Objective To investigate the effects of cinobufacini on TGF-β1-induced epithelial-to-mesenchymal transition of human colorectal carcinoma cells in vitro.Methods The cultured colorectal carcinoma cell line(SW480) was divided into the control group,TGF-β1 (10 ng/mL) individual treatment group and co-treatment groups with TGF-β1 (10 ng/mL) + cinobufacini (2.5,5,10,20,40,80 mg/mL),which were cultured in vitro for 48 h.The proliferation of the cells were measured by CCK8 assay.The morphological changes were observed by inverted phase contrast microscope.The ability of cell invasion and migration was detected by Transwell assay.The mRNA and protein expressions of E-cadherin and Vimentin were detected with QRT-PCR and Western Blot.Results (1)Compared with the TGF-β1 (10 ng/mL) individual treatment group,TGF-β1 (10 ng/mL) + cinobufacini (10,20,40,80 mg/mL) co-treatment groups significantly had significantly proliferation inhibitory effect on SW480 (P＜0.05).(2) Compared with the normal control group,TGF-β1 individual treatment group and TGF-β1 (10 ng/mL) + cinobufacini(2.5 mg/mL) group exhibited classical mesenchymal phenotype,while the TGF-β1 (10 ng/mL) + cinobufacini (5 mg/mL) co-treatment group showed classical epithelial phenotype.(3) The ability of invasion and migration in the TGF-β1(10 ng/mL)+ cinobufacini(2.5,5 mg/mL) co-treatment group were significantly weakened compared with the TGF-β1 individual treatment group (P＜0.01).(4) QRT-PCR and Western Blot results indicated that compared with the normal control group,the Vimentin expression in the in the TGF-β1 individual treatment group was significantly increased and the E-cadherin expression was significantly decreased.Furthermore,compared with the TGF-β1 individual treatment group and control group,the Vimentin expression level in the TGF-β1 (10 ng/mL) + cinobufacini (2.5,5 mg/mL) groups was significantly decreased and E-cadherin expression was significantly increased.Conclusion Cinobufacini can inhibit TGF-β1-induced cell proliferation in human colorectal carcinoma SW480 cells,and its mechanism may be related with promoting E-cadherin expression increase,meanwhile decreasing the vimentin expression,thus inhibiting the EMT process induced by TGF-β1.

With the development of modern medicine,to see a doctor is no longer a simple process of diagnosis or taking drugs.Human have deeply recognized medical science and health from a simple biomedical model to a social,psychological and biomedical model.On the one hand,people seek help from medical science.On the other hand,people fear that malpractice could occur.It has been in a love-hate mentality for people to seek medical and legal relief.In the new era,physician-patient relationship needs to be recognized again from medical staff,patients and legal angles.The combination of medicine and law is the requirement of development of the times.No matter how strong the law is,and no matter how much we hope the legislation is from strength to strength,moderation is still the virtues of the law.Medical perspective is still required in the examination of medical activity.

AIM: To investigate the melatonin receptor (MR) gene and protein expression in the human peripheral blood lymphocytes. METHODS: Total RNA of human peripheral lymphocytes was isolated by single step method of acid guanidinium-thiocyanate-phenol-chloform, mt 1 and MT 2 mRNA were determined by the reverse transcription-polymerase chain reaction(RT-PCR). Immunohistochemistry was applied to identify and localize mt 1 and MT 2 protein. RESULTS: 1 1 kb mt 1 mRNA was detected by Northern blot, but 1 1 kb MT 2 mRNA was not detected. The mt 1 cDNA fragment of the expected size of 370 bp was determined and the MT 2 cDNA fragment of the expected size of 320 bp was not determined by RT-PCR. Sequencing result indicated that the positive product coincided with the cDNA of human mt 1. The mt 1 protein was observed by immunohistochemistry. These buffy positive granules were scattered with some areas stronger than the others, and primarily located in cytoplasm and membrane, with rare location in nucleus; meanwhile the MT 2 protein was not observed. CONCLUSION: These results demonstrated the mt 1 mRNA and protein expression in human peripheral leukocytes. It is indicated that melatonin has directly immune-regulative effects on peripheral lymphocytes.

Objective To construct prokaryotic expression systems of genes of intact rotavirus non -structural protein 4 (NSP4) and 86 to 175 amino acid peptide of NSP4 (NSP486-175) and to identify the bio-activities of the recombinant expression products .Methods Colloidal gold assay was used to screen group A rotavirus antibody-positive stools , from which the viral RNA was extracted and transcribed into cDNA .Then NSP4 gene and NSP486-175 gene were amplified by PCR and inserted into pET-28a(+) vector to construct the prokaryotic expression plasmids of pET-28a(+)-NSP4 and pET-28a(+)-NSP486-175 , which were transformed into E.coli BL21( DE3) and inducted by IPTG .The expressed NSP486-175 protein was purified by Ni-NTA af-finity chromatography and examined by Western blot .Fluo-3 AM ester was used to label intracellular free Ca2+( [ Ca2+] i) in Caco-2 cells and laser scanning confocal microscopy was performed to detect the effect of NSP486-175 protein on [Ca2+]i.Results DNA sequencing indicated that NSP4 gene and NSP486-175 gene were consistent with the corresponding sequences in NCBI .The intact NSP4 had difficulty in expressing in E.coli. NSP486-175 protein mainly existed in soluble form with a relative molecular weight of 10×103 .Fluorescence in-tensity and distribution were changed by exogenous addition of NSP 486-175 to cultured Caco-2 cells.Conclu-sion The expression vectors of pET-28a(+)-NSP4 and pET-28a(+)-NSP486-175 were successfully construc-ted and the target protein could induce [Ca2+]i imbalance.This phenomenon preliminarily indicates that the prokaryotic expressed NSP486-175 protein has biological activity and can be further used to elucidate rotavirus pathogenic mechanisms and perform vaccine research .

Four weeks after SD diabetic rats were induced by streptozotocin,skin thickness was obviously reduced with obscure multilayer epithelium features.Moreover,the thickness of epidermic layers in diabetic rat skin was significantly thinner than that ofnornlal rat skin at the eighth week[(0.016±0.006 vs 0.041±0.007)mm,P<0.01].The percentage of G2/M phase cells in epidermic layers of diabetic group was significantly lower than that in the normal group.At the twelfth week,skin microangiopathy was easily detected in the diabetic group.The blood levels of advanced glycation end products(AGEs)and malonialdehyde were significantly increased and glutathione decreased in diabetic rats compared with control rats(aU P<0.01),along with the increased contents of local glucose and AGEs in the skin of diabetic rats.These results suggest that the local accumulation of glucose and AGEs seems to be one of the important mechanisms in the pathogenesis of diabetic skin lesions.

AIM: To explore the histochemical changes o f diabetic skin and the pathogenesis of impaired wound healing in diabetes. METHODS: 54 male Sprague-Dawley (SD) rats weighing 200-220 g wer e randomized into control and STZ-induced diabetic groups. The shaved skin speci mens from the back of rats were collected in 4, 8 and 12 weeks post STZ-inductio n, respectively. Hematoxylin-eosin dye was used for histological examination. Me a nwhile, the skin glucose contents were measured by Beckman's autoanalyzer. Skin AGEs concentrations were assessed by detecting total fluorescence in tissue coll agen and immunohistochemistry assay. RESULTS: The skin thickness in diabetic animals was decreased, w ith the features of multilayer epithelium structure disappeared in epidermis and collagen fibers atrophied, swollen and degenerated in dermis; The inflammatory responses in the dermis of diabetic animals were increased obviously. The result s also revealed that skin glucose contents in diabetic rats [(2.64?1.03)mg/ g skin] were 2-3 times higher than those in the controls [(0.74?0.33)mg/g s kin] (P

Objective:To analysis the correlation of eating speed with obesity.Methods:A total of 644 people aged 40-65 from Caihe Community in Hangzhou were enrolled to collect clinical and demographic data, undergo extensive physical examination and laboratory tests. Participants were divided into two groups according to their eating speed (non-fast and fast). Obesity-related parameters were compared between two groups. Multivariable logistic regression was conducted to explore the relationship between eating speed and obesity after adjusting confounders.Results:Body mass index, waist circumference, and visceral fat area were greater in the fast eating group than non-fast eating group(all P<0.01). After adjusting for age, gender, smoking, alcohol drinking, physical activity level per week, and principal food intake, logistic regression analysis showed that eating fast was correlated with abdominal obesity( OR=1.66, 95% CI 1.11-2.48, P=0.014) and visceral obesity( OR=1.65, 95% CI 1.14-2.39, P=0.007). After stratified by gender, in the group of men, eating fast was correlated with abdominal obesity( OR=2.04, 95% CI 1.07-4.04, P=0.032) and visceral obesity( OR=1.85, 95% CI 1.04-3.31, P=0.037); In the group of women, eating fast was correlated with overweight and obesity( OR=1.59, 95% CI 1.04-2.42, P=0.031). Conclusion:Eating fast is positively associated with obesity. Interventions for reducing eating speed may be effective for weight control.

Objective:To summarize the patient profiles and therapeutic efficacies of ABO-incompatible living-related kidney transplantations at 19 domestic transplant centers and provide rationales for clinical application of ABOi-KT.Methods:Clinical cases of ABO-incompatible/compatible kidney transplantation (ABOi-KT/ABOc-KT) from December 2006 to December 2009 were collected. Then, statistical analyses were conducted from the aspects of tissue matching, perioperative managements, complications and survival rates of renal allograft or recipients.Results:Clinical data of 342 ABOi-KT and 779 ABOc-KT indicated that (1) no inter-group differences existed in age, body mass index (BMI), donor-recipient relationship or waiting time of pre-operative dialysis; (2) ABO blood type: blood type O recipients had the longest waiting list and transplantations from blood type A to blood type O accounted for the largest proportion; (3) HLA matching: no statistical significance existed in mismatch rate or positive rate of PRA I/II between two types of surgery; (4) CD20 should be properly used on the basis of different phrases; (5) hemorrhage was a common complication during an early postoperative period and microthrombosis appeared later; (6) no difference existed in postoperative incidence of complications or survival rate of renal allograft and recipients at 1/3/5/10 years between ABOi-KT and ABOc-KT. The acute rejection rate and serum creatinine levels of ABOi-KT recipients were comparable to those of ABOc-KT recipients within 1 year.Conclusions:ABOi-KT is both safe and effective so that it may be applied at all transplant centers as needed.