Objective:To investigate expression and role of nesprins gene during mouse embryonic stem cell(ESC)differentiation into cardiomyocyte in vitro.Methods:Mouse ESC cultrued in combining 5-azacytidine as inducer with hanging drop cultures and the differentiated cells were identified with cardiac special genes and proteins;The mRNA expression of nesprin-1 and nesprin-2 genes were detected by reverse transcription-polymerase chain reaction(RT-PCR),and sub-cellular location of nesprin-1 protein in nucleus was detected by immunofluorescence in d7,d7+3,d7+6,d7+9 and d7+20.Results:The expression of nesprin-1 and nesprin-2 genes mRNA can be found in the period of mouse ESC differentiation into cardiomyocyte.There is expression peak of nesprin-1 gene mRNA in d7,d7+3,d7+6,then declined gradually to the lowest in d7+20;expression peak of nesprin-2 gene mRNA in d7,then also declined gradually to the lowest in d7+20;The protein of nesprin-1 can be detected in the period of mouse ESC differentiation into cardiomyocyte,and mainly located in nuclear envelope and space around nuclei.Conclusion:The expression peak of nesprin-1 and nesprin-2 gene mRNA is interrelated with the phase of inducing and specialization of ESC differentiation into cardiac muscle cell,it can be speculated that nesprin genes has an important role in maintaining ESC differentiation into cardiac muscle cell.

Acupuncture mechanics in acupuncture manipulation of traditional Chinese medicine is important information for elucidating the essence of acupuncture manipulation and clinically observed acupuncture effects and has some morphological, histochemical and biochemical effects on fibroblasts and cytoskeletons in the tissue. This article generalizes from the present newest research advances, which is of important significance for revealing the mechanism of the therapeutic effect of acupuncture.

Objective To study the effects of Batroxobin( BX) on the intimal proliferation of graft veins. Methods In this study 25 dogs were selected and evenly divided into experimental group, control group and sham operation group. In experiment and control group, a segment of auto- femoral vein were grafted into femoral artery by clean microsurgery technique. In experimental group, BX was given at the dosage of 0. 1 BU/kg, dayly?2 preoperatively, and once a day for consecutive 6 days postoperatively. Plasma NO, ET was determined in the three groups. Computer image analysis system was applied to calculate the thickness of neointima and media in the vein grafts, immunohistochemistry was used to identify PCNA and C-myc. Result The experimental group had a higher level of NO and lower level of ET compared with control group and sham operation group(P 0. 05 ). The PCNA expression in experimental group was statistically different from that of the control group(P

Objective:To establish a method to determine the metabolites in rat kidney tissues by gas chromatography-mass spectrometry (GC-MS) combined with chemometric techniques. Methods:Metabolites were separated and identiifed on HP-5MS column (30 m × 0.25 μm × 0.25 mm). The initial column temperature was 100℃lasting 3 min, and then programmed at 8℃/min to 300℃, maintaining at this temperature for 6 min. hTe internal standard was heptadecanoic acid. hTe grinded kidney tissue was exacted by methanol. hTe supernatant was dried by nitrogen. Atfer the oximation and derivation, the supernatant was analyzed by GC-MS. hTe overlapped peaks were resolved into pure chromatogram and mass spectra with chemometric techniques. Qualitative analysis was performed by comparing the obtained pure mass spectra with those in NIST mass spectra database and certiifcated by the standards and the references. hTe internal method was used for semi-quantitation. Results:A total of 53 compounds were identiifed. hTe main constitutions in the kidney tissue were amino acids, saccharides, fatty acids and urea. Conclusion:hTe combination of methods is rapid and accurate for the analysis of metabolites in the kidney tissue, which provides more information for further study of metabonomics in kidney tissues.

BACKGROUND:Osteopontin, a kind of extracellular matrix glycoprotein, has been found to participate in synthesis and catabolism of osteoarthritic chondrocyte extracellular matrix. However, the effect of osteopontin on the proliferation ability of osteoarthritic chondrocytes is little reported. OBJECTIVE:To investigate the effect of osteopontin on the chondrocyte proliferation in human knee osteoarthritis. METHODS: Cartilage samples were obtained from the patients with knee osteoarthritis undergoing knee arthroplasty at the Xiangya Hospital from January 2012 to June 2012. The chondrocytes were isolated and cultured, and then divided into four groups: blank control, osteopontin, Con-shRNA and osteopontin-shRNA groups. The cell proliferation was detected by MTT and BrdU assays. RESULTS AND CONCLUSION: After transfection of osteopontin-shRNA lentivirus, the infection rate was up to 80%. Compared with the blank control group, osteopontin group showed a significant increase in the absorbance value of osteoarthritic chondrocytes, while after osteopontin-shRNA lentivirus transfection, the absorbance value was significantly decreased (bothP < 0.05). Additionally, after osteopontin-shRNA transfection, the expression level of osteopontin was significantly downregulated (P < 0.05). To conclude, osteopontin can promote the proliferation of osteoarthritic chondrocytes, which is considered as a new treatment target for osteoarthritis.

Objective To observe the cerebral protective effect of mild hypothermia by semiconductor cooling device on the liver surface in rabbits after cardiac arrest (CA). Methods Eighteen healthy male New Zealand white rabbits were randomly and equally divided into CA control group, ice saline group and semiconductor group. CA was induced by rapid intravenous injection of potassium chloride. Five minutes after onset of CA, CPR was initiated. Compared to the control group, which was not treated by hypothermia intervention after CPR, the ice saline group was treated by 4 ℃ ice saline infusion and the semiconductor group was treated by the semiconductor refrigeration piece device cooling on the liver surface for hypothermia intervention after CPR. We recorded the changes of temperature (tympanic temperature and anus temperature), heart rate (HR), mean arterial pressure (MAP) of rabbits in each group, neurological deficit scores (NDS) at 24, 48, 72 hours after the return of spontaneous circulation (ROSC) and the changes of serum neuron specific enolase (NSE) by enzyme linked immunosorbent assay (ELISA). Pathological changes of the hippocampus tissue, liver tissue and skin tissue were obtained by HE staining. Results There was no significant difference in ROSC time in each group. Two rabbits died at 55 hours and 67 hours after ROSC respectively in the control group. The remaining rabbits survived to 72 hours after challenge. There was no significant difference in the overall survival time in groups. Two hypothermia intervention groups had significantly lower level of serum NSE at 24 hours after ROSC and lower DNS scores at 24, 48, 72 hours after ROSC than control group. And the level of serum NSE after 24 hours of ROSC in the semiconductor group were significantly lower than the ice saline group (μg/L: 6.916±1.161 vs. 8.615±1.430, P < 0.05). DNS scores at 24, 48, 72 hours after ROSC in the semiconductor group were all significantly lower than the ice saline group (scores: 1.33±0.52 vs. 2.00±0.01, 1.01±0.41 vs. 2.00±0.01, 0.92±0.40 vs. 2.10±0.52 respectively, all P < 0.05). Two hypothermia intervention groups had more minor damage of neuronal cell in hippocampus than the control group. And the semiconductor group had more minor damage than the ice saline group. There were no obvious hepatic and subcutaneous tissue injury through which the semiconductor induced hypothermia was performed at corresponding liver surface skin. Conclusion The hypothermia by semiconductor cooling device on the liver surface is a new safe way of protecting brain tissue after CA, which has better cerebral protective effect than ice saline infusion.

Objective To investigate the value of individualized thyroid stimulating hormone (TSH) inhibition therapy in postoperative patients with differentiated thyroid carcinoma.Methods The medical record and follow-up data of the 556 patients with differentiated thyroid carcinoma after total or neartotal thyroidectomy were retrospectively reviewed.Patients were divided into two groups:Group A (304 cases) received TSH suppression therapy without risk assessment.Group B (252 cases) were given TSH suppression therapy in accord with risk assessment of both differentiated thyroid cancer recurrence risk stratification condition and the side effects of TSH suppression therapy risk stratification.Results The 3-year non-recurrence and (or) non-metastasis rate in group B was 99.2％ which was higher than 96.8％ in group A (P =0.044).The hospitalization rate caused by postoperative cardiovascular events or other morbidities in group B decreased 89％ than that in group A.Conclusions Individualized TSH suppression therapy can significantly decrease the recurrence and metastasis rate as well as concurrent morbidities caused by unnecessary TSH inhibition.

OBJECTIVETo conclude the possible interaction between gingko extract and simvastatin, by studing the effect of gingko extract on vitro metabolism of simvastatin and involving critical metabolic enzyme, which can guide the clinicians to use them rationally (propose good guideline for rational using of the two medicine mentioned above).

METHODThirty-two female SD rats were randomly separated into 4 groups, including negative control group. High dose of gingko extract. Low dose of gingko extract and positive control group. All groups were administered for 10 days with stomach tube, and then the quantity of liver microsome protein, the activity of CYP3A were determined by spectrophotograph simvastatin was incubated with the liver microsome, and the effect of gingko extract on its metabolism was estimated by measuring the amount of simvastatin by HPLC.

RESULTComparing with the negative contrast group: the quantity of liver microsome protein, the activity of CYP3A and the metabolism of simvastatin in positive control group were all increased markly (P < 0.05). In high dose group, the quantity of liver microsome protein and the activity of CYP3A were both increased significantly (P < 0.05), and the metabolism of simvastatin also accelerate obviously (P < 0.05). But in the low dose group significant distinction of every index was not found.

CONCLUSIONHigh dose of gingko extract can induce the activity of CYP3A, and promote the metabolism of simvastatin, so the medical interaction should be focused when gingko extract is coadministered with simvastatin and other substracts of CYP3A.

Animals ; Cytochrome P-450 CYP3A ; metabolism ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; administration & dosage ; Female ; Ginkgo biloba ; chemistry ; Microsomes, Liver ; drug effects ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Simvastatin ; metabolism

OBJECTIVETo identify a nuclear factor κB (NF-κB) responsive element within the Smad ubiquitination regulatory factor 1 (SMURF1) gene promoter, and to demonstrate its role in the regulation of SMURF1 expression.

METHODSA series of truncated luciferase reporter plasmids of the SMURF1 promoter were constructed and transfected into hepatic cancer Hep G2 cells. Luciferase assays were carried out to assess the activities of such promoters. DNA binding and chromatin immunoprecipitation (ChIP) assays were used to identify an NF-κB responsive element within the SMURF1 promoter. Lucifease plasmid with mutated NF-κB site was constructed and its activity was assessed. The expression of SMURF1 in Hep G2 cells was detected after transfection of NF-κB specific small interfering RNA (siRNA).

RESULTSThe SMURF1 promoter showed a high transcription activity, and the region of -519 to -378 was demonstrated to be a positive regulatory region. -411 to -420 of the SMURF1 promoter was an NF-κB responsive element, and NF-κB may specifically bind to this site. Mutation of this element may prominently decrease the activity of the promoter. Transfection of NF-κB siRNA evidently down-regulated SMURF1 expression.

CONCLUSIONNF-κB can specifically bind to the -411 to -420 region of the SMURF1 promoter and plays an essential role in the expression of SMURF1.

Gene Expression Regulation ; Hep G2 Cells ; Humans ; NF-kappa B ; genetics ; metabolism ; Promoter Regions, Genetic ; Protein Binding ; Transcription, Genetic ; Ubiquitin-Protein Ligases ; genetics ; metabolism

Objective:To investigate the risk factors of non-alcoholic fatty liver disease (NAFLD) in patients with hormone receptor positive (HR+), human epidermal growth factor receptor 2 negative (HER2-) breast cancer (HR+/HER2-BC) and the impact of NAFLD on the survival of patients.Methods:54 HR+BC patients were enrolled in this study. The liver fat accumulation was examined by magnetic resonance imaging (MRI). The patients were divided into two groups: non-NAFLD and NAFLD. Student's t test or Fisher's test was used to analyze the clinical indicators of the two groups. Logistic univariate and multivariate tests were used to analyze the clinical risk factors related to NAFLD. Receiver operating characteristic curve (ROC curve) was used to further analyze the sensitivity of clinical risk factors to predict the diagnosis of NAFLD. The Disease-free survival (DFS) and Overall survival (OS) of the two groups were analyzed by Log-rank (Mantel-Cox) test. Results:There were 22 NAFLD patients and 32 non-NAFLD patients diagnosed by MRI. Student's t test or Fisher's test showed that BMI, waist circumference, AST, ALT, GGT, TG, LDL and HDL were statistically different between the two groups (all P<0.05). Logistic univariate and multivariate analysis showed that AST ( OR=1.05, 95% CI: 1.02-1.10, P=0.007), GGT ( OR=1.04, 95% CI: 1.01-1.09, P=0.038), TG ( OR=1.03, 95% CI: 1.01-1.06, P=0.011) and HDL ( OR=1.06, 95% CI: 1.01-1.12, P=0.037) were the risk factors associated with NAFLD. ROC curve analysis showed that the combination of AST, GGT, TG and HDL had high sensitivity in predicting NAFLD (AUC=0.869, P<0.05). There was no difference in DFS ( HR=1.830, 95% CI: 0.983-3.409, P=0.057) or OS ( HR=2.482, 95% CI: 0.761-8.093, P=0.132) between the two groups. Conclusion:AST, GGT, TG and HDL are the independent risk factors for NAFLD in HR+BC patients during treatment, but concurrent NAFLD has no significant effect on DFS or OS.