Objective To investigate the features of immune response of human immunodeficiency virus type-1 (HIV-1) antigen specific T lymphocytes in HIV-1 monoinfected or HIV 1/hepatitis C virus (HCV) coinfected individuals.Methods Twenty-six HIV-1 monoinfected and 23 HIV-1/HCV coinfected individuals were enrolled.Immunomagnetic microbeads were used to isolate T lymphocyte subpopulation CD4+ T cells and CD8+ T cells from human peripheral blood mononuclear cells (PBMC).Frequencies of interferon-γ (IFN-γ) secreting cells of CD4+,CD8+ T lymphocytes and PBMC stimulated by a peptide pool containing 12 overlapping peptides in HIV-1 P24 from 49 patients were assessed by enzyme-linked immunospot (ELISPOT) assay.HIV-1 RNA levels of these patients were also detected by real-time fluorescence quantitative polymerase chain reaction.The data were compared by one-way ANOVA and Mann-Whitney U test,and Spearman test was used for correlation analysis.Results Frequencies of HIV-1 antigen specific CD4+ T lymphocytes [median =25 spot-forming cells (SFC)/106 cells] were significantly lower than those of CD8+T lymphocytes (median=38SFC/106 cells,F=4.592,P=0.037) and PBMC (median=53 SFC/106 cells,F=5.436,P=0.025) in HIV-1 monoinfected group.Frequencies of HIV-1 antigen specific CD4+ T lymphocytes (median=5 SFC/106 cells,Z=-2.432,P=0.015),CD8+T lymphocytes (median=5 SFC/106 cells,Z=-1.996,P=0.046) and PBMC (median=10 SFC/106 cells,Z=-2.306,P=0.021) in HIV-1/HCV coinfected group were significantly lower than those in HIV-1 monoinfected group.Conclusions In HIV-1 infection,antigen specific immune response of CD4+ T cells can be activated,but weaker than that of CD8+ T cells.Co-infection with HCV might down-regulate the responses of HIV-1 antigen specific T lymphocytes in HIV-1 infected individuals.

Objective To modify the techniques for establishment of an abdominal working heart transplantation model in rats and to sum up the key factors to success.Methods A total of 180 12-week old Brown Norway rats ( donor) and Lewis rats ( recipient) were used in this study:50 BN rats and 50 Lewis rats for pilot experiment, and 40 BN rats and 50 Lewis rats for the formal experiment.The rat model of working heart heterotopic transplantation was adopted and estab-lished by Wiedemann’ s mode.We transplanted the heart from BN rats to Lewis rats and analyzed the survival rate, causes of death and histological changes of the heart ( HE staining) in this experiment.Results After exercise and modification, the survival rate was increased to 77.5%, and the mean total duration of operation was 71 ±11 min, and the mean ische-mic time of the donor hearts was 34 ±5 min.Histological examination ( HE staining) of the cardiac allograft showed a mild inflammatory cell infiltration in the graft at 24 h after transplantation, indicating that the model was reliable.Conclusions A variety of factors may affect the final operation success rate in the establishment of this heart transplantation model.A-mong them, the major affecting factors include: healthy animals, donor heart protection, rapid and effective vascular su-ture, and postoperative animal management.

To investigate the effects of osthole on chondrocyte proliferation in vitro.

Objective To investigate the clinical characteristics,treatment effect,long-term follow up results,guanosine triphosphate (GTP) cyrclohydrolase Ⅰ (GCH Ⅰ)gene and tyrosine hydroxylase(TH) gene mutations in patients with dopa-responsive dystonia (DRD).Methods The clinical features of 3 families with 4 affected members were analyzed and all of 4 patients were screened for mutations of the GCH Ⅰ gene and TH gene with DNA sequences.Results Four patients were females,average age at onset was (15.3 ± 5.6) years (range:from 9 to 20 years).The initial symptoms were a gait disorder,stiffness or tremor of the lower limbs in all patients presented with diurnal fluctuation.As the increase of disease duration,bilateral hand tremor was found in three patients,systemic torsion was found in one patient and torticollis was found in one patient.All patients' symptoms were in complete remission after administration of low dose of levodopa.Four patients were followed up for 0.5 to 10.0 years,and all were still responsive to the levodopa treatment and effective dosage was decreased as the increase of the disease duration.No longterm side effects of levodopa had occurred after long-term treatment.One patient was found to have c.607G ＞A(p,Gly203Arg) heterogenetic mutation in GCH I gene.Molecular analysis revealed a compound heterozygous mutation in the TH gene (p.Y447Ter and p.V468M) in one patient.No point mutations in both genes were found in other patients.Conclusions DRD patients have dramatic and sustained response to levodopa and no long-term side effects of levodopa after long-term treatment.The detection of GCH Ⅰ and TH gene mutations is helpful in early diagnosis but the negative results could not exclude the diagnosis of DRD.

Objective To explore the effects of H2S on cerebral injury after cardiopulmonary resuscitation (CPR) and its mechanism.Methods Forty-five healthy Sprague-Dawley (SD) rats were randomly (random number) divided into shame-operated group ( group A,n =5 ),resuscitation group ( group B,further divided into four subgroups as per rats sacrificed 6 h,12 h,24 h,and 72 h after resuscitation,n =5),and NaHS pretreatment group ( group C,further divided into 4 subgroups as done in group B).The ratio of water content in brain tissue was calculated.The content of H2S in cerebral cortex of rats in all groups was determined by using universal microplate reader. Immunohistochemistry method was used to count the Nestin-positive cells. Results The content of H2S in hippocampus area of brain showed dramatic changes from rising up at first and then to lowering down to the minimum and finally returning to the original level in 72 h in B group.Compare to group B,brain water content was lesser ( P ＜0.05 or P ＜ 0.01 ) and the levels of Nestin in hippocampus increased in group C(P＜0.05 or P ＜0.01).The neurological deficit score (NDS) was improved (P ＜0.05 or P ＜0.01) and pathological changes in hippocampus of rat brain detected by using hemotoxylin - eosin staining were slighter in group C in comparison with group B.Conclusions Endogenous H2S may involve in the course of formation and progress of cerebral injury after CPR and small dose of NaHS (exogenous H2S) can improve NDS by decreasing cerebral edema and up-regulating Nestin level in hippocampus of brain,playing a protection role in cerebral injury after CPR.

Objective To compare the effect of glucosamine (Virtral-s) on the cartilage oligomeric matrix protein (COMP) secretion of chondrocytes and synoviocytes in vitro.Methods Chondrocytes and synoviocytes isolated from knee cartilage of osteoarthritic patients were cultured by phased enzymatic digestion.Sera containing Virtral-s of the experimental animals were obtained after orally administrated Virtral-s at the dosages that equal to human.Cells were cultured in the medium with Virtral-s containing sera.Super-natant COMP level was tested by enzyme-linked immunoabsorbent assays (ELISA).Results COMP conceu-tration of synoviocytes cultured in vitro was significantly higher than that of chondrocytes (P＜0.05).Virtral-s could significantly increase COMP secretion in cultured chondrocytes in vitro (P＜0.05),however,it had a weaker role on synoviocytes,ie,it could only mildly reduce COMP secretion of synoviocytes.Conclusion Glucosamine (Virtral-s)-containing serum can promote COMP secretion of chondrocytes in vitro,and it has no significant effect on synoviocytes in vitro.

Objective To study the expression of microRNA-301 in pancreatic carcinoma andvalidate the significance of miR-301 in invasion and metastasis of pancreatic carcinoma.Methods miR-301 expression were detected by FQ-PCR in 5 pancreatic cancer eell lines(PANC-1,PaCa-2,AsPC-1,Hs766T.BxPC-3).Further immunohistochemistry in pancreatic cancer tissue microarrays was detected miR-301 expression,which contained 60 pancreatic cancer specimens along with 10 normal adjacent tissues and 10 normal pancreas tissues.After high expression of miR-301 in pancreatic carcinoma being confirmed.the clinical significance of high expression of miR-301 in invasion and metastasis of pancreatic carcinoma were studed.Pancreatic cancer cell lines(PANC-1.PaCa-2)were transfected by 100 nmoml/L miR-301 inhibitor(anti-miR-301)or negative eontrol(Anti-miR~(TM) Negative Control#1).COX-2 and MMP-2 protein expression in pancreatic cancer cell lines were detected by WB.and cell migration assays were performed using transwell technology.Results FQ-PCR resuhs indicated that miR-301 expression was higher in pancreatic cancer cell lines than normal pancreatic cells.The relative level of miR-301 in 5 pancreatic cancer cell lines(PANC-1,PaCa-2,AsPC-1,Hs-766T,BxPC-3)and normal pancreatic cell were 33.09± 4.21,30.76±3.18,47.57±3.56,20.20 ±1.21,76.75±13.51 and 1.00±0.08 respectively.The miR-301 level in all 5 pancreatic cancer cells were significantly higher than those of normal pancreatic cell(t=8.86,9.53,6.39,6.77,11.18,P<0.01).Immunohistochemistry results also showed miR-301 expression was higher in pancreatic carcinoma tissues than those in the cancer adjacent tissues and normal pancreatic tissues.The relative levels of miR-301 in pancreatic carcinoma tissues.normal adjacent tissues and normal pancreas tissues were 0.88±0.09,0.22±0.04 and 0.14±0.05 respectively.The miR-301 levels in pancreatic carcinoma tissues were significantly higher than those of normal adjacent tissues and normal pancreatic tissues(t=15.1,10.6,P<0.01).There was no significant difference between normal adjacent tissues and normal pancreas tissues(t=1.32,P=0.22).After miR-301 inhibitor was introduced into pancreatic cancer cells PANC-1 and PaCa-2.miR-301 levels were reduced while the protein levels of COX-2 and MMP-2.which were invasion and metastasis related factors,were down-regulated.The cell migration assay indicated the numbers of PANC-1 and PaCa-2 cells,which migrated to lower chamber.were 587±27 and 363±13 respectively after miR-301 inhibitor was applied.The numbers of migrated cells were 1091 4-15.737±44 when the netative control was applied.The cell invasion ability was decreased significantly in the inhibitor group compared with the negative group(t=7.89,7.56,P<0.01).Conclusions miR-301 is highly expressed in pancreatic cancer cell lines and pancreatic cancer tissues.Inhibition of miR-301 expression can effectively supress the invasion of pancreatic cancer cells.miR-301 may serve as a new biomarker for early detection of pancreatic cancer and molecular target for early treatment of pancreatic cancer.

Molecularly imprinted polymers (MIPs) with high selectivity to tilmicosin (TIM) were prepared using tylosin(TYL) as dummy template, methacrylic acid(MAA) as monomer, ethylene glycol dimethacrylate (EGDMA) as cross-linker.The effects of 4 porogens including dimethyl formamide, methanol, acetone, and chloroform on the recognition capability of MIPs were investigated.Orthogonal test was used to optimize the preparation of MIPs, and the optimal composition was as follows; 1.0 mmol TYL, 8.0 mmol MAA, 20.0 mmol EGDMA, 6.0 mL chloroform, 20.0 mg azobisisobutyronitrile.The solid phase extraction condi tions and characteristics of MIPs as adsorptive material for the selective extraction and enrichment of TIM were also studied.The recovery of TIM was above 90% when the following procedure was applied to MIPs cartridge: conditioning with methanol and water(pH 9.0), loading with acetonitrile, cleaning with methanol and chloroform respectively, and eluting with 3 mL methanol-ammonia(95:5, V/V).The recovery of TIM on non-imprinted polymers cartridge was only 32%.

Objective To investigate the effect of different Chinese herbs on cell proliferation and cartilage oligomeric matrix protein(COMP)expression in chondrocyte culture.Methods Chondrocytes isolated from rabbit knee cartilage were cultured for 3 generations with the density of 2?10~4/cm~2 and were verified by collagenⅡimmunohistochemical staining.Rabbit sera containing herbs were obtained after animals orally ad- ministrated herbs at the dosage equivalent to human.At 5% and 10% serum density,cells were cultured in the medium that contained liver-softening herbal compound sera.Subgroups setting at 1,3 and 5 hours after herb intervention were observed.Rabbit and bovine sera were control groups.Seven days after intervention,chon- drocytes proliferation was observed using the MTT assay kit.For the study of COMP expression,chondrocytes were isolated from human knee cartilage supematant.Superuatant COMP level was tested by enzyme-linked immunoabsorbent assays(ELISA)after directly adding compound and extract from liver-softening herbs to the culture at the final concentration of 10 mg/ml for 3 days.Results Liver-softening herbal compound group had significant effect on cell proliferation compared to control,of which,3-hour subgroup was more significant than 1-and 5-hour subgroups(P

Background and purpose:It has beenreported that miR-1284 is associated with gastric cancer lymph node metastasis in the research of microRNA microarray in human gastric cancer tissues. But the specific role of miR-1284 in gastric cancer has not been reported. The aim of this study was to investigate the effect of miR-1284 over-expression on the gene expression profiling and invasion/metastasis of human gastric cancer SGC-7901 cells. Methods:Gastric cancer SGC-7901 cells of LV-miR-1284 group were transfected with lentiviral vectors of miR-1284, cells of LV-NC-GFP group were transfected with lentiviral vectors without miR-1284, and cells of control group were not transfected with lentiviral vectors. The expression of miR-1284 was detected by the real-time fluorescent quantitative PCR. Differential expression genes were detected by the microRNA chip. Target genes of miR-1284 were predicted by the bioinformatics. Invasive ability was detected by the Transwell invasion assay. Metastasis ability was detected by subcutane-ously transplanted tumor model of nude mice.Results:Compared with LV-NC-GFP and control groups, the expressions of miR-1284 and 20 genes were up-regulated, and the expression of 17 genes was down-regulated in LV-miR-1284 group. One hundred and thirty-eight target genes of miR-1284 were predicted by the bioinformatics website. Compared with invasive cell number of LV-NC-GFP group (168.67±4.55) and control group (170.33±3.08), the ability of invasion ofcells was weakened in LV-miR-1284 group (70.00±2.37). Compared with the liver metastasis rate of LV-NC-GFP group (85.71%) and control group (85.71%), the ability of metastasis of cells was weakened in LV-miR-1284 group (14.29%). Conclusion:The ability of invasion and metastasis of SGC-7901 cells is suppressed by over-expression of miR-1284. The mechanism may be related to regulating the expression ofSUMO1 andJUNgenes.