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Objective To evaluate the effect of morphine preconditioning on the expression of miR-133b-Sp and Fas in rat cardiomyocytes subjected to hypoxia/reoxygenation (H/R).Methods Cardiomyocytes were isolated from healthy adult male Sprague-Dawley rats by using Langendorff perfusion.The cells were seeded into 24-well plates or 60 mm diameter dishes and randomly divided into 3 groups (n =24 each) using a random number table:control group (group C),group H/R,and morphine preconditioning group (group MPC).The cells in group C were cultured in normal culture atmosphere.In H/R and MPC groups,the cells were exposed to 95％ N2-5％ CO2 for 90 min followed by 120 min reoxygenation.In group MPC,the cells were cultured for 10 min in serum-free DMEM liquid culture medium containing morphine 1 μmol/L,and then were cultured for 30 min in morphine-free DMEM liquid culture medium before hypoxia.At 120 min of reoxygenation,the cells in 24-well plates were selected to detect the cell viability (by MTT),lactate dehydrogenase (LDH) activity in the culture medium,and cell apoptosis (by Hoechst 33234 staining).Apoptosis rate was calculated.Total RNA and protein were extracted from the cells in 60 mm dishes to detect the expression of miR-133b-5p and Fas mRNA (by quantitative real-time PCR) and Fas protein (by Western blot).Results Compared with C group,the cell viability was significantly decreased,LDH activity and apoptosis rate were increased,the expression of miR-133b-Sp was down-regulated,and the expression of Fas mRNA and protein was up-regulated in H/R group.Compared with H/R group,the cell viability was significantly increased,LDH activity and apoptosis rate were decreased,the expression of miR-133b-5p was up-regulatcd,and the expression of Fas mRNA and protein was down-regulated in MPC group.Conclusion The mechanism by which morphine preconditioning reduces H/R injury to rat cardiomyocytesis related to up-regulation of the expression of miR-133b-Sp and down-regulation of the expression of Fas.
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Objective To assess the diagnostic value of cytokeratin 18 fragment M30 (CK18-M30) and Fas in patients with nonalcoholic fatty liver disease (NAFLD),especially nonalcoholic steatohepatitis (NASH).Methods Among 58 patients with NAFLD,36 patients with NAFLD received liver biopsy.According to NAFLD activity score (NAS) and liver fibrosis score,patients were divided into NASH group (24 cases) and non-NASH group (12 cases).And at the same period,15 healthy individuals were set as healthy control group.The serum level of CK18 M30 and Fas were measured with enzyme-linked immunosorbant assay (ELISA).Rank sum test was performed to analyze the differences in the level of CK18-M30 and Fas between groups.The diagnostic value of CK18 M30 and Fas were assessed by the receiver operating characteristic (ROC) curves.Results The level of serum CK18-M30 of NAFLD group was significantly higher than that of healthy control group (97.24 U/L (86.06 to 113.12 U/L) vs 78.41 U/L (74.29 to 80.76 U/L),Z=-4.206,P＜0.01)).The level of serum CK18-M30 of NASH group was higher than that of non-NASH group (111.06 U/L (94.30 to 142.68 U/L) vs 89.00 U/L (83.56 to 106.50 U/L),Z=-2.233,P＜0.05)).The area under the ROC curve (AUC) of CK18-M30 in the diagnosis of NASH was 0.73 (0.56,0.90),and the sensitivity and specificity of CK18-M30 in diagnosis of NASH was 79.2％ and 58.3％,respectively.The AUC of Fas in diagnosis of NASH was 0.58 (0.38,0.77),while the sensitivity and specificity of Fas in diagnosis of NASH was 54.2％ and 66.7 ％.The serum level of Fas increased in FAFLD group compared with healthy control group,and in NASH group compared with non-NASH group,however the differences were not signifincant (both P＞ 0.05).Conclusions The level of CK18-M30 has certain value in the diagnosis of NASH.The diagnostic value of Fas in NASH needs more samples in further study.
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Objective To investigate the effects of penehyclidine hydrochloride on Fas/FasL expression during acute lung injury induced by blunt chest trauma-hemorrhagic shock and resuscitation (HSR) in rats.Methods Thirty male SPF Sprague-Dawley rats, aged 8 weeks, weighing 245-275 g, were randomly assigned into 3 equal groups using a random number table: sham operation group (group Sham) , blunt chest trauma-HSR group (group THSR) and penehyclidine hydrochloric group (group PHCD).The model of acute lung injury induced by blunt chest trauma-HSR was induced by dropping a 300 g weight onto a precordium in anesthetized rats.Blood was withdrawn via the femoral artery 5 min later until mean arterial pressure was decreased to 35-45 mmHg within 15 min, and maintained at this level for 60 min, followed by resuscitation.In PHCD group, PHCD 2 mg/kg was injected intravenously at 60 min after hemorrhagic shock.At 6 h after the model was established, the rats were sacrificed, the lungs were then removed for microscopic examination of pathologic changes and for determination of Fas, FasL and caspase-8 expression, and interleukin-6 (IL-6) and IL-1β contents in lung tissues.Apoptotic index was calculated.Results Compared with group Sham, the expression of Fas, FasL and caspase-8 was significantly up-regulated, and AI and contents of IL-6 and IL-1β were increased in THSR and PHCD groups (P＜O.05).Compared with group THSR, the expression of Fas, FasL and caspase-8 was significantly down-regulated,and AI and contents of IL-6 and IL-1β were decreased in group PHCD (P＜0.05).The pathologic changes of lungs were significantly reduced in group PHCD compared with group THSR.Conclusion The mechanism by which penehyclidine hydrochloride inhibits lung cell apoptosis induced by blunt chest trauma-HSR is associated with inhibition of Fas/FasL expression in rats.
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Objective To evaluate the role of Fas/FasL signaling pathway in ulinastatin postconditioning-induced attenuation of apoptosis in the myocardial cells of patients undergoing cardiac valve replacement with cardiopulmonary bypass (CPB).Methods Forty patients of both sexes,aged 21-59 yr,of ASA physical status Ⅱ or Ⅲ (NYHA class Ⅱ or Ⅲ),scheduled for elective cardiac valve replacement with CPB,were randomly divided into 2 groups (n =20 each):control group (group C),and ulinastatin postconditioning group (group U).In group U,ulinastatin 10 000 U/kg was perfused via the aortic root at 4 000-5 000 U·kg-1 ·min-1 starting from 5 min before aortic unclamping.In group C,the equal volume of normal saline was infused instead of ulinastatin.Myocardial specimens were taken from the right auricle at 45 min after aortic unclamping for determination of Fas,Fas ligand (FasL),caspase-8,Bcl-2 and Bax expression and cell apoptosis.The ratio of Bcl-2 expression to/Bax expression (Bcl-2/Bax) and apoptotic index were calculated.Results Fas,FasL,caspase-8 and Bax expression and apoptotic index were significantly lower,and Bcl-2 expression and Bcl-2/Bax were higher in group U than in group C.Conclusion Ulinastatin postconditioning attenuates apoptosis in the myocardial cells through inhibiting Fas/FasL signaling pathway in the patients undergoing cardiac valve replacement with CPB.
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Fas ligand (FasL) is critical for maintaining immune privilege status of central nervous system in physiological conditions.FasL binds Fas via extrinsic pathway mediated ischemic penumbra neuronal apoptosis after the onset of cerebral ischemia.In addition,FasL is involved in the regulation of T lymphocyte subsets by inducing the expression of inflammatory cytokines and chemokines,promoting intracerebral microglia activation and peripheral neutrophil infiltration,and thus aggravating immune inflammatory response after stroke.FasL-Fas system has a certain role for promoting the recovery of neural function during the convalescence of ischemic stroke.Taking FasL-Fas system as a target,it may become a new way for brain protection treatment of ischemic stroke.
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Objective To observe the changing levels of serum sFas before and after intravenous i mmunoglobulin (IVIG) treatment of incomplete Kawasaki disease (IKD),to explore the roles of sFas in the pathogenesis of IKD and IVIG treatment mechanism.Methods Thirty eight cases of IKD children were selected as experimental group and 20 examples of the same age of children as the control group.The IKD children were treated by IVIG in combination with aspirin (ASP) ; and blood test was performed before treatment,3 days after treatment,and 14 days after treatment,respectively.Dual-resistant sandwich enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of serum sFas,plasma Fibrinogen (PT-D),d-dimer (D-D),and c-reactive protein (CRP).Results The levels of serum sFas,PT-D,D-D,and CRP were significantly higher than the control group for IKD children before treatment[(0.55 ± 0.14)ng/L vs (0.24 ±0.04) ng/L,(552.3 ± 147.2) mg/dl vs (277.3 ±82.5)mg/dl,(649.0 ±201.6) μg/L vs (315.4 ±91.8)μg/L,and(72.2 ±28.7)mg/L vs (7.2 ±2.9)mg/L; t' =12.41,9.11,8.64,13.82;All P ＜ 0.05] ;3 days after treatment,compared with those before treatment and control group,the sFas level of IKD children at the third day after treatment was significantly decreased compared to that before treatment and control groups,respectively [(0.43 ± 0.09) ng/L vs (0.55 ± 0.14) ng/L,(0.24 ± 0.04) ng/L,F =47.624,All P ＜0.05] ;For the level of sFas at the 14th day after treatment,no statistical significance was found between IKD children and the control group[(0.24 ±0.05) ng/L vs (0.24 ±0.04) ng/L,t =0.596,P ＞ 0.05].Conclusions The abnormally increased serum sFas level before IVIG treatment suggests that dysfunction of apoptosis be involved in the pathogenesis of the IKD.Intravenous immunoglobulin treatment may be involved in the apoptosis process.
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BACKGROUND:The clinical research have found that the interbervebral disc herniation often occurs in several members or even al the members of a family, and the location, reason and symptom are basical y the same, indicating that genes play an important role in this kind of disease. OBJECTIVE:To analyze the apoptosis Fas gene expression characteristics of lumbar disc in the familial patients with intervertebral disc herniation. METHODS:Semi-quantitative reverse transcription-PCR was used to test Fas gene expression of vertebral pulp and cartilage endplate in the intervertebral disc among 15 familial patients, 21 ordinary patients and five fresh cadavers. RESULTS AND CONCLUSION:Fas gene expression level of endplate of familial and ordinary patients with intervertebral disc herniation was higher than that of fresh cadavers, and there was no significant difference (P0.05). Compared with the vertebral pulps of ordinary patients with intervertebral disc herniation and fresh cadavers, there was no significant difference in the Fas expression of vertebral pulps of familial patients with intervertebral disc herniation (P>0.05). The increasing Fas gene expression may be secondary in the endplates of familial patients with intervertebral disc herniation, which can prevent intervertebral disc degeneration through preventing the endplate degeneration.
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ObjectiveTo compare the levels of sFas in the sera among Kawasaki disease (KD),incomplete Kawasaki disease (IKD),and normal control groups,and to analyze the relationship of sFas with IKD children.MethodsA total of 32 cases of acute KD and acute IKD children,and 20 cases of the control children were selected,respectively.The levels of serum sFas among three groups were measured using ELISA kits.Each child among the three groups was examined by echocardiography.Results(1)The levels of serum sFas among the three groups were[ (0.54±0.20)ng/L in KD,(0.55±0.16)ng/L in IKD,and (0.24 ± 0.04) ng/L] in control group,respectively.The overall means of sFas in the KD and IKD groups were higher than the control group,and the differences were statistically significant( F=29.276,P＜0.05 ).(2)The levels of serum sFas among echocardiography abnormal and normal groups were[ (0.65±0.19) ng/L and (0.49±0.10)ng/L],respectively; and the difference between two groups were statistically significant ( t=3.139,P ＜ 0.05 ).ConclusionsThe expression levels of sFas in the peripheral serum of IKD children were increased,and there was a close association of overexpression of sFas with the cardiovascular damage in IKD children.
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Objective To investigate the effects of arsenic on liver Fas/FasL expression in mice and to observe the antagonize effect of zinc. Methods Sixty health Kunming mice were divided into five groups according to their body mass: negative control group(no arsenic and no zinc), arsenic group(55 mg/L NaAsO2 solution), low dose zinc intervention group(55 mg/L NaAsO2 solution and 20 mg/L ZnSO4 solution), middle dose zinc intervention group (55 mg/L NaAsO2 solution and 40 mg/L ZnSO4 solution), and high dose zinc intervention group (55 mg/LNaAsO2 solution and 80 mg/L ZnSO4 solution). Everyday the solution was given by oral gavage at a dose of 0.02 ml/g body weight for six weeks. The expression of Fas/FasL in mice liver was examined by immunohistochemistry.Results With the amount of zinc increasing, the expression of both Fas and FasL in mice liver decreased gradually. The expression rates of Fas[83.33%(10/12), 50%(6/12)] and FasL[66.67%(8/12), 41.67%(5/12)]in low dose zinc intervention group and middle dose zinc intervention group, respectively, were different from the expression rate of Fas[8.33%(1/12)] and FasL[0.00%(0/12)] in the negative control group(all P < 0.05). The Fas expression rate of middle dose zinc intervention group[50%(6/12)] and the high dose zinc intervention group [25%(3/12)] was compared with the arsenic group[8.33%(1/12)], and the difference was statistically significant (all P < 0.01 ). The FasL expression rate of the middle dose zinc intervention group [41.67% (5/12 )] and the high dose zinc intervention group[16.67%(2/12)] was compared with positive control group[91.67%(11/12)], and the difference was statistically significant (all P < 0.05). Conclusions Arsenic can increase the expression of Fas and FasL in mice liver, and promote apoptosis in liver, zinc may antagonize the effect of arsenic.
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Objective To investigate the relation between Fas-mediated apoptosis and glucocorticoid treatment in myasthenia gravis (MG). Methods In 17 patients with MG, 6 patients received glucocorticoid treatment (glucocorticoid treatment group),and 11 patients were treated without glucocorticoid (nonglucocorticoid treatment group). Meanwhile, 13 healthy cases were selected as healthy control group. CD4,CD8 and Fas expressions in peripheral blood T lymphocyte were detected by flow cytometry in three groups and analyzed. Results The percentage of CD4-CD8+ cells in peripheral blood T lymphocyte in glucocorticoid treatment group was significantly higher than that in healthy control group[(36.75 ± 11.56)% vs. (26.31 ±9.00)%, P = 0.027], while the percentage of CD4-CD8- cells was significantly lower [(30.56 ± 9.72)% vs.(42.96 ± 11.54)%, P =0.018]. The percentage of CD4-CD8+ cells in peripheral blood T lymphocyte in glucocorticoid treatment group was significantly higher than that in non-glucocorticoid treatment group [(36.75 ± 11.56)% vs. (25.24 ±7.63)% ,P =0.019]. The percentages of Fas+ and CD8 +Fas+ cells in peripheral blood T lymphocyte in glucocorticoid treatment group were significantly higher than those in healthy control group[(46.10 ± 7.13)% vs. (31.22 ± 13.00)%, P=0.006; (62.86 ± 12.29)% vs. (45.59 ±11.50)%, P = 0.003]. The percentage of CD8+ Fas+ cells in peripheral blood T lymphocyte in glucocorticoid treatment group was significantly higher than that in non-glucocorticoid treatment group [(62.86 ± 12.29)%vs (50.84 ± 8.31 )%, P = 0.034]. Conclusions Glucocorticoid treatment may have influence on peripheral blood T lymphocyte subsets in patients with MG. Fas-mediated apoptosis may be involved in the mechanism of glucocorticoid treatment in MG.
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Objective To investigate the changes of serum soluble Fas (sFas) and soluble Fas-ligand (sFasL), and the relationship betweenthe level of serum sFas or sFasL and the infarct volume in patients with acute cerebral infarction (ACI). Methods Sixty patients with ACI (female 28, male 32) served as study group and 30 healthy subjects (female 18, male 12) served as control group. An enzyme-linked immunosorbent assay was used to detect the levels of serum sFas and sFasL in both groups, and the differences of the sFas and sFasL concentration were compared between the two groups. Results The levels of serum sFas at 48 hours, at day 7 and 14 in the ACl group were 6. 27 ± 1.48 ng/L, 4. 99 ± 1.15 ng/L, and 3.74 ± 0.58 ng/L,respectively, and they were all significantly higher than 3.00 ± 0. 38 ng/L in the control group (P <0. 05). The levels of serum sFasL at 48 hours, at day 7 and 14 in the ACI group were 4.40 ± 1.32 ng/L, 3. 19 ± 0.94 ng/L, and 1.91±0.45 ng/L, respectively. They were significantly higher than 1.15 ±0.21 ng/L in the control group (P<0.01). The levels of sFas (1.91 ± 0.45) ng/L, respectively, and they were all significantly higher than (4.98 ±0.91) ng/L(t = 12.12 ,P <0. 01)and (3.58 ±0. 87) ng/L(t =5.35 ,P <0.01) in the small infarction group. The levels of serum sFas and sFasL in patients with ACI showed positive correlation (r =0. 748, P =0. 01). Conclusions High serum sFas and sFasL may indicate larger infarct volume in patients with ACI.
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Objective:To assess the expression levels of CD95 and OX40 ligand(OX40L)messenger RNA(mRNA)in hepatocellular carcinoma(HCC),and their clinical values thereof.Methods:The lymphocytes of research objects were mixed with corresponding fluorescence labeled monoclonal antibody(McAb);CD95 expression by CD3 positive T ceils was quantitatively measured by dual color flow cytometry.The expression of OX40L mRNA was detected in peripheral blood mononuclear cells by fluorescence quantitative reverse transcription polymerase chain reaction(FQ-RT-PCR).Results:The CD95 expression by CD3 positive T cells was significantly higher in patients with HCC(34±20)% compared with that of normal controls (20±7)%(t=2.960,P ＜ 0.01).The expression level of OX40L mRNA was significantly decreased in patients with HCC compared with that of normal controls(t=2.302,P ＜ 0.05).Conclusion:These results suggest that abnormal expressions of CD95 and OX40L play crucial roles in peripheral blood of HCC patients at the stage of human hepatocareinogenesis.
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Objective To study the expression of PTEN,Fas and DCR3 in human gastric cancer and evaluate their effects on carcinogenesis,cell proliferation,invasion and metastasis of gastric carcinoma,as well as clinical significance.Methods The expression of PTEN,Fas and DCR3 were detected by immunohistochemical method (streptavidin-peroxidase) in the tissues of 75 primary gastric carcinomas and 15 gastric ulcers.Results The positive expression rate of DCR3 in gastric carcinoma tissues was significantly higher than that in normal tissues,and the expression of PTEN and Fas was obviously lower than that in normal tissues (P<0.01).The expression of DCR3 was negatively correlated with Fas and PTEN respectively (r=-0.720,P<0.001; r=-0.336,P<0.001).There was a positive correlation between PTEN and Fas.The expression of PTEN,Fas and DCR3 was closely correlated with lymph node metastasis,clinical stage,histological differentiation grade and tumor depth of gastric cancer,but not related to sex,age and tumor size (P>0.05).Conclusion PTEN,DCR3 and Fas have mutual restrictive relationships in the process of invasion and metastasis in gastric cancer.Detecting the expression of PTEN,Fas and DCR3 in gastric cancer is helpful to judge the malignant biological behavior of gastric cancer at a different angle,which may be considered as reliability index in clinic diagnosis,judging curative effect and monitoring the prognosis of gastric carcinoma.
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Objective To investigate the expression of caspase-3 and FasL in the hippocampus of the infantile rats with recurrent sei-zures. Methods 72 of 20-day-old Sprague-Dawley rats were randomly divided into two groups, control group and seizure group. Seizures in rats were induced by inhalant flurothyl daily in six consecutive days. Brain tissue was sampled at different time points (the 1st day, 3rd day and 7th day) after last seizure. The expressions of caspase-3 and FasL proteins in the hippocampus were detected by immunohistochemistry, and the expression of caspase-3 mRNA was measured by reverse transcription- polymerase chain reaction (RT-PCR). Results The caspase-3 protein, FasL protein and caspnse-3 mRNA levels were obviously increased at the 1st, 3rd and 7th day after recurrent seizure in the hippocampus of the rat(P<0.01). Conclusions Caspase-3 and FasL are participated in the infantile brain injury after recurrent seizures.
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Objective To observe the protective effect of magnesium gluconate on myocardial apoptosis by ischemia reperfusion injury in isolated rat hearts, and study the possible mechanism. Methods The hearts of 48 Sprague-Dawely rats were isolated, linked to Lange-ndorff perfusion apparatus, and randomly divided into 3 equal groups(n = 16 each) : Control group, ischemia/reperfusion (I/R) group and magnesium gheonate group. 8 rats in each group were perfused. Control group was pedused with modified KH buffer for 110min. I/B group was perfuesd with modified KH buffer for 20 min, then exposed to iscbemia for 30 min, and then reperfused with modified KH buffer for 60 min. Magnesium gheonate group was perfumed with modified KH buffer with magnesium gluconate for 20 min, then exposed to isohemia for 30 min and then reperfused with modified KH buffer with magnesium glueonate for 60 min. Lacate dehydrogenase (LDH) and ereatine kinase (CK) in the effluent liquid from the heart were measured after reperfusion. The concentration of Ca2+ and NO in the left ventricle were determined. The other 8 rats in each group were reperfused for 120 minutes as the method described before. After repeffusion, the myoeyte apoptosis was examined by Annexin-V-FITC/PI. After the two experiments the incidence of ventrieular arrhytlunias during reperfusion was assessed. Results Compared with I/R, magnesium glueonate decreased the incidence of ventricular an'hythmias(P <0. 01). The contents of CK and LDH in the effluent liquid from the heart in magnesium glueonate group was lower than that of I/R group (P <0. 01). The contents of Ca2+ and NO in the left ventricle in magnesium gluconate group was decreased than that of I/R group (P <0. 01). The index of myocyte apoptosis were significanfly lower in magnesium glueonate group than that of I/R group (apoptosis index :27.79±1.59 vs 33.61±2.10, P < 0. 01) . Conclusion Magnesium glueonate has protective effect on myocardial isohemia reperfusion injury in rats. The protective effect may be related to decreasing myocyte apoptosis by increasing the content of NO and relieving calcium overload.
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Objective To investigate the dynamic expressions and clinical significance of CD141, CD31 and CD95 on circulating endothelial cells (CEC) in febrile and polyuria phases of patients with hemorrhagic fever with renal syndrome (HFRS). Methods Expressions of CD141, CD31 and CD95 in the peripheral blood of patients with HFRS in febrile and polyuria phases were detected by flow cytometry. Comparisons among groups were done by one-factor analysis of variance. Results The percentages of CD141+ CD31+ cells in the peripheral blood cells from patients with HFRS in febrile and polyuria phases were 9.47% ±1.98 % and 8. 26% ±1.55 %, respectively, which were both higher than that (7.05%±1.45%) in healthy controls (F=8. 42; P=0. 000 and P=0. 029, respectively), and that in febrile phase was higher than that in polyuria phase (P = 0. 048). The mean fluorescent intensity (MFI) of CD95 on CEC of HFRS patients in febrile and polyuria phases were both significantly higher than that in healthy controls (F=19. 93; P=0. 000 and P=0. 000 respectively), and that in febrile phase was higher than that in polyuria phase (P=0. 049). In the febrile phase of HFRS,the MFI of CD95+ on CEC in patients with all clinical types were all higher than that in healthy controls (F= 17. 36; all P=0. 000), and that in severe (critical) type was the highest and higher than those in mild type and moderate type (P=0. 002 and P=0. 009, respectively). Conclusion The proportion of CEC and expression of CD95 on CEC are possibly related with the phase and severity of HFRS.
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Objective To study the effect of dehydroepiandrosterone (DHEA) on the immune system by measuring the apoptosis and expression of Fas/FasL mRNA of thymocytes. Methods The cell viability of thymocytes was determined by MTT assay. Thymocyte apoptosis was detected by agrese gel electrophoresis. The expression of Fas and Fas-L was analyzed by a semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) method. Results DHEA decreased the cell viability of thymocytes and induced thymocytes apoptosis and increased the levels of mRNA of Fas/FasL. Conclusion DHEA can induce apoptosis of thymocytes through Fas/Fas-L pathway. It suggests that DHEA may play a role in certain autoimmune diseases related to apoptosis disturbance.
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icating that Fas/FasL plays an impertant role in the regulation of immune/inflammation response in the CNS, and such role does not depend on the apoptotic pathway, This article reviews its progress in research on the inflammatory response in the CNS.
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Objective To investigate the coordinate repression of angiostatin(AS)and Fas gene on human colon carcinoma LOVO.Methods Plasmid pcDNA3-AS and pcDNA3-Fas were constructed,and AS,Fas,AS and Fas gene were transfected to human colon carcinoma LOVO cells by liposome method.The expressions of target protein were detected by Western blot.The effects of transfection of AS and Fas gene on the growth of human colon carcinoma cells were detected by MTr methods.AS,Fas,AS and Fas gene were transfected to the human colon carcinoma subcutaneously implanted in nude mice by direct injection into the tumor.The tumor sizes were detected after 14 days of the first gene transfection.Results The effect of gene transfeetion in LOVO cell after twelve hours,24 h,48 h and 72 h were observed and compared with control group,co-transfection of Fas and AS gene group and Fas group significantly inhibited the growth of human colon carcinoma LOVO line cells(P＜0.01).Fourteen days after plasmid transfection,the tumor volume in group Fas and group co-transfection were signifieandy smaller than that of control,AS and Fas group(P＜0.05).The tumor volume in AS group and Fas group were significantly smaller than that of control(P＜0.05).Conclusion Angiostatin and Fas gene have coordinate repression effect on human colon carcinoma in vivo.
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Objective To establish the method for isolation and culture of porcine Sertoli cell sand detect the expression of the immune privilege-related molecules in the cultured cells. MethodsTestes were aseptically removed from the 10-to 15-days old Large White piglets. Testes were decap-sulated, minced and then digested with 0.2% (W/V) collagenase type V, 0.25% (W/V) trypsin and 0.05% (W/V) DNaseI. In the primary isolation, Sertoli cells were cultured at 37℃with 5% CO2.Inverted phase contrast microscopy and HE staining were used to observe the morphology of Sertoliceils, and the Sertoli cells were identified under an electron microscope. Viability and apoptosis ofcultured cells were measured with the AnnexinV-PI staining by flow cytometry. The expression of sox9, FasL, TGF-β and clusterin in Sertoli cells was detected by RT-PCR. The viability of long-termcultured Sertoli cells was assayed by MTT. Results In the cultured total cells, Sertoli cells accountedfor more than 90%. The apoptosis rate and mortality of Sertoli cells was (2.61±0.96)% and (2.12±0.74)% respectively. RT-PCR revealed that the expression of sox9, TGF-β and clusterin in theSertoli cells was strongly positive, but FasL was weakly positive. Viability of cultured cells measuredby MTT conformed that the sertloli cells could survive more than 21 days. Conclusion The isolationand culture methods of the neonatal porcine Sertoli cells was established. Under the culturedconditions, the Sertoli cells can express the immune privilege molecules and successfully survive at least 21 days.