OBJECTIVETo investigate the effects of tributyltin chloride (TBT) and triphenyltin chloride (TPT) on rat testicular Leydig cells.

METHODSThe rat Leydig cells (LC-540) were incubated with 0 to 80 nmol/L TBT and TPT for 24 to approximately 96 h, and then the cell viability was determined by MTT. DNA fragmentation ladder formation of cell apoptosis was examined by agarose electrophoresis. Effects of chelator of intracellular Ca2+ (BAPTA) and the inhibitors of PKA, PKC and TPK on cell apoptosis induced by TBT were observed. Effects of TBT on testosterone production in primary cultured rat Leydig cells treated with or without hCG were detected.

RESULTSTBT and TPT suppressed Leydig cell survival in a time- and dose-dependent manner. The suppressive effects of TBT and TPT on the cell survival was caused by apoptosis which was determined by DNA ladder formation. The apoptotic effect of TBT was possibly mediated by the rise in intracellular Ca2+ because it could be blocked by BAPTA, the chelator of intracellular Ca2+; PKA, PKC and TPK inhibitors did not prevent the apoptotic effects induced by TBT. TBT markedly suppressed testosterone production of primary cultured rat Leydig cells with or without hCG stimulation.

CONCLUSIONTBT and TPT induced apoptosis in rat testicular Leydig cells possibly through increasing intracellular Ca2+. TBT reduced the testosterone production of rat Leydig cells.

Animals ; Apoptosis ; drug effects ; Calcium ; metabolism ; Cell Line ; Dose-Response Relationship, Drug ; Environmental Pollutants ; toxicity ; Leydig Cells ; drug effects ; metabolism ; secretion ; Male ; Organotin Compounds ; toxicity ; Rats ; Testosterone ; secretion ; Trialkyltin Compounds ; toxicity

A growing body of evidence suggests that p300 histone acetyltransferase plays important roles in cancer cell differentiation and proliferation. Here, we employed structure-based hierarchical virtual screening method to identify novel lead compounds of p300 histone acetyltransferase. From a screening library containing approximate 100 000 diverse druglike compounds, 33 compounds were chosen for experimental testing and one compound, 4-acetyl-2-methyl-N-morpholino-3,4-dihydro-2H-benzo[b][1, 4]thiazine-7-sulfonamide (17), showed as micromolar inhibitor. Based on its predicted binding pose, we investigated its binding characteristics by designing two series of structural modifications. The obtained structure-activity relationship results are consistent with the predicted binding model. We expect that the identified novel p300 histone acetyltransferase inhibitors will serve as starting points for further development of more potent and specific histone acetyltransferase inhibitors.
Drug Design ; Enzyme Inhibitors ; chemical synthesis ; chemistry ; Molecular Structure ; Morpholines ; chemical synthesis ; chemistry ; Structure-Activity Relationship ; Sulfonamides ; chemical synthesis ; chemistry ; p300-CBP Transcription Factors ; antagonists & inhibitors ; chemistry

The expression vectors of the gene encoding ScFv-2F3 were transformed into E. coli BL21(DE3). Clones of higher expression were first selected, then were grown in the presence of IPTG at 37 degrees C to induce its expression. The culture conditions were carefully optimized. It was found that optimal conditions were as follows: the induction was started as OD590 reached to 1.0-1.8; the concentration of IPTG was 0.3-0.5 mmol/L and induction time is 7 h. The yield of ScFv-2F3 expressed in the selected clones is about 20% of the total proteins. The optimal culture conditions were successfully applied to fermenter of 50 L. The conditions of washing the inclusion bodies were also optimized. A two-step method was used to renature the inclusion body. The expression product of interest and its biological activities were characterized with Western blotting and ELISA. A novel selenium-containing single-chain abzyme with GPX activity was prepared.
Antibodies, Catalytic ; biosynthesis ; chemistry ; genetics ; isolation & purification ; Bioreactors ; microbiology ; Cloning, Molecular ; Escherichia coli ; Gene Expression ; Immunoglobulin Fragments ; biosynthesis ; chemistry ; genetics ; isolation & purification ; Inclusion Bodies ; metabolism ; Protein Folding ; Protein Renaturation ; Recombinant Proteins ; biosynthesis ; chemistry ; genetics ; isolation & purification ; Selenium ; metabolism

OBJECTIVETo evaluate the effect of bridged free latissimus dorsi musculo-cutaneous flap on repairing of soft tissue defect in the lower extremity.

METHODSSeven patients with extensive soft tissue defects in the lower extremities were enrolled in the clinical investigation. The defects were all repaired with bridged free latissimus dorsi musculo-cutaneous flaps. The condition of the blood vessels in the flaps and the healthy extremities was examined with ultrasound Doppler before the operation to assure the blood circulation of grafted flap. After debridement, the flap was designed in accordance with the size and the depth of the wound. Then the transplantation of the flaps were done. The operative indication and points for attention were summarized thereafter.

RESULTSAll the 7 flaps survived. All patients recovered well with satisfactory function and external appearance, except flap reduction was done in 2 patients due to undue thickness of the flaps. Indications for operation: (1) Patients with anterior or posterior tibial artery injury in the injured lower extremity in which arterial transplantation was not possible to allow the free transplantation of a skin flap. (2) The injury was extensive and deep, with the injurious condition of the blood vessels indeterminable and no healthy artery could be found for anastomosis with a donor artery. (3) No vascular injury could be identified in the contralateral healthy extremity. Points for attention included that the blood supply of the flap to be transferred should be adequate, and the survival of the flap after division of the pedicle should be assured. The length of the flap to be transferred should be longer by 10% than the distance between the site of transplantation in the lower extremities and the donor area; and the donor area should be larger by 20% than the recipient area. The skin area of the flap to be transferred should be broad enough avoid tension so that there would be no pressure on the blood vessels. Pay attention to the blood supply of the flap after operation, and the recipient limb should be properly immobilized.

CONCLUSIONThe repair of extensive soft tissue defect in the lower extremity with bridged free latissimus dorsi musculo-cutaneous flap could be satisfactory. Proper wound management, broad flap, stable immobilization were the pivotal points for the success of the operation.

Adolescent ; Adult ; Back ; Child ; Child, Preschool ; Female ; Humans ; Leg Injuries ; surgery ; Lower Extremity ; Male ; Middle Aged ; Muscle, Skeletal ; transplantation ; Reconstructive Surgical Procedures ; Soft Tissue Injuries ; surgery ; Surgical Flaps ; Tissue Transplantation

OBJECTIVEThe aim of this study was to observe the coexistence of periodontal bacteria DNA (Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Tannerella. forsythensis) in coronary atheromatous plaques and subgingival plaques in patients scheduled for coronary artery bypass graft.

METHODSCoronary atheromatous plaque and subgingival plaque samples were obtained from 51 patients and examined by polymerase chain reaction (PCR) technique with specific primers for periodontal bacteria.

RESULTSP. gingivalis (33%), T. forsythensis (31%), P. intermedia (18%) and F. nucleatum (12%) were detected while A. actinomycetemcomitans was negative and not found in coronary atheromatous plaques; T. forsythensis (84%), F. nucleatum (78%), P. intermedia (59%), P. gingivalis (39%) and A. actinomycetemcomitans (22%) were detected in subgingival plaque samples. Coexistence of periodontal bacteria DNA in coronary atheromatous and subgingival plaque samples was evidenced in 32 patients.

CONCLUSIONSThe coexistence of T. forsythensis, F. nucleatum, P. intermedia, P. gingivalis in coronary atheromatous plaques and the subgingival plaque samples in CAD patients could suggest a potential role for periodontal pathogenic bacteria in atherosclerosis disease process.

Adult ; Aged ; Bacteroides ; isolation & purification ; Coronary Artery Bypass ; Coronary Artery Disease ; microbiology ; surgery ; DNA, Bacterial ; analysis ; Dental Plaque ; microbiology ; Female ; Fusobacterium nucleatum ; isolation & purification ; Gingiva ; microbiology ; Humans ; Male ; Middle Aged ; Polymerase Chain Reaction

Epilepsy is one of the most common and debilitating neurological diseases that affects more than 40 million people worldwide. Genetic factors contribute to the pathogenesis of epilepsy. Molecular genetic studies have identified 15 disease-causing genes for epilepsy. The majority of the genes encode ion channels, including voltage-gated potassium channels KCNQ2 and KCNQ3, sodium channels SCN1A, SCN2A, and SCN1B, chloride channels CLCN2, and ligand-gated ion channels CHRNA4, CHRNB2, GABRG2, and GABRA1. Interestingly, non-ion channel genes have also been identified as epilepsy genes, and these genes include G-protein-coupled receptor MASS1/VLGR1, GM3 synthase, and proteins with unknown functions such as LGI1, NHLRC1, and EFHC1. These studies make genetic testing possible in some patients, and further characterization of the identified epilepsy genes may lead to the development of new drugs and new treatments for patients with epilepsy.
Chloride Channels ; genetics ; Epilepsies, Myoclonic ; genetics ; Epilepsy ; genetics ; Epilepsy, Absence ; genetics ; Humans ; KCNQ2 Potassium Channel ; genetics ; KCNQ3 Potassium Channel ; genetics ; NAV1.1 Voltage-Gated Sodium Channel ; NAV1.2 Voltage-Gated Sodium Channel ; Nerve Tissue Proteins ; genetics ; Sodium Channels ; genetics

OBJECTIVETo investigate the possibility of marrow derived multipotent adult progenitor cells (MAPCs) differentiating into hepatocytes by co-culturing with human hepatocyte line L02, and to evaluate the potential use of MAPCs in tissue-engineering either experimentally or clinically.

METHODS(1) Co-culturing without cell-to-cell contact: MAPCs and L02 hepatocytes were spread on coverslips separately (both with a cell density of 1x10(5)/ml), and then they were put in a culture dish (10 cm). The expressions of Alb, AFP, CK18, and CK19 in MAPCs were detected by immunocytochemistry at different time points. A separate culture of L02 hepatocytes served as a positive control and a separate culture of MAPCs served as a negative control. (2) Co-culturing with cell-to-cell contact: MAPCs labeled with CFSE were mixed with L02 hepatocytes (both with a cell density of 1x10(4)/ml), and then the mixed cells were seeded on specific dishes for detection by laser scanning confocal microscope (LSCM). Five days later, the cells were double-stained with SABC-Cy3. The expressions of Alb, AFP, CK18 in MAPCs were observed under LSCM. Similarly, separately cultured L02 hepatocytes served as a positive control and separately cultured MAPCs served as a negative control.

RESULTS(1) Results of co-culturing without cell-to-cell contact: On the first day, the MAPCs expressed a high level of AFP. Then AFP expression tapered daily and there was hardly any expression of AFP on day 7. The expression of Alb was very weak on day 1, but increased significantly by day 3, reached its peak on day 5, and still maintained a high level on day 7. The initial expression of CK18 appeared on day 5 and reached a higher level on day 7. The expression of CK19 was always negative. The positive control cells had a high expression of Alb and CK18, while there was a weak expression of AFP and a negative expression of CK19. The negative control cells had no expressions for the four markers. (2) Results of co-culturing with cell-to-cell contact: On day 5, there were three colors of fluorescence under LSCM: yellow cells were MAPCs differentiating into hepatocytes; green cells were undifferentiated MAPCs; red cells were L02 hepatocytes. The result showed that Alb and CK18 were expressed in many cells and AFP appeared in only a few cells.

CONCLUSIONHuman MAPCs can be induced to differentiate into mature hepatocyte-like cells by co-culturing with L02 hepatocytes, either with or without cell-to-cell contact, but the former way may be more effective.

Bone Marrow Cells ; cytology ; Cell Differentiation ; Cells, Cultured ; Coculture Techniques ; Hepatocytes ; cytology ; Humans ; Multipotent Stem Cells ; cytology

OBJECTIVETo explore the association between metabolic syndrome (MS) and risk of cardiovascular disease events (CVD) in patients with ischemic stroke.

METHODA total of 1087 patients with ischemic stroke were enrolled from 5 community-based medical centres and underwent baseline evaluation on risk factors of stroke during the period of Jan. 2003 to Dec. 2006. After baseline survey, all patients were followed up until Dec 31, 2008 and new CVD events were recorded. MS was defined using CDS criteria. Proportional hazard models were used to assess the HRs and 95% CI of CVD events associated with MS and other components.

RESULTSThe prevalence of MS was 40.4% at baseline. During an average follow-up of 3.5 years, 178 patients developed new CVD events. After adjusted for age, gender, smoking, drinking, marriage status, education level, hospitalization, recurrence of stroke, stroke duration, depression, cognition impairment and ADL, MS remains the independent predictor for the risk of CVD events. Compared with patients with non-MS, the risk of CVD events increased by 44% (HR: 1.44, 95%CI: 1.06 - 1.95). The risk of CVD also increased with the number of MS components. Compared with patients with 1 or less than 1 components of MS, the risk of CVD events increased by 30% (HR: 1.30, 95% CI: 0.83 - 2.04) in those with 2 components and by 69% (HR: 1.69, 95%CI: 1.11 - 2.56) in those with 3 or more components of MS. Hypertension and hyperglycemia and impaired fasting glucose also served as independent risk factors for CVD event (all P < 0.001).

CONCLUSIONSMS was independently associated with increased risk of CVD events in patients with ischemic stroke. There was a dose-response relationship between the numbers of MS components and the risk of CVD event.

Aged ; Brain Ischemia ; complications ; epidemiology ; Cardiovascular Diseases ; complications ; epidemiology ; metabolism ; China ; epidemiology ; Female ; Humans ; Male ; Metabolic Syndrome ; complications ; epidemiology ; Middle Aged ; Prevalence ; Prospective Studies ; Risk Factors ; Stroke ; complications ; epidemiology

Achondroplasia ; genetics ; Adult ; Amino Acid Sequence ; Genes, Dominant ; Humans ; Male ; Molecular Sequence Data ; Mutation ; Polymorphism, Restriction Fragment Length ; Receptor, Fibroblast Growth Factor, Type 3 ; chemistry ; genetics ; physiology

Adult ; Aged ; Bacteria ; isolation & purification ; Coronary Artery Disease ; microbiology ; DNA, Bacterial ; analysis ; Humans ; Middle Aged ; Periodontitis ; microbiology