Objective:To explore the effect of tectorigenin on myocardial fibrosis(MF) in the rats and clarify the related mechanism,and to provide reference for its clinical application. Methods:Sixty Wistar rats were randomly divided into normal control group,model group,positive drug (capropril) control group, and low,middle,high doses of tectorigenin groups(n=10).Except normal control group, the rats in other groups were used to construct MF models by subcutaneous injection of 5 mg·kg -1·d -1 isoproterenol (Iso) for 7 d.The rats in tectorigenin groups and captopril group were intragastricly administrated with different doses of tectorigenin (25,50,100 mg·kg-1·d-1)and captopril(10 mg·kg -1·d -1) from the second day after modeling for consecutive 28 d.Bl-420E+ biological function experiment system was used to detect the heart function;Heart mass index (HMI) and left ventricular mass index (LVMI) were measured after experiment.UV detection was used to measure the levels of malondialdehyde (MDA) and the activities of superoxide dismutase (SOD) in myocardial tissue.Microplate reader was used to measure the activities of lactic dehydrogenase(LDH) and creatine kinase(CK) and the levels of nitric oxide (NO)in serum.ELISA were used to detect the levels of collagen typeⅠ(ColⅠ) and collagen type Ⅲ (Col Ⅲ) in myocardium tissue of the rats.The pathological changes of myocardium tissue of the rats in various groups were observed by HE staining.Results:Compared with normal control group,the HR of rats in model groups was increased,and the left ventricular systolic pressure (LVSP) was decreased(P<0.01);the HMI and LVMI were increased(P<0.05),the levels of MDA in left ventricular myocardial tissue was increased(P<0.01),and the activity of SOD was decreased,the levels of serum ColⅠ,Col Ⅲ and the activities of LDH , CK were also increased(P<0.01);the level of NO in serum was decreased(P<0.01).Compared with model groups, the HR were decreased,LVSP were increased, and HMI and LVMI of the rats in different doses of tectorigenin groups were decreased in a dose-dependent manner;the levels of MDA were reduced;the activities of SOD were increased in myocardium tissue,and the CK activities and the ColⅠ and ColⅢ levels were decreased(P<0.05 or P<0.01);the LDH activities in middle and high doses of tectorgenin groups were decreased(P<0.01);and the levels of NO in serum in different doses of tectorigenin groups were significantly increased(P<0.05 or P<0.01) .Conclusion:Tectorigenin could inhibit the MF induced by Iso in the rats, and its mechanism may be related to antioxidation,scavenging free radical and inhibition of collagen synthesis.

Objective To test and verify the performance of analyzing IgG using nephelometry assay,and discuss reasonable model of performance verification of this system.Methods According to related documents and standards,this study verified the precision,accuracy,assay measurement range(AMR)and reference interval.Results The within-run precision in low level was 2.24%,while it was 2.73% in high level.The overall precision in low level was 2.25%,while it was 2.68%.The relative bias between the results of analyzing the calibrator with a different lot from that used for calibrating and its concen-tration printed was 5.18%.The AMR of the original dilution was 2.44~33.5 g/L.The results of reference interval verifica-tion identified with what the manufactur declares.Conclusion The major performances of analyzing IgG by this system are identifies with the manufactur declares.The reference interval offered by the manufactur is acceptable.The verification and calculation methods are simple and convenient,with strong operability.

Objective To investigate the effect of CTLA4-Ig chimera protein on mice mortality, histopathological changes, viral fiters, expression of CTLA4 protein on infiltrated T lymphocyte and the balance of Thl/Th2 in mice myocarditis caused by coxsackie virus B3 (CVB3). Methods A total of 106 four to six week-old male BALB/c mice were used in the experiments, which were divided into CTLA4-Ig group (n = 16), CVB3 group (n=40), IgG group (n =40) and normal control group(n = 10) randomly. The mice in CVB3 group, IgG group and CTLA4-Ig group were inoculated intraperitoneally with 0. 15 ml CVB3 and the mice in norreal control group with 0. 15 ml Eagle. The mice in IgG group and CTLA4-1g group were inoculated with IgG (0. I mg/kg) and CTLA4-Ig(0. 1 mg/kg) at 6 h and 72 h post inoculation(p, i. ), respectively, The surplus mice in each group were sacrificed at day 7 p.i. Light microscope was used to quantify the inflammation. The expression of CVB3 mRNA in mycardium were semi-quantified by real-time quantitative polymerase chain reaction (RQ-PCR). The expression of CTLA4 protein were analyzed by immunohistochemistry. The levels of IL-2, IL-4 and 1FN-γ in serum were measured by ELISA. Results The mice mortality, histopathological score and CVB3 mRNA in CTLA4-Ig group were lower than that in CVB3 group ( P < 0.05, P < 0.01, P < 0. 05, respectively). The expression of CTLA4 was significantly increased in CTLA4-Ig therapy group (P < 0.05 ). The serum level of IFN-γ of mice in CVB3 group were significantly higher than that in normal control group( P < 0.01 ). The serum level of IL-4 of mice in CVB3 group were much lower than that in normal control group( P < 0.01 ). The serum level of IL-2 in CVB3 group had no statistical significance with that in normal control group ( P > 0.05 ). The serum level of IFN-γ in mice of CTLA4-Ig group were much lower than that in CVB3 group ( P <0.01 ) and lgG group (P < 0. 01 ). The serum level of IL-4 of mice in CTLA4-Ig group were significantly higher than that in CVB3 group (P<0.01) and IgG group (P<0.01). The serum level of IL-2 in CTLA4-Ig group had no statistical significance with that in CVB 3 control group and lgG group ( P > 0. 0 5 ) . Conclusion CTLA4-Ig may relieve inflammation and reduce mice mortality by blocking the costimulation signals for T lymphocyte activation and reinforcing Th2 response.

Four primers against the genes encoding(cpa,cpb,etx,and iA) four major toxins(?,?,?,?) of Cl. perfringens were designed and the colony multiplex PCR of identification and genotyping of Cl. perfringens strains were developed. Cl. perfringens reference strains stored in china institute of veterinary drug control including A,B,C,D and E genotyping were genotyped using the colony muitiplex PCR assay. The expected sequences were obtained successfully by the colony multiplex PCR assay. But the sequences were not obtained from Cl. novyi,Cl. septicum and Cl. tetani. The expected sequences were obtained from Cl. perfringens individual colony diluted to 100 times with 0.85% saline solution.13 Cl. perfringens strains isolated from diferent animals were genotyped using the colony multiplex PCR assay,and the results were comparaed with the results of toxins neutranization test in mice. The two assays showed good accordance. These results showed that the development of the colony multiplex PCR is very important for early and fast identification and genotyping of Cl. perfringens in china.

ObjectiveTo evaluate the relationship of human papillomavirus(HPV) intection with expressions of cell cycle regulatory proteins cyclin D1,E and their dependent kinase inhibitor p27 in bowenoid papulosis(BP).MethodsTissue specimens were obtained from the lesions of 44 patients with BP,and circumcised foreskin tissue from 10 males served as the control.Gene chip was used to determine the genotypes of HPV,and immunohistochemistry to quantify the expressions of cyclin D1,E,p27,in these specimens.Results Of the 44 BP specimens,all were positive for HPV DNA,38(86.36％) for high risk types of HPV,and 6 for low risk types of HPV.Of the high risk HPV-positive specimens,30(68.18％) harbored HPV16,16 harbored single HPV 16,14 harbored other types of HPV besides HPV 16,8 harbored other high risk types of HPV.HPV 6 predominated in low risk HPV-positive specimens.The expression of cyclin D1 was significantly higher in patients with high-risk HPV(u =53.00,P＜ 0.05),with both high and low risk HPV(u =5.00,P＜ 0.01) and with low risk HPV (u =22.50,P＜ 0.05) than in normal human controls,and higher in patients with both high and low risk HPV (u =44.00,P＜ 0.01) and with low risk HPV (u =22.50,P＜ 0.05) than those with high risk HPV.In the case of cyclin E expression,patients with high risk HPV (u =0.00,P ＜ 0.01 ),with both high and low risk HPV (u =4.00,P ＜ 0.01 ),and with low risk HPV(u =1.50,P ＜ 0.01 ) were higher than normal human controls,and patients with high risk HPV were higher than those with low risk HPV(u =11.00,P ＜ 0.01).No significant difference was observed in the expression of p27 between patients with high and low risk types of HPV.A significant increase was observed in the expression of p27 in patients aged ＞ 50 years compared with patients aged 20-30 years(u =47.00,P＜ 0.05) and aged 31-50 years (u =55.50,P＜ 0.05),as well as in the expression of cyclin E in patients aged ＞ 50 years compared with those aged 20-30 years(u =45.50,P ＜ 0.05),and in female patients compared with male patients (u =137.50,P＜ 0.05).ConlusionThere is a significant difference in the expression of cyclin D1,E and p27 among patients with BP infected with different types of HPV.

BACKGROUND: Recent trials and clinical studies have shown that intracoronary transplantation of bone marrow-derived mesenchymal stem cells (MSCs) improves cardiac function following acute myocardial infarction (AMI). However, whether homing of MSCs into the infarcted myocardium or not is still unknown.OBJECTIVE: To study the homing of MSCs intracoronary administration in porcine myocardial infarction model using in vivo magnetic resonance imaging tracking.METHODS: Porcine MSCs were isolated and cultured by the whole bone marrow method. Following labeling by superparamagnetic iron oxide (SPIO), MSCs were treated with trypsinization to adjust the concentration at 10~(10)/L. Myocardial infarction was induced in all 10 pigs. At one week after modeling, the labeled MSCs were delivered via intracoronary infusion with standard over-the-wire (OTW) balloon angioplasty catheters. Prussian blue staining was used to evaluate labeling efficiency, and double echo steady state was used to scan four-chamber and cor biloculare at long axis view, which was considered as locating phase to obtain image of left ventricle at short axis view. RESULTS AND CONCLUSION: MSCs could be efficiently and safely labeled with SPIO. Intracoronary transplantation of MSCs is able to home the sites of myocardial injury and the border between infarcted and normal tissue. MRI can track SPIO-labeled MSCs delivered through intracoronary and were confirmed on pathology. After 5 weeks the injected labeled cells could still be detected with MRI.

Aim To observe effects of Seabuckthorn fatty acids on old rat ovarian granulosa cell apoptosis and expression of bcl-2 and fas,then discuss the adjustive effect of Seabuckthorn fatty acids on ovarian function of menopause.Methods 22 months old rats were randomly divided into 5 groups,estrogen control group,old control group,Seabuckthorn fatty acids group of high dose,middle dose and low dose.In addition,one young control group was made up of 3 months old rats.Rat ovarian granulosa cell apoptosis was detected by TUNEL.The expression of protein bcl-2 and protein fas were detected by immunohistochemistry,and bcl-2 mRNA and fas mRNA by in situ hybridization(ISH).Grey analyse was done using Q550CW image analyse system.Results Compared with the old rats control group,Seabuckthorn fatty acids of high dose,middle dose and low dose can differently restrain rat ovarian granulosa cell apoptosis,enhance the expression of protein bcl-2 and bcl-2 mRNA,weakened the expression of protein fas and fas mRNA.Conclusion Seabuckthorn fatty acids restrains ovarian granulosa cell apoptosis through adjusting the expression of bcl-2 and fas.Sequentially,it adjusts estrogen level and treats menopause syndrome.

Objective To construct and identity pSIREN-HIF-1α/shRNA expression vector in order to make foundation of gene therapy for further exploration of RNA interference to nasopharyngeal darcinoma. Methods According to HIF-1αcDNA gene sequence in the gene bank (NM_001530/NM_181054), a pair of 60 nt oligonucleotides each containing the sites of restriction endonuclease at both ends,were designed and synthesized by Reynolds design principles. Oligonucleotides were annealed and ligated with linedrized RNAi-Ready pSIREN-RetroQ-ZsGreen.Transfected into JM109, the recombinants were finally sequenced and identified by 1%agarose gel electrophoresis. Results The size of the target gene fragment amplified by PCR was 470 bp and in accordance with the expected result.pSIREN-HIF-1α was successfully constructed and identitfied by 1%agarose gel electrophoresis.Sequence analysis of inserted fragment revealed the same sequence as synthesized shRNA Oligonucleotides. Conclusion pSIREN-HIF-1α /shRNA expression vector has been successfully constructed, and can make the foundation of research using liposome packaging transfectiing nasopharyngeal darcinoma cell for the next step .

Objective To evaluate the effect of dexmedetomidine on spinal purinergic receptor 2X-4 (P2X4)/nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) pathway in a rat model of diabetic neuropathic pain (DNP).Methods Twenty-four pathogenfree adult male Sprague-Dawley rats,weighing 200-220 g,aged 8 weeks,were allocated into 3 groups (n =8 each) using a random number table:control group (C group),DNP group and DNP plus dexmedetomidine group (DNP+D group).Diabetes mellitus was induced by intraperitoneal 1％ streptozotocin 60 mg/kg and confirmed by blood glucose ≥ 16.7 mmol/L 3 days later.In group DNP+D,dexmedetomidine 50 μg/kg was intraperitoneally injected once a day for 6 consecutive weeks starting from 3 days after the model was successfully established.The mechanical paw withdrawal threshold (MWT) and sciatic nerve conduction velocity (SNCV) were measured at 2,4 and 6 weeks after injection of dexmedetomidine.The rats were sacrificed at 6 weeks after injection of dexmedetomidine,and the L4-6 segments of the spinal cord were removed for examination of the pathological changes (with a light microscope) and for determination of P2X4,NLRP3 and interleukin-lbeta (IL-1β3) expression (by Western blot).The sural nerve was obtained for examination of the ultrastructure by electron microscopy.Results Compared with group C,the MWT was significantly decreased at 2,4 and 6 weeks after injection of dexmedetomidine,the SNCV was decreased at 6 weeks after injection of dexmedetomidine,the expression of P2X4,NLRP3 and IL-1β in the spinal cord was up-regulated (P＜0.05),and the pathological changes of the spinal cord and sural nerve were marked in DNP and DNP+D groups.Compared with group DNP,the MWT was significantly increased at 2,4 and 6 weeks after injection of dexmedetomidine,the SNCV was increased at 6 weeks after injection of dexmedetomidine,the expression of P2X4,NLRP3 and IL-1β in the spinal cord was down-regulated (P＜0.05),and the pathological changes of the spinal cord and sural nerve were significantly attenuated in group DNP+D.Conclusion The mechanism by which dexmedetomidine mitigates DNP is probably related to inhibition of P2X4/NLRP3 pathway in rats.

A white-rot fungi Rigidoporus sp.W-1 which could produce laccase was isolated. The fermentation conditions in shaking flasks were investigated. The optimal carbon source was wheat bran and (NH 4) 2SO 4 was the optimal nitrogen source. The components of the medium were optimized by orthogonal experiment. When W-1 was cultured under the optimum conditions, the activity of laccase could get to 7.1U/mL in 7 days.A great amount of crude laccase could be obtained by adding fresh medium to the 7 days old mycelium.