Objective To investigate mechanism of cyclooxygenase-2 ( COX-2) in bone loss in a postmenopausal osteoporosis (PMOP) rat mode with ovarietomy (OVX).Methods Forty female Sprague Dawley adult rats at age of 3 months were randomly divided into 4 groups,10 in each group,including shamoperated (sham) group,OVX group,OVX treated with nilesteriol (OVX + E) group and OVX treated with aspirin ( OVX + P) group.All rats in OVX,OVX + E and OVX + P groups underwent ovarietomy under abdominal anesthesia with 10％ chloral hydrate.Rats in sham group were only taken with fat tissue with same weight under bilateral ovary.After surgery,penicillin was administered to prevent infection.At day 7 after surgery,agents were given by intragastric administration for 12 weeks.Nilestriol at 1.0 mg/kg was used in OVX + E group once a week,aspirin at 45 mg · kg - 1 · d- 1 was used in OVX + P group once a day.Saline with same volume was used in rats in sham and OVX groups.All agents were administered one time per day.Dose of agents were adjusted by weight per week.At end of study,bone mineral density (BMD) of right femurs and lumbar vertebrae 3 -5 (L3-5) were measured.Morphology of bone was detected by hematoxylineosin,and expression of COX-2 was determined by immunohistochemistry staining.Results ( 1 ) BMD:BMD of right femur and L3-5 was (0.209 ±0.010) g/cm2 and (0.230 ±0.012) g/cm2 in sham group and (0.181 ±0.008) g/cm2 and (0.201 ± 0.016) g/cm2 in OVX group,which reached statistical difference (P＜0.01).BMD of right femur and L3-5 was (0.203 ±0.009) g/cm2 and (0.224 ±0.028) g/cm2 in OVX + E group and (0.200 ± 0.011 ) g/cm2 and (0.204 ± 0.003 ) g/cm2 in OVX + P group,which were all higher than those in OVX group (P ＜0.01,P ＜0.05).However,there was no statistical difference in BMD between OVX + E and OVX + P group ( P ＞ 0.05).(2) Morphology of bone:bone trabeculae became fewer and degenerated in OVX group.However,bone trabeculae were regular and dense in OVX + P group and OVX + E group,which were similar to those in sham group.(3) Expression of COX-2:cells with COX-2 positive and expression of COX-2 around bone trabeculae in OVX group were more than those in sham,OVX + E and OVX + P group.Conclusion COX-2 plays an important role in PMOP.Aspirin could prevent bone loss by decreasing COX-2 expression in OVX rats.

Objective To investigate the efficacy and safety of Bevacizumab (Bev) combined with chemotherapy protocols in the patients with metastatic colorectal cancer (mCRC).Methods 43 patients with mCRC were treated with Bev combined with various of chemotherapy protocols.The efficacy was assessed based on Response Evaluation Criterion for Solid Tumors (RECIST),and the adverse events were evaluated according to National Cancer Institute-Common Toxicity Criterion for Adverse Events (NCI-CTCAE) 3.0.Results The data of 43 patients (16 males and 27 females) were analyzed.17 patients achieved partial remission (PR),19 patients stable disease (SD) and 7 patients progressive disease (PD).The median progression-free survival (PFS) time was 10.3 months.The major of 3-4 adverse events included leucocytopenia,neutropenic febrile,and nausea/vomiting.Bey-associated adverse events were proteinuria,hypertension,rhinorrhagia,hemorrhagic hemorrhoid,menstrual blood increased,gastrointestinal perforation and venous thrombosis.Conclusions Bey combined with various chemotherapy protocols is more effective for patients with mCRC.Most patients can tolerate the side effects of the combined treatment.The long-term effects of the combined treatment need to be followed up.

Objective To study the differentially expressed microRNA (miRNA) between pancreatic ductal adenocarcinoma (PDAC) and para-cancerous tissues,and determine related target genes.Methods Nine fresh PDAC tumor tissues and 3 adjacent normal pancreatic tissues were collected,then Agilent miRNAmicroarray with 713 miRNA loci was used to identify the differentially expressed miRNA.Real-time PCR method was applied to verify the up-regulated miRNA.TargetScan 5.1 and miRandaV5 software were used to analyze the related target genes.Results miRNA microarray identified 11 PDAC related miRNAs,among them,the expressions of miR-194*,miR-192*,miR-602,miR-194 were up-regulated,while the expressions of miR-139-3p,miR-513a-5p,miR-630,miR-30c-1 *,miR-887,miR-508-5p,miR-516a-5p were down-regulated.The expressions of miR-192,miR-194 and their homolog were verified in 31 PDAC tumor tissues.After software analysis,it was found that target genes of miR-192 were ZEB2,CXCL-2,EEF1A1,ERCC3,and target genes of miR-192 * were DCC,SMAD4,FAS,and target genes of miR-194 included DACHI,IGSF11,PTPN2,RBBP4,while target genes of miR-194 * included CD40LG,CIDEB,FHL1.Conclusions Eleven differentially expressed miRNAs are present in PDC,and they may be involved in the occurrence and development of PDC.

Objective To study the aberrant DNA methylation of H19 and MEST in infertile male sperm.Methods Routine semen analysis and morphological analysis were performed on 15 control individuals and 33 oligoastheno-teratozoospermias.Sperms were purified by density gradient centrifugation and DNA was extracted and treated with bisulfite.After PCR,the purified PCR products were cloned into pMD (R)18-T vector and proceed to chemical transformation,restriction enzyme reaction.For each sample,positive clones were selected for sequencing analysis and DNA methylation analysis.Results Five control individuals had high degree of H19 and methylation rate was 100.0％,while the methylation rate of 93.0％ in 10 oligoastheno-teratozoospermias was significantly lower.Significant difference was found in methylation rate between the control individuals and oligoastheno-teratozoospermias (P ＜ 0.01).As to MEST,10 control individuals had a low degree of MEST and the methylation rate was 1.6％,while the methylation rate in 23 oligoastheno-teratozoospermias was 4.8％.No significant difference was found in the methylation rate between control individuals and oligoastheno-teratozoospermias (P ＞ 0.05).Conclusion Male infertility is associated with aberrant methylation of H19 differentially methylated region,and it may be a biomarker of human spermatogenesis defect.

AIM: To study the chemical constituents from the leaves of Phyllanthus emblica L. METHODS: The constituents were extracted by percolation with 95% ethanol.Then the extract was separated by systemic solvent separation methods.The ethyl acetate portion from the leaves were separated and purified by silica gel column chromatography and polyamide column chromatography with gradient elution,liquid preparation and recrystal methods.The structures of crystals were identified by physiochemical properties,spectrum analysis and literatures ontrast. RESULTS: Five compounds were isolated and identified.They were ?-sitosterol(Ⅰ);?-carotene(Ⅱ);kaemferol(Ⅲ);quercetin(Ⅳ);avicularin(Ⅴ). CONCLUSION: Chemical compound Ⅴ is isolated from this plant for the first time.

Objective To study the influence of site-directed deglycosylation of the HIV-1 envelope (Env) on its immunogenicity and assembly of functional pseudovirus. Methods Site-directed deglycosylation were performed using cycling mutagenesis and selection of mutants with DpnⅠ. Single-cycle infection assay was employed to analyze the effect of the mutations on the ability of functional pseudovirus assembly. The influence of deglycosylations on the immunodeficiency of Env was evaluated using pseudovirusbased neutralization assay and ELISPOT assay. Results Mutant N197Q induced higher neutralization activities for both pseudoviruses, but lower Env-specific T-cell response. And N197Q rendered the Env to lose the ability of functional pseudovirus assembly. Mutant G2 induced higher neutralization activities for pseudovirus 74-2 but lower for pseudovirus Wt, and had almost no influence on Env-specific T-cell response and functional pseudovirus forming. Conclusion The site-directed deglycosylation of the HIV-1 Env affects the pseudovirus forming and its immunogenicity.

Objective To compare cellular immune responses in mice elicited by Chinese different AIDS candidate vaccines.Methods According to their different immunization procedures,BALB/c mice were immunized with 6 AIDS candidate vaccines,separately.Spleen cells were isolated for the detection of cellular immune response to HIV-specific peptides using enzyme-linked immunosorbent spot(ELISPOT)assay and intracellular cytokine staining(ICS)method.Results AIDS vaccines were evaluated by using potential T-cell epitopes(PTE)Gag,Env and Pol peptides pool and ELISPOT.The positive conversion rates for cellular immune response of 1#-6# vaccines fluctuated from 70% to 100%.The vaccine-induced cellular immune responses to specific peptides pool are different not only in magnitude but also in breadth.The Th1type cytokines,IFN-γand IL-2,were detected with ELISPOT in 1# and 2# vaccines.The productions of IFN-γand IL-2 induced by both of the two vaccines showed a moderate correlation(r1 =0.62,P1 ＜0.01 ;r2=0.79,P2 ＜ 0.01).The positive conversion rate of IFN-γ secreting cells of 1 # vaccine was 66.7%(10/15)mice detected with both ELISPOT and ICS.And the results tested by ELISPOT and ICS showed moderate correlation(r = 0.55,P ＜ 0.05).Conclusion The magnitude and breadth of cellular immune responses induced by different AIDS candidate vaccines are different.Being induced by different AIDS candidate vaccines,the IFN-γand other Th1 type cytokines detected by ELISPOT or ICS could be used to evaluate the cellular immune responses in mice.

Objective To analyze the practicability of using ELISA kit for the detection of hepati-tis E virus antigen ( HEV-Ag) in plasma donations and Biomex HEV seroconversion panels. Methods The HEV-Ag positive samples were screened out from 36 340 donated blood plasma samples. Real-time fluores-cent PCR was performed for the detection of HEV RNA in HEV-Ag positive samples. The open reading frame 2 (ORF2) in HEV RNA was amplified by nested RT-PCR and the amplified products were confirmed by sequencing analysis. Phylogenetic tree was constructed for HEV genotyping. Five Biomex HEV serocon-version panels were used in this study for the detection of HEV-Ag, anti-HEV antibody and HEV RNA as well as the correlation analysis between HEV-Ag and HEV RNA. Results Twenty-six out of 36 340 plasma samples (0. 07%) were positive for HEV-Ag. Of the 26 samples, 25 samples were positive for HEV RNA as indicated by the results of nested RT-PCR and 23 positive samples were confirmed by sequencing analysis. The positive rate of HEV RNA in blood plasma donators was 1 ∶ 1 580 (0. 06%). There were 17 samples of genotype 1 (74%) and 6 samples of genotype 4 (26%) according to the phylogenetic tree analysis. All of the HEV-Ag positive samples were also positive for HEV RNA as indicated by the analysis of Biomex sero-conversion panels. HEV-Ag was consistent with the peak of the HEV RNA concentration. Conclusion A close relationship between HEV-Ag and HEV RNA was observed. HEV-Ag screening could be used as a measure to reduce the risk of HEV transmission by blood transfusion.

Objective To investigate the expression of HSPB1 in pancreatic cancer and its relationship with clinicopathological characterization.Methods Pathological specimens from 47 cases of pancreatic cancers,13 cases of para-cancerous normal pancreatic tissues and 3 cases of chronic pancreatitis tissues were selected,and tissue microarray construction instrument was used to prepare tissue microarray,then the expression of HSPB1 was determined by immunohistochemistry SP method.The scores of the proportion of positive cells and staining intensity were multiplied to make the judgment.Results The expression of HSPB1 in malignant,para-cancerous and chronic pancreatitis tissues was 80.9％ (38/47),12.5％ (2/16),respectively; and the difference between the two groups was statistically significant (X2 =24.058,P =0.000).The expression of HSPB1 in pancreatic cancer tissue was not significantly related to sex,age,location,differentiation degree and nerve infiltration of tumor ( P ＞ 0.05 ).Conclusions The expression of HSPB1 is related to development and progression of pancreatic cancer,and may be an early molecular event.

Objective To evaluate the trimester specific reference ranges and dynamic changes of thyroid functional indicators during gestation of healthy pregnant women.Methods According to the selection criteria,139 cases were selected as the normal pregnant population to establish a self-sequential longitudinal reference range.We collected samples eight times in every case throughout the gestation(including the 8th,12th,16th,20th,28th,36th week of prenatal,the 3rd and the 6th month postpartum) and detected the indicators of thyroid funciton.Meanwhile,301 healthy non-pregnant women of childbearing age were selected as the control group.Results The median of TSH was lowest in the first trimester,and then was increased in the second and third trimesters.The reference ranges of TSH were 0.20-4.01 mIU/L,0.43-3.97 mIU/L,and 0.57-3.96 mIU/L in the above trimesters.The free thyroxine (FT4) level from the 12th week of gestation to the 6th month of postpartum was gradually decreased and was lower than the non-pregnant level.The total thyroxine (TT4) was significantly increased at the 12th week of gestation and thereafter it was stable and returned to normal after childbirth.The free triiodothyronine(FT3) was gradually declined from the 12th week to the 28th week and returned to normal postpartum.Total triiodothyronine(TT3) was significantly increased at the 12th week and thereafter was stable,and returned to normal after childbirth.There was a significant negative correlation between TSH and FT4 in the first trimester(P＜0.01).There was no correlation between TSH and TT4 during the whole gestation.There were positive correlations between FT4 and TT4,FT3 and TT3 since the 12th week of gestation (P ＜ 0.01).Conclusions The gestational specific reference range of TSH based on the selfsequential longitudinal study was narrow,however,the upper limit was still higher than that recommended in foreign guidelines.