Objective To explore the relationship between drug resistance of leukemic cells and Caspase-3,this study took adriamycin(ADR)-resistant human chronic granulocytic leukemic cell strain K562/AO_2 as research subject,observing the cell survival and the morphological change of cell apoptosis under the action of ADR and arsenic sulfide and the Caspase-3 activity before and after putting in the Caspase-3 inhibitor.Methods ① The 3-(4,5-dimethylthiazo-2-yl)-2,5-diphenyl-tetrazolium bromide(MTT) method was used to determine the cell survival(A value) of K562/AO_(2)cell strain under the action of ADR and arsenic sulfide.② DNA agarose gel electrophoresis was performed to observe the DNA cleavage of apoptotic cells.③ The enzyme colorimetric activity assay(CAA) method was used to measure the change of the Caspase-3 activity of K562/AO_(2) cell strain.Results ① The A value of K562/AO_(2) cells had a time and dosage dependent relation with arsenic sulfide.② Apoptosis occurred in the K562/AO_(2) cell strain affected by arsenic sulfide.③ Compared with the cell strains with the Caspase-3 inhibitor added,the Caspase-3 activity of those without the Caspase-3 inhibitor increased remarkably(P

Purpose To explore the pathological feature of urinary exfoliated cell examination and influence factors by retrospectively comparing the coincidence of diagnosis between urinary exfoliated cell examination and histopathologic results of cystoscopic biopsy. Methods 735 patients underwent both urinary exfoliated cell examination and histopathologic biopsy of cystoscope evaluation from No-vember 2010 to July 2014 in Peking University Shougang Hospital were enrolled in this study. The urinary exfoliated cells were treated with Pap staining, while the histopathologic biopsy were dealt with HE staining. All cases were divided into three groups according to the diagnosis of urinary exfoliated cell examination:negative group ( no cancer or atypical cell detected) , suspicious group ( atypical cell detected) and positive group ( cancer cell detected) . These above diagnoses were confirmed with the histopathologic biopsy. ROC curve analysis, Cochran-Armitage trend test and logistic regression analysis were performed to evaluate the sensitivity and the specificity of urinary exfoliated cell examination as well as the relationship between diagnoses with age and sex. Results The age range of 735 patients (551 male and 184 female) was 28 ~91 years and the median age was 69 years. There were 187 patients in the positive group, including 184 malignant and 3 false-positive cases. The suspicious group, including 186 cases, consisted of 67 malignant, 119 benign reactive changes. Of all 362 cases in the negative group, malignant tumor was detected in 90 cases. For histologic diagnosis, the AUC of ROC(95%CI)was 0. 800 (0. 767~0. 834), displaying significant difference as compared to the histological pathological diagnostic results(P<0. 001). As the cyto-histologic diagnostic level elevated from negative, suspicious to positive, the results of Co-chran-Armitage trend test showed significant differences(Z=15. 83, P<0. 001). If standardized with the histopathologic biopsy re-sults, the AUC (area under curve) of urinary exfoliated cell examination was 0. 800 (0. 767~0. 834) in ROC curve analysis was sig-nificantly larger (P<0. 001). Furthermore, we also found in Logistic regression that the incidence of cancer was 1. 04 (1. 03~1. 05) times higher if aged one year older ( P<0. 001 ) , while there was no significant relationship between the incidence and the sex ( P=0. 655). Conclusions The coincidence rate of urinary exfoliated cell examination increases with the malignant degree. A positive cor-relation is detected between age and the incidence of malignant tumor. Detailed clinical material can markedly improve the sensitivity and accuracy of cyto-histologic diagnosis.

Aim To investigate the modulation effects of NaHS on arterial vasodilatation functions of renal hy-pertensive rats .Methods Two-kidney , one-c lip ( 2K1C ) renovascular hypertension was induced .Rats were randomly divided into four group:sham group , two-kidney one-clip model ( 2K1C ) group, 2KIC +NaSH( H2 S donor ) group, PPG group.The systolic blood pressure ( SBP ) was measured before the opera-tion and each week after the operation .The carotid ar-tery was collected for morphometric parameters ( outer radius, wall thickness, the radio of wall thickness to outer radius) and the tension of the carotid artery was observed with the isolated artery ring technique .Immu-nohistochemistry was used to determine the protein ex-pression of eNOS , ET-1 protein in carotid artery .Re-sults The blood pressure of 2K1C group and PPG group was higher than that of sham group ( P<0.05) . Compared with 2K1C group,the blood pressure and the rat arteria carotis communis of the radio of wall thick-ness to outer radius of 2K1C+NaHS group decreased significantly , while the relaxation of carotid artery to ACh in NaHS group increased .According to the immu-nohistochemistry results , eNOS expression was upregu-lated while ET-1 was downregulated in 2K1C +NaHS group as compared with 2K1C group.Conclusions Chronical administration of NaHS can decrease blood pressure in renovasocular hypertensive rats .The anti-hypertensive effect of H 2 S maybe associated with im-provement of the arterial functions .

BACKGROUND: Metabotropic glutamate receptor(mGluR) is G-protein coupled membrane receptors, which participate in various physiology or pathology process in brain, but how it induce brain ischemic tolerance(BIT)is unclear.OBJECTIVE: To study roles of mGluR2/3 and mGluR1/5 in the BIT induction.DESIGN: Randomized controlled study based on experimental animals.SETTING: Neurological department of provincial hospital and pathophysiological department of basic institute in a university.MATERIALS: The study was conducted at the Pathophysiological Department, Institute of Basic Medicine, Hebei Medical University from May 2002 to May 2003. Totally 64 healthy male SD rats were selected from the Experimental Animal Center of Hebei Medical University. Glial fibrillary acidic protein (GFAP) antibody, MTPG and(s)-4C3HPG were got from Sigma Company.INTERVENTIONS: 4 vessel occlusion(4VO) brain ischemic models in rats stained with thionine staining and GFAP immunohistochemistry staining. were used. Sixty-four rats, of which bilateral vertebral arteries were occluded permanently by electrocautery, were divided into the following 8groups: sham operation group, cerebral ischemic preconditioning(CIP)group, ischemic insult group; BIT group; MTPG + sham operation group;MTPG+BIT group; MTPG+ischemia group and(s) -4C3HPG+BIT coup. All the rats were killed 7 days after the operation or the final ischemic treatment. Cerebral sections were selected and stained with thionine staining and GFAP immunohistochemistry staining.MAIN OUTCOME MEASURE: The changes of the morphologic hippocampal pyramidal cell and GFAP expression of astrocyte.RESULTS: ① The 8 minutes ischemic insult increased the histological grade(HG) in CA1 area, decreased the pyramidal neuronal density(ND)and increased the expression of GFAP significantly( P ＜ 0.05) . ② The above changes were not observed in the BIT group, indicating that the CIP could protect pyramidal neurons against the 8-minute ischemic insult. ③The protective effects of the CIP were blocked by MTPG or(s)-4C3HPG, as manifested by significant increases in HG and decreases in ND in the groups of MTPG + BIT, MTPG + ischemia and(s)-4C3HPG + BIT( P ＜ 0.05).CONCLUSION: MTPG or (s) -4C3HPG could block the induction of BIT induced by CIP, but mGluR2/3 or mGluR1/5 could participate in the induction of BIT by which protect effect of mGluR is further induced.

AIM: To explore the role of NO/ inducible nitric oxide synthase (iNOS) in the metabotromi glutamate receptor 2/3C (mGluR2/3) mediated-brain ischemic tolerance induced by cerebral ischemic preconditioning (CIP), and to observe the influences of α-methyl- (4-tetrazolyl- phenyl) glycine (MTPG), an antagonist of mGluR2/3, on the expression of iNOS during the induction of brain ischemic tolerance. METHODS: Thirty-six Sprague-Dawley rats were subjected to four vessel occluding global brain ischemic model. Thionin staining and immunohistochemistry were used for neuropathological evaluation and assay of iNOS expression in the hippocampal CA1 subregion of the rats. RESULTS: In the sham group, weak expression of iNOS was detected. The expression of iNOS in the CIP and CIP+ischemic insult groups were increased significantly compared with that in the sham group. Administration of MTPG via lateral cerebral ventricle 20 min before CIP blocked the up-regulation of iNOS induced by CIP, but had no influence on the pyramidal neuron survival. However, in the MTPG+CIP+ischemic insult group, the expression of iNOS was extremely intensive compared to that in CIP and MTPG+CIP groups. Importantly, this up-regulation was accompanied with obvious delayed neuronal death. CONCLUSION: NO/iNOS pathway plays an important role in the process of mGluR2/3 mediated-brain ischemic tolerance induced by CIP.

AIM:To investigate the formation of membrane pore in PC 12 cells induced by exogenous adenosine triphosphate ( ATP) and to identify the key molecular targets .METHODS:PC12 cells were treated with different concen-trations of ATP to establish the injury model .The morphological change was observed under an inverted phase -contrast mi-croscope.The viability of the PC12 cells was measured by CCK-8 assay.Fluorescent dye YO-PRO-1 was used to detect the membrane permeability.The expression of P2X7 receptor and pannexin 1 (Panx1) at mRNA and protein levels was as-sessed by real-time PCR and Western blotting .RESULTS:After exposed to ATP (1 mmol/L, 3 mmol/L and 5 mmol/L) for 3 h, the PC12 cells became edematous , and the number of adherent cells decreased gradually in a dose-dependent man-ner .The cell viabilities in 3 mmol/L ATP group and 5 mmol/L ATP group were significantly decreased compared with con-trol group (P<0.05).YO-PRO-1 uptake in the PC12 cells exposed to ATP (0, 1, 3 and 5 mmol/L) for 15 min, 30 min and 60 min increased in a dose-dependent and time-dependent manner .The cell viability increased and the intracellular fluorescence intensity induced by ATP were significantly antagonized in brilliant blue G ( a P2X7 receptor inhibitor ) pre-treatment group (P<0.05), whereas it did not change in carbenoxolone (a Panx1 inhibitor) pretreatment group (P>0. 05).The expression of P2X7 receptor at mRNA and protein levels was significantly increased (P<0.05), but the expres-sion of Panx1 was not changed ( P>0.05) when PC12 cells were exposed to ATP for 3 h.CONCLUSION:Extracellular ATP at high concentration may induce membrane pore formation with the expression and activation of P 2X7 receptor in PC12 cells.

Aim To investigate the protective effects of hydrogen sulfide ( H2 S) on focal cerebral ischemia/reperfusion injury in rats and the possible mechanisms. Methods Male Sprague Dawley rats were divided into three groups randomly: sham-operated group, cerebral ischemia/ reperfusion ( I/R) group and sodium hydro-sulfide ( NaHS ) + I/R group. The left temporary middle cerebral artery occlusion ( MCAO ) model was established by the line-embolism method. After rats were suffered 2h/24h ischemia/reperfusion stress, the mortality rate was evaluated, and the nervous function-al defect degree was evaluated by Longe scoring, the volumes of cerebral infarction was evaluated by 2 ,3 ,5-triphenyltetrazolium chloride ( TTC) staining, and the expression of P2X7 receptor protein in brain tissue was detected by the immunofluorescence method. Results The mortality rate in NaHS + I/R rats ( 29.41%) was obviously lower than those of I/R group ( 42 . 86%) . The nervous defect scores in NaHS + I/R rats were significant lower than those of I/R group ( P <0.05 ) . The volumes of cerebral infarction in NaHS +I/R group (21.88% ±3.53%) were significant lower than those of I/R group ( 36.71% ±3.73%) ( P <0.01 ) . The results of immunofluorescence showed that the positive expression cells of P2X7 receptor protein in cerebral cortex and hippocampal CA1 area of I/R group were significantly higher than those of sham-op-erated group(P<0. 01). However, compared with I/R group, the positive expression cells of P2X7 receptor protein in cerebral cortex and hippocampal CA1 area of NaHS + I/R group were significantly decreased ( P<0. 01). Conclusions H2S exerts the neuroprotective effect on focal cerebral ischemia/reperfusion injury in rats, and the protective mechanism might be associated with down-regulating the expression of P2X7 receptor protein in brain tissue.

[ABSTRACT]AIM:ToexaminetheeffectsofhighconcentrationofextracellularATPonhumanneuroblastoma SH-SY5Y cell injury.METHODS: Cultured SH-SY5Y cells were grouped according to the concentrations of ATP and treatment time.The cell viability was detected by CCK-8 assay.The variation of autophagic vacuoles was observed with monodansylcadaverine staining .The cell apoptosis was analyzed by Hoechst 33258 staining.Meanwhile, apoptotic rate was detected by flow cytometry .The levels of caspase-3 and microtubule-associated protein 1 light chain 3-Ⅱ ( LC3-Ⅱ) were determined by Western blotting .RESULTS:Compared with control group , the survival rate of SH-SY5Y cells was signifi-cantly reduced by ATP at different concentrations (3, 6, 9, 12 and 15 mmol/L for 3 h) and different treatment time (1, 2, 3 and 6 h with 6 mmol/L ATP, peaking at 3 h).The autophagic vacuoles of SH-SY5Y cells were significantly increased at 1 h with ATP treatment , trended to decrease over time and returned to control level at 6 h.The protein expression of LC3-Ⅱwas significantly increased at 1 h with ATP treatment , which was consistent with the time points of increasing auto-phagic vacuoles .LC3-Ⅱexpression level gradually decreased at 2~3 h with ATP treatment , and returned to control level at 6 h.Compared with control group , the apoptotic rate and the expression level of caspase-3 were enhanced synchronously . The peak of apoptotic rate occurred at 3 h, and kept until 6 h.The level of cleaved caspase-3 expression peaked at 6 h. CONCLUSION:High concentration of extracellular ATP induces the autophagy and apoptosis of SH -SY5Y cells.The in-creased autophagy shows up , followed by the climax of apoptosis until 6 h.With the prolonged duration of ATP , apoptosis is the main process in the cells .

Objective To assess the value of double balloon enteroscopy (DBE) in etiological diagnosis of small bowel ulcer.Methods The clinical and endoscopic data of patients undergoing DBE at the Department of Gastroenterology, Renmin Hospital of Wuhan University from January 2010 to January 2016 were reviewed.For patients with small bowel ulcer revealed by DBE, the etiologic diagnosis was retrospectively analyzed.Results One hundred and three patients with small bowel ulcer were included in the analysis.The initial diagnosis based on morphology of ulcer was Crohn's disease (58.3%, 60/103), intestinal tuberculosis (18.4%,19/103), etiology unknown (24.3%, 25/103).The final diagnosis comprehensively based on pathology, clinical data and disease outcome was Crohn's disease (44.7%,46/103), intestinal tuberculosis (18.4%,19/103), lymphoma (3.8%,4/103), eosinophils enteritis (1.9%,2/103), non-steroidal anti-inflammatory drug enteritis (1.9%,2/103), adenocarcinoma (1.0%,1/103), stroma tumor (1.0%,1/103),ischemic enteritis (1.0%,1/103), vascular malformation (1.0%,1/103), Henoch-Schonlein purpura (1.0%,1/103), etiology unknown (24.3,25/103).The accurate diagnosis rate and misdiagnosis rate for Crohn's disease by DBE were 73.3%(44/60)and 10.0%(6/60).The accurate diagnosis rate and misdiagnosis rate for intestinal tuberculosis by DBE were 14/18 and 2/18.Conclusion There are diversity and complexity in etiologic diagnosis of small bowel ulcer.The diagnosis should be based on the comprehensive analysis of morphology of ulcer, endoscopic and surgical pathology and clinical data.Follow up of small bowel ulcer by DBE may provide important information for etiologic diagnosis.