ObjectiveTo observe the effect of Batroxobin and Urokinase on brain of diabetic rats following focal cerebral ischemia and reperfusion injury. To investigate the preventive mechanism of Batroxobin following focal cerebral ischemia and reperfusion injury in diabetic rats after thrombolysis therapy.MethodsDiabetic rat was induced by administrating streptozotocin intraperitoneally. Rat model of focal cerebral ischemia and reperfusion injury induced by intraluminal filament occlusion of middle cerebral artery(MCA) that removed 2h later was used. Batroxobin and Urokinase were administrated intravenously in different groups. Infarct volume,cerebral hemorrhage and matrix metalloproteinase (MMP)-2,MMP-9 were detected at 2h,24h,48h after ischemia and reperfusion injury.ResultsThe significant decrease of infarct volume were observed in Batroxobin and Urokinase groups. There were 5 rats observed cerebral hemorrhage in Urokinase group and no cerebral hemorrhage in Batroxobin group. The number of MMP-2 and MMP-9 positive cells in Batroxobin and Batroxobin Urokinase groups decreased compared with saline and Urokinase groups. ConclusionBatroxobin can decrease the infarct volume significantly without the complication of cerebral hemorrhage after ischemia and reperfusion injury in diabetic rats, which maybe relate to down regulation of the expression of MMP-2 and MMP-9.

Objective To investigate the effect of acupuncture decompression operation on the quality of life of patients with mild to moderate essential hypertension. Methods 74 cases of mild to moderate essential hypertension were divided into experimental group and control group by random number table, 4 cases were lost. Finally, a total of 70 patients were included in the data analysis. In the control group, 35 cases were given routine drug taking and health guidance, the experimental group of 35 patients increase the treatment of acupuncture decompression operation points on this basis. The two groups of patients were measured with blood pressure before and after intervention, and World Health Organization Quality of Life (WHOQOL-BREF) was used to evaluate the quality of life, and the data were analyzed. Results The scores of physiological and psychological fields in the experimental group were (15.23 ± 2.66) points and (16.68 ± 2.23) points while the control group were (13.98 ± 2.18) points, (15.25 ± 2.52) points, the difference was statistically significant (t=2.305, 2.522, P<0.05). The experimental group of social relations, environmental score were respectively (1.37 ± 2.24) points, (14.16 ± 1.24) points, while the control group were (13.29 ± 2.30) points, (13.56 ± 1.36) points, 2 groups were not statistically significant (t=0.770, 3.063, P > 0.05). The experimental group systolic blood pressure, diastolic blood pressure were (121.43 ± 6.43) mmHg (1 mmHg=0.133 kPa), (71.74 ± 3.88) mmHg, compared with the control group were (128.34±5.72) mmHg, (76.06±4.43) mmHg. The difference was statistically significant (t=-4.753,-4.210, P<0.05). Conclusions acupuncture decompression operation can improve blood pressure in young and middle-aged patients with hypertension, improve the quality of life.

Total mRNA was extracted from lymphocytes separated from the peripheral blood of allergic patients, and then variable region of heavy chain (VH) and variable region of light chain (VL) cDNA library were constructed by RT-PCR. Human scFv templates for rabbit reticulocyte lysate ribosome display were assembled by primers and linker peptide (Gly4Ser)3. mRNA bound in antibody-ribosome-mRNA complexes was recovered using in-situ single primer RT-PCR, and three rounds of anti-IgE scFv DNA were enriched. The target DNA fragments were double enzyme digested and ligated into plasmid pET22b (+), followed by transformation in E. coli Rosseta (DE3). Positive clones were screened using clone PCR, Dot blotting and antigen ELISA. The correct lengths of VH (400 bp) and VL (710 bp) PCR products were obtained. The expected 1,000 bp ribosome display templates were also observed in agarose gel electrophoresis. After three rounds of ribosome display target sequences were effectively enriched, leading to a library of 10(13) members. Antibodies with the highest ELISA value for IgE were generated in the strain pET-IgE-6. A human anti-IgE scFv library was successfully constructed as described herein. Ribosome display using single primer in-situ RT-PCR as the recovery procedure effectively enriched target sequences. Anti-IgE scFv with high affinity and specificity were identified. The prepared human anti-IgE scFv fragment might be self-developed to a lead drug for treating asthma. Our study provides an alternative method for rapid discovery of human antibodies of therapeutic importance.

SUMMARY Our work focused on the studies on the expression and function of transient receptor poten -tial vanilloid subtype 1 ( TRPV1) in the submandibular gland .By using reverse transcription-polymerase chain reaction ( RT-PCR ) , Western blotting , and immunofluorescence , our data demonstrated the expression and distribution characteristics of TRPV 1 in rabbit and human submandibular glands , as well as rat submandibular gland cell line SMG-C6.Furthermore, the possible intracellular signal molecules involved in the TRPV1-modulated saliva secretion were explored .Activation of TRPV1 increased the intracellular Ca2+concentration, upregulated the expression of aquaporin 5 (AQP5), the main transpor-ter that mediate water secretion through transcellular pathway , and led to AQP5 redistribution .Extracel-lular signal-regulated kinase 1/2 ( ERK1/2 ) was involved in the TRPV1-regulated AQP5 content. Besides, TRPV1 activation also modulated the expression , distribution, and function of tight junction protein, and increased paracellular permeability .ERK1/2 and myosin light chain 2 ( MLC2 ) were responsible for the regulation of TRPV1on tight junction properties.Taken together, our work suggested that TRPV1 was a potential target to promote saliva secretion , and activation of TRPV1 might provide a new and safe therapeutic strategy to ameliorate submandibular gland hypofunction .

To observe different expression patterns of β-catenin and its clinical significance in colorectal cancer (CRC).Methods A total of 181 cases of CRC tissues and 30 cases of normal colorectal tissue were investigated by immunohistochemistry for the expression of β-catenin.Results The expression rate of β-catenin was 56.9％ (103/181) in CRC,and higher than that in normal colorectal tissue (P ＜ 0.05).The overexpression of nuclear β-catenin was significantly correlated with histological differentiation,lymph node metastasis and Dukes' stage in CRC (P ＜ 0.05),and no relationship with other pathological parameters,such as age,gender and the depth of infiltration.The incomplete membranous expression of β-catenin was significantly correlated with histological differentiation,the depth of infiltration,lymph node metastasis and Dukes' stage in CRC (P ＜ 0.05).The high expression of nuclear β-catenin related to histological differentiation and Dukes' stage in CRC (P ＜ 0.05).In the follow-up data of 82 cases of CRC,the expression of nuclear β-catenin was associated with poor prognosis,and the 5-year survival rate was significantly lower than that of self-control groups (P ＜ 0.05).Conclusion β-catenin plays important roles in colorectal carcinogenesis.Abnormal expression of β-catenin was related to the aggressive progression of CRC and may be helpful for evaluating the prognosis of patients with CRC.β-catenin is expected to become a new target for diagnosis and treatment of CRC in future.

Purpose To investigate the expression and clinical significance of microRNA-21 (miR-21) in patients with prostate cancer (PCa).Methods The human tissue specimens were paraffin-embedded blocks which were collected from the Department of Pathology.miR-21 was detected by in situ hybridization (ISH) in 90 cases of PCa and 69 cases of benign prostatic hyperplasia (BPH).Results The ISH data showed that the positive rate of miR-21 in PCa (71.1％) was significantly higher than that in BPH (24.6％) (P ＜ 0.05),the expression of miR-21 correlated with the Gleason grade (P ＜ 0.05),and no relationship was observed with other clinicpathological features (P ＞ 0.05).Conclusion The miR-21 plays an important role in the pathogenesis of PCa,which can be a new diagnostic target for PCa.

AIM: To observe the effects of puerarin on blood pressure(BP),serum lipid in a rat model of insulin resistance and the mechanism.METHODS: Adult SD rats were maintained on high-fat-sugar-salt diet for 12 weeks.Puerarin was administered to the rats from 9th week for 4 weeks by intramuscular injection.BP was measured at the end of 0,8th,12th week.The levels of serum glucose,serum lipid,fasting serum insulin and the levels of MDA,TNF-?,plasma rennin,angiotensinⅡ(AngⅡ) and the activity of SOD were measured and the insulin-sensitivity index was also calculated at the end of the experiment.RESULTS: High and middle dosage of puerarin significantly decreased blood pressure,reduced the levels of serum lipid and AngⅡ,and also increased insulin-sensitivity index.The levels of MDA and TNF-? were significantly decreased by high dosage of puerarin.The activity of SOD was increased significantly.CONCLUSIONS: Puerarin possesses the effects of decreasing the blood pressure and serum lipid in a rat model of insulin resistance,which may be concerned with the changes of rennin-angiotensin system,the levels of oxygen-derived free radicals and TNF-?.

Objective:To study the pathological mechanism of the inducible co-stimulator molecular and ligand ( ICOS/ICOSL) in Graves disease animal.Methods:45 out-bred BALB/c mice were randomly divided into three groups with 15 rats in each group;using gene gun to deliver different plasmid injection.Group A was delivered with pCDNA3.0-mICOSL and pCDNA3.0-hTSHR, Group B with pCDNA3.0-hTSHR and null pCDNA3.0 with Group C for immunization as the control group.The concentration of serum free thyroxine immunization was deter mined with immunoassay and serum thyrotropin receptor antibody ( TRAb ) with ELISA, supernatant of IFN-γconcentration in mouse spleen cells was measured with radioimmunoassay,and hTSHR transected CHO cells were incubated to detect the concentration of cAMP to deter mine autoantibody TRAb activity.Results: After plasmid injection serum FT4 level in Group A (0.49±0.25) pg/ml ( q=6.571,P=0.023) was higher than that in Group C,the standard rate was higher than Group B and C (χ2=14.47,P=0.005).IFN-γconcentration of mice spleen cultured supernatant in Group A (1.88±0.41) pmol/L was significantly higher than the other two groups.The activity of autoantibody TRAb in Group A 188.3 (179.7-260.2) %was higher than that in the other two groups ( P=0.027 ) .Conclusion: Exogenous delivery of pCDNA3.0-mICOSL plasmid in GD mice could stimulate the spleen lymphocytes to secrete more IFN-γ,increase the activity of TRAb autoantibodies and might lead to upregulation of immune response in Graves animal model in vivo.

Objective To study the effects of prokaryotic ubiquitin-like protein ( Pup)-proteasome system on the growth of Mycobacterium strains.Methods The genes encoding Pup ( pup gene) and protea-someβsubunit ( prcB gene) were respectively knocked out from Mycobacterium smegmatis ( M.sm) strains by homologous recombination.The growth and viability of the wild-type and mutant strains of M.sm were an-alyzed under normal culture condition and under hypoxia as well as anaerobic conditions.Results The pup and prcB genes were completely and precisely knocked out from M.sm strains and the mutant strains were named △SM-Pup and△SM-prcB, respectively.The△SM-Pup strains grew faster than the wild type ( WT) and△SM-prcB strains.No significantly differences in the growth of M.sm were found between the WT and△SM-prcB strains.Conclusion The Pup-proteasome system was involved in the growth of M.sm, espe-cially the pup gene.There was difference between pup and prcB genes in regulating the growth of M.sm.The functions and influences of Pup-proteasome system still need further investigation.

Objective To explore the role and correlation of peroxisome proliferator-activated receptor γ (PPAR-γ) and Toll-like receptor 4 (TLR4) on pulmonary vascular remodeling of chronic obstructive pulmonary disease(COPD).Methods Ninety male patients who underwent surgery for squamous cell carcinoma were enrolled as our subjects.All patients were divided into COPD group and uon-COPD group based on lung function,and 45 cases in each group.Peripheral lung tissues without tumor infiltrated after lobectomy were taken to assess the degree of arterial inflammation,percentage of wall thickness to vessel diamater (WT％) and percentage of wall area to total vascular area(WA％) were measured through Hematoxylin-Eosin(HE) staining under light microscope.The expression of PPARγand TLR4 were determined by immunohistochemistry.Results The distribution of WA％ and WT％ in COPD group were (43.98 ±6.43)％ and (27.37 ±3.34)％,higher than that of non-COPD group ((26.09 ± 2.82) ％,t =-13.949,P =0.000 ; (15.57 ± 1.75) ％,t =-7.140,P =0.000).The expression of PPAR-γand TLR4 in COPD group were (10.74 ± 8.81) ％,(3 1.41 ± 14.67) ％ respectively,and (28.22 ± 15.08)％,(4.67 ± 4.47)％ in non-COPD group.The differences were statistic significantly(t =5.483,P =0.000; t =-9.555,P =0.000).And there was negative correlation between the expression of PPARγand TLR4 (r =-0.404,P ＜ 0.01).Conclusion The pulmonary vascular of COPD patients showed the obviously inflammatory cell infiltration,fibrosis and proliferation,and PPAR-γ and TLR4 participate in the regulation of pulmonary vascular remodeling.