In order to approach the mechanisms of Chinese drug Bu Sui Sheng Xue Instant Granules for tonifying kidney inhibiting apoptosis in the patient of chronic aplastic anemia (CAA), levels of soluble Fas (sFas), tumor necrosis factor (TNF-a), interleukin - 2 (IL - 2) before and after treatment were determined in 30 cases of CAA and Zai Zang Sheng Xue Tablet was used as control. Results indicated that expression of serum sFas, TNF - a and IL - 2 before treatment was significantly higher than those of the control group (P0. 05), and the IL-2 level still was higher than the normal (P

BACKGROUND: Results from clinical trials suggested that clopidogrel and ticlopidine had side effects of granulopenia, and aspirin could inhibit endothelial progenitor cell proliferation. There is no report of effects of these drugs on human bone marrow mesenchymal stem cells (hBMSCs) in stem cell transplantation. OBJECTIVE: To investigate the effects of antiplatelet drugs including clopidogrel, ticlopidine and aspirin on hBMSC proliferation and secretion. DESIGN, TIME AND SETTING: The cytology in vitro observation was performed at the Laboratory of Toxicology, Shanghai Municipal Center for Disease Control and Prevention from March to December 2006.MATERIALS: The second passage of hBMSCs was kindly donated from Shanghai Tissue Engineering Research & Development Center, Shanghai Ninth People's Hospital. Clopidogrel (Lot number J20040006) and ticlopidine (Lot number H19980186) were obtained from Hangzhou Sanofi-Synthelabo Minsheng Pharmaceutical CO., Ltd. Aspirin (Lot number 20050059) was obtained from Bayer Vital GmbH. METHODS: The standard culture medium consisted of DMEM-LG, 10% heat-inactivated FBS, 100 U/mL penicillin and 100 μg/mL streptomycin. After being cultured in vitro expanded out to passage 6, hBMSCs were treated with antiplatelet drugs of different concentrations and compared with control group. MAIN OUTCOME MEASURES: Cell proliferation was assessed by 3- (4, 5-dimethylthiazol -2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay, level of vascular endothelial growth factor (VEGF) of culture medium was detected by enzyme-linked immunoadsordent assay (ELISA), and surface antigens of hBMSCs were analyzed by the flow cytometry. RESULTS: A570 values of hBMSCs treated by clopidogrel or ticlopidine of 0.02,0.1,0.4,2,10,40 μmol/L were higher than control group (P < 0.01), while A570 values of aspirin group of 60, 600, 2 000 μmol/L were lower than control group(P < 0.05). Antiplatelet drugs had no obvious effect on cell surface antigens(CD34, CD105, CD106)expressed by hBMSCs. Treated by high dose clopidogrel or ticlopidine (40 μmol/L), VEGF level from hMSCs was lower than that of control group(P < 0.01), but VEGF level of low dose (0.02 μmol/L) ticlopidine group was higher than control group(P < 0.01), and there was no significantly difference of VEGF level among low dose clopidogrel group (0.02 μmol/L), aspirin group (5, 2 000 μmol/L), and control group(P > 0.05). CONCLUSION: Clopidogrel and ticlopidine improve proliferation of hBMSCs, but aspirin inhibits proliferation of hBMSCs. High dose of clopidogrel and ticlopidine suppress VEGF secretion of hBMSCs, while low dose of ticlopidine promote it. Antiplatelet drugs have no obvious effect on hBMSCs differentiation.

Seven genes (fabD, fabG, fabH, fabA, fabZ, fabB and fabI) of E.coli fatty acid biosynthetic enzymes were cloned by PCR amplifying and appropriate expression vectors were constructed. Under induction assay expression of plasmid encoded proteins was carried out in strain BL21(DE3) and seven enzymes were purified using Ni-NTA agarose resin. In the absence of [2-14C] malonyl-CoA fatty acid synthetic reaction was reconstituted in vitro by adding seven enzymes and co-factors. And several model reactions were established for identification of special fatty acid biosynthetic enzymes. Meanwhile Clostridium acetobutylicium FabZ function was characterized by this method.

Objective To investigate the expression of paired box 2 (Pax2) and E-cadherin in human renal cell carcinoma (RCC) and their correlation with clinicopathologic parameters.Methods The RCC tissue microarrays containing 85 renal cell carcinoma specimens and 35 normal kidney tissue specimens were used to detect Pax2 and E-cadherin expressions by immunohistochemistry.Results The positive expression rate of Pax2 was 77.6％ (66/85) in RCC specimens,which was significantly higher than that in 35 normal kidney tissue specimens (P ＜ 0.01).The positive expression rate of E-cadherin was 30.6％ (26/85) in RCC specimens,which was significantly lower than that in 35 normal kidney tissue specimens (P ＜ 0.01).The expression of Pax2 in RCC was significantly related to histological classification and pathological grade (P ＜ 0.05),while it was not related to the size of tumor and clinical stage (P ＞ 0.05).The expression of E-cadherin in RCC was significantly related to the size of tumor,histological classification,pathological grade and clinical stage (P ＜ 0.05).The positive expression rate of E-cadherin in survival group was significantly higher than that in death group (P ＜ 0.05).Conclusions The abnormal expressions of Pax2 and E-cadherin may play a crucial stage in development of human RCC.Detection of the expressions of Pax2 and E-cadherin can be used in the evaluation of malignant degree and prognosis of RCC.

Objective To study the activation of TLRs/NF-κB signal pathway and production of different functional cytokines during invasive pulmonary aspergillosis( IPA) , in order to probe the pathogene-sis of IPA. Methods Mouse were randomly divided into normal, normal + inoculated with Aspergillus fumigatus( normal inoculation group), and immune suppression + inoculation with Aspergillus fumigatus (IPAmodel group) , the mouse were killed at different time points after inhaling Aspergillus fumigatus spores by nose. Removing the lung tissue in a sterile manner and making pathological section respectively, counting Aspergillus fumigatus colony, dynamiclly detecting the expression of TLR2, TLR4 mRNA, variation of NF-κB p65 protein, pro-inflammatory cytokines TNF-α, IL-1β and anti-inflammatory cytokines IL-10 levels in the lung tissue by RT-PCR and Western blot method during Aspergillus fumigatus infection in mouse. Results (1) When it's 72 h after inhaling Aspergillus fumigatus by nose, IPA model emerged severe lung tissue inflammation, and generated a large number of hyphae, meanwhile, burthen of Aspergillus fumigatus was higher than normal inoculation group at each time point. (2)Compared with the normal inoculation group, IPA group whose TLR2 mRNA was low expression at early stage of infection (24 h), and emerged high expression at late stage of infection (120 h, 144 h); and TLR4 mRNA has been at a state of low expression in the infection process; NF-κB p65 suddenly increased at early stage of infection(24 h) and then continued to decline. (3) After infected by Aspergillus fumigatus in normal mouse, proinflammatory cytokine TNF-α, IL-1β in lung exhibited high expression at the early stages of infection, and the highest expression levels appeared at 48 h or 72 h, then decreased and recovered to normal level. And the expression level of anti-inflammatory cytokine IL-10 rised at late stage of infection; The IP A mouse released a lot of anti-inflammatory cytokine IL-10 at early stage of infection, which significantly reduced at late stage, and released pro-inflammatory cyto-kines TNF-α, IL-1β at slow and low level. Conclusion The abnormal activation of TLRs/NF-β signaling pathway caused the loss of dynamic balance between pro-inflammatory cytokines and anti-inflammatory cytokines, leading to the occurrence and development of IPA.

Objective To investigate a new selection criteria for lower instrumentation vertebrae (LIV) in thoracolumbar adolescent idiopathic scoliosis (AIS).Methods Fifty-two cases of fusion thoracolumbar bend and more than 1.5 years follow up were included.The cases conformed to the AIS selection criteria,i.e.,the first vertebral body touched at the scoliosis distal end in the central sacral vertical line (CSVL) of erect orthotopic X-ray film served as the touch vertebrae (TV),moreover which should meet the following requirements:the Nash-Moe rotation ≤ degree Ⅱ;CSVL located between bilateral vertebral pedicle in the concave side Bending image;no kyphosis of thoracolumbar and lumbar segments existed;the patients with the distance 3-4 mm from CSVL to TV were also included into the study.The operation of each case was carried out by an experienced spine surgeon team.The internal fixation orthopedic fusion of total pedicle screws was adopted by posterior approach.The spine full length of the anterior-posterior and lateral X-ray film and left and right sides Bending images at clinostatism were taken before operation,instantly after operation and at final follow up.The Cobb angle of main bending,trunk shift (TS) at coronal plane,LIV tilting (LIVT) and LIV distal disc angulation (LIVA) were measured.The positions of TV and stable spine were observed,recorded and performed the statistical analysis.Results All cases were followed up for more than 18 months,with an average follow-up period of (23 ± 3) months.The main bending Cobb angles before operation,instantly after operation and at final follow up were (49.32±11.37)°,(9.08 ± 6.78)°and (10.65±6.68)°respectively,LIVT were (21.76±4.68)°,(5.17±4.09)° and (5.16±3.08)° respectively and LIVA were (7.19±5.16)°,(3.16±2.78)° and (4.17±3.28)° respectively;the above indicators had statistical differences between before operation with instantly after operation and at final follow up (P＜0.05).Twenty seven cases were preoperative trunk decompensation,and 5 cases were complicating trunk decompensation at final follow up,which were not increased compared with those before operation.Comparing TV as LIV with adopting stable vertebrae as LIV,the former could save (1.42 ±0.45) fusion segments.Conclusion Adopting the new criteria to conduct thoracolumbar AIS operation saves the fusion segments as compared with commonly used method in clinic.

Objective To summarize the endoscopic appearance,endoscopic ultrasound findings and histopathological characteristics of gastric inflammatory fibroid polyp (IFP) in order to improve diagnosis of IFP.Methods From September 2011 to November 2016,49 patients with pathologically comfirmed gastric IFP,who underwent endoscopy in Taizhou Hospital of Zhejiang Province,were enrolled.The medical history,endoscopic examination,treatment and follow-up were retrospectively enalyzed.Results Among 49 patients with gastric IFP (16 males and 33 females;average age 54 years) the maximum diameter of 33 cases (67.3％) was between 1.0 cm and 2.0 cm.Forty-eight cases had single lesion and one case had multiple lesions.The lesions of 17 cases (34.7％) were located at the anterior wall of gastric antrum,the lesions of seven cases (14.3％) were at the posterior wall of gastric antrum,the lesions of seven cases (14.3％) were at the lesser curvature of gastric antrum and the lesions of seven cases (14.3％) were at the great curvature of gastric antrum.Among the lesions of 41 patients who received endoscopic ultrasonography,28 lesions were located in the submucosa of stomach,13 lesions were located in mucosa and muscularis mucosa.The rate of misdiagnosis of endoscopic ultrasonography was 29.3％ (12/41).The endoscopic ultrasound findings of the lesions included 26 hypoechoic structures,11 hyperechoic structures and four slightly hypoechoic structures.The lesions of all the patients were successfully removed by endoscopic polypectomy without any complication.Thirty-seven lesions were treated by endoscopic submucosal dissection (ESD) and 12 lesions by endoscopic mucosal resection (EMR).All the patients were not clearly diagnosed before operation and were finally diagnosed by pathological examination.Postoperative pathological examination showed that in the suhmucosa and mucosa lamina propria,spindle-shaped cells proliferated and arranged in an interwoven pattern or cells around vessels or mucosal glands formed vortex-like or onion skin like pattern.Forty-seven patients were followed up and the median follow-up time was 31 months.All patients survived withont recurrence or metastasis until the submission of this paper.Conclusions The rate of misdiagnosis of gastric IFP is high before operation,and the diagnosis is depended on histopathological examination.Endoscopic resection is the first choice because the diameter of most lesions are less than 5 cm.

AIM:To investigate the RNA oxidative damage in human gastric cancer tissue and para-carcinoma tissue for exploring the role of RNA oxidation in the occurrence of gastric cancer .METHODS:Immunohistochemical ob-servation and LC-MS/MS analysis were performed in 61 cases of gastric carcinoma .The position and concentration of 8-oxoguanosine ( 8-oxoGsn ) were detected , respectively . RESULTS: The results of immunohistochemical observation showed that 8-oxoGsn was obviously up-regulated in the gastric cancer .The positive staining mainly accumulated in the cy-toplasm of the tumor cells .The results of mass spectrometry showed that the level of 8-oxoGsn in the gastric cancer tissues was higher than that in the para-carcinoma tissues (P<0.05).CONCLUSION:8-oxoGsn is up-regulated in gastric canc-er.RNA oxidative damage may play important roles in the occurrence of gastric cancer .

Objective To investigate the relationship between inflammation and blood coagulation function in the patients with acute exacerbation of chronic cor pulmonale (AECCP) and discuss the potential mechanism and influence on the patients. Methods The present study was based on 30 healthy controls and 141 cases of AECCP in our hospital from January 2011 to June 2014.Levels of white blood cell (WBC), neutrophil (NEUT), high-sensitivity C-reactive protein (hs-CRP, Complement 3 (C3), prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (FIB) and thrombin time (TT) in the patients were determined. Results Compared with the healthy controls, the patients had higher levels of WBC, NEUT, hs-CRP, PT, APTT, FIB, TT (all P < 0.001) and lower level of C3 (P < 0.001). Significant positive correlations were found between the levels of WBC, NEUT and FIB (r = 0.196 and r = 0.199, both P < 0.05); hs-CRP and APTT, FIB(r = 0.234, P < 0.01 and r = 0.466, P < 0.001); C3 and FIB(r = 0.466, P < 0.001), and significant negative correlations were observed between the levels of C3 and PT, APTT, TT (r=-0.258, P<0.01;r=-0.279, P < 0.01 and r = -0.168, P < 0.05, respectively). Compared with the survival patients, the cases of death had higher levels of WBC and NEUT (both P < 0.01). The area under receiver operating characteristic curve of WBC and NEUT, predicting the prognosis, was 0.666 (95% CI 0.552, 0.780; P < 0.01) and 0.695 (95% CI 0.558, 0.801; P = 0.001) respectively. Conclusions Inflammation and blood coagulation function disorder usually coexist in the patients with AECCP, and are closely associated with the severity. Levels of both WBC and NEUT can be used as prognosis predictors for the patients.

Objective To evaluate the inductive effect of interferon-γ(IFN-γ) combined with tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) on programmed necrosis of the human immortalized keratinocyte cell line HaCaT,and to explore its mechanisms.Methods In vitro cultured HaCaT cells were divided into several groups:negative control group receiving no treatment,IFN-γ group treated with 50 μg/L IFN-γ,TRAIL group treated with 4 μg/L TRAIL,and cytokine combination group treated with 50 μg/L IFN-γ and 4 μg/L TRAIL or zVAD combination group pretreated with 40 μmo/L zVAD for 1 hour followed by the treatment with 50 μg/L IFN-γand 4 μg/L TRAIL.After 48-hour treatment,the morphology of HaCaT cells were observed under a light microscope,methyl-thiazolyl-tetrazolium assay was performed to evaluate the inhibitory effect of the treatment on the proliferation of HaCaT cells,and double staining flow cytometry to detect the necrosis of HaCaT cells.Meanwhile,real-time fluorescence-based quantitative PCR (qPCR) was conducted to determine the mRNA expression of receptor interaction protein kinase 3 (RIP3) and mixed lineage kinase domain-like protein (MLKL),Western blot analysis to determine the expression of RIP1,RIP3,MLKL proteins and their phosphorylated forms (pRIP1,pRIP3,pMLKL),immunofluorescent staining to observe the distribution of pRIP3 and pMLKL in HaCaT cells,and the level of reactive oxygen species (ROS) in HaCaT cells in the above groups was detected by the fluorescence probe DCFH-DA.Statistical analysis was carried out with SPSS 22 software by using one-way analysis of variance (ANOVA) for comparing indices among different groups,and least significant difference (LSD)-t test for multiple comparisons.Results After 48-hour treatment,HaCaT cells in the cytokine combination group and zVAD combination group showed necrosis-like morphologic features.Methyl-thiazolyl-tetrazoliumassay revealed significant differences in the survival rate of HaCaT cells among the IFN-γgroup,TRAIL group,cytokine combination group,zVAD combination group and negative control group (73.16％ ± 5.71％,81.46％ ± 4.68％,72.18％ ± 2.93％,69.67％ ± 3.24％ and 100％,respectively;F =24.34,P ＜ 0.001).The necrosis rate of HaCaT cells was notably higher in the cytokine combination group and zVAD combination group (9.86％ ± 1.31％,10.33％ ± 2.16％,respectively) than in the negative control group (5.26％ ± 0.91％,t =4.61,5.07,respectively,both P ＜ 0.05).qPCR revealed that the mRNA expression of RIP3 and MLKL significantly increased in the cytokine combination group and zVAD combination group compared with the negative control group (tRIP3 =0.99,1.84,tMLKL =1.51,2.17,respectively,all P ＜ 0.05).Western blot analysis suggested that the protein expression of RIP1,RIP3,MLKL,pRIP1,pRIP3 and pMLKL significantly increased in the cytokine combination group compared with the negative control group (all P ＜ 0.05),and the zVAD combination group showed significantly decreased caspase 8 expression and increased expression of the above proteins compared with the cytokine combination group.Fluorescence microscopy showed that enhanced green dot-like or clump-like fluorescent spots (representing pRIP3) could be observed in the cytoplasm,and red fluorescent spots (representing pMLKL) could be seen on the cell membrane in the cytokine combination group.The average fluorescence intensity of ROS was significantly higher in the cytokine combination group than in the negative control group (t =702.00,P ＜ 0.05).Conclusion IFN-γcombined with TRAIL can induce the programmed necrosis of HaCaT cells with increased level of ROS.