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1.
Apoptosis phenomenon in the schistosomulum and adult worm life cycle stages of Schistosoma japonicum
Hongxiao Han Jinbiao Peng Geoffrey N. Gobert Yang Hong Min Zhang Yanhui Han Zhiqiang Fu Yaojun Shi Jinjun Xu Jiaojiao Lin Jianping Tao 《Parasitology international》2013,62(2):100-108
Apoptosis is an important aspect of a number of biological processes, from embryogenesis to the stress–injury response. It plays a central role in balancing cell proliferation and tissue remodeling activity in many organisms. In the present study, apoptosis in 14 days post infection schistosomula was evaluated using TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling) assays and DAPI staining. Additionally, flow cytometry using the Annexin V-FITC/propidium iodide (PI) (Annexin V/PI) assay confirmed the percentage of early apoptotic, late apoptotic, and necrotic cells in 14 and 23 days post infection worms. Conserved Domain Database (CDD) BLAST analysis and alignment analysis of known schistosome proteins demonstrated the feasibility of detecting the activity of caspase-3 and -7 using the caspase-3/7 Glo analysis assay. Analysis of caspase-3 and -7 activities in schistosome demonstrated that both caspases were active in each developmental stage of Schistosoma japonicum, but was highest in the 14 days post infection schistosomula. Additionally, the caspase peptide inhibitor (Z-VAD-FMK) inhibited the caspase-3/7 activity at all developmental stages examined. Therefore, we hypothesized that two main signaling pathways are involved in apoptosis in S. japonicum, the caspase cascade and the mitochondrial-initiated pathway. We have constructed a model of these two pathways, including how they may interact and their biological outcomes. qRT-PCR analyses of the gene expression profiles of apoptosis-related genes supported our hypothesis of the relationship between the apoptotic pathway and parasite development. The data presented here demonstrates that apoptosis is an important biological process for the survival and development of the schistosome, and identifies potential novel therapeutic targets. 相似文献
2.
Analysis of radiation-induced apoptosis in human lymphocytes: flow cytometry using Annexin V and propidium iodide versus the neutral comet assay 总被引:15,自引:0,他引:15
BACKGROUND: The neutral comet assay was devised to measure double-stranded DNA breaks, but it has also been used to measure apoptosis based on its characteristic DNA fragmentation patterns. There is still uncertainty about the reliability of this method. By comparing the comet assay with a flow cytometry method that uses Annexin V binding to apoptotic cells, we have provided further evidence for evaluating the usefulness of the comet assay for detecting apoptosis. METHODS: Apoptosis was induced in human peripheral blood mononuclear cells (PBMC) by ionizing radiation and measured using the comet assay and a flow cytometry method that measures Annexin V and propidium iodide (PI) staining. RESULTS: The Annexin V flow cytometry assay distinguished among early apoptosis, late apoptosis, and an apoptotic or necrotic phase in which the cells were labeled with both Annexin V and PI. The comet assay detected only the latter two phases of apoptosis. CONCLUSIONS: The comet assay is a useful tool for measuring the late stages of apoptosis whereas the Annexin V assay measures higher amounts of apoptosis because it can detect cells in an earlier stage of the apoptotic pathway. 相似文献
3.
ABSTRACT: BACKGROUND: Buddleja davidii is widely distributed in the southwestern region of China. We have undertaken a systematic analysis of B. davidii as a Chinese traditional medicine with anticancer activity by isolating natural products for their activity against the human gastric cancer cell line Mgc-803 and the human breast cancer cell line Bcap-37. RESULTS: Ten compounds were extracted and isolated from B. davidii, among which colchicine was identified in B. davidii for the first time. The inhibitory activities of these compounds were investigated in Mgc-803, Bcap-37 cells in vitro by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay, and the results showed that luteolin and colchicine had potent inhibitory activities against the growth of Mgc-803 cells. Subsequent fluorescence staining and flow cytometry analysis indicated that these two compounds could induce apoptosis in Mgc-803 cells. The results also showed that the percentages of early apoptotic cells (Annexin V+/PI-, where PI is propidium iodide) and late apoptotic cells (Annexin V+/PI+) increased in a dose- and time-dependent manner. After 36 h of incubation with luteolin at 20 muM, the percentages of cells were approximately 15.4% in early apoptosis and 43.7% in late apoptosis; after 36 h of incubation with colchicine at 20 muM, the corresponding values were 7.7% and 35.2%, respectively. CONCLUSIONS: Colchicine and luteolin from B. davidii have potential applications as adjuvant therapies for treating human carcinoma cells. These compounds could also induce apoptosis in tumor cells. 相似文献
4.
Rapid cold-hardening protects <Emphasis Type="Italic">Drosophila melanogaster</Emphasis> from cold-induced apoptosis 总被引:1,自引:0,他引:1
Yi SX Moore CW Lee RE 《Apoptosis : an international journal on programmed cell death》2007,12(7):1183-1193
The rapid cold-hardening (RCH) response increases the cold tolerance of insects by protecting against non-freezing, cold-shock
injury. Apoptosis, or programmed cell death, plays important roles in development and the elimination of sub-lethally damaged
cells. Our objectives were to determine whether apoptosis plays a role in cold-shock injury and, if so, whether the RCH response
protects against cold-induced apoptosis in Drosophila melanogaster. The present study confirmed that RCH increased the cold tolerance of the adults at the organismal level. No flies in the
cold-shocked group survived direct exposure to ‒7°C for 2 h, whereas significantly more flies in the RCH group survived exposure
to ‒7°C for 2 h after a 2-h exposure to 5°C. We used a TUNEL assay to detect and quantify apoptotic cell death in five groups
of flies including control, cold-shocked, RCH, heat-shocked (37.5°C, 30 min), and frozen (‒20°C, 24 h) and found that apoptosis
was induced by cold shock, heat shock, and freezing. The RCH treatment significantly improved cell viability by 38% compared
to the cold-shocked group. Cold shock-induced DNA fragmentation shown by electrophoresis provided further evidence for apoptosis.
SDS-PAGE analysis revealed an RCH-specific protein band with molecular mass of ∼150 kDa. Western-blotting revealed three proteins
that play key roles in the apoptotic pathway: caspase-9-like (apoptotic initiator), caspase-3-like (apoptotic executioner)
and Bcl-2 (anti-apoptotic protein). Consequently, the results of this study support the hypothesis that the RCH response protects
against cold-shock-induced apoptosis. 相似文献
5.
A new four-color flow cytometric assay to detect apoptosis in lymphocyte subsets of cultured peripheral blood cells 总被引:1,自引:0,他引:1
Hasper HJ Weghorst RM Richel DJ Meerwaldt JH Olthuis FM Schenkeveld CE 《Cytometry》2000,40(2):167-171
BACKGROUND: Human peripheral blood lymphocytes kept in culture after isolation die by an apoptotic process. Detection of apoptosis with labeled Annexin V to demonstrate loss of plasma membrane asymmetry is sensitive, specific, and easy using flow cytometry. This is true in lymphoblastic cell lines when combining Annexin V-fluorescein isothiocyanate (FITC) and propidium iodide (PI). However, measurement of apoptosis by flow cytometry in isolated human lymphocytes using Annexin V-FITC/PI is disturbed by the presence of a variable percentage of erythrocytes in the isolated lymphocyte population. To overcome this problem, we have developed and tested a new four-color flow cytometric assay to detect apoptosis in lymphocyte subsets of cultured peripheral blood cells. METHODS: Peripheral blood lymphocytes are isolated by density gradient centrifugation. Nucleus-containing cells are selected using CD45-phycoerythrin (PE). The lymphocyte subset of interest is selected using CD4, CD8, or CD19 energy-coupled dye (ECD) labeling. Apoptosis is detected using Annexin V-FITC with 7-amino-Actinomycin-D (7-AAD) to distinguish early apoptotic from late apoptotic lymphocytes. RESULTS: We have developed a new technique to detect apoptosis in isolated human peripheral blood lymphocyte subsets with good reproducibility, coefficient of variation < 17%. CONCLUSIONS: We now have a validated tool to study apoptosis in subsets of isolated human lymphocytes to increase our knowledge of pathogenesis and therapies in lymphoreticular malignancies. 相似文献
6.
《Saudi Journal of Biological Sciences》2021,28(9):5214-5220
Erythropoietin (EPO) is widely used to treat anemia in patients undergoing chemotherapy for cancers. The main objective of this study was to investigate the effect of rHuEPO on the response of spheroid breast cancer, MCF-7, cells to tamoxifen treatment. The MCF-7 spheroids were treated with 10 mg/mL tamoxifen in combination with either 0, 10, 100 or 200 IU/mL rHuEPO for 24, 48 or 72 h. The viability of the MCF-7 cells was determined using the annexin-V, cell cycle, caspases activation and acridine orange/propidium iodide staining. rHuEPO-tamoxifen combination significantly (p greater than 0.05) increased the number of spheroid MCF-7 cells entering early apoptotic phase after 12 h and late apoptotic phase after 24 h of treatment; primarily the result of the antiproliferative effect tamoxifen. Tamoxifen alone significantly (p < 0.05) increased the caspase-3 and −9 activities in the spheroid MCF-7 cells by 200 to 550% of the control. Combination rHuEPO and tamoxifen produced much lesser effect on the caspase-8 activity. The rHuEPO in the combination treatment had concentration-dependently caused decrease in the caspase activities. rHuEPO-tamoxifen combination markedly increased MCF-7 cells entering the SubG0/G1 phase of the cell cycle by more than 500% of the control, while decreasing those entering the G2 + M and S phases by 50%. After 72 h, the combination treatment produced greater (p < 0.05) change in the SubG0/G1 phase than tamoxifen treatment alone. Morphologically, spheroid MCF-7 cells subjected to combination rHuEPO-tamoxifen treatment showed nuclear condensation and margination, cytoplasmic blebbing, necrosis, and early and late apoptosis. Thus, the study showed that rHuEPO-tamoxifen combination induced apoptosis in the spheroid MCF-7 cells. The apoptotic effect of the rHuEPO-tamoxifen combination treatment on the MCF-7 cells was greater than that produced by tamoxifen alone. The rHuEPO-tamoxifen treatment enhanced the caspase-independent apoptotic effects of tamoxifen on the spheroid MCF-7 cells. 相似文献
7.
BACKGROUND: Chemical induction of apoptosis in cells is believed to contribute to toxicity. Techniques for measuring apoptosis have increased in both sensitivity and number and in many cases can be readily extended to nontraditional research species. A comparison of established assays for measuring apoptosis of lymphoid cells has thus far not been performed in the fish and thus would be efficacious in assessing immunotoxicity. METHODS: The present study evaluated chemical-induced immune cell apoptosis in fish (tilapia, Oreochromis niloticus) exposed to two known immunotoxic chemicals, azathioprine and T-2 toxin. Cytocentrifugation and light microscopy of leukocyte-enriched cell samples from the pronephros (i.e., the fish primary hematopoietic compartment) demonstrated chemical-related increases in apoptotic bodies. This observation was examined further with the ApoAlert Annexin V Apoptosis kit and two DNA-binding dyes employed for detecting apoptosis, 7-aminoactinomycin D (7-AAD) and propidium iodide (PI). RESULTS: The apoptotic probes confirmed the microscopic observations of increased apoptosis in the chemical-exposed fish. The ApoAlerttrade mark annexin V and 7-AAD assays, which discriminate early and late apoptosis/necrosis, compared well in identifying apoptotic populations. PI staining in Vindelov's solution was unable to detect early apoptosis. CONCLUSIONS: The present data suggest that apoptotic immune cells may be a useful marker for certain immunotoxicant exposures in fish. These findings agree with those of previous reports that fish may respond immunologically in a manner similar to mammals after immunotoxicant challenge. 相似文献
8.
We describe a microfluidic device with microgrooved patterns for studying cellular behavior. This microfluidic platform consists of a top fluidic channel and a bottom microgrooved substrate. To fabricate the microgrooved channels, a top poly(dimethylsiloxane) (PDMS) mold containing the impression of the microfluidic channels was aligned and bonded to a microgrooved substrate. Using this device, mouse fibroblast cells were immobilized and patterned within microgrooved substrates (25, 50, 75, and 100 microm wide). To study apoptosis in a microfluidic device, media containing hydrogen peroxide, Annexin V, and propidium iodide was perfused into the fluidic channel for 2 hours. We found that cells exposed to the oxidative stress became apoptotic. These apoptotic cells were confirmed by Annexin V that bound to phosphatidylserine at the outer leaflet of the plasma membrane during the apoptosis process. Using this microfluidic device with microgrooved patterns, the apoptosis process was observed in real-time and analyzed by using an inverted microscope containing an incubation chamber (37 degrees C, 5% CO(2)). Therefore, this microfluidic device incorporated with microgrooved substrates could be useful for studying the cellular behavior and performing high-throughput drug screening. 相似文献
9.
Schöier J Högdahl M Söderlund G Kihlström E 《FEMS immunology and medical microbiology》2006,47(2):207-216
Atherosclerosis and coronary heart disease are causing high morbidity and mortality worldwide. Different risk factors have been demonstrated, but the exact mechanisms behind these diseases are still not fully understood. Recent studies have suggested Chlamydia pneumoniae to be involved in the pathogenesis, and increased apoptotic indexes in atherosclerotic plaques have been documented. In this study, we show that C. pneumoniae induces apoptosis and necrosis in populations of human coronary artery endothelial cells. Apoptosis was determined by TUNEL and flow cytometry after staining of cells with annexin V and propidium iodide, and defined as TUNEL-reactive or annexin V-positive, propidium iodide-negative cells. The apoptosis was induced within 2 h postinfection and increased with inoculation dose. The general caspase inhibitor z-VAD-fmk did not affect apoptotic frequencies. By immunochemistry and immunoblot, we demonstrated activation and subcellular translocation of the proapoptotic protein Bax, and translocation of apoptosis-inducing factor from the cytosol to the nucleus. These results indicate that C. pneumoniae-induced apoptosis in human coronary artery endothelial cells is caspase-independent and regulated by Bax and apoptosis-inducing factor. 相似文献
10.
At present, the current therapeutic strategy for apoptosis induction mainly relies on the administration of pharmacological apoptotic modulators. Apart from that, apoptosis can be induced by various external stimuli such as hyperthermia, ionizing radiation, and electric fields. Despite advantages, both physical and pharmacological approaches bear some limitations as well. The rationale of this study was to overcome the limitations by combining hyperthermia and apoptotic modulator ‘bortezomib’ (Velcade). Two types of human blood cancer cell lines were utilized: human leukemic monocyte lymphoma cell U937 line and peripheral blood mononuclear cells (PMBCs) derived from the patient diagnosed with acute myeloid leukemia. Prior to apoptosis experiments, cytotoxicity tests were performed at three types of temperature regimes (40 °, 42 ° and 44 °C). We observed a gradual inhibition of cell viability correlating with an increase of temperature and drug concentration in both cell lines. However, there was no significant difference between sham group and groups of leukemic PMBCs treated by high temperature (44 °C) and bortezomib. In U937 cells, combined treatment by heat shock and bortezomib led to an increase the number of cells underwent the late apoptosis stage. At the same time, similar treatment of PMBCs resulted in the stimulation of early apoptosis. Our data suggest that combination of bortezomib and hyperthermia enhances apoptosis induction in human cancer white blood cells, indicating a therapeutic potential for blood cancer therapy. 相似文献
11.
The effects of insulin-like growth factor-1 (IGF-1) on the cytotoxicity and apoptosis induced by okadaic acid (OA) in SH-SY5Y cells were investigated. Cell viability was measured using the MTT (3-(4,5-dimethylthiazolyl-2)-2,-5-diphenyltetrazolium bromide) assay. Early and late apoptosis/necrosis were analyzed by flow cytometry using Annexin V and propidium iodide (PI) double-staining. Caspase-3 activation was detected by Western blot analysis. Preincubation with IGF-1 for 24 h prevented cytotoxicity induced by 40 nM OA given for 24 h, and the MTT value significantly increased. Incubation with 20 nM OA for 24 h caused a marked increase in the percentage of early apoptotic and late apoptotic/necrotic cells, which was not dependent on the activation of caspase-3. OA-induced apoptosis was significantly decreased by pretreatment with 10 ng/ml of IGF-1 for 24 h. The results supported the hypothesis that IGF-1 may be useful in the treatment of Alzheimer's disease. 相似文献
12.
Rosales-Torres AM Avalos-Rodríguez A Vergara-Onofre M Hernández-Pérez O Ballesteros LM García-Macedo R Ortíz-Navarrete V Rosado A 《Molecular reproduction and development》2000,55(3):270-281
The differential quantitative participation of apoptosis and necrosis in ewe antral follicles of two different sizes, separated in four stages of atresia using macroscopic, histologic, and esteroid quantification methods was assessed. Annexin V binding and propidium iodide (PI) uptake was used to detect healthy live cells (Annexin V negative/PI negative), early apoptotic cells (Annexin V+/PI-), and necrotic or late apoptotic cells (PI+). Additionally we used internucleosomal DNA fragmentation as a quantitative estimate of apoptosis. Presence and distribution of lysosomal enzymes in follicular fluid and granulosa cells was used as a measure of necrotic cell death. DNA flow cytometry and gel electrophoresis were positively correlated with the progression of atresia, small atretic follicles tend to have higher percentages of internucleosomal cleaved DNA than follicles >6 mm. Annexin/PI binding also indicates that apoptosis and necrosis increase with atresia progression, generally apoptosis outweighs necrosis in small follicles. Acid phosphatase and glucosaminidase in follicular fluid of 3-6 mm follicles showed no significant modifications between healthy and initially atretic follicles, and only a small, but significant increase in activity in advancedly atretic follicles. On the contrary, lysosomal enzyme activity in follicles >6 mm showed positive correlation between atresia stages and the activities of acid phosphatase and glucosaminidase in follicular fluid. A similar size-differential behavior was found in free or membrane-bound lysosomal enzyme activity of granulosa cells. Necrosis, but principally apoptosis, were present during all stages of follicular maturation indicating that growth and maturation of ovarian follicles involves a continuous renewal of granulosa cells, regulated by apoptosis. Mechanisms regulating this equilibrium may participate in the final destiny, whether ovulation or atresia of ovarian follicles. 相似文献
13.
To investigate whether Annexin 1 can protect a retinal ganglion cells line (RGC-5) from apoptosis as induced by serum deprivation.
Annexin 1 location in RGC-5 cells was determined using an indirect immunofluorescent assay. Expression of Annexin 1 in RGC-5
cultures deprived of serum for 0, 2 days was semi-quantified by western blot and RT-PCR. Effects of varying concentrations
of the Annexin 1 peptide fragment, Ac2-26, on the survival of the RGC-5 cells was determined, and apoptotic cells were quantified
by flow cytometry. Immunoblot and RT-PCR analysis was preformed to identify caspase 3, bax and bcl-2 in RGC extracts. Annexin
1 was localized in the cytoplasm of RGC-5 cells and the expression of Annexin 1, caspase 3 and bax was upregulated in serum-deprived
RGC-5 cells. Ac2-26 attenuated the apoptosis resulting from serum deprivation of RGC-5 in a concentration-dependent manner,
decreased caspase 3 and bax levels and produced an increase of bcl-2 in cell lysates. Annexin 1, in specific the peptide fragment
Ac2-26, may play an important role in decreasing apoptosis in serum-deprived RGC-5 cells. 相似文献
14.
Liang Zhang Ji Li ZhenZhou Jiang LiXin Sun Xue Mei Bian Yong LuYong Zhang 《Biochemical and biophysical research communications》2011,(1):68
The aim of this study was to investigate the protective effect of inhibition of aquaporin-1 (AQP1) expression against aristolochic acid I (AA-I)-induced apoptosis. HK-2 cells impaired by AA-I were used in this study as the cell model of aristolochic acid nephropathy. Apoptosis was studied by different methods, including 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assays, flow cytometry, and caspase 3 activity assays. We compared AA-I-mediated apoptosis in HK-2 cells with or without knockdown of AQP1 expression by RNA interference. MTT assays showed that AA-I inhibited the viability of HK-2 cells in a time- and concentration-dependent manner. Apoptosis was evidenced by the results of the Annexin V/propidium iodide assay and the occurrence of a sub-G1 peak in cell-cycle analysis. The activity of caspase 3 was found to have been increased by AA-I in a concentration-dependent manner. However, AQP1 RNA interference provided protection against injury in cells treated with AA-I (40 μM) for 24 h and attenuated the number of apoptotic cells. These results suggested that AQP1 plays an important role in AA-I-induced apoptosis and that inhibition of AQP1 expression may protect HK-2 cells from AA-I-induced apoptotic damage. 相似文献
15.
John S. Thompson Reto Asmis Andrea A. Tapp Brandy Nelson Yanxia Chu Judith Glass Micheal Moneyhon Stephen A. Brown 《Apoptosis : an international journal on programmed cell death》2010,15(6):705-714
Ionizing radiation (IR) is a pro-oxidant that kills cells by both apoptotic and necrotic mechanisms. Pyrrolidine dithiocarbamate
(PDTC) is a thiol-containing compound that may act either as a pro- or anti-oxidant depending on the experimental conditions.
This study was designed to determine whether PDTC would reduce or enhance IR-induced cell death of freshly-isolated normal
mouse B6/129 spleen cells (NMSC). We determined the effect of increasing doses of IR, PDTC alone and PDTC followed by IR on
the viability of NMSC. Annexin V and propidium iodide (Annexin V/PI) staining demonstrated a dose and time-dependent relationship
in which PDTC enhanced the percentage of IR-induced apoptotic/necrotic NMSC. Trypan blue dye inclusion confirmed that a loss
of membrane integrity was occurring 1 h after incubation with PDTC plus IR. Reduction in the glutathione (GSH)/glutathione
disulfide (GSSG) ratio and GSH demonstrated that both IR (8.5 Gy) and PDTC acted as pro-oxidants, but their mechanisms of
action differed: In contrast to IR, which promoted p53 activation and caspase 3/7-mediated apoptosis, PDTC inhibited IR-induced
p53 and caspase 3/7 activity. However, PDTC increased H2O2 formation and necrosis, resulting in an overall increase in IR-induced cell death. Catalase prevented the PDTC-induced increase
in IR cytotoxicity implicating the generation of H2O2 as a major factor in this mechanism. These results demonstrate that in NMSC PDTC acts as pro-oxidant and enhances IR-induced
cell cytotoxicity by increasing H2O2formation and thiol oxidation. As such, they strongly suggest that the use of PDTC as an adjunct to reduce radiation toxicity
should be avoided. 相似文献
16.
Apoptotic cell death eventually results in secondary necrotic cell death, whereas caspase-independent primary necrotic cell death has been reported and its mechanism involving RIP1 and RIP3 has been recently elucidated. Dual staining with fluorescent Annexin V and propidium iodide (PI) has been used to discriminate apoptotic and necrotic cell death, in which Annexin V-positive/PI-negative staining is regarded as apoptosis and PI-positive staining as necrosis. Here we demonstrate that primary necrotic cells unexpectedly show Annexin V-positive/PI-negative staining before they become PI-positive, and that primary necrotic and apoptotic Annexin V-positive/PI-negative cells can be discriminated by necrostatin-1, an inhibitor of primary necrosis by inhibition of RIP1. 相似文献
17.
《Cryobiology》2020
Boar spermatozoa are extremely sensitive to low temperatures and the cryopreservation causes dramatic changes in sperm survivability, but it is not clear which part of the cryopreservation process affects the most. The aim of this work was to assess early events of apoptotic changes as damage indicators in boar sperm cooled to 5 °C and exposed to different glycerol (GLY) concentrations. For this purpose, progressive sperm motility (CASA), plasmatic and acrosome membranes integrity (CFDA/PI; phase contrast), plasma membrane functionality (HOS), phosphatidylserine translocation (Annexin-V/FITC) and reduction of mitochondrial membrane potential (Ψm) (JC-10) were carried out at 37 °C, 17 °C and 5 °C in eight boar sperm pools. Afterwards, three aliquots were diluted in different freezing extenders (control: 0% GLY; A: 2% GLY and B: 3% GLY); sperm quality and early apoptotic changes were assessed. Motility was negatively affected during cooling to 5 °C. Furthermore, plasma membrane functionality was the most affected by cooling. The number of necrotic cells was higher at 5 °C. However, no differences were observed in phosphatidylserine translocation. The extender with 3% GLY at 5 °C presented better Ψm than 0 and 2% GLY. Based on this analysis, boar sperm cooling to 5 °C does not modify the rate of early apoptotic changes, although alterations in the Ѱm were evident. 相似文献
18.
Under conditions of extreme heat stress, the process of autophagy has previously been shown to protect human cells, but the exact body temperature at which autophagic activation occurs is largely unknown. Further, the interplay between autophagy, the heat shock response (HSR), inflammation, and apoptosis have yet to be examined together under temperature conditions representative of human internal body temperatures at rest (37 °C) or under severe heat stress conditions (41 °C). Thus, the purpose of this study was to examine threshold changes in autophagy, the HSR, inflammation, and apoptosis to increasing levels of ex vivo heat stress. Whole blood was collected from 20 young (23 ± 4 years; 10 men, 10 women) physically active participants. Peripheral blood mononuclear cells (PBMCs) were isolated immediately (baseline) and after 90-min of whole blood heating in 37, 39, and 41 °C water baths, representative of normal resting (non-heat stress) as well as moderate and severe heat stress conditions in humans, respectively. At 37 °C, increased autophagic activity was demonstrated, with no change in the HSR, and inflammation. Subsequently, responses of autophagy, the HSR, and inflammation increased with a moderate heat stress (39 °C), with further increases in only autophagy and the HSR under a severe heat stress of 41 °C. We observed no increase in apoptosis under any temperature condition. Our findings show that in human PBMCs, the autophagy and HSR systems may act cooperatively to suppress apoptotic signaling following heat stress, which may in part be mediated by an acute inflammatory response. 相似文献
19.
Apoptosis in human embryo development: 1. Cerebral cortex 总被引:3,自引:2,他引:1
We investigated the apoptosis at the beginning of human cerebral cortex development, in the 6th week of embryogenesis, Carnegie stages 16 and 17. Attention was focused on the dorsal wall of the telencephalon to the ventricular zone of proliferation and to the postmitotic zone with beginning of neuronal migration. We identified apoptotic cells in tissue sections by propidium iodide staining, TUNEL and immunohistochemistry for Fas(APO-1/CD95). We determined the distribution and the percentage (reported to the propidium iodide stained nuclei) of apoptotic TUNEL-positive and Fas(APO-1/CD95)-positive cells. TUNEL-positive apoptotic cells in the proliferative zone were 20% in stage 16 and 60% in stage 17. TUNEL-positive apoptotic cells in the postmitotic zone were 8% in stage 16 and 30% in stage 17. CD95-positive apoptotic cells in the proliferative zone were 5% in stage 16 and 2% in stage 17. There were no CD95-positive cells in the postmitotic zone. We evidentiated the presence of the suicide receptor Fas(APO-1/CD95) only on a small population of apoptotic neuroblasts in the proliferative zone. The differences between apoptotic distribution and receptors in early corticogenesis suggest that different apoptotic pathways drive the selection of neuronal populations. 相似文献
20.
Phosphatidylserine (PS) exposure on propidium iodide negative cells using FITC labelled annexin-V has been used to quantify apoptosis in vitro and in vivo. Detection of PS within cells undergoing necrosis is also possible if labelled annexin-V specific for PS enters the cell following early membrane damage. Necrotic or late apoptotic cells can be excluded from flow cytometric analysis using propidium iodide which enters and stains cells with compromised membrane integrity. Here we show that thymocytes undergoing death exclusively by necrosis show early exposure of PS prior to loss of membrane integrity. This early exposure of PS occurs in cells treated with agents which both raise intracellular calcium levels and are also capable of interacting with protein thiol groups. We also demonstrate that PS exposure in thymocytes induced to undergo apoptosis by three different agents does not correlate with calcium rises but correlates with and precedes DNA fragmentation. 相似文献