Objective:To construct a mutant of human proinsulin with signal peptide code,so that it may be expressed in eukaryotic system to produce mature human insulin in COS-7 cells.Methods:Human proinsulin gene was mutated by the polymerase chain reaction (PCR) for the site-directed mutagenesis method.Two furin recognized sites of Arg-X-Lys-Arg-motifs were introduced into human proinsulin.Signal peptide,Kozak sequence and restriction enzyme cutting site were added and cloned into pcDNA3.1(+) vector and verified by sequencing analysis.Eukaryotic expression vector was constructed and transfected into COS-7 cells.Insulin were evaluated respectively with RT-PCR,radioimmunoassay and Western blot analysis.Results:The sequencing analysis showed that the mutation sites were correct,and the mutant of human proinsulin was constructed successfully.Human insulin mRNA was highly expressed in transfected COS-7 cells,and secretive recombinant human insulin was detected in the culture supernatant of transfected COS-7 cells by Western blotting.Conclusion:A mutant human proinsulin eukaryotic expression vector is obtained and secretive recombinant human insulin is successfully expressed in COS-7 cells.

OBJECTIVE:To discuss the accuracy of detecting tacrolimus(FK506) concentration in whole blood by ELISA and its application in organ transplantation.METHODS:The ELISA was used to detect FK506 valley point concentrations in 5 liver transplanted recipients at different postoperative periods .Based on the results combined with clinical findings, the dosages were adjusted.The accuracy of the method and guidance action in FK506 clinical application were described.RESULTS: In 131 detections of 350 samples, the results of quality control samples were all in the control range .The detecting RSDs of high and low concentration quality control samples were 9.09% and 11.90%, respectively.The control range of FK506 concentrations in whole blood were similar to that reported in literature .CONCLUSION :ELISA is a sensitive and specific method for routine FK506 concentration monitoring in whole blood,which is suitable for application in hospital.

Objective To evaluate the effects of exogenous adenovirus-mediated gene transfer of human apM1 gene (Ad-apM1) on malonaldehyde (MAD) and superoxide dismutase (SOD) levels in human umbilical vein endothelial cells (HUVEC). Methods HUVEC proliferation was measured by MTT after infection by AdapM1. Adiponetin protein level in cell culture medium of HUVEC cells infected with Ad-apM1 was detected by double antibody sandwich ELISA. The levels of MAD and SOD were measured by chromatometry. Results MTT assay showed that Ad-apMl had no effect on HUVEC proliferation. Human adiponectin protein levels in cell culture medium significantly increased after HUVEC was infected with Ad-apM1 for 24, 48, and 72 h, along with decreased MAD and increased SOD ( all P<0.05 ). MAD levels markedly increased and SOD levels decreased after HUVEC were incubated with 100 μmol/L H2O2 for 6, 12, and 24 h, and these reactions were reversed by AdapM1 transfection (all P<0.05 ). Conclusions HUVEC infected with Ad-apM1 effectively secrete human adiponectin. Ad-apM1 exerts antioxidation effect and antagonizes H2O2 -induced endothelial injury.

Objective To study the effect of phosphodiesterase inhibitor pentoxifylline (PTX) on type 1 diabetes in NOD mice and its possible mechanism.Methods Blood glucose, urinary glucose, insulitis and incidence of diabetes were investigated in NOD mice treated with PTX, insulitis was observed by HE staining and the expressions of IFN ?, TNF ?, IL 10 in pancreas were measured by RT PCR technique. Results The incidence of diabetes in the PTX group was 30.0% which was significantly lower than 67.9% in the control group (P

Type 2 diabetic rats were treated with recombiriant adenovirus expressing human adiponectin (Ad-APN).The results showed that the levels of both plasma C-reactive protein and soluble intercellular adhesion molecule-1 were significantly decreased in Ad-APN-treated diabetic rats compared with those of diabetic rats [(18.73±4.66 vs 23.60±4.25)mg/L and (14.91±1.79 vs 19.09±2.95) ng/ml,both P＜0.01].These results suggest that increased serum concentration of adiponectin may improve the status of inflammation in diabetes mellitus.

Metabolic syndrome (MS) is a series of metabolic disorder syndrome relating to insulin resistance, which is a risk factor for atherosclerosis and vascular disease. It includes central obesity, lipid metabolism disorders, high blood pressure, high blood sugar and glucose tolerance. Studies have shown that the human's hypothalamus, kidneys, liver and adipose tissue can express functional thyroid stimulating hormone receptor (TSHR) besides thyroid tissue. Thyroid stimulating hormone(TSH) bind to these receptors, involving in the regulation of blood sugar, blood lipids, blood pressure and the formation of fat through multiple targets and signal pathways.

Amyloidosis ; Cardiomyopathy, Hypertrophic ; Humans

Objective To analyze the overactive bladder symptom score (OABSS) in evaluating bladder dysfunction in the different stages among type 2 diabetes mellitus patients,and to explore the value of bladder hyperactivity symptom score in screening early diabetic bladder dysfunction.Methods A total of 1 157 patients with type 2 diabetes mellitus,aged 40-88 yearswith mean age of 60.2 years,were enrolled from October 2013 to October 2016.The survey included the patients' characteristics,past history,current history,OABSS and quality of life (QOL) index scores.T test,single factor analysis of variance and multiple regression analysis are used to analyze the results.Results As many as 1 157 were qualified for final statistical analysis.OABSS is 1.94 ± 1.23 in group with diabetes duration ＜ 10 years,3.24 ± 1.45in group with diabetes duration 10-20 years,and 4.00 ± 1.72 in group with diabetes duration ＞ 20 years.The differences of OABSS in the different duration of diabetes was statistically significant.As diabetic duration increased,OABSS value increased (F =48.419,P ＜ 0.001).The difference of OABSS in the different HbA1c level,age and concurrent peripheral neuropathy was statistically significant.There was no significant difference of OABSS in the different BMI and distinct therapies.There was no significant difference of OABSS in diabetes with hypertension and without hypertension,with cardiopathy and without cardiopathy,with cerebrovascular disease (CVD) and without CVD,with hyperhpemia and without hyperlipemia.The significant factors were used to make multivariate analysis.The results showed that the duration of diabetes,HbA1 c level,age,peripheral neuropathy were still statistically significant.Standardized partial regression coefficient of diabetic duration was 0.366.OABSS was positively correlated with QOL score (r =0.434,P ＜ 0.001).Conclusions The related symptoms in OABSS with diabetic bladder dysfunction is correlated with the duration of diabetes,HbA1c level,age,concurrent peripheral neuropathy among type 2 diabetes.The duration of diabetes was the most significant factor.OABSS is likely to be one of the tools to assess the early symptoms of diabetic bladder dysfunction.

Objective:To analyze the clinical symptoms and laboratory characteristics of patients with brucellosis combined with osteoarthritis.Methods:A retrospective analysis was conducted on the medical records of 168 patients with brucellosis who were hospitalized at the Jining Public Health Medical Center, Shandong Province from January 2021 to December 2022. Based on the imaging examination results, they were divided into combined osteoarthritis group and non combined osteoarthritis group. The demographic characteristics, clinical symptoms, and laboratory test results of the two groups of patients were compared and analyzed.Results:Among 168 patients with brucellosis, there were 83 patients with concurrent osteoarthritis, 85 patients without concurrent osteoarthritis, the males and females ratio was 2.73 ∶ 1.00 (123 ∶ 45), and the age was 56 (46, 64) years old. The median age of patients in combined osteoarthritis group was higher than that of patients in non combined osteoarthritis group (58 years vs 53 years, Z = - 2.89, P = 0.004). In clinical symptoms, the incidence of back pain, fatigue, and joint muscle pain in patients of combined osteoarthritis group was significantly higher than that in the non combined osteoarthritis group [75.9% (63/83) vs 56.5% (48/85), 30.1% (25/83) vs 15.3% (13/85), 47.0% (39/83) vs 17.6% (15/85), χ 2 = 7.07, 5.27, 16.58, P < 0.05]. In laboratory tests, the erythrocyte sedimentation rate (ESR) and hypersensitive C-reactive protein (hs-CRP) levels in patients of combined osteoarthritis group were higher than those in the non combined osteoarthritis group [27.0 (17.0, 34.0) mm/h vs 21.0 (10.5, 34.0) mm/h, 22.7 (14.3, 43.4) mg/L vs 17.9 (10.8, 34.2) mg/L, Z = - 2.51, - 2.00, P < 0.05]. Among patients with combined osteoarthritis, combined spondylitis accounted for the highest proportion (55.4%, 46/83), followed by peripheral arthritis (51.8%, 43/83). Conclusion:Osteoarthritis is a common complication of brucellosis, and patients of brucellosis with concurrent osteoarthritis have clinical features such as lower back pain, fatigue, and joint muscle pain, with significantly elevated of ESR and hs-CRP level.