Objective To explore the relationship between methylene tetrahydrofolate reductase (MTHFR) gene polymorphism and homocysteine (Hcy),and further to investigate their correlation to coronary heart disease (CHD) in females.Methods This study involved 29 premenopausal women with CHD,57 postmenopausal women with CHD,50 men with CHD and 46 postmenopausal women with non-CHD.All patients underwent coronary angiography.The patients' gender,age,history of hypertension,diabetes and smoking were recorded.The plasma Hcy level,blood lipids and MTHFR gene polymorphism were examined.Results The level of Hcy in postmenopausal women with CHD was (18.11±7.21)μmol/L,which was significantly higher than (15.01±6.53)μmol/L in premenopausal women with CHD,(12.98±5.46)μmol/L in postmenopausal women with non-CHD group and (14.32±6.01)μmol/L in men with CHD group (t=5.23,3.65,5.08,all P<0.05).The frequency of the MTHFR TT genotype in postmenopausal women with CHD was 28.1% which was significantly higher than postmenopausal women with non-CHD(17.4%)(χ2=5.13,P<0.05),but there was no statistically significant difference with the other two groups.The plasma Hcy level in patients with TT genotype was significantly higher than that with CT or CC genotype in all groups.Multivariate logistic regression analysis showed a significant positive correlation in Hcy and TT genotype.Multivariate logistic regression analysis showed that smoking,Hcy,hypertension and diabetes were independent risk factors for CHD.The odds ratios(OR) of Hcy for CHD was 2.601 (95% confidence interval,1.326 to 5.745).Conclusion Hcy may be an independent risk factor for CHD in females.MTHFR C677T genotype is significantly associated with hyperhomocysteinemia,and its mutation frequency increases significantly in patients with CHD,but MTHFR mutation is not related with the incidence of CHD.

Objective To evaluate the effect of antler polypeptides from Cervus elaphus yarkandensis on MC3T3-E1 cells. Methods The concentration of antler polypeptides of Cervus elaphus yarkandensis was measured by BCA protein assay kit. MC3T3-E1 cells were cultured and analyzed by BCIP/NBT chromogenic alkaline phosphatase (ALP) staining kit. After being induced to form mineralized knot, the cells were stained by using Alizarin red staining. Three different concentrations (10, 1.0, 0.1 μg/mL) of antler polypeptides were analyzed by MTT method and micronutrients enzymes standard method to determine the effect of cell proliferation and ALP synthesis. Results The concentration of antler polypeptides was 0.07 mg/mL. The results of in vitro cell activity analysis showed that the positive rate of ALP was 90%and the mineralization knot was stained red. Compared with the control group, the different concentrations of antler polypeptides all showed the function of cell proliferation and the effect was dose-dependent after 3 d and 7 d. Compared with the control group, at the 3 d, three groups of antler polypeptides promoted synthesis and secretion of ALP (P<0.01) and the results showed a dose-dependent effect. Conclusion Antler polypeptides could obviously promote the proliferation of MC3T3-E1 cells and secretion of ALP, which indicated that antler polypeptides have certain effect on osteoporosis.

Objective To conduct extensive quality control tests on Madin-Darby Canine Kidney ( MDCK) cells used for the production of influenza vaccine .Methods Tests for characteristics , extraneous agents, endogenous agents and tumorigenicity were performed on MDCK cells according to Chinese Pharma -copeia Book III .Cell lysate and DNA of MDCK cells were tested for oncogenicity in the light of new interna -tional requirements .Results The MDCK cells extracted from canis were adherent cells with an epithelial morphology, whose average number of chromosome was 80±1.No bacteria, fungi and mycoplasma contami-nation were detected . The detection for extraneous and endogenous virus showed that there was no nonspecific virus causing cytopathic effect , hemadsorption , hemagglutination or animal death .Tests for re-verse transcriptase , bovine viruses and canine viruses were all negative .Each nude mouse was injected with 107 viable cells to observe their tumorigenicity .Twelve weeks after cell injection , no node was found at the injection site and in large organs by gross anatomy .There was no significant difference between test group and negative control group .The test for tumorigenicity of viable cells was negative .Cell lysate and cellular DNA collected from equivalent amount of cells were respectively injected into nude mice , and no node forma-tion was found.There was no significant difference between the test cells and negative controls in pathology indicating that the tested MDCK cells were non-oncogenic .Conclusion It showed the possibility of using MDCK cells for the production of influenza vaccine .

Objective To investigate the production and mechanism of chemokine (C-C motif) ligand 5 (CCL5) by macrophages in U14 cervical cancer-bearing mice during infection. Methods The U14 cervical cancer cells were injected in C57BL/6 mice to induce tumor-bearing condition. Lipopolysaccharide (LPS) was injected into C57BL/6 mice to induce infection. The protein expression of CCL5 in the serum and the CCL5 mRNA expression in inflammatory cells were measured by ELISA and fluorescence quantitative-PCR in four groups. Macrophages were induced in the tumor conditioned medium (TCM) which extracted from mice serum. The protein expression levels of CCL5, prostaglandin E2 (PGE2) and cyclic adenosine monophosphate (cAMP) in the medium and CCL5, PGE2 and cAMP mRNA expression in the macrophages were detected in different groups. In order to determine whether the inhibition was related to PGE2, selective cyclooxygenase 2(COX-2) inhibitor NS398 was used to reverse this phenomenon and protein kinase A (PKA) inhibitor H89 demonstrated the mechanism through blocking cAMP/PKA signaling pathway. Results (1) The protein and mRNA level of CCL5 in tumor-bearing mice were respectively (151±35) pg/ml and 1.0, which were lower than those in the tumor-free mice (691 ± 85) pg/ml and 4.5 ± 0.8, there were significant difference between them (all P<0.05). The protein and mRNA level of PGE2 in tumor-bearing mice were (1 198±83) pg/ml and 5.8±0.8, which were higher than those in the tumor-free mice (187±25) pg/ml and 1.0, the difference were significant (all P<0.05). The protein and mRNA level of CCL5 in tumor-free+LPS mice were (4 049±141) pg/ml and 31.5±2.0, which were higher than those in the tumor-bearing+LPS mice (1 951±71) pg/ml and 12.1±2.8, the difference were also significant (P<0.05). The protein and mRNA level of PGE2 in tumor-free+LPS mice were (676±70) pg/ml and 3.4±0.4, which were lower than those in tumor-bearing+LPS mice (2 550±382) pg/ml and 11.6±0.9, the difference were also significant (all P<0.05). (2) Macrophages were cultured in vitro using TCM derived from mice. The protein and mRNA level of CCL5 in tumor-bearing mice TCM were respectively (1 626 ± 177) pg/ml and 28.6 ± 1.2, which were higher than those in the tumor-free mice TCM [(27 ± 3) pg/ml and 1.0], there were significant difference (P<0.05). The protein and mRNA level of PGE2 in tumor-bearing mice TCM were (790 ± 156) pg/ml and 1.7 ± 0.3, which were higher than those in the tumor-free mice TCM [(448 ± 115) pg/ml, 1.0], the difference were significant (all P<0.05). The protein and mRNA level of cAMP in tumor-bearing mice TCM were (164 ± 30) pg/ml and 1.6 ± 0.3, which weres higher than those in the tumor-free mice TCM [(118 ± 25) pg/ml,1.0], the difference were significant (all P<0.05). The protein and mRNA level of CCL5 in tumor-free + LPS mice TCM were (10 475 ± 742) pg/ml and 212.0 ± 5.7, which were higher than those in the tumor-bearing+LPS mice TCM [(6 375±530) pg/ml, 142.3±2.5], the difference were significant (all P<0.05). The protein and mRNA level of PGE2 in tumor-free+LPS mice TCM were (2 438±95) pg/ml and 4.3±0.7, which weres lower than those in the tumor-bearing + LPS mice TCM [(3 441 ± 163) pg/ml, 5.9 ± 0.3], the difference were significant (all P<0.05). The protein and mRNA level of cAMP in tumor-free+LPS mice TCM were (340 ± 13) pg/ml and 4.1 ± 0.4, which were lower than those in the tumor-bearing + LPS mice TCM [(542 ± 42) pg/ml, 5.4 ± 0.5], the difference were significant (all P<0.05). (3) Using COX-2 inhibitor NS398 in the tumor-bearing+LPS mice, the protein and mRNA level of CCL5, PGE2 and cAMP were (7 691±269) pg/ml and 159.0±8.9, (2 820±152) pg/ml and 4.9 ± 0.3, (465 ± 8) pg/ml and 4.3 ± 0.4, respectively, and there were significant difference (all P<0.05), compared to before treatment. Using PKA inhibitor H89 in the tumor-bearing+LPS mice, the protein and mRNA level of CCL5, PGE2 and cAMP were (8 375±520) pg/ml and 177.0±8.8, (2 650±35) pg/ml and 4.7 ± 0.4, (368 ± 13) pg/ml and 3.1 ± 0.7, respectively, and there were significant difference (all P<0.05), compared to before treatment. Conclusion TCM of U14 cells activated macrophages to release PGE2 could inhibit the expression of CCL5 levels by cAMP/PKA signaling pathway.

Objective To study the safety and feasibility of endobiliary intraductal radiofrequency ablation (RFA) to reopen occluded self-expandable metal stents in patients with malignant biliary obstruction.Methods 11 patients with malignant biliary obstruction and blocked metal stents were prospectively studied.During ERCP, after biliary cannulation, the blocked metal stents underwent RFA using a bipolar radiofrequency probe which was introduced into the stenotic bile duct via a guide wire.This was followed by a balloon to repeatedly remove debris and then endoscopic nasobiliary drainage.The patients were closely observed and followed up.Results RFA was successfully carried out in all the patients and patencies were achieved when compared with pre-RFA.The median post-RFA luminal diameter of the strictures showed significant improvement: 6 (4 ～ 10) mm versus 2 (0 ～ 5) mm, and the mean post-RFA total bilirubin level decreased sharply : (39.4 ± 8.7) μ mol/L versus (130.1 ± 38.2) μmol/L.Following this intervention, 3 patients developed fever, which were controlled with conservative therapy.There was no mortality, haemorrhage, bile duct perforation or bile leak.Of the 11 patients, 3 were dead and 6 were alive at a median follow-up of 187 (75 ～ 304) days.The median stent patency was 135 (75 ～ 203) days and the median survival was 278 (75 ～ 304) days.Four patients had their stents patent at the time of the last follow-up or death.Seven patients had their stents blocked on 113, 124, 154, 203, 96, 135 and 112 days post-procedure.Condusions Endobiliary intraductal RFA is technically feasible and safe to reopen occluded metal stents in malignant biliary obstruction.This efficacy needs to be confirmed by future randomized studies.

Objective To investigate the chemokine 12 (CXCL12) and chemokine receptor 4 (CXCR4) expressions in hypopharyngeal carcinoma and its place in the disease development,invasion and metastasis of significance.Methods Immunohistochemistry was used to detect the expressions of CXCL12 and CXCR4 in 35 cases of hypopharyngeal cancer tissues and in 28 cases of tumor-adjacent non-tumor tissues.Results The expressions of CXCL12 and CXCR4 in the hypopharynx carcinomas were significantly higher (P ＜ 0.05).Both expressed in hypopharyngeal carcinomas was significantly positively correlated (P ＜ 0.01).Both hypopharynx cancer in lymph node metastasis group were significantly higher than the expression of cervical lymph node metastasis group,the difference was significant (P ＜ 0.05).Conclusions CXCL12 and CXCR4 are involved in hypopharynx cancer development,invasion and metastasis,and there is a positive feedback regulation mechanism between two factors.Moreover,CXCL12 and CXCR4 have synergistic effect in development,invasion and metastasis of hypopharynx cancers.

Objective To evaluate the diagnostic significance of blood cells and electrolyte in preoperative diagnosis and surgical treatment of Cushing's disease (CD).Methods 116 csses of CD and 21 cases of adrenocortical adenoma (ACA) patients pathologically proven adrenocorticotropic hormone (ACTH) adenoma from Jan.2003 to Dec.2010 were enrolled into the study.They were given transsphenoidal surgery (TSS) and laparoscopic resection of adrenal adenoma (LRAA).Patients were divided into remission group ( group A),non-remission group (group B) and ACA group (group C) according to the remission criteria of CD.The preoperative and postoperative level of blood cells and electrolyte were determined.Results TSS treatment improved the abnormal level of blood cells and electrolyte in patients with CD.Group A had significant change in preoperative and postoperative level of blood cells and electrolyte,while group B and group C didn't.Conclusions Compared with the endocrine examination items which need huge expense and harsh conditions,the change of blood cells and electrolyte level is more superior in assessing diagnostic and cruative effects on CD patients.

Objective To determine the clinical value of cytological diagnosis of the impressed specimens in the radical gastrectomy for patients with gastric cancer. Methods During radical gastrectomies for 52 patients with gastric cancer, 232 impressed specimens were taken from the tumor tissues,paratumor tissues, proximal and distal incision margin,and partial lymph nodes respectively, to detect if there was tumor cells by cytological examination and compared with the pathology. Results In the tumor tissues,there were no pseudo positive specimen,and the pseudo negative specimen occurred in 1.In the incision margin,there was no pseudo negative specimen,while 12 pseudo positive specimens were found.In the paratumor tissues,there were 3 pseudo negative and 10 pseudo positive specimens.In all specimens,the sensibility was 92.8%; specificity was 85.8%; accuracy rate was 87.9%. Conclusions Cytological examination of the impressed specimens in radical gastrectomy is helpful in detecting residue tumor tissue in the incision margin, especially in hospitals without rapid pathologic examination.If cytological examination is combined with rapid pathological examination,the results will be better.

Objective To obtain the data of injury among children aged 0~6 years in Haidian area and Pinggu area of Beijing.Methods A total of 2970 children aged 0~6 years in Haidian District(city area) and 2558 in Pinggu District (rural area)were investigated by cluster sampling method.Results A total of 5528 children were investigated,the incidence of injury was 8.64%.The incidence rate of aged 0~6 years child injury in Haidian District was10.54%,which was much higher than that in Pinggu District (6.45%).The top five causes of injury were falls,animal bites,burn/scalds,injury by sharp articles and struck/hit by falling object in Pinggu District.The top five causes of injury were falls,animal bites,injury by sharp articles,burn/scalds and blunt in Haidian District.the sequences of injury were similar between the two districts.The incidence rate of burns/scalds was higher in Pinggu District than that in Haidian District.The types of injury were little different between children who lived at home and children who lived in kindergarten.The types of injury were not too different between genders.Falls was the leading cause of injury for all children(in cities or in countries,boys and girls,in different age group).Conclusion The incidence rates of injury and types of injury are different in different areas,genders and age groups.So the intervention of injury should be different.

Objective To detect the immune effect of FbaAmAb2 against the surface protein of group A Straptococcus (GAS),and explore the pathogenesis and therapy of GAS infections.Methods By subclonal and bacterial ELISA,the positive hybridoma cells were screened that can produce better titers of FbaAmAb2 against GAS-surface FbaA protein,and were injected into the peritoneal cavities of BALB/c mice to produce ascites.The collected ascites were performed to dilute,as follows,original ascite,1:2,1:4,1:8,and 1:16 to test tube agglutination.Based on the results,we selected appropriate dilution to passively immunize mice,and then challenged the mice with GAS,evaluating FbaAmAb2 neutralizing ability with GAS in mice by the survival rate of the immunized mice.Whether FbaAmAb2 could inhibit the binding of factor H to GAS was confirmed by the invasive inhibition assay.Results The IgG titer of bacteria solution ELISA is 1:160 and the titer of tube agglutination is 1∶8.The protect rates of FbaAmAb2 on preventing mice with GAS infections are as follows:66.67％ in original ascite and 1:2 diluted groups,and 50％ in 1:4 diluted group.Mice in each experimental group were evoked significantly protective immune responses compared with the PBS control by SPSS analysis.FbaAmAb2 can competitively inhibit factor H binding to the surface proteins FbaA of GAS,which decreased the entry of GAS into the cytoplasm of human epithelial cells through the binding of factor H.Conclusion FbaAmAb2 is promising to be used in emergent prevention or the clinical therapy for GAS infection and it is promising starting points for pharmacologic targeting and further development of new therapeutic agents for GAS.