limitations of this technique at the early state of PCa recurrence/metastatic spread are still not well characterized. The aim of this study was to determine the quantitative properties and the fundamental imaging limits of Ga68-PSMA-11-PET/CT in localizing small PCa cell deposits. The human PCa LNCaP cells (PSMA expressing) were grown and collected as single cell suspension or as 3D-spheroids at different cell numbers and incubated with Ga68-PSMA-11. Thereafter, human HCT116 cells (PSMA negative) were added to a total cell number of 2 × 10 5 cells per tube. The tubes were then pelleted and the supernatant aspirated. A whole-body PET/CT scanner with a clinical routine protocol was used for imaging the pellets inside of a cylindrical water phantom with increasing amounts of background activity. The actual activity bound to the cells was also measured in an automatic gamma counter. Imaging detection limits and activity recovery coefficients as a function of LNCaP cell number were obtained. The effect of Ga68-PSMA-11 mass concentration on cell binding was also investigated in samples of LnCaP cells incubated with increasing concentrations of radioligand. A total of 1 × 10 4 LNCaP cells mixed in a pellet of 2 × 10 5 cells were required to reach a 50% detection probability with Ga68-PSMA-11-PET/CT without background. With a background level of 1 kBq/ml, between 4 × 10 5 and 1 × 10 6 cells are required. The radioligand equilibrium dissociation constant was 27.05 nM, indicating high binding affinity. Hence, the specific activity of the radioligand has a profound effect on image quantification. Ga68-PSMA-11-PET detects a small number of LNCaP cells even when they are mixed in a population of non-PSMA expressing cells and in the presence of background. The obtained image detection limits and characteristic quantification properties of Ga68-PSMA-11-PET/CT are essential hallmarks for the individualization of patient management. The use of the standardized uptake value for Ga68-PSMA-11-PET/CT image quantification should be precluded.

Improved Murine Blastocyst Quality and Development in a Single Culture Medium Compared to Sequential Culture Media

PubMed Central

Hennings, Justin M.; Zimmer, Randall L.; Nabli, Henda; Davis, J. Wade; Sutovsky, Peter; Sutovsky, Miriam; Sharpe-Timms, Kathy L.

2015-01-01

Objective: Validate single versus sequential culture media for murine embryo development. Design: Prospective laboratory experiment. Setting: Assisted Reproduction Laboratory. Animals: Murine embryos. Interventions: Thawed murine zygotes cultured for 3 or 5 days (d3 or d5) in single or sequential embryo culture media developed for human in vitro fertilization. Main Outcome Measures: On d3, zygotes developing to the 8 cell (8C) stage or greater were quantified using 4’,6-diamidino-2-phenylindole (DAPI), and quality was assessed by morphological analysis. On d5, the number of embryos reaching the blastocyst stage was counted. DAPI was used to quantify total nuclei and inner cell mass nuclei. Localization of ubiquitin C-terminal hydrolase L1 (UCHL1) and ubiquitin C-terminal hydrolase L3 (UCHL3) was reference points for evaluating cell quality. Results: Comparing outcomes in single versus to sequential media, the odds of embryos developing to the 8C stage on d3 were 2.34 time greater (P = .06). On d5, more embryos reached the blastocyst stage (P = <.0001), hatched, and had significantly more trophoblast cells (P = .005) contributing to the increased total cell number. Also at d5, localization of distinct cytoplasmic UCHL1 and nuclear UCHL3 was found in high-quality hatching blastocysts. Localization of UCHL1 and UCHL3 was diffuse and inappropriately dispersed throughout the cytoplasm in low-quality nonhatching blastocysts. Conclusions: Single medium yields greater cell numbers, an increased growth rate, and more hatching of murine embryos. Cytoplasmic UCHL1 and nuclear UHCL3 localization patterns were indicative of embryo quality. Our conclusions are limited to murine embryos but one might speculate that single medium may also be more beneficial for human embryo culture. Human embryo studies are needed. PMID:26668049

Improved Murine Blastocyst Quality and Development in a Single Culture Medium Compared to Sequential Culture Media.

PubMed

Hennings, Justin M; Zimmer, Randall L; Nabli, Henda; Davis, J Wade; Sutovsky, Peter; Sutovsky, Miriam; Sharpe-Timms, Kathy L

2016-03-01

Validate single versus sequential culture media for murine embryo development. Prospective laboratory experiment. Assisted Reproduction Laboratory. Murine embryos. Thawed murine zygotes cultured for 3 or 5 days (d3 or d5) in single or sequential embryo culture media developed for human in vitro fertilization. On d3, zygotes developing to the 8 cell (8C) stage or greater were quantified using 4',6-diamidino-2-phenylindole (DAPI), and quality was assessed by morphological analysis. On d5, the number of embryos reaching the blastocyst stage was counted. DAPI was used to quantify total nuclei and inner cell mass nuclei. Localization of ubiquitin C-terminal hydrolase L1 (UCHL1) and ubiquitin C-terminal hydrolase L3 (UCHL3) was reference points for evaluating cell quality. Comparing outcomes in single versus to sequential media, the odds of embryos developing to the 8C stage on d3 were 2.34 time greater (P = .06). On d5, more embryos reached the blastocyst stage (P = <.0001), hatched, and had significantly more trophoblast cells (P = .005) contributing to the increased total cell number. Also at d5, localization of distinct cytoplasmic UCHL1 and nuclear UCHL3 was found in high-quality hatching blastocysts. Localization of UCHL1 and UCHL3 was diffuse and inappropriately dispersed throughout the cytoplasm in low-quality nonhatching blastocysts. Single medium yields greater cell numbers, an increased growth rate, and more hatching of murine embryos. Cytoplasmic UCHL1 and nuclear UHCL3 localization patterns were indicative of embryo quality. Our conclusions are limited to murine embryos but one might speculate that single medium may also be more beneficial for human embryo culture. Human embryo studies are needed. © The Author(s) 2015.

EFFECT OF THAI SARAPHI FLOWER EXTRACTS ON WT1 AND BCR/ABL PROTEIN EXPRESSION IN LEUKEMIC CELL LINES.

PubMed

Sangkaruk, Rungkarn; Rungrojsakul, Methee; Tima, Singkome; Anuchapreeda, Songyot

2017-01-01

Saraphi (Mammea siamensis) is a Thai traditional herb. In this study, the cytotoxic effects of crude ethanolic and fractional extracts including hexane, ethyl acetate, and methanol fractions from M. siamensis flowers were investigated in order to determine their effect on WT1 expression in Molt4 and K562 cells and Bcr/Abl expression in K562 cells. The flowers of M. siamensis were extracted using ethanol. The ethanol flower extract was further fractionated with hexane, ethyl acetate, and methanol. Cytotoxic effects were measured by the MTT assay. Bcr/Abl and WT1 protein levels after treatments were determined by Western blotting. The total cell number was determined via the typan blue exclusion method. The hexane fraction showed the strongest cytotoxic activity on Molt4 and K562 cells, with IC 50 values of 2.6 and 77.6 μg/ml, respectively. The hexane extract decreased Bcr/Abl protein expression in K562 cells by 74.6% and WT1 protein expressions in Molt4 and K562 cells by 68.4 and 72.1%, respectively. Total cell numbers were decreased by 66.2 and 48.7% in Molt4 and K562 cells, respectively. Mammea E/BB (main active compound) significantly decreased both Bcr/Abl and WTlprotein expressions by 75 and 49.5%, respectively when compared to vehicle control. The hexane fraction from M. siamensis flowers inhibited cell proliferation via the suppression of WT1 expression in Molt4 and K562 cells and Bcr/Abl expression in K562 cells. The active compound may be mammea E/BB. Extracts from M. siamensis flowers show promise as naturally occurring anti-cancer drugs.

Escherichia coli growth under modeled reduced gravity

NASA Technical Reports Server (NTRS)

Baker, Paul W.; Meyer, Michelle L.; Leff, Laura G.

2004-01-01

Bacteria exhibit varying responses to modeled reduced gravity that can be simulated by clino-rotation. When Escherichia coli was subjected to different rotation speeds during clino-rotation, significant differences between modeled reduced gravity and normal gravity controls were observed only at higher speeds (30-50 rpm). There was no apparent affect of removing samples on the results obtained. When E. coli was grown in minimal medium (at 40 rpm), cell size was not affected by modeled reduced gravity and there were few differences in cell numbers. However, in higher nutrient conditions (i.e., dilute nutrient broth), total cell numbers were higher and cells were smaller under reduced gravity compared to normal gravity controls. Overall, the responses to modeled reduced gravity varied with nutrient conditions; larger surface to volume ratios may help compensate for the zone of nutrient depletion around the cells under modeled reduced gravity.

Mast cell inhibition as a therapeutic approach in fibrodysplasia ossificans progressiva (FOP).

PubMed

Brennan, Tracy A; Lindborg, Carter M; Bergbauer, Christian R; Wang, Haitao; Kaplan, Frederick S; Pignolo, Robert J

2018-04-01

Episodic flare-ups of fibrodysplasia ossificans progressiva (FOP) are characterized clinically by severe, often posttraumatic, connective tissue swelling and intramuscular edema, followed histologically by an intense and highly angiogenic fibroproliferative reaction. This early inflammatory and angiogenic fibroproliferative response is accompanied by the presence of abundant mast cells far in excess of other reported myopathies. Using an injury-induced, constitutively-active transgenic mouse model of FOP we show that mast cell inhibition by cromolyn, but not aprepitant, results in a dramatic reduction of heterotopic ossification. Cromolyn, but not aprepitant, significantly decreases the total number of mast cells in FOP lesions. Furthermore, cromolyn specifically diminishes the number of degranulating and resting degranulated mast cells in pre-osseous lesions. This work demonstrates that consideration of FOP as a type of localized mastocytosis may offer new therapeutic interventions for treatment of this devastating condition. Copyright © 2017 Elsevier Inc. All rights reserved.

Heritability of semen traits in German Warmblood stallions.

PubMed

Gottschalk, M; Sieme, H; Martinsson, G; Distl, O

2016-07-01

The objectives of the present study were to evaluate genetic parameters for semen quality traits of 241 fertile German Warmblood stallions regularly employed in artificial insemination (AI). Stallions were owned by the National Studs Celle and Warendorf in Germany. Semen traits analyzed were gel-free volume, sperm concentration, total number of sperm, progressive motility and total number of progressively motile sperm. Semen protocols from a total of 63,972 ejaculates were collected between the years 2001 and 2014 for the present analysis. A multivariate linear animal model was employed for estimation of additive genetic and permanent environmental variances among stallions and breeding values (EBVs) for semen traits. Heritabilities estimated for all German Warmblood stallions were highest for gel-free volume (h(2)=0.28) and lowest for total number of progressively motile sperm (h(2)=0.13). The additive genetic correlation among gel-free volume and sperm concentration was highly negative (rg=-0.76). Average reliabilities of EBVs were at 0.37-0.68 for the 241 stallions with own records. The inter-stallion variance explained between 33 and 61% of the trait variance, underlining the major impact of the individual stallion on semen quality traits analyzed here. Recording of semen traits from stallions employed in AI may be recommended because EBVs achieve sufficient accuracies to improve semen quality in future generations. Due to favorable genetic correlations, sperm concentration, total number of sperm and total number of progressively motile sperm may be increased simultaneously. Copyright © 2016 Elsevier B.V. All rights reserved.

Increased HIV-1 transcriptional activity and infectious burden in peripheral blood and gut-associated CD4+ T cells expressing CD30

PubMed Central

Carvidi, Alexander B.; Smith, Louis C. B.; Khan, Shahzada; Trapecar, Martin; Stoddart, Cheryl A.; Kuritzkes, Daniel R.

2018-01-01

HIV-1-infected cells persist indefinitely despite the use of combination antiretroviral therapy (ART), and novel therapeutic strategies to target and purge residual infected cells in individuals on ART are urgently needed. Here, we demonstrate that CD4+ T cell-associated HIV-1 RNA is often highly enriched in cells expressing CD30, and that cells expressing this marker considerably contribute to the total pool of transcriptionally active CD4+ lymphocytes in individuals on suppressive ART. Using in situ RNA hybridization studies, we show co-localization of CD30 with HIV-1 transcriptional activity in gut-associated lymphoid tissues. We also demonstrate that ex vivo treatment with brentuximab vedotin, an antibody-drug conjugate (ADC) that targets CD30, significantly reduces the total amount of HIV-1 DNA in peripheral blood mononuclear cells obtained from infected, ART-suppressed individuals. Finally, we observed that an HIV-1-infected individual, who received repeated brentuximab vedotin infusions for lymphoma, had no detectable virus in peripheral blood mononuclear cells. Overall, CD30 may be a marker of residual, transcriptionally active HIV-1 infected cells in the setting of suppressive ART. Given that CD30 is only expressed on a small number of total mononuclear cells, it is a potential therapeutic target of persistent HIV-1 infection. PMID:29470552

Side population cells in the human vocal fold.

PubMed

Yamashita, Masaru; Hirano, Shigeru; Kanemaru, Shin-ichi; Tsuji, Shunichiro; Suehiro, Atsushi; Ito, Juichi

2007-11-01

The regenerative processes of the vocal fold, or the existence of stem cells in the folds, are unknown. Side population (SP) cells are defined as cells that have the ability to exclude the DNA binding dye, Hoechst 33342. They are regarded as a cell population enriched with stem cells and can be isolated from non-SP cells by a fluorescence-activated cell sorter. This study was designed to determine whether SP cells exist in the human vocal fold, as a first step in elucidating the regenerative mechanisms of the vocal fold. Seven human excised larynges were used in this study. Two were used for fluorescence-activated cell sorter analysis, and 5 were subjected to immunohistochemical analysis with antibodies against an adenosine triphosphate binding cassette transporter family member, ABCG2, which is expressed in SP cells. The number of SP cells in the human vocal fold was about 0.2% of the total number of cells. ABCG2-positive cells were identified in both the epithelium and subepithelial tissue throughout the entire vocal fold. This preliminary study demonstrated the existence of SP cells in the human vocal fold. Further studies are warranted to clarify how these cells work in the vocal fold, particularly in the regenerative process.

The Protective Effects of Astaxanthin on the OVA-Induced Asthma Mice Model.

PubMed

Hwang, Yun-Ho; Hong, Seong-Gyeol; Mun, Seul-Ki; Kim, Su-Jin; Lee, Sung-Ju; Kim, Jong-Jin; Kang, Kyung-Yun; Yee, Sung-Tae

2017-11-21

Although astaxanthin has a variety of biological activities such as anti-oxidant effects, inhibitory effects on skin deterioration and anti-inflammatory effects, its effect on asthma has not been studied. In this paper, the inhibitory effect of astaxanthin on airway inflammation in a mouse model of ovalbumin (OVA)-induced asthma was investigated. We evaluated the number of total cells, Th1/2 mediated inflammatory cytokines in bronchoalveolar lavage fluid (BALF) and airway hyperresponsiveness as well as histological structure. The level of total IgE, IgG1, IgG2a, OVA-specific IgG1, and OVA-specific IgG2a were also examined. The oral administration of 50 mg/mL astaxanthin inhibited the respiratory system resistance, elastance, newtonian resistance, tissue damping, and tissue elastance. Also, astaxanthin suppressed the total cell number, IL-4, and IL-5, and increased the IFN-γ in the BALF. In the sera, total IgE, IgG1, and OVA-specific IgG1 were reduced by astaxanthin exposure and IgG2a and OVA-specific IgG2a were enhanced via oral administration of astaxanthin. Infiltration of inflammatory cells in the lung, production of mucus, lung fibrosis, and expression of caspase-1 or caspase-3 were suppressed in OVA-induced asthmatic animal treated with astaxanthin. These results suggest that astaxanthin may have therapeutic potential for treating asthma via inhibiting Th2-mediated cytokine and enhancing Th1-mediated cytokine.

Differential effects of Cytomegalovirus carriage on the immune phenotype of middle-aged males and females

PubMed Central

van der Heiden, Marieke; van Zelm, Menno C.; Bartol, Sophinus J. W.; de Rond, Lia G. H.; Berbers, Guy A. M.; Boots, Annemieke M. H.; Buisman, Anne-Marie

2016-01-01

The elderly population is more susceptible to infections as a result of an altered immune response, commonly referred to as immunosenescence. Cytomegalovirus (CMV)-infection associated changes in blood lymphocytes are known to impact this process, but the interaction with gender remains unclear. Therefore, we analysed the effects and interaction of gender and CMV on the absolute numbers of a comprehensive set of naive and memory T- and B-cell subsets in people between 50 and 65 years of age. Enumeration and characterisation of lymphocyte subsets by flow cytometry was performed on fresh whole blood samples from 255 middle-aged persons. CMV-IgG serostatus was determined by ELISA. Gender was a major factor affecting immune cell numbers. CMV infection was mainly associated with an expansion of late-differentiated T-cell subsets. CMV+ males carried lower numbers of total CD4+, CD4+ central memory (CM) and follicular helper T-cells than females and CMV− males. Moreover, CMV+ males had significantly lower numbers of regulatory T (Treg)-cells and memory B-cells than CMV+ females. We here demonstrate an interaction between the effects of CMV infection and gender on T- and B-cells in middle-aged individuals. These differential effects on adaptive immunity between males and females may have implications for vaccination strategies at middle-age. PMID:27243552

Analysis of Cell Turnover in the Bronchiolar Epithelium Through the Normal Aging Process.

PubMed

Ortega-Martínez, Marta; Rodríguez-Flores, Laura E; Ancer-Arellano, Adriana; Cerda-Flores, Ricardo M; de-la-Garza-González, Carlos; Ancer-Rodríguez, Jesús; Jaramillo-Rangel, Gilberto

2016-08-01

Aging is associated with changes in the lung that leads to a decrease in its function. Alterations in structure and function in the small airways are well recognized in chronic lung diseases. The aim of this study was the assessment of cell turnover in the bronchiolar epithelium of mouse through the normal aging process. Lungs from CD1 mice at the age of 2, 6, 12, 18, or 24 months were fixed in neutral-buffered formalin and paraffin-embedded. Proliferating cell nuclear antigen was examined by immunohistochemistry. Apoptosis was analyzed by in situ end-labeling of fragmented DNA. Epithelial dimensions were analyzed by morphometry. The 2-month-old mice showed significantly higher number of proliferating cells when compared with mice at all other age groups. The number of apoptotic cells in mice at 24 months of age was significantly greater than in mice at all other age groups. Thus, the number of epithelial cells decreased as the age of the subject increased. We also found reductions in both area and height of the bronchiolar epithelium in mice at 18 and 24 months of age. We found a decrease in the total number of epithelial cells in the aged mice, which was accompanied by a thinning of the epithelium. These changes reflect a dysregulated tissue regeneration process in the bronchiolar epithelium that might predispose to respiratory diseases in elderly subjects.

Culturability as an indicator of succession in microbial communities

NASA Technical Reports Server (NTRS)

Garland, J. L.; Cook, K. L.; Adams, J. L.; Kerkhof, L.

2001-01-01

Successional theory predicts that opportunistic species with high investment of energy in reproduction and wide niche width will be replaced by equilibrium species with relatively higher investment of energy in maintenance and narrower niche width as communities develop. Since the ability to rapidly grow into a detectable colony on nonselective agar medium could be considered as characteristic of opportunistic types of bacteria, the percentage of culturable cells may be an indicator of successional state in microbial communities. The ratios of culturable cells (colony forming units on R2A agar) to total cells (acridine orange direct microscopic counts) and culturable cells to active cells (reduction of 5-cyano-2,3-ditolyl tetrazolium chloride) were measured over time in two types of laboratory microcosms (the rhizosphere of hydroponically grown wheat and aerobic, continuously stirred tank reactors containing plant biomass) to determine the effectiveness of culturabilty as an index of successional state. The culturable cell:total cell ratio in the rhizosphere decreased from approximately 0.25 to less than 0.05 during the first 30-50 days of plant growth, and from 0.65 to 0.14 during the first 7 days of operation of the bioreactor. The culturable cell:active cell ratio followed similar trends, but the values were consistently greater than the culturable cell:total cell ratio, and even exceeded I in early samples. Follow-up studies used a cultivation-independent method, terminal restriction fragment length polymorphisms (TRFLP) from whole community DNA, to assess community structure. The number of TRFLP peaks increased with time, while the number of culturable types did not, indicating that the general decrease in culturability is associated with a shift in community structure. The ratio of respired to assimilated C-14-labeled amino acids increased with the age of rhizosphere communities, supporting the hypothesis that a shift in resource allocation from growth to maintenance occurs with time. Results from this work indicate that the percentage of culturable cells may be a useful method for assessing the successional state of microbial communities.

Morphometric analysis of the neuronal numbers and densities of the inferior olivary complex in the donkey (Equus asinus).

PubMed

Alkafafy, Mohamed; Rashed, Reda; Attia, Hossam

2011-07-01

The morphometric interrelations between the compartments of the inferior olivary complex (IOC) in the donkey (Equus asinus) were ascertained by examining serial sections throughout the entire length of the IOC for both sides. Nissl-stained celloidin sections of four brainstems of donkeys were used. The IOC consisted of three major nuclei and four small cell groups. The total neuronal count in both sides of the IOC was 202,040±8480 cells. The medial accessory olivary nucleus (MAO) had the largest relative area (46%) and the highest number of neurons (90,800±7600). The dorsal accessory olivary nucleus (DAO) had the second largest relative area (33%), while the principal olivary nucleus (PO) had the lowest relative area (21%). However, the total neuron count in the PO was larger (60,840±1840) than DAO (50,360±4040). The average neuronal density was 2700±400 cells/mm(3). The numerical values of the current study of the IOC in the donkey were similar to those of other mammals. Copyright © 2010 Elsevier GmbH. All rights reserved.

Kaposi’s sarcoma–associated herpesvirus stably clusters its genomes across generations to maintain itself extrachromosomally

PubMed Central

Chiu, Ya-Fang; Sugden, Arthur U.

2017-01-01

Genetic elements that replicate extrachromosomally are rare in mammals; however, several human tumor viruses, including the papillomaviruses and the gammaherpesviruses, maintain their plasmid genomes by tethering them to cellular chromosomes. We have uncovered an unprecedented mechanism of viral replication: Kaposi’s sarcoma–associated herpesvirus (KSHV) stably clusters its genomes across generations to maintain itself extrachromosomally. To identify and characterize this mechanism, we developed two complementary, independent approaches: live-cell imaging and a predictive computational model. The clustering of KSHV requires the viral protein, LANA1, to bind viral genomes to nucleosomes arrayed on both cellular and viral DNA. Clustering affects both viral partitioning and viral genome numbers of KSHV. The clustering of KSHV plasmids provides it with an effective evolutionary strategy to rapidly increase copy numbers of genomes per cell at the expense of the total numbers of cells infected. PMID:28696226

Kaposi’s sarcoma–associated herpesvirus stably clusters its genomes across generations to maintain itself extrachromosomally

DOE PAGES

Chiu, Ya-Fang; Sugden, Arthur U.; Fox, Kathryn; ...

2017-07-10

Genetic elements that replicate extrachromosomally are rare in mammals; however, several human tumor viruses, including the papillomaviruses and the gammaherpesviruses, maintain their plasmid genomes by tethering them to cellular chromosomes. We have uncovered an unprecedented mechanism of viral replication: Kaposi’s sarcoma–associated herpesvirus (KSHV) stably clusters its genomes across generations to maintain itself extrachromosomally. To identify and characterize this mechanism, we developed two complementary, independent approaches: live-cell imaging and a predictive computational model. The clustering of KSHV requires the viral protein, LANA1, to bind viral genomes to nucleosomes arrayed on both cellular and viral DNA. Clustering affects both viral partitioning andmore » viral genome numbers of KSHV. The clustering of KSHV plasmids provides it with an effective evolutionary strategy to rapidly increase copy numbers of genomes per cell at the expense of the total numbers of cells infected.« less

Kaposi’s sarcoma–associated herpesvirus stably clusters its genomes across generations to maintain itself extrachromosomally

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

Chiu, Ya-Fang; Sugden, Arthur U.; Fox, Kathryn

Genetic elements that replicate extrachromosomally are rare in mammals; however, several human tumor viruses, including the papillomaviruses and the gammaherpesviruses, maintain their plasmid genomes by tethering them to cellular chromosomes. We have uncovered an unprecedented mechanism of viral replication: Kaposi’s sarcoma–associated herpesvirus (KSHV) stably clusters its genomes across generations to maintain itself extrachromosomally. To identify and characterize this mechanism, we developed two complementary, independent approaches: live-cell imaging and a predictive computational model. The clustering of KSHV requires the viral protein, LANA1, to bind viral genomes to nucleosomes arrayed on both cellular and viral DNA. Clustering affects both viral partitioning andmore » viral genome numbers of KSHV. The clustering of KSHV plasmids provides it with an effective evolutionary strategy to rapidly increase copy numbers of genomes per cell at the expense of the total numbers of cells infected.« less

Chronically reinforced, operant olfactory conditioning increases the number of newborn GABAergic olfactory periglomerular neurons in the adult rat.

PubMed

Tapia-Rodríguez, Miguel; Esquivelzeta-Rabell, José F; Gutiérrez-Ospina, Gabriel

2012-12-01

The mammalian brain preserves the ability to replace olfactory periglomerular cells (PGC) throughout life. Even though we have detailed a great deal the mechanisms underlying stem and amplifying cells maintenance and proliferation, as well as those modulating migration and differentiation, our knowledge on PGC phenotypic plasticity is at best fragmented and controversial. Here we explored whether chronically reinforced olfactory conditioning influences the phenotype of newborn PGC. Accordingly, olfactory conditioned rats showed increased numbers of GAD 65/67 positive PGC. Because such phenotypic change was not accompanied neither by increments in the total number of PGC, or periglomerular cell nuclei labeled with bromodeoxyuridine, nor by reductions in the number of tyrosine hydroxylase (TH), calbindin (CB) or calretinin (CR) immunoreactive PGC, we speculate that increments in the number of GABAergic PGC occur at the expense of other PGC phenotypes. In any event, these results support that adult newborn PGC phenotype may be subjected to phenotypic plasticity influenced by sensory stimulation. Copyright © 2012 Elsevier B.V. All rights reserved.

A new hand-held microfluidic cytometer for evaluating irradiation damage by analysis of the damaged cells distribution.

PubMed

Wang, Junsheng; Fan, Zhiqiang; Zhao, Yile; Song, Younan; Chu, Hui; Song, Wendong; Song, Yongxin; Pan, Xinxiang; Sun, Yeqing; Li, Dongqing

2016-03-17

Space radiation brings uneven damages to cells. The detection of the distribution of cell damage plays a very important role in radiation medicine and the related research. In this paper, a new hand-held microfluidic flow cytometer was developed to evaluate the degree of radiation damage of cells. The device we propose overcomes the shortcomings (e.g., large volume and high cost) of commercial flow cytometers and can evaluate the radiation damage of cells accurately and quickly with potential for onsite applications. The distribution of radiation-damaged cells is analyzed by a simultaneous detection of immunofluorescence intensity of γ-H2AX and resistance pulse sensor (RPS) signal. The γ-H2AX fluorescence intensity provides information of the degree of radiation damage in cells. The ratio of the number of cells with γ-H2AX fluorescence signals to the total numbers of cells detected by RPS indicates the percentage of the cells that are damaged by radiation. The comparison experiment between the developed hand-held microfluidic flow cytometer and a commercial confocal microscope indicates a consistent and comparable detection performance.

A new hand-held microfluidic cytometer for evaluating irradiation damage by analysis of the damaged cells distribution

NASA Astrophysics Data System (ADS)

Wang, Junsheng; Fan, Zhiqiang; Zhao, Yile; Song, Younan; Chu, Hui; Song, Wendong; Song, Yongxin; Pan, Xinxiang; Sun, Yeqing; Li, Dongqing

2016-03-01

Space radiation brings uneven damages to cells. The detection of the distribution of cell damage plays a very important role in radiation medicine and the related research. In this paper, a new hand-held microfluidic flow cytometer was developed to evaluate the degree of radiation damage of cells. The device we propose overcomes the shortcomings (e.g., large volume and high cost) of commercial flow cytometers and can evaluate the radiation damage of cells accurately and quickly with potential for onsite applications. The distribution of radiation-damaged cells is analyzed by a simultaneous detection of immunofluorescence intensity of γ-H2AX and resistance pulse sensor (RPS) signal. The γ-H2AX fluorescence intensity provides information of the degree of radiation damage in cells. The ratio of the number of cells with γ-H2AX fluorescence signals to the total numbers of cells detected by RPS indicates the percentage of the cells that are damaged by radiation. The comparison experiment between the developed hand-held microfluidic flow cytometer and a commercial confocal microscope indicates a consistent and comparable detection performance.

Up-regulated Th17 cell function is associated with increased peptic ulcer disease in Helicobacter pylori-infection.

PubMed

Bagheri, Nader; Razavi, Alireza; Pourgheysari, Batoul; Azadegan-Dehkordi, Fatemeh; Rahimian, Ghorbanali; Pirayesh, Ashkan; Shafigh, Mohammedhadi; Rafieian-Kopaei, Mahmoud; Fereidani, Rana; Tahmasbi, Kamran; Shirzad, Hedayatollah

2018-06-01

During Helicobacter pylori (H. pylori) infection CD4 + T cells in the gastric lamina propria are hyporesponsive and polarized by Th1/Th17 cell responses controlled by Treg cells. The objective of this study was to determine the number of Th17 cells in gastric mucosa of patients with gastritis and peptic ulcer and determined the relationship between main virulence factor of H. pylori and Th17 cells. A total of 89 H. pylori-infected gastritis patients, 63 H. pylori-infected peptic ulcer patients and 48 H. pylori-negative non-ulcer dysplasia patients were enrolled in this study. The number of Th17 was determined by immunohistochemistry. IL-8 and IL-17A expressions were determined by real-time polymerase chain reaction (qPCR). Also, the grade of chronic and active inflammation was investigated for involvement according to the density of neutrophils and mononuclear in gastric mucosal crypts, from one to all crypts. The number of Th17 cells and the expression of IL-8 and IL-17A in infected patients were significantly higher than uninfected subjects. The number of Th17 cells and the expression of IL-8 and IL-17A in infected patients with peptic ulcer were significantly higher than patients with gastritis. Additionally, the numbers of Th17 cells as well as the expression of IL-8 and IL-17A were positively correlated with the degree of H. pylori density in infected patients with peptic ulcer, while this correlation was negative in infected patients with gastritis. The numbers of Th17 cells as well as the expression of IL-8 and IL-17A were positively correlated with the degree of chronic inflammation. The predominant Th17 cell responses may play a role in the pathogenesis of peptic ulcers disease in infected patients. Copyright © 2018 Elsevier B.V. All rights reserved.

Factors for C-Kit Expression in Cardiac Outgrowth Cells and Human Heart Tissue.

PubMed

Matsushita, Satoshi; Minematsu, Kazuo; Yamamoto, Taira; Inaba, Hirotaka; Kuwaki, Kenji; Shimada, Akie; Yokoyama, Yasutaka; Amano, Atsushi

2017-12-12

We determined the factors associated with the expression of c-kit in the heart and the proliferation of c-kit-positive (c-kit pos ) cardiac stem cells among the outgrowth cells cultured from human cardiac explants.Samples of the right atrium (RA), left atrium (LA), and left ventricle obtained from patients during open-heart surgery were processed for cell culture of outgrowth cells and tissue analysis. The total number of growing cells and the population of c-kit pos cells were measured and compared with c-kit expression in native tissues and characteristics of the patients according to the region of the heart.We analyzed 452 samples from 334 patients. Atrial fibrillation (AF) in the patients reduced the number of outgrowth cells from the RA and LA, and aging was a co-factor for the LA. The c-kit pos population from the RA was associated with serum brain natriuretic peptide (BNP). C-kit expression in native tissue was also associated with BNP expression. However, we observed no relationship in expression between outgrowth cells and native tissue. In addition, the RA tissue provided the highest number of c-kit pos cells, and the left ventricle provided the lowest.C-kit was weakly expressed in response to damage. In addition, no correlation between outgrowth cells and native tissue was found for c-kit expression.

New method for estimating digestion of Paracoccidioides brasiliensis by phagocytic cells in vitro.

PubMed Central

Goihman-Yahr, M; Essenfeld-Yahr, E; Albornoz, M C; Yarzábal, L; de Gómez, M H; San Martín, B; Ocanto, A; Convit, J

1979-01-01

We describe a method by which phagocytosis and digestion of Paracoccidioides brasiliensis yeast cells by polymorphonuclear leukocytes or other phagocytic cells may be estimated. Suspensions of P. brasiliensis in its yeastlike phase were sonicated, counted, and incubated with known numbers of peripheral blood polymorphonuclear leukocytes. At given intervals, cytocentrifuge droplets were stained by a variation of Papanicolaou's method. Stained preparations were examined with phase-contrast optics. Digested organisms showed total or partial disappearance of protoplasm. Green-stained cell walls resisted digestion. The proportion of digested cells as a function of time was estimated. Images PMID:90683

Ganoderma lucidum total triterpenes induce apoptosis in MCF-7 cells and attenuate DMBA induced mammary and skin carcinomas in experimental animals.

PubMed

Smina, T P; Nitha, B; Devasagayam, T P A; Janardhanan, K K

2017-01-01

Ganoderma lucidum total triterpenes were evaluated for its apoptosis-inducing and anti-cancer activities. Cytotoxicity and pro-apoptotic effect of total triterpenes were evaluated in human breast adenocarcinoma (MCF-7) cell line using MTT assay and DNA fragmentation analysis. Total triterpenes induced apoptosis in MCF-7 cells by down-regulating the levels of cyclin D1, Bcl-2, Bcl-xL and also by up-regulating the levels of Bax and caspase-9. Anti-carcinogenicity of total triterpenes was analysed using dimethyl benz [a] anthracene (DMBA) induced skin papilloma and mammary adenocarcinoma in Swiss albino mice and Wistar rats respectively. Topical application of 5mg, 10mg and 20mg total triterpenes reduced the incidence of skin papilloma by 62.5, 37.5 and 12.5% respectively. Incidence of the mammary tumour was also reduced significantly by 33.33, 66.67 and 16.67% in 10, 50 and 100mg/kg b.wt. total triterpenes treated animals respectively. Total triterpenes were also found to reduce the average number of tumours per animal and extended the tumour latency period in both the models. The results indicate the potential cytotoxicity and anti-cancerous activity of total triterpenes, there by opens up a path to the development of a safe and successive chemo preventive agent of natural origin. Copyright © 2016 Elsevier B.V. All rights reserved.

Hematopoietic stem cell transplantation in Europe 1998.

PubMed

Gratwohl, A; Passweg, J; Baldomero, H; Hermans, J; Urbano-Ispizua, A

2000-01-01

Transplantation of hematopoietic stem cells from blood or bone marrow has become accepted therapy for many diseases. Numbers of transplants have increased significantly and stem cell source, donor type and indications have changed during this decade. Information on these changes is essential for interpretation of current data, patient counseling and health care planning. Since 1990, members of the European Group for Blood and Marrow Transplantation and teams known to perform blood or marrow transplants have been invited annually to report their transplant numbers by indication, donor type and stem cell source. Data from these surveys have been used to present data for 1998, to assess current status and to give numbers of transplants per participating country, coefficients of variation between countries for individual indications and changes in indication, stem cell source and donor type over the past decade. In 1998, a total of 20 892 transplants were performed by 528 teams in 31 European countries. Of these transplants 18 400 were first transplants, 5308 (29%) were allogenic, and 13 092 (71%) were autologous. Of the autologous transplants, 809 (6%) were bone marrow derived, and 12 283 (94%) were from peripheral blood stems cells. Of the allogeneic transplants, 3372 (64%) were bone marrow derived, and 1936 (36%) were peripheral blood stem cell transplants. In 1990, the respective figures were 2137 allogeneic (50%) and 2097 (50%) autologous transplants, all exclusively bone marrow derived. Main indications in 1998 were leukemias with 6015 transplants (33%), 68% thereof allogeneic transplants; lymphomas with 7492 transplants (41%), 94% thereof autologous transplants; solid tumors with 4025 transplants (22%), 99% thereof autologous transplants; non-malignant disorders with 868 transplants (5%), 80% thereof allogeneic transplants. Absolute numbers of transplants per year did increase from 4234 in 1990 to 20 892 in 1998. Increase is higher for autologous, than for allogeneic transplants. There were differences in absolute or relative increase over time for individual indications. Transplant rates per number of inhabitants varied between countries, ranging from 0 to >500 total transplants per 10 million inhabitants with a clear correlation between number of teams and transplants per 10 million inhabitants (r=0.61, P<0.001). The least variation between countries was observed for acute leukemias, chronic myeloid leukemia and severe aplastic anemia in allogeneic transplants, for Hodgkin's disease and non-Hodgkin's lymphoma in autologous transplants. These data reflect the current status of blood and marrow transplantation in Europe. They show the continuing increase in utilization, highlight the change from bone marrow to blood as stem cell source and give an objective assessment on presence or absence of trends.

Natural killer cell dysfunction in hepatocellular carcinoma and NK cell-based immunotherapy

PubMed Central

Sun, Cheng; Sun, Hao-yu; Xiao, Wei-hua; Zhang, Cai; Tian, Zhi-gang

2015-01-01

The mechanisms linking hepatitis B virus (HBV) and hepatitis C virus (HCV) infection to hepatocellular carcinoma (HCC) remain largely unknown. Natural killer (NK) cells account for 25%–50% of the total number of liver lymphocytes, suggesting that NK cells play an important role in liver immunity. The number of NK cells in the blood and tumor tissues of HCC patients is positively correlated with their survival and prognosis. Furthermore, a group of NK cell-associated genes in HCC tissues is positively associated with the prolonged survival. These facts suggest that NK cells and HCC progression are strongly associated. In this review, we describe the abnormal NK cells and their functional impairment in patients with chronic HBV and HCV infection, which contribute to the progression of HCC. Then, we summarize the association of NK cells with HCC based on the abnormalities in the numbers and phenotypes of blood and liver NK cells in HCC patients. In particular, the exhaustion of NK cells that represents lower cytotoxicity and impaired cytokine production may serve as a predictor for the occurrence of HCC. Finally, we present the current achievements in NK cell immunotherapy conducted in mouse models of liver cancer and in clinical trials, highlighting how chemoimmunotherapy, NK cell transfer, gene therapy, cytokine therapy and mAb therapy improve NK cell function in HCC treatment. It is conceivable that NK cell-based anti-HCC therapeutic strategies alone or in combination with other therapies will be great promise for HCC treatment. PMID:26073325

Significance of Fractionated Administration of Thalidomide Combined With γ-Ray Irradiation in Terms of Local Tumor Response and Lung Metastasis

PubMed Central

Masunaga, Shin-ichiro; Sanada, Yu; Moriwaki, Takahiro; Tano, Keizo; Sakurai, Yoshinori; Tanaka, Hiroki; Suzuki, Minoru; Kondo, Natsuko; Narabayashi, Masaru; Watanabe, Tsubasa; Nakagawa, Yosuke; Maruhashi, Akira; Ono, Koji

2014-01-01

Background The aim of this study was to evaluate the significance of fractionated administration of thalidomide combined with γ-ray irradiation in terms of local tumor response and lung metastatic potential, referring to the response of intratumor quiescent (Q) cells. Methods B16-BL6 melanoma tumor-bearing C57BL/6 mice were continuously given 5-bromo-2’-deoxyuridine (BrdU) to label all proliferating (P) cells. The tumor-bearing mice then received γ-ray irradiation after thalidomide treatment through a single or two consecutive daily intraperitoneal administrations up to a total dose of 400 mg/kg in combination with an acute hypoxia-releasing agent (nicotinamide) or mild temperature hyperthermia (MTH). Immediately after the irradiation, cells from some tumors were isolated and incubated with a cytokinesis blocker. The responses of the Q and total (= P + Q) cell populations were assessed based on the frequency of micronuclei using immunofluorescence staining for BrdU. In other tumor-bearing mice, 17 days after irradiation, macroscopic lung metastases were enumerated. Results Thalidomide raised the sensitivity of the total cell population more remarkably than Q cells in both single and daily administrations. Daily administration of thalidomide elevated the sensitivity of both the total and Q cell populations, but especially the total cell population, compared with single administration. Daily administration, especially combined with MTH, decreased the number of lung metastases. Conclusion Daily fractionated administration of thalidomide in combination with γ-ray irradiation was thought to be more promising than single administration because of its potential to enhance local tumor response and repress lung metastatic potential. PMID:29147396

Xylem Development of Loblolly Pine During Irrigation and Simulated Drought

Treesearch

David M. Moehring; Charles X. Grano; John R. Bassett

1975-01-01

Induced drought decreased the total number of tracheid cells produced in a season and the radial width of late wood tracheids. Transition from earlywood to late woodo ccurred in early June and was little affected by irrigation.

H2O2-induced mild stress in relation with in vitro ovine oocyte developmental competence: implications for blastocyst apoptosis and related genes expression.

PubMed

Nikdel, K; Aminafshar, M; Mohammadi-Sangcheshmeh, A; EmamJomeh-Kashan, N; Seyedjafari, E

2017-05-20

In this study, in vitro maturation was performed in presence of various concentrations (0, 10, 100, or 1000 µM) of H2O2. The intracellular glutathione (GSH) level, fertilization, cleavage, and blastocyst rates, total cell number, and apoptotic cell number and expression of Bax, Bcl-2, and p53 genes in blastocyst-stage embryos were studied. At 10 μM H2O2 concentration, a higher GSH level was detected in comparison to the other groups while oocytes exposed to 1000 μM H2O2 had the lowest GSH level. Treatment of oocytes with 1000 μM H2O2 decreased the rate of two pronuclei formation as compared with other groups. A higher rate of blastocyst formation was seen in 100 μM H2O2 group as compared with the control group. However, exogenous H2O2 in maturation medium did not affect total cell numbers and apoptotic cell ratio at the blastocyst stage. Moreover, mRNA transcript abundance of Bax, Bcl-2, and p53 genes was similar between blastocysts derived from H2O2-induced oocytes and control blastocysts. Treatment of oocytes with H2O2 at mild level during in vitro maturation had a positive effect on GSH level and this, in turn, may lead to improvement in preimplantation embryonic development.

Numerical analysis of a dielectrophoresis field-flow fractionation device for the separation of multiple cell types.

PubMed

Shamloo, Amir; Kamali, Ali

2017-10-01

In this study, a dielectrophoresis field-flow fractionation device was analyzed using a numerical simulation method and the behaviors of a set of different cells were investigated. By reducing the alternating current frequency of the electrodes from the value used in the original setup configuration and increasing the number of exit channels, total discrimination in cell trajectories and subsequent separation of four cell types were achieved. Cells were differentiated based on their size and dielectric response that are represented in their real part of Clausius-Mossotti factor at different frequencies. A number of novel designs were also proposed based on the original setup configuration. It was seen that by reducing the length of the main channel and the number of electrodes at low frequencies and not changing the inlet flow velocities, cell separation was still achieved successfully, although with a slightly larger electrode voltage. The shorter main channel decreased the residence time for the cells on the chip and also reduced the overall size of the device-these were improvements over the original design. The obtained results can be used to analyze other cell types by knowing their size and dielectric properties to design geometries that can ensure separation. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Prediction of Response to Therapy and Clinical Outcome through a Pilot Study of Complete Genetic Assessment of Ovarian Cancer

DTIC Science & Technology

2015-12-01

Oncology program supported by this grant consented patients to 11-104. OncoPanel is a cancer genomic assay that detects somatic mutations, copy number...KMT2D, EP300, FANCD2 Sertoli Leydig cell DICER1 Copy number variants: In addition, 219 patients were analyzed for copy-number variations ( CNV ) in...OncoPanel genes. >12,000 total CNV were reported in the cohort (Figure 2). Single- copy deletions (n=5558) and copy-number gains (low amplification) (n

Interaction between phloretin and the red blood cell membrane

PubMed Central

1976-01-01

Phloretin binding to red blood cell components has been characterized at pH6, where binding and inhibitory potency are maximal. Binding to intact red cells and to purified hemoglobin are nonsaturated processes approximately equal in magnitude, which strongly suggests that most of the red cell binding may be ascribed to hemoglobin. This conclusion is supported by the fact that homoglobin-free red cell ghosts can bind only 10% as much phloretin as an equivalent number of red cells. The permeability of the red cell membrane to phloretin has been determined by a direct measurement at the time-course of the phloretin uptake. At a 2% hematocrit, the half time for phloretin uptake is 8.7s, corresponding to a permeability coefficient of 2 x 10(-4) cm/s. The concentration dependence of the binding to ghosts reveals two saturable components. Phloretin binds with high affinity (K diss = 1.5 muM) to about 2.5 x 10(6) sites per cell; it also binds with lower affinity (Kdiss = 54 muM) to a second (5.5 x 10(7) per cell) set of sites. In sonicated total lipid extracts of red cell ghosts, phloretin binding consists of a single, saturable component. Its affinity and total number of sites are not significantly different from those of the low affinity binding process in ghosts. No high affinity binding of phloretin is exhibited by the red cell lipid extracts. Therefore, the high affinity phloretin binding sites are related to membrane proteins, and the low affinity sites result from phloretin binding to lipid. The identification of these two types of binding sites allows phloretin effects on protein-mediated transport processes to be distinguished from effects on the lipid region of the membrane. PMID:5575

Microbial Community Response during the Iron Fertilization Experiment LOHAFEX

PubMed Central

Thiele, Stefan; Ramaiah, Nagappa; Amann, Rudolf

2012-01-01

Iron fertilization experiments in high-nutrient, low-chlorophyll areas are known to induce phytoplankton blooms. However, little is known about the response of the microbial community upon iron fertilization. As part of the LOHAFEX experiment in the southern Atlantic Ocean, Bacteria and Archaea were monitored within and outside an induced bloom, dominated by Phaeocystis-like nanoplankton, during the 38 days of the experiment. The microbial production increased 1.6-fold (thymidine uptake) and 2.1-fold (leucine uptake), while total cell numbers increased only slightly over the course of the experiment. 454 tag pyrosequencing of partial 16S rRNA genes and catalyzed reporter deposition fluorescence in situ hybridization (CARD FISH) showed that the composition and abundance of the bacterial and archaeal community in the iron-fertilized water body were remarkably constant without development of typical bloom-related succession patterns. Members of groups usually found in phytoplankton blooms, such as Roseobacter and Gammaproteobacteria, showed no response or only a minor response to the bloom. However, sequence numbers and total cell numbers of the SAR11 and SAR86 clades increased slightly but significantly toward the end of the experiment. It seems that although microbial productivity was enhanced within the fertilized area, a succession-like response of the microbial community upon the algal bloom was averted by highly effective grazing. Only small-celled members like the SAR11 and SAR86 clades could possibly escape the grazing pressure, explaining a net increase of those clades in numbers. PMID:23064339

In vitro expansion of Lin+ and Lin- mononuclear cells from human peripheral blood

NASA Astrophysics Data System (ADS)

Norhaiza, H. Siti; Rohaya, M. A. W.; Zarina, Z. A. Intan; Hisham, Z. A. Shahrul

2013-11-01

Haematopoietic stem cells (HSCs) are used in the therapy of blood disorders due to the ability of these cells to reconstitute haematopoietic lineage cells when transplanted into myeloablative recipients. However, substantial number of cells is required in order for the reconstitution to take place. Since HSCs present in low frequency, larger number of donor is required to accommodate the demand of transplantable HSCs. Therefore, in vitro expansion of HSCs will have profound impact on clinical purposes. The aim of this study was to expand lineage negative (Lin-) stem cells from human peripheral blood. Total peripheral blood mononuclear cells (PBMNCs) were fractionated from human blood by density gradient centrifugation. Subsequently, PBMNCs were subjected to magnetic assisted cell sorter (MACS) which depletes lineage positive (Lin+) mononuclear cells expressing lineage positive markers such as CD2, CD3, CD11b, CD14, CD15, CD16, CD19, CD56, CD123, and CD235a to obtained Lin- cell population. The ability of Lin+ and Lin- to survive in vitro was explored by culturing both cell populations in complete medium consisting of Alpha-Minimal Essential Medium (AMEM) +10% (v/v) Newborn Calf Serum (NBCS)+ 2% (v/v) pen/strep. In another experiment, Lin+ and Lin- were cultured with complete medium supplemented with 10ng/mL of the following growth factors: stem cell factor (SCF), interleukin (IL)-3, granulocyte-macrophage colony stimulating factor (GM-CSF), 2IU/mL of Erythropoietin (Epo) and 20ng/mL of IL-6. Three samples were monitored in static culture for 22 days. The expansion potential was assessed by the number of total viable cells, counted by trypan blue exclusion assay. It was found that Lin+ mononuclear cells were not able to survive either in normal proliferation medium or proliferation medium supplemented with cytokines. Similarly, Lin- stem cells were not able to survive in proliferation medium however, addition of cytokines into the proliferation medium support Lin- stem cells for at least 18 days. The Lin- stem cells started to response to the cytokines added as early as Day 2 of culture. It is concluded that Lin- stem cells can be expanded in vitro by culturing in proliferation medium supplemented with cytokines.

Stem cell transplantation for treating Parkinson's disease: Literature analysis based on the Web of Science.

PubMed

Li, Runhui

2012-06-05

To identify global research trends of stem cell transplantation for treating Parkinson's disease using a bibliometric analysis of the Web of Science. We performed a bibliometric analysis of data retrievals for stem cell transplantation for treating Parkinson's disease from 2002 to 2011 using the Web of Science. (a) peer-reviewed articles on stem cell transplantation for treating Parkinson's disease which were published and indexed in the Web of Science; (b) type of articles: original research articles, reviews, meeting abstracts, proceedings papers, book chapters, editorial material and news items; (c) year of publication: 2002-2011. (a) articles that required manual searching or telephone access; (b) we excluded documents that were not published in the public domain; (c) we excluded a number of corrected papers from the total number of articles. (1) Type of literature; (2) annual publication output; (3) distribution according to journals; (4) distribution according to subject areas; (5) distribution according to country; (6) distribution according to institution; (7) comparison of countries that published the most papers on stem cell transplantation from different cell sources for treating Parkinson's disease; (8) comparison of institutions that published the most papers on stem cell transplantation from different cell sources for treating Parkinson's disease in the Web of Science from 2002 to 2011; (9) comparison of studies on stem cell transplantation from different cell sources for treating Parkinson's disease. In total, 1 062 studies on stem cell transplantation for treating Parkinson's disease appeared in the Web of Science from 2002 to 2011, almost one third of which were from American authors and institutes. The number of studies on stem cell transplantation for treating Parkinson's disease had gradually increased over the past 10 years. Papers on stem cell transplantation for treating Parkinson's disease appeared in journals such as Stem Cells and Experimental Neurology. Although the United States published more articles addressing neural stem cell and embryonic stem cell transplantation for treating Parkinson's disease, China ranked first for articles published on bone marrow mesenchymal stem cell transplantation for treating Parkinson's disease. From our analysis of the literature and research trends, we found that stem cell transplantation for treating Parkinson's disease may offer further benefits in regenerative medicine.

Characteristics of viral protein expression by Epstein-Barr virus-infected B cells in peripheral blood of patients with infectious mononucleosis.

PubMed

Wagner, H J; Hornef, M; Middeldorp, J; Kirchner, H

1995-11-01

The frequency of Epstein-Barr virus (EBV) antigen-positive B cells in the peripheral blood of patients with infectious mononucleosis compared with that for latently EBV-infected individuals was examined by immunocytochemistry. B cells positive for Epstein-Barr nuclear antigen (EBNA) 1, EBNA2, and latent membrane protein were frequently found in all peripheral B lymphocyte preparations from 25 patients suffering for 3 to 28 days from infectious mononucleosis by using monoclonal antibodies and the alkaline phosphatase anti-alkaline technique. There was a significant decrease in the number of positive B cells during the course of disease. EBNA1-positive B cells were detected in 0.01 to 2.5% of total B cells (median, 0.8%), EBNA2-positive B cells were detected in 0.01 to 4.5% of total B cells (median, 0.9%), and latent membrane protein-positive B cells were detected in 0.01 to 1.8% of total B cells (median, 0.5%), depending on the duration of clinical signs. In contrast, we did not find any EBNA1- or EBNA2-positive B cells in 2 x 10(6) peripheral blood B lymphocytes of 10 latently EBV-infected individuals, whereas aliquots of the same cell preparations were EBV DNA positive by a PCR assay. Therefore, it appears to be possible to detect infectious mononucleosis by immunocytochemical determination of latent EBV products, which might be of relevance for the diagnosis of EBV reactivations in immunosuppressed patients.

Cutaneous CD30+ cells in children with atopic dermatitis.

PubMed

Cavagni, G; Caffarelli, C; Facchetti, F; Brugnoni, D; Notarangelo, L D; Tosoni, C; Altobelli, R

2000-03-01

CD30 expression can be considered a marker of Th2 cells. We investigated the presence of CD30+ cells in the lesional skin of children with atopic dermatitis (AD). We also analyzed the possible relationship between CD30+ cells and serum soluble CD 30 (sCD30) levels, and IgE, soluble interleukin-2 (IL-2) receptor (sIL-2R) or soluble CD23 (sCD23) levels in the blood, and clinical score. Ten eczematous children (4 males, 6 females; median age: 4 years and 5 months; range: 11 months to 14 years), 9 sex- and age-matched control children and an adult control group were studied. A clinical score (SCORAD index), was given to eczematous lesions. Blood was taken for the determination of IgE, sCD30, sIL-2R and sCD23 levels. Punch biopsies of lesional skin were stained with hematoxylin and eosin or incubated with anti-CD30 monoclonal antibodies. Skin prick tests (SPTs) were also performed. In the biopsy specimens, CD30 expression was observed in high proportions of infiltrating cells. In children with AD, total serum IgE, sCD30, sIL-2R, sCD23 and eosinophils were significantly elevated compared to controls. CD30+ cells were not associated with serum IgE, sCD30, sIL-2R, sCD23, or SPT results, score of inflammatory cells in lesional skin or clinical score. Children with AD who had high total IgE and specific IgE antibodies did not differ from those with normal total IgE and negative specific IgE in respect of age, clinical score, number of CD30+ cells, sCD30, sIL-2R and sCD23 levels, score of inflammatory cells in skin or clinical score. Our results showed remarkable numbers of CD30+ cells in the lesional skin and high sCD30 in the serum of children with AD. CD30+ cells did not correlate with systemic markers of IgE reaction. Copyright 2000 S. Karger AG, Basel

Establishment of a long-term spiral ganglion neuron culture with reduced glial cell number: Effects of AraC on cell composition and neurons.

PubMed

Schwieger, Jana; Esser, Karl-Heinz; Lenarz, Thomas; Scheper, Verena

2016-08-01

Sensorineural deafness is mainly caused by damage to hair cells and degeneration of the spiral ganglion neurons (SGN). Cochlear implants can functionally replace lost hair cells and stimulate the SGN electrically. The benefit from cochlear implantation depends on the number and excitability of these neurons. To identify potential therapies for SGN protection, in vitro tests are carried out on spiral ganglion cells (SGC). A glial cell-reduced and neuron-enhanced culture of neonatal rat SGC under mitotic inhibition (cytarabine (AraC)) for up to seven days is presented. Serum containing and neurotrophin-enriched cultures with and without AraC-addition were analyzed after 4 and 7 days. The total number of cells was significantly reduced, while the proportion of neurons was greatly increased by AraC-treatment. Cell type-specific labeling demonstrated that nearly all fibroblasts and most of the glial cells were removed. Neither the neuronal survival, nor the neurite outgrowth or soma diameter were negatively affected. Additionally neurites remain partly free of surrounding non-neuronal cells. Recent culture conditions allow only for short-term cultivation of neonatal SGC and lack information on the influence of non-neuronal cells on SGN and of direct contact of neurites with test-materials. AraC-addition reduces the number of non-neuronal cells and increases the ratio of SGN in culture, without negative impact on neuronal viability. This treatment allows longer-term cultivation of SGC and provides deeper insight into SGN-glial cell interaction and the attachment of neurites on test-material surfaces. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Changes in T-lymphocyte distribution associated with ingestion of aldicarb-contaminated drinking water: a follow-up study.

PubMed

Mirkin, I R; Anderson, H A; Hanrahan, L; Hong, R; Golubjatnikov, R; Belluck, D

1990-02-01

The carbamate pesticide, aldicarb, is the most commonly found man-made groundwater contaminant in Wisconsin. A 1985 study linked ingestion of aldicarb-contaminated drinking water with altered T-cell distributions, specifically an increase in the mean number of CD8+ (T8) T cells. To further evaluate this finding, a follow-up study was done in 1987. Of the 50 Portage County, Wisconsin, women who participated in the first study, 45 participated in the follow-up: 18 formerly exposed and 27 formerly unexposed. In our follow-up study, only 5 women were found to be currently exposed to aldicarb. This group of 5 women, compared to 39 unexposed women who had peripheral blood specimens taken, had an increased percentage of lymphocytes and an increased number of CD2+ T cells, due to an increased number of total CD8+ T cells. Although the number of exposed persons was small, the increases in percentage lymphocytes and absolute numbers of CD2+ and CD8+ T cells were consistent with a dose-response relationship. No identified drinking water contaminant other than aldicarb could explain these findings. These results support earlier evidence linking aldicarb exposure and lymphocyte distribution changes. Although adverse clinical effects have not been documented, the widespread use of this chemical and consequent potential for widespread exposure indicate a clear need for further research on this issue.

Whole body proton irradiation causes acute damage to bone marrow hematopoietic progenitor and stem cells in mice.

PubMed

Chang, Jianhui; Wang, Yingying; Pathak, Rupak; Sridharan, Vijayalakshmi; Jones, Tamako; Mao, Xiao Wen; Nelson, Gregory; Boerma, Marjan; Hauer-Jensen, Martin; Zhou, Daohong; Shao, Lijian

2017-12-01

Exposure to proton irradiation during missions in deep space can lead to bone marrow injury. The acute effects of proton irradiation on hematopoietic stem and progenitor cells remain undefined and thus were investigated. We exposed male C57BL/6 mice to 0.5 and 1.0 Gy proton total body irradiation (proton-TBI, 150 MeV) and examined changes in peripheral blood cells and bone marrow (BM) progenitors and LSK cells 2 weeks after exposure. 1.0 Gy proton-TBI significantly reduced the numbers of peripheral blood cells compared to 0.5 Gy proton-TBI and unirradiated animals, while the numbers of peripheral blood cell counts were comparable between 0.5 Gy proton-TBI and unirradiated mice. The frequencies and numbers of LSK cells and CMPs in BM of 0.5 and 1.0 Gy irradiated mice were decreased in comparison to those of normal controls. LSK cells and CMPs and their progeny exhibited a radiation-induced impairment in clonogenic function. Exposure to 1.0 Gy increased cellular apoptosis but not the production of reactive oxygen species (ROS) in CMPs two weeks after irradiation. LSK cells from irradiated mice exhibited an increase in ROS production and apoptosis. Exposure to proton-TBI can induce acute damage to BM progenitors and LSK cells.

Flow Cytometry of Human Primary Epidermal and Follicular Keratinocytes

PubMed Central

Gragnani, Alfredo; Ipolito, Michelle Zampieri; Sobral, Christiane S; Brunialti, Milena Karina Coló; Salomão, Reinaldo; Ferreira, Lydia Masako

2008-01-01

Objective: The aim of this study was to characterize using flow cytometry cultured human primary keratinocytes isolated from the epidermis and hair follicles by different methods. Methods: Human keratinocytes derived from discarded fragments of total skin and scalp hair follicles from patients who underwent plastic surgery in the Plastic Surgery Division at UNIFESP were used. The epidermal keratinocytes were isolated by using 3 different methods: the standard method, upon exposure to trypsin for 30 minutes; the second, by treatment with dispase for 18 hours and with trypsin for 10 minutes; and the third, by treatment with dispase for 18 hours and with trypsin for 30 minutes. Follicular keratinocytes were isolated using the standard method. Results: On comparing the group treated with dispase for 18 hours and with trypsin for 10 minutes with the group treated with dispase for 18 hours and with trypsin for 30 minutes, it was observed that the first group presented the largest number of viable cells, the smallest number of cells in late apoptosis and necrosis with statistical significance, and no difference in apoptosis. When we compared the group treated with dispase for 18 hours and with trypsin for 10 minutes with the group treated with trypsin, the first group presented the largest number of viable cells, the smallest number of cells in apoptosis with statistical significance, and no difference in late apoptosis and necrosis. When we compared the results of the group treated with dispase for 18 hours and with trypsin for 10 minutes with the results for follical isolation, there was a statistical difference in apoptosis and viable cells. Conclusion: The isolation method of treatment with dispase for 18 hours and with trypsin for 10 minutes produced the largest number of viable cells and the smallest number of cells in apoptosis/necrosis. PMID:18350110

Flow cytometry of human primary epidermal and follicular keratinocytes.

PubMed

Gragnani, Alfredo; Ipolito, Michelle Zampieri; Sobral, Christiane S; Brunialti, Milena Karina Coló; Salomão, Reinaldo; Ferreira, Lydia Masako

2008-02-19

The aim of this study was to characterize using flow cytometry cultured human primary keratinocytes isolated from the epidermis and hair follicles by different methods. Human keratinocytes derived from discarded fragments of total skin and scalp hair follicles from patients who underwent plastic surgery in the Plastic Surgery Division at UNIFESP were used. The epidermal keratinocytes were isolated by using 3 different methods: the standard method, upon exposure to trypsin for 30 minutes; the second, by treatment with dispase for 18 hours and with trypsin for 10 minutes; and the third, by treatment with dispase for 18 hours and with trypsin for 30 minutes. Follicular keratinocytes were isolated using the standard method. On comparing the group treated with dispase for 18 hours and with trypsin for 10 minutes with the group treated with dispase for 18 hours and with trypsin for 30 minutes, it was observed that the first group presented the largest number of viable cells, the smallest number of cells in late apoptosis and necrosis with statistical significance, and no difference in apoptosis. When we compared the group treated with dispase for 18 hours and with trypsin for 10 minutes with the group treated with trypsin, the first group presented the largest number of viable cells, the smallest number of cells in apoptosis with statistical significance, and no difference in late apoptosis and necrosis. When we compared the results of the group treated with dispase for 18 hours and with trypsin for 10 minutes with the results for follical isolation, there was a statistical difference in apoptosis and viable cells. The isolation method of treatment with dispase for 18 hours and with trypsin for 10 minutes produced the largest number of viable cells and the smallest number of cells in apoptosis/necrosis.

Number of circulating endothelial progenitor cells and intratumoral microvessel density in non-small cell lung cancer patients: differences in angiogenic status between adenocarcinoma histologic subtypes.

PubMed

Maeda, Ryo; Ishii, Genichiro; Ito, Masami; Hishida, Tomoyuki; Yoshida, Junji; Nishimura, Mitsuyo; Haga, Hironori; Nagai, Kanji; Ochiai, Atsushi

2012-03-01

Angiogenesis plays a significant role in tumor progression. This study examined the association between the number of circulating endothelial progenitor cells (EPCs), intratumoral microvessel density (MVD) (both of which may be markers for neovascularization), and lung cancer histological types, particularly adenocarcinoma histological subtypes. A total of 83 stage I non-small cell lung cancer (NSCLC) patients underwent complete tumor resection between November 2009 and July 2010. The number of EPCs from the pulmonary artery of the resected lungs was measured by assaying CD34/vascular endothelial growth factor receptor 2 positive cells, and the MVD was assessed immunohistochemically in tumor specimens by staining for CD34. A statistically significant correlation between the number of EPCs from pulmonary artery and intratumoral MVD was found (p < 0.001). No statistically significant differences in the number of EPCs and the MVD were observed between the adenocarcinomas and the squamous cell carcinomas. Among the adenocarcinoma histological subtypes, a higher number of EPCs and MVD were found significantly more frequently in solid adenocarcinomas than in nonsolid adenocarcinomas (p < 0.001 and p = 0.011, respectively). In addition, solid adenocarcinomas showed higher levels of vascular endothelial growth factor using quantitative real-time polymerase chain reaction in the tumor tissue samples than in the nonsolid adenocarcinomas (p = 0.005). The higher number of circulating EPCs and the MVD of solid adenocarcinoma may indicate the presence of differences in the tumor angiogenic status between early-stage adenocarcinoma histological subtypes. Among adenocarcinoma patients, patients with solid adenocarcinoma may be the best candidates for antiangiogenic therapies.

Stem cell transplantation for treating stroke: status, trends and development.

PubMed

Huo, Wenxin; Liu, Xiaoyang; Tan, Cheng; Han, Yingying; Kang, Chunyang; Quan, Wei; Chen, Jiajun

2014-09-01

The developing approaches of thrombolytic therapy, endovascular treatment, neuroprotective therapy, and stem cell therapy have enabled breakthroughs in stroke treatment. In this study, we summarize and analyze trends and progress in stem cell transplantation for stroke treatment by retrieval of literature from Thomson Reuters Web of Science database, the NIH Clinical Trial Planning Grant Program, and Clinical Trials Registration Center in North America. In the last 10 years, there has been an increasing number of published articles on stem cell transplantation for stroke treatment. In particular, research from the USA and China has focused on stem cell transplantation. A total of 2,167 articles addressing stem cell transplantation for stroke treatment from 2004 to 2013 were retrieved from the Thomson Reuters Web of Science database. The majority of these articles were from the USA (854, 39.4%), with the journal Stroke publishing the most articles (145, 6.7%). Of the published articles, 143 were funded by the National Institutes of Health (accounting for 6.6% of total publications), and 91 by the National Natural Science Foundation of China. Between 2013 and 2014, the National Institutes of Health provided financial support ($130 million subsidy) for 329 research projects on stroke therapy using stem cell transplantation. In 2014, 215 new projects were approved, receiving grants of up to $70,440,000. Ninety clinical trials focusing on stem cell transplantation for stroke were registered in the Clinical Trial Registration Center in North America, with 40 trials registered in the USA (ranked first place). China had the maximum number of registered research or clinical trials (10 projects).

An optimized protocol for the generation and functional analysis of human mast cells from CD34+ enriched cell populations.

PubMed

Yin, Yuzhi; Bai, Yun; Olivera, Ana; Desai, Avanti; Metcalfe, Dean D

2017-09-01

The culture of mast cells from human tissues such a cord blood, peripheral blood or bone marrow aspirates has advanced our understanding of human mast cells (huMC) degranulation, mediator production and response to pharmacologic agents. However, existing methods for huMC culture tend to be laborious and expensive. Combining technical approaches from several of these protocols, we designed a simplified and more cost effective approach to the culture of mast cells from human cell populations including peripheral blood and cryopreserved cells from lymphocytapheresis. On average, we reduced by 30-50 fold the amount of culture media compared to our previously reported method, while the total MC number generated by this method (2.46±0.63×10 6 vs. 2.4±0.28×10 6 , respectively, from 1.0×10 8 lymphocytapheresis or peripheral blood mononuclear blood cells [PBMCs]) was similar to our previous method (2.36±0.70×10 6 ), resulting in significant budgetary savings. In addition, we compared the yield of huMCs with or without IL-3 added to early cultures in the presence of stem cell factor (SCF) and interlukin-6 (IL-6) and found that the total MC number generated, while higher with IL-3 in the culture, did not reach statistical significance, suggesting that IL-3, often recommended in the culture of huMCs, is not absolutely required. We then performed a functional analysis by flow cytometry using standard methods and which maximized the data we could obtain from cultured cells. We believe these approaches will allow more laboratories to culture and examine huMC behavior going forward. Published by Elsevier B.V.

Development of a flow cytometric method to analyze subpopulations of bacteria in probiotic products and dairy starters.

PubMed

Bunthof, Christine J; Abee, Tjakko

2002-06-01

Flow cytometry (FCM) is a rapid and sensitive technique that can determine cell numbers and measure various physiological characteristics of individual cells by using appropriate fluorescent probes. Previously, we developed an FCM assay with the viability probes carboxyfluorescein diacetate (cFDA) and TOTO-1 [1'-(4,4,7,7-tetramethyl-4,7-diazaundecamethylene)-bis-4-[3-methyl-2,3dihydro(benzo-1,3-oxazole)-2-methylidene]-1-(3'-trimethylammoniumpropyl)-pyridinium tetraiodide] for (stressed) lactic acid bacteria (C. J. Bunthof, K. Bloemen, P. Breeuwer, F. M. Rombouts, and T. Abee, Appl. Environ. Microbiol. 67:2326-2335, 2001). cFDA stains intact cells with enzymatic activity, and TOTO-1 stains membrane-permeabilized cells. Here we used this assay to study the viability of bacterial suspensions in milk, dairy fermentation starters, and probiotic products. To facilitate FCM analysis of bacteria in milk, a commercially available milk-clearing solution was used. The procedure was optimized to increase the signal-to-noise ratio. FCM enumerations were accurate down to a concentration of 10(5) cells ml(-1). The level of retrieval of Lactobacillus plantarum WCFS 1 suspended in milk was high, and viability was not affected by the procedure. The plate counts for cleared samples of untreated cell suspensions were nearly as high as the total FCM counts, and the correlation was strong (r > 0.99). In dairy fermentation starters and in probiotic products the FCM total cell counts were substantially higher than the numbers of CFU. Three functional populations could be distinguished: culturable cells, cells that are intact and metabolically active but not culturable, and permeabilized cells. The proportions of the populations differed in the products tested. This FCM method provides tools to assess the functionality of different populations in fermentation starters and probiotic products.

Development of a Flow Cytometric Method To Analyze Subpopulations of Bacteria in Probiotic Products and Dairy Starters

PubMed Central

Bunthof, Christine J.; Abee, Tjakko

2002-01-01

Flow cytometry (FCM) is a rapid and sensitive technique that can determine cell numbers and measure various physiological characteristics of individual cells by using appropriate fluorescent probes. Previously, we developed an FCM assay with the viability probes carboxyfluorescein diacetate (cFDA) and TOTO-1 {1′-(4,4,7,7-tetramethyl-4,7-diazaundecamethylene)-bis-4-[3-methyl-2,3dihydro(benzo-1,3-oxazole)-2-methylidene]-1-(3′-trimethylammoniumpropyl)-pyridinium tetraiodide} for (stressed) lactic acid bacteria (C. J. Bunthof, K. Bloemen, P. Breeuwer, F. M. Rombouts, and T. Abee, Appl. Environ. Microbiol. 67:2326-2335, 2001). cFDA stains intact cells with enzymatic activity, and TOTO-1 stains membrane-permeabilized cells. Here we used this assay to study the viability of bacterial suspensions in milk, dairy fermentation starters, and probiotic products. To facilitate FCM analysis of bacteria in milk, a commercially available milk-clearing solution was used. The procedure was optimized to increase the signal-to-noise ratio. FCM enumerations were accurate down to a concentration of 105 cells ml−1. The level of retrieval of Lactobacillus plantarum WCFS 1 suspended in milk was high, and viability was not affected by the procedure. The plate counts for cleared samples of untreated cell suspensions were nearly as high as the total FCM counts, and the correlation was strong (r > 0.99). In dairy fermentation starters and in probiotic products the FCM total cell counts were substantially higher than the numbers of CFU. Three functional populations could be distinguished: culturable cells, cells that are intact and metabolically active but not culturable, and permeabilized cells. The proportions of the populations differed in the products tested. This FCM method provides tools to assess the functionality of different populations in fermentation starters and probiotic products. PMID:12039752

Combining laser-assisted microdissection (LAM) and RNA-seq allows to perform a comprehensive transcriptomic analysis of epidermal cells of Arabidopsis embryo.

PubMed

Sakai, Kaori; Taconnat, Ludivine; Borrega, Nero; Yansouni, Jennifer; Brunaud, Véronique; Paysant-Le Roux, Christine; Delannoy, Etienne; Martin Magniette, Marie-Laure; Lepiniec, Loïc; Faure, Jean Denis; Balzergue, Sandrine; Dubreucq, Bertrand

2018-01-01

Genome-wide characterization of tissue- or cell-specific gene expression is a recurrent bottleneck in biology. We have developed a sensitive approach based on ultra-low RNA sequencing coupled to laser assisted microdissection for analyzing different tissues of the small Arabidopsis embryo. We first characterized the number of genes detected according to the quantity of tissue yield and total RNA extracted. Our results revealed that as low as 0.02 mm 2 of tissue and 50 pg of total RNA can be used without compromising the number of genes detected. The optimised protocol was used to compare the epidermal versus mesophyll cell transcriptomes of cotyledons at the torpedo-shaped stage of embryo development. The approach was validated by the recovery of well-known epidermal genes such AtML1 or AtPDF2 and genes involved in flavonoid and cuticular waxes pathways. Moreover, the interest and sensitivity of this approach were highlighted by the characterization of several transcription factors preferentially expressed in epidermal cells. This technical advance unlocks some current limitations of transcriptomic analyses and allows to investigate further and efficiently new biological questions for which only a very small amounts of cells need to be isolated. For instance, it paves the way to increasing the spatial accuracy of regulatory networks in developing small embryo of Arabidopsis or other plant tissues.

Advanced glycation end product-induced astrocytic differentiation of cultured neurospheres through inhibition of Notch-Hes1 pathway-mediated neurogenesis.

PubMed

Guo, Yijing; Wang, Pin; Sun, Haixia; Cai, Rongrong; Xia, Wenqing; Wang, Shaohua

2013-12-23

This study aims to investigate the roles of the Notch-Hes1 pathway in the advanced glycation end product (AGE)-mediated differentiation of neural stem cells (NSCs). We prepared pLentiLox3.7 lentiviral vectors that express short hairpin RNA (shRNA) against Notch1 and transfected it into NSCs. Cell differentiation was analyzed under confocal laser-scanning microscopy. The percentage of neurons and astrocytes was quantified by normalizing the total number of TUJ1+ (Neuron-specific class III β-tubulin) and GFAP+ (Glial fibrillary acidic protein) cells to the total number of Hoechst 33342-labeled cell nuclei. The protein and gene expression of Notch-Hes1 pathway components was examined via western blot analysis and real-time PCR. After 1 week of incubation, we found that AGE-bovine serum albumin (BSA) (400 μg/mL) induced the astrocytic differentiation of cultured neurospheres and inhibited neuronal formation. The expression of Notch-Hes1 pathway components was upregulated in the cells in the AGE-BSA culture medium. Immunoblot analysis indicated that shRNA silencing of Notch1 expression in NSCs significantly increases neurogenesis and suppresses astrocytic differentiation in NSCs incubated with AGE-BSA. AGEs promote the astrocytic differentiation of cultured neurospheres by inhibiting neurogenesis through the Notch-Hes1 pathway, providing a potential therapeutic target for hyperglycemia-related cognitive deficits.

Braided and Stacked Electrospun Nanofibrous Scaffolds for Tendon and Ligament Tissue Engineering

PubMed Central

Rothrauff, Benjamin B.; Lauro, Brian B.; Yang, Guang; Debski, Richard E.; Musahl, Volker

2017-01-01

Tendon and ligament injuries are a persistent orthopedic challenge given their poor innate healing capacity. Nonwoven electrospun nanofibrous scaffolds composed of polyesters have been used to mimic the mechanics and topographical cues of native tendons and ligaments. However, nonwoven nanofibers have several limitations that prevent broader clinical application, including poor cell infiltration, as well as tensile and suture-retention strengths that are inferior to native tissues. In this study, multilayered scaffolds of aligned electrospun nanofibers of two designs–stacked or braided–were fabricated. Mechanical properties, including structural and mechanical properties and suture-retention strength, were determined using acellular scaffolds. Human bone marrow-derived mesenchymal stem cells (MSCs) were seeded on scaffolds for up to 28 days, and assays for tenogenic differentiation, histology, and biochemical composition were performed. Braided scaffolds exhibited improved tensile and suture-retention strengths, but reduced moduli. Both scaffold designs supported expression of tenogenic markers, although the effect was greater on braided scaffolds. Conversely, cell infiltration was superior in stacked constructs, resulting in enhanced cell number, total collagen content, and total sulfated glycosaminoglycan content. However, when normalized against cell number, both designs modulated extracellular matrix protein deposition to a similar degree. Taken together, this study demonstrates that multilayered scaffolds of aligned electrospun nanofibers supported tenogenic differentiation of seeded MSCs, but the macroarchitecture is an important consideration for applications of tendon and ligament tissue engineering. PMID:28071988

Braided and Stacked Electrospun Nanofibrous Scaffolds for Tendon and Ligament Tissue Engineering.

PubMed

Rothrauff, Benjamin B; Lauro, Brian B; Yang, Guang; Debski, Richard E; Musahl, Volker; Tuan, Rocky S

2017-05-01

Tendon and ligament injuries are a persistent orthopedic challenge given their poor innate healing capacity. Nonwoven electrospun nanofibrous scaffolds composed of polyesters have been used to mimic the mechanics and topographical cues of native tendons and ligaments. However, nonwoven nanofibers have several limitations that prevent broader clinical application, including poor cell infiltration, as well as tensile and suture-retention strengths that are inferior to native tissues. In this study, multilayered scaffolds of aligned electrospun nanofibers of two designs-stacked or braided-were fabricated. Mechanical properties, including structural and mechanical properties and suture-retention strength, were determined using acellular scaffolds. Human bone marrow-derived mesenchymal stem cells (MSCs) were seeded on scaffolds for up to 28 days, and assays for tenogenic differentiation, histology, and biochemical composition were performed. Braided scaffolds exhibited improved tensile and suture-retention strengths, but reduced moduli. Both scaffold designs supported expression of tenogenic markers, although the effect was greater on braided scaffolds. Conversely, cell infiltration was superior in stacked constructs, resulting in enhanced cell number, total collagen content, and total sulfated glycosaminoglycan content. However, when normalized against cell number, both designs modulated extracellular matrix protein deposition to a similar degree. Taken together, this study demonstrates that multilayered scaffolds of aligned electrospun nanofibers supported tenogenic differentiation of seeded MSCs, but the macroarchitecture is an important consideration for applications of tendon and ligament tissue engineering.

[Anti-aging action of the total lactones of ginkgo on aging mice].

PubMed

Dong, Liu-yi; Fan, Li; Li, Gui-fang; Guo, Yan; Pan, Jian; Chen, Zhi-wu

2004-03-01

To investigate the effects of total lactones of ginkgo on aging by using D-galactose induced aging mice and natural aging mice. By using D-galactose induced aging mice, to detect the LF content in heart and liver, the Hyp content in liver, the MAO, GSH-Px activities and the NO content in cerebrum. The apoptosis of cerebral cell was determined by terminal deoxy-nucleotidyl transforase-mediated dUTP-digoxigenin nick end-labeling (Tunel) in natural aging mice. TLG was shown to increase the GSH-Px activities, reduce the NO content and decrease the MAO activity in cerebrum. Meanwhile, TLG was found to reduce the LF content in liver and heart and raise the Hyp content in liver. TLG was shown to inhibit apoptosis of cerebral cell and decrease the number of apoptotic cells in the brain. TLG possesses effect on antiaging via attenuating lipid peroxidation and NO and apoptosis of cerebral cells.

An Evaluation of the Impact of Goal Setting and Cell Phone Calls on Juvenile Rearrests.

PubMed

Bahr, Stephen J; Cherrington, David J; Erickson, Lance D

2016-12-01

Using a sample of 256 juvenile offenders who were randomly assigned to treatment or control groups, this study evaluates a cognitive-behavioral program that combines cognitive training, goal setting, and a phone-coach follow-up. The training involved six classroom sessions where participants received instruction and help in creating individualized goals. After attending the classes, participants received automated phone calls twice a day for up to a year. During the year following the program, the treatment and control groups were not significantly different in whether or not they were rearrested or in total rearrests. However, the total number of calls received had a significant negative association with whether or not they were rearrested for a felony and with the total number of felony rearrests.

Inhibition of Aldehyde Dehydrogenase-Activity Expands Multipotent Myeloid Progenitor Cells with Vascular Regenerative Function.

PubMed

Cooper, Tyler T; Sherman, Stephen E; Kuljanin, Miljan; Bell, Gillian I; Lajoie, Gilles A; Hess, David A

2018-05-01

Blood-derived progenitor cell transplantation holds potential for the treatment of severe vascular diseases. Human umbilical cord blood (UCB)-derived hematopoietic progenitor cells purified using high aldehyde dehydrogenase (ALDH hi ) activity demonstrate pro-angiogenic functions following intramuscular (i.m.) transplantation into immunodeficient mice with hind-limb ischemia. Unfortunately, UCB ALDH hi cells are rare and prolonged ex vivo expansion leads to loss of high ALDH-activity and diminished vascular regenerative function. ALDH-activity generates retinoic acid, a potent driver of hematopoietic differentiation, creating a paradoxical challenge to expand UCB ALDH hi cells while limiting differentiation and retaining pro-angiogenic functions. We investigated whether inhibition of ALDH-activity during ex vivo expansion of UCB ALDH hi cells would prevent differentiation and expand progeny that retained pro-angiogenic functions after transplantation into non-obese diabetic/severe combined immunodeficient mice with femoral artery ligation-induced unilateral hind-limb ischemia. Human UCB ALDH hi cells were cultured under serum-free conditions for 9 days, with or without the reversible ALDH-inhibitor, diethylaminobenzaldehyde (DEAB). Although total cell numbers were increased >70-fold, the frequency of cells that retained ALDH hi /CD34+ phenotype was significantly diminished under basal conditions. In contrast, DEAB-inhibition increased total ALDH hi /CD34+ cell number by ≥10-fold, reduced differentiation marker (CD38) expression, and enhanced the retention of multipotent colony-forming cells in vitro. Proteomic analysis revealed that DEAB-treated cells upregulated anti-apoptotic protein expression and diminished production of proteins implicated with megakaryocyte differentiation. The i.m. transplantation of DEAB-treated cells into mice with hind-limb ischemia stimulated endothelial cell proliferation and augmented recovery of hind-limb perfusion. DEAB-inhibition of ALDH-activity delayed hematopoietic differentiation and expanded multipotent myeloid cells that accelerated vascular regeneration following i.m. transplantation in vivo. Stem Cells 2018;36:723-736. © AlphaMed Press 2018.

Lower airways inflammation in patients with ARDS measured using endotracheal aspirates: a pilot study.

PubMed

Spadaro, Savino; Kozhevnikova, Iryna; Casolari, Paolo; Ruggeri, Paolo; Bellini, Tiziana; Ragazzi, Riccardo; Barbieri, Federica; Marangoni, Elisabetta; Caramori, Gaetano; Volta, Carlo Alberto

2017-01-01

Our knowledge of acute respiratory distress syndrome (ARDS) pathogenesis is incomplete. The goal of this pilot study is to investigate the feasibility of measuring lower airways inflammation in patients with ARDS using repeated endotracheal aspirates (ETAs). ETAs were obtained within 24 hours by intensive care unit admission from 25 mechanically ventilated patients with ARDS and 10 of them underwent a second ETA within 96 hours after the first sampling. In each sample, cell viability was assessed using trypan blue exclusion method and the total and differential cell counts were measured using Neubauer-improved cell counting chamber and cytospins stained with Diff-Quik. The median cell viability was 89 (IQR 80-93)%, with a median total cell count of 305 (IQR 130-1270)×10 3 /mL and a median macrophage, neutrophil, lymphocyte and eosinophil count, respectively, of 19.8 (IQR 5.4-71.6)×10 3 /mL; 279 (IQR 109-1213)×10 3 /mL; 0 (IQR 0-0.188)×10 3 /mL; 0 (IQR 0-1.050)×10 3 /mL. Eosinophil count in the ETA correlated with the number of blood eosinophils (r=0.4840, p=0.0142). Cell viability and total and differential cell counts were neither significantly different in the second ETA compared with the first ETA nor were unaffected by the presence or absence of bacteria in the blood and/or ETA, or by the ARDS aetiology, apart from the macrophage count which was significantly increased in patients with ARDS associated with acute pancreatitis compared with those associated with pneumonia (p=0.0143). ETA can be used to investigate the cellularity of the lower airways in patients with ARDS and it is an easy-to-perform and non-invasive procedure. Eosinophil counts in ETA and blood are significantly correlated. The number of macrophages in ETA may be affected by the aetiology of the ARDS.

Exogenous proline enhances the sensitivity of Tobacco BY-2 cells to arsenate.

PubMed

Nahar, Mst Nur-E-Nazmun; Islam, Mohammad Muzahidul; Hoque, Md Anamul; Yonezawa, Anna; Prodhan, Md Yeasin; Nakamura, Toshiyuki; Nakamura, Yoshimasa; Munemasa, Shintaro; Murata, Yoshiyuki

2017-09-01

Arsenic causes physiological and structural disorders in plants. Proline is accumulated as a compatible solute in plants under various stress conditions and mitigates stresses. Here, we investigated the effects of exogenous proline on tobacco Bright Yellow-2 (BY-2) cultured cells under [Formula: see text] stress. Arsenate did not inhibit BY-2 cell growth at 40 and 50 μM but did it at 60 μM. Proline at 0.5 to 10 mM did not affect the cell growth but delayed it at 20 mM. At 40 μM [Formula: see text], neither 0.5 mM nor 1 mM proline affected the cell growth but 10 mM proline inhibited it. In the presence of [Formula: see text], 10 mM proline increased the number of Evans Blue-stained (dead) cells and decreased the number of total cells. Together, our results suggest that exogenous proline does not alleviate arsenate toxicity but enhances the sensitivity of BY-2 cells to arsenate.

Effects of limonene stress on the growth of and microcystin release by the freshwater cyanobacterium Microcystis aeruginosa FACHB-905.

PubMed

Hu, Xi; Liu, Yunguo; Zeng, Guangming; Hu, Xinjiang; Wang, Yaqin; Zeng, Xiaoxia

2014-07-01

The effects of limonene exposure on the growth of Microcystisaeruginosa and the release of toxic intracellular microcystin (MCY) were tested by evaluating the results obtained from the batch culture experiments with M. aeruginosa FACHB-905. The time series of cell as well as intracellular and extracellular MCY concentrations were evaluated during 5d of the incubation. After exposure to limonene, the number of cells gradually diminished; the net log cell reduction after 5d of the exposure was 3.0, 3.6, and 3.8log when the initial cell densities were set at 1.6×10(7), 1.1×10(6) and 4.1×10(5)cell/mL, respectively. Limonene was found to significantly influence the production and release of MCY. As the limonene exposure could inhibit the increase in the number of cells, the increase in the total MCY concentration in the medium was also inhibited. In the presence of limonene, the intracellular MCY was gradually released into the medium through a gradual reduction in the number of cells. The extracellular MCY concentration in the medium was significantly higher in the limonene-exposed samples than in the control samples, which confirmed that limonene cannot decompose the extracellular MCY. Copyright © 2014 Elsevier Inc. All rights reserved.

Protective effects on vascular endothelial cell in N'-nitro-L-arginine (L-NNA)-induced hypertensive rats from the combination of effective components of Uncaria rhynchophylla and Semen Raphani.

PubMed

Li, Yunlun; Yang, Wenqing; Zhu, Qingjun; Yang, Jinguo; Wang, Zhen

2015-08-01

Endothelial dysfunction is closely associated with hypertension. Protection of vascular endothelial cell is the key to prevention and treatment of hypertension. Uncaria rhynchophylla total alkaloids and Semen Raphani soluble alkaloid, isolated from traditional Chinese medicine Uncaria rbyncbopbylla and Semen Raphani respectively, exhibit properties of anti-hypertension and protection of blood vessels. In the present study, we observed the protective effect of the combined use of Uncaria rhynchophylla total alkaloids and Semen Raphani soluble alkaloid to the vascular endothelial cell in N'-nitro-L-arginine-induced hypertensive rats and investigate the preliminary mechanism. Blood pressure was detected by non-invasive rats tail method to observe the anti-hypertension effect of drugs. Scanning electron microscopy was used to observe the integrity or shedding state of vascular endothelial cell. The amount of circulating endothelial cells and CD54 and CD62P expression on circulating endothelial cells were tested to evaluate the endothelium function. In this study, we found that the Uncaria rhynchophylla total alkaloids and Semen Raphani soluble alkaloid compatibility can effectively lower the blood pressure, improve the structural integrity of vascular endothelium, and significantly reduce the number of circulating endothelial cells. Furthermore, the mean fluorescence intensity of CD54 and CD62P expressed showed decrease after the intervention of Uncaria rhynchophylla total alkaloids and Semen Raphani soluble alkaloid compatibility. In conclusion, the combination of effective components of the Uncaria rhynchophylla total alkaloids and Semen Raphani soluble alkaloid demonstrated good antihypertension effect and vascular endothelium protective effect. The preliminary mechanism of the protective effect may attribute to relieve the overall low-grade inflammation.

General statistics of stochastic process of gene expression in eukaryotic cells.

PubMed Central

Kuznetsov, V A; Knott, G D; Bonner, R F

2002-01-01

Thousands of genes are expressed at such very low levels (< or =1 copy per cell) that global gene expression analysis of rarer transcripts remains problematic. Ambiguity in identification of rarer transcripts creates considerable uncertainty in fundamental questions such as the total number of genes expressed in an organism and the biological significance of rarer transcripts. Knowing the distribution of the true number of genes expressed at each level and the corresponding gene expression level probability function (GELPF) could help resolve these uncertainties. We found that all observed large-scale gene expression data sets in yeast, mouse, and human cells follow a Pareto-like distribution model skewed by many low-abundance transcripts. A novel stochastic model of the gene expression process predicts the universality of the GELPF both across different cell types within a multicellular organism and across different organisms. This model allows us to predict the frequency distribution of all gene expression levels within a single cell and to estimate the number of expressed genes in a single cell and in a population of cells. A random "basal" transcription mechanism for protein-coding genes in all or almost all eukaryotic cell types is predicted. This fundamental mechanism might enhance the expression of rarely expressed genes and, thus, provide a basic level of phenotypic diversity, adaptability, and random monoallelic expression in cell populations. PMID:12136033

Ontogeny of serotonin-immunoreactive cells in the gut epithelium of the cloudy dogfish, Scyliorhinus torazame, with reference to coexistence of serotonin and neuropeptide Y.

PubMed

Chiba, A

1998-09-01

The ontogeny of serotonin (5-HT)-immunoreactive (IR) cells in the gut epithelium of an oviparous elasmobranch, Scyliorhinus torazame, was examined immunohistochemically. 5-HT-IR cells first appeared in the proximal part of the vitellointestinal duct (VID) and in the anterior part of the midgut of the embryo (30 mm in total length). At the 40-mm stage, the cells slightly increased in number and spread to the posterior part of the midgut, but no labeled cells were found in the foregut or hindgut. By the late embryonic (74- and 80-mm) stages, 5-HT-IR cells were markedly increased in number in the spiral intestine and stomach, whereas they were few in the VID and rectum. During these stages, the density of the cells in the inner yolk sac, the derivative of the VID, tended to be increased. This tendency seemed to be consistent in the posthatching juveniles at the 95-mm stage. In juveniles, 125 mm in length and 1.7 months after hatching, the cells further increased in number in the spiral intestine, reaching their adult value. Double immunostaining by the use of anti-5-HT and -neuropeptide Y (NPY) antisera demonstrated that some of the 5-HT-IR cells were also positive for NPY. Copyright 1998 Academic Press.

The Isolation and Enrichment of Large Numbers of Highly Purified Mouse Spleen Dendritic Cell Populations and Their In Vitro Equivalents.

PubMed

Vremec, David

2016-01-01

Dendritic cells (DCs) form a complex network of cells that initiate and orchestrate immune responses against a vast array of pathogenic challenges. Developmentally and functionally distinct DC subtypes differentially regulate T-cell function. Importantly it is the ability of DC to capture and process antigen, whether from pathogens, vaccines, or self-components, and present it to naive T cells that is the key to their ability to initiate an immune response. Our typical isolation procedure for DC from murine spleen was designed to efficiently extract all DC subtypes, without bias and without alteration to their in vivo phenotype, and involves a short collagenase digestion of the tissue, followed by selection for cells of light density and finally negative selection for DC. The isolation procedure can accommodate DC numbers that have been artificially increased via administration of fms-like tyrosine kinase 3 ligand (Flt3L), either directly through a series of subcutaneous injections or by seeding with an Flt3L secreting murine melanoma. Flt3L may also be added to bone marrow cultures to produce large numbers of in vitro equivalents of the spleen DC subsets. Total DC, or their subsets, may be further purified using immunofluorescent labeling and flow cytometric cell sorting. Cell sorting may be completely bypassed by separating DC subsets using a combination of fluorescent antibody labeling and anti-fluorochrome magnetic beads. Our procedure enables efficient separation of the distinct DC subsets, even in cases where mouse numbers or flow cytometric cell sorting time is limiting.

Assessment of Regenerative Capacity in the Dolphin

DTIC Science & Technology

2010-10-10

liposuction ; cells released during the digestion of the adipose tissue were analyzed for cytology, assayed for the total number of colony-forming cells... liposuction was used to harvest subcutaneous adipose from the nuchal pad of six dolphins at the Navy Marine Mammal Program (NMMP). Adipose samples...and aid in the placement of the liposuction cannula in the adipose depot. The difference between the various layers of muscle and adipose tissue

Photoperiod affects the expression of sex and species differences in leukocyte number and leukocyte trafficking in congeneric hamsters.

PubMed

Bilbo, S D; Dhabhar, F S; Viswanathan, K; Saul, A; Nelson, R J

2003-11-01

Sex differences in immune function are well documented. These sex differences may be modulated by social and environmental factors. Individuals of polygynous species generally exhibit more pronounced sex differences in immune parameters than individuals of monogamous species, often displaying an energetic trade-off between enhanced immunity and high mating success. During winter, animals contend with environmental conditions (e.g. low temperatures and decreased food availability) that evoke energetic-stress responses; many mammals restrict reproduction in response to photoperiod as part of an annual winter coping strategy. To test the hypothesis that extant sex and species differences in immune surveillance may be modulated by photoperiod, we examined leukocyte numbers in males and females of two closely related hamster species (Phodopus). As predicted, uniparental P. sungorus exhibited a robust sex difference, with total white blood cells, total lymphocytes, T cells, and B cells higher in females than males, during long days when reproduction occurs, but not during short days when reproduction usually stops. In contrast, biparental male and female P. campbelli exhibited comparable leukocyte numbers during both long and short days. To study sex differences in stress responses, we also examined immune cell trafficking in response to an acute (2 h) restraint stressor. During stressful challenges, it appears beneficial for immune cells to exit the blood and move to primary immune defense areas such as the skin, in preparation for potential injury or infection. Acute stress moved lymphocytes and monocytes out of the blood in all animals. Blood cortisol concentrations were increased in P. sungorus females compared to males at baseline (52%) and in response to restraint stress (38%), but only in long days. P. campbelli males and females exhibited comparable blood cortisol and stress responses during both long and short days. Our results suggest that interactions among social factors and the environment play a significant role in modulating sex and seasonal alterations in leukocyte numbers and stress responses.

Type 2 porcine reproductive and respiratory syndrome virus infection increases apoptosis at the maternal-fetal interface in late gestation pregnant gilts

PubMed Central

Harding, John C. S.; Al-Dissi, Ahmad N; Detmer, Susan E.

2017-01-01

The pathogenesis of fetal death associated with porcine reproductive and respiratory syndrome (PRRS) is hypothesized to be a consequence of PRRS virus-induced apoptosis at the maternal-fetal interface (MFI). The objectives of this study were to evaluate distribution and degree of apoptosis in the uterine and fetal placental tissues during the experimental type 2 PRRS virus (PRRSV) infection and determine associations between apoptosis at the MFI, PRRSV RNA concentration and antigen staining intensity, PRRSV-induced microscopic lesions, and fetal preservation status. A total of 114 naïve, high-health pregnant gilts were inoculated with type 2 PRRSV on gestation day 85±1 with euthanasia 21 days later; 19 sham-inoculated gilts served as controls. Two hundred and fifty samples of uterine tissue with fetal placenta were selected based on negative, low PRRSV RNA, and high PRRSV RNA concentration (0, < or > 2.7 log10 copies/mg, respectively). TUNEL assay was used to detect apoptosis in the endometrium and at the MFI. PRRSV RNA concentration and numbers of PRRSV immunopositive cells in uterine and placental tissue were positively associated with the severity of apoptosis in the endometrium and the MFI (P<0.001, P<0.05 and P<0.001, respectively). The number of TUNEL positive cells at the MFI was also positively associated with the severity (P<0.001) of vasculitis, but not total numbers of inflammatory cells in the endometrium. Increased numbers of TUNEL positive cells at the MFI were associated with PRRSV load in the fetal thymus, and greater odds of meconium staining of the fetus at 21 days post infection (P<0.001 for both). These findings suggest an important role of apoptosis in the pathogenesis of uterine epithelial and trophoblastic cell death at the MFI. Moreover, apoptosis at the MFI is significantly associated with fetal demise during in utero type 2 PRRSV infection. PMID:28253336

Quantitative Assessment of Proliferative Effects of Oral Vanadium on Pancreatic Islet Volumes and Beta Cell Numbers of Diabetic Rats

PubMed Central

Pirmoradi, Leila; Noorafshan, Ali; Safaee, Akbar; Dehghani, Gholam Abbas

2016-01-01

Background: Oral vanadyl sulfate (vanadium) induces normoglycemia, proliferates beta cells and prevents pancreatic islet atrophy in streptozotocin-induced diabetic rats. Soteriological method is used to quantitate the proliferative effects of vanadium on beta-cell numbers and islet volumes of normal and diabetic rats. Methods: Adult male Sprague-Dawley rats were made diabetic with intravenous streptozotocin injection (40 mg/kg). Normal and diabetic rats were divided into four groups. While control normal and diabetic (CD) groups used water, vanadium-treated normal (VTN) and diabetic (VTD) groups used solutions containing vanadyl sulfate (0.5-1 mg/mL, VOSO4+5H2O). Tail blood samples were used to measure blood glucose (BG) and plasma insulin. Two months after treatment, rats were sacrificed, pancreata prepared, and stereology method was used to quantitatively evaluate total beta cell numbers (TBCN) and total islet volumes (TISVOL). Results: Normoglycemia persisted in VTN with significantly decreased plasma insulin (0.190.08 vs. 0.970.27 ng/dL, P<0.002). The respective high BG (53249 vs. 14446 mg/dL, P<0.0001) and reduced plasma insulin (0.260.15 vs. 0.540.19 ng/dL, P<0.002) seen in CD were reversed in VTD during vanadium treatment or withdrawal. While the induction of diabetes, compared to their control, significantly decreased TISVOL (1.90.2 vs. 3.030.6 mm3, P<0.003) and TBCN (0.990.1 vs. 3.20.2 x 106, P<0.003), vanadium treatment significantly increased TISVOL (2.90.8 and 4.071.0 mm3, P<0.003) and TBCN (1.50.3 and 3.80.6 x 106, P<0.03). Conclusion: Two-month oral vanadium therapy in STZ-diabetic rats ameliorated hyperglycemia by partially restoring plasma insulin. This action was through proliferative actions of vanadium in preventing islet atrophy by increasing beta-cell numbers. PMID:26459400

The effects of perfluorinated chemicals on adipocyte differentiation in vitro.

PubMed

Watkins, Andrew M; Wood, Carmen R; Lin, Mimi T; Abbott, Barbara D

2015-01-15

The 3T3-L1 preadipocyte culture system has been used to examine numerous compounds that influence adipocyte differentiation or function. The perfluoroalkyl acids (PFAAs), used as surfactants in a variety of industrial applications, are of concern as environmental contaminants that are detected worldwide in human serum and animal tissues. This study was designed to evaluate the potential for PFAAs to affect adipocyte differentiation and lipid accumulation using mouse 3T3-L1 cells. Cells were treated with perfluorooctanoic acid (PFOA) (5-100 µM), perfluorononanoic acid (PFNA) (5-100 µM), perfluorooctane sulfonate (PFOS) (50-300 µM), perfluorohexane sulfonate (PFHxS) (40-250 µM), the peroxisome proliferator activated receptor (PPAR) PPARα agonist Wyeth-14,643 (WY-14,643), and the PPARγ agonist rosiglitazone. The PPARγ agonist was included as a positive control as this pathway is critical to adipocyte differentiation. The PPARα agonist was included as the PFAA compounds are known activators of this pathway. Cells were assessed morphometrically and biochemically for number, size, and lipid content. RNA was extracted for qPCR analysis of 13 genes selected for their importance in adipocyte differentiation and lipid metabolism. There was a significant concentration-related increase in cell number and decreased cell size after exposure to PFOA, PFHxS, PFOS, and PFNA. All four PFAA treatments produced a concentration-related decrease in the calculated average area occupied by lipid per cell. However, total triglyceride levels per well increased with a concentration-related trend for all compounds, likely due to the increased cell number. Expression of mRNA for the selected genes was affected by all exposures and the specific impacts depended on the particular compound and concentration. Acox1 and Gapdh were upregulated by all six compounds. The strongest overall effect was a nearly 10-fold induction of Scd1 by PFHxS. The sulfonated PFAAs produced numerous, strong changes in gene expression similar to the effects after treatment with the PPARγ agonist rosiglitazone. By comparison, the effects on gene expression were muted for the carboxylated PFAAs and for the PPARα agonist WY-14,643. In summary, all perfluorinated compounds increased cell number, decreased cell size, increased total triglyceride, and altered expression of genes associated with adipocyte differentiation and lipid metabolism. Published by Elsevier Ireland Ltd.

Morphological Changes of Myoepithelial Cells in the Rat Submandibular Gland Following the Application of Surgical Stimuli.

PubMed

Kawabe, Yoshihiro; Mizobe, Kenich; Bando, Yasuhiko; Sakiyama, Koji; Taira, Fuyoko; Tomomura, Akito; Araki, Hisao; Amano, Osamu

2016-12-28

Myoepithelial cells (MECs) exist on the basal surface of acini in major exocrine glands, include myofilaments and various constructive proteins, and share characteristics with smooth muscle and epithelial cells. MECs project several ramified processes to invest acini, and possibly contract to compress acini to support the secretion by the glandular cells. However, the functional roles of MECs in salivary secretion are still unclear. We investigated morphological changes in immunostained MECs using the anti-α-smooth muscle actin (αSMA) antibody in operated or non-operated contralateral (NC) submandibular glands after partial or total resection. Furthermore, we investigated and discuss other salivary glands of rats. MECs in the parotid, sublingual and submandibular gland of adult rats exhibited different shapes and localizations. After surgery, in both operated and NC glands, the number of MECs and αSMA-immunopositive areas increased significantly. Three-dimensional analysis using a confocal laser-scanning microscope revealed that substantial and significant enhancement became evident in the number, length, and thickness of MEC-processes covering acini of the operated and NC submandibular glands. The preset findings indicate that MECs alter the morphology of their processes in operated and NC glands after surgery of the partial or total resection. It is suggested that MECs promote salivary secretion using elongated, thickened, and more ramified processes.

Structural convergence properties of amorphous InGaZnO4 from simulated liquid-quench methods.

PubMed

Buchanan, Jacob C; Fast, Dylan B; Hanken, Benjamin E; Mustard, Thomas J L; Laurita, Geneva; Chiang, Tsung-Han; Keszler, Douglas A; Subramanian, Mas A; Wager, John F; Dolgos, Michelle R; Rustad, James R; Cheong, Paul Ha-Yeon

2017-11-14

The study of structural properties of amorphous structures is complicated by the lack of long-range order and necessitates the use of both cutting-edge computer modeling and experimental techniques. With regards to the computer modeling, many questions on convergence arise when trying to assess the accuracy of a simulated system. What cell size maximizes the accuracy while remaining computationally efficient? More importantly, does averaging multiple smaller cells adequately describe features found in bulk amorphous materials? How small is too small? The aims of this work are: (1) to report a newly developed set of pair potentials for InGaZnO 4 and (2) to explore the effects of structural parameters such as simulation cell size and numbers on the structural convergence of amorphous InGaZnO 4 . The total number of formula units considered over all runs is found to be the critical factor in convergence as long as the cell considered contains a minimum of circa fifteen formula units. There is qualitative agreement between these simulations and X-ray total scattering data - peak trends and locations are consistently reproduced while intensities are weaker. These new IGZO pair potentials are a valuable starting point for future structural refinement efforts.

Adenylate Energy Pool and Energy Charge in Maturing Rape Seeds 1

PubMed Central

Ching, Te May; Crane, Jim M.; Stamp, David L.

1974-01-01

A study of energy state and chemical composition of pod walls and seeds of maturing rape (Brassica napus L.) was conducted on two varieties, Victor and Gorczanski. Total adenosine phosphates, ATP, and adenylate energy charge increased with increasing cell number and cellular synthesis during the early stages, remained high at maximum dry weight accumulation and maximum substrate influx time, and decreased with ripening. A temporal control of energy supply and ATP concentration is evident in developing tissues with determined functions; whereas the association of a high energy charge and active cellular biosynthesis occurs only in tissues with a stabilized cell number. PMID:16658964

Development of a new score to estimate clinical East Coast Fever in experimentally infected cattle.

PubMed

Schetters, Th P M; Arts, G; Niessen, R; Schaap, D

2010-02-10

East Coast Fever is a tick-transmitted disease in cattle caused by Theileria parva protozoan parasites. Quantification of the clinical disease can be done by determining a number of variables, derived from parasitological, haematological and rectal temperature measurements as described by Rowlands et al. (2000). From a total of 13 parameters a single ECF-score is calculated that allows categorization of infected cattle in five different classes that correlate with the severity of clinical signs. This score is complicated not only by the fact that it requires estimation of 13 parameters but also because of the subsequent mathematics. The fact that the values are normalised over a range of 0-10 for each experiment makes it impossible to compare results from different experiments. Here we present an alternative score based on the packed cell volume and the number of piroplasms in the circulation and that is calculated using a simple equation; ECF-score=PCV(relday0)/log(PE+10). In this equation the packed cell volume is expressed as a value relative to that of the day on infection (PCV(relday0)) and the number of piroplasms is expressed as the logarithmic value of the number of infected red blood cells (=PE) in a total of 1000 red blood cells. To allow PE to be 0, +10 is added in the denominator. We analysed a data set of 54 cattle from a previous experiment and found a statistically significant linear correlation between the ECF-score value reached during the post-infection period and the Rowlands' score value. The new score is much more practical than the Rowlands score as it only requires daily blood sampling. From these blood samples both PCV and number of piroplasms can be determined, and the score can be calculated daily. This allows monitoring the development of ECF after infection, which was hitherto not possible. In addition, the new score allows for easy comparison of results from different experiments.

Sertoli cell androgen receptor expression regulates temporal fetal and adult Leydig cell differentiation, function, and population size.

PubMed

Hazra, Rasmani; Jimenez, Mark; Desai, Reena; Handelsman, David J; Allan, Charles M

2013-09-01

We recently created a mouse model displaying precocious Sertoli cell (SC) and spermatogenic development induced by SC-specific transgenic androgen receptor expression (TgSCAR). Here we reveal that TgSCAR regulates the development, function, and absolute number of Leydig cells (LCs). Total fetal and adult type LC numbers were reduced in postnatal and adult TgSCAR vs control testes, despite normal circulating LH levels. Normal LC to SC ratios found in TgSCAR testes indicate that SC androgen receptor (SCAR)-mediated activity confers a quorum-dependent relationship between total SC and LC numbers. TgSCAR enhanced LC differentiation, shown by elevated ratios of advanced to immature LC types, and reduced LC proliferation in postnatal TgSCAR vs control testes. Postnatal TgSCAR testes displayed up-regulated expression of coupled ligand-receptor transcripts (Amh-Amhr2, Dhh-Ptch1, Pdgfa-Pdgfra) for potential SCAR-stimulated paracrine pathways, which may coordinate LC differentiation. Neonatal TgSCAR testes displayed normal T and dihydrotestosterone levels despite differential changes to steroidogenic gene expression, with down-regulated Star, Cyp11a1, and Cyp17a1 expression contrasting with up-regulated Hsd3b1, Hsd17b3, and Srd5a1 expression. TgSCAR males also displayed elevated postnatal and normal adult serum testosterone levels, despite reduced LC numbers. Enhanced adult-type LC steroidogenic output was revealed by increased pubertal testicular T, dihydrotestosterone, 3α-diol and 3β-diol levels per LC and up-regulated steroidogenic gene (Nr5a1, Lhr, Cyp11a1, Cyp17a1, Hsd3b6, Srd5a1) expression in pubertal or adult TgSCAR vs control males, suggesting regulatory mechanisms maintain androgen levels independently of absolute LC numbers. Our unique gain-of-function TgSCAR model has revealed that SCAR activity controls temporal LC differentiation, steroidogenic function, and population size.

Tandem repeats of the 5' non-transcribed spacer of Tetrahymena rDNA function as high copy number autonomous replicons in the macronucleus but do not prevent rRNA gene dosage regulation.

PubMed Central

Pan, W J; Blackburn, E H

1995-01-01

The rRNA genes in the somatic macronucleus of Tetrahymena thermophila are normally on 21 kb linear palindromic molecules (rDNA). We examined the effect on rRNA gene dosage of transforming T.thermophila macronuclei with plasmid constructs containing a pair of tandemly repeated rDNA replication origin regions unlinked to the rRNA gene. A significant proportion of the plasmid sequences were maintained as high copy circular molecules, eventually consisting solely of tandem arrays of origin regions. As reported previously for cells transformed by a construct in which the same tandem rDNA origins were linked to the rRNA gene [Yu, G.-L. and Blackburn, E. H. (1990) Mol. Cell. Biol., 10, 2070-2080], origin sequences recombined to form linear molecules bearing several tandem repeats of the origin region, as well as rRNA genes. The total number of rDNA origin sequences eventually exceeded rRNA gene copies by approximately 20- to 40-fold and the number of circular replicons carrying only rDNA origin sequences exceeded rRNA gene copies by 2- to 3-fold. However, the rRNA gene dosage was unchanged. Hence, simply monitoring the total number of rDNA origin regions is not sufficient to regulate rRNA gene copy number. Images PMID:7784211

A vegan or vegetarian diet substantially alters the human colonic faecal microbiota.

PubMed

Zimmer, J; Lange, B; Frick, J-S; Sauer, H; Zimmermann, K; Schwiertz, A; Rusch, K; Klosterhalfen, S; Enck, P

2012-01-01

Consisting of ≈10(14) microbial cells, the intestinal microbiota represents the largest and the most complex microbial community inhabiting the human body. However, the influence of regular diets on the microbiota is widely unknown. We examined faecal samples of vegetarians (n=144), vegans (n=105) and an equal number of control subjects consuming ordinary omnivorous diet who were matched for age and gender. We used classical bacteriological isolation, identification and enumeration of the main anaerobic and aerobic bacterial genera and computed absolute and relative numbers that were compared between groups. Total counts of Bacteroides spp., Bifidobacterium spp., Escherichia coli and Enterobacteriaceae spp. were significantly lower (P=0.001, P=0.002, P=0.006 and P=0.008, respectively) in vegan samples than in controls, whereas others (E. coli biovars, Klebsiella spp., Enterobacter spp., other Enterobacteriaceae, Enterococcus spp., Lactobacillus spp., Citrobacter spp. and Clostridium spp.) were not. Subjects on a vegetarian diet ranked between vegans and controls. The total microbial count did not differ between the groups. In addition, subjects on a vegan or vegetarian diet showed significantly (P=0.0001) lower stool pH than did controls, and stool pH and counts of E. coli and Enterobacteriaceae were significantly correlated across all subgroups. Maintaining a strict vegan or vegetarian diet results in a significant shift in the microbiota while total cell numbers remain unaltered.

Total neuron numbers in CA1-4 sectors of the dog hippocampus.

PubMed

Rağbetli, Murat Cetin; Aydinlioğlu, Atif; Koyun, Necat; Yayici, Recep; Arslan, Kadir

2010-06-01

Early reports addressed morphological asymmetry in the cross-sectional width of the rat hippocampus. The present study was aimed at counting total number of neurons in CA1-4 sectors and the subiculum of the dog hippocampus as well as investigating possible left /right and male/female asymmetry. Adult mongrel dogs (8 female and 5 male) were assessed by the right and left pawedness and sacrificed by exsanguinations. In each hippocampus dissected, the total neuron numbers of CAs and subiculum were estimated by the physical fractioning method. Significant hemispheric asymmetries were found in the number of pyramidal cells of CA1, CA3/2, CA4 and the subiculum. Sex difference was also found in the subiculum, in favour of the males. Our study indicated a left dominant asymmetry in males and right dominancy in females as well as no functional asymmetry in specific regions of the dog hippocampus. Further investigations are necessary to verify the hypothesis that hippocampal morphological asymmetries in normal subjects are functionally related in memory or in cognitive skills.

Question 7: Biosynthesis of Phosphatidic Acid in Liposome Compartments Toward the Self-Reproduction of Minimal Cells

NASA Astrophysics Data System (ADS)

Kuruma, Yutetsu

2007-10-01

Self-reproduction is one of main properties that define living cells. In order to explore the self-reproduction process for the study of early cells, and to develop a research line somehow connected to the origin of life, we have built up a constructive ‘synthetic cells (minimal cells)’ approach. The minimal cells approach consists in the investigation of the minimal number of elements to accomplish simple cell-like processes like self-reproduction. Such approach belongs to the field of synthetic biology. The minimal cells are reconstructed from a totally reconstituted cell-free protein synthesis system (PURESYSTEM) and liposome compartments as containers. Based on this approach, we synthesized two membrane proteins (enzymes), GPAT and LPAAT, which are involved in the phosphatidic acid biosynthesis in bacteria. Both membrane proteins were successfully synthesized by PURESYSTEM encapsulated inside POPC liposomes. Additionally, the enzymatic activity of GPAT was restored by mixing the expressed enzyme with lipid and by forming liposomes in situ. Through these experimental evidences, here we present a possible model to achieve self-reproduction in minimal cells. Our results would contribute to the idea that early cells could have been built by an extremely small number of genes.

Tbo-Filgrastim versus Filgrastim during Mobilization and Neutrophil Engraftment for Autologous Stem Cell Transplantation.

PubMed

Elayan, Mohammed M; Horowitz, Justin G; Magraner, Jose M; Shaughnessy, Paul J; Bachier, Carlos

2015-11-01

There are limited data available supporting the use of the recombinant granulocyte colony-stimulating factor (G-CSF), tbo-filgrastim, rather than traditionally used filgrastim to mobilize peripheral blood stem cells (PBSC) or to accelerate engraftment after autologous stem cell transplantation (ASCT). We sought to compare the efficacy and cost of tbo-filgrastim to filgrastim in these settings. Patients diagnosed with lymphoma or plasma cell disorders undergoing G-CSF mobilization, with or without plerixafor, were included in this retrospective analysis. The primary outcome was total collected CD34(+) cells/kg. Secondary mobilization endpoints included peripheral CD34(+) cells/μL on days 4 and 5 of mobilization, adjunctive use of plerixafor, CD34(+) cells/kg collected on day 5, number of collection days and volumes processed, number of collections reaching 5 million CD34(+) cells/kg, and percent reaching target collection goal in 1 day. Secondary engraftment endpoints included time to neutrophil and platelet engraftment, number of blood product transfusions required before engraftment, events of febrile neutropenia, and length of stay. A total of 185 patients were included in the final analysis. Patients receiving filgrastim (n = 86) collected a median of 5.56 × 10(6) CD34(+) cells/kg, compared with a median of 5.85 × 10(6) CD34(+) cells/kg in the tbo-filgrastim group (n = 99; P = .58). There were no statistically significant differences in all secondary endpoints with the exception of apheresis volumes processed (tbo-filgrastim, 17.0 liters versus filgrastim, 19.7 liters; P < .01) and mean platelet transfusions (tbo-filgrastim, 1.7 units versus filgrastim, 1.4 units; P = .04). In conclusion, tbo-filgrastim demonstrated similar CD34(+) yield compared with filgrastim in mobilization and post-transplantation settings, with no clinically meaningful differences in secondary efficacy and safety endpoints. Furthermore, tbo-filgrastim utilization was associated with cost savings of approximately $1406 per patient utilizing average wholesale price. Copyright © 2015 American Society for Blood and Marrow Transplantation. Published by Elsevier Inc. All rights reserved.

Aging-associated shifts in functional status of mast cells located by adult and aged mesenteric lymphatic vessels

PubMed Central

Chatterjee, Victor

2012-01-01

We had previously proposed the presence of permanent stimulatory influences in the tissue microenvironment surrounding the aged mesenteric lymphatic vessels (MLV), which influence aged lymphatic function. In this study, we performed immunohistochemical labeling of proteins known to be present in mast cells (mast cell tryptase, c-kit, prostaglandin D2 synthase, histidine decarboxylase, histamine, transmembrane protein 16A, and TNF-α) with double verification of mast cells in the same segment of rat mesentery containing MLV by labeling with Alexa Fluor 488-conjugated avidin followed by toluidine blue staining. Additionally, we evaluated the aging-associated changes in the number of mast cells located by MLV and in their functional status by inducing mast cell activation by various activators (substance P; anti-rat DNP Immunoglobulin E; peptidoglycan from Staphyloccus aureus and compound 48/80) in the presence of ruthenium red followed by subsequent staining by toluidine blue. We found that there was a 27% aging-associated increase in the total number of mast cells, with an ∼400% increase in the number of activated mast cells in aged mesenteric tissue in resting conditions with diminished ability of mast cells to be newly activated in the presence of inflammatory or chemical stimuli. We conclude that higher degree of preactivation of mast cells in aged mesenteric tissue is important for development of aging-associated impairment of function of mesenteric lymphatic vessels. The limited number of intact aged mast cells located close to the mesenteric lymphatic compartments to react to the presence of acute stimuli may be considered contributory to the aging-associated deteriorations in immune response. PMID:22796537

Culture of bovine embryos on a polydimethylsiloxane (PDMS) microwell plate.

PubMed

Akagi, Satoshi; Hosoe, Misa; Matsukawa, Kazutsugu; Ichikawa, Akihiko; Tanikawa, Tamio; Takahashi, Seiya

2010-08-01

We fabricated a polydimethylsiloxane (PDMS)-based microwell plate (PDMS-MP) containing 100 microwells with a rounded bottom and examined whether it can be used for culture of individual in vitro fertilized (IVF) embryos or parthenogenetically activated zona-free embryos in cattle. In Experiment 1, we examined the in vitro developmental ability of IVF embryos cultured individually on PDMS-MP. After IVF, 20 embryos were transferred into 100 microl drops on PDMS-MP and cultured individually in each well of PDMS-MP (PDMS group). After 7 days of culture, the embryos in the PDMS group developed to the blastocyst stage at the same rate of those in the control group cultured in a group of 20 embryos without PDMS-MP. There were no differences in total number of cells and the ratio of inner cell mass to total cells between the PDMS and control groups. In Experiment 2, we examined the in vitro developmental ability of parthenogenetically activated zona-free bovine embryos cultured individually on PDMS-MP. The zona-free embryos were cultured individually in each well of a PDMS-MP or in each well produced by pressing a darning needle onto the bottom of a culture dish (WOW group). After 7 days of culture, the blastocyst formation rate and cell number of blastocysts in the PDMS group did not differ from those of the zona-intact embryos in the control group. Also, there were no differences in the blastocyst formation rate and cell number of blastocysts between the WOW and PDMS groups. These results suggest that the culture system using PDMS-MP is useful for individual embryos or zona-free embryos in cattle.

Therapeutic Effect of External Application of Ligustrazine Combined with Holistic Nursing on Pressure Sores.

PubMed

Niu, Junzhi; Han, Lin; Gong, Fen

2016-08-15

BACKGROUND This study aimed to explore the therapeutic effect of external application of ligustrazine combined with holistic nursing on pressure sores, as well as the underlying mechanism. MATERIAL AND METHODS From February 2014 to March 2015, a total of 32 patients with Phase II and Phase III pressure sores were enrolled and randomly assigned to an experimental group or a control group. The clinical data were comparable between the 2 groups. In addition to holistic nursing, the patients in the experimental group received 4 weeks of continuous external application of ligustrazine, whereas patients in the control group received compound clotrimazole cream. Therapeutic effect and healing time were recorded. HaCaT cells were used as an in vitro model for mechanism analysis of the effect of ligustrazine in treating pressure sores. After culturing with different concentrations of ligustrazine or the inhibitor of AKT (LY294002) for 72 h, cell viability, clone formation numbers, and levels of phosphatidyl inositol 3-kinase (PI3K), p-AKT, and p-mammalian target of rapamycin (mTOR) were determined. RESULTS Compared to the control group, the total effective rate in the experimental group was significantly higher, and the healing time was significantly reduced. Cell viability and clone formation numbers were significantly upregulated by ligustrazine in a dose-dependent manner. Both the cell viability and clone formation numbers were significantly inhibited by application of LY294002. CONCLUSIONS Our results suggest that ligustrazine combined with holistic nursing is an effective treatment of pressure sores. The protective effect may be associated with the promotion of cell growth by activation of the PI3K/AKT pathway.

Therapeutic Effect of External Application of Ligustrazine Combined with Holistic Nursing on Pressure Sores

PubMed Central

Niu, Junzhi; Han, Lin; Gong, Fen

2016-01-01

Background This study aimed to explore the therapeutic effect of external application of ligustrazine combined with holistic nursing on pressure sores, as well as the underlying mechanism. Material/Methods From February 2014 to March 2015, a total of 32 patients with Phase II and Phase III pressure sores were enrolled and randomly assigned to an experimental group or a control group. The clinical data were comparable between the 2 groups. In addition to holistic nursing, the patients in the experimental group received 4 weeks of continuous external application of ligustrazine, whereas patients in the control group received compound clotrimazole cream. Therapeutic effect and healing time were recorded. HaCaT cells were used as an in vitro model for mechanism analysis of the effect of ligustrazine in treating pressure sores. After culturing with different concentrations of ligustrazine or the inhibitor of AKT (LY294002) for 72 h, cell viability, clone formation numbers, and levels of phosphatidyl inositol 3-kinase (PI3K), p-AKT, and p-mammalian target of rapamycin (mTOR) were determined. Results Compared to the control group, the total effective rate in the experimental group was significantly higher, and the healing time was significantly reduced. Cell viability and clone formation numbers were significantly upregulated by ligustrazine in a dose-dependent manner. Both the cell viability and clone formation numbers were significantly inhibited by application of LY294002. Conclusions Our results suggest that ligustrazine combined with holistic nursing is an effective treatment of pressure sores. The protective effect may be associated with the promotion of cell growth by activation of the PI3K/AKT pathway. PMID:27523814

BMS 493 Modulates Retinoic Acid-Induced Differentiation During Expansion of Human Hematopoietic Progenitor Cells for Islet Regeneration.

PubMed

Elgamal, Ruth M; Bell, Gillian I; Krause, Sarah C T; Hess, David A

2018-06-06

Cellular therapies are emerging as a novel treatment strategy for diabetes. Thus, the induction of endogenous islet regeneration in situ represents a feasible goal for diabetes therapy. Umbilical cord blood-derived hematopoietic progenitor cells (HPCs), isolated by high aldehyde dehydrogenase activity (ALDH hi ), have previously been shown to reduce hyperglycemia after intrapancreatic (iPan) transplantation into streptozotocin (STZ)-treated nonobese diabetic (NOD)/severe combined immunodeficiency (SCID) mice. However, these cells are rare and require ex vivo expansion to reach clinically applicable numbers for human therapy. Therefore, we investigated whether BMS 493, an inverse retinoic acid receptor agonist, could prevent retinoic acid-induced differentiation and preserve islet regenerative functions during expansion. After 6-day expansion, BMS 493-treated cells showed a twofold increase in the number of ALDH hi cells available for transplantation compared with untreated controls. Newly expanded ALDH hi cells showed increased numbers of CD34 and CD133-positive cells, as well as a reduction in CD38 expression, a marker of hematopoietic cell differentiation. BMS 493-treated cells showed similar hematopoietic colony-forming capacity compared with untreated cells, with ALDH hi subpopulations producing more colonies than low aldehyde dehydrogenase activity subpopulations for expanded cells. To determine if the secreted proteins of these cells could augment the survival and/or proliferation of β-cells in vitro, conditioned media (CM) from cells expanded with or without BMS 493 was added to human islet cultures. The total number of proliferating β-cells was increased after 3- or 7-day culture with CM generated from BMS 493-treated cells. In contrast to freshly isolated ALDH hi cells, 6-day expansion with or without BMS 493 generated progeny that were unable to reduce hyperglycemia after iPan transplantation into STZ-treated NOD/SCID mice. Further strategies to reduce retinoic acid differentiation during HPC expansion is required to expand ALDH hi cells without the loss of islet regenerative functions.

Changes in human bone marrow fat content associated with changes in hematopoietic stem cell numbers and cytokine levels with aging

PubMed Central

Tuljapurkar, Sonal R; McGuire, Timothy R; Brusnahan, Susan K; Jackson, John D; Garvin, Kevin L; Kessinger, Margaret A; Lane, Judy T; O' Kane, Barbara J; Sharp, John G

2011-01-01

Hematological deficiencies increase with aging, including anemias, reduced responses to hematopoietic stress and myelodysplasias. This investigation tested the hypothesis that increased bone marrow (BM) fat content in humans with age was associated with decreased numbers of side population (SP) hematopoietic stem cells, and this decrease correlated with changes in cytokine levels. BM was obtained from the femoral head and trochanteric region of the femur removed at surgery for total hip replacement (N = 100 subjects). In addition, BM from cadavers (N = 36), with no evidence of hip disease, was evaluated for fat content. Whole trabecular marrow samples were ground in a sterile mortar and pestle, and cellularity and lipid content determined. Marrow cells were stained with Hoechst dye and SP profiles were acquired. Plasma levels of insulin-like growth factor (IGF)-1, stromal-derived factor (SDF)-1 and interleukin (IL)-6 were measured using ELISA. Fat content in the BM of human subjects and cadavers increased with age. The numbers of SP stem cells in BM as well as plasma IGF-1 and SDF-1 levels decreased in correlation with increased BM fat. IL-6 had no relationship to changes in marrow fat. These data suggest that increased BM fat may be associated with a decreased number of SP stem cells and IGF-1 and SDF-1 levels with aging. These data further raise a more general question as to the role of adipose cells in the regulation of tissue stem cells. PMID:21923862

Effect of Haemoproteus columbae infection on the hemogram of the Pigeons (Columba livia domestica).

PubMed

Samani, Amir Dehghani; Kheirabadi, Khodadad Pirali; Mohebbi, Abdonnaser

2016-12-01

It has been reported that parasites affect the health and productivity of birds, initiate excessive preening which interrupts feeding, as the birds spend much time preening rather than being involved in other essential life activities. Haemoproteus columbae occurs in pigeons widely in tropical and subtropical regions. The present investigation was concentrated on the effect of H. columbae infection on the hemogram and changes in the number of white blood cells in infected pigeons. Blood samples taked from 100 pigeons in Shahrekord. Blood smears stained and observed for H. columbae infection. Packed cell volume (PCV) and total white blood cells (WBC) count and differential WBC count determinate for every smears. This study show that PCV increased in infected pigeons by H. columbae than non infected pigeons. There were increasing in the number of total WBC in the infected pigeons by H. columbae than non infected pigeons. Also there were significant differences between the number of lymphocytes and eosinophils in the infected pigeons by H. columbae than non infected pigeons. This study and same studies show the part of the effects of blood parasites on the hemogram, but there were many questions about the effects of blood parasites such as H. columbae on the other parameters of host's blood, so we advise to focus on the other hematological parameters in the future studies.

A Phenotypic Change But Not Proliferation Underlies Glial Responses in Alzheimer Disease

PubMed Central

Serrano-Pozo, Alberto; Gómez-Isla, Teresa; Growdon, John H.; Frosch, Matthew P.; Hyman, Bradley T.

2014-01-01

Classical immunohistochemical studies in the Alzheimer disease (AD) brain reveal prominent glial reactions, but whether this pathological feature is due primarily to cell proliferation or to a phenotypic change of existing resting cells remains controversial. We performed double-fluorescence immunohistochemical studies of astrocytes and microglia, followed by unbiased stereology-based quantitation in temporal cortex of 40 AD patients and 32 age-matched nondemented subjects. Glial fibrillary acidic protein (GFAP) and major histocompatibility complex II (MHC2) were used as markers of astrocytic and microglial activation, respectively. Aldehyde dehydrogenase 1 L1 and glutamine synthetase were used as constitutive astrocytic markers, and ionized calcium-binding adaptor molecule 1 (IBA1) as a constitutive microglial marker. As expected, AD patients had higher numbers of GFAP+ astrocytes and MHC2+ microglia than the nondemented subjects. However, both groups had similar numbers of total astrocytes and microglia and, in the AD group, these total numbers remained essentially constant over the clinical course of the disease. The GFAP immunoreactivity of astrocytes, but not the MHC2 immunoreactivity of microglia, increased in parallel with the duration of the clinical illness in the AD group. Cortical atrophy contributed to the perception of increased glia density. We conclude that a phenotypic change of existing glial cells, rather than a marked proliferation of glial precursors, accounts for the majority of the glial responses observed in the AD brain. PMID:23602650

Decoy receptor 3 attenuates collagen-induced arthritis by modulating T cell activation and B cell expansion.

PubMed

Cheng, Chia-Pi; Sytwu, Huey-Kang; Chang, Deh-Ming

2011-12-01

To investigate the immune-modulated effects of decoy receptor 3 (DCR3) in an experimental model of rheumatoid arthritis (RA). We delivered DCR3 plasmid into collagen-induced arthritis (CIA) mice using the hydrodynamic method and evaluated the serum level of DCR3 protein by ELISA. After immunization, we assessed disease severity of arthritis incidence, arthritis scores, paw thickness, and means of arthritic limbs, and used hematoxylin and eosin staining to observe synovial hyperplasia. We analyzed numbers of murine splenocytes and inguinal lymphocyte cells, cell populations, and serum proinflammatory cytokines by flow cytometry. We investigated B cell proliferation by carboxyfluorescein succinimidyl ester assay. We evaluated serum levels of total IgG2a and type II collagen-specific IgG and IgG2a using ELISA. DCR3 expression in sera significantly attenuated disease severity in CIA mice. We found that DCR3 inhibited the volume of inguinal lymph nodes, numbers of CD19+ B cells, and populations of interferon-γ, interleukin 4 (IL-4), IL-17A, and Foxp3-producing CD4+ T cell in vivo. We found that DCR3 inhibited Pam3CSK4 (Toll-like receptor 1/2 ligand)-induced B220+ B cell proliferation in vitro. DCR3 treatment reduced the serum level of IL-6, total IgG2a, and CII-specific IgG2a antibody. We postulated that the protective effects of DCR3 in CIA resulted from modulation of the immune system by maintaining the B/T cell balance and decreasing lymphocyte expansion. We suggest DCR3 as a prophylactic and potential therapeutic agent in the treatment of RA.

Rat pancreatic B-cells after chronic alcohol feeding. A morphometric and fine structural study.

PubMed

Koko, V; Todorović, V; Nikolić, J A; Glisić, R; Cakić, M; Lacković, V; Petronijević, L; Stojković, M; Varagić, J; Janić, B

1995-04-01

Quantitative analysis of the light microscopic and fine structure of rat islet B-cells was carried out in chronic alcoholism. Absolute pancreatic weight and volume were similar in groups C (control) and E (ethanol), but relative pancreatic weight in group E rat was decreased. The results for fasting blood glucose and insulin levels were similar in the two groups of animals. There was a significantly reduced total pancreatic islet volume in E rats. The total number of endocrine cells both per islet and per microns2 of islet was similar in the two groups of animals. The volume density and number of B-cells per islet and per microns2 of islet were not changed in ethanol-treated rats as compared with the control. On the other hand, diameter, surface area and volume of the B-cells and their nuclei were found to be statistically significantly decreased. Histological examination revealed that islet blood vessels were dilated in alcoholic rats. Over the 4-month period of ethanol intake a significant decrease in cell profile area, nuclear profile area and volume density of cytoplasmic granules and an increase in the profile area and volume density of endoplasmic reticulum occurred. The gross histological alteration seen in most B-cells of the ethanol-treated rats was irregularity of the nuclear envelope with deep invagination and with margination of heterochromatin and many empty granules or granules without clear electron dense crystals of insulin. The present results indicate some optical and structural abnormalities of B-cells in chronic alcoholism that may be related to cell dysfunction and may contribute, at least in part, to the endocrine pancreas functional disturbance.

Chemotaxis of Dictyostelium discoideum: Collective Oscillation of Cellular Contacts

PubMed Central

Schäfer, Edith; Tarantola, Marco; Polo, Elena; Westendorf, Christian; Oikawa, Noriko; Bodenschatz, Eberhard; Geil, Burkhard; Janshoff, Andreas

2013-01-01

Chemotactic responses of Dictyostelium discoideum cells to periodic self-generated signals of extracellular cAMP comprise a large number of intricate morphological changes on different length scales. Here, we scrutinized chemotaxis of single Dictyostelium discoideum cells under conditions of starvation using a variety of optical, electrical and acoustic methods. Amebas were seeded on gold electrodes displaying impedance oscillations that were simultaneously analyzed by optical video microscopy to relate synchronous changes in cell density, morphology, and distance from the surface to the transient impedance signal. We found that starved amebas periodically reduce their overall distance from the surface producing a larger impedance and higher total fluorescence intensity in total internal reflection fluorescence microscopy. Therefore, we propose that the dominant sources of the observed impedance oscillations observed on electric cell-substrate impedance sensing electrodes are periodic changes of the overall cell-substrate distance of a cell. These synchronous changes of the cell-electrode distance were also observed in the oscillating signal of acoustic resonators covered with amebas. We also found that periodic cell-cell aggregation into transient clusters correlates with changes in the cell-substrate distance and might also contribute to the impedance signal. It turned out that cell-cell contacts as well as cell-substrate contacts form synchronously during chemotaxis of Dictyostelium discoideum cells. PMID:23349816

MC3T3-E1 Cells on Titanium Surfaces with Nanometer Smoothness and Fibronectin Immobilization

PubMed Central

Hayakawa, Tohru; Yoshida, Eiji; Yoshimura, Yoshitaka; Uo, Motohiro; Yoshinari, Masao

2012-01-01

The present study was aimed to evaluate the viability and total protein contents of osteoblast-like cells on the titanium surface with different surface mechanical treatment, namely, nanometer smoothing (Ra: approximately 2.0 nm) and sandblasting (Ra: approximately 1.0 μm), and biochemical treatment, namely, with or without fibronectin immobilization. Fibronectin could be easily immobilized by tresyl chloride-activation technique. MC3T3-E1 cells were seeded on the different titanium surfaces. Cell viability was determined by MTT assay. At 1 day of cell culture, there were no significant differences in cell viability among four different titanium surfaces. At 11 days, sandblasted titanium surface with fibronectin immobilization showed the significantly highest cell viability than other titanium surface. No significant differences existed for total protein contents among four different titanium surfaces at 11 days of cell culture. Scanning electron microscopy observation revealed that smoothness of titanium surface produced more spread cell morphologies, but that fibronectin immobilization did not cause any changes of the morphologies of attached cells. Fibronectin immobilization provided greater amount of the number of attached cells and better arrangement of attached cells. In conclusion, the combination of sandblasting and fibronectin immobilization enhanced the cell viability and fibronectin immobilization providing better arrangements of attached cells. PMID:22675359

Develop and test fuel cell powered on site integrated total energy systems. Phase 3: Full-scale power plant development

NASA Technical Reports Server (NTRS)

Kaufman, A.; Olson, B.; Pudick, S.; Wang, C. L.; Werth, J.; Whelan, J. A.

1986-01-01

The third in a series of 4kW stacks, consisting of 24 cells of the 13 inch x 23 inch cell size, has been on test for about 1600 hours. This stack is similar to the first two stacks, which ran 7000 and 8400 hours, respectively. The present stack incorporates technology improvements relating to the electrolyte-matrix, the current-collector assembly, and a reduction in the number of cooling plates. Performance is currently averaging about 0.64 per cell at 161 mA sq cm.

Voltage equalization of an ultracapacitor module by cell grouping using number partitioning algorithm

NASA Astrophysics Data System (ADS)

Oyarbide, E.; Bernal, C.; Molina, P.; Jiménez, L. A.; Gálvez, R.; Martínez, A.

2016-01-01

Ultracapacitors are low voltage devices and therefore, for practical applications, they need to be used in modules of series-connected cells. Because of the inherent manufacturing tolerance of the capacitance parameter of each cell, and as the maximum voltage value cannot be exceeded, the module requires inter-cell voltage equalization. If the intended application suffers repeated fast charging/discharging cycles, active equalization circuits must be rated to full power, and thus the module becomes expensive. Previous work shows that a series connection of several sets of paralleled ultracapacitors minimizes the dispersion of equivalent capacitance values, and also the voltage differences between capacitors. Thus the overall life expectancy is improved. This paper proposes a method to distribute ultracapacitors with a number partitioning-based strategy to reduce the dispersion between equivalent submodule capacitances. Thereafter, the total amount of stored energy and/or the life expectancy of the device can be considerably improved.

Copy number variation is a fundamental aspect of the placental genome.

PubMed

Hannibal, Roberta L; Chuong, Edward B; Rivera-Mulia, Juan Carlos; Gilbert, David M; Valouev, Anton; Baker, Julie C

2014-05-01

Discovery of lineage-specific somatic copy number variation (CNV) in mammals has led to debate over whether CNVs are mutations that propagate disease or whether they are a normal, and even essential, aspect of cell biology. We show that 1,000 N polyploid trophoblast giant cells (TGCs) of the mouse placenta contain 47 regions, totaling 138 Megabases, where genomic copies are underrepresented (UR). UR domains originate from a subset of late-replicating heterochromatic regions containing gene deserts and genes involved in cell adhesion and neurogenesis. While lineage-specific CNVs have been identified in mammalian cells, classically in the immune system where V(D)J recombination occurs, we demonstrate that CNVs form during gestation in the placenta by an underreplication mechanism, not by recombination nor deletion. Our results reveal that large scale CNVs are a normal feature of the mammalian placental genome, which are regulated systematically during embryogenesis and are propagated by a mechanism of underreplication.

Mechanism of radiosensitization by porphyrins.

PubMed

Luksiene, Zivile; Labeikyte, Danute; Juodka, Benediktas; Moan, Johan

2006-01-01

According to our previous data, hematoporphyrin dimethyl ether (HPde) at concentrations useful for photodynamic therapy can radiosensitize aggressive Ehrlich ascite carcinoma (EAT) to 2Gy irradiation inducing total tumour growth inhibition. The aim of this study was to further investigate the possible mechanism of radiosensitization of EAT by dicarboxylic porphyrin-HPde. Our results reveal that HPde is inducing several rearrangements in the EAT cells: 1.2 x 10-6 M of the photosensitizer diminishes the number of cells in mitosis by a factor of 3, increases the number of cells in the S phase of the cell cycle, modifies the activities of antioxidant enzymes glutation S-transferase (GST) and DT-diaphorase (DTD), and eventually induces slight apoptosis. Moreover, it was shown that HPde is a ligand of peripheral benzodiazepine receptor (PBR). Named "house keeper," PBR is usually responsible for all these perturbations, which, in our case, act in concert with the following ionizing radiation, producing the interaction of two antiproliferative/destructive factors.

Regeneration of the epidermis and basement membrane of the planarian Dugesia japonica after total-body x irradiation

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

Hori, I.

1979-03-01

Fresh-water planarians were studied to examine effects of x rays on regeneration of the epidermis and basement membrane. During early stages of regeneration, free rhabdite-forming cells were associated with the wound epidermis and recruited it. In later stages, however, a gradual degeneration occurred in the epidermis and cells undergoing epithelization decreased in number. Eventually epidermal cells on the wound surface appeared necrotic as evidenced by pyknotic nuclei and vacuolized dense cytoplasm. The entire basement membrane could not be reconstituted in any stage after wounding though its precursor-like material was secreted in the interspace between epidermis and parenchyma. Morphological changes inmore » extracellular products and in the cells surrounding the products suggest that epidermal cells which have covered the wound surface synthesize precursors of the basement membrane. Possible factors of a characteristic perturbation in epithelization and basement membrane formation after total-body irradiation are discussed.« less

Development, glycolytic activity, and viability of preimplantation mouse embryos subjected to different periods of glucose starvation.

PubMed

Leppens-Luisier, G; Sakkas, D

1997-03-01

After compaction, the preimplantation mouse embryo switches to a glucose-based metabolism, whereas for the 2- to 4-cell stage embryo, glucose can be inhibitory. In this study, we investigated the adaptability of preimplantation embryos to different periods of glucose starvation by culturing in vitro fertilized (IVF) and in vivo-fertilized 1-cell OF1 mouse embryos. Blastocysts obtained from exposure to glucose starvation for different periods of time were examined for the number of cells in the trophectoderm and inner cell mass, and for glycolytic activity and viability. A high percentage of blastocysts was obtained when 1-cell embryos fertilized in vitro or in vivo were cultured in M16 until the 2-cell stage, were transferred to M16 without glucose (M16-G) until the 4- or 8-cell stage, and then were transferred to fresh M16-G. When in vivo-fertilized 1-cell embryos were cultured to the 2-cell stage and then left in M16, less than 5% formed blastocysts compared to 26% of those transferred into M16-G. Blastocysts obtained when in vivo-fertilized 1-cell embryos were left in M16-G after the 2-cell stage, however, showed a significantly elevated glycolytic activity compared to those transferred to fresh M16 or M16-G medium at the 4- or 8-cell stage. Interestingly, even though these embryos displayed elevated glycolytic activity, they did not exhibit differences in the numbers of inner cell mass and trophectoderm cells or in viability compared to embryos cultured according to other protocols. Blastocysts from all cultured protocols had a significantly lower total cell number and a lower trophectoderm, but not inner cell mass, cell number compared to blastocysts developed in vivo. This study documents the metabolic adaptability of the preimplantation embryo by highlighting its ability to proceed with development and retain viability when challenged with glucose starvation at different periods.

Association between length of storage of red blood cell units and outcome of critically ill children: a prospective observational study

PubMed Central

2010-01-01

Introduction Transfusion is a common treatment in pediatric intensive care units (PICUs). Studies in adults suggest that prolonged storage of red blood cell units is associated with worse clinical outcome. No prospective study has been conducted in children. Our objectives were to assess the clinical impact of the length of storage of red blood cell units on clinical outcome of critically ill children. Methods Prospective, observational study conducted in 30 North American centers, in consecutive patients aged <18 years with a stay ≥ 48 hours in a PICU. The primary outcome measure was the incidence of multiple organ dysfunction syndrome after transfusion. The secondary outcomes were 28-day mortality and PICU length of stay. Odds ratios were adjusted for gender, age, number of organ dysfunctions at admission, total number of transfusions, and total dose of transfusion, using a multiple logistic regression model. Results The median length of storage was 14 days in 296 patients with documented length of storage. For patients receiving blood stored ≥ 14 days, the adjusted odds ratio for an increased incidence of multiple organ dysfunction syndrome was 1.87 (95% CI 1.04;3.27, P = 0.03). There was also a significant difference in the total PICU length of stay (adjusted median difference +3.7 days, P < 0.001) and no significant change in mortality. Conclusions In critically ill children, transfusion of red blood cell units stored for ≥ 14 days is independently associated with an increased occurrence of multiple organ dysfunction syndrome and prolonged PICU stay. PMID:20377853

"allometry" Deterministic Approaches in Cell Size, Cell Number and Crude Fiber Content Related to the Physical Quality of Kangkong (Ipomoea reptans) Grown Under Different Plant Density Pressures

NASA Astrophysics Data System (ADS)

Selamat, A.; Atiman, S. A.; Puteh, A.; Abdullah, N. A. P.; Mohamed, M. T. M.; Zulkeefli, A. A.; Othman, S.

Kangkong, especially the upland type (Ipomoea reptans) is popularly consumed as a vegetable dish in the South East Asian countries for its quality related to Vitamins (A and C) and crude fiber contents. Higher fiber contents would prevent from the occurrence of colon cancer and diverticular disease. With young stem edible portion, its cell number and size contribute to the stem crude fiber content. The mathematical approach of allometry of cell size, number, and fiber content of stem could be used in determining the 'best' plant density pressure in producing the quality young stem to be consumed. Basically, allometry is the ratio of relative increment (growth or change) rates of two parameters, or the change rate associated to the log of measured variables relationship. Kangkog grown equal or lower than 55 plants m-2 produced bigger individual plant and good quality (physical) kangkong leafy vegetable, but with lower total yield per unit area as compared to those grown at higher densities.

Treatment of lupus nephritis with total lymphoid irradiation. Observations during a 12-79-month followup

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

Strober, S.; Farinas, M.C.; Field, E.H.

1988-07-01

Seventeen patients with intractable lupus nephritis and nephrotic syndrome were treated with total lymphoid irradiation. Statistically significant improvement in mean renal disease and serologic activity parameters occurred within 3 months and persisted for at least 3 years. Although there was a marked reduction of T helper cell numbers and function after total lymphoid irradiation, recovery of these parameters was not associated with a return of disease activity. Risks of sterility, severe infections, and hematologic malignancy appeared to be lower than with alkylating agents.

Bone-marrow-derived mesenchymal stem cells inhibit gastric aspiration lung injury and inflammation in rats.

PubMed

Zhou, Jing; Jiang, Liyan; Long, Xuan; Fu, Cuiping; Wang, Xiangdong; Wu, Xiaodan; Liu, Zilong; Zhu, Fen; Shi, Jindong; Li, Shanqun

2016-09-01

Gastric aspiration lung injury is one of the most common clinical events. This study investigated the effects of bone-marrow-derived mesenchymal stem cells (BMSCs) on combined acid plus small non-acidified particle (CASP)-induced aspiration lung injury. Enhanced green fluorescent protein (EGFP(+) ) or EGFP(-) BMSCs or 15d-PGJ2 were injected via the tail vein into rats immediately after CASP-induced aspiration lung injury. Pathological changes in lung tissues, blood gas analysis, the wet/dry weight ratio (W/D) of the lung, levels of total proteins and number of total cells and neutrophils in bronchoalveolar lavage fluid (BALF) were determined. The cytokine levels were measured using ELISA. Protein expression was determined by Western blot. Bone-marrow-derived mesenchymal stem cells treatment significantly reduced alveolar oedema, exudation and lung inflammation; increased the arterial partial pressure of oxygen; and decreased the W/D of the lung, the levels of total proteins and the number of total cells and neutrophils in BALF in the rats with CASP-induced lung injury. Bone-marrow-derived mesenchymal stem cells treatment decreased the levels of tumour necrosis factor-α and Cytokine-induced neutrophil chemoattractant (CINC)-1 and the expression of p-p65 and increased the levels of interleukin-10 and 15d-PGJ2 and the expression of peroxisome proliferator-activated receptor (PPAR)-γ in the lung tissue in CASP-induced rats. Tumour necrosis factor-α stimulated BMSCs to secrete 15d-PGJ2 . A tracking experiment showed that EGFP(+) BMSCs were able to migrate to local lung tissues. Treatment with 15d-PGJ2 also significantly inhibited CASP-induced lung inflammation and the production of pro-inflammatory cytokines. Our results show that BMSCs can protect lung tissues from gastric aspiration injury and inhibit lung inflammation in rats. A beneficial effect might be achieved through BMSC-derived 15d-PGJ2 activation of the PPAR-γ receptor, reducing the production of proinflammatory cytokines. © 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Synergy of bone marrow transplantation and curcumin ensue protective effects at early onset of diabetes in mice.

PubMed

Arivazhagan, Arivarasan; Krishna, Soni; Yadav, Shivangi; Shah, Harshit Rajesh; Kumar, Pravir; Ambasta, Rashmi Kumar

2015-07-01

The aim of this study was to investigate the early onset effects of diabetes on pro-angiogenic signaling pathway, total number of bone marrow cells, organs (pancreas and kidney) damage and the reversal effect of diabetes by combinatorial treatment of curcumin and bone marrow transplantation in streptozotocin (STZ) induced diabetic mice. In the present study, Streptozotocin induced diabetic mice were transplanted with bone marrow cells (2 × 10(6) ) followed by the administration of curcumin (80 mg/kg bodyweight). Effect of diabetes on the different organs was studied by H&E, Western blotting and immunofluorescence using vascular endothelial growth factor (VEGF), platelet/endothelial cell adhesion molecule (PECAM), insulin, Caspase-9 and Caspase-3 antibodies. The effect of diabetes results in the reduction of the total cell number and viability of the bone marrow cells, organ degeneration and lower VEGF/PECAM expression. However, transplantation with normal bone marrow cells significantly reduced the blood glucose levels (above normal range) and initiated the organ regeneration via the VEGF/PECAM mediated manner. Curcumin treatment further reduced the blood glucose level (near normal); and accelerated the organ regeneration, enhanced VEGF/PECAM expression and decreased caspase expression level in the organs. Curcumin also had a protective role against the glucotoxicity test performed on the bone marrow cells. This study suggests that bone marrow transplantation and curcumin administration is an effective treatment in reversing the early onset effects of diabetes via the VEGF/PECAM signaling pathway. © 2014 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and Wiley Publishing Asia Pty Ltd.

Effect of Weight Reduction on Cardiovascular Risk Factors and CD34-positive Cells in Circulation

PubMed Central

Mikirova, Nina A; Casciari, Joseph J; Hunninghake, Ronald E; Beezley, Margaret M

2011-01-01

Being overweight or obese is associated with an increased risk for the development of non-insulin-dependent diabetes mellitus, hypertension, and cardiovascular disease. Dyslipidemia of obesity is characterized by elevated fasting triglycerides and decreased high-density lipoprotein-cholesterol concentrations. Endothelial damage and dysfunction is considered to be a major underlying mechanism for the elevated cardiovascular risk associated with increased adiposity. Alterations in endothelial cells and stem/endothelial progenitor cell function associated with overweight and obesity predispose to atherosclerosis and thrombosis. In our study, we analyzed the effect of a low calorie diet in combination with oral supplementation by vitamins, minerals, probiotics and human chorionic gonadotropin (hCG, 125-180 IUs) on the body composition, lipid profile and CD34-positive cells in circulation. During this dieting program, the following parameters were assessed weekly for all participants: fat free mass, body fat, BMI, extracellular/intracellular water, total body water and basal metabolic rate. For part of participants blood chemistry parameters and circulating CD34-positive cells were determined before and after dieting. The data indicated that the treatments not only reduced body fat mass and total mass but also improved the lipid profile. The changes in body composition correlated with the level of lipoproteins responsible for the increased cardiovascular risk factors. These changes in body composition and lipid profile parameters coincided with the improvement of circulatory progenitor cell numbers. As the result of our study, we concluded that the improvement of body composition affects the number of stem/progenitor cells in circulation. PMID:21850193

Effect of weight reduction on cardiovascular risk factors and CD34-positive cells in circulation.

PubMed

Mikirova, Nina A; Casciari, Joseph J; Hunninghake, Ronald E; Beezley, Margaret M

2011-01-01

Being overweight or obese is associated with an increased risk for the development of non-insulin-dependent diabetes mellitus, hypertension, and cardiovascular disease. Dyslipidemia of obesity is characterized by elevated fasting triglycerides and decreased high-density lipoprotein-cholesterol concentrations. Endothelial damage and dysfunction is considered to be a major underlying mechanism for the elevated cardiovascular risk associated with increased adiposity. Alterations in endothelial cells and stem/endothelial progenitor cell function associated with overweight and obesity predispose to atherosclerosis and thrombosis. In our study, we analyzed the effect of a low calorie diet in combination with oral supplementation by vitamins, minerals, probiotics and human chorionic gonadotropin (hCG, 125-180 IUs) on the body composition, lipid profile and CD34-positive cells in circulation. During this dieting program, the following parameters were assessed weekly for all participants: fat free mass, body fat, BMI, extracellular/intracellular water, total body water and basal metabolic rate. For part of participants blood chemistry parameters and circulating CD34-positive cells were determined before and after dieting. The data indicated that the treatments not only reduced body fat mass and total mass but also improved the lipid profile. The changes in body composition correlated with the level of lipoproteins responsible for the increased cardiovascular risk factors. These changes in body composition and lipid profile parameters coincided with the improvement of circulatory progenitor cell numbers. As the result of our study, we concluded that the improvement of body composition affects the number of stem/progenitor cells in circulation.

Polymorphisms in O-methyltransferase genes are associated with stover cell wall digestibility in European maize (Zea mays L.).

PubMed

Brenner, Everton A; Zein, Imad; Chen, Yongsheng; Andersen, Jeppe R; Wenzel, Gerhard; Ouzunova, Milena; Eder, Joachim; Darnhofer, Birte; Frei, Uschi; Barrière, Yves; Lübberstedt, Thomas

2010-02-12

OMT (O-methyltransferase) genes are involved in lignin biosynthesis, which relates to stover cell wall digestibility. Reduced lignin content is an important determinant of both forage quality and ethanol conversion efficiency of maize stover. Variation in genomic sequences coding for COMT, CCoAOMT1, and CCoAOMT2 was analyzed in relation to stover cell wall digestibility for a panel of 40 European forage maize inbred lines, and re-analyzed for a panel of 34 lines from a published French study. Different methodologies for association analysis were performed and compared. Across association methodologies, a total number of 25, 12, 1, 6 COMT polymorphic sites were significantly associated with DNDF, OMD, NDF, and WSC, respectively. Association analysis for CCoAOMT1 and CCoAOMT2 identified substantially fewer polymorphic sites (3 and 2, respectively) associated with the investigated traits. Our re-analysis on the 34 lines from a published French dataset identified 14 polymorphic sites significantly associated with cell wall digestibility, two of them were consistent with our study. Promising polymorphisms putatively causally associated with variability of cell wall digestibility were inferred from the total number of significantly associated SNPs/Indels. Several polymorphic sites for three O-methyltransferase loci were associated with stover cell wall digestibility. All three tested genes seem to be involved in controlling DNDF, in particular COMT. Thus, considerable variation among Bm3 wildtype alleles can be exploited for improving cell-wall digestibility. Target sites for functional markers were identified enabling development of efficient marker-based selection strategies.

Polymorphisms in O-methyltransferase genes are associated with stover cell wall digestibility in European maize (Zea mays L.)

PubMed Central

2010-01-01

Background OMT (O-methyltransferase) genes are involved in lignin biosynthesis, which relates to stover cell wall digestibility. Reduced lignin content is an important determinant of both forage quality and ethanol conversion efficiency of maize stover. Results Variation in genomic sequences coding for COMT, CCoAOMT1, and CCoAOMT2 was analyzed in relation to stover cell wall digestibility for a panel of 40 European forage maize inbred lines, and re-analyzed for a panel of 34 lines from a published French study. Different methodologies for association analysis were performed and compared. Across association methodologies, a total number of 25, 12, 1, 6 COMT polymorphic sites were significantly associated with DNDF, OMD, NDF, and WSC, respectively. Association analysis for CCoAOMT1 and CCoAOMT2 identified substantially fewer polymorphic sites (3 and 2, respectively) associated with the investigated traits. Our re-analysis on the 34 lines from a published French dataset identified 14 polymorphic sites significantly associated with cell wall digestibility, two of them were consistent with our study. Promising polymorphisms putatively causally associated with variability of cell wall digestibility were inferred from the total number of significantly associated SNPs/Indels. Conclusions Several polymorphic sites for three O-methyltransferase loci were associated with stover cell wall digestibility. All three tested genes seem to be involved in controlling DNDF, in particular COMT. Thus, considerable variation among Bm3 wildtype alleles can be exploited for improving cell-wall digestibility. Target sites for functional markers were identified enabling development of efficient marker-based selection strategies. PMID:20152036

Umbilical Cord Blood-Derived Mononuclear Cells Exhibit Pericyte-Like Phenotype and Support Network Formation of Endothelial Progenitor Cells In Vitro.

PubMed

Peters, Erica B; Liu, Betty; Christoforou, Nicolas; West, Jennifer L; Truskey, George A

2015-10-01

Umbilical cord blood represents a promising cell source for pro-angiogenic therapies. The present study examined the potential of mononuclear cells (MNCs) from umbilical cord blood to support endothelial progenitor cell (EPC) microvessel formation. MNCs were isolated from the cord blood of 20 separate donors and selected for further characterization based upon their proliferation potential and morphological resemblance to human vascular pericytes (HVPs). MNCs were screened for their ability to support EPC network formation using an in vitro assay (Matrigel™) as well as a reductionist, coculture system consisting of no additional angiogenic cytokines beyond those present in serum. In less than 15% of the isolations, we identified a population of highly proliferative MNCs that phenotypically resembled HVPs as assessed by expression of PDGFR-β, NG2, α-SMA, and ephrin-B2. Within a Matrigel™ system, MNCs demonstrated pericyte-like function through colocalization to EPC networks and similar effects as HVPs upon total EPC tubule length (p = 0.95) and number of branch points (p = 0.93). In a reductionist coculture system, MNCs served as pro-angiogenic mural cells by supporting EPC network formation to a significantly greater extent than HVP cocultures, by day 14 of coculture, as evidenced through EPC total tubule length (p < 0.0001) and number of branch points (p < 0.0001). Our findings are significant as we demonstrate mural cell progenitors can be isolated from umbilical cord blood and develop culture conditions to support their use in microvascular tissue engineering applications.

Umbilical Cord Blood-Derived Mononuclear Cells Exhibit Pericyte-Like Phenotype and Support Network Formation of Endothelial Progenitor Cells In Vitro

PubMed Central

Peters, Erica B.; Liu, Betty; Christoforou, Nicolas; West, Jennifer L.; Truskey, George A.

2015-01-01

Umbilical cord blood represents a promising cell source for pro-angiogenic therapies. The present study examined the potential of mononuclear cells (MNCs) from umbilical cord blood to support endothelial progenitor cell (EPC) microvessel formation. MNCs were isolated from the cord blood of 20 separate donors and selected for further characterization based upon their proliferation potential and morphological resemblance to human vascular pericytes (HVPs). MNCs were screened for their ability to support EPC network formation using an in vitro assay (Matrigel™) as well as a reductionist, coculture system consisting of no additional angiogenic cytokines beyond those present in serum. In less than 15% of the isolations, we identified a population of highly proliferative MNCs that phenotypically resembled HVPs as assessed by expression of PDGFR-β, NG2, α-SMA, and ephrin-B2. Within a Matrigel™ system, MNCs demonstrated pericyte-like function through colocalization to EPC networks and similar effects as HVPs upon total EPC tubule length (p = 0.95) and number of branch points (p = 0.93). In a reductionist coculture system, MNCs served as pro-angiogenic mural cells by supporting EPC network formation to a significantly greater extent than HVP cocultures, by day 14 of coculture, as evidenced through EPC total tubule length (p <0.0001) and number of branch points (p < 0.0001). Our findings are significant as we demonstrate mural cell progenitors can be isolated from umbilical cord blood and develop culture conditions to support their use in microvascular tissue engineering applications. PMID:25777295

Evaluation of biological effects of intermediate frequency magnetic field on differentiation of embryonic stem cell.

PubMed

Yoshie, Sachiko; Ogasawara, Yuki; Ikehata, Masateru; Ishii, Kazuyuki; Suzuki, Yukihisa; Wada, Keiji; Wake, Kanako; Nakasono, Satoshi; Taki, Masao; Ohkubo, Chiyoji

2016-01-01

The embryotoxic effect of intermediate frequency (IF) magnetic field (MF) was evaluated using murine embryonic stem (ES) cells and fibroblast cells based on the embryonic stem cell test (EST). The cells were exposed to 21 kHz IF-MF up to magnetic flux density of 3.9 mT during the cell proliferation process (7 days) or the cell differentiation process (10 days) during which an embryonic body differentiated into myocardial cells. As a result, there was no significant difference in the cell proliferation between sham- and IF-MF-exposed cells for both ES and fibroblast cells. Similarly, the ratio of the number of ES-derived cell aggregates differentiated to myocardial cells to total number of cell aggregates was not changed by IF-MF exposure. In addition, the expressions of a cardiomyocytes-specific gene, Myl2 , and an early developmental gene, Hba-x , in the exposed cell aggregate were not altered. Since the magnetic flux density adopted in this study is much higher than that generated by an inverter of the electrical railway, an induction heating (IH) cooktop, etc . in our daily lives, these results suggested that IF-MF in which the public is exposed to in general living environment would not have embryotoxic effect.

Resistance Training Enhances Skeletal Muscle Innervation Without Modifying the Number of Satellite Cells or their Myofiber Association in Obese Older Adults

PubMed Central

Messi, María Laura; Li, Tao; Wang, Zhong-Min; Marsh, Anthony P.; Nicklas, Barbara

2016-01-01

Studies in humans and animal models provide compelling evidence for age-related skeletal muscle denervation, which may contribute to muscle fiber atrophy and loss. Skeletal muscle denervation seems relentless; however, long-term, high-intensity physical activity appears to promote muscle reinnervation. Whether 5-month resistance training (RT) enhances skeletal muscle innervation in obese older adults is unknown. This study found that neural cell-adhesion molecule, NCAM+ muscle area decreased with RT and was inversely correlated with muscle strength. NCAM1 and RUNX1 gene transcripts significantly decreased with the intervention. Type I and type II fiber grouping in the vastus lateralis did not change significantly but increases in leg press and knee extensor strength inversely correlated with type I, but not with type II, fiber grouping. RT did not modify the total number of satellite cells, their number per area, or the number associated with specific fiber subtypes or innervated/denervated fibers. Our results suggest that RT has a beneficial impact on skeletal innervation, even when started late in life by sedentary obese older adults. PMID:26447161

Lack of effectiveness of laser therapy applied to the nerve course and the correspondent medullary roots

PubMed Central

Sousa, Fausto Fernandes de Almeida; Ribeiro, Thaís Lopes; Fazan, Valéria Paula Sassoli; Barbieri, Claudio Henrique

2013-01-01

OBJECTIVE: To investigate the influence of low intensity laser irradiation on the regeneration of the fibular nerve of rats after crush injury. METHODS: Twenty-five rats were used, divided into three groups: 1) intact nerve, no treatment; 2) crushed nerve, no treatment; 3) crush injury, laser irradiation applied on the medullary region corresponding to the roots of the sciatic nerve and subsequently on the course of the damaged nerve. Laser irradiation was carried out for 14 consecutive days. RESULTS: Animals were evaluated by functional gait analysis with the peroneal functional index and by histomorphometric analysis using the total number of myelinated nerve fibers and their density, total number of Schwann cells, total number of blood vessels and the occupied area, minimum diameter of the fiber diameter and G-quotient. CONCLUSION: According to the statistical analysis there was no significant difference among groups and the authors conclude that low intensity laser irradiation has little or no influence on nerve regeneration and functional recovery. Laboratory investigation. PMID:24453650

The induction of menadione stress tolerance in the marine microalga, Dunaliella viridis, through cold pretreatment and modulation of the ascorbate and glutathione pools.

PubMed

Madadkar Haghjou, Maryam; Colville, Louise; Smirnoff, Nicholas

2014-11-01

The effect of cold pretreatment on menadione tolerance was investigated in the cells of the marine microalga, Dunaliella viridis. In addition, the involvement of ascorbate and glutathione in the response to menadione stress was tested by treating cell suspensions with l-galactono-1,4-lactone, an ascorbate precursor, and buthionine sulfoximine, an inhibitor of glutathione synthesis. Menadione was highly toxic to non cold-pretreated cells, and caused a large decrease in cell number. Cold pretreatment alleviated menadione toxicity and cold pretreated cells accumulated lower levels of reactive oxygen species, and had enhanced antioxidant capacity due to increased levels of β-carotene, reduced ascorbate and total glutathione compared to non cold-pretreated cells. Cold pretreatment also altered the response to l-galactono-1,4-lactone and buthionine sulfoximine treatments. Combined l-galactono-1,4-lactone and menadione treatment was lethal in non-cold pretreated cells, but in cold-pretreated cells it had a positive effect on cell numbers compared to menadione alone. Overall, exposure of Dunaliella cells to cold stress enhanced tolerance to subsequent oxidative stress induced by menadione. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Evaluation of umbilical cord blood CD34+ hematopoietic stem cells expansion with inhibition of TGF-β receptorII in co-culture with bone marrow mesenchymal stromal cells.

PubMed

Sohrabi Akhkand, Saman; Amirizadeh, Naser; Nikougoftar, Mahin; Alizadeh, Javad; Zaker, Farhad; Sarveazad, Arash; Joghataei, Mohammad Taghi; Faramarzi, Mahmood

2016-08-01

Umbilical cord blood (UCB) is an important source of hematopoietic stem cells (HSCs). However, low number of HSCs in UCB has been an obstacle for adult hematopoietic stem cell transplantation. The expansion of HSCs in culture is one approach to overcome this problem. In this study, we investigated the expansion of UCB-HSCs by using human bone marrow mesenchymal stromal cells (MSCs) as feeder layer as well as inhibiting the TGF-β signaling pathway through reduction of TGF-βRII expression. CD34(+) cells were isolated from UCB and transfected by SiRNA targeting TGF-βRII mRNA. CD34(+) cells were expanded in four culture media with different conditions, including 1) expansion of CD34(+) cells in serum free medium containing growth factors, 2) expansion of cells transfected with SiRNA targeting TGF-βRII in medium containing growth factors, 3) expansion of cells in presence of growth factors and MSCs, 4) expansion of cells transfected with SiRNA targeting TGF-βRII on MSCs feeder layer in medium containing growth factors. These culture conditions were evaluated for the number of total nucleated cells (TNCs), CD34 surface marker as well as using CFU assay on 8th day after culture. The fold increase in CD34(+) cells, TNCs, and colony numbers (71.8±6.9, 93.2±10.2 and 128±10, respectively) was observed to be highest in fourth culture medium compared to other culture conditions. The difference between number of cells in four culture media in 8th day compared to unexpanded cells (0day) before expansion was statistically significant (P<0.05). The results showed that transfection of CD34(+) cells with SiRNA targeting TGF-βRII and their co-culture with MSCs could considerably increase the number of progenitors. Therefore, this method could be useful for UCB-HSCs expansion. Copyright © 2016 Elsevier Ltd. All rights reserved.

Does stinging nettle (Urtica dioica) have an effect on bone formation in the expanded inter-premaxillary suture?

PubMed

Irgin, Celal; Çörekçi, Bayram; Ozan, Fatih; Halicioğlu, Koray; Toptaş, Orçun; Birinci Yildirim, Arzu; Türker, Arzu; Yilmaz, Fahri

2016-09-01

To determine whether systemically given stinging nettle (SN) has an effect on bone formation in response to expansion of the rat inter-premaxillary suture. A total of 28 male Wistar albino rats were randomly divided into 4 equal groups: control (C), only expansion (OE), SN extract given only during the expansion and retention periods (SN group; a total of 17days), and SN extract given during the nursery phase before expansion (a period of 40days) and during the expansion and retention periods (N+SN group; a total of 57days). After the 5-day expansion period was completed, the rats in the OE, SN, and N+SN groups underwent 12days of mechanical retention, after which they were sacrificed, and their premaxilla were dissected and fixed. A histologic evaluation was done to determine the number of osteoblasts, osteoclasts, and capillaries, as well as the number and intensity of inflammatory cells and new bone formation. Statistically significant differences were found between the groups in all histologic parameters except the ratio of intensities of inflammatory cells. New bone formation and the number of capillaries were significantly higher in the SN groups than in the other groups. The statistical analysis also showed that the numbers of osteoblasts, osteoclasts, and capillaries were highest in the N+SN group. Systemic administration of SN may be effective in accelerating new bone formation and reducing inflammation in the maxillary expansion procedure. It may also be beneficial in preventing relapse after the expansion procedure. Copyright © 2016 Elsevier Ltd. All rights reserved.

Automated assembling of single fuel cell units for use in a fuel cell stack

NASA Astrophysics Data System (ADS)

Jalba, C. K.; Muminovic, A.; Barz, C.; Nasui, V.

2017-05-01

The manufacturing of PEMFC stacks (POLYMER ELEKTROLYT MEMBRAN Fuel Cell) is nowadays still done by hand. Over hundreds of identical single components have to be placed accurate together for the construction of a fuel cell stack. Beside logistic problems, higher total costs and disadvantages in weight the high number of components produce a higher statistic interference because of faulty erection or material defects and summation of manufacturing tolerances. The saving of costs is about 20 - 25 %. Furthermore, the total weight of the fuel cells will be reduced because of a new sealing technology. Overall a one minute cycle time has to be aimed per cell at the manufacturing of these single components. The change of the existing sealing concept to a bonded sealing is one of the important requisites to get an automated manufacturing of single cell units. One of the important steps for an automated gluing process is the checking of the glue application by using of an image processing system. After bonding the single fuel cell the sealing and electrical function can be checked, so that only functional and high qualitative cells can get into further manufacturing processes.

[Impact of diabetes in children and adolescents].

PubMed

Pera, Pilar Isla

2012-06-01

DM 1 can occur at any age, but its higher incidence seen in under 15 years of age with greater frequency in age preschool and especially prepuberal. It represents about 10 of the total number of forms of DM and is one of chronic disturbances more frequent in children and adolescents. The destruction of the pancreatic cells involves a total deficit of insulin in these patients by requiring treatment with insulin from the time of diagnosis.

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

Degowin, R.L.; Lass, S.L.

Our studies show that the induction of a chronic inflammatory lesion in the left hind legs of mice by administration of 5000 rad produced distinct abnormalities of the hematopoietic system. A peripheral neutrophilia accompanied reduced numbers of total nucleated cells, stem cells, stromal cells, erythroblasts, and lymphocytes in the unirradiated femoral marrow, and the spleen was enlarged. Mice with these hematopoietic abnormalities promptly succumbed with bone marrow failure to a sublethal dose of total body irradiation (600 rad TB). Acute inflammation associated with a sterile abscess also impaired survival after 600 rad TB. Hematopoietic abnormalities resembling those in mice withmore » inflammation had been reported in mice bearing a solid extramedullary tumor of sarcoma-180. Concomitant studies showed that bone marrow failure and impaired survival after 600 rad TB administered to mice bearing sarcoma-180 occurred at the same time as that in mice with chronic inflammation. We concluded that chronic inflammation or tumor produced similar abnormalities in the bone marrow and spleen that led to markedly impaired survival and death from bone marrow failure after a sublethal dose of total body irradiation. Although the extramedullary hematopoiesis in the enlarged spleen indicated that its microenvironment supported hematopoiesis, whereas that in marrow was reduced, it was insufficient to compensate for a total body deficit of functional stem cells.« less

Screening of bioactive peptides using an embryonic stem cell-based neurodifferentiation assay.

PubMed

Xu, Ruodan; Feyeux, Maxime; Julien, Stéphanie; Nemes, Csilla; Albrechtsen, Morten; Dinnyés, Andras; Krause, Karl-Heinz

2014-05-01

Differentiation of pluripotent stem cells, PSCs, towards neural lineages has attracted significant attention, given the potential use of such cells for in vitro studies and for regenerative medicine. The present experiments were designed to identify bioactive peptides which direct PSC differentiation towards neural cells. Fifteen peptides were designed based on NCAM, FGFR, and growth factors sequences. The effect of peptides was screened using a mouse embryonic stem cell line expressing luciferase dual reporter construct driven by promoters for neural tubulin and for elongation factor 1. Cell number was estimated by measuring total cellular DNA. We identified five peptides which enhanced activities of both promoters without relevant changes in cell number. We selected the two most potent peptides for further analysis: the NCAM-derived mimetic FGLL and the synthetic NCAM ligand, Plannexin. Both compounds induced phenotypic neuronal differentiation, as evidenced by increased neurite outgrowth. In summary, we used a simple, but sensitive screening approach to identify the neurogenic peptides. These peptides will not only provide new clues concerning pathways of neurogenesis, but they may also be interesting biotechnology tools for in vitro generation of neurons.

Comparison of mesenchymal stem cells obtained by suspended culture of synovium from patients with rheumatoid arthritis and osteoarthritis.

PubMed

Kohno, Yuji; Mizuno, Mitsuru; Ozeki, Nobutake; Katano, Hisako; Otabe, Koji; Koga, Hideyuki; Matsumoto, Mikio; Kaneko, Haruka; Takazawa, Yuji; Sekiya, Ichiro

2018-03-09

Mobilization of mesenchymal stem cells (MSCs) from the synovium was revealed using a "suspended synovium culture model" of osteoarthritis (OA). The pathology of rheumatoid arthritis (RA) differs from that of OA. We investigated whether mobilization of MSCs from the synovium also occurred in RA, and we compared the properties of synovial MSCs collected from suspended synovium culture models of RA and OA. Human synovium was harvested during total knee arthroplasty from the knee joints of patients with RA (n = 8) and OA (n = 6). The synovium was suspended in a bottle containing culture medium and a culture dish at the bottom. Cells were harvested from the dish and analyzed. No significant difference was observed between RA and OA in the harvested cell numbers per g of synovium. However, the variation in the number of cells harvested from each donor was greater for RA than for OA. The harvested cells were multipotent and no difference was observed in the cartilage pellet weight between RA and OA. The surface epitopes of the cells in RA and OA were similar to those of MSCs. Mobilization of MSCs from the synovium was demonstrated using a suspended synovium culture model for RA. The harvested cell numbers, chondrogenic potentials, and surface epitope profiles were comparable between the RA and OA models.

Ethanol intoxication prolongs post-burn pulmonary inflammation: role of alveolar macrophages

PubMed Central

Shults, Jill A.; Curtis, Brenda J.; Boe, Devin M.; Ramirez, Luis; Kovacs, Elizabeth J.

2016-01-01

In this study, the role and fate of AMs were examined in pulmonary inflammation after intoxication and injury. Clinical evidence has revealed that half of all burn patients brought to the emergency department are intoxicated at the time of injury. This combined insult results in amplified neutrophil accumulation and pulmonary edema, with an increased risk of lung failure and mortality, relative to either insult alone. We believe that this excessive pulmonary inflammation, which also parallels decreased lung function, is mediated in part by AMs. Restoration of lung tissue homeostasis is dependent on the eradication of neutrophils and removal of apoptotic cells, both major functions of AMs. Thirty minutes after binge ethanol intoxication, mice were anesthetized and given a 15% total body surface area dorsal scald injury. At 24 h, we found a 50% decrease in the total number of AMs (P < 0.05) and observed a proinflammatory phenotype on the remaining lung AMs. Loss of AMs paralleled a 6-fold increase in the number of TUNEL+ lung apoptotic cells (P < 0.05) and a 3.5-fold increase in the percentage of annexin V+ apoptotic cells in BAL (P < 0.05), after intoxication and injury, relative to controls. In contrast to the reduction in the number of cells, AMs from intoxicated and injured mice had a 4-fold increase in efferocytosis (P < 0.05). In summary, these data suggest that loss of AMs may delay resolution of inflammation, resulting in the pulmonary complications and elevated mortality rates observed in intoxicated and burn-injured patients. PMID:27531926

Allyl isothiocyanate-induced changes in the distribution of white blood cells in rats.

PubMed

Imaizumi, Kazuhiko; Sato, Shogo; Sakakibara, Yuko; Mori, Sayuri; Ohkuma, Masaki; Kawashima, Yu; Ban, Takamasa; Sasaki, Hiromi; Tachiyashiki, Kaoru

2010-08-01

The main pungent component of wasabi (Eutrema japonica) is known to be isothiocyanate and its derivatives, volatile substances. Allyl isothiocyanate (AITC) accounts for more than half of isothiocyanate derivatives. However, there is a little information on the effects of AITC on the immune system by analyzing the number of white blood cells (WBCs) over the course of days of AITC administration. In the present study, we studied the effects of AITC (dose=20 mg/kg body weight/day for 10 days, subcutaneous: s.c.) on the number of WBCs (total WBCs, lymphocytes, monocyte, neutrophil, basophil and eosinophil) and plasma corticosterone concentrations in adult male rats. Administration of AITC decreased significantly the number of total WBCs on days 1-4 post s.c. injection by 25-27%. Administration of AITC also decreased the number of lymphocytes on days 1-10 by 21-36% and monocyte on days 1-8 by 28-78%. However, administration of AITC increased the number of neutrophil on days 8-10 by 61-112%. AITC did not change the number of eosinophil and basophil. Plasma corticosterone concentrations during the experimental period were 4.7-8.4 times significantly higher in the AITC group than in the control group, indicating that AITC induced stress-responses. The relative weights of thymus and adrenals per body weight were significantly lower and clearly higher in the AITC group than in the control group, respectively. These results suggest that AITC-mediated stress-responses are at least in part attributable to changes in the number of circulating WBCs.

Expansion of Endothelial Progenitor Cells in High Density Dot Culture of Rat Bone Marrow Cells

PubMed Central

Wang, Ling; Kretlow, James D.; Zhou, Guangdong; Cao, Yilin; Liu, Wei; Zhang, Wen Jie

2014-01-01

In vitro expansion of endothelial progenitor cells (EPCs) remains a challenge in stem cell research and its application. We hypothesize that high density culture is able to expand EPCs from bone marrow by mimicking cell-cell interactions of the bone marrow niche. To test the hypothesis, rat bone marrow cells were either cultured in high density (2×105 cells/cm2) by seeding total 9×105 cells into six high density dots or cultured in regular density (1.6×104 cells/cm2) with the same total number of cells. Flow cytometric analyses of the cells cultured for 15 days showed that high density cells exhibited smaller cell size and higher levels of marker expression related to EPCs when compared to regular density cultured cells. Functionally, these cells exhibited strong angiogenic potentials with better tubal formation in vitro and potent rescue of mouse ischemic limbs in vivo with their integration into neo-capillary structure. Global gene chip and ELISA analyses revealed up-regulated gene expression of adhesion molecules and enhanced protein release of pro-angiogenic growth factors in high density cultured cells. In summary, high density cell culture promotes expansion of bone marrow contained EPCs that are able to enhance tissue angiogenesis via paracrine growth factors and direct differentiation into endothelial cells. PMID:25254487

A replacement for islet equivalents with improved reliability and validity.

PubMed

Huang, Han-Hung; Ramachandran, Karthik; Stehno-Bittel, Lisa

2013-10-01

Islet equivalent (IE), the standard estimate of isolated islet volume, is an essential measure to determine the amount of transplanted islet tissue in the clinic and is used in research laboratories to normalize results, yet it is based on the false assumption that all islets are spherical. Here, we developed and tested a new easy-to-use method to quantify islet volume with greater accuracy. Isolated rat islets were dissociated into single cells, and the total cell number per islet was determined by using computer-assisted cytometry. Based on the cell number per islet, we created a regression model to convert islet diameter to cell number with a high R2 value (0.8) and good validity and reliability with the same model applicable to young and old rats and males or females. Conventional IE measurements overestimated the tissue volume of islets. To compare results obtained using IE or our new method, we compared Glut2 protein levels determined by Western Blot and proinsulin content via ELISA between small (diameter≤100 μm) and large (diameter≥200 μm) islets. When normalized by IE, large islets showed significantly lower Glut2 level and proinsulin content. However, when normalized by cell number, large and small islets had no difference in Glut2 levels, but large islets contained more proinsulin. In conclusion, normalizing islet volume by IE overestimated the tissue volume, which may lead to erroneous results. Normalizing by cell number is a more accurate method to quantify tissue amounts used in islet transplantation and research.

Comparative Analysis of Transcriptomes of Macrophage Revealing the Mechanism of the Immunoregulatory Activities of a Novel Polysaccharide Isolated from Boletus speciosus Frost.

PubMed

Ding, Xiang; Zhu, Hongqing; Hou, Yiling; Hou, Wanru; Zhang, Nan; Fu, Lei

2017-01-01

The mechanism of the immunoregulatory activities of polysaccharide is still not clear. Here, we performed the B-cell, T-cell, and macrophage cell proliferation, the cell cycle analysis of macrophage cells, sequenced the transcriptomes of control group macrophages, and Boletus speciosus Frost polysaccharide (BSF-1) group macrophages using Illumina sequencing technology to identify differentially expressed genes (DEGs) to determine the molecular mechanisms of immunomodulatory activity of BSF-1 in macrophages. These results suggested that BSF-1 could promote the proliferation of B-cell, T-cell, and macrophages, promote the proliferation of macrophage cells by abolishing cell cycle arrests in the G0/G1 phases, and promote cell cycle progression in S-phase and G2/M phase, which might induce cell division. A total of 12,498,414 and 11,840,624 bp paired-end reads were obtained for the control group and BSF-1 group, respectively, and they corresponded to a total size of 12.5 G bp and 11.8 G bp, respectively, after the low-quality reads and adapter sequences were removed. Approximately 81.83% of the total number of genes (8,257) were expressed reads per kilobase per million mapped reads (RPKM ≥1) and more than 1366 genes were highly expressed (RPKM >60) in the BSF-1 group. A gene ontology-enrichment analysis generated 13,042 assignments to cellular components, 13,094 assignments to biological processes, and 13,135 assignments to molecular functions. A Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed that the mitogen-activated protein kinase (MAPK) signaling pathways are significantly enriched for DEGs between the two cell groups. An analysis of transcriptome resources enabled us to examine gene expression profiles, verify differential gene expression, and select candidate signaling pathways as the mechanisms of the immunomodulatory activity of BSF-1. Based on the experimental data, we believe that the significant antitumor activities of BSF-1 in vivo mainly involve the MAPK signaling pathways. Boletus speciosus Frost-1 (BSF-1) could promote the proliferation of B-cell, T-cell, and macrophages, promote the proliferation of macrophage cells by abolishing cell cycle arrests in the G0/G1 phases, and promote cell cycle progression in S-phase and G2/M phase, which might induce cell divisionApproximately 81.83% of the total number of genes (8257) were expressed (reads per kilobase per million mapped reads [RPKM] =1) and more than 1366 genes were highly expressed (RPKM >60) in the BSF-1 groupA gene ontology-enrichment analysis generated 13,042 assignments to cellular components, 13,094 assignments to biological processes, and 13,135 assignments to molecular functionsA Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed that the mitogen-activated protein kinase signaling pathways are significantly enriched for DEGs between the two cell groups. Abbreviations used: BSF-1: Boletus speciosus Frost polysaccharide.

Experimentally Induced Repeated Anhydrobiosis in the Eutardigrade Richtersius coronifer.

PubMed

Czernekova, Michaela; Jönsson, K Ingemar

2016-01-01

Tardigrades represent one of the main animal groups with anhydrobiotic capacity at any stage of their life cycle. The ability of tardigrades to survive repeated cycles of anhydrobiosis has rarely been studied but is of interest to understand the factors constraining anhydrobiotic survival. The main objective of this study was to investigate the patterns of survival of the eutardigrade Richtersius coronifer under repeated cycles of desiccation, and the potential effect of repeated desiccation on size, shape and number of storage cells. We also analyzed potential change in body size, gut content and frequency of mitotic storage cells. Specimens were kept under non-cultured conditions and desiccated under controlled relative humidity. After each desiccation cycle 10 specimens were selected for analysis of morphometric characteristics and mitosis. The study demonstrates that tardigrades may survive up to 6 repeated desiccations, with declining survival rates with increased number of desiccations. We found a significantly higher proportion of animals that were unable to contract properly into a tun stage during the desiccation process at the 5th and 6th desiccations. Also total number of storage cells declined at the 5th and 6th desiccations, while no effect on storage cell size was observed. The frequency of mitotic storage cells tended to decline with higher number of desiccation cycles. Our study shows that the number of consecutive cycles of anhydrobiosis that R. coronifer may undergo is limited, with increased inability for tun formation and energetic constraints as possible causal factors.

Experimentally Induced Repeated Anhydrobiosis in the Eutardigrade Richtersius coronifer

PubMed Central

2016-01-01

Tardigrades represent one of the main animal groups with anhydrobiotic capacity at any stage of their life cycle. The ability of tardigrades to survive repeated cycles of anhydrobiosis has rarely been studied but is of interest to understand the factors constraining anhydrobiotic survival. The main objective of this study was to investigate the patterns of survival of the eutardigrade Richtersius coronifer under repeated cycles of desiccation, and the potential effect of repeated desiccation on size, shape and number of storage cells. We also analyzed potential change in body size, gut content and frequency of mitotic storage cells. Specimens were kept under non-cultured conditions and desiccated under controlled relative humidity. After each desiccation cycle 10 specimens were selected for analysis of morphometric characteristics and mitosis. The study demonstrates that tardigrades may survive up to 6 repeated desiccations, with declining survival rates with increased number of desiccations. We found a significantly higher proportion of animals that were unable to contract properly into a tun stage during the desiccation process at the 5th and 6th desiccations. Also total number of storage cells declined at the 5th and 6th desiccations, while no effect on storage cell size was observed. The frequency of mitotic storage cells tended to decline with higher number of desiccation cycles. Our study shows that the number of consecutive cycles of anhydrobiosis that R. coronifer may undergo is limited, with increased inability for tun formation and energetic constraints as possible causal factors. PMID:27828978

Effect of blood transfusion on outcome after major burn injury: a multicenter study.

PubMed

Palmieri, Tina L; Caruso, Daniel M; Foster, Kevin N; Cairns, Bruce A; Peck, Michael D; Gamelli, Richard L; Mozingo, David W; Kagan, Richard J; Wahl, Wendy; Kemalyan, Nathan A; Fish, Joel S; Gomez, Manuel; Sheridan, Robert L; Faucher, Lee D; Latenser, Barbara A; Gibran, Nicole S; Klein, Robert L; Solem, Lynn D; Saffle, Jeffrey R; Morris, Stephen E; Jeng, James C; Voigt, David; Howard, Pamela A; Molitor, Fred; Greenhalgh, David G

2006-06-01

To delineate blood transfusion practices and outcomes in patients with major burn injury. Patients with major burn injury frequently require multiple blood transfusions; however, the effect of blood transfusion after major burn injury has had limited study. Multicenter retrospective cohort analysis. Regional burn centers throughout the United States and Canada. Patients admitted to a participating burn center from January 1 through December 31, 2002, with acute burn injuries of >or=20% total body surface area. Outcome measurements included mortality, number of infections, length of stay, units of blood transfused in and out of the operating room, number of operations, and anticoagulant use. A total of 21 burn centers contributed data on 666 patients; 79% of patients survived and received a mean of 14 units of packed red blood cells during their hospitalization. Mortality was related to patient age, total body surface area burn, inhalation injury, number of units of blood transfused outside the operating room, and total number of transfusions. The number of infections per patient increased with each unit of blood transfused (odds ratio, 1.13; p<.001). Patients on anticoagulation during hospitalization received more blood than patients not on anticoagulation (16.3+/-1.5 vs. 12.3+/-1.5, p<.001). The number of transfusions received was associated with mortality and infectious episodes in patients with major burns even after factoring for indices of burn severity. The utilization of blood products in the treatment of major burn injury should be reserved for patients with a demonstrated physiologic need.

High levels of FOXP3⁺ regulatory T cells in gastric MALT lymphoma predict responsiveness to Helicobacter pylori eradication.

PubMed

Iwaya, Yugo; Kobayashi, Motohiro; Momose, Masanobu; Hiraoka, Nobuyoshi; Sakai, Yasuhiro; Akamatsu, Taiji; Tanaka, Eiji; Ohtani, Haruo; Fukuda, Minoru; Nakayama, Jun

2013-10-01

Although Helicobacter pylori eradication is a first-line treatment of gastric MALT lymphoma, roughly 25% of patients do not respond to treatment. CD4⁺ FOXP3⁺ regulatory T (Treg) cells regulate immune responses in physiological conditions and various inflammatory conditions, including H. pylori-associated diseases. Our goal was to determine how Treg cells affect responsiveness to H. pylori eradication therapy. We performed dual immunohistochemistry for CD4 and FOXP3 to evaluate the prevalence of FOXP3⁺ Treg cells in the stomach of 63 patients with MALT lymphoma and 55 patients with chronic active gastritis. Receiver operating characteristic analysis was carried out to determine the best cut-off point in differentiating H. pylori eradication responders from nonresponders. Both the FOXP3⁺/CD4⁺ cell ratio and the absolute number of FOXP3⁺ cells per high-power field in MALT lymphoma were significantly greater in H. pylori eradication responders compared with nonresponders, suggesting that Treg cells function in regression mechanisms of MALT lymphomas. Cut-off points with good sensitivities and specificities were obtained to predict eradication outcome. A high number of Treg cells or a high ratio of Treg cells to the total number of CD4⁺ T cells in gastric MALT lymphoma could predict responsiveness to eradication therapy. © 2013 John Wiley & Sons Ltd.

Targeted Ablation of CML Stem Cells

DTIC Science & Technology

2007-01-01

centuries .12 More recently, PTL has been found to have several other properties, including antitumor activity, inhibition of DNA synthesis, and...as a chemopreventive agent in a UVB-induced skin cancer animal model. 21 PTL is a potent inhibitor of NF-B activation and has been shown to directly...diluted in phosphate buffer saline (PBS). Ara-C was obtained from Sigma ( St Louis, MO). Total cell numbers were determined before and after culture for

Multigenerational Breast Cancer Risk Factors in African-American Women

DTIC Science & Technology

1996-10-01

Estrogen is of special interest in breast cancer etiology. It’s role in breast cell multiplication, the total number of menstrual cycles and possible...genetic alterations in cells during this replication are thought to be important in disease development. The typical woman is thought to have...as a key source to collect information. Introduction activities, such as the use of tobacco and alcohol; and have less access to health care services

Infections in the tissue material and their impact on the loss of transplants in the Laboratory of in vitro Cell and Tissue Culture with Tissue Bank in the years 2011-2015.

PubMed

Kitala, D; Klama-Baryła, A; Kawecki, M; Kraut, M; Łabuś, W; Glik, J; Ples, M; Tomanek, E; Nowak, M

2017-03-01

Radiation sterilization eliminates microbiological infections but causes the degradation of the cell factor. The negative result of microbiological examination for tissue transplants is one of the conditions for approval for distribution in patients. The study attempts to verify impact of the presence of microbes onto material for transplant loss. In the 2011-2015 period, we analyzed 293 donors of skin and amnion. Microbiological sampling was performed. The total of 21 strains of bacteria, molds and fungi was identified in collected tissue. The widest spectrum of strains was found in skin (17), followed by amnia (8). The total number of positive findings was 147 and was again highest in skin (129), while the number of positive findings in amnia was 18 only. The general percentage of fungal infections was very low. The presence of fungal strains was only observed in allogeneic skin (2%). Large number of microorganisms isolated from the skin before sterilization was observed, so it seems impossible to use allogeneic intravital skin. However, the intravital application of allogeneic amnion obtained from cesarean section remains to be considered.

Distribution of hydrocarbon-degrading bacteria in the soil environment and their contribution to bioremediation.

PubMed

Fukuhara, Yuki; Horii, Sachie; Matsuno, Toshihide; Matsumiya, Yoshiki; Mukai, Masaki; Kubo, Motoki

2013-05-01

A real-time PCR quantification method for indigenous hydrocarbon-degrading bacteria (HDB) carrying the alkB gene in the soil environment was developed to investigate their distribution in soil. The detection limit of indigenous HDB by the method was 1 × 10(6) cells/g-soil. The indigenous HDB were widely distributed throughout the soil environment and ranged from 3.7 × 10(7) to 5.0 × 10(8) cells/g-soil, and the ratio to total bacteria was 0.1-4.3 %. The dynamics of total bacteria, indigenous HDB, and Rhodococcus erythropolis NDKK6 (carrying alkB R2) during bioremediation were analyzed. During bioremediation with an inorganic nutrient treatment, the numbers of these bacteria were slightly increased. The numbers of HDB (both indigenous bacteria and strain NDKK6) were gradually decreased from the middle stage of bioremediation. Meanwhile, the numbers of these bacteria were highly increased and were maintained during bioremediation with an organic nutrient. The organic treatment led to activation of not only the soil bacteria but also the HDB, so an efficient bioremediation was carried out.

Capability of the two microorganisms Bifidobacterium breve B632 and Bifidobacterium breve BR03 to colonize the intestinal microbiota of children.

PubMed

Mogna, Luca; Del Piano, Mario; Mogna, Giovanni

2014-01-01

The total number of bacteria present in the gut microbiota of a newborn is consistently lower than the average found in adults, with the extent of this difference being directly related to body weight and age. It could be assumed that a lower number of viable probiotic cells is necessary to achieve significant gut colonization in infants and children. This study assessed the capability of Bifidobacterium breve B632 (DSM 24706) and Bifidobacterium breve BR03 (DSM 16604), 2 strains able to significantly inhibit some gram-negative bacteria in vitro, to integrate into the intestinal microbiota of children. Ten healthy children aged an average of 5.7±2.6 were given an oily suspension containing B. breve B632 and B. breve BR03 for 21 consecutive days. The daily dose was 100 million live cells of each strain. Fecal specimens were collected and analyzed at the beginning (d0) and at the end of the study (d21). Total fecal bifidobacteria and coliforms have been quantified by microbiological plate counts. A significant increase in total fecal bifidobacteria (from 8.99 to 9.47 log10 CFU/g, P=0.042) and a parallel decrease in total coliforms (from 8.60 to 7.93 log10 CFU/g, P=0.048) was recorded after 21 days of supplementation. An oily suspension has proved an effective way of providing probiotics to children. A lower viable cells concentration was sufficient to mediate this effect in the light of the fact that the intestinal microbiota of children harbors a considerably smaller amount of total bacteria compared with adults. In addition to gut colonization in healthy children, B. breve B632 and B. breve BR03 were able to decrease total fecal coliforms, therefore supporting their potential specific use in colicky infants.

Computerized image analysis of cell-cell interactions in human renal tissue by using multi-channel immunoflourescent confocal microscopy

NASA Astrophysics Data System (ADS)

Peng, Yahui; Jiang, Yulei; Liarski, Vladimir M.; Kaverina, Natalya; Clark, Marcus R.; Giger, Maryellen L.

2012-03-01

Analysis of interactions between B and T cells in tubulointerstitial inflammation is important for understanding human lupus nephritis. We developed a computer technique to perform this analysis, and compared it with manual analysis. Multi-channel immunoflourescent-microscopy images were acquired from 207 regions of interest in 40 renal tissue sections of 19 patients diagnosed with lupus nephritis. Fresh-frozen renal tissue sections were stained with combinations of immunoflourescent antibodies to membrane proteins and counter-stained with a cell nuclear marker. Manual delineation of the antibodies was considered as the reference standard. We first segmented cell nuclei and cell membrane markers, and then determined corresponding cell types based on the distances between cell nuclei and specific cell-membrane marker combinations. Subsequently, the distribution of the shortest distance from T cell nuclei to B cell nuclei was obtained and used as a surrogate indicator of cell-cell interactions. The computer and manual analyses results were concordant. The average absolute difference was 1.1+/-1.2% between the computer and manual analysis results in the number of cell-cell distances of 3 μm or less as a percentage of the total number of cell-cell distances. Our computerized analysis of cell-cell distances could be used as a surrogate for quantifying cell-cell interactions as either an automated and quantitative analysis or for independent confirmation of manual analysis.

Computational model for chromosomal instabilty

NASA Astrophysics Data System (ADS)

Zapperi, Stefano; Bertalan, Zsolt; Budrikis, Zoe; La Porta, Caterina

2015-03-01

Faithful segregation of genetic material during cell division requires alignment of the chromosomes between the spindle poles and attachment of their kinetochores to each of the poles. Failure of these complex dynamical processes leads to chromosomal instability (CIN), a characteristic feature of several diseases including cancer. While a multitude of biological factors regulating chromosome congression and bi-orientation have been identified, it is still unclear how they are integrated into a coherent picture. Here we address this issue by a three dimensional computational model of motor-driven chromosome congression and bi-orientation. Our model reveals that successful cell division requires control of the total number of microtubules: if this number is too small bi-orientation fails, while if it is too large not all the chromosomes are able to congress. The optimal number of microtubules predicted by our model compares well with early observations in mammalian cell spindles. Our results shed new light on the origin of several pathological conditions related to chromosomal instability.

[Malignant ovarian tumors and second look operations].

PubMed

Yasuda, M

1983-08-01

There are many problems that, the timing of S.L.O. the range of reduction, and side effects etc., we studied retrospectively for the S.L.O. The number of subjects were 302 of common epithelial and 251 of germinal cell tumors, totalling 553 cases. Among them, 41 cases were common epithelial and 49 cases germ cell origine respectively, totalling 90 cases of S.L.O. were carried out. Conclusion The 5 year survival rate of S.L.O. was observed to a significant extent. The mean survival time of S.L.O. group: were significantly prolonged. Complete surgery at the S.L.O. of the patient with epithelial and germinal affected the survival rate significantly.

Propolis extracts from the northern region of Thailand suppress cancer cell growth through induction of apoptosis pathways.

PubMed

Khacha-Ananda, Supakit; Tragoolpua, Khajornsak; Chantawannakul, Panuwan; Tragoolpua, Yingmanee

2016-12-01

The continual increase in mortality rates and number of cancer cases is a matter of serious concern in developing countries. The incorporation of natural products into classical cancer treatment approaches is a promising direction. The mechanisms of A549 and HeLa cancer cell death induction by ethanolic extracts of propolis samples from Phayao, Chiang Mai, and Nan provinces in northern Thailand were investigated in this study. The propolis extract from Chiang Mai showed the highest antioxidant activity and the greatest total phenolic content. The propolis extract from Nan also exhibited the highest total flavonoid content. The proliferation of A549 and HeLa cells grown in the presence of the propolis extracts was suppressed in a dose- and time-dependent manner. Moreover, treatment of both cancer cells with the propolis extracts showed DNA fragmentation and significantly increased the number of the apoptotic cells. On A549 cells, the extrinsic and intrinsic pathways of caspase enzymes were activated by the propolis extracts from Phayao and Chiang Mai. In the case of the propolis extract from Nan, the mechanisms involved apoptosis on the A549 cells were caspase-independent pathway. The extrinsic pathway of the caspase enzyme was triggered by all of the propolis extracts on HeLa cells. Finally, oral administration of the propolis granule produced from the propolis extract from Nan resulted in extended survival of tumour-bearing mice. Therefore, propolis extracts from the northern region of Thailand demonstrated pharmacological properties, both antioxidant and anticancer activities. From these findings, it is evident that propolis extracts can be considered as a naturally obtained agent extremely useful in cancer treatment.

Influence of copper ions on the viability and cytotoxicity of Pseudomonas aeruginosa under conditions relevant to drinking water environments.

PubMed

Dwidjosiswojo, Zenyta; Richard, Jessica; Moritz, Miriam M; Dopp, Elke; Flemming, Hans-Curt; Wingender, Jost

2011-11-01

Copper plumbing materials can be the source of copper ions in drinking water supplies. The aim of the current study was to investigate the influence of copper ions on the viability and cytotoxicity of the potential pathogen Pseudomonas aeruginosa that presents a health hazard when occurring in building plumbing systems. In batch experiments, exposure of P. aeruginosa (10(6)cells/mL) for 24h at 20°C to copper-containing drinking water from domestic plumbing systems resulted in a loss of culturability, while total cell numbers determined microscopically did not decrease. Addition of the chelator diethyldithiocarbamate (DDTC) to copper-containing water prevented the loss of culturability. When suspended in deionized water with added copper sulfate (10 μM), the culturability of P. aeruginosa decreased by more than 6 log units, while total cell counts, the concentration of cells with intact cytoplasmic membranes, determined with the LIVE/DEAD BacLight kit, and the number of cells with intact 16S ribosomal RNA, determined by fluorescent in situ hybridization, remained unchanged. When the chelator DDTC was added to copper-stressed bacteria, complete restoration of culturability was observed to occur within 14 d. Copper-stressed bacteria were not cytotoxic towards Chinese hamster ovary (CHO-9) cells, while untreated and resuscitated bacteria caused an almost complete decrease of the concentration of viable CHO-9 cells within 24 h. Thus, copper ions in concentrations relevant to drinking water in plumbing systems seem to induce a viable but non-culturable (VBNC) state in P. aeruginosa accompanied by a loss of culturability and cytotoxicity, and VBNC cells can regain both culturability and cytotoxicity, when copper stress is abolished. Copyright © 2011 Elsevier GmbH. All rights reserved.

Clinostat rotation induces apoptosis in luteal cells of the pregnant rat

NASA Technical Reports Server (NTRS)

Yang, Hyunwon; Bhat, Ganapathy K.; Sridaran, Rajagopala

2002-01-01

Recent studies have shown that microgravity induces changes at the cellular level, including apoptosis. However, it is unknown whether microgravity affects luteal cell function. This study was performed to assess whether microgravity conditions generated by clinostat rotation induce apoptosis and affect steroidogenesis by luteal cells. Luteal cells isolated from the corpora lutea of Day 8 pregnant rats were placed in equal numbers in slide flasks (chamber slides). One slide flask was placed in the clinostat and the other served as a stationary control. At 48 h in the clinostat, whereas the levels of progesterone and total cellular protein decreased, the number of shrunken cells increased. To determine whether apoptosis occurred in shrunken cells, Comet and TUNEL assays were performed. At 48 h, the percentage of apoptotic cells in the clinostat increased compared with that in the control. To investigate how the microgravity conditions induce apoptosis, the active mitochondria in luteal cells were detected with JC-1 dye. Cells in the control consisted of many active mitochondria, which were evenly distributed throughout the cell. In contrast, cells in the clinostat displayed fewer active mitochondria, which were distributed either to the outer edge of the cell or around the nucleus. These results suggest that mitochondrial dysfunction induced by clinostat rotation could lead to apoptosis in luteal cells and suppression of progesterone production.

Treatment with at Homeopathic Complex Medication Modulates Mononuclear Bone Marrow Cell Differentiation

PubMed Central

Cesar, Beatriz; Abud, Ana Paula R.; de Oliveira, Carolina C.; Cardoso, Francolino; Bernardi, Raffaello Popa Di; Guimarães, Fernando S. F.; Gabardo, Juarez; de Freitas Buchi, Dorly

2011-01-01

A homeopathic complex medication (HCM), with immunomodulatory properties, is recommended for patients with depressed immune systems. Previous studies demonstrated that the medication induces an increase in leukocyte number. The bone marrow microenvironment is composed of growth factors, stromal cells, an extracellular matrix and progenitor cells that differentiate into mature blood cells. Mice were our biological model used in this research. We now report in vivo immunophenotyping of total bone marrow cells and ex vivo effects of the medication on mononuclear cell differentiation at different times. Cells were examined by light microscopy and cytokine levels were measured in vitro. After in vivo treatment with HCM, a pool of cells from the new marrow microenvironment was analyzed by flow cytometry to detect any trend in cell alteration. The results showed decreases, mainly, in CD11b and TER-119 markers compared with controls. Mononuclear cells were used to analyze the effects of ex vivo HCM treatment and the number of cells showing ring nuclei, niche cells and activated macrophages increased in culture, even in the absence of macrophage colony-stimulating factor. Cytokines favoring stromal cell survival and differentiation in culture were induced in vitro. Thus, we observe that HCM is immunomodulatory, either alone or in association with other products. PMID:19736221

In vitro expansion of Lin{sup +} and Lin{sup −} mononuclear cells from human peripheral blood

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

Norhaiza, H. Siti; Zarina, Z. A. Intan; Hisham, Z. A. Shahrul

2013-11-27

Haematopoietic stem cells (HSCs) are used in the therapy of blood disorders due to the ability of these cells to reconstitute haematopoietic lineage cells when transplanted into myeloablative recipients. However, substantial number of cells is required in order for the reconstitution to take place. Since HSCs present in low frequency, larger number of donor is required to accommodate the demand of transplantable HSCs. Therefore, in vitro expansion of HSCs will have profound impact on clinical purposes. The aim of this study was to expand lineage negative (Lin{sup −}) stem cells from human peripheral blood. Total peripheral blood mononuclear cells (PBMNCs)more » were fractionated from human blood by density gradient centrifugation. Subsequently, PBMNCs were subjected to magnetic assisted cell sorter (MACS) which depletes lineage positive (Lin{sup +}) mononuclear cells expressing lineage positive markers such as CD2, CD3, CD11b, CD14, CD15, CD16, CD19, CD56, CD123, and CD235a to obtained Lin{sup −} cell population. The ability of Lin{sup +} and Lin{sup −} to survive in vitro was explored by culturing both cell populations in complete medium consisting of Alpha-Minimal Essential Medium (AMEM) +10% (v/v) Newborn Calf Serum (NBCS)+ 2% (v/v) pen/strep. In another experiment, Lin{sup +} and Lin{sup −} were cultured with complete medium supplemented with 10ng/mL of the following growth factors: stem cell factor (SCF), interleukin (IL)-3, granulocyte-macrophage colony stimulating factor (GM-CSF), 2IU/mL of Erythropoietin (Epo) and 20ng/mL of IL-6. Three samples were monitored in static culture for 22 days. The expansion potential was assessed by the number of total viable cells, counted by trypan blue exclusion assay. It was found that Lin{sup +} mononuclear cells were not able to survive either in normal proliferation medium or proliferation medium supplemented with cytokines. Similarly, Lin{sup −} stem cells were not able to survive in proliferation medium however, addition of cytokines into the proliferation medium support Lin{sup −} stem cells for at least 18 days. The Lin{sup −} stem cells started to response to the cytokines added as early as Day 2 of culture. It is concluded that Lin{sup −} stem cells can be expanded in vitro by culturing in proliferation medium supplemented with cytokines.« less

Casein-specific IL-4- and IL-13-secreting T cells: a tool to implement diagnosis of cow's milk allergy.

PubMed

Michaud, B; Aroulandom, J; Baiz, N; Amat, F; Gouvis-Echraghi, R; Candon, S; Foray, A-P; Couderc, R; Bach, J-F; Chatenoud, L; Just, J

2014-11-01

Cow's milk allergy (CMA) is a frequent food allergy in young children. The oral food challenge is the gold standard for diagnosis, and there is currently no reliable biological test. Our aim was to evaluate the diagnostic potential of a functional assay quantifying allergen-specific Th2 cells in CMA children. A total of 29 children aged 2.8-10.5 years underwent a double-blind, placebo-controlled food challenge (DBPCFC) to cow's milk. Blood was collected before performing the DBPCFC, and peripheral mononuclear cells were cultured in an 18-h ELISpot assay with casein, α-lactalbumin, or β-lactoglobulin. Numbers of antigen-specific IL-4- and IL-13-secreting lymphocytes and serum-specific IgE, IgG4, and total IgE levels were assessed. Receiver operating characteristic (ROC) curves were generated. A total of 17 (59%) children reacted to cow's milk and were therefore considered as allergic to cow's milk (CMA). The mean number of casein-specific IL-4- and IL-13-secreting T cells was higher in CMA than in non-CMA children (P = 0.009, 0.004, respectively). Moreover, it was inversely correlated with the cumulative dose of cow's milk tolerated (P = 0.003, 0.0009, respectively). ROC curve of combined IL-4 and IL-13 analysis showed an area under the curve of 0.98 (95% CI 0.90-1.06). For a cutoff of 10 IL-4- and 12 IL-13-secreting T cells, sensitivity and negative predictive value were 100%. Enumeration of casein-specific IL-4- and IL-13-secreting T cells appears a promising tool to improve diagnosis and, if confirmed in larger studies, could permit less frequent use of the oral food challenge. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

High IFN-gamma and TNF production by peripheral NK cells of Colombian patients with different clinical presentation of Plasmodium falciparum

PubMed Central

2012-01-01

Background In Colombia, Plasmodium falciparum infection rarely results in severe disease or mortality compared to infections in African populations. During natural infection NK cells exhibit a cytolytic effect and regulate dendritic cells, macrophages, neutrophils as well as affect antigen specific T and B cell responses. To characterize the NK cells in P. falciparum infected patients of a highly endemic region of Colombia, the degree of NK proliferation and production of IFN gamma and TNF production in these cells were explored. Methods Seventeen patients with acute and three with severe P. falciparum malaria patients from the Northwest region of the country were recruited in the study. In addition, 20 healthy controls were included: 10 from Medellin (no-transmission area) and 10 from the Uraba region (a malaria endemic area). Immunophenotypic analysis of peripheral mononuclear cells was performed by FACS to detect total number of NK cells, subtypes and intracellular IFNγ and TNF production by NK cells in the different patient groups. Results The total mean CD56+/CD3- NK cell proportions in acute and severe malaria subjects were 9.14% (7.15%CD56dim, 2.01%CD56bright) and 19.62% (16.05%CD56dim, 3.58%CD56bright), respectively, in contrast to healthy controls from endemic (total mean CD56+/CD3-1.2%) and non-endemic area (total mean CD56+/CD3- 0.67%). Analysis of basal IFNγ and TNF levels confirmed the CD56bright NK population as the main cytokine producer (p < 0.0001) in the groups affected with malaria, with the CD56dim NK cell exhibiting the highest potential of TNF production after stimulus in the acute malaria group. Conclusions The results confirm the important role of not only CD56bright but also of CD56dim NK cell populations as producers of the two cytokines in malaria patients in Colombia. PMID:22316273

Free energy analysis of cell spreading.

PubMed

McEvoy, Eóin; Deshpande, Vikram S; McGarry, Patrick

2017-10-01

In this study we present a steady-state adaptation of the thermodynamically motivated stress fiber (SF) model of Vigliotti et al. (2015). We implement this steady-state formulation in a non-local finite element setting where we also consider global conservation of the total number of cytoskeletal proteins within the cell, global conservation of the number of binding integrins on the cell membrane, and adhesion limiting ligand density on the substrate surface. We present a number of simulations of cell spreading in which we consider a limited subset of the possible deformed spread-states assumed by the cell in order to examine the hypothesis that free energy minimization drives the process of cell spreading. Simulations suggest that cell spreading can be viewed as a competition between (i) decreasing cytoskeletal free energy due to strain induced assembly of cytoskeletal proteins into contractile SFs, and (ii) increasing elastic free energy due to stretching of the mechanically passive components of the cell. The computed minimum free energy spread area is shown to be lower for a cell on a compliant substrate than on a rigid substrate. Furthermore, a low substrate ligand density is found to limit cell spreading. The predicted dependence of cell spread area on substrate stiffness and ligand density is in agreement with the experiments of Engler et al. (2003). We also simulate the experiments of Théry et al. (2006), whereby initially circular cells deform and adhere to "V-shaped" and "Y-shaped" ligand patches. Analysis of a number of different spread states reveals that deformed configurations with the lowest free energy exhibit a SF distribution that corresponds to experimental observations, i.e. a high concentration of highly aligned SFs occurs along free edges, with lower SF concentrations in the interior of the cell. In summary, the results of this study suggest that cell spreading is driven by free energy minimization based on a competition between decreasing cytoskeletal free energy and increasing passive elastic free energy. Copyright © 2017 Elsevier Ltd. All rights reserved.

Association between memory B-cells and clinical and immunological features of primary Sjögren's syndrome and Sicca patients.

PubMed

Barcelos, Filipe; Martins, Catarina; Papoila, Ana; Geraldes, Carlos; Cardigos, Joana; Nunes, Glória; Lopes, Teresa; Alves, Nuno; Vaz-Patto, José; Branco, Jaime; Borrego, Luís-Miguel

2018-06-01

B-cells play a pivotal role in primary Sjögren's syndrome (pSS) pathogenesis. We aim to (1) evaluate the distribution of B-lymphocyte subpopulations in pSS and Sicca patients, (2) establish cut-off points that discriminate pSS from controls, (3) evaluate the association between memory B-cells and phenotypic features in pSS. We included 57 pSS patients, 68 Sicca and 24 healthy controls. Circulating B-cells were characterized by flow cytometry as naïve and memory subsets and classified from Bm1 to Bm5. Compared to controls, pSS patients had lower percentages (29.5 vs 44.4%) and absolute numbers (47 vs 106 cells/µl) of memory B-cells. Through ROC curves, a cut-off of ≤ 58 total memory B-cells/µl yielded a specificity of 0.88 and a sensitivity of 0.60 for pSS, and was met by 59.6% of pSS patients, 38.8% of Sicca and 12.5% of controls. A cut-off of < 23.5 Switched-memory B-cells/µl yielded a specificity of 0.88 and a sensitivity of 0.54 and was met by 54.4% of pSS patients, 37.3% of Sicca and 12.5% of controls. In pSS, lower total memory B-cells count was associated with longer disease duration (14.3 vs 8.1 years, p = 0.006) and more active disease profile, as evaluated by the European League Against Rheumatism (EULAR) Sjögren's Syndrome Disease Activity Index (ESSDAI) (3.1 vs 1.4, p = 0.043). Decreased numbers of memory B-cells clearly discriminated pSS from controls and can also have prognostic value. It remains to be clarified whether Sicca patients with decreased memory B-cells represent pSS and if B-cell profiling could help in the diagnosis of pSS.

Cysteinyl Leukotriene 1 Receptor Expression Associated With Bronchial Inflammation in Severe Exacerbations of COPD

PubMed Central

Zhu, Jie; Bandi, Venkata; Qiu, Shengyang; Figueroa, David J.; Evans, Jilly F.; Barnes, Neil; Guntupalli, Kay K.

2012-01-01

Background: Cysteinyl leukotriene 1 (CysLT1) receptor expression is known to be increased in the airway mucosa of patients with asthma, especially during exacerbations; however, nothing is known of its expression in COPD. Methods: We applied immunohistochemistry and in situ hybridization to endobronchial biopsies to determine inflammatory cell CysLT1 receptor protein and mRNA expression in the following: (1) 15 nonsmoker control subjects (NSC), (2) 16 smokers with moderate to severe COPD in its stable phase (S-COPD), and (3) 15 smokers with COPD hospitalized for a severe exacerbation (SE-COPD). Results: The total number of bronchial mucosal inflammatory cells (CD45+) and those expressing CysLT1 receptor protein were significantly greater in SE-COPD (CysLT1 receptor protein: median [range] = 139 [31-634]) as compared with S-COPD (32 [6-114]) or NSC (16 [4-66]) (P < .001 for both). CysLT1 receptor gene expression showed similar differences. A greater proportion of CD451 cells expressed CysLT1 receptor protein in SE-COPD (median [range] = 22% [8-81]) compared with S-COPD (10% [4-32]) (P < .03) or NSC (7% [1-19]) (P < .002). In SE-COPD, the relative frequencies of CysLT1 receptor-expressing cells were as follows: tryptase1 mast cells > CD681 monocytes/macrophage > neutrophils > CD201 B lymphocytes = EG21 eosinophils. Moreover, there were positive correlations between the numbers of cells expressing CysLT1 receptor protein and the numbers of CD451 cells (r = 0.78; P < .003) and tryptase1 mast cells (r = 0.62; P < .02). Conclusions: Bronchial mucosal CysLT1 receptor-positive inflammatory cells are present in the bronchial mucosa in COPD in greatest number in those experiencing a severe exacerbation. PMID:22871757

Immunoexpression of tryptase-positive mast cells in periapical granulomas and radicular cysts.

PubMed

Costa Neto, H; de Andrade, A L D L; Gordón-Núñez, M A; Freitas, R de A; Galvão, H C

2015-08-01

To evaluate and compare the immunoexpression of tryptase in samples of periapical granulomas (PGs) and radicular cysts (RCs) correlating it with the type of lesion, localization, intensity of the inflammatory infiltrate and thickness of the cystic epithelial lining, in order to gain insight into the phlogistic role of these cells in the lesions studied. Twenty-five PGs and twenty-five RCs obtained from human teeth without endodontic treatment were submitted to morphological and immunohistochemical analysis using anti-tryptase antibody. Mast cells were identified and counted in three regions: intra-epithelial, central/superficial and deep portions. The data were analysed using the Mann-Whitney U-test (P < 0.05). In comparison with RCs, PGs exhibited higher immunoexpression of tryptase-positive mast cells located in both central/superficial and deep regions (P < 0.001 and P < 0.001, respectively). When considering the total number of mast cells and disregarding the location, the number of tryptase-positive mast cells increased gradually from RCs to PGs (P < 0.001). Lesions with inflammatory infiltrate grade III had greater number of tryptase-positive mast cells located in both central/superficial and deep regions than lesions with inflammatory infiltrates grade II (P = 0.045 and P = 0.025). When the location was ignored, the lesions with inflammatory infiltrate grade III also exhibited higher immunostaining of tryptase-positive mast cells (P = 0.01). Tryptase-positive mast cells were present in chronic periapical lesions in a larger number in periapical granulomas than in radicular cysts, in both central/superficial and deep regions. © 2014 International Endodontic Journal. Published by John Wiley & Sons Ltd.

Optimised cord blood sample selection for small‑scale CD34+ cell immunomagnetic isolation.

PubMed

Perdomo-Arciniegas, Ana-María; Vernot, Jean-Paul

2012-03-01

Haematopoietic stem cells (HSCs) are defined as multipotential cells, capable of self-renewal and reconstituting in vivo the haematopoietic compartment. The CD34 antigen is considered an important HSCs marker in humans. Immunomagnetic isolation, by targeting CD34 antigen, is widely used for human HSC separation. This method allows the enrichment of human HSCs that are present at low frequencies in umbilical cord blood (CB). Immunomagnetic CD34+-cell isolation reproducibility, regarding cell yield and purity, is affected by the CD34+ cell frequency and total cell numbers present in a given sample; CB HSC purification may thus yield variable results, which also depend on the volume and density fractionation-derived cell loss of a CB sample. The uncertainty of such an outcome and associated technical costs call for a cost-effective sample screening strategy. A correlation analysis using clinical and laboratory data from 59 CB samples was performed to establish predictive variables for CD34+-immunomagnetic HSCs isolation. This study described the positive association of CD34+-cell isolation with white and red cell numbers present after cell fractionation. Furthermore, purity has been correlated with lymphocyte percentages. Predictive variable cut-off values, which are particularly useful in situations involving low CB volumes being collected (such as prevalent late umbilical cord clamping clinical practice), were proposed for HSC isolation sampling. Using the simple and cost-effective CB sample screening criteria described here would lead to avoiding costly inefficient sample purification, thereby ensuring that pure CD34+ cells are obtained in the desired numbers following CD34 immunomagnetic isolation.

Dietary Restriction and Fasting Arrest B and T Cell Development and Increase Mature B and T Cell Numbers in Bone Marrow

PubMed Central

Shushimita, Shushimita; de Bruijn, Marjolein J. W.; de Bruin, Ron W. F.; IJzermans, Jan N. M.; Hendriks, Rudi W.; Dor, Frank J. M. F.

2014-01-01

Dietary restriction (DR) delays ageing and extends life span. Both long- and short-term DR, as well as short-term fasting provide robust protection against many “neuronal and surgery related damaging phenomena” such as Parkinson’s disease and ischemia-reperfusion injury. The exact mechanism behind this phenomenon has not yet been elucidated. Its anti-inflammatory actions prompted us to thoroughly investigate the consequences of DR and fasting on B and T cell compartments in primary and secondary lymphoid organs of male C57Bl/6 mice. In BM we found that DR and fasting cause a decrease in the total B cell population and arrest early B cell development, while increasing the number of recirculating mature B cells. In the fasting group, a significant reduction in peripheral B cell counts was observed in both spleen and mesenteric lymph nodes (mLN). Thymopoiesis was arrested significantly at double negative DN2 stage due to fasting, whereas DR resulted in a partial arrest of thymocyte development at the DN4 stage. Mature CD3+ T cell populations were increased in BM and decreased in both spleen and mLN. Thus, DR arrests B cell development in the BM but increases the number of recirculating mature B cells. DR also arrests maturation of T cells in thymus, resulting in depletion of mature T cells from spleen and mLN while recruiting them to the BM. The functional relevance in relation to protection against organ damage needs to be determined. PMID:24504160

Clustering single cells: a review of approaches on high-and low-depth single-cell RNA-seq data.

PubMed

Menon, Vilas

2017-12-11

Advances in single-cell RNA-sequencing technology have resulted in a wealth of studies aiming to identify transcriptomic cell types in various biological systems. There are multiple experimental approaches to isolate and profile single cells, which provide different levels of cellular and tissue coverage. In addition, multiple computational strategies have been proposed to identify putative cell types from single-cell data. From a data generation perspective, recent single-cell studies can be classified into two groups: those that distribute reads shallowly over large numbers of cells and those that distribute reads more deeply over a smaller cell population. Although there are advantages to both approaches in terms of cellular and tissue coverage, it is unclear whether different computational cell type identification methods are better suited to one or the other experimental paradigm. This study reviews three cell type clustering algorithms, each representing one of three broad approaches, and finds that PCA-based algorithms appear most suited to low read depth data sets, whereas gene clustering-based and biclustering algorithms perform better on high read depth data sets. In addition, highly related cell classes are better distinguished by higher-depth data, given the same total number of reads; however, simultaneous discovery of distinct and similar types is better served by lower-depth, higher cell number data. Overall, this study suggests that the depth of profiling should be determined by initial assumptions about the diversity of cells in the population, and that the selection of clustering algorithm(s) is subsequently based on the depth of profiling will allow for better identification of putative transcriptomic cell types. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Cyclin-dependent kinase 5 mediates adult OPC maturation and myelin repair through modulation of Akt and GsK-3β signaling.

PubMed

Luo, FuCheng; Burke, Kathryn; Kantor, Christopher; Miller, Robert H; Yang, Yan

2014-07-30

Failure of remyelination in diseases, such as multiple sclerosis (MS), leads to permanent axonal damage and irreversible functional loss. The mechanisms controlling remyelination are currently poorly understood. Recent studies implicate the cyclin-dependent kinase 5 (Cdk5) in regulating oligodendrocyte (OL) development and myelination in CNS. In this study, we show that Cdk5 is also an important regulator of remyelination. Pharmacological inhibition of Cdk5 inhibits repair of lysolecithin lesions. This inhibition is a consequence of Cdk5 disruption in neural cells because remyelination in slice cultures is blocked by Cdk5 inhibitors, whereas specific deletion of Cdk5 in OLs inhibits myelin repair. In CNP-Cre;Cdk5(fl/fl) conditional knock-out mouse (Cdk5 cKO), myelin repair was delayed significantly in response to focal demyelinating lesions compared with wild-type animals. The lack of myelin repair was reflected in decreased expression of MBP and proteolipid protein and a reduction in the total number of myelinated axons in the lesion. The number of CC1(+) cells in the lesion sites was significantly reduced in Cdk5 cKO compared with wild-type animals although the total number of oligodendrocyte lineage cells (Olig2(+) cells) was increased, suggesting that Cdk5 loss perturbs the transition of early OL lineage cell into mature OL and subsequent remyelination. The failure of remyelination in Cdk5 cKO animals was associated with a reduction in signaling through the Akt pathway and an enhancement of Gsk-3β signaling pathways. Together, these data suggest that Cdk5 is critical in regulating the transition of adult oligodendrocyte precursor cells to mature OLs that is essential for myelin repair in adult CNS. Copyright © 2014 the authors 0270-6474/14/3410415-15$15.00/0.

Dietary supplementation of probiotic Bacillus polyfermenticus, Bispan strain, modulates natural killer cell and T cell subset populations and immunoglobulin G levels in human subjects.

PubMed

Kim, Hyun-Sook; Park, Hyunjin; Cho, In-Young; Paik, Hyun-Dong; Park, Eunju

2006-01-01

A probiotic is a viable microbial dietary supplement that has beneficial effects such as prevention and treatment of specific gastrointestinal disorders, including counteracting gut barrier dysfunction associated with inflammation and infection. Probiotic Bacillus polyfermenticus, which is commonly called Bispan strain, has been appropriately used for the treatment of long-term intestinal disorders. The use of B. polyfermenticus for immune-related chronic intestinal disease may be appropriate considering that about 80% of the body's immune system is localized in the gastrointestinal tract. The current study aimed to evaluate the effect of probiotic B. polyfermenticus on the immune response of human subjects through the quantification of immune cell population and serum levels of immunoglobulins (Igs). Twenty-five male subjects, 20-35 years of age, were randomly assigned to either a control group (n =12) supplemented with a placebo or the experimental group (n = 13) supplemented with B. polyfermenticus tablets at a dose of 3.1 x 10(8) colony-forming units/day for 8 weeks. Dietary intake analyses from 3-day dietary records from three consecutive days including one weekend day and two weekdays revealed no significant differences in total energy and nutrient intakes between the two groups. The humoral immune response was monitored by the number of total B lymphocytes and serum concentrations of IgG, IgA, and IgM. To investigate the changes in immune cell populations, percentages of total T lymphocytes, CD4+ helper T cells, CD8+ cytotoxic T cells, and CD56+ natural killer (NK) cells were quantified. The concentration of IgG in the experimental group was 12% higher than in the placebo group after 8 weeks of Bispan supplementation. Also, the percentages of CD4+ helper T cells, CD8+cytotoxic T cells, and CD56+ NK cells in the Bispan strain-supplemented group were 32%, 28%, and 35% higher, respectively, compared with the control group. Because of a higher increment of the CD4+ T cell subset than CD8+ T cells, the ratio of CD4+/CD8+ T cells was greater in the experimental group. This study suggests that the supplementation of B. polyfermenticus has a potentially positive effect on immune function by enhancing IgG production as well as by modulating the number of immune cell population such as CD4+ and CD8+ T cells and NK cells.

Proliferation of protease-enriched mast cells in sarcoptic skin lesions of raccoon dogs.

PubMed

Noviana, D; W Harjanti, D; Otsuka, Y; Horii, Y

2004-07-01

Skin sites, tongue, lung, liver, jejunum and rectum from two raccoon dogs with Sarcoptes scabiei infestation and five normal (control) raccoon dogs were examined in terms of the distribution, proteoglycan properties and protease activity of mast cells. Infestation with S. scabiei caused a significant increase in the number of dermal mast cells. While the number of mast cells (average +/- standard deviation) in specimens of skin from the dorsum, dorsal neck, dorsal hind foot and dorsal fore foot was 40.0 +/- 19.8/mm2 in control animals, it was 236.1 +/- 58.9/mm2 in the skin of mange-infested animals. Histochemical analysis revealed the glycosaminoglycan, heparin, within the mast cells of all organs examined in both control and affected animals. Enzyme-histochemical detection of serine proteases demonstrated an increase in mast-cell-specific protease activity (i.e., chymase and tryptase) in the skin of infested animals. The percentage of mast cells demonstrating chymase activity was 53.0 +/- 27.4% in control animals and 73.8 +/- 19.4% in mite-infested animals. The corresponding results for tryptase activity were 53.5 +/- 25.2% and 89.4 +/- 9.8%. Increases in mast cell chymase or tryptase activity, or both, were also observed within other organs of the infected animals, but the total number of mast cells found at such sites (with the exception of liver and ventrolateral pinna) did not differ from those of control animals. Copyright 2004 Elsevier Ltd.

Adverse early life environment increases hippocampal microglia abundance in conjunction with decreased neural stem cells in juvenile mice.

PubMed

Cohen, Susan; Ke, Xingrao; Liu, Qiuli; Fu, Qi; Majnik, Amber; Lane, Robert

2016-12-01

Adverse maternal lifestyle resulting in adverse early life environment (AELE) increases risks for neuropsychiatric disorders in offspring. Neuropsychiatric disorders are associated with impaired neurogenesis and neuro-inflammation in the hippocampus (HP). Microglia are neuro-inflammatory cells in the brain that regulate neurogenesis via toll-like receptors (TLR). TLR-9 is implicated in neurogenesis inhibition and is responsible for stress-related inflammatory responses. We hypothesized that AELE would increase microglia cell count and increase TLR-9 expression in juvenile mouse HP. These increases in microglia cell count and TLR-9 expression would be associated with decrease neural stem cell count and neuronal cell count. We developed a mouse model of AELE combining Western diet and a stress environment. Stress environment consisted of random change from embryonic day 13 (E13) to E17 as well as static change in maternal environment from E13 to postnatal day 21(P21). At P21, we measured hippocampal cell numbers of microglia, neural stem cell and neuron, as well as hippocampal TLR-9 expression. AELE significantly increased total microglia number and TLR-9 expression in the hippocampus. Concurrently, AELE significantly decreased neural stem cell and neuronal numbers. AELE increased the neuro-inflammatory cellular response in the juvenile HP. We speculate that increased neuro-inflammatory responses may contribute to impaired neurogenesis seen in this model. Copyright © 2016 ISDN. Published by Elsevier Ltd. All rights reserved.

Dependence of the photovoltaic performance of pseudomorphic InGaN/GaN multiple-quantum-well solar cells on the active region thickness

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

Mukhtarova, Anna; Valdueza-Felip, Sirona; Redaelli, Luca

2016-04-18

We investigate the photovoltaic performance of pseudomorphic In{sub 0.1}Ga{sub 0.9}N/GaN multiple-quantum well (MQW) solar cells as a function of the total active region thickness. An increase in the number of wells from 5 to 40 improves the short-circuit current and the open-circuit voltage, resulting in a 10-fold enhancement of the overall conversion efficiency. Further increasing the number of wells leads to carrier collection losses due to an incomplete depletion of the active region. Capacitance-voltage measurements point to a hole diffusion length of 48 nm in the MQW region.

Identification and quantitation of morphological cell types in electrophoretically separated human embryonic kidney cell cultures

NASA Technical Reports Server (NTRS)

Williams, K. B.; Kunze, M. E.; Todd, P. W.

1985-01-01

Four major cell types were identified by phase microscopy in early passage human embryonic kidney cell cultures. They are small and large epithelioid, domed, and fenestrated cells. Fibroblasts are also present in some explants. The percent of each cell type changes with passage number as any given culture grows. As a general rule, the fraction of small epithelioid cells increases, while the fraction of fenestrated cells, always small, decreases further. When fibroblasts are present, they always increase in percentage of the total cell population. Electrophoretic separation of early passage cells showed that the domed cells have the highest electrophoretic mobility, fibroblasts have an intermediate high mobility, small epithelioid cells have a low mobility, broadly distributed, and fenestrated cells have the lowest mobility. All cell types were broadly distributed among electrophoretic subfractions, which were never pure but only enriched with respect to a given cell type.

Direct Determination of Activities for Microorganisms of Chesapeake Bay Populations

PubMed Central

Tabor, Paul S.; Neihof, Rex A.

1984-01-01

We used three methods in determination of the metabolically active individual microorganisms for Chesapeake Bay surface and near-bottom populations over a period of a year. Synthetically active bacteria were recognized as enlarged cells in samples amended with nalidixic acid and yeast extract and incubated for 6 h. Microorganisms with active electron transport systems were identified by the reduction of a tetrazolium salt electron acceptor. Microorganisms active in uptake of amino acids, thymidine, and acetate were determined by microautoradiography. In conjunction with enumeration of active organisms, a total direct count was made for each sample preparation by epifluorescence microscopy. For the majority of samples, numbers of amino acid uptake-active organisms were greater than numbers of organisms determined to be active by other direct measurements. Within a sample, the numbers of uptake-active organisms (amino acids or thymidine) and electron transport system-active organisms were significantly different for 68% of the samples. Numbers of synthetically active bacteria were generally less than numbers determined by the other direct activity measurements. The distribution of total counts in the 11 samplings showed a seasonal pattern, with significant dependence on in situ water temperature, increasing from March to September and then decreasing through February. Synthetically active bacteria and amino acid uptake-active organisms showed a significant dependence on in situ temperature, independent of the function of temperature on total counts. Numbers of active organisms determined by at least one of the methods used exceeded 25% of the total population of all samplings, and from June through September, >85% of the total population was found to be active by at least one direct activity measurement. Thus, active rather than dormant organisms compose a major portion of the microbial population in this region of Chesapeake Bay. PMID:16346659

Direct determination of activities for microorganisms of chesapeake bay populations.

PubMed

Tabor, P S; Neihof, R A

1984-11-01

We used three methods in determination of the metabolically active individual microorganisms for Chesapeake Bay surface and near-bottom populations over a period of a year. Synthetically active bacteria were recognized as enlarged cells in samples amended with nalidixic acid and yeast extract and incubated for 6 h. Microorganisms with active electron transport systems were identified by the reduction of a tetrazolium salt electron acceptor. Microorganisms active in uptake of amino acids, thymidine, and acetate were determined by microautoradiography. In conjunction with enumeration of active organisms, a total direct count was made for each sample preparation by epifluorescence microscopy. For the majority of samples, numbers of amino acid uptake-active organisms were greater than numbers of organisms determined to be active by other direct measurements. Within a sample, the numbers of uptake-active organisms (amino acids or thymidine) and electron transport system-active organisms were significantly different for 68% of the samples. Numbers of synthetically active bacteria were generally less than numbers determined by the other direct activity measurements. The distribution of total counts in the 11 samplings showed a seasonal pattern, with significant dependence on in situ water temperature, increasing from March to September and then decreasing through February. Synthetically active bacteria and amino acid uptake-active organisms showed a significant dependence on in situ temperature, independent of the function of temperature on total counts. Numbers of active organisms determined by at least one of the methods used exceeded 25% of the total population of all samplings, and from June through September, >85% of the total population was found to be active by at least one direct activity measurement. Thus, active rather than dormant organisms compose a major portion of the microbial population in this region of Chesapeake Bay.

Actual and Simulated Weightlessness Inhibit Osteogenesis in Long Bone Metaphysis by Different Mechanisms

NASA Technical Reports Server (NTRS)

Roberts, W. E.

1985-01-01

Weightlessness and simulated weightlessness inhibit the rate of periosteal bone formation in long bones. Formation of preosteoblasts is suppressed in periodontal ligament (PDL) of maxillary molars, which suggests a generalized block in osteoblast histogenesis. Growth in length of long bones is decreased by simulated weightlessness, but there are no reliable data on the influence of actual weightlessness on metaphyseal growth. The nuclear size assay for assessing relative numbers of osteoblast precursor cells was utilized in the primary spongiosa of growing long bones subjected to actual and simulated weightlessness. It is found that: (1) Actual weightlessness decreases total number of osteogenic cells and inhibits differentiation of osteoblast precursor cells, (2) Simulated weightlessness suppresses only osteoblast differentation; and (3) The nuclear morphometric assay is an effective means of assessing osteogenic activity in the growing metaphysis or long bones.

SU-E-T-352: Why Is the Survival Rate Low in Oropharyngeal Squamous Cell Carcinoma?

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

Huang, Z; Feng, Y; Rasmussen, K

2014-06-01

Purpose: Tumors are composed of a large number of clonogens that have the capability of indefinite reproduction. Even when there is complete clinical or radiographic regression of the gross tumor mass after treatment, tumor recurrence can occur if the clonogens are not completely eradicated by radiotherapy. This study was to investigate the colonogen number and its association with the tumor control probability (TCP) in oropharyngeal squamous cell carcinoma (OSCCA). Methods: A literature search was conducted to collect clinical information of patients with OSCCA, including the prescription dose, tumor volume and survival rate. The linear-quadratic (LQ) model was incorporated into TCPmore » model for clinical data analysis. The total dose ranged from 60 to 70 Gy and tumor volume ranged from 10 to 50 cc. The TCP was calculated for each group according to tumor size and dose. The least χ{sup 2} method was used to fit the TCP calculation to clinical data while other LQ model parameters (α, β) were adopted from the literature, due to the limited patient data. Results: A total of 190 patients with T2–T4 OSCCA were included. The association with HPV was not available for all the patients. The 3-year survival rate was about 82% for T2 squamous cell carcinoma and 40% for advanced tumors. Fitting the TCP model to the survival data, the average clonogen number was 1.56×10{sup 12}. For the prescription dose of 70 Gy, the calculated TCP ranged from 40% to 90% when the tumor volume varied from 10 to 50 cc. Conclusion: Our data suggests variation between the clonogen number and TCP in OSCCA. Tumors with larger colonogen number tend to have lower TCP and therefore dose escalation above 70 Gy may be indicated in order to improve the TCP and survival rate. Our result will require future confirmation with a large number of patients.« less

Parts-Per-Billion Mass Measurement Accuracy Achieved through the Combination of Multiple Linear Regression and Automatic Gain Control in a Fourier Transform Ion Cyclotron Resonance Mass Spectrometer

PubMed Central

Williams, D. Keith; Muddiman, David C.

2008-01-01

Fourier transform ion cyclotron resonance mass spectrometry has the ability to achieve unprecedented mass measurement accuracy (MMA); MMA is one of the most significant attributes of mass spectrometric measurements as it affords extraordinary molecular specificity. However, due to space-charge effects, the achievable MMA significantly depends on the total number of ions trapped in the ICR cell for a particular measurement. Even through the use of automatic gain control (AGC), the total ion population is not constant between spectra. Multiple linear regression calibration in conjunction with AGC is utilized in these experiments to formally account for the differences in total ion population in the ICR cell between the external calibration spectra and experimental spectra. This ability allows for the extension of dynamic range of the instrument while allowing mean MMA values to remain less than 1 ppm. In addition, multiple linear regression calibration is used to account for both differences in total ion population in the ICR cell as well as relative ion abundance of a given species, which also affords mean MMA values at the parts-per-billion level. PMID:17539605

Enrichment of denitrifying methane-oxidizing microorganisms using up-flow continuous reactors and batch cultures.

PubMed

Hatamoto, Masashi; Kimura, Masafumi; Sato, Takafumi; Koizumi, Masato; Takahashi, Masanobu; Kawakami, Shuji; Araki, Nobuo; Yamaguchi, Takashi

2014-01-01

Denitrifying anaerobic methane oxidizing (DAMO) microorganisms were enriched from paddy field soils using continuous-flow and batch cultures fed with nitrate or nitrite as a sole electron acceptor. After several months of cultivation, the continuous-flow cultures using nitrite showed remarkable simultaneous methane oxidation and nitrite reduction and DAMO bacteria belonging to phylum NC10 were enriched. A maximum volumetric nitrite consumption rate of 70.4±3.4 mg-N·L(-1)·day(-1) was achieved with very short hydraulic retention time of 2.1 hour. In the culture, about 68% of total microbial cells were bacteria and no archaeal cells were detected by fluorescence in situ hybridization. In the nitrate-fed continuous-flow cultures, 58% of total microbial cells were bacteria while archaeal cells accounted for 7% of total cell numbers. Phylogenetic analysis of pmoA gene sequence showed that enriched DAMO bacteria in the continuous-flow cultivation had over 98% sequence similarity to DAMO bacteria in the inoculum. In contrast, for batch culture, the enriched pmoA gene sequences had 89-91% sequence similarity to DAMO bacteria in the inoculum. These results indicate that electron acceptor and cultivation method strongly affect the microbial community structures of DAMO consortia.

Changes in human bone marrow fat content associated with changes in hematopoietic stem cell numbers and cytokine levels with aging.

PubMed

Tuljapurkar, Sonal R; McGuire, Timothy R; Brusnahan, Susan K; Jackson, John D; Garvin, Kevin L; Kessinger, Margaret A; Lane, Judy T; O' Kane, Barbara J; Sharp, John G

2011-11-01

Hematological deficiencies increase with aging, including anemias, reduced responses to hematopoietic stress and myelodysplasias. This investigation tested the hypothesis that increased bone marrow (BM) fat content in humans with age was associated with decreased numbers of side population (SP) hematopoietic stem cells, and this decrease correlated with changes in cytokine levels. BM was obtained from the femoral head and trochanteric region of the femur removed at surgery for total hip replacement (N = 100 subjects). In addition, BM from cadavers (N = 36), with no evidence of hip disease, was evaluated for fat content. Whole trabecular marrow samples were ground in a sterile mortar and pestle, and cellularity and lipid content determined. Marrow cells were stained with Hoechst dye and SP profiles were acquired. Plasma levels of insulin-like growth factor (IGF)-1, stromal-derived factor (SDF)-1 and interleukin (IL)-6 were measured using ELISA. Fat content in the BM of human subjects and cadavers increased with age. The numbers of SP stem cells in BM as well as plasma IGF-1 and SDF-1 levels decreased in correlation with increased BM fat. IL-6 had no relationship to changes in marrow fat. These data suggest that increased BM fat may be associated with a decreased number of SP stem cells and IGF-1 and SDF-1 levels with aging. These data further raise a more general question as to the role of adipose cells in the regulation of tissue stem cells. © 2011 The Authors. Journal of Anatomy © 2011 Anatomical Society of Great Britain and Ireland.

Genetic profiling of putative breast cancer stem cells from malignant pleural effusions.

PubMed

Tiran, Verena; Stanzer, Stefanie; Heitzer, Ellen; Meilinger, Michael; Rossmann, Christopher; Lax, Sigurd; Tsybrovskyy, Oleksiy; Dandachi, Nadia; Balic, Marija

2017-01-01

A common symptom during late stage breast cancer disease is pleural effusion, which is related to poor prognosis. Malignant cells can be detected in pleural effusions indicating metastatic spread from the primary tumor site. Pleural effusions have been shown to be a useful source for studying metastasis and for isolating cells with putative cancer stem cell (CSC) properties. For the present study, pleural effusion aspirates from 17 metastatic breast cancer patients were processed to propagate CSCs in vitro. Patient-derived aspirates were cultured under sphere forming conditions and isolated primary cultures were further sorted for cancer stem cell subpopulations ALDH1+ and CD44+CD24-/low. Additionally, sphere forming efficiency of CSC and non-CSC subpopulations was determined. In order to genetically characterize the different tumor subpopulations, DNA was isolated from pleural effusions before and after cell sorting, and compared with corresponding DNA copy number profiles from primary tumors or bone metastasis using low-coverage whole genome sequencing (SCNA-seq). In general, unsorted cells had a higher potential to form spheres when compared to CSC subpopulations. In most cases, cell sorting did not yield sufficient cells for copy number analysis. A total of five from nine analyzed unsorted pleura samples (55%) showed aberrant copy number profiles similar to the respective primary tumor. However, most sorted subpopulations showed a balanced profile indicating an insufficient amount of tumor cells and low sensitivity of the sequencing method. Finally, we were able to establish a long term cell culture from one pleural effusion sample, which was characterized in detail. In conclusion, we confirm that pleural effusions are a suitable source for enrichment of putative CSC. However, sequencing based molecular characterization is impeded due to insufficient sensitivity along with a high number of normal contaminating cells, which are masking genetic alterations of rare cancer (stem) cells.

Effect of colorectal cancer on the number of normal stem cells circulating in peripheral blood.

PubMed

Marlicz, Wojciech; Sielatycka, Katarzyna; Serwin, Karol; Kubis, Ewa; Tkacz, Marta; Głuszko, Rafał; Białek, Andrzej; Starzyńska, Teresa; Ratajczak, Mariusz Z

2016-12-01

Bone marrow (BM) residing stem cells are mobilized from their BM niches into peripheral blood (PB) in several pathological situations including tissue organ injury and systemic inflammation. We recently reported that the number of BM-derived stem cells (SCs) increases in patients with pancreatic and stomach cancer. Accordingly, we observed higher numbers of circulating very small embryonic/epiblast‑like stem cells (VSELs) and mesenchymal stem cells (MSCs) that were associated with the activation of pro-mobilizing complement cascade and an elevated level of sphingosine-1 phosphate (S1P) in PB plasma. We wondered if a similar correlation occurs in patients with colorectal cancer (CRC). A total of 46 patients were enrolled in this study: 17 with CRC, 18 with benign colonic adenomas (BCA) and 11 healthy individuals. By employing fluorescence-activated cell sorting (FACS) we evaluated the number of BM-derived SCs circulating in PB: i) CD34+/Lin-/CD45- and CD133-/Lin-/CD45- VSELs; ii) CD45-/CD105+/CD90+/CD29+ MSCs; iii) CD45-/CD34+/CD133+/KDR+ endothelial progenitor cells (EPCs); and iv) CD133+/Lin-/CD45+ or CD34+/Lin-/CD45+ cells enriched for hematopoietic stem/progenitor cells (HSPCs). In parallel, we measured in the PB parameters regulating the egress of SCs from BM into PB. In contrast to pancreatic and gastric cancer patients, CRC subjects presented neither an increase in the number of circulating SCs nor the activation of pro-mobilizing factors such as complement, coagulation and fibrinolytic cascade, circulating stromal derived factor 1 (SDF‑1), vascular endothelial growth factor (VEGF) and intestinal permeability marker (zonulin). In conclusion, mobilization of SCs in cancer patients depends on the type of malignancy and its ability to activate pro-mobilization cascades.

Nickel hydrogen common pressure vessel battery development

NASA Technical Reports Server (NTRS)

Jones, Kenneth R.; Zagrodnik, Jeffrey P.

1992-01-01

Our present design for a common pressure vessel (CPV) battery, a nickel hydrogen battery system to combine all of the cells into a common pressure vessel, uses an open disk which allows the cell to be set into a shallow cavity; subsequent cells are stacked on each other with the total number based on the battery voltage required. This approach not only eliminates the assembly error threat, but also more readily assures equal contact pressure to the heat fin between each cell, which further assures balanced heat transfer. These heat fin dishes with their appropriate cell stacks are held together with tie bars which in turn are connected to the pressure vessel weld rings at each end of the tube.

Evaluation of plasma chemistry and haematological studies on chickens infected with Eimeria tenella and E acervulina.

PubMed

Fukata, T; Komba, Y; Sasai, K; Baba, E; Arakawa, A

1997-07-12

Plasma chemistry and haematological studies were conducted on chickens with coccidiosis. Male White Leghorn chickens, of two weeks old, were inoculated with 5 x 10(4) Eimeria tenella sporulated oocysts or with 1 x 10(6) E acervulina sporulated oocysts. Blood samples were taken four, seven and 11 days after inoculation. A wet chemistry system was applied to measure the plasma activities of aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, gamma glutamyltransferase, creatine kinase, amylase and lactate dehydrogenase and the concentrations of creatine, total bilirubin, urate, total cholesterol, total protein, albumin, glucose and triglycerides. A dry chemistry system was applied to measure sodium, potassium, chloride and calcium. The number of red blood cells and packed cell volume were determined by a micro cell counter and blood pH was measured with a blood gas analyser. The erythrocyte count, packed cell volume, sodium and chloride levels in the chickens infected with E tenella were significantly (P < 0.05) lower than those of the uninfected controls. The significant decrease in blood pH of the chickens infected with E acervulina suggests malabsorption associated with duodenal lesions induced by the infection.

Influence of IL-3 functional fragment on cord blood stem cell ex vivo expansion and differentiation.

PubMed

Ren, Zhihua; Zhang, Yu; Zhang, Yanxi; Jiang, Wenhong; Dai, Wei; Ding, Xinxin; Jiang, Yongping

2016-01-01

Recombinant human interleukin-3 (rhIL-3) is a multiple hematopoietic growth factor, which enhances stem cell expansion and hematopoiesis regeneration in vitro and in vivo, when administrated in combination with other cytokines. However, the structure-function study of rhIL-3 remains rarely studied, so far. The purpose of this study was to recognize the short peptide with similar function as rhIL-3, and assess the hematopoietic efficacy in umbilical cord blood (UCB) stem cell culture as well. Two novel monoclonal antibodies (mAb) (C1 and E1) were generated against rhIL-3 using hybridoma technique. Eleven short peptides were depicted and synthesized to overlap covering the full length sequence of rhIL-3. ELISA was employed to distinguish the antibody-binding peptide from the negative peptides. In addition, the multi-potential hematopoiesis capabilities of the positive peptides were evaluated by adding 25 ng/mL of each peptide to the culture medium of hematopoietic stem cells (HSCs) derived from UCB. Total nucleated cell number and the CD34(+) cell number from each individual treatment group were calculated on day 7. Correlated antibodies at 0.5 or 2 molar fold to each peptide were also tested in the stem cell expansion experiment, to further confirm the bioactivity of the peptides. Two peptides were recognized by the novel generated antibodies, using ELISA. Peptide 3 and 8 exhibited comparable hematopoiesis potentials, with 25.01±0.14 fold, and 19.89±0.12 fold increase of total nucleated cell number on day 7, respectively, compared with the basal medium control (4.93±0.55 fold). These biological effects were neutralized by adding the corresponding mAb at a dose dependent manner. Our results identified two specific regions of rhIL-3 responsible for HSC proliferation and differentiation, which were located from 28 to 49 amino acids (P3), and 107 to 127 amino acids (P8), respectively. The short peptide 3 and 8 might act synergistically, which could serve as an economic substitute to rhIL-3 in research laboratory.

[Effect of electroacupuncture on differentiation and proliferation of hippocampal nerve stem cells in splenic asthenia pedo-rats].

PubMed

Zhuo, Yuan-yuan; Yang, Zhuo-xin; Wu, Jia-man

2011-10-01

To observe the effect of electroacupuncture (EA) on the differentiation and proliferation of nerve stem cells in the hippocampal dentate gyrus (DG) in splenic asthenia pedo-rats so as to study its central mechanism. A total of 72 SD male rats were randomly assigned to normal control group (n=24), model group (n=24) and EA group (n=24) which were further divided into 7 d, 14 d, 28 d and 49 d time-points (n=6). Splenic asthenia model was established by intraperitoneal injection of reserpine and gavage of Dahuang (Radix et Rhizoma Rhei) fluid. EA was applied to bilateral "Zusanli" (ST 36) and "Sanyinjiao" (SP 6) for 20 min, once daily for 7, 14, 28 and 49 days respectively. Brdu, Nestin, glial fibrillary acidic protein (GFAP), and neuron-specific enolase (NSE) expression in the DG of hippocampus were detected by immunohistochemistry double staining. Compared with the normal control group, the numbers of Brdu, Brdu/GFAP, Brdu/NSE Immunoreactive (IR) positive cells in the DG of hippocampus on day 7 and 14, and that of Brdu/Nestin IR-positive cells on day 7 were decreased considerably in the model group (P < 0.05). Compared to the model group, the numbers of hippocampal Brdu IR-positive cells at the 4 time-points, Brdu/Nestin and Brdu/GFAP on day 7, 14 and 49, and Brdu/NSE on day 7, 14 and 28 were increased significantly in the EA group (P < 0.05). No significant differences were found between the model and control groups in the numbers of hippocampal Brdu and Brdu/NSE IR-positive cells on day 28 and 49, in the number of Brdu/Nestin IR-positive cells on day 14 and 49, in the number of Brdu/GFAP IR-positive cells on day 28; and between the EA and model groups in the numbers of Brdu/Nestin and Brdu/GFAP IR-positive cells on day 28, and in the number of Brdu/NSE IR-positive cells on day 49 (P > 0.05). EA of ST 36 and SP 6 can effectively suppress splenic asthenia syndrome-induced decrease of the numbers of Brdu, Brdu/GFAP, Brdu/Nestin and Brdu/NSE IR-positive cells in the DG of hippocampus at the early stage in the splenic asthenia rats, which may contribute to its effect in improving splenic asthenia symptoms in clinic by promoting the proliferation and differentiation of some nerve stem cells in the hippocampus.

Sparse grid techniques for particle-in-cell schemes

NASA Astrophysics Data System (ADS)

Ricketson, L. F.; Cerfon, A. J.

2017-02-01

We propose the use of sparse grids to accelerate particle-in-cell (PIC) schemes. By using the so-called ‘combination technique’ from the sparse grids literature, we are able to dramatically increase the size of the spatial cells in multi-dimensional PIC schemes while paying only a slight penalty in grid-based error. The resulting increase in cell size allows us to reduce the statistical noise in the simulation without increasing total particle number. We present initial proof-of-principle results from test cases in two and three dimensions that demonstrate the new scheme’s efficiency, both in terms of computation time and memory usage.

Stretching and movement of fibroblasts and osteoblasts cultured in microchannel and micropit arrays

NASA Astrophysics Data System (ADS)

Kikuchi, Hiroko E.; Kikuchi, Yuji; Kuboki, Yoshinori

1999-06-01

Tissue cells bind to extracellular matrix (ECM), and this attachment to ECM plays an essential role in their growth, function, and even survival. Furthermore, the geometry of ECM is known to play an additional role in regulation for these cells to proliferate and differentiate so that tissues with normal morphologies can be formed or maintained. We attempted to culture fibroblast, osteoblast, and bone marrow derived cells in previously described microchannel arrays and newly created micropit arrays, both coated with ECM protein collagen, to examine usefulness of microfabricated structures for elucidating 'what is geometry?' for cells. Cells were inseminated in the well in front of a microchannel array and their movement and stretching behavior against the microchannel array including the entrance and exit terraces were observed using a microscope-TV camera-time lapse video recorder system for 24 hours. Cells entered into the entrance terrace and showed active motions including extending pseudopodia into the channels and whole cell passage through the channels, and fully stretched in the entrance terrace in 8 hours or so. Those cells, however, voluntarily detached themselves from the area in another 8 hours or so probably because of worsening condition of nutrient supply there. Micropit arrays used in the present study consist of a regular arrangement of circular or square pits of diameter or side length of 25, 50, 100, 200, 400, and 600 micrometer and depth of 10 micrometer with one size per array or chip. The total area of the pits was designed to be equal to the rest surface area. After four days of incubation of bone marrow derived cells, the total number of cells and the number of cells in the pits were counted. The former and the ratio of the latter to the former appeared to become maximal when the pits of diameter or side length of 100 and 50 micrometer were used, respectively.

Novel platform for minimizing cell loss on separation process: Droplet-based magnetically activated cell separator

NASA Astrophysics Data System (ADS)

Kim, Youngho; Hong, Su; Lee, Sang Ho; Lee, Kangsun; Yun, Seok; Kang, Yuri; Paek, Kyeong-Kap; Ju, Byeong-Kwon; Kim, Byungkyu

2007-07-01

To reduce the problem of cell loss due to adhesion, one of the basic phenomena in microchannel, we proposed the droplet-based magnetically activated cell separator (DMACS). Based on the platform of the DMACS—which consists of permanent magnets, a coverslip with a circle-shaped boundary, and an injection tube—we could collect magnetically (CD45)-labeled (positive) cells with high purity and minimize cell loss due to adhesion. To compare separation efficiency between the MACS and the DMACS, the total number of cells before and after separation with both the separators was counted by flow cytometry. We could find that the number (3241/59940) of cells lost in the DMACS is much less than that (22360/59940) in the MACS while the efficiency of cell separation in the DMACS (96.07%) is almost the same as that in the MACS (96.72%). Practically, with fluorescent images, it was visually confirmed that the statistical data are reliable. From the viability test by using Hoechst 33 342, it was also demonstrated that there was no cell damage on a gas-liquid interface. Conclusively, DMACS will be a powerful tool to separate rare cells and applicable as a separator, key component of lab-on-a-chip.

Development of an Aquatic Bioassay using the Medaka (Oryzias latipes) to Assess Human Health Risk: Tumor Immunodiagnosis.

DTIC Science & Technology

1995-01-10

C-8029) (5) Type Exposure group and total number of each tumor type Adenoma II (1), III (1) Cholangioma II (1) Hepatocellular carcinoma I (1), II (2...antitrypsin Common marker for hepatocellular carcinoma Factor VIII/UEA-I Markers for endothelial cell differentiation S-100 protein Marker for nerve sheath...cells *Different cytokeratins can be used to detect different types of epithelium, i.e. to differentiate hepatocellular carcinoma from cholangiocellular

Composition of Fatty Acids and Carbohydrates in Leptospira1

PubMed Central

Kondo, Eiko; Ueta, Nobuo

1972-01-01

The fatty acid and monosaccharide composition of four pathogenic and two saprophytic strains of Leptospira was analyzed by gas chromatography (GC) and GC-mass spectrometry. Among the fatty acids, palmitic acid was most abundant and constituted 30 to 50% of the total fatty acids. Even-numbered unsaturated acids including octadecenoic, hexadecenoic, octadecadienoic, and tetradecadienoic acids comprised 40 to 60% of the total fatty acids. Tetradecanoic acid was about 5% in saprophytic strains, but 1% or less in pathogenic strains. The amount of chloroform-methanol extract of L. biflexa strain Ancona was 14 to 20% of the dry weight of the cell. Tetradecadienoic acid was found in the chloroform-methanol insoluble fraction, suggesting the presence of the acid in a bound form. GC analysis of monosaccharides revealed the existence of arabinose, xylose, rhamnose, mannose, galactose, glucose, glucosamine, and muramic acid in the cells. Among the neutral sugars, glucose was a minor component and was especially low in pathogenic strains. Total pentose content was about two to three times greater than total hexose. PMID:5022167

Maternal hCG concentrations in early IVF pregnancies: associations with number of cells in the Day 2 embryo and oocytes retrieved.

PubMed

Tanbo, T G; Eskild, A

2015-12-01

Do number of cells in the transferred cleavage stage embryo and number of oocytes retrieved for IVF influence maternal hCG concentrations in early pregnancies? Compared with transfer of a 2-cell embryo, transfer of a 4-cell embryo results in higher hCG concentrations on Day 12 after transfer, and more than 20 oocytes retrieved were associated with low hCG concentrations. Maternal hCG concentration in very early pregnancy varies considerably among women, but is likely to be an indicator of time since implantation of the embryo into the endometrium, in addition to number and function of trophoblast cells. We followed 1047 pregnancies after IVF/ICSI from oocyte retrieval until Day 12 after embryo transfer. Women were recruited in Norway during the years 2005-2013. Successful pregnancies after transfer of one single embryo that had been cultured for 2 days were included. Maternal hCG was quantified on Day 12 after embryo transfer by chemiluminescence immunoassay, which measures intact hCG and the free β-hCG chain. Information on a successful pregnancy, defined as birth after >16 weeks, was obtained by linkage to the Medical Birth Registry of Norway. Transfer of a 4-cell embryo resulted in higher maternal hCG concentrations compared with transfer of a 2-cell embryo (134.8 versus 87.8 IU/l, P < 0.05). A high number of oocytes retrieved (>20) was associated with low hCG concentrations (P < 0.05). The factors studied explain a limited part of the total variation of hCG concentrations in early pregnancy. Although embryo transfer was performed at the same time after fertilization, we do not know the exact time of implantation. A further limitation to our study is that the number of pregnancies after transfer of a 2-cell embryo was small (27 cases). Number of cells in the transferred embryo and number of oocytes retrieved may influence the conditions and timing for embryo implantation in different ways and thereby influence maternal hCG concentrations. Such knowledge may be important for interpretation of hCG concentrations in early pregnancy. © The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Supplementation of Diabetic Rats with Leucine, Zinc, and Chromium: Effects on Function and Histological Structure of Testes.

PubMed

Kolahian, Saeed; Sadri, Hassan; Larijani, Amir; Hamidian, Gholamreza; Davasaz, Afshin

2015-12-01

The objective was to study whether leucine, zinc, and chromium supplementations influence function and histological structure of testes in a rat model of type 2 diabetes. Seventy seven adult male rats were categorized into 11 groups of 7 animals each: (1) nondiabetic (negative control); (2) non-treated (positive control); (3) treated with insulin; (4) treated with glibenclamide; (5) treated with leucine; (6) treated with zinc; (7) treated with chromium; (8) treated with leucine + zinc; (9) treated with leucine + chromium; (10) treated with zinc + chromium; (11) treated with leucine + zinc + chromium. In the non-treated group, hyperglycemia severely damaged testes morphology as well as the spermatogenic process. Diabetes induction decreased testicular length, height, width, volume, total number of epididymal sperm, and number of live sperm. Seminiferous tubules of diabetic rats showed a decrease in diameter of tubules and height of epithelium. Diabetes induction decreased the number of cells (spermatogonia, spermatocyte, spermatid, and Sertoli) in cross sections of seminiferous tubules. Administration of nutritional supplements to the diabetic rats improved testes morphology and reversed, although not completely, impairment of spermatogenesis. Treatment with nutritional supplements increased testicular length, height, width, and volume. All treatments increased the number of live sperm and the total number of epididymal sperm. Furthermore, nutritional supplements increased diameter of tubules, height of epithelium, and the number of cells in seminiferous tubules. These alleviating effects were more pronounced in animals treated with the leucine-zinc-chromium combination. The present results demonstrate beneficial effects of zinc, leucine, and chromium supplements to improve testes morphology and to restore spermatogenesis in type 2 diabetic rats.

Inorganic mercury accumulation in brain following waterborne exposure elicits a deficit on the number of brain cells and impairs swimming behavior in fish (white seabream-Diplodus sargus).

PubMed

Pereira, Patrícia; Puga, Sónia; Cardoso, Vera; Pinto-Ribeiro, Filipa; Raimundo, Joana; Barata, Marisa; Pousão-Ferreira, Pedro; Pacheco, Mário; Almeida, Armando

2016-01-01

The current study contributes to fill the knowledge gap on the neurotoxicity of inorganic mercury (iHg) in fish through the implementation of a combined evaluation of brain morphometric alterations (volume and total number of neurons plus glial cells in specific regions of the brain) and swimming behavior (endpoints related with the motor activity and mood/anxiety-like status). White seabream (Diplodus sargus) was exposed to realistic levels of iHg in water (2μgL(-1)) during 7 (E7) and 14 days (E14). After that, fish were allowed to recover for 28 days (PE28) in order to evaluate brain regeneration and reversibility of behavioral syndromes. A significant reduction in the number of cells in hypothalamus, optic tectum and cerebellum was found at E7, accompanied by relevant changes on swimming behavior. Moreover, the decrease in the number of neurons and glia in the molecular layer of the cerebellum was followed by a contraction of its volume. This is the first time that a deficit on the number of cells is reported in fish brain after iHg exposure. Interestingly, a recovery of hypothalamus and cerebellum occurred at E14, as evidenced by the identical number of cells found in exposed and control fish, and volume of cerebellum, which might be associated with an adaptive phenomenon. After 28 days post-exposure, the optic tectum continued to show a decrease in the number of cells, pointing out a higher vulnerability of this region. These morphometric alterations coincided with numerous changes on swimming behavior, related both with fish motor function and mood/anxiety-like status. Overall, current data pointed out the iHg potential to induce brain morphometric alterations, emphasizing a long-lasting neurobehavioral hazard. Copyright © 2015 Elsevier B.V. All rights reserved.

Cancer Prevention and Control Research Manpower Development

DTIC Science & Technology

1996-10-01

Acute Epstein-Barr Viral Mononucleosis . Annals of Allergy, Vol. 54, Number 4, 1985. 10. Shacks, S.J. and Johnson, C.S.: Serum Concentration of Total...production in sickle cell disease patients with acute pneumonia, Pediatric Infectious Disease Journal, Vol. 15, Issue 4: 340-344, 1996. PHS 398 (Rev. 5

The Fifth Cell: Correlation Bias in U.S. Census Adjustment.

ERIC Educational Resources Information Center

Wachter, Kenneth W.; Freedman, David A.

2000-01-01

Presents a method for estimating the total national number of doubly missing people (missing from Census counts and adjusted counts as well) and their distribution by race and sex. Application to the 1990 U.S. Census yields an estimate of three million doubly-missing people. (SLD)

A Bereavement Support Group Intervention Is Longitudinally Associated with Salutary Effects on the CD4 Cell Count and Number of Physician Visits

PubMed Central

Goodkin, Karl; Feaster, Daniel J.; Asthana, Deshratn; Blaney, Nancy T.; Kumar, Mahendra; Baldewicz, Teri; Tuttle, Raymond S.; Maher, Kevin J.; Baum, Marianna K.; Shapshak, Paul; Fletcher, Mary Ann

1998-01-01

A randomized, controlled, clinical trial was conducted to examine the impact of a semistructured, 10-week, once weekly, 90-min/session bereavement support group intervention on immunological, neuroendocrine, and clinical health status in human immunodeficiency virus type 1-seropositive (HIV-1+) and HIV-1-seronegative (HIV-1−) homosexual men, compared to a standard of care control condition. A total of 119 homosexual men (74 HIV-1+ and 45 HIV-1−) were assessed at baseline, 10 weeks, and 6 months follow-up. At the 6-month follow-up assessment, the intervention groups exhibited significant beneficial effects compared to controls on changes in CD4 cell, total T-lymphocyte, and total lymphocyte counts, when baseline levels, antiretroviral medication use, CDC stage of disease, and other potentially confounding factors were accounted for. There was no statistically significant effect on the CD4/CD8 ratio or on the CD8 cell count. The effect on CD4 cell count was associated with group attendance and with changes in plasma cortisol level. Plasma cortisol levels decreased significantly among intervention subjects, compared to controls. A significantly reduced number of health care visits over the 6-month follow-up period among the intervention subjects supported the clinical relevance of the immunological changes observed for both HIV-1+ and HIV-1− individuals. These results indicate that behavioral interventions may have salutary immunological and clinical health effects following bereavement among HIV-1-infected individuals. The effect in HIV-1− individuals suggests that this bereavement support group intervention might have similar salutary effects in the general population. Potential effects of such interventions on clinical HIV disease progression are of interest and should be studied. PMID:9605995

Hepatic Oval Cells Have the Side Population Phenotype Defined by Expression of ATP-Binding Cassette Transporter ABCG2/BCRP1

PubMed Central

Shimano, Koichi; Satake, Makoto; Okaya, Atsuhito; Kitanaka, Junichi; Kitanaka, Nobue; Takemura, Motohiko; Sakagami, Masafumi; Terada, Nobuyuki; Tsujimura, Tohru

2003-01-01

Organ-specific stem cells can be identified by the side population (SP) phenotype, which is defined by the property to effectively exclude the Hoechst 33342 dye. The ATP-binding cassette transporter ABCG2/BCRP1 mediates the SP phenotype. Because hepatic oval cells possess several characteristics of stem cells, we examined whether they have the SP phenotype using the 2-acetylaminofluorene/partial hepatectomy (PH) model. Fluorescence-activated cell sorting analysis showed that a population of non-parenchymal cells containing oval cells, prepared on day 7 after PH, carried a significant number of SP cells, whereas that of non-parenchymal cells without oval cells, prepared on day 0 after PH, did not. Northern blot analysis using total liver RNA obtained on various days after PH showed that the expression of ABCG2/BCRP1 mRNA increased after PH, reaching the highest level on day 7, and then gradually decreased. This pattern of changes in the ABCG2/BCRP1 mRNA level was well correlated to that in the number of oval cells. Furthermore, in situ hybridization revealed that oval cells were the sites of expression of ABCG2/BCRP1 mRNA. These results indicate that oval cells have the SP phenotype defined by expression of ABCG2/BCRP1, suggesting that oval cells may represent stem cells in the liver. PMID:12819005

Factors affecting reconstitution of the T cell compartment in allogeneic haematopoietic cell transplant recipients.

PubMed

Fallen, P R; McGreavey, L; Madrigal, J A; Potter, M; Ethell, M; Prentice, H G; Guimarães, A; Travers, P J

2003-11-01

The factors affecting T cell reconstitution post haematopoietic cell transplantation (HCT) are not well characterised. We carried out a longitudinal analysis of T cell reconstitution in 32 HCT recipients during the first 12 months post transplant. We analysed reconstitution of naïve, memory and effector T cells, their diversity and monitored thymic output using TCR rearrangement excision circles (TRECs). Thymic-independent pathways were responsible for the rapid reconstitution of memory and effector T cells less than 6 months post HCT. Thymic-dependent pathways were activated between 6 and 12 months in the majority of patients with naïve T cell numbers increasing in parallel with TREC levels. Increasing patient age, chronic GVHD and T cell depletion (with or without pretransplant Campath-1H) predicted low TREC levels and slow naïve T cell recovery. Furthermore, increasing patient age also predicted high memory and effector T cell numbers. The effects of post HCT immunosuppression, total body irradiation, donor leucocyte infusions, T cell dose and post HCT infections on T cell recovery were also analysed. However, no effects of these single variables across a variety of different age, GVHD and T cell depletion groups were apparent. This study suggests that future analysis of the factors affecting T cell reconstitution and studies aimed at reactivating the thymus through therapeutic intervention should be analysed in age-, GVHD- and TCD-matched patient groups.

Stage and season effects on cell cycle and apoptotic activities of germ cells and Sertoli cells during spermatogenesis in the spiny dogfish (Squalus acanthias).

PubMed

McClusky, L M

2005-01-01

To understand the processes involved in the spatial and temporal maturation of testicular cells in Squalus acanthias, we used standard morphometry, proliferating-cell nuclear antigen (PCNA) and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL) immunohistochemistry. Except for immature spermatocysts (germinal zone, GZ; early-stage pre-meiotic, E-PrM), the number of cysts in all subsequent stages and the total number of cysts in the spermatogenic progression varied seasonally. The spermatogenic cycle spans about 2 years and is interrupted by germcell clone deletion via apoptosis at the mitosis-meiosis transition in April/May, manifesting as a zone of degeneration (ZD). Rate of displacement of the ZD across the testis diameter indicates that late-stage premeiotic (L-PrM) generations 12-13 require 9-10 months to reach the mature-spermatid stage. Also, the number of cysts completing spermatogenesis is approximately 4-5-fold less than the number that entered spermatogenesis proper 2 years earlier. Pronounced gonocytogenesis in the germinal ridge was coincident with ZD formation in April/May, but it was absent in the fall when mature spermatogonial and meiotic activities had resumed. Whereas strong Sertoli cell PCNA immunoreactivity dominated the GZ cyst cell-cycle activities throughout the year, except during the spring/summer months, the spermatogonial- and Sertoli-cell PCNA indices in E-PrM cysts were inversely related. PCNA immunoreactivity in spermatocytes was seasonal and dependent on the stage of meiosis. TUNEL labelling was limited to spermatogonia and increased stage-dependently in the PrM region (L-PrM = mid-stage PrM >E-PrM >GZ), correlating with ZD formation, in a season-dependent manner. Results imply that effects of normal regulatory factors in Squalus are stage- and process-specific.

Left-right asymmetry in neck lymph nodes distribution in patients with bilateral laryngeal cancer.

PubMed

Yoruk, Ozgur; Yuksel, Ramazan; Yuksel, Yasemin; Dane, Senol

2014-04-01

We aimed to examine left-right asymmetry in involved and total neck lymph nodes distribution in patients with bilateral laryngeal cancer in the present study. Forty-six patients with bilateral laryngeal cancer was included the study. The oncologic database of our otorhinolaryngology department was used. The right and left lymph node with and without involvement by cancer cells counts were retrieved from pathological reports. The numbers of both involved and total neck lymph nodes were significantly higher on right side than on left side for all neck levels in laryngeal malignancies. The results of the present study suggest the existence of a left-right asymmetry in neck lymph node distribution and in the neck lymph node distribution involved by laryngeal cancer cells. The stronger cell-mediated immune activity in the left side of humans may be associated with the blocking of the metastatic invasion of cancer cells from laryngeal malignancies in the left body side.

Iron loading in HFE p.C282Y homozygotes found by population screening: relationships to HLA-type and T-lymphocyte subsets.

PubMed

Thorstensen, Ketil; Kvitland, Mona A; Irgens, Wenche Ø; Åsberg, Arne; Borch-Iohnsen, Berit; Moen, Torolf; Hveem, Kristian

2017-11-01

Iron loading in p.C282Y homozygous HFE hemochromatosis subjects is highly variable, and it is unclear what factors cause this variability. Finding such factors could aid in predicting which patients are at highest risk and require closest follow-up. The degree of iron loading has previously been associated with certain HLA-types and with abnormally low CD8 + cell counts in peripheral blood. In 183 Norwegian, p.C282Y homozygotes (104 men, 79 women) originally found through population screening we determined HLA type and measured total T-lymphocytes, CD4 + and CD8 + cells, and compared this with data on iron loading. In p.C282Y homozygous men, but not in homozygous women, we found that the presence of two HLA-A*03 alleles increased the iron load on average by approximately 2-fold compared to p.C282Y homozygous men carrying zero or one A*03 allele. On the other hand, the presence of two HLA-A*01 alleles, in male subjects, apparently reduced the iron loading. In p.C282Y homozygous individuals, the iron loading was increased if the CD8 + cell number was below the 25 percentile or if the CD4 + cell number was above the 75 percentile. This effect appeared to be additive to the effect of the number of HLA-A*03 alleles. Our data indicate that homozygosity for the HLA-A*03 allele significantly increases the risk of excessive iron loading in Norwegian p.C282Y homozygous male patients. In addition, low CD8 + cell number or high CD4 + cell number further increases the risk of excessive iron loading.

Expansion of 3D human induced pluripotent stem cell aggregates in bioreactors: Bioprocess intensification and scaling-up approaches.

PubMed

Abecasis, Bernardo; Aguiar, Tiago; Arnault, Émilie; Costa, Rita; Gomes-Alves, Patricia; Aspegren, Anders; Serra, Margarida; Alves, Paula M

2017-03-20

Human induced pluripotent stem cells (hiPSC) are attractive tools for drug screening and disease modeling and promising candidates for cell therapy applications. However, to achieve the high numbers of cells required for these purposes, scalable and clinical-grade technologies must be established. In this study, we use environmentally controlled stirred-tank bioreactors operating in perfusion as a powerful tool for bioprocess intensification of hiPSC production. We demonstrate the importance of controlling the dissolved oxygen concentration at low levels (4%) and perfusion at 1.3day -1 dilution rate to improve hiPSC growth as aggregates in a xeno-free medium. This strategy allowed for increased cell specific growth rate, maximum volumetric concentrations (4.7×10 6 cell/mL) and expansion factors (approximately 19 in total cells), resulting in a 2.6-fold overall improvement in cell yields. Extensive cell characterization, including whole proteomic analysis, was performed to confirm that cells' pluripotent phenotype was maintained during culture. A scalable protocol for continuous expansion of hiPSC aggregates in bioreactors was implemented using mechanical dissociation for aggregate disruption and cell passaging. A total expansion factor of 1100 in viable cells was obtained in 11days of culture, while cells maintained their proliferation capacity, pluripotent phenotype and potential as well as genomic stability after 3 sequential passages in bioreactors. Copyright © 2017 Elsevier B.V. All rights reserved.

Relating tumor score to hematology in green turtles with fibropapillomatosis in Hawaii

USGS Publications Warehouse

Work, Thierry M.; Balazs, George H.

1999-01-01

The relationship between hematologic status and severity of tumor affliction in green turtles (Chelonia mydas) with fibropapillomatosis (FP) was examined. During 1 wk periods in July 1997 and July 1998, we bled 108 free-ranging green turtles from Pala'au (Molokai, Hawaii, USA) where FP is endemic. Blood was analyzed for hematocrit, estimated total solids, total white blood cell (WBC) count and differential WBC count. Each turtle was assigned a subjective tumor score ranging from 0 (no visible external tumors) to 3 (heavily tumored) that indicated the severity of FP. There was a progressive increase in monocytes and a decrease in all other hematologic parameters except heterophils and total numbers of white blood cells as tumor score increased. These data indicate that tumor score can relate to physiologic status of green turtles afflicted with FP, and that tumor score is a useful field monitor of severity of FP in this species.

Enhanced alveolar monocytic phagocyte (macrophage) proliferation in tobacco and marijuana smokers

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

Barbers, R.G.; Evans, M.J.; Gong, H. Jr.

We tested the hypothesis that enhanced cell division accounted for the augmented numbers of monocytic phagocytes with characteristics attributed to alveolar macrophages (AM) found in the lungs of habitual tobacco (T) and marijuana (M) smokers. The monocytic phagocytes, that is, alveolar macrophages, were obtained by bronchoalveolar lavage (BAL) from 12 nonsmoking subjects; 10 subjects who smoked T only (TS); 13 subjects who smoked M only (MS); and 6 smokers of both T and M (MTS). The replication of these cells was determined by measuring the incorporation of ({sup 3}H)thymidine into the DNA of dividing cells and visually counting 2,000 cellsmore » on autoradiographically prepared cytocentrifuge cell preparations. This study demonstrated that the number of ({sup 3}H)thymidine-labeled monocytic phagocytes with characteristics of alveolar macrophages from either TS or MS have a higher proliferative index compared to cells (macrophages) from nonsmokers, p less than 0.05 by one-way ANOVA. The total number of BAL macrophages that are in mitosis in TS (17.90 +/- 4.50 labeled AM x 10(3)/ml) or MTS (10.50 +/- 4.20 labeled AM x 10(3)/ml) are 18- and 10-fold greater, respectively, than the number obtained from nonsmokers (1.01 +/- 0.18 labeled AM x 10(3)/ml). Interestingly, the number of ({sup 3}H)thymidine-labeled macrophages from MS (2.90 +/- 0.66 labeled AM x 10(3)/ml) are also greater than the number obtained from nonsmokers, although this is not statistically significant. The stimulus augmenting alveolar macrophage replication is as yet unknown but may likely be found in the T or M smoke.« less

Analysis and IbM simulation of the stages in bacterial lag phase: basis for an updated definition.

PubMed

Prats, Clara; Giró, Antoni; Ferrer, Jordi; López, Daniel; Vives-Rego, Josep

2008-05-07

The lag phase is the initial phase of a culture that precedes exponential growth and occurs when the conditions of the culture medium differ from the pre-inoculation conditions. It is usually defined by means of cell density because the number of individuals remains approximately constant or slowly increases, and it is quantified with the lag parameter lambda. The lag phase has been studied through mathematical modelling and by means of specific experiments. In recent years, Individual-based Modelling (IbM) has provided helpful insights into lag phase studies. In this paper, the definition of lag phase is thoroughly examined. Evolution of the total biomass and the total number of bacteria during lag phase is tackled separately. The lag phase lasts until the culture reaches a maximum growth rate both in biomass and cell density. Once in the exponential phase, both rates are constant over time and equal to each other. Both evolutions are split into an initial phase and a transition phase, according to their growth rates. A population-level mathematical model is presented to describe the transitional phase in cell density. INDividual DIScrete SIMulation (INDISIM) is used to check the outcomes of this analysis. Simulations allow the separate study of the evolution of cell density and total biomass in a batch culture, they provide a depiction of different observed cases in lag evolution at the individual-cell level, and are used to test the population-level model. The results show that the geometrical lag parameter lambda is not appropriate as a universal definition for the lag phase. Moreover, the lag phase cannot be characterized by a single parameter. For the studied cases, the lag phases of both the total biomass and the population are required to fully characterize the evolution of bacterial cultures. The results presented prove once more that the lag phase is a complex process that requires a more complete definition. This will be possible only after the phenomena governing the population dynamics at an individual level of description, and occurring during the lag and exponential growth phases, are well understood.

Adenylyl cyclase localization to the uropod of aggregating Dictyostelium cells requires RacC

PubMed Central

Wang, C.; Jung, D.; Cao, Z.; Chung, C. Y.

2015-01-01

The localization of adenylyl cyclase A (ACA) to uropod of cells is required for the stream formation during Dictyostelium development. RacC is a Dictyostelium orthologue of Cdc42. We identified a streaming defect of racC− cells as they are clearly less polarized and form smaller and fragmented streams. ACA-YFP is mainly associated with intracellular vesicular structures, but not with the plasma membrane in racC− cells. racC− cells have a slightly higher number of vesicles than Ax3 cells, suggesting that the defect of ACA trafficking is not simply due to the lack of vesicle formation. While the ACA-YFP vesicles traveled with an average velocity of 9.1 µm/min in Ax3 cells, a slow and diffusional movement without direction with an average velocity of 4 µm/min was maintained in racC− cells. Images acquired by using total internal reflection fluorescence (TIRF) microscopy and fluorescence recovery after photobleaching (FRAP) analysis revealed that a significantly decreased number of ACA-YFP vesicles appeared near the cell membrane, indicating a defect in ACA-YFP vesicle trafficking. These results suggest an important role of RacC in the rapid and directional movements of ACA vesicles on microtubules to the plasma membrane, especially to the back of polarized cell. PMID:26315268

In-depth Characterization of a TCR-specific Tracer for Sensitive Detection of Tumor-directed Transgenic T Cells by Immuno-PET.

PubMed

Yusufi, Nahid; Mall, Sabine; Bianchi, Henrique de Oliveira; Steiger, Katja; Reder, Sybille; Klar, Richard; Audehm, Stefan; Mustafa, Mona; Nekolla, Stephan; Peschel, Christian; Schwaiger, Markus; Krackhardt, Angela M; D'Alessandria, Calogero

2017-01-01

A number of different technologies have been developed to monitor in vivo the distribution of gene-modified T cells used in immunotherapy. Nevertheless, in-depth characterization of novel approaches with respect to sensitivity and clinical applicability are so far missing. We have previously described a novel method to track engineered human T cells in tumors using 89 Zr-Df-aTCRmu-F(ab') 2 targeting the murinized part of the TCR beta domain (TCRmu) of a transgenic TCR. Here, we performed an in-depth in vitro characterization of the tracer in terms of antigen affinity, immunoreactivity, influence on T-cell functionality and stability in vitro and in vivo . Of particular interest, we have developed diverse experimental settings to quantify TCR-transgenic T cells in vivo . Local application of 89 Zr-Df-aTCRmu-F(ab') 2 -labeled T cells in a spot-assay revealed signal detection down to approximately 1.8x10 4 cells. In a more clinically relevant model, NSG mice were intravenously injected with different numbers of transgenic T cells, followed by injection of the 89 Zr-Df-aTCRmu-F(ab') 2 tracer, PET/CT imaging and subsequent ex vivo T-cell quantification in the tumor. Using this setting, we defined a comparable detection limit of 1.0x10 4 T cells. PET signals correlated well to total numbers of transgenic T cells detected ex vivo independently of the engraftment rates observed in different individual experiments. Thus, these findings confirm the high sensitivity of our novel PET/CT T-cell tracking method and provide critical information about the quantity of transgenic T cells in the tumor environment suggesting our technology being highly suitable for further clinical translation.

Modulation of osteoblast attachment and growth in vitro by inertial forces

NASA Astrophysics Data System (ADS)

Kacena, Melissa Ann

1999-11-01

Spaceflight exploration and associated experiments show that human bones lose in density during inertial unloading, due principally to their demineralization. This research project examines the effect of gravity on osteoblast attachment and function in various inertial environments. Chicken calvarial osteoblasts were cultured under the following inertial conditions: spaceflight, simulated shuttle launch accelerations and vibrations, centrifugation, clino-rotation, and inversion. Cultures exposed to these conditions were compared with cultures grown in the laboratory as static 1G controls. Electron and light microscopy revealed the number of total osteoblasts attached to their substrate. Biochemical assays discerned changes in viable cell number, alkaline phosphatase levels, and mineralization. Immunohistochemical assays were used to investigate differences in cytoskeletal and extracellular matrix protein concentrations in the cultures, the percentage of proliferative cells, and cell viability. Compared to controls, spaceflight results indicated that the number of attached osteoblast cells was reduced. Launch simulation data indicated that the associated accelerations and vibrations may contribute to the reduction of attached osteoblasts in spaceflight cultures. Following centrifugation, the number of attached cells was unaltered; however, immunostaining of actin, fibronectin, and vinculin did show alterations in cultures exposed to hypergravity. Confluent cultures that were right side up, inverted, and clino-rotated contained a comparable number of attached cells and functioned similarly on the basis of measured alkaline phosphatase and bound calcium content. Sparse clino-rotated or inverted cultures showed an immediate response of diminished viable osteoblast numbers, but this effect disappeared with time and all remaining attached cells functioned similarly (APase and bound calcium). On the basis of these data osteoblast attachment and function in confluent cultures is minimally, if at all, affected by alterations in inertial environments. However, in sparse cultures about half as many cells are found attached initially. The remaining attached cells appear to multiply and function normally. These results suggest that the effects of spaceflight on bone are thus not likely to be caused by direct intrinsic effects of gravity on single osteoblasts that can be simulated in laboratory experiments in vitro experiments.

Expression of mRNA encoding leukaemia inhibitory factor (LIF) and its receptor (LIFRβ) in buffalo preimplantation embryos produced in vitro: markers of successful embryo implantation.

PubMed

Eswari, S; Sai Kumar, G; Sharma, G Taru

2013-05-01

Summary The objective of this study was to evaluate the effect of supplementation of recombinant leukaemia inhibitory factor (LIF) in culture media on blastocyst development, total cell number and blastocyst hatching rates and the reverse transcription-polymerase chain reaction analysis of preimplantation buffalo embryos to determine whether they contain the LIF-encoding mRNA and its beta receptor (LIFRβ) genes in different stages of preimplantation buffalo embryos. Cumulus-oocyte complexes retrieved from slaughterhouse buffalo ovaries were matured in vitro and fertilized using frozen buffalo semen. After 18 h of co-incubation with sperm, the presumptive zygotes were cultured in modified synthetic oviductal fluid without (control) or with rhLIF (100 ng/ml). There was no significant difference in the overall cleavage rate up to morula stage however the development of blastocysts, hatching rate and total cell numbers were significantly higher in the LIF-treated group than control. Transcripts for LIFRβ were detected from immature, in vitro-matured oocytes and in the embryos up to blastocyst stage, while transcripts for the LIF were detected from 8-16-cell stage up to blastocyst, which indicated that embryo-derived LIF can act in an autocrine manner on differentiation process and blastocyst formation. This study indicated that the addition of LIF to the embryo culture medium improved development of blastocysts, functional (hatching) and morphological (number of cells) quality of the blastocysts produced in vitro. The stage-specific expression pattern of LIF and LIFRβ mRNA transcripts in buffalo embryos indicated that LIF might play an important role in the preimplantation development and subsequent implantation of buffalo embryos.

Hospitalizations of children with sickle cell disease in the Brazilian Unified Health System in the state of Minas Gerais.

PubMed

Fernandes, Ana Paula P C; Avendanha, Fernanda A; Viana, Marcos B

To identify and characterize hospital admissions and readmissions in the Brazilian Unified Public Health System (Sistema Único de Saúde [SUS]) in children with sickle cell disease diagnosed by the Minas Gerais Newborn Screening Program between 1999 and 2012. Hospital Admission Authorizations with the D57 (International Classification of Diseases-10) code in the fields of primary or secondary diagnosis were retrieved from the SUS Databank (1999-2012). There were 2991 hospitalizations for 969 children. 73.2% of children had hemoglobin SS/Sβ 0 -thalassemia and 48% were girls. The mean age was 4.3±3.2 years, the mean number of hospitalizations, 3.1±3.3, and the hospital length of stay, 5±3.9 days. Hospital readmissions occurred for 16.7% of children; 10% of admissions were associated with readmission within 30 days after discharge; 33% of readmissions occurred within seven days post-discharge. There were 41 deaths, 95% of which were in-hospital. Secondary diagnoses were not recorded in 96% of admissions, making it impossible to know the reason for admission. In 62% of cases, hospitalizations occurred in the child's county of residence. The total number of hospitalizations of children under 14 with sickle cell disease relative to the total of pediatric hospitalizations increased from 0.12% in 1999 to 0.37% in 2012. A high demand for hospital care in children with sickle cell disease was evident. The number of hospitalizations increased from 1999 to 2012, suggesting that the disease has become more "visible." Knowledge of the characteristics of these admissions can help in the planning of care for these children in the SUS. Copyright © 2016 Sociedade Brasileira de Pediatria. Published by Elsevier Editora Ltda. All rights reserved.

Stromal and Hematopoietic Progenitors from C57/BI/6N Murine Bone Marrow After 30-Day "BION-M1" Spaceflight.

PubMed

Markina, Elena; Andreeva, Elena; Andrianova, Irina; Sotnezova, Elena; Buravkova, Ludmila

2018-05-02

Elucidation of the spaceflight (SF) effects on the adult stem and progenitor cells is an important goal in space biology and medicine. A unique opportunity for this was provided by project "BION-M1". The purpose of this study was to evaluate the effects of 30-day SF on biosatellite, 7-day recovery (SFR), and subsequent ground control (GC) experiment on the mononuclear cells (MNCs) from C57/BI/6N murine tibia bone marrow. Also, hematopoietic and stromal precursor functions were characterized ex vivo. There was no significant difference in the total MNC number between experimental groups. After SF, immunophenotyping revealed an increase of large-sized CD45 + MNCs corresponded to committed hematopoietic progenitors. The total hematopoietic colony-forming unit (CFU) number decreased after SF and did not restore after 7 day of recovery due to predominant reduction of bi- and multipotent CFUs and primitive burst-forming units in favor of unipotent CFUs. Functional activity of stromal precursors in vitro was only slightly altered. SF cells displayed the enhanced expression of alkaline phosphatase. The data of the GC experiment demonstrated the preservation of the functional activity of progenitor cells from mice bone marrow. The activation of erythropoiesis in expense of burst-forming units of erythrocytes elevation was detected. After 7 days of recovery, the number of colony-forming units of fibroblast (CFUs-f) was similar to the vivarium control, while the proliferative activity of bone marrow stromal precursors decreased. The present study demonstrated that certain hematopoietic progenitors are susceptible to SF factors, while the stromal precursors displayed a certain degree of resistance. These data indicate mild and reversible alterations of bone marrow progenitors after SF.

Auditory hair cell innervational patterns in lizards.

PubMed

Miller, M R; Beck, J

1988-05-22

The pattern of afferent and efferent innervation of two to four unidirectional (UHC) and two to nine bidirectional (BHC) hair cells of five different types of lizard auditory papillae was determined by reconstruction of serial TEM sections. The species studies were Crotaphytus wislizeni (iguanid), Podarcis (Lacerta) sicula and P. muralis (lacertids), Ameiva ameiva (teiid), Coleonyx variegatus (gekkonid), and Mabuya multifasciata (scincid). The main object was to determine in which species and in which hair cell types the nerve fibers were innervating only one (exclusive innervation), or two or more hair cells (nonexclusive innervation); how many nerve fibers were supplying each hair cell; how many synapses were made by the innervating fibers; and the total number of synapses on each hair cell. In the species studies, efferent innervation was limited to the UHC, and except for the iguanid, C. wislizeni, it was nonexclusive, each fiber supplying two or more hair cells. Afferent innervation varied both with the species and the hair cell types. In Crotaphytus, both the UHC and the BHC were exclusively innervated. In Podarcis and Ameiva, the UHC were innervated exclusively by some fibers but nonexclusively by others (mixed pattern). In Coleonyx, the UHC were exclusively innervated but the BHC were nonexclusively innervated. In Mabuya, both the UHC and BHC were nonexclusively innervated. The number of afferent nerve fibers and the number of afferent synapses were always larger in the UHC than in the BHC. In Ameiva, Podarcis, and Mabuya, groups of bidirectionally oriented hair cells occur in regions of cytologically distinct UHC, and in Ameiva, unidirectionally oriented hair cells occur in cytologically distinct BHC regions.

Hippo circuitry and the redox modulation of hippo components in cancer cell fate decisions.

PubMed

Ashraf, Asma; Pervaiz, Shazib

2015-12-01

Meticulous and precise control of organ size is undoubtedly one of the most pivotal processes in mammalian development and regeneration along with cell differentiation, morphogenesis and programmed cell death. These processes are strictly regulated by complex and highly coordinated mechanisms to maintain a steady growth state. There are a number of extrinsic and intrinsic factors that dictate the total number and/or size of cells by influencing growth, proliferation, differentiation and cell death. Multiple pathways, such as those involved in promoting organ size and others that restrict disproportionate tissue growth act simultaneously to maintain cellular and tissue homeostasis. Aberrations at any level in these organ size-regulating processes can lead to various pathological states with cancers being the most formidable one (Yin and Zhang, 2011). Extensive research in the realm of growth control has led to the identification of the Hippo-signaling pathway as a critical network in modulating tissue growth via its effect on multiple signaling pathways and through intricate crosstalk with proteins that regulate cell polarity, adhesion and cell-cell interactions (Zhao et al., 2011b). The Hippo pathway controls cell number and organ size by transducing signals from the plasma membrane to the nucleus to regulate the expression of genes involved in cell fate determination (Shi et al., 2015). In this review, we summarize the recent discoveries concerning Hippo pathway, its diversiform regulation in mammals as well as its implications in cancers, and highlight the possible role of oxidative stress in Hippo pathway regulation. Copyright © 2015 Elsevier Ltd. All rights reserved.

Mesomere-derived glutamate decarboxylase-expressing blastocoelar mesenchyme cells of sea urchin larvae

PubMed Central

Katow, Hideki; Katow, Tomoko; Abe, Kouki; Ooka, Shioh; Kiyomoto, Masato; Hamanaka, Gen

2014-01-01

Summary The ontogenetic origin of blastocoelar glutamate decarboxylase (GAD)-expressing cells (GADCs) in larvae of the sea urchin Hemicentrotus pulcherrimus was elucidated. Whole-mount in situ hybridisation (WISH) detected transcription of the gene that encodes GAD in H. pulcherrimus (Hp-gad) in unfertilised eggs and all blastomeres in morulae. However, at and after the swimming blastula stage, the transcript accumulation was particularly prominent in clumps of ectodermal cells throughout the embryonic surface. During the gastrula stage, the transcripts also accumulated in the endomesoderm and certain blastocoelar cells. Consistent with the increasing number of Hp-gad transcribing cells, immunoblot analysis indicated that the relative abundance of Hp-Gad increased considerably from the early gastrula stage until the prism stage. The expression pattern of GADCs determined by immunohistochemistry was identical to the pattern of Hp-gad transcript accumulation determined using WISH. In early gastrulae, GADCs formed blastocoelar cell aggregates around the blastopore with primary mesenchyme cells. The increase in the number of blastocoelar GADCs was inversely proportional to the number of ectodermal GADCs ranging from a few percent of total GADCs in early gastrulae to 80% in late prism larvae; this depended on ingression of ectodermal GADCs into the blastocoel. Some of the blastocoelar GADCs were fluorescein-positive in the larvae that developed from the 16-cell stage chimeric embryos; these comprised fluorescein-labeled mesomeres and unlabelled macromeres and micromeres. Our finding indicates that some of the blastocoelar GADCs are derived from the mesomeres and thus they are the new group of mesenchyme cells, the tertiary mesenchyme cells. PMID:24357228

Screening for Developmental Neurotoxicity Using PC12 Cells: Comparisons of Organophosphates with a Carbamate, an Organochlorine, and Divalent Nickel

PubMed Central

Slotkin, Theodore A.; MacKillop, Emiko A.; Ryde, Ian T.; Tate, Charlotte A.; Seidler, Frederic J.

2007-01-01

Background In light of the large number of chemicals that are potential developmental neurotoxicants, there is a need to develop rapid screening techniques. Objectives We exposed undifferentiated and differentiating neuronotypic PC12 cells to different organophosphates (chlorpyrifos, diazinon, parathion), a carbamate (physostigmine), an organochlorine (dieldrin), and a metal (divalent nickel; Ni2+) and examined indices of cell replication and differentiation for both short- and long-term exposures. Results In undifferentiated cells, all the agents inhibited DNA synthesis, with the greatest effect for diazinon, but physostigmine eventually produced the largest deficits in the total number of cells after prolonged exposure. The onset of differentiation intensified the adverse effects on DNA synthesis and changed the rank order in keeping with a shift away from noncholinergic mechanisms and toward cholinergic mechanisms. Differentiation also worsened the effects of each agent on cell number after prolonged exposure, whereas cell growth was not suppressed, nor were there any effects on viability as assessed with trypan blue. Nevertheless, differentiating cells displayed signs of oxidative stress from all of the test compounds except Ni2+, as evidenced by measurements of lipid peroxidation. Finally, all of the toxicants shifted the transmitter fate of the cells away from the cholinergic phenotype and toward the catecholaminergic phenotype. Conclusions These studies point out the feasibility of developing cell-based screening methods that enable the detection of multiple end points that may relate to mechanisms associated with developmental neurotoxicity, revealing some common targets for disparate agents. PMID:17366826

Microbiological tap water profile of a medium-sized building and effect of water stagnation.

PubMed

Lipphaus, Patrick; Hammes, Frederik; Kötzsch, Stefan; Green, James; Gillespie, Simon; Nocker, Andreas

2014-01-01

Whereas microbiological quality of drinking water in water distribution systems is routinely monitored for reasons of legal compliance, microbial numbers in tap water are grossly understudied. Motivated by gross differences in water from private households, we applied in this study flow cytometry as a rapid analytical method to quantify microbial concentrations in water sampled at diverse taps in a medium size research building receiving chlorinated water. Taps differed considerably in frequency of usage and were located in laboratories, bathrooms, and a coffee kitchen. Substantial differences were observed between taps with concentrations (per mL) in the range from 6.29 x 10(3) to 7.74 x 10(5) for total cells and from 1.66 x 10(3) to 4.31 x 10(5) for intact cells. The percentage of intact cells varied between 7% and 96%. Water from taps with very infrequent use showed the highest bacterial numbers and the highest proportions of intact cells. Stagnation tended to increase microbial numbers in water from those taps which were otherwise frequently used. Microbial numbers in other taps that were rarely opened were not affected by stagnation as their water is probably mostly stagnant. For cold water taps, microbial numbers and the percentage of intact cells tended to decline with flushing with the greatest decline for taps used least frequently whereas microbial concentrations in water from hot water taps tended to be somewhat more stable. We conclude that microbiological water quality is mainly determined by building-specific parameters. Tap water profiling can provide valuable insight into plumbing system hygiene and maintenance.

bcl-2 transgene inhibits T cell death and perturbs thymic self-censorship.

PubMed

Strasser, A; Harris, A W; Cory, S

1991-11-29

Early death is the fate of most developing T lymphocytes. Because bcl-2 can promote cell survival, we tested its impact in mice expressing an E mu-bcl-2 transgene within the T lymphoid compartment. The T cells showed remarkably sustained viability and some spontaneous differentiation in vitro. They also resisted killing by lymphotoxic agents. Although total T cell numbers and the rate of thymic involution were unaltered, the response to immunization was enhanced, consistent with reduced death of activated T cells. No T cells reactive with self-superantigens appeared in the lymph nodes, but an excess was found in the thymus. These observations, together with previous findings on B cells, suggest that modulated bcl-2 expression is a determinant of life and death in normal lymphocytes.

Parallel RNA extraction using magnetic beads and a droplet array.

PubMed

Shi, Xu; Chen, Chun-Hong; Gao, Weimin; Chao, Shih-Hui; Meldrum, Deirdre R

2015-02-21

Nucleic acid extraction is a necessary step for most genomic/transcriptomic analyses, but it often requires complicated mechanisms to be integrated into a lab-on-a-chip device. Here, we present a simple, effective configuration for rapidly obtaining purified RNA from low concentration cell medium. This Total RNA Extraction Droplet Array (TREDA) utilizes an array of surface-adhering droplets to facilitate the transportation of magnetic purification beads seamlessly through individual buffer solutions without solid structures. The fabrication of TREDA chips is rapid and does not require a microfabrication facility or expertise. The process takes less than 5 minutes. When purifying mRNA from bulk marine diatom samples, its repeatability and extraction efficiency are comparable to conventional tube-based operations. We demonstrate that TREDA can extract the total mRNA of about 10 marine diatom cells, indicating that the sensitivity of TREDA approaches single-digit cell numbers.

Parallel RNA extraction using magnetic beads and a droplet array

PubMed Central

Shi, Xu; Chen, Chun-Hong; Gao, Weimin; Meldrum, Deirdre R.

2015-01-01

Nucleic acid extraction is a necessary step for most genomic/transcriptomic analyses, but it often requires complicated mechanisms to be integrated into a lab-on-a-chip device. Here, we present a simple, effective configuration for rapidly obtaining purified RNA from low concentration cell medium. This Total RNA Extraction Droplet Array (TREDA) utilizes an array of surface-adhering droplets to facilitate the transportation of magnetic purification beads seamlessly through individual buffer solutions without solid structures. The fabrication of TREDA chips is rapid and does not require a microfabrication facility or expertise. The process takes less than 5 minutes. When purifying mRNA from bulk marine diatom samples, its repeatability and extraction efficiency are comparable to conventional tube-based operations. We demonstrate that TREDA can extract the total mRNA of about 10 marine diatom cells, indicating that the sensitivity of TREDA approaches single-digit cell numbers. PMID:25519439

Somatic mutations in cancer: Stochastic versus predictable.

PubMed

Gold, Barry

2017-02-01

The origins of human cancers remain unclear except for a limited number of potent environmental mutagens, such as tobacco and UV light, and in rare cases, familial germ line mutations that affect tumor suppressor genes or oncogenes. A significant component of cancer etiology has been deemed stochastic and correlated with the number of stem cells in a tissue, the number of times the stem cells divide and a low incidence of random DNA polymerase errors that occur during each cell division. While somatic mutations occur during each round of DNA replication, mutations in cancer driver genes are not stochastic. Out of a total of 2843 codons, 1031 can be changed to stop codons by a single base substitution in the tumor suppressor APC gene, which is mutated in 76% of colorectal cancers (CRC). However, the nonsense mutations, which comprise 65% of all the APC driver mutations in CRC, are not random: 43% occur at Arg CGA codons, although they represent <3% of the codons. In TP53, CGA codons comprise <3% of the total 393 codons but they account for 72% and 39% of the mutations in CRC and ovarian cancer OVC, respectively. This mutation pattern is consistent with the kinetically slow, but not stochastic, hydrolytic deamination of 5-methylcytosine residues at specific methylated CpG sites to afford T·G mismatches that lead to C→T transitions and stop codons at CGA. Analysis of nonsense mutations in CRC, OVC and a number of other cancers indicates the need to expand the predictable risk factors for cancer to include, in addition to random polymerase errors, the methylation status of gene body CGA codons in tumor suppressor genes. Copyright © 2017. Published by Elsevier B.V.

Natural killer cell activity, lymphocyte proliferation, and cytokine profile in tumor-bearing mice treated with MAPA, a magnesium aggregated polymer from Aspergillus oryzae.

PubMed

Justo, G Z; Durán, N; Queiroz, M L S

2003-08-01

The present study examined the effects of MAPA, an antitumor aggregated polymer of protein magnesium ammonium phospholinoleate-palmitoleate anhydride, isolated from Aspergillus oryzae, on concanavalin A (Con A)-induced spleen cell proliferation, cytokine production and on natural killer (NK) cell activity in Ehrlich ascites tumor-bearing mice. The Ehrlich ascites tumor (EAT) growth led to diminished mitogen-induced expansion of spleen cell populations and total NK activity. This was accompanied by striking spleen enlargement, with a marked increase in total cell counts. Moreover, a substantial enhancement in IL-10 levels, paralleled by a significant decrease in IL-2 was observed, while production of IL-4 and interferon-gamma (IFN-gamma) was not altered. Treatment of mice with 5 mg/kg MAPA for 7 days promoted spleen cell proliferation, IL-2 production and NK cell activity regardless of tumor outgrowth. In addition, MAPA treatment markedly enhanced IFN-gamma levels and reduced IL-10 production relative to EAT mice. A 35% reduction in splenomegaly with normal number of nucleated cells was also found. Altogether, our results suggest that MAPA directly and/or indirectly modulates immune cell activity, and probably disengages tumor-induced suppression of these responses. Clearly, MAPA has an impact and may delay tumor outgrowth through immunotherapeutic mechanisms.

Time related variations in stem cell harvesting of umbilical cord blood

NASA Astrophysics Data System (ADS)

Mazzoccoli, Gianluigi; Miscio, Giuseppe; Fontana, Andrea; Copetti, Massimiliano; Francavilla, Massimo; Bosi, Alberto; Perfetto, Federico; Valoriani, Alice; de Cata, Angelo; Santodirocco, Michele; Totaro, Angela; Rubino, Rosa; di Mauro, Lazzaro; Tarquini, Roberto

2016-02-01

Umbilical cord blood (UCB) contains hematopoietic stem cells and multipotent mesenchymal cells useful for treatment in malignant/nonmalignant hematologic-immunologic diseases and regenerative medicine. Transplantation outcome is correlated with cord blood volume (CBV), number of total nucleated cells (TNC), CD34+ progenitor cells and colony forming units in UCB donations. Several studies have addressed the role of maternal/neonatal factors associated with the hematopoietic reconstruction potential of UCB, including: gestational age, maternal parity, newborn sex and birth weight, placental weight, labor duration and mode of delivery. Few data exist regarding as to how time influences UCB collection and banking patterns. We retrospectively analyzed 17.936 cord blood donations collected from 1999 to 2011 from Tuscany and Apulia Cord Blood Banks. Results from generalized multivariable linear mixed models showed that CBV, TNC and CD34+ cell were associated with known obstetric and neonatal parameters and showed rhythmic patterns in different time domains and frequency ranges. The present findings confirm that volume, total nucleated cells and stem cells of the UCB donations are hallmarked by rhythmic patterns in different time domains and frequency ranges and suggest that temporal rhythms in addition to known obstetric and neonatal parameters influence CBV, TNC and CD34+ cell content in UBC units.

Analysis of interphase node proteins in fission yeast by quantitative and superresolution fluorescence microscopy

PubMed Central

Akamatsu, Matthew; Lin, Yu; Bewersdorf, Joerg; Pollard, Thomas D.

2017-01-01

We used quantitative confocal microscopy and FPALM superresolution microscopy of live fission yeast to investigate the structures and assembly of two types of interphase nodes—multiprotein complexes associated with the plasma membrane that merge together and mature into the precursors of the cytokinetic contractile ring. During the long G2 phase of the cell cycle, seven different interphase node proteins maintain constant concentrations as they accumulate in proportion to cell volume. During mitosis, the total numbers of type 1 node proteins (cell cycle kinases Cdr1p, Cdr2p, Wee1p, and anillin Mid1p) are constant even when the nodes disassemble. Quantitative measurements provide strong evidence that both types of nodes have defined sizes and numbers of constituent proteins, as observed for cytokinesis nodes. Type 1 nodes assemble in two phases—a burst at the end of mitosis, followed by steady increase during interphase to double the initial number. Type 2 nodes containing Blt1p, Rho-GEF Gef2p, and kinesin Klp8p remain intact throughout the cell cycle and are constituents of the contractile ring. They are released from the contractile ring as it disassembles and then associate with type 1 nodes around the equator of the cell during interphase. PMID:28539404

Modeling Cancer Cell Growth Dynamics In vitro in Response to Antimitotic Drug Treatment

PubMed Central

Lorz, Alexander; Botesteanu, Dana-Adriana; Levy, Doron

2017-01-01

Investigating the role of intrinsic cell heterogeneity emerging from variations in cell-cycle parameters and apoptosis is a crucial step toward better informing drug administration. Antimitotic agents, widely used in chemotherapy, target exclusively proliferative cells and commonly induce a prolonged mitotic arrest followed by cell death via apoptosis. In this paper, we developed a physiologically motivated mathematical framework for describing cancer cell growth dynamics that incorporates the intrinsic heterogeneity in the time individual cells spend in the cell-cycle and apoptosis process. More precisely, our model comprises two age-structured partial differential equations for the proliferative and apoptotic cell compartments and one ordinary differential equation for the quiescent compartment. To reflect the intrinsic cell heterogeneity that governs the growth dynamics, proliferative and apoptotic cells are structured in “age,” i.e., the amount of time remaining to be spent in each respective compartment. In our model, we considered an antimitotic drug whose effect on the cellular dynamics is to induce mitotic arrest, extending the average cell-cycle length. The prolonged mitotic arrest induced by the drug can trigger apoptosis if the time a cell will spend in the cell cycle is greater than the mitotic arrest threshold. We studied the drug’s effect on the long-term cancer cell growth dynamics using different durations of prolonged mitotic arrest induced by the drug. Our numerical simulations suggest that at confluence and in the absence of the drug, quiescence is the long-term asymptotic behavior emerging from the cancer cell growth dynamics. This pattern is maintained in the presence of small increases in the average cell-cycle length. However, intermediate increases in cell-cycle length markedly decrease the total number of cells and can drive the cancer population to extinction. Intriguingly, a large “switch-on/switch-off” increase in the average cell-cycle length maintains an active cell population in the long term, with oscillating numbers of proliferative cells and a relatively constant quiescent cell number. PMID:28913178

Insulin-Like Growth Factor 1 Mitigates Hematopoietic Toxicity After Lethal Total Body Irradiation

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

Zhou, Dunhua; Deoliveira, Divino; Kang, Yubin

2013-03-15

Purpose: To investigate whether and how insulin-like growth factor 1 (IGF-1) mitigates hematopoietic toxicity after total body irradiation. Methods and Materials: BALB/c mice were irradiated with a lethal dose of radiation (7.5 Gy) and treated with IGF-1 at a dose of 100 μg/dose intravenously once a day for 5 consecutive days starting within 1 hour after exposure. Survival and hematopoietic recovery were monitored. The mechanisms by which IGF-1 promotes hematopoietic recovery were also studied by use of an in vitro culture system. Results: IGF-1 protected 8 of 20 mice (40%) from lethal irradiation, whereas only 2 of 20 mice (10%) inmore » the saline control group survived for more than 100 days after irradiation. A single dose of IGF-1 (500 μg) was as effective as daily dosing for 5 days. Positive effects were noted even when the initiation of treatment was delayed as long as 6 hours after irradiation. In comparison with the saline control group, treatment with IGF-1 significantly accelerated the recovery of both platelets and red blood cells in peripheral blood, total cell numbers, hematopoietic stem cells, and progenitor cells in the bone marrow when measured at day 14 after irradiation. IGF-1 protected both hematopoietic stem cells and progenitor cells from radiation-induced apoptosis and cell death. In addition, IGF-1 was able to facilitate the proliferation and differentiation of nonirradiated and irradiated hematopoietic progenitor cells. Conclusions: IGF-1 mitigates radiation-induced hematopoietic toxicity through protecting hematopoietic stem cells and progenitor cells from apoptosis and enhancing proliferation and differentiation of the surviving hematopoietic progenitor cells.« less

[Neoplastic transformation of mouse fibroblasts under the influence of high-energy protons and gamma-rays].

PubMed

Voskanian, K Sh

2004-01-01

Oncoginic transformations of mouse fibroblasts C3H10T1/2 after exposure to proton energies 150 and 584 MeV were compared with fibroblast effects of gamma-radiation. Prior to exposure, cell populations (2.7 x 10(3) cells/cm2) were inoculated in plastic vials with the surface area of 75 cm2 and cultivated 11 days. Survivability was determined by comparing the number of cell colonies in irradiated and non-irradiated (control) vials. Transformation rate was calculated by dividing the total transformation focus number by the number of survived cells in a vial. Rate of oncogenic transformations after gamma- and proton (584 MeV) irradiation was essentially identical, i.e. the parameter grew rapidly at the doses < 1 Gy and slowed down at the doses > 1 Gy. In the dose interval between 1 and 5 Gy, transformation rate for proton energy 150 MeV was found low compared with gamma-radiation and proton energy 584 MeV. It is hypothesized that the different transformation rate after exposure to proton energy 150 MeV is linked with the high linear energy transfer as compared with the proton energy of 584 MeV and gamma-radiation.

Better Quality and More Usable Embryos Obtained on Day 3 Cultured in 5% Than 20% Oxygen: A Controlled and Randomized Study Using the Sibling Oocytes.

PubMed

Peng, Huixia; Shi, Wenhao; Zhang, Wei; Xue, Xia; Li, Na; Li, Wei; Shi, Juanzi

2016-03-01

To evaluate the effect of atmospheric oxygen (O2) concentration on embryonic development, a controlled and randomized study using the sibling oocytes was carried out. A total of 147 patients were studied. Embryos were cultured in O2 concentration 20% versus 5% during the gamete, zygote, and first 3 days. The mean cell numbers of embryo (7.69 ± 1.91 vs 7.20 ± 1.82, P = .011) and rate of clinically useable embryos (81.62% vs 77.22%, P = .017) were significantly higher in 5% O2 than in 20% O2. There was no difference in the zygote developmental stage, day 2, day 4, and blastocyst stage. The quality of blastocyst (both inner cell mass and trophectoderm) showed no difference. Also, there was no increase in embryos fragmentation and uneven cells in 20% O2 culture condition. In conclusion, 20% O2 reduced the mean cell numbers of embryo and the number of clinically useable embryos on day 3. However, there was no subsequent negative impact on development of day 4 and blastocyst stage. © The Author(s) 2015.

Clonorchis sinensis-derived total protein attenuates airway inflammation in murine asthma model by inducing regulatory T cells and modulating dendritic cell functions

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

Jeong, Young-Il; Kim, Seung Hyun; Ju, Jung Won

Highlights: {yields} Treatment with Clonorchis sinensis-derived total protein attenuates OVA-induced airway inflammation and AHR to methacholine. {yields} Induction of CD4{sup +}CD25{sup +}Foxp3{sup +} T cells and IL-10 along with suppression of splenocyte proliferation by C. sinensis-derived total protein. {yields} C. sinensis-derived total protein interferes with the expression of co-stimulatory molecules in DCs. -- Abstract: Asthma is characterized by Th2-mediated inflammation, resulting in airway hyperresponsiveness (AHR) through airway remodeling. Recent epidemiological and experimental reports have suggested an inverse relationship between the development of allergy and helminth infections. Infection by Clonorchis sinensis, a liver fluke that resides in the bile duct ofmore » humans, is endemic predominantly in Asia including Korea and China. Using a murine model for asthma, we investigated the effects of C. sinensis-derived total protein (Cs-TP) on allergen-induced airway inflammation and the mechanism underlying the protective effects of Cs-TP administration on asthma. Treatment with Cs-TP attenuated OVA-induced airway inflammation and methacholine-induced AHR, as well as eosinophilia development, lymphocyte infiltration into the lung, and goblet cell metaplasia. This protective effect of Cs-TP is associated with markedly reduced OVA-specific IgE and Th1/Th2 cytokine production. Moreover, Cs-TP increased the number of CD4{sup +}CD25{sup +}Foxp3{sup +} regulatory T (Treg) cells as well as their suppressive activity. In fact, proliferation of OVA-restimulated splenocytes was suppressed significantly. Cs-TP also inhibited the expression of such co-stimulatory molecules as CD80, CD86, and CD40 in LPS- or OVA-stimulated dendritic cells (DCs), suggesting that Cs-TP could interfere with the capacity of airway DCs to prime naive T cells. These data demonstrate the capacity of C. sinensis to ameliorate allergic asthma and broaden our understanding of the paradoxical relationship between the allergic immune response and helminth infection.« less

Comparative Analysis of Transcriptomes of Macrophage Revealing the Mechanism of the Immunoregulatory Activities of a Novel Polysaccharide Isolated from Boletus speciosus Frost

PubMed Central

Ding, Xiang; Zhu, Hongqing; Hou, Yiling; Hou, Wanru; Zhang, Nan; Fu, Lei

2017-01-01

Background: The mechanism of the immunoregulatory activities of polysaccharide is still not clear. Materials and Methods: Here, we performed the B-cell, T-cell, and macrophage cell proliferation, the cell cycle analysis of macrophage cells, sequenced the transcriptomes of control group macrophages, and Boletus speciosus Frost polysaccharide (BSF-1) group macrophages using Illumina sequencing technology to identify differentially expressed genes (DEGs) to determine the molecular mechanisms of immunomodulatory activity of BSF-1 in macrophages. Results: These results suggested that BSF-1 could promote the proliferation of B-cell, T-cell, and macrophages, promote the proliferation of macrophage cells by abolishing cell cycle arrests in the G0/G1 phases, and promote cell cycle progression in S-phase and G2/M phase, which might induce cell division. A total of 12,498,414 and 11,840,624 bp paired-end reads were obtained for the control group and BSF-1 group, respectively, and they corresponded to a total size of 12.5 G bp and 11.8 G bp, respectively, after the low-quality reads and adapter sequences were removed. Approximately 81.83% of the total number of genes (8,257) were expressed reads per kilobase per million mapped reads (RPKM ≥1) and more than 1366 genes were highly expressed (RPKM >60) in the BSF-1 group. A gene ontology-enrichment analysis generated 13,042 assignments to cellular components, 13,094 assignments to biological processes, and 13,135 assignments to molecular functions. A Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed that the mitogen-activated protein kinase (MAPK) signaling pathways are significantly enriched for DEGs between the two cell groups. Conclusion: An analysis of transcriptome resources enabled us to examine gene expression profiles, verify differential gene expression, and select candidate signaling pathways as the mechanisms of the immunomodulatory activity of BSF-1. Based on the experimental data, we believe that the significant antitumor activities of BSF-1 in vivo mainly involve the MAPK signaling pathways. SUMMARY Boletus speciosus Frost-1 (BSF-1) could promote the proliferation of B-cell, T-cell, and macrophages, promote the proliferation of macrophage cells by abolishing cell cycle arrests in the G0/G1 phases, and promote cell cycle progression in S-phase and G2/M phase, which might induce cell divisionApproximately 81.83% of the total number of genes (8257) were expressed (reads per kilobase per million mapped reads [RPKM] =1) and more than 1366 genes were highly expressed (RPKM >60) in the BSF-1 groupA gene ontology-enrichment analysis generated 13,042 assignments to cellular components, 13,094 assignments to biological processes, and 13,135 assignments to molecular functionsA Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed that the mitogen-activated protein kinase signaling pathways are significantly enriched for DEGs between the two cell groups. Abbreviations used: BSF-1: Boletus speciosus Frost polysaccharide. PMID:28839373

Ganglion cell distribution and retinal resolution in the Florida manatee, Trichechus manatus latirostris.

PubMed

Mass, Alla M; Ketten, Darlene R; Odell, Daniel K; Supin, Alexander Ya

2012-01-01

The topographic organization of retinal ganglion cells was examined in the Florida manatee (Trichechus manatus latirostris) to assess ganglion cell size and distribution and to estimate retinal resolution. The ganglion cell layer of the manatee's retina was comprised primarily of large neurons with broad intercellular spaces. Cell sizes varied from 10 to 60 μm in diameter (mean 24.3 μm). The retinal wholemounts from adult animals measured 446-501 mm(2) in area with total ganglion cell counts of 62,000-81,800 (mean 70,200). The cell density changed across the retina, with the maximum in the area below the optic disc and decreasing toward the retinal edges and in the immediate vicinity of the optic disc. The maximum cell density ranged from 235 to 337 cells per millimeter square in the adult retinae. Two wholemounts obtained from juvenile animals were 271 and 282 mm(2) in area with total cell numbers of 70,900 and 68,700, respectively (mean 69,800), that is, nearly equivalent to those of adults, but juvenile retinae consequently had maximum cell densities that were higher than those of adults: 478 and 491 cells per millimeter square. Calculations indicate a retinal resolution of ∼19' (1.6 cycles per degree) in both adult and juvenile retinae. Copyright © 2011 Wiley Periodicals, Inc.

In Vivo Chemoprotective Activity of Bovine Dialyzable Leukocyte Extract in Mouse Bone Marrow Cells against Damage Induced by 5-Fluorouracil

PubMed Central

Coronado-Cerda, Erika Evangelina; Franco-Molina, Moisés Armides; Mendoza-Gamboa, Edgar; Prado-García, Heriberto; Rivera-Morales, Lydia Guadalupe; Zapata-Benavides, Pablo; Rodríguez-Salazar, María del Carmen; Caballero-Hernandez, Diana; Tamez-Guerra, Reyes Silvestre; Rodríguez-Padilla, Cristina

2016-01-01

Chemotherapy treatments induce a number of side effects, such as leukopenia neutropenia, peripheral erythropenia, and thrombocytopenia, affecting the quality of life for cancer patients. 5-Fluorouracil (5-FU) is wieldy used as myeloablative model in mice. The bovine dialyzable leukocyte extract (bDLE) or IMMUNEPOTENT CRP® (ICRP) is an immunomodulatory compound that has antioxidants and anti-inflammatory effects. In order to investigate the chemoprotection effect of ICRP on bone marrow cells in 5-FU treated mice, total bone marrow (BM) cell count, bone marrow colony forming units-granulocyte/macrophage (CFU-GM), cell cycle, immunophenotypification, ROS/superoxide and Nrf2 by flow cytometry, and histological and hematological analyses were performed. Our results demonstrated that ICRP increased BM cell count and CFU-GM number, arrested BM cells in G0/G1 phase, increased the percentage of leukocyte, granulocytic, and erythroid populations, reduced ROS/superoxide formation and Nrf2 activation, and also improved hematological levels and weight gain in 5-FU treated mice. These results suggest that ICRP has a chemoprotective effect against 5-FU in BM cells that can be used in cancer patients. PMID:27191003

Study on mobile phone use while driving in a sample of Iranian drivers.

PubMed

Arvin, Ramin; Khademi, Mostafa; Razi-Ardakani, Hesamoddin

2017-06-01

The use of cell phone is a significant source of driver distraction. Phone use while driving can impair a number of factors critical for safe driving which can cause serious traffic safety problems. The objective of this paper was to investigate the frequency of using cell phones while driving in Iran's roads through an observational survey with a random sample of drivers, to recognize contributing factors to cell phone usage and to understand the magnitude of the problem. A total of 1794 observations were collected from 12 sites at controlled intersections, entrance and exit points of highways. The cell phone use rate among drivers (talking or texting) was estimated at 10% which is significantly higher than that in other countries such as Australia, USA and Canada. Rate of cell phone use among younger drivers (14.15%) was higher in comparison with other groups. In order to identify factors affecting cell phone use while driving, a binary logit model is estimated. Variables which significantly contribute to the rate of using cell phone were found to be the age of driver, number of passengers, presence of kids under the age of 8, time of observation, vehicle price and type of car.

Tetrahydrocannabinol-induced suppression of macrophage spreading and phagocytic activity in vitro.

PubMed

Lopez-Cepero, M; Friedman, M; Klein, T; Friedman, H

1986-06-01

The effects of tetrahydrocannabinol (THC) on several parameters of macrophage function in vitro were assessed. Delta 9 THC added to cultures of normal mouse peritoneal cells in vitro affected the ability of the cells to spread on glass surfaces and also had some effect on their ability to phagocytize yeast. These effects were dose related. A concentration of 20 micrograms of THC almost completely inhibited macrophage spreading, but it also decreased viability and the total number of these cells. Doses of 10 or 5 micrograms of THC also inhibited spreading but had little effect on cell viability or number. A dose of 1.0 microgram of THC had some inhibitory effect on spreading and the lowest dose affecting spreading appeared to be about 0.05 micrograms per culture. Higher doses of THC were necessary to inhibit phagocytosis of yeast particles as determined by direct microscopic examination or use of radiolabeled yeast as the test particles. These results indicate that several readily measured functions of macrophages may be suppressed by THC.

Copy Number Variation Is a Fundamental Aspect of the Placental Genome

PubMed Central

Hannibal, Roberta L.; Chuong, Edward B.; Rivera-Mulia, Juan Carlos; Gilbert, David M.; Valouev, Anton; Baker, Julie C.

2014-01-01

Discovery of lineage-specific somatic copy number variation (CNV) in mammals has led to debate over whether CNVs are mutations that propagate disease or whether they are a normal, and even essential, aspect of cell biology. We show that 1,000N polyploid trophoblast giant cells (TGCs) of the mouse placenta contain 47 regions, totaling 138 Megabases, where genomic copies are underrepresented (UR). UR domains originate from a subset of late-replicating heterochromatic regions containing gene deserts and genes involved in cell adhesion and neurogenesis. While lineage-specific CNVs have been identified in mammalian cells, classically in the immune system where V(D)J recombination occurs, we demonstrate that CNVs form during gestation in the placenta by an underreplication mechanism, not by recombination nor deletion. Our results reveal that large scale CNVs are a normal feature of the mammalian placental genome, which are regulated systematically during embryogenesis and are propagated by a mechanism of underreplication. PMID:24785991

Stereochemistry Balances Cell Permeability and Solubility in the Naturally Derived Phepropeptin Cyclic Peptides.

PubMed

Schwochert, Joshua; Lao, Yongtong; Pye, Cameron R; Naylor, Matthew R; Desai, Prashant V; Gonzalez Valcarcel, Isabel C; Barrett, Jaclyn A; Sawada, Geri; Blanco, Maria-Jesus; Lokey, R Scott

2016-08-11

Cyclic peptide (CP) natural products provide useful model systems for mapping "beyond-Rule-of-5" (bRo5) space. We identified the phepropeptins as natural product CPs with potential cell permeability. Synthesis of the phepropeptins and epimeric analogues revealed much more rapid cellular permeability for the natural stereochemical pattern. Despite being more cell permeable, the natural compounds exhibited similar aqueous solubility as the corresponding epimers, a phenomenon explained by solvent-dependent conformational flexibility among the natural compounds. When analyzing the polarity of the solution structures we found that neither the number of hydrogen bonds nor the total polar surface area accurately represents the solvation energies of the high and low dielectric conformations. This work adds to a growing number of natural CPs whose solvent-dependent conformational behavior allows for a balance between aqueous solubility and cell permeability, highlighting structural flexibility as an important consideration in the design of molecules in bRo5 chemical space.

Interferon regulatory factor 4 (IRF4) controls myeloid-derived suppressor cell (MDSC) differentiation and function.

PubMed

Nam, Sorim; Kang, Kyeongah; Cha, Jae Seon; Kim, Jung Woo; Lee, Hee Gu; Kim, Yonghwan; Yang, Young; Lee, Myeong-Sok; Lim, Jong-Seok

2016-12-01

Myeloid-derived suppressor cells (MDSCs) are immature cells that do not differentiate into mature myeloid cells. Two major populations of PMN-MDSCs (Ly6G high Ly6C low Gr1 high CD11b + ) and MO-MDSCs (Ly6G - Ly6C high Gr-1 int CD11b + ) have an immune suppressive function. Interferon regulatory factor 4 (IRF4) has a role in the negative regulation of TLR signaling and is associated with lymphoid cell development. However, the roles of IRF4 in myeloid cell differentiation are unclear. In this study, we found that IRF4 expression was remarkably suppressed during the development of MDSCs in the tumor microenvironment. Both the mRNA and protein levels of IRF4 in MDSCs were gradually reduced, depending on the development of tumors in the 4T1 model. siRNA-mediated knockdown of IRF4 in bone marrow cells promoted the differentiation of PMN-MDSCs. Similarly, IRF4 inhibition in bone marrow cells using simvastatin, which has been known to inhibit IRF4 expression, increased PMN-MDSC numbers. In contrast, IRF4 overexpression in bone marrow cells inhibited the total numbers of MDSCs, especially PMN-MDSCs. Notably, treatment with IL-4, an upstream regulator of IRF4, induced IRF4 expression in the bone marrow cells, and consequently, IL-4-induced IRF4 expression resulted in a decrease in PMN-MDSC numbers. Finally, we confirmed that IRF4 expression in MDSCs can modulate their activity to inhibit T cell proliferation through IL-10 production and ROS generation, and myeloid-specific deletion of IRF4 leads to the increase of MDSC differentiation. Our present findings indicate that IRF4 reduction induced by tumor formation can increase the number of MDSCs, and increases in the IRF4 expression in MDSCs may infringe on the immune-suppressive function of MDSCs. © Society for Leukocyte Biology.

Preterm Cord Blood Contains a Higher Proportion of Immature Hematopoietic Progenitors Compared to Term Samples

PubMed Central

Podestà, Marina; Bruschettini, Matteo; Cossu, Claudia; Sabatini, Federica; Dagnino, Monica; Romantsik, Olga; Spaggiari, Grazia Maria; Ramenghi, Luca Antonio; Frassoni, Francesco

2015-01-01

Background Cord blood contains high number of hematopoietic cells that after birth disappear. In this paper we have studied the functional properties of the umbilical cord blood progenitor cells collected from term and preterm neonates to establish whether quantitative and/or qualitative differences exist between the two groups. Methods and Results Our results indicate that the percentage of total CD34+ cells was significantly higher in preterm infants compared to full term: 0.61% (range 0.15–4.8) vs 0.3% (0.032–2.23) p = 0.0001 and in neonates <32 weeks of gestational age (GA) compared to those ≥32 wks GA: 0.95% (range 0.18–4.8) and 0.36% (0.15–3.2) respectively p = 0.0025. The majority of CD34+ cells co-expressed CD71 antigen (p<0.05 preterm vs term) and grew in vitro large BFU-E, mostly in the second generation. The subpopulations CD34+CD38- and CD34+CD45- resulted more represented in preterm samples compared to term, conversely, Side Population (SP) did not show any difference between the two group. The absolute number of preterm colonies (CFCs/10microL) resulted higher compared to term (p = 0.004) and these progenitors were able to grow until the third generation maintaining an higher proportion of CD34+ cells (p = 0.0017). The number of colony also inversely correlated with the gestational age (Pearson r = -0.3001 p<0.0168). Conclusions We found no differences in the isolation and expansion capacity of Endothelial Colony Forming Cells (ECFCs) from cord blood of term and preterm neonates: both groups grew in vitro large number of endothelial cells until the third generation and showed a transitional phenotype between mesenchymal stem cells and endothelial progenitors (CD73, CD31, CD34 and CD144)The presence, in the cord blood of preterm babies, of high number of immature hematopoietic progenitors and endothelial/mesenchymal stem cells with high proliferative potential makes this tissue an important source of cells for developing new cells therapies. PMID:26417990

Preterm Cord Blood Contains a Higher Proportion of Immature Hematopoietic Progenitors Compared to Term Samples.

PubMed

Podestà, Marina; Bruschettini, Matteo; Cossu, Claudia; Sabatini, Federica; Dagnino, Monica; Romantsik, Olga; Spaggiari, Grazia Maria; Ramenghi, Luca Antonio; Frassoni, Francesco

2015-01-01

Cord blood contains high number of hematopoietic cells that after birth disappear. In this paper we have studied the functional properties of the umbilical cord blood progenitor cells collected from term and preterm neonates to establish whether quantitative and/or qualitative differences exist between the two groups. Our results indicate that the percentage of total CD34+ cells was significantly higher in preterm infants compared to full term: 0.61% (range 0.15-4.8) vs 0.3% (0.032-2.23) p = 0.0001 and in neonates <32 weeks of gestational age (GA) compared to those ≥32 wks GA: 0.95% (range 0.18-4.8) and 0.36% (0.15-3.2) respectively p = 0.0025. The majority of CD34+ cells co-expressed CD71 antigen (p<0.05 preterm vs term) and grew in vitro large BFU-E, mostly in the second generation. The subpopulations CD34+CD38- and CD34+CD45- resulted more represented in preterm samples compared to term, conversely, Side Population (SP) did not show any difference between the two group. The absolute number of preterm colonies (CFCs/10microL) resulted higher compared to term (p = 0.004) and these progenitors were able to grow until the third generation maintaining an higher proportion of CD34+ cells (p = 0.0017). The number of colony also inversely correlated with the gestational age (Pearson r = -0.3001 p<0.0168). We found no differences in the isolation and expansion capacity of Endothelial Colony Forming Cells (ECFCs) from cord blood of term and preterm neonates: both groups grew in vitro large number of endothelial cells until the third generation and showed a transitional phenotype between mesenchymal stem cells and endothelial progenitors (CD73, CD31, CD34 and CD144)The presence, in the cord blood of preterm babies, of high number of immature hematopoietic progenitors and endothelial/mesenchymal stem cells with high proliferative potential makes this tissue an important source of cells for developing new cells therapies.

Increased number of mast cells in the dermis in actinic keratosis lesions effectively treated with imiquimod.

PubMed

Oyama, Satomi; Funasaka, Yoko; Tsuchiya, Shin-Ichi; Kawana, Seiji; Saeki, Hidehisa

2017-08-01

Actinic keratosis (AK) is a cutaneous cancer in situ which develops as a result of excessive exposure to ultraviolet (UV). Toll-like receptor (TLR)7 agonist imiquimod is a topical immune response modifier and is effective for the treatment of non-melanoma skin cancers. Recently, the diagnostic role of the dermatoscope has been reported in the course of treatment of AK. In addition, mast cells are now considered to contribute to both the innate and adaptive immune systems in topical imiquimod therapy. We assessed the effect of imiquimod treatment by dermatoscopic and immunohistochemical findings in 14 patients with a total of 21 AK lesions. With the dermatoscope, though the mean erythema score was not significantly different between the cured lesions and the unresponsive lesions, the erythema/red pseudo-network ("strawberry") pattern was decreased significantly in the cured lesions. By immunohistochemistry, the number of Ki-67-positive proliferative cells in the epidermis was decreased and that of CD117-positive mast cells in the dermis was increased in the responding lesions. To the best of our knowledge, this is the first study demonstrating that the number of mast cells in the dermis was increased in AK lesions effectively treated with imiquimod. Our present result suggests that mast cells may contribute an antitumor effect in human skin treated with topical imiquimod. © 2017 Japanese Dermatological Association.

Th17 cells and CD4(+) multifunctional T cells in patients with systemic lupus erythematosus.

PubMed

Araújo, Júlio Antônio Pereira; Mesquita, Danilo; de Melo Cruvinel, Wilson; Salmazi, Karina Inácio; Kallás, Esper Georges; Andrade, Luis Eduardo Coelho

2016-01-01

Recent evidence suggests that abnormalities involving Th17 lymphocytes are associated with the pathophysiology of systemic lupus erythematosus (SLE). In addition, multifunctional T cells (MFT), i.e., those producing multiple cytokines simultaneously, are present in the inflammatory milieu and may be implicated in the autoimmune process observed in SLE. In the present study, we aimed to characterize the functional status of CD4(+) T cells in SLE by simultaneously determining the concentration of IL-2, IFN-γ and IL-17 in lymphocyte cultures under exogenous and self-antigenic stimuli. Eighteen patients with active disease, 18 with inactive disease, and 14 healthy controls had functional status of CD4(+) T cells analyzed. We found that SLE patients presented a decreased number of total CD4(+) cells, an increased number of activated T cells, and an increased frequency of Th17 cells compared to healthy controls (HC). MFT cells had increased frequency in SLE patients and there was an increased frequency of tri-functional MFT in patients with active SLE compared with those with inactive SLE. Interestingly, MTF cells produced larger amounts of IFNγ than mono-functional T cells in patients and controls. Taken together these data indicate the participation of recently activated Th17 cells and MTF cells in the SLE pathophysiology. Copyright © 2015 Elsevier Editora Ltda. All rights reserved.

Quantitative Assessment of Proliferative Effects of Oral Vanadium on Pancreatic Islet Volumes and Beta Cell Numbers of Diabetic Rats.

PubMed

Pirmoradi, Leila; Noorafshan, Ali; Safaee, Akbar; Dehghani, Gholam Abbas

2016-01-01

Oral vanadyl sulfate (vanadium) induces normoglycemia, proliferates beta cells and prevents pancreatic islet atrophy in streptozotocin-induced diabetic rats. Soteriological method is used to quantitate the proliferative effects of vanadium on beta-cell numbers and islet volumes of normal and diabetic rats. Adult male Sprague-Dawley rats were made diabetic with intravenous streptozotocin injection (40 mg/kg). Normal and diabetic rats were divided into four groups. While control normal and diabetic (CD) groups used water, vanadium-treated normal (VTN) and diabetic (VTD) groups used solutions containing vanadyl sulfate (0.5-1 mg/mL, VOSO4+5H2O). Tail blood samples were used to measure blood glucose (BG) and plasma insulin. Two months after treatment, rats were sacrificed, pancreata prepared, and stereology method was used to quantitatively evaluate total beta cell numbers (TBCN) and total islet volumes (TISVOL). Normoglycemia persisted in VTN with significantly decreased plasma insulin (0.19±0.08 vs. 0.97±0.27 ng/dL, P<0.002). The respective high BG (532±49 vs. 144±46 mg/dL, P<0.0001) and reduced plasma insulin (0.26±0.15 vs. 0.54±0.19 ng/dL, P<0.002) seen in CD were reversed in VTD during vanadium treatment or withdrawal. While the induction of diabetes, compared to their control, significantly decreased TISVOL (1.9±0.2 vs. 3.03±0.6 mm3, P<0.003) and TBCN (0.99±0.1 vs. 3.2±0.2 x 106, P<0.003), vanadium treatment significantly increased TISVOL (2.9±0.8 and 4.07±1.0 mm3, P<0.003) and TBCN (1.5±0.3 and 3.8±0.6 x 106, P<0.03). Two-month oral vanadium therapy in STZ-diabetic rats ameliorated hyperglycemia by partially restoring plasma insulin. This action was through proliferative actions of vanadium in preventing islet atrophy by increasing beta-cell numbers.

T-cell immunity and cytokine production in cosmonauts after long-duration space flights

NASA Astrophysics Data System (ADS)

Morukov, B.; Rykova, M.; Antropova, E.; Berendeeva, T.; Ponomaryov, S.; Larina, I.

2011-04-01

Long-duration spaceflight effects on T-cell immunity and cytokine production were studied in 12 Russian cosmonauts flown onto the International Space Station. Specific assays were performed before launch and after landing and included analysis of peripheral leukocyte distribution, analysis of T-cell phenotype, expression of activation markers, apoptosis, proliferation of T cells in response to a mitogen, concentrations of cytokines in supernatants of cell cultures. Statistically significant increase was observed in leukocytes', lymphocytes', monocytes' and granulocytes' total number, increase in percentage and absolutely number of CD3 +CD4 +-cells, CD4 +CD45RA +-cells and CD4 +CD45RA +/CD4 +CD45RО + ratio, CD4 +CD25 +Bright regulatory cells ( p<0,05) in peripheral blood after landing. T-lymphocytes' capacity to present CD69 and CD25 on its own surfaces was increased for the majority of crewmembers. Analysis of T-cell response to PHA-stimulation in vitro revealed there were some trends toward reduced proliferation of stimulated T-lymphocytes. There was an apparent post flight decrease in secreted IFN-g for the majority of crewmembers and in most instances there was elevation in secreted IL-10. It revealed depression of IFN-g/IL-10 ratio after flight. Correlation analysis according to Spearman's rank correlation test established significant positive correlations ( p<0.05) between cytokine production and T-cell activation (CD25+, CD38+) and negative correlation ( p<0.05) between cytokine production and number of bulk memory CD4+T-cells (CD45RO+). Thus, these results suggest that T-cell dysfunction can be conditioned by cytokine dysbalance and could lead to development of disease after long-duration space flights.

Angels and demons: Th17 cells represent a beneficial response, while neutrophil IL-17 is associated with poor prognosis in squamous cervical cancer.

PubMed

Punt, Simone; Fleuren, Gert Jan; Kritikou, Eva; Lubberts, Erik; Trimbos, J Baptist; Jordanova, Ekaterina S; Gorter, Arko

2015-01-01

The role of interleukin (IL)-17 in cancer remains controversial. In view of the growing interest in the targeting of IL-17, knowing its cellular sources and clinical implications is crucial. In the present study, we unraveled the phenotype of IL-17 expressing cells in cervical cancer using immunohistochemical double and immunofluorescent triple stainings. In the tumor stroma, IL-17 was found to be predominantly expressed by neutrophils (66%), mast cells (23%), and innate lymphoid cells (8%). Remarkably, T-helper 17 (Th17) cells were a minor IL-17 expressing population (4%). A similar distribution was observed in the tumor epithelium. The Th17 and granulocyte fractions were confirmed in head and neck, ovarian, endometrial, prostate, breast, lung, and colon carcinoma. An above median number of total IL-17 expressing cells was an independent prognostic factor for poor disease-specific survival in early stage disease ( p = 0.016). While a high number of neutrophils showed at trend toward poor survival, the lowest quartile of mast cells correlated with poor survival ( p = 0.011). IL-17 expressing cells and neutrophils were also correlated with the absence of vaso-invasion ( p < 0.01). IL-17 was found to increase cell growth or tightness of cervical cancer cell lines, which may be a mechanism for tumorigenesis in early stage disease. These data suggest that IL-17, primarily expressed by neutrophils, predominantly promotes tumor growth, correlated with poor prognosis in early stage disease. Strikingly, a high number of Th17 cells was an independent prognostic factor for improved survival ( p = 0.026), suggesting Th17 cells are part of a tumor suppressing immune response.

Angels and demons: Th17 cells represent a beneficial response, while neutrophil IL-17 is associated with poor prognosis in squamous cervical cancer

PubMed Central

Punt, Simone; Fleuren, Gert Jan; Kritikou, Eva; Lubberts, Erik; Trimbos, J. Baptist; Jordanova, Ekaterina S.; Gorter, Arko

2015-01-01

The role of interleukin (IL)-17 in cancer remains controversial. In view of the growing interest in the targeting of IL-17, knowing its cellular sources and clinical implications is crucial. In the present study, we unraveled the phenotype of IL-17 expressing cells in cervical cancer using immunohistochemical double and immunofluorescent triple stainings. In the tumor stroma, IL-17 was found to be predominantly expressed by neutrophils (66%), mast cells (23%), and innate lymphoid cells (8%). Remarkably, T-helper 17 (Th17) cells were a minor IL-17 expressing population (4%). A similar distribution was observed in the tumor epithelium. The Th17 and granulocyte fractions were confirmed in head and neck, ovarian, endometrial, prostate, breast, lung, and colon carcinoma. An above median number of total IL-17 expressing cells was an independent prognostic factor for poor disease-specific survival in early stage disease (p = 0.016). While a high number of neutrophils showed at trend toward poor survival, the lowest quartile of mast cells correlated with poor survival (p = 0.011). IL-17 expressing cells and neutrophils were also correlated with the absence of vaso-invasion (p < 0.01). IL-17 was found to increase cell growth or tightness of cervical cancer cell lines, which may be a mechanism for tumorigenesis in early stage disease. These data suggest that IL-17, primarily expressed by neutrophils, predominantly promotes tumor growth, correlated with poor prognosis in early stage disease. Strikingly, a high number of Th17 cells was an independent prognostic factor for improved survival (p = 0.026), suggesting Th17 cells are part of a tumor suppressing immune response. PMID:25949866

Breast cancer genome and transcriptome integration implicates specific mutational signatures with immune cell infiltration

PubMed Central

Smid, Marcel; Rodríguez-González, F. Germán; Sieuwerts, Anieta M.; Salgado, Roberto; Prager-Van der Smissen, Wendy J. C.; Vlugt-Daane, Michelle van der; van Galen, Anne; Nik-Zainal, Serena; Staaf, Johan; Brinkman, Arie B.; van de Vijver, Marc J.; Richardson, Andrea L.; Fatima, Aquila; Berentsen, Kim; Butler, Adam; Martin, Sancha; Davies, Helen R.; Debets, Reno; Gelder, Marion E. Meijer-Van; van Deurzen, Carolien H. M.; MacGrogan, Gaëtan; Van den Eynden, Gert G. G. M.; Purdie, Colin; Thompson, Alastair M.; Caldas, Carlos; Span, Paul N.; Simpson, Peter T.; Lakhani, Sunil R.; Van Laere, Steven; Desmedt, Christine; Ringnér, Markus; Tommasi, Stefania; Eyford, Jorunn; Broeks, Annegien; Vincent-Salomon, Anne; Futreal, P. Andrew; Knappskog, Stian; King, Tari; Thomas, Gilles; Viari, Alain; Langerød, Anita; Børresen-Dale, Anne-Lise; Birney, Ewan; Stunnenberg, Hendrik G.; Stratton, Mike; Foekens, John A.; Martens, John W. M.

2016-01-01

A recent comprehensive whole genome analysis of a large breast cancer cohort was used to link known and novel drivers and substitution signatures to the transcriptome of 266 cases. Here, we validate that subtype-specific aberrations show concordant expression changes for, for example, TP53, PIK3CA, PTEN, CCND1 and CDH1. We find that CCND3 expression levels do not correlate with amplification, while increased GATA3 expression in mutant GATA3 cancers suggests GATA3 is an oncogene. In luminal cases the total number of substitutions, irrespective of type, associates with cell cycle gene expression and adverse outcome, whereas the number of mutations of signatures 3 and 13 associates with immune-response specific gene expression, increased numbers of tumour-infiltrating lymphocytes and better outcome. Thus, while earlier reports imply that the sheer number of somatic aberrations could trigger an immune-response, our data suggests that substitutions of a particular type are more effective in doing so than others. PMID:27666519

Impact of an emergency department pain management protocol on the pattern of visits by patients with sickle cell disease.

PubMed

Givens, Melissa; Rutherford, Cynthia; Joshi, Girish; Delaney, Kathleen

2007-04-01

This study explores how implementation of pain management guidelines in concert with clinic case management affected emergency department (ED) utilization, clinic visits, and hospital admissions for patients with sickle cell disease. A pain management guideline that eliminated meperidine and encouraged timely use of morphine or hydromorphone for pain control in sickle cell crisis was introduced as a quality improvement project. This study is a retrospective review of ED visits, clinic visits, and admissions from 1 year before and 3 years after the guideline implementation. Working with the ED, the Hematology Clinic began to proactively seek the return of their patients for clinic follow-up. A formal case management program for sickle cell patients was initiated in June 2003. A total of 1584 visits by 223 patients were collected, 1097 to the ED and 487 to the Hematology Clinic. Total hospital visits did not change significantly in any of the 4 years, p > 0.10 for each comparison. Total ED visits decreased significantly over the 4-year study period (p < 0.001), whereas clinic visits steadily increased (p < 0.001). Return visits to the ED within 30 days also declined significantly, p < 0.001. Both the absolute number of admissions per year and the total admissions per hospital visit per year declined significantly over the study period, p = 0.001. Although total admissions per hospital visit did not change, the proportion of ED visits that resulted in admission in year 1 (29%) was significantly lower than the proportion admitted in year 2 (43%), p = 0.04. A pain protocol using morphine or hydromorphone coupled with increased access to outpatient clinics decreased ED visits, hospitalizations, and increased utilization of a more stable primary care clinic setting by patients with sickle cell disease.

Stable transformation of a mosquito cell line results in extraordinarily high copy numbers of the plasmid.

PubMed Central

Monroe, T J; Muhlmann-Diaz, M C; Kovach, M J; Carlson, J O; Bedford, J S; Beaty, B J

1992-01-01

Stable incorporation of high copy numbers (greater than 10,000 per cell) of a plasmid vector containing a gene conferring resistance to the antibiotic hygromycin was achieved in a cell line derived from the Aedes albopictus mosquito. Plasmid sequences were readily observed by ethidium bromide staining of cellular DNA after restriction endonuclease digestion and agarose gel electrophoresis. The plasmid was demonstrated by in situ hybridization to be present in large arrays integrated in metaphase chromosomes and in minute and double-minute replicating elements. In one subclone, approximately 60,000 copies of the plasmid were organized in a large array that resembles a chromosome, morphologically and in the segregation of its chromatids during anaphase. The original as well as modified versions of the plasmid were rescued by transformation of Escherichia coli using total cellular DNA. Southern blot analyses of recovered plasmids indicate the presence of mosquito-derived sequences. Images PMID:1631052

Platelet collection efficiencies of three different platelet-rich plasma preparation systems.

PubMed

Aydin, Fatma; Pancar Yuksel, Esra; Albayrak, Davut

2015-06-01

Different systems have been used for the preparation of platelet-rich plasma (PRP), but platelet collection efficiencies of these systems are not clear. To evaluate the platelet collection efficiencies of three different PRP preparation systems. Blood samples were obtained from the same 16 volunteers for each system. The samples were centrifuged and PRP was prepared by three systems. The ratio of the total number of platelets in PRP to the total number of platelets of the venous blood sample of the patient expressed in percentage was named as platelet collection efficiency and calculated for each system. Mean platelet collection efficiencies were 66.6 (min: 56.9, max: 76.9), 58.3 (min: 27.3, max: 102.8), 50.8 (min: 27.2, max: 73) for top and bottom bag system, system using citrated tube, and the system using tube with Ficoll and cell extraction kit, respectively. Statistically significant difference was found only between the platelet collection efficiencies of systems using the tube with ficoll and cell extraction kit and the top and bottom bag system (p = 0.002). All three systems could be used for PRP preparation, but top and bottom bag system offers a slight advantage over the system using Ficoll and cell extraction kit regarding the platelet collection efficiency.

Excluding Anti-cytomegalovirus Immunoglobulin M-Positive Cord Blood Units Has a Minimal Impact on the Korean Public Cord Blood Bank Inventory

PubMed Central

Shin, Sue; Roh, Eun Youn; Oh, Sohee; Song, Eun Young; Kim, Eui Chong; Yoon, Jong Hyun

2017-01-01

Cord blood units (CBUs) for transplantation should be free of communicable disease and must contain a specific amount of total nucleated cells and CD34+ cells. Although posttransplantation cytomegalovirus (CMV) infections are from latent infection in patients, ensuring CMV-free CBUs by performing CMV-specific IgM and nucleic acid amplification testing (NAT) is one of the mandatory procedures for the safety of CBUs. However, the exclusion policies (based on these test results) vary among nations and institutions. We tested 28,000 processed CBUs between May 2006 and June 2014. The cord blood leukocytes from CMV IgM-positive samples were then subjected to NAT. The total nucleated cell and CD34+ cell counts were measured for each CBU, and the results were compared to the CMV IgM and IgG results. The seroprevalence of CMV among pregnant women was 98.1% (18,459/18,818) for IgG and 1.7% (441/25,293) for IgM. The concentration and the total number of CD34+ cells were significantly higher in CBUs from IgM-negative mothers compared to those from IgM-positive mothers (72.4/μl vs. 57.2/μl, respectively, p < 0.0001; 1.45 × 106/unit vs. 1.15 × 106/unit, respectively, p < 0.0001). Among CBUs with positive CMV IgM in their mothers' plasma or cord blood plasma, only 0.58% of the samples (3/517) had a positive NAT. The number of excluded CBUs from inventory due to positive CMV IgM in the cord blood was 54 of 18,326 (0.3%). For inventory purposes, it is appropriate to remove CBUs with positive cord blood CMV IgM findings irrespective of the NAT status as well as positive maternal CMV IgM in South Korea. PMID:27524276

Excluding Anti-cytomegalovirus Immunoglobulin M-Positive Cord Blood Units Has a Minimal Impact on the Korean Public Cord Blood Bank Inventory.

PubMed

Shin, Sue; Roh, Eun Youn; Oh, Sohee; Song, Eun Young; Kim, Eui Chong; Yoon, Jong Hyun

2017-01-24

Cord blood units (CBUs) for transplantation should be free of communicable disease and must contain a specific amount of total nucleated cells and CD34+ cells. Although posttransplantation cytomegalovirus (CMV) infections are from latent infection in patients, ensuring CMV-free CBUs by performing CMV-specific IgM and nucleic acid amplification testing (NAT) is one of the mandatory procedures for the safety of CBUs. However, the exclusion policies (based on these test results) vary among nations and institutions. We tested 28,000 processed CBUs between May 2006 and June 2014. The cord blood leukocytes from CMV IgM-positive samples were then subjected to NAT. The total nucleated cell and CD34+ cell counts were measured for each CBU, and the results were compared to the CMV IgM and IgG results. The seroprevalence of CMV among pregnant women was 98.1% (18,459/18,818) for IgG and 1.7% (441/25,293) for IgM. The concentration and the total number of CD34+ cells were significantly higher in CBUs from IgM-negative mothers compared to those from IgM-positive mothers (72.4/μl vs. 57.2/μl, respectively, p < 0.0001; 1.45 × 106/unit vs. 1.15 × 106/unit, respectively, p < 0.0001). Among CBUs with positive CMV IgM in their mothers' plasma or cord blood plasma, only 0.58% of the samples (3/517) had a positive NAT. The number of excluded CBUs from inventory due to positive CMV IgM in the cord blood was 54 of 18,326 (0.3%). For inventory purposes, it is appropriate to remove CBUs with positive cord blood CMV IgM findings irrespective of the NAT status as well as positive maternal CMV IgM in South Korea.

Effect of timing of development on total cell number and expression profile of HSP-70.1 and GLUT-1 in buffalo (Bubalus bubalis) oocytes and preimplantation embryos produced in vitro.

PubMed

Rajhans, Rajib; Kumar, G Sai; Dubey, Pawan K; Sharma, G Taru

2010-03-29

The present study was designed to compare the expression profile of two developmentally important genes (HSP-70.1 and GLUT-1) and TCN (total cell number) count in fast (group A) and slow (group B) cleaved buffalo embryos to access their in vitro developmental competence. Buffalo COCs (cumulus oocyte complexes) were collected from local abattoir ovaries and subjected to in vitro maturation in: TCM-199 supplemented with 10% FBS (fetal bovine serum), BSA (3 mg/ml), sodium pyruvate (0.25 mM) and 20 ng/ml EGF (epidermal growth factor) at 38.5 degrees C under 5% CO2. In vitro derived embryos were collected at 4-8, 8-16 cell, morula and blastocyst stages at specific time points for gene expression analysis and total cell count. A semiquantitative RT-PCR (reverse transcriptase-PCR) assay was used to determine the HSP-70.1 and GLUT-1 transcripts. Results showed that developmental competence and TCN count in fast (group A)-cleaving embryos was significantly (P<0.05) higher than in the slow group (group B). The gene transcript of HSP-70.1 and GLUT-1 was expressed in oocytes (immature and mature) and throughout the embryonic developmental stages in the fast group (group A), while in the slow (group B) cleaving embryos, the expression of HSP-70.1 was absent in all the embryonic developmental stages, and expression of GLUT-1 was absent after 8-16 cell stage. In conclusion, TCN count and expression profile of HSP-70.1 and GLUT-1 genes in buffalo embryos are different taking into account the cleavage rate. Quality of such embryos for research purposes, TCN and expression profiling of developmentally important genes should be employed to optimize the in vitro culture system to produce superior quality of embryos.

Flow analysis of human chromosome sets by means of mixing-stirring device

NASA Astrophysics Data System (ADS)

Zenin, Valeri V.; Aksenov, Nicolay D.; Shatrova, Alla N.; Klopov, Nicolay V.; Cram, L. Scott; Poletaev, Andrey I.

1997-05-01

A new mixing and stirring device (MSD) was used to perform flow karyotype analysis of single human mitotic chromosomes analyzed so as to maintain the identity of chromosomes derived from the same cell. An improved method for cell preparation and intracellular staining of chromosomes was developed. The method includes enzyme treatment, incubation with saponin and separation of prestained cells from debris on a sucrose gradient. Mitotic cells are injected one by one in the MSD which is located inside the flow chamber where cells are ruptured, thereby releasing chromosomes. The set of chromosomes proceeds to flow in single file fashion to the point of analysis. The device works in a stepwise manner. The concentration of cells in the sample must be kept low to ensure that only one cell at a time enters the breaking chamber. Time-gated accumulation of data in listmode files makes it possible to separate chromosome sets comprising of single cells. The software that was developed classifies chromosome sets according to different criteria: total number of chromosomes, overall DNA content in the set, and the number of chromosomes of certain types. This approach combines the high performance of flow cytometry with the advantages of image analysis. Examples obtained with different human cell lines are presented.

Detection of cell proliferation in adults of the water bear Hypsibius dujardini (Tardigrada) via incorporation of a thymidine analog.

PubMed

Gross, V; Bährle, R; Mayer, G

2018-04-01

The taxon Tardigrada, commonly called "water bears", consists of microscopic, eight-legged invertebrates that are well known for their ability to tolerate extreme environmental conditions. Their miniscule body size means that tardigrades possess a small total number of cells, the number and arrangement of which may be highly conserved in some organs. Although mitoses have been observed in several organs, the rate and pattern of cell divisions in adult tardigrades has never been characterized. In this study, we incubated live tardigrades over a period of several days with a thymidine analog in order to visualize all cells that had divided during this time. We focus on the midgut, the largest part of the digestive system. Our results show that new cells in the midgut arise from the anterior and posterior ends of this organ and either migrate or divide toward its middle. These cells divide at a constant rate and all cells of the midgut epithelium are replaced in approximately one week. On the other hand, we found no cell divisions in the nervous system or any other major organs, suggesting that the cell turnover of these organs may be extremely slow or dependent on changing environmental conditions. Copyright © 2018 Elsevier Ltd. All rights reserved.

Biochemical changes to fibroblast cells subjected to ionizing radiation.

PubMed

Jones, Pamala; Benghuzzi, Hamed; Tucci, Michelle; Richards, Latoya; Harrison, George; Patel, Ramesh

2008-01-01

High energy X-rays are capable of interacting with biological membranes to cause both functional and structural modifications. The goal of the present study was to investigate the effects human fibroblast cells exposed multiple times to 10 Gy over time. Following exposures of 2, 3, or 4 times to 10 Gy/10min the cells were evaluated for cell number changes, membrane damage, and intracellular glutathione content after 24, 48 and 72 hours. Twenty-four hours following exposure the cell numbers were reduced and increased levels of cellular membrane damage was evident. This trend was observed for the duration of the study. Interestingly, there was not an exposure dependent increase in cell damage or cell loss with time. Intracellular antioxidant systems were activated as indicated by anincrease in total cellular glutathione content. Additional studies are needed to determine if the cellular reduction is caused by a direct effect of the X-rays targeting the DNA or an indirect effect of the X-ray targeting the cellular membrane, which then generates radicals that target cell cycle checkpoints or DNA damage. In conclusion, fibroblast cells can be used to determine early and late events of cellular function following exposure to harmful levels of radiation exposure and results of exposure can be seen within twenty four hours.

Synergistic Integration of Mesenchymal Stem Cells and Hydrostatic Pressure in the Expansion and Maintenance of Human Hematopoietic/Progenitor Cells

PubMed Central

2018-01-01

Ex vivo expansion of hematopoietic stem/progenitor cell (HSPC) has been investigated to improve the clinical outcome of HSPC transplantation. However, ex vivo expansion of HSPCs still faces a major obstacle in that HPSCs tend to differentiate when proliferating. Here, we cocultured HSPCs with mesenchymal stem cells (MSCs) and divided the HSPCs into two fractions according to whether they came into adherent to MSCs or not. Additionally, we used hydrostatic pressure (HP) to mimic the physical conditions in vivo. Even nonadherent cells expanded to yield a significantly larger number of total nucleated cells (TNCs), adherent cells maintained the HSPC phenotype (CD34+, CD34+CD38−, and CD133+CD38−) to a greater extent than nonadherent cells and had superior clonogenic potential. Moreover, applying HP significantly increased the number of TNCs, the frequency of the immature HSPC phenotype, and the clonogenic potential. Furthermore, the genetic markers for the HSPC niche were significantly increased under HP. Our data suggest that the nonadherent fraction is the predominant site of HSPC expansion, whereas the adherent fraction seems to mimic the HSPC niche for immature cells. Moreover, HP has a synergistic effect on expansion and functional maintenance. This first study utilizing HP has a potential of designing clinically applicable expansion systems. PMID:29681947

A Division-Dependent Compartmental Model for Computing Cell Numbers in CFSE-based Lymphocyte Proliferation Assays

DTIC Science & Technology

2012-02-12

is the total number of data points, is an approximately unbiased estimate of the “expected relative Kullback - Leibler distance” ( information loss...possible models). Thus, after each model from Table 2 is fit to a data set, we can compute the Akaike weights for the set of candidate models and use ...computed from the OLS best- fit model solution (top), from a deconvolution of the data using normal curves (middle) and from a deconvolution of the data

k-Space Image Correlation Spectroscopy: A Method for Accurate Transport Measurements Independent of Fluorophore Photophysics

PubMed Central

Kolin, David L.; Ronis, David; Wiseman, Paul W.

2006-01-01

We present the theory and application of reciprocal space image correlation spectroscopy (kICS). This technique measures the number density, diffusion coefficient, and velocity of fluorescently labeled macromolecules in a cell membrane imaged on a confocal, two-photon, or total internal reflection fluorescence microscope. In contrast to r-space correlation techniques, we show kICS can recover accurate dynamics even in the presence of complex fluorophore photobleaching and/or “blinking”. Furthermore, these quantities can be calculated without nonlinear curve fitting, or any knowledge of the beam radius of the exciting laser. The number densities calculated by kICS are less sensitive to spatial inhomogeneity of the fluorophore distribution than densities measured using image correlation spectroscopy. We use simulations as a proof-of-principle to show that number densities and transport coefficients can be extracted using this technique. We present calibration measurements with fluorescent microspheres imaged on a confocal microscope, which recover Stokes-Einstein diffusion coefficients, and flow velocities that agree with single particle tracking measurements. We also show the application of kICS to measurements of the transport dynamics of α5-integrin/enhanced green fluorescent protein constructs in a transfected CHO cell imaged on a total internal reflection fluorescence microscope using charge-coupled device area detection. PMID:16861272

Effect of Cypermethrin on the Growth of Ciliate Protozoan Paramecium caudatum.

PubMed

Dutta, Joydeep

2015-01-01

The objective of this study is to assess the effect of cypermethrin on the growth of ciliate protozoan Paramecium caudatum. Monoxenic culture of P. caudatum, were exposed to different doses (0.01, 0.05, 0.1, 0.15, and 0.2 µg/L) of cypermethrin along with control for 24, 48, 72, and 96 h time interval. The total numbers of live and dead cells were counted after trypan blue staining in Neubauer hemocytometer. Marked decrease in the number of living cells with the increase in the concentration of cypermethrin and with increasing exposure time intervals was recorded. The results indicate that cypermethrin is toxic to P. caudatum even at low concentrations when it enters in the aquatic system through runoff.

Myostatin inhibition induces muscle fibre hypertrophy prior to satellite cell activation.

PubMed

Wang, Qian; McPherron, Alexandra C

2012-05-01

Muscle fibres are multinucleated post-mitotic cells that can change dramatically in size during adulthood. It has been debated whether muscle fibre hypertrophy requires activation and fusion of muscle stem cells, the satellite cells. Myostatin (MSTN) is a negative regulator of skeletal muscle growth during development and in the adult, and MSTN inhibition is therefore a potential therapy for muscle wasting diseases, some of which are associated with a depletion of satellite cells. Conflicting results have been obtained in previous analyses of the role of MSTN on satellite cell quiescence. Here, we inhibited MSTN in adult mice with a soluble activin receptor type IIB and analysed the incorporation of new nuclei using 5-bromo-2-deoxyuridine (BrdU) labelling by isolating individual myofibres. We found that satellite cells are activated by MSTN inhibition. By varying the dose and time course for MSTN inhibition, however, we found that myofibre hypertrophy precedes the incorporation of new nuclei, and that the overall number of new nuclei is relatively low compared to the number of total myonuclei. These results reconcile some of the previous work obtained by other methods. In contrast with previous reports, we also found that Mstn null mice do not have increased satellite cell numbers during adulthood and are not resistant to sarcopaenia. Our results support a previously proposed model of hypertrophy in which hypertrophy can precede satellite cell activation. Studies of the metabolic and functional effects of postnatal MSTN inhibition are needed to determine the consequences of increasing the cytoplasm/myonuclear ratio after MSTN inhibition.

Retinal ganglion cell topography and spatial resolving power in penguins.

PubMed

Coimbra, João Paulo; Nolan, Paul M; Collin, Shaun P; Hart, Nathan S

2012-01-01

Penguins are a group of flightless seabirds that exhibit numerous morphological, behavioral and ecological adaptations to their amphibious lifestyle, but little is known about the topographic organization of neurons in their retinas. In this study, we used retinal wholemounts and stereological methods to estimate the total number and topographic distribution of retinal ganglion cells in addition to an anatomical estimate of spatial resolving power in two species of penguins: the little penguin, Eudyptula minor, and the king penguin, Aptenodytes patagonicus. The total number of ganglion cells per retina was approximately 1,200,000 in the little penguin and 1,110,000 in the king penguin. The topographic distribution of retinal ganglion cells in both species revealed the presence of a prominent horizontal visual streak with steeper gradients in the little penguin. The little penguin retinas showed ganglion cell density peaks of 21,867 cells/mm², affording spatial resolution in water of 17.07-17.46 cycles/degree (12.81-13.09 cycles/degree in air). In contrast, the king penguin showed a relatively lower peak density of ganglion cells of 14,222 cells/mm², but--due to its larger eye--slightly higher spatial resolution in water of 20.40 cycles/degree (15.30 cycles/degree in air). In addition, we mapped the distribution of giant ganglion cells in both penguin species using Nissl-stained wholemounts. In both species, topographic mapping of this cell type revealed the presence of an area gigantocellularis with a concentric organization of isodensity contours showing a peak in the far temporal retina of approximately 70 cells/mm² in the little penguin and 39 cells/mm² in the king penguin. Giant ganglion cell densities gradually fall towards the outermost isodensity contours revealing the presence of a vertically organized streak. In the little penguin, we confirmed our cytological characterization of giant ganglion cells using immunohistochemistry for microtubule-associated protein 2. This suite of retinal specializations, which is also observed in the closely related procellariiform seabirds, affords the eyes of the little and king penguins panoramic surveillance of the horizon and motion detection in the frontal visual field. Copyright © 2012 S. Karger AG, Basel.

Cytotoxicity control of silicon nanoparticles by biopolymer coating and ultrasound irradiation for cancer theranostic applications

NASA Astrophysics Data System (ADS)

Sviridov, A. P.; Osminkina, L. A.; Kharin, A. Yu; Gongalsky, M. B.; Kargina, J. V.; Kudryavtsev, A. A.; Bezsudnova, Yu I.; Perova, T. S.; Geloen, A.; Lysenko, V.; Timoshenko, V. Yu

2017-03-01

Silicon nanoparticles (SiNPs) prepared by mechanical grinding of luminescent porous silicon were coated with a biopolymer (dextran) and investigated as a potential theranostic agent for bioimaging and sonodynamic therapy. Transmission electron microscopy, photoluminescence and Raman scattering measurements of dextran-coated SiNPs gave evidence of their enhanced stability in water. In vitro experiments confirmed the lower cytotoxicity of the dextran-coated NPs in comparison with uncoated ones, especially for high concentrations of about 2 mg ml-1. Efficient uptake of the NPs by cancer cells was found using bioimaging in the optical transmittance and photoluminescence modes. Treatment of the cells with uptaken SiNPs by therapeutic ultrasound for 5-20 min resulted in a strong decrease in the number of living cells, while the total number of cells remained nearly unchanged. The obtained data indicate a ‘mild’ effect of the combined action of ultrasonic irradiation and SiNPs on cancer cells. The observed results reveal new opportunities for controlling the photoluminescent and sonosensitizing properties of silicon-based NPs for applications in the diagnostics and mild therapy of cancer.

Cytotoxicity of dimethyl sulfoxide (DMSO) in direct contact with odontoblast-like cells.

PubMed

Hebling, J; Bianchi, L; Basso, F G; Scheffel, D L; Soares, D G; Carrilho, M R O; Pashley, D H; Tjäderhane, L; de Souza Costa, C A

2015-04-01

To evaluate the cytotoxicity of dimethyl sulfoxide (DMSO) on the repair-related activity of cultured odontoblast-like MDPC-23 cells. Solutions with different concentrations of DMSO (0.05, 0.1, 0.3, 0.5 and 1.0 mM), diluted in culture medium (DMEM), were placed in contact with MDPC-23 cells (5 × 104 cells/cm(2)) for 24 h. Eight replicates (n = 8) were prepared for each solutions for the following methods of analysis: violet crystal dye for cell adhesion (CA), quantification of total protein (TP), alizarin red for mineralization nodules formation (MN) and cell death by necrosis (flow cytometry); while twelve replicates (n = 12) were prepared for viable cell number (Trypan Blue) and cell viability (MTT assay). Data were analyzed by ANOVA and Tukey or Kruskal-Wallis and Mann-Whitney's tests (p < 0.05). Cell viability, adhesion and percentage of cell death by necrosis were not affected by DMSO at any concentration, with no statistical significant difference among the groups. A significant reduction in total protein production was observed for 0.5 and 1.0 mM of DMSO compared to the control while increased mineralized nodules formation was seen only for 1.0 mM DMSO. DMSO caused no or minor cytotoxic effects on the pulp tissue repair-related activity of odontoblast-like cells. Copyright © 2015 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

Structural Complexity of Non-acid Glycosphingolipids in Human Embryonic Stem Cells Grown under Feeder-free Conditions*

PubMed Central

Barone, Angela; Benktander, John; Ångström, Jonas; Aspegren, Anders; Björquist, Petter; Teneberg, Susann; Breimer, Michael. E.

2013-01-01

Due to their pluripotency and growth capability, there are great expectations for human embryonic stem cells, both as a resource for functional studies of early human development and as a renewable source of cells for use in regenerative medicine and transplantation. However, to bring human embryonic stem cells into clinical applications, their cell surface antigen expression and its chemical structural complexity have to be defined. In the present study, total non-acid glycosphingolipid fractions were isolated from two human embryonic stem cell lines (SA121 and SA181) originating from leftover in vitro fertilized human embryos, using large amounts of starting material (1 × 109 cells/cell line). The total non-acid glycosphingolipid fractions were characterized by antibody and lectin binding, mass spectrometry, and proton NMR. In addition to the globo-series and type 1 core chain glycosphingolipids previously described in human embryonic stem cells, a number of type 2 core chain glycosphingolipids (neo-lactotetraosylceramide, the H type 2 pentaosylceramide, the Lex pentaosylceramide, and the Ley hexaosylceramide) were identified as well as the blood group A type 1 hexaosylceramide. Finally, the mono-, di-, and triglycosylceramides were characterized as galactosylceramide, glucosylceramide, lactosylceramide, galabiaosylceramide, globotriaosylceramide, and lactotriaosylceramide. Thus, the glycan diversity of human embryonic stem cells, including cell surface immune determinants, is more complex than previously appreciated. PMID:23404501

Picking Cell Lines for High-Throughput Transcriptomic Toxicity ...

EPA Pesticide Factsheets

High throughput, whole genome transcriptomic profiling is a promising approach to comprehensively evaluate chemicals for potential biological effects. To be useful for in vitro toxicity screening, gene expression must be quantified in a set of representative cell types that captures the diversity of potential responses across chemicals. The ideal dataset to select these cell types would consist of hundreds of cell types treated with thousands of chemicals, but does not yet exist. However, basal gene expression data may be useful as a surrogate for representing the relevant biological space necessary for cell type selection. The goal of this study was to identify a small (< 20) number of cell types that capture a large, quantifiable fraction of basal gene expression diversity. Three publicly available collections of Affymetrix U133+2.0 cellular gene expression data were used: 1) 59 cell lines from the NCI60 set; 2) 303 primary cell types from the Mabbott et al (2013) expression atlas; and 3) 1036 cell lines from the Cancer Cell Line Encyclopedia. The data were RMA normalized, log-transformed, and the probe sets mapped to HUGO gene identifiers. The results showed that <20 cell lines capture only a small fraction of the total diversity in basal gene expression when evaluated using either the entire set of 20960 HUGO genes or a subset of druggable genes likely to be chemical targets. The fraction of the total gene expression variation explained was consistent when

Retinal ganglion cells in the eastern newt Notophthalmus viridescens: topography, morphology, and diversity.

PubMed

Pushchin, Igor I; Karetin, Yuriy A

2009-10-20

The topography and morphology of retinal ganglion cells (RGCs) in the eastern newt were studied. Cells were retrogradely labeled with tetramethylrhodamine-conjugated dextran amines or horseradish peroxidase and examined in retinal wholemounts. Their total number was 18,025 +/- 3,602 (mean +/- SEM). The spatial density of RGCs varied from 2,100 cells/mm(2) in the retinal periphery to 4,500 cells/mm(2) in the dorsotemporal retina. No prominent retinal specializations were found. The spatial resolution estimated from the spatial density of RGCs varied from 1.4 cycles per degree in the periphery to 1.95 cycles per degree in the region of the peak RGC density. A sample of 68 cells was camera lucida drawn and subjected to quantitative analysis. A total of 21 parameters related to RGC morphology and stratification in the retina were estimated. Partitionings obtained by using different clustering algorithms combined with automatic variable weighting and dimensionality reduction techniques were compared, and an effective solution was found by using silhouette analysis. A total of seven clusters were identified and associated with potential cell types. Kruskal-Wallis ANOVA-on-Ranks with post hoc Mann-Whitney U tests showed significant pairwise between-cluster differences in one or more of the clustering variables. The average silhouette values of the clusters were reasonably high, ranging from 0.52 to 0.79. Cells assigned to the same cluster displayed similar morphology and stratification in the retina. The advantages and limitations of the methodology adopted are discussed. The present classification is compared with known morphological and physiological RGC classifications in other salamanders.

Improvement of Cell Survival During Human Pluripotent Stem Cell Definitive Endoderm Differentiation

PubMed Central

Wang, Han; Luo, Xie; Yao, Li; Lehman, Donna M.

2015-01-01

Definitive endoderm (DE) is a vital precursor for internal organs such as liver and pancreas. Efficient protocol to differentiate human embryonic stem cells (hESCs) or induced pluripotent stem cells (iPSCs) to DE is essential for regenerative medicine and for modeling diseases; yet, poor cell survival during DE differentiation remains unsolved. In this study, our use of B27 supplement in modified differentiation protocols has led to a substantial improvement. We used an SOX17-enhanced green fluorescent protein (eGFP) reporter hESC line to compare and modify established DE differentiation protocols. Both total live cell numbers and the percentages of eGFP-positive cells were used to assess differentiation efficiency. Among tested protocols, three modified protocols with serum-free B27 supplement were developed to generate a high number of DE cells. Massive cell death was avoided during DE differentiation and the percentage of DE cells remained high. When the resulting DE cells were further differentiated toward the pancreatic lineage, the expression of pancreatic-specific markers was significantly increased. Similar high DE differentiation efficiency was observed in H1 hESCs and iPSCs through the modified protocols. In B27 components, bovine serum albumin was found to facilitate DE differentiation and cell survival. Using our modified DE differentiation protocols, satisfactory quantities of quality DE can be produced as primary material for further endoderm lineage differentiation. PMID:26132288

Change in the Content of Immunoproteasomes and Macrophages in Rat Liver At the Induction of Donor-Specific Tolerance.

PubMed

Karpova, Ya D; Ustichenko, V D; Alabedal'karim, N M; Stepanova, A A; Lyupina, Yu V; Boguslavski, K I; Bozhok, G A; Sharova, N P

2017-01-01

Induction of donor specific tolerance (DST) by the introduction of donor cells into a recipient's portal vein is one of the approaches used to solve the problem of transplant engraftment. However, the mechanism of DST development remains unclear to this moment. In the present work, we first studied the change in the content of immunoproteasomes and macrophages of the liver at early stages of the development of allospecific portal tolerance in rats by Western blotting and flow cytofluorimetry. On the basis of the data obtained, we can conclude that the induction of DST is an active process characterized by two phases during which the level of the proteasome immune subunits LMP2 and LMP7 in liver mononuclear cells, including Kupffer cells, and the number of Kupffer cells change. The first phase lasts up to 5 days after the beginning of DST induction; the second phase - from 5 to 14 days. In both phases, the level of the subunits LMP2 and LMP7 in the total pool of mononuclear cells and Kupffer cells increases, with maximum values on days 1 and 7. In addition, the total number of Kupffer cells increases in both phases with a shift in several days. The most noticeable changes take place in the second phase. The third day is characterized by a lower content of mononuclear cells expressing immunoproteasomes compared to the control value in native animals. Presumably, at this time point a "window of opportunity" appears for subsequent filling of an empty niche with cells of different subpopulations and, depending on this fact, the development of tolerance or rejection. The results obtained raise the new tasks of finding ways to influence the cellular composition in the liver and the expression of immunoproteasomes on the third day after the beginning of DST induction to block the development of rejection.

Glioma Invasiveness Responds Variably to Irradiation in a Co-Culture Model

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

Nakamura, Jean L.; Haas-Kogan, Daphne A.; Department of Neurological Surgery, University of California-San Francisco, San Francisco, CA

2007-11-01

Purpose: We developed a co-culture system to quantitate the growth and invasion of human malignant gliomas into a background of confluent normal human astrocytes, then used this assay to assess independently the effects of irradiating both cell types on glioma invasion. Methods and Materials: Enhanced green fluorescent protein (EGFP)-labeled immortalized human astrocytes, human malignant glioma cells, or transformed human astrocytes were focally plated onto a confluent layer of normal human astrocytes, and the invasiveness of EGFP-labeled cells was scored after 96 h. To address the consequences of irradiation on glioma invasion, the invasiveness of irradiated glioma cell lines and irradiatedmore » astrocytic backgrounds was assessed. Fluorescence-activated cell sorting was used to quantitate the total number of EGFP-labeled cells. Results: Growth in the co-culture assay consistently reflected transformation states of the plated cells. Immortalized, but untransformed human astrocytes failed even to establish growth on confluent normal human astrocytes. In contrast, all malignant human glioma cell lines and transformed human astrocytes demonstrated various degrees of infiltration into the astrocytic bed. Irradiation failed to alter the invasiveness of U87, A172, and U373. A 1-Gy dose slightly reduced the invasiveness of U251 MG by 75% (p < 0.05 by one-way analysis of variance and post hoc Neuman-Keuls), without reducing total cell numbers. Independently irradiating the human astrocytic bed did not alter the invasiveness of nonirradiated U251, whereas the matrix metalloproteinase (MMP) inhibitor GM6001 reduced U251 invasiveness in the co-culture assay. Conclusions: Growth in the co-culture assay reflects the transformation status and provides a useful in vitro model for assessing invasiveness. Human glioma invasiveness in the co-culture model responds variably to single low-dose fractions. MMP activity promotes invasiveness in the co-culture model. Reduced invasiveness in irradiated U251 appears to be mediated by MMP-independent mechanisms.« less

Early Virological and Immunological Events in Asymptomatic Epstein-Barr Virus Infection in African Children

PubMed Central

Jayasooriya, Shamanthi; de Silva, Thushan I.; Njie-jobe, Jainaba; Sanyang, Chilel; Leese, Alison M.; Bell, Andrew I.; McAulay, Karen A.; Yanchun, Peng; Long, Heather M.; Dong, Tao; Whittle, Hilton C.; Rickinson, Alan B.; Rowland-Jones, Sarah L.; Hislop, Andrew D.; Flanagan, Katie L.

2015-01-01

Epstein-Barr virus (EBV) infection often occurs in early childhood and is asymptomatic. However, if delayed until adolescence, primary infection may manifest as acute infectious mononucleosis (AIM), a febrile illness characterised by global CD8+ T-cell lymphocytosis, much of it reflecting a huge expansion of activated EBV-specific CD8+ T-cells. While the events of AIM have been intensely studied, little is known about how these relate to asymptomatic primary infection. Here Gambian children (14–18 months old, an age at which many acquire the virus) were followed for the ensuing six months, monitoring circulating EBV loads, antibody status against virus capsid antigen (VCA) and both total and virus-specific CD8+ T-cell numbers. Many children were IgG anti-VCA-positive and, though no longer IgM-positive, still retained high virus loads comparable to AIM patients and had detectable EBV-specific T-cells, some still expressing activation markers. Virus loads and the frequency/activation status of specific T-cells decreased over time, consistent with resolution of a relatively recent primary infection. Six children with similarly high EBV loads were IgM anti-VCA-positive, indicating very recent infection. In three of these donors with HLA types allowing MHC-tetramer analysis, highly activated EBV-specific T-cells were detectable in the blood with one individual epitope response reaching 15% of all CD8+ T-cells. That response was culled and the cells lost activation markers over time, just as seen in AIM. However, unlike AIM, these events occurred without marked expansion of total CD8+ numbers. Thus asymptomatic EBV infection in children elicits a virus-specific CD8+ T-cell response that can control the infection without over-expansion; conversely, in AIM it appears the CD8 over-expansion, rather than virus load per se, is the cause of disease symptoms. PMID:25816224

Pulmonary neutrophil recruitment and bronchial reactivity in formaldehyde-exposed rats are modulated by mast cells and differentially by neuropeptides and nitric oxide

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

Lino dos Santos Franco, Adriana; Damazo, Amilcar Sabino; Post-Graduation in Morphology, UNIFESP, EPM, Sao Paulo

2006-07-01

We have used a pharmacological approach to study the mechanisms underlying the rat lung injury and the airway reactivity changes induced by inhalation of formaldehyde (FA) (1% formalin solution, 90 min once a day, 4 days). The reactivity of isolated tracheae and intrapulmonary bronchi were assessed in dose-response curves to methacholine (MCh). Local and systemic inflammatory phenomena were evaluated in terms of leukocyte countings in bronchoalveolar lavage (BAL) fluid, blood, bone marrow lavage and spleen. Whereas the tracheal reactivity to MCh did not change, a significant bronchial hyporesponsiveness (BHR) was found after FA inhalation as compared with naive rats. Also,more » FA exposure significantly increased the total cell numbers in BAL, in peripheral blood and in the spleen, but did not modify the counts in bone marrow. Capsaicin hindered the increase of leukocyte number recovered in BAL fluid after FA exposure. Both compound 48/80 and indomethacin were able to prevent the lung neutrophil influx after FA, but indomethacin had no effect on that of mononuclear cells. Following FA inhalation, the treatment with sodium cromoglycate (SCG), but not with the nitric oxide (NO) synthase inhibitor L-NAME, significantly reduced the total cell number in BAL. Compound 48/80, L-NAME and SCG significantly prevented BHR to MCh after FA inhalation, whereas capsaicin was inactive in this regard. On the other hand, indomethacin exacerbated BHR. These data suggest that after FA inhalation, the resulting lung leukocyte influx and BHR may involve nitric oxide, airway sensory fibers and mast cell-derived mediators. The effect of NO seemed to be largely restricted to the bronchial tonus, whereas neuropeptides appeared to be linked to the inflammatory response, therefore indicating that the mechanisms responsible for the changes of airway responsiveness caused by FA may be separate from those underlying its inflammatory lung effects.« less

Stereology techniques in radiation biology

NASA Technical Reports Server (NTRS)

Kubinova, Lucie; Mao, XiaoWen; Janacek, Jiri; Archambeau, John O.; Nelson, G. A. (Principal Investigator)

2003-01-01

Clinicians involved in conventional radiation therapy are very concerned about the dose-response relationships of normal tissues. Before proceeding to new clinical protocols, radiation biologists involved with conformal proton therapy believe it is necessary to quantify the dose response and tolerance of the organs and tissues that will be irradiated. An important focus is on the vasculature. This presentation reviews the methodology and format of using confocal microscopy and stereological methods to quantify tissue parameters, cell number, tissue volume and surface area, and vessel length using the microvasculature as a model tissue. Stereological methods and their concepts are illustrated using an ongoing study of the dose response of the microvessels in proton-irradiated hemibrain. Methods for estimating the volume of the brain and the brain cortex, the total number of endothelial cells in cortical microvessels, the length of cortical microvessels, and the total surface area of cortical microvessel walls are presented step by step in a way understandable for readers with little mathematical background. It is shown that stereological techniques, based on a sound theoretical basis, are powerful and reliable and have been used successfully.

Design optimization of large-size format edge-lit light guide units

NASA Astrophysics Data System (ADS)

Hastanin, J.; Lenaerts, C.; Fleury-Frenette, K.

2016-04-01

In this paper, we present an original method of dot pattern generation dedicated to large-size format light guide plate (LGP) design optimization, such as photo-bioreactors, the number of dots greatly exceeds the maximum allowable number of optical objects supported by most common ray-tracing software. In the proposed method, in order to simplify the computational problem, the original optical system is replaced by an equivalent one. Accordingly, an original dot pattern is splitted into multiple small sections, inside which the dot size variation is less than the ink dots printing typical resolution. Then, these sections are replaced by equivalent cells with continuous diffusing film. After that, we adjust the TIS (Total Integrated Scatter) two-dimensional distribution over the grid of equivalent cells, using an iterative optimization procedure. Finally, the obtained optimal TIS distribution is converted into the dot size distribution by applying an appropriate conversion rule. An original semi-empirical equation dedicated to rectangular large-size LGPs is proposed for the initial guess of TIS distribution. It allows significantly reduce the total time needed to dot pattern optimization.

Calibrations between the variables of microbial TTI response and ground pork qualities.

PubMed

Kim, Eunji; Choi, Dong Yeol; Kim, Hyun Chul; Kim, Keehyuk; Lee, Seung Ju

2013-10-01

A time-temperature indicator (TTI) based on a lactic acid bacterium, Weissella cibaria CIFP009, was applied to ground pork packaging. Calibration curves between TTI response and pork qualities were obtained from storage tests at 2°C, 10°C, and 13°C. The curves of the TTI vs. total cell number at different temperatures coincided to the greatest extent, indicating the highest representativeness of calibration, by showing the least coefficient of variance (CV=11%) of the quality variables at a given TTI response (titratable acidity) on the curves, followed by pH (23%), volatile basic nitrogen (VBN) (25%), and thiobarbituric acid-reactive substances (TBARS) (47%). Similarity of Arrhenius activation energy (Ea) could also reflect the representativeness of calibration. The total cell number (104.9 kJ/mol) was found to be the most similar to that of the TTI response (106.2 kJ/mol), followed by pH (113.6 kJ/mol), VBN (77.4 kJ/mol), and TBARS (55.0 kJ/mol). Copyright © 2013 Elsevier Ltd. All rights reserved.

Study of oxygen gas production phenomenon during stand and discharge in silver-zinc batteries

NASA Technical Reports Server (NTRS)

1973-01-01

The effects of a number of cell process and performance variables upon the oxygen evolution rate of silver/silver oxide cathodes are studied to predict and measure the conditions which would result in the production of a minimum of oxygen. The following five tasks comprise the study: the design and fabrication of two pilot test cells to be used for electrode testing; the determination of the sensitivity and accuracy of the test cell; the determination of total volumes and rates of generation by cathodes of standard production procedures; the construction of a sequential test plan; and the construction of a series of positive formation cells in which formation process factors can be controlled.

Specific subpopulations of hypothalamic leptin receptor-expressing neurons mediate the effects of early developmental leptin receptor deletion on energy balance.

PubMed

Rupp, Alan C; Allison, Margaret B; Jones, Justin C; Patterson, Christa M; Faber, Chelsea L; Bozadjieva, Nadejda; Heisler, Lora K; Seeley, Randy J; Olson, David P; Myers, Martin G

2018-06-06

To date, early developmental ablation of leptin receptor (LepRb) expression from circumscribed populations of hypothalamic neurons (e.g., arcuate nucleus (ARC) Pomc- or Agrp-expressing cells) has only minimally affected energy balance. In contrast, removal of LepRb from at least two large populations (expressing vGat or Nos1) spanning multiple hypothalamic regions produced profound obesity and metabolic dysfunction. Thus, we tested the notion that the total number of leptin-responsive hypothalamic neurons (rather than specific subsets of cells with a particular molecular or anatomical signature) subjected to early LepRb deletion might determine energy balance. We generated new mouse lines deleted for LepRb in ARC Ghrh Cre neurons or in Htr2c Cre neurons (representing roughly half of all hypothalamic LepRb neurons, distributed across many nuclei). We compared the phenotypes of these mice to previously-reported models lacking LepRb in Pomc, Agrp, vGat or Nos1 cells. The early developmental deletion of LepRb from vGat or Nos1 neurons produced dramatic obesity, but deletion of LepRb from Pomc, Agrp, Ghrh, or Htr2c neurons minimally altered energy balance. Although early developmental deletion of LepRb from known populations of ARC neurons fails to substantially alter body weight, the minimal phenotype of mice lacking LepRb in Htr2c cells suggests that the phenotype that results from early developmental LepRb deficiency depends not simply upon the total number of leptin-responsive hypothalamic LepRb cells. Rather, specific populations of LepRb neurons must play particularly important roles in body energy homeostasis; these as yet unidentified LepRb cells likely reside in the DMH. Copyright © 2018 The Authors. Published by Elsevier GmbH.. All rights reserved.

Contrasting intra- and extracellular distribution of catalytic ferrous iron in ovalbumin-induced peritonitis.

PubMed

Ito, Fumiya; Nishiyama, Takahiro; Shi, Lei; Mori, Masahiko; Hirayama, Tasuku; Nagasawa, Hideko; Yasui, Hiroyuki; Toyokuni, Shinya

2016-08-05

Iron is an essential nutrient for every type of life on earth. However, excess iron is cytotoxic and can lead to an increased cancer risk in humans. Catalytic ferrous iron [Fe(II)] is an initiator of the Fenton reaction, which causes oxidative stress by generating hydroxyl radicals. Recently, it became possible to localize catalytic Fe(II) in situ with a turn-on fluorescent probe, RhoNox-1. Here, we screened each organ/cell of rats to globally evaluate the distribution of catalytic Fe(II) and found that eosinophils showed the highest abundance. In various cells, lysosomes were the major organelle, sharing ∼40-80% of RhoNox-1 fluorescence. We then used an ovalbumin-induced allergic peritonitis model to study the dynamics of catalytic Fe(II). Peritoneal lavage revealed that the total iron contents per cell were significantly decreased, whereas an increase in the number of inflammatory cells (macrophages, neutrophils, eosinophils and lymphocytes) resulted in an increased total iron content of the peritoneal inflammatory cells. Notably, macrophages, eosinophils and neutrophils exhibited significantly increased catalytic Fe(II) with increased DMT1 expression and decreased ferritin expression, though catalytic Fe(II) was significantly decreased in the peritoneal lavage fluid. In conclusion, catalytic Fe(II) in situ more directly reflects cellular activity and the accompanying pathology than total iron does. Copyright © 2016 Elsevier Inc. All rights reserved.

Allergen-specific transforming growth factor-β-producing CD19+CD5+ regulatory B-cell (Br3) responses in human late eczematous allergic reactions to cow's milk.

PubMed

Lee, Jae Ho; Noh, Joonyong; Noh, Geunwoong; Choi, Wahn Soo; Cho, Sunheui; Lee, Sang Sun

2011-05-01

CD19(+)CD5(+) regulatory B cells produce transforming growth factor β (TGF-β) in both mouse and human B-cell leukemias. In this study, TGF-β was uniquely produced by normal human regulatory B cells. TGF-β-producing regulatory B-cell (Br3) responses were characterized through allergic responses to cow's milk. In total, 10 subjects allergic to milk and 13 milk-tolerant subjects were selected following double-blinded, placebo-controlled food challenges. Their peripheral blood mononuclear cells were stimulated in vitro with casein. Following allergen stimulation, the percentage of Br3s among CD5(+) B cells decreased from 11.5% ± 13.7% to 8.0% ± 9.6% (P = 0.042, n = 5) in the milk-allergy group and increased from 14.7% ± 15.6% to 18.9% ± 20.1% (P = 0.006, n = 7) in the milk-tolerant group. However, the numbers of Br3s increased only in the milk-tolerant group, from 1,954 ± 1,058 to 4,548 ± 1,846 per well (P = 0.026), whereas the numbers of Br3s in the milk-allergy group were unchanged [2,596 ± 823 to 2,777 ± 802 per well (P = 0.734)]. The numbers of apoptotic events were similar to the numbers of total Br3 responses. The percentage of non-TGF-β-producing CD5(+) B cells with apoptotic changes increased from 13.4% ± 17.1% to 16.4% ± 20.3% (P = 0.047, n = 5) in the milk-allergy group and remained unchanged [from 9.9% ± 11.9% to 9.3% ± 11.4% (P = 0.099, n = 7)] in the milk-tolerant group. Using carboxyfluorescein succinimidyl ester labeling, we observed that the percentage of proliferating Br3s among CD5(+) B cells was unchanged [from 6.1% ± 2.8% to 6.4% ± 2.9% (P = 0.145)] in the milk-allergy group and increased from 6.8% ± 3.9% to 10.2% ± 5.3% (P = 0.024) in the milk-tolerant group. In conclusion, Br3s proliferated in response to allergen stimulation in the milk-tolerant group and not in the milk-allergy group. TGF-β-producing regulatory B cells (Br3) may be involved in allergy tolerance by negatively regulating the immune system with TGF-β, and this negative regulation may be controlled by apoptosis.

A system for counting fetal and maternal red blood cells.

PubMed

Ge, Ji; Gong, Zheng; Chen, Jun; Liu, Jun; Nguyen, John; Yang, Zongyi; Wang, Chen; Sun, Yu

2014-12-01

The Kleihauer-Betke (KB) test is the standard method for quantitating fetal-maternal hemorrhage in maternal care. In hospitals, the KB test is performed by a certified technologist to count a minimum of 2000 fetal and maternal red blood cells (RBCs) on a blood smear. Manual counting suffers from inherent inconsistency and unreliability. This paper describes a system for automated counting and distinguishing fetal and maternal RBCs on clinical KB slides. A custom-adapted hardware platform is used for KB slide scanning and image capturing. Spatial-color pixel classification with spectral clustering is proposed to separate overlapping cells. Optimal clustering number and total cell number are obtained through maximizing cluster validity index. To accurately identify fetal RBCs from maternal RBCs, multiple features including cell size, roundness, gradient, and saturation difference between cell and whole slide are used in supervised learning to generate feature vectors, to tackle cell color, shape, and contrast variations across clinical KB slides. The results show that the automated system is capable of completing the counting of over 60,000 cells (versus ∼2000 by technologists) within 5 min (versus ∼15 min by technologists). The throughput is improved by approximately 90 times compared to manual reading by technologists. The counting results are highly accurate and correlate strongly with those from benchmarking flow cytometry measurement.

Neutral dynamics and cell renewal of colonic crypts in homeostatic regime

NASA Astrophysics Data System (ADS)

Fendrik, A. J.; Romanelli, L.; Rotondo, E.

2018-05-01

The self renewal process in colonic crypts is the object of several studies. We present here a new compartment model with the following characteristics: (a) we distinguish different classes of cells: stem cells, six generations of transit amplifying cells and the differentiated cells; (b) in order to take into account the monoclonal character of crypts in homeostatic regimes we include symmetric divisions of the stem cells. We first consider the dynamic differential equations that describe the evolution of the mean values of the populations, but the small observed value of the total number of cells involved plus the huge dispersion of experimental data found in the literature leads us to study the stochastic discrete process. This analysis allows us to study fluctuations, the neutral drift that leads to monoclonality, and the effects of the fixation of mutant clones.

Multicolor flow-cytometric analysis of milk allergen-specific T-helper type 2 cells revealed coexpression of interleukin-4 with Foxp3.

PubMed

Yamawaki, Kazuo; Inuo, Chisato; Nomura, Takayasu; Tanaka, Kenichi; Nakajima, Yoichi; Kondo, Yasuto; Yoshikawa, Tetsushi; Urisu, Atsuo; Tsuge, Ikuya

2015-12-01

Allergen-specific T-helper type 2 (TH2) cells play an important role in the development of allergic inflammation; however, investigations of the properties of allergen-specific T cells have been challenging in humans. Despite clear evidence that forkhead box p3 (Foxp3) is expressed in conventional effector T cells, its function has remained unknown. To characterize allergen-specific TH2 cells in milk allergy, with particular focus on the expression of Foxp3. Twenty-one children with milk allergy and 11 children without milk allergy were studied. Peripheral blood mononuclear cells from subjects were stimulated with milk allergen for 6 hours and analyzed using multicolor flow cytometry to identify CD154(+) allergen-specific T-helper cells. Simultaneously, the expression of intracellular cytokines and Foxp3 was analyzed. The milk allergy group had significantly larger numbers of milk allergen-specific interleukin (IL)-4- and IL-5-producing CD4(+) T cells than the control group. Subjects in the milk allergy group had significantly more CD154(+)CD4(+) IL-10-producing cells and CD154(+)Foxp3(+)CD4(+) cells than those in the control group. In addition, the number of milk allergen-specific CD154(+)Foxp3(+)CD4(+) cells strongly correlated with that of CD154(+)IL4(+)CD4(+) cells. Bcl-2 expression in CD154(+)IL-4(+)Foxp3(+) T-helper cells was significantly lower compared with that in total CD4 cells. Increased numbers of IL-4-producing allergen-specific T-helper cells were found in patients with milk allergy. In addition, Foxp3 was coexpressed with IL-4 in allergen-specific TH2 cells from patients. This coexpression was associated with lower Bcl-2 levels and could contribute to the phenotype and function of TH2 cells. Copyright © 2015 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Aqueous Extract of Agaricus blazei Murrill Prevents Age-Related Changes in the Myenteric Plexus of the Jejunum in Rats

PubMed Central

de Santi-Rampazzo, Ana Paula; Schoffen, João Paulo Ferreira; Cirilo, Carla Possani; Zapater, Mariana Cristina Vicente Umada; Vicentini, Fernando Augusto; Soares, Andréia Assunção; Peralta, Rosane Marina; Bracht, Adelar; Buttow, Nilza Cristina; Natali, Maria Raquel Marçal

2015-01-01

This study evaluated the effects of the supplementation with aqueous extract of Agaricus blazei Murrill (ABM) on biometric and blood parameters and quantitative morphology of the myenteric plexus and jejunal wall in aging Wistar rats. The animals were euthanized at 7 (C7), 12 (C12 and CA12), and 23 months of age (C23 and CA23). The CA12 and CA23 groups received a daily dose of ABM extract (26 mg/animal) via gavage, beginning at 7 months of age. A reduction in food intake was observed with aging, with increases in the Lee index, retroperitoneal fat, intestinal length, and levels of total cholesterol and total proteins. Aging led to a reduction of the total wall thickness, mucosa tunic, villus height, crypt depth, and number of goblet cells. In the myenteric plexus, aging quantitatively decreased the population of HuC/D+ neuronal and S100+ glial cells, with maintenance of the nNOS+ nitrergic subpopulation and increase in the cell body area of these populations. Supplementation with the ABM extract preserved the myenteric plexus in old animals, in which no differences were detected in the density and cell body profile of neurons and glial cells in the CA12 and CA23 groups, compared with C7 group. The supplementation with the aqueous extract of ABM efficiently maintained myenteric plexus homeostasis, which positively influenced the physiology and prevented the death of the neurons and glial cells. PMID:25960748

Size and cell number of the utricle in kinetotically swimming fish: a parabolic aircraft flight study

NASA Astrophysics Data System (ADS)

Bäuerle, A.; Anken, R. H.; Hilbig, R.; Baumhauer, N.; Rahmann, H.

2004-01-01

Humans taking part in parabolic aircraft flights (PAFs) may suffer from space motion sickness (SMS, a kinetosis). Since it has been repeatedly shown earlier that some fish of a given batch also reveal a kinetotic behavior during PAFs (especially so-called spinning movements and looping responses) and due to the homology of the vestibular apparatus among all vertebrates, fish can be used as model systems to investigate the origin of susceptibility to motion sickness. Therefore, we examined the utricular maculae (they are responsible for the internalization of gravity in teleosteans) of fish swimming kinetotically at microgravity in comparison with animals from the same batch who swam normally. On the histological level, it was found that the total number of both sensory and supporting cells of the utricular maculae did not differ between kinetotic animals as compared to normally swimming fish. Cell density (sensory and supporting cells/100 μm 2), however, was reduced in kinetotic animals ( p < 0.0001), which seemed to be due to malformed epithelial cells (increase in cell size) of the kinetotic specimens. Susceptibility to kinetoses may therefore originate in malformed sensory epithelia.

Immune and Inflammatory Cell Composition of Human Lung Cancer Stroma

PubMed Central

Banat, G-Andre; Tretyn, Aleksandra; Pullamsetti, Soni Savai; Wilhelm, Jochen; Weigert, Andreas; Olesch, Catherine; Ebel, Katharina; Stiewe, Thorsten; Grimminger, Friedrich; Seeger, Werner; Fink, Ludger; Savai, Rajkumar

2015-01-01

Recent studies indicate that the abnormal microenvironment of tumors may play a critical role in carcinogenesis, including lung cancer. We comprehensively assessed the number of stromal cells, especially immune/inflammatory cells, in lung cancer and evaluated their infiltration in cancers of different stages, types and metastatic characteristics potential. Immunohistochemical analysis of lung cancer tissue arrays containing normal and lung cancer sections was performed. This analysis was combined with cyto-/histomorphological assessment and quantification of cells to classify/subclassify tumors accurately and to perform a high throughput analysis of stromal cell composition in different types of lung cancer. In human lung cancer sections we observed a significant elevation/infiltration of total-T lymphocytes (CD3+), cytotoxic-T cells (CD8+), T-helper cells (CD4+), B cells (CD20+), macrophages (CD68+), mast cells (CD117+), mononuclear cells (CD11c+), plasma cells, activated-T cells (MUM1+), B cells, myeloid cells (PD1+) and neutrophilic granulocytes (myeloperoxidase+) compared with healthy donor specimens. We observed all of these immune cell markers in different types of lung cancers including squamous cell carcinoma, adenocarcinoma, adenosquamous cell carcinoma, small cell carcinoma, papillary adenocarcinoma, metastatic adenocarcinoma, and bronchioloalveolar carcinoma. The numbers of all tumor-associated immune cells (except MUM1+ cells) in stage III cancer specimens was significantly greater than those in stage I samples. We observed substantial stage-dependent immune cell infiltration in human lung tumors suggesting that the tumor microenvironment plays a critical role during lung carcinogenesis. Strategies for therapeutic interference with lung cancer microenvironment should consider the complexity of its immune cell composition. PMID:26413839

Nutritional deprivation and LPS exposure as feasible methods for induction of cellular - A methodology to validate for vitro photobiomodulation studies.

PubMed

Basso, F G; Turrioni, A P S; Almeida, L F; Soares, D G; Oliveira, C F; Hebling, J; de Souza Costa, C A

2016-06-01

Previous studies have demonstrated that high biostimulation takes place when cells under stress are subjected to phototherapy by laser or light-emitting-diode (LED) devices. Several studies selected nutritional deprivation by reducing the concentration of fetal bovine serum (FBS) in the culture medium or the exposure of cultured cells to lipopolysaccharide (LPS) as an in vitro cellular stress condition. However, there are no data certifying that these stimuli cause stressful conditions for cultured cells. This investigation assessed the induction of cellular stress by decreasing the concentration of FBS or adding LPS to culture medium. Odontoblast-like cells (MDPC-23) were cultured in complete culture medium (DMEM) containing 10% FBS. After a 12-hour incubation period, the DMEM was replaced by fresh medium containing 10% FBS (control), low concentrations of FBS (0, 0.2, 0.5, 2, or 5%) or LPS from Escherichia coli (10μg/ml). After an additional 12-hour incubation, cell viability, total cell-counting, total protein production, and gene expression of heat shock protein 70 (HSP70) were assessed. Data were statistically analyzed by ANOVA complemented by the Tukey test, with 5% considered significant. Cell viability was negatively affected only for 0% FBS, while reduced viable cell numbers and total protein production were detected for FBS concentrations lower than 2%. Higher HSP70 gene expression was also observed for FBS concentrations lower than 2% and for cells exposed to LPS. The nutritional deprivation model with culture medium lower than 2% of FBS can be safely used to induce cellular stress for in vitro photobiomodulation studies. Copyright © 2016 Elsevier B.V. All rights reserved.

Aromatase mRNA in the brain of adult green anole lizards: effects of sex and season.

PubMed

Cohen, R E; Wade, J

2011-03-01

Neural testosterone metabolism, particularly the synthesis of oestradiol (E(2)) via the aromatase enzyme, is important for sexual behaviours in many vertebrates. In green anole lizards, E(2) metabolised from testosterone facilitates female receptivity and increases sexual motivation in males. Testosterone treatment increases aromatase activity in the whole brain homogenates of gonadectomised male, but not female, anoles, which is an effect limited to the breeding season (BS). To investigate the potential for local effects of this enzyme in reproductive behaviour, we used in situ hybridisation for aromatase mRNA to examine expression during the BS and nonbreeding season (NBS) in areas of the brain that control male sexual behaviours [preoptic area (POA) and amygdala (AMY)], as well as one regulating female reproductive behaviours ventromedial hypothalamus (VMH). Males had a greater total number of aromatase-expressing cells in the POA than females, and the density of aromatase-expressing cells (number per unit volume) was greater in the VMH and AMY of females. This density was also higher during the BS than NBS in the POA. Expression of aromatase in the AMY appeared to be lateralised because trends were detected for the left side to have more total cells and more cells per unit volume than the right. These results suggest that, similar to other vertebrates, regional aromatisation of testosterone may be important for the control of sex-specific reproductive behaviours. © 2011 The Authors. Journal of Neuroendocrinology © 2011 Blackwell Publishing Ltd.

Influence of coronary artery diameter on eNOS protein content

NASA Technical Reports Server (NTRS)

Laughlin, M. H.; Turk, J. R.; Schrage, W. G.; Woodman, C. R.; Price, E. M.

2003-01-01

The purpose of this study was to test the hypothesis that the content of endothelial nitric oxide synthase (eNOS) protein (eNOS protein/g total artery protein) increases with decreasing artery diameter in the coronary arterial tree. Content of eNOS protein was determined in porcine coronary arteries with immunoblot analysis. Arteries were isolated in six size categories from each heart: large arteries [301- to 2,500-microm internal diameter (ID)], small arteries (201- to 300-microm ID), resistance arteries (151- to 200-microm ID), large arterioles (101- to 150-microm ID), intermediate arterioles (51- to 100-microm ID), and small arterioles(<50-microm ID). To obtain sufficient protein for analysis from small- and intermediate-sized arterioles, five to seven arterioles 1-2 mm in length were pooled into one sample for each animal. Results establish that the number of smooth muscle cells per endothelial cell decreases from a number of 10 to 15 in large coronary arteries to 1 in the smallest arterioles. Immunohistochemistry revealed that eNOS is located only in endothelial cells in all sizes of coronary artery and in coronary capillaries. Contrary to our hypothesis, eNOS protein content did not increase with decreasing size of coronary artery. Indeed, the smallest coronary arterioles had less eNOS protein per gram of total protein than the large coronary arteries. These results indicate that eNOS protein content is greater in the endothelial cells of conduit arteries, resistance arteries, and large arterioles than in small coronary arterioles.

Statistical Characterization of 18650-Format Lithium-Ion Cell Thermal Runaway Energy Distributions

NASA Technical Reports Server (NTRS)

Walker, William Q.; Rickman, Steven; Darst, John; Finegan, Donal; Bayles, Gary; Darcy, Eric

2017-01-01

Effective thermal management systems, designed to handle the impacts of thermal runaway (TR) and to prevent cell-to-cell propagation, are key to safe operation of lithium-ion (Li-ion) battery assemblies. Critical factors for optimizing these systems include the total energy released during a single cell TR event and the fraction of the total energy that is released through the cell casing vs. through the ejecta material. A unique calorimeter was utilized to examine the TR behavior of a statistically significant number of 18650-format Li-ion cells with varying manufacturers, chemistries, and capacities. The calorimeter was designed to contain the TR energy in a format conducive to discerning the fractions of energy released through the cell casing vs. through the ejecta material. Other benefits of this calorimeter included the ability to rapidly test of large quantities of cells and the intentional minimization of secondary combustion effects. High energy (270 Wh/kg) and moderate energy (200 Wh/kg) 18650 cells were tested. Some of the cells had an imbedded short circuit (ISC) device installed to aid in the examination of TR mechanisms under more realistic conditions. Other variations included cells with bottom vent (BV) features and cells with thin casings (0.22 1/4m). After combining the data gathered with the calorimeter, a statistical approach was used to examine the probability of certain TR behavior, and the associated energy distributions, as a function of capacity, venting features, cell casing thickness and temperature.

The number of tumor infiltrating T-cell subsets in lymph nodes from patients with Hodgkin lymphoma is associated with the outcome after first line ABVD therapy.

PubMed

Alonso-Álvarez, Sara; Vidriales, Maria Belén; Caballero, Maria Dolores; Blanco, Oscar; Puig, Noemí; Martin, Alejandro; Peñarrubia, Maria Jesús; Zato, Esther; Galende, Josefina; Bárez, Abelardo; Alcoceba, Miguel; Orfão, Alberto; González, Marcos; García-Sanz, Ramón

2017-05-01

Prognostic factors in Hodgkin lymphoma (HL) still fail to accurately identify high-risk patients. Tumor microenvironment in HL is a current focus of research for risk definition but few studies have focused on infiltrating lymphocytes. Here, we analyzed the number of tumor infiltrating lymphocytes by flow cytometry in diagnostic biopsies from 96 HL homogeneously treated patients with ABVD with or without radiotherapy. Most lymph node cells were lymphocytes (90 ± 17), with a median T/B/NK distribution of 74%/26%/0.7%, and CD4 + T-cell predominance. The amount of CD19 + B cells, and NK cells did not show association with disease features. However, high numbers of CD8 + and CD4 + cells were associated with better and poorer outcomes, respectively. Patients with ≥15% cytotoxic CD8 + cells among the total cell population had a longer 10-year freedom from treatment failure (FFTF) (93% vs. 73%, p=.04). In turn, cases with ≥75% of CD4 + infiltrating cells showed a significantly decreased FFTF (73% vs. 96%, p=.021). Consequently, CD4/CD8 ratio ≥5 associated with a poorer 10-year FFTF (69.5% vs. 94%, p=.02). This deleterious effect was particularly prominent in advanced disease (n = 58, p=.01). In multivariate analysis, a CD4/CD8 ratio ≥5 was the only independent variable to predict for treatment failure (HR = 4.5, 95% confidence interval, 1.2-16.8). In conclusion, our study shows that high CD4 + and low CD8 + T-cells infiltrates of tumor specimens associate with poor prognosis in HL patients, and CD4/CD8 ratio might be potentially useful for tailoring therapy.