Objective To investigate the prevalence of mtDNA A3243G mutation in the early onset diabetics in Tianjin for exploring the relationship between mtDNA mutation and diabetes. Methods 348 kinship-free diabetics whose ages at onset were less than 45y were randomly recruited, with 207 control subjects. The PCR-RFLP and cloning techniques were applied to screen the A to G substitution at nucleotide 3243 of mtDNA tRNALEU（UUR）. Meanwhile, the clinical and genetic analysis was done from one pedigree. Results We detected two diabetics harboring the well-known a3243g mutation with the mutation frequency of 0.6%. While the mutation frequency in diabetics with （positive） family history was 1.2%, and zero in the control subject. The proband showed a typical （picture） of mitochondrial diabetes mellitus, while the other family members harboring the same （mutation） had heterogeneous presentation. Conclusion The prevalence of mtDNA A3243G mutation in early onset diabetes of Tianjin is low, but relatively higher in diabetes complicated with other （symptoms） of mitochondrial disease. Furthermore, its heterogeneity would decrease with aging in （mitotic） tissue

Objective To explore the relationship between MTHFR gene polymorphism and ~diabetic cardiovascular disease. Methods The MTHFR C677T and MTHFR A1298C gene polymorphisms were detectd by PCR-RFLP to compare MTHFR C677T and MTHFR A1298C genotype distribution and allele frequencies among three groups. Enrolled were 84 healthy individuals (NC) and 158 T2DM subjects who were classified into two groups, i.e. non-diabetes complications (NDC) and ~diabetic cardiovascular disease (DC). Results Patients with diabetic cardiovascular disease had higher frequencies of MTHFR C677T mutation genotype and T allele than control and non-diabetes complications subjects, and had lower folate levels (P<0.001). There were no significant differences in MTHFR A1298C genotype distribution and allele frequencies among three groups (P>0.05). A logistic regression analysis demonstrated that MTHFR C677T and age were the independent predictors of DC. Conclution There is a strong correlation between the polymorphism of MTHFR C677T and diabetic cardiovascular disease.

Objective To investigate clinical traits of 2 families members habouring mtDNA 12026A→G mutation based on our previous studies.Methods 25 members in 2 families with probands with mtDNA 12026 mutation were examined.All their clinical and biochemical data were collected.Total genome was extracted conventionally from peripheral leucocytes of all participants,and PCR-RFLP techniques were applied to screen A to G substitution at nucleotide 12026 of mtDNA in ND4 region.Results We found 13 individuals habouring the 12026 A→G mutation in 2 pedigrees,all without deafness.Among them,5 with diabetes were found.Interestingly,we found 3 individuals with hyperthyroidism in one family(one also combined with diabetes).Conclusions Our findings suggest that diabetic families with mtDNA 12026 A→G mutation in ND4 region can have different clinical pictures,and may involve in autoimmune diseases

Objective To investigate clinical traits of 2 families members habouring mtDNA 12026A→G mutation based on our previous studies.Methods 25 members in 2 families with probands with mtDNA 12026 mutation were examined.All their clinical and biochemical data were collected.Total genome was extracted conventionally from peripheral leucocytes of all participants,and PCR-RFLP techniques were applied to screen A to G substitution at nucleotide 12026 of mtDNA in ND4 region.Results We found 13 individuals habouring the 12026 A→G mutation in 2 pedigrees,all without deafness.Among them,5 with diabetes were found.Interestingly,we found 3 individuals with hyperthyroidism in one family(one also combined with diabetes).Conclusions Our findings suggest that diabetic families with mtDNA 12026 A→G mutation in ND4 region can have different clinical pictures,and may involve in autoimmune diseases.

Objective To investigate the relationship between point mutation of nt3394 T→C of mitochondrial DNA ND1 and type 2 diabetes mellitus(DM) in the elderly. Methods Two hundred and eight patients with type 2 diabetes mellitus and 180 subjects with normal glucose tolerance without DM family history were included. The mutation was determined by polymerase chain reaction and restriction fragment length polymorphism(PCR RFLP). Results The point mutation of nt3394 T→C of mitochondrial DNA ND1 was found in 9 of 208 elderly patients with type 2 DM(4 3%), and 1 of 180 controls (0 6%). The incidence of this mutation was significantly different between the two groups ( P

Objective To investigate the expression of RCAS1 in the tissues of normal cervix,cervical intraepithelial neoplasia(CIN)and cervical cancer,and to explore their role in the pathogenesis of cervical carcinoma.Methods The expression of RCAS1 protein was analyzed by immunohistochemical S-P method in 89 cases of cervical carcinoma,82 cases of CIN,and 24 cases of normal cervical specimens.Results Immunohistochemistry showed that the expression rate of RCAS1 in normal cervix,CINⅠ,CINⅡ-Ⅲ and cervical cancer were 0.00,0.314,0.447 and 0.775,respectively.RCAS1 expression tended to increase along with tumor progression.The expression of RCAS1 in cervical cancer was significantly related to the histological grade,the tumor in different sizes(≥4cm or 0.05)betwan patients' age,clinical stage and histological classification.Conclusions The data suggested that RCAS1 might play an important role in the pathoqenesis and development of cervical carcinoma.Over expressed RCAS1 could be regard as an important parameter in malignant degree and poor prognosis of cervical cancer.

Objective A easier and faster rat model of organ injury caused by cardiopulmonary bypass was developed to provide a stable and economical experimental platform for the research of CPB ( cardiopulmonary bypass ) .Methods Twenty male Sprague-Dawley rats weight between (288.8 ±26.1)g were randomly divided into sham group and CPB group .The right external jugular vein and bilateral femoral arteries were punctured in sham group ,while the rats in CPB group underwent 60 minutes cardiopulmoanry bypass with the perfusion flow no less than 120 mL· kg-1· min-1 .And the weight ,blood pressure ,blood gas and electrolytes were monitored and analyzed throughout the experi -ment.The serum,lung,liver and kidney were collected at 72 h after the operation for further detection .Results The rat model of cardiopul-monary bypass was established successfully with 90%survival rate .The changes of blood gas during intraoperative and postoperative were in accordance with clinical reality and the injury of lung ,liver and kidney were confirmed by serum results and pathological changes .Conclu-sion The model has the characteristics of lower weight ,less priming solution ,no need for endotracheal intubation and easier to assemble the bypass line ,furthermore the methods of puncture and CPB were exhibited through a large number of pictures .As a consequence ,the difficul-ties of establishing the rat model of CPB were reduced significantly .The changes of pathophysiology during intraoperative and postoperative were similar to clinical realities ,the survival rate and organ injury were satisfactory .So it can be used as an effective tool for the research of organ injury and the evaluation of therapeutic strategies associated with CPB .

Objective Thoracic cavity fistula following esophagus carcinoma resection is a serious complication with a high mortality.This study aims at a better therapy for thoracic cavity fistula following esophagus carcinoma resection by summarizing the ex-perience with the four-tube strategy ( jejunal fistula tube, stomach tube, chest drainage tube, and nasal fistula tube) in the treatment of the complication. Methods We retrospectively analyzed the clinical data about 62 cases of thoracic cavity fistula following esopha-gus carcinoma resection, 35 treated with the four-tube strategy ( treatment group) and the other 27 with the three-tube ( stomach tube, chest drainage tube, and nasal fistula tube) method ( control group) .We compared the hospital days, wound healing time, mortality, and incidence of anastomotic stenosis at 6 months after operation between the two groups of patients. Results Compared with the controls, the treatment group showed remarkable decreases in the hospital days (P<0.05), wound healing time (P<0.05), and mortality (P<0.05), but no statistically significant difference was observed in the incidence rate of anastomotic stenosis at 6 months after operation between the two groups of patients ( P >0.05 ) . Conclusion Compared with the three-tube method, the four-tube strategy has the advantages of shorter healing time and lower mortali-ty, and therefore is preferable for the treatment of thoracic cavity fis-tula following esophagus carcinoma resection.

Objective This study investigates feasibility of replacing urinary epidermal cells with foreskin epidermal cells to reconstruct engineered anterior urethra with acellular collagen matrix. Methods A cellular collagen matrices were generated from allogeneic rabbit bladder submucosa.In 6 rabbits,autologous foreskin epidermal cells isolated,in vitro expanded and labeled with Brdu before seeding onto a tubular collagen matrix of 2 cm long.In 12 male rabbits, urethral mucous defect was created and urethroplasty was performed with tubular collagen matrix seeded with epidermal cells (n=6) in experimental group or matrix without cells seeding (n=6) in control group.Urethrography was performed at 1,2,and 6 months postoperatively.Urethras grafts were harvested and analyzed grossly and histologically at the same time points. Results In control group, gross and urethrography demonstrated urethral stricture of repaired defects at all time points.Histology showed a single layer of epidermal cell with unorganized and crassitude muscle fiber bundles in submucosa layer.In contrast, urethrography showed a patency of repaired urethral defect with the maintenance of a wide urethral caliber in experimental group without sign of strictures at any time points. Histologically,the graft seeded with epidermal cell had a multi-layers epidermal cells by 1 month,which demonstrated the presence of a epidermal cells layer with a normal submucosa structure.The immunoflorescent staining of Brdu confirmed the presence of implanted epidermal cells.In addition, the implanted cells also express keratin when stained with anti-pancytokeratins AE1/AE3 antibody.At 2 month,histology showed relatively normal urethral architecture with vessel abundance in submucosa structure.The staining of Brdu showed decreased at implanted epidermal cells and epithelial phenotypes were confirmed immunocytochemically using pancytokeratins AE1/AE3.At 6 month,the graft structure show multi-layers epidermal cells without staining of Brdu. Conclusions Urethra reconstruction was better achieved with cell seeded acellular collagen matrix when compare with acellular matrices alone.Foreskin epidermal cells seems succeeded in replacing urethral epidermal cells for urethra reconstruction.

Objective To investigate the expression of recombinant human adiponectin(ADPN) using baculovirus expression system in Sf 9 insect cell line.Methods PCR method was used to amplify human ADPN gene,and then the gene was inserted into plasmid pFastBacl.Recombinant plasmid was transformed into E.coli DH10Bac containing a shuttle vector,Bacmid.Recombinant shuttle vector Bacmid-ADPN was obtained by site-specific transposition and transfect Sf 9 cells.To identify the expressed product by SDS-PAGE、Western blot.Results Sf 9 cells infected by the recombinant baculovirus was observed for morphological changes.The Mw of recombinant protein was about 30 ku and Western blot proved that expressed protein could combine with ADPN polyclonal antibody.ConclusionHuman ADPN gene was successfully expressed in Sf 9 cells,and thus provided the material for studying its bioactivity and mechanism of action.