Objective To investigate the effect of rh-resistin on glucose metabolism in HepG2 cells and to elucidate whether the underlying mechanisms are related to AMPK pathway. Methods Cells transfected with control siRNA or AMPKα2 siRNA were cultured in 6-well plates and then treated with 50 ng/mL rh-resistin for 24 hours , while untransfected cells were treated with or without 50 ng/mL rh-resistin on the same conditions , followed by serum-starving in glucose-free DMEM for 3 ~ 5 hours in the continued absence or presence of rh-re-sistin. Then the cells were treated with or without insulin for 2 hours. AMPKα2, G6Pase, PEPCK and Glut2 mRNA expression levels were determined by quantitative RT-PCR. The phosphorylation state of AMPK was deter-mined by Western blotting. Glycogen synthesis was measured by the incorporation of D-[U-14C] glucose to glycogen. Results Rh-resistin suppressed the AMPKα2, Glut2 mRNA expressions, and reduced the phosphory-lation level of AMPK and glycogen synthesis on both basal and insulin-stimulated conditions (P < 0.05), while it accelerated G6Pase and PEPCK mRNA expressions on the same conditions (P < 0.05). The mRNA expression levels of G6Pase , PEPCK , Glut2 and the phosphorylation level of AMPK and glycogen synthesis were signifi-cantly different between the rh-resistin group and the rh-resistin in conjunction with AMPKα2 siRNA-treated group. Conclusion Rh-resistin may affect glucose metabolism in HepG2 cell via AMPK pathway.

Objective To investigate the effect of rh-resistin on lipid metabolism in HepG2 cells and to elucidate its relation to AMPK pathway. Methods We treated the HepG2 cells with 50 ng/ml rh-resistin and 0.5 mmol/L palmitic acid,used siRNA technique to inhibite α2 subunite expression of AMPK in HepG2 cells and quantitative RT-PCR to detect ACC1,ACC2,and HL mRNA expression levels of related lipid metabolism genes. The P-AMPK-Thr172 of AMPK and P-ACC-Ser79 of ACC were determined by Western blotting. The Lipid accumu-lation in cells was determined by images of Laser Scanning Confocal Microscope after Nile red staining. Results Rh-resistin decreased the AMPKα2,HL mRNA expressions and the phosphorylation level of AMPK and ACC in both basal and insulin-stimulated conditions (P < 0.05),had no influence on ACC2 mRNA expressions (P >0.05),while it increased ACC2 mRNA expressions and cytoplasmic lipid droplets in the same conditions (P <0.05). Conclusion Rh-resistin may affect lipid metabolism via AMPK pathway with increase of fatty acid synthe-sis and inhibition of triglyceride catabolism,which leading to lipid accumulation in HepG2 cells.

BACKGROUND: The experimental results showed that insulin sensitivity and glucose uptake in skeletal muscle could be improved by activating adenosine monophosphate-activated protein kinase a2 (AMPKα2). AMPKa2 is expected to become a new physiological and pharmacological target for the prevention and treatment of type 2 diabetes mellitus. OBJECTIVE: To clone human AMPKa2 subunit gene and to construct its wild-type and mutant eukaryotic expression vectors. DESIGN: A single sample observation.TIME AND SETTING: The experiment was performed in the Clinical Molecular Biology Laboratory, the Second Affiliated Hospital of Sun Yet-sen University from April 2007 to January 2008.MATERIALS: QuikChange II Site-Directed Mutagenesis Kit was produced by Stratagene. Eukaryotic expression vector pcDNA3.1(+) and E. coll DH5a were provided by the laboratory. Human skeletal muscle tissue was from patients who received amputation surgery in the Second Affiliated Hospital of Sun Yat-sen University. Informed consent was obtained from the patients, and fresh samples were collected and frozen in liquid nitrogen.METHODS: The human AMPKo2 subunit gone was amplified from human skeletal muscle by RT-PCR, cloned into T vector, and the recombinant plasmid was confirmed by sequencing. In vitro site-directed mutagenesis was carded out with Quickchange site-directed mutagenesis kit. The wild-type and mutant coding genes were subcloned into eukaryotic expression vector pcDNA3.1, and the recombinant plasmids were validated by enzyme digestion and sequencing.MAIN OUTCOME MEASURES: ①The cloning of aim gone; ②site-directed mutagenesis; ③ eukaryotic expression plasmid. RESULTS: The human AMPKα2 subunit gene (about 1700 bp) was successfully cloned, with 99％ homology to the reported AMPK α2 gene. A GenBank accession number was EF056019, The achieved mutation of the 45<'th> Lysine (AAA) was found to Arginine(AGA). The wild-type and mutant pcDNA-AMPKα2 recombinant plasmids were constructed successfully. CONCLUSIONS: The human AMPKα2 subunit gene was cloned successfully from human skeletal muscle and its wild-type and mutant eukeryotic expression vectors were constructed successfully in the experiment.

Objective: To observe the clinical efficacy of combined electroacupuncture and nerve growth factor (NGF) injection at acupoints in the treatment of common fibular nerve paralysis and provide evidences for integrative Chinese & western medicine against diabetic peripheral neuropathy (DPN). Methods: Forty subjects were randomized into two groups and NGF injection; and control group was given herbal suffocation, oral Dibazol and compound vitamin B and Mecobalamin Injection. The clinical symptoms and nerve conduction velocity were observed and compared. Results: The cure rate was higher in treatment group than in control group (P<0.05); after treatment, the nerve conduction velocity was improved in both groups (P<0.01), with a significant improvement in treatment group than in control group (P<0.01). Conclusion: Combined electro-acupuncture and NGF injection at acupoints is quite effective in the treatment of common fibular nerve paralysis.

Objective To explore the effects of donor dendritic cells(DC)treated with Ad-IL- 12p35siRNA on the survival of allogragft recipients.Methods The recombinant adenoviral vectors carrying IL-12p35siRNA and HKsiRNA were transfected into bone marrow derived DC of BALB/C murine.C57BL/6 recipients were infused with DG(Ad-IL-12p35siRNA DC,Ad-HKsiRNA DC and control DC)from BALB/C donors 7 days before cardiac allograft,the survival time of murine and the change of T_H 1 and T_H2(IL-2,IL-4,IL-10 and IFN-?)cytokine were observed.Results The survival time of p35 group(20.17?2.71)days was longer than that of control group(7.81?1.61)days and HK group(7.17?1.60)days.The concentration of IL-2 and IFN-?in p35 group were significantly lower than those of control group and HK group,otherwise were the concentration of IL-4 and IL-10. Conclusion Pretreatment of dondor dendritic ceils with Ad-IL-12p35siRNA could prolonged cardiac allograft survival in recipicents.

Objective To explore the levels of serum leptin and insulin and their relationships with growth and development in premature infants.Methods The serum leptin and insulin concentrations of 37 premature infants were examined by the radioimmunoassay(day 1 and day 10,respectively),and anthropometric measurements were performed at birth.The blood sugar and triglyeride,cholesterol were tested at the same time.Twenty full-term newborn without severe diseases were controlled in 24 h after delivery and the level of leptin and insulin of them were detected,too.The results were analyzed by SPSS 10.0 software.Results The leptin and insulin levels in preterm infants were significantly lower than those in full-term infants[(6.45?2.24)?g/L vs(8.56?1.33)?g/L,(5.84?1.07)IU/L vs(10.85?5.24)IU/L Pa0.05).Serum leptin was positive correlation with serum insulin(Pa0.05).Conclusions Serum leptin and insulin can be the markers that reflect intrauterine nutrition status of preterm infants and the growth and development of fetus.A dipoinsular axis may be functional before 34 weeks gestation without maturity.The changes of leptin and insulin concentrations are useful for metabolism and growth in preterm infants.

This study is to establish the HPLC fingerprint and determine eight components of Callicarpa nudiflora, and provide a scientific basis for the identification and quality control. The Waters sunfire C18 column (4.6 mm x 250 mm, 5 µm) was used and the detection wavelength was 330 nm . The column temperature was 30 °C. The mobile phases were acetonitrile (A) and 0.1% formic acid (B) eluting in a gradient program at a flow rate of 1.0 mL · min(-1). The chromatographic fingerprint similarity evaluation system for tradition Chinese medicine(2012) was used for analysis. C. nudiflora from different samples were of high similarity in fingerprint and the separation of ten components was good. There was an obvious difference between other samples and C. nudiflora leaves. In quantitative analysis, the ten components showed good regression(R2 > 0 999 0) with linear ranges, and their recoveries were in the range of 96.0%-105.0%. The established qualitative and quantitative methods are highly specific, simple and accurate, which can be used for the identification and quality control of C. nudiflora.
Callicarpa ; chemistry ; Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; analysis ; isolation & purification ; Plants, Medicinal ; chemistry

To investigate the metabolic rate and metabolites of 9-dehydro-17-dehydro-andrographolide, which is the main active ingredient in Xiyanping injection, by using the in vitro rat liver microsome incubation system. 9-dehydro-17-dehydro-andrographolide was incubated together with liver microsome mixed with NADPH. Its metabolic rate was studied by determining its residual concentrations with the UHPLC-MS/MS method; Its metabolites were identified by the UPLC-TOF-MS(E) method. The results showed that 9-dehydro-17-dehydro-andrographolide was metabolized faster than rat liver microsomes mixed with coenzymes, with t½ and CL of (19.7 ± 0.5) min and (35.1 ± 0.8) mL x min(-1) x g(-1) (protein), respectively. Based on the high resolution mass spectrum data and information from literatures, altogether nine metabolites of 9-dehydro-17-dehydro-andrographolide were identified in the incubation system, particularly hydroxylated and dehydrogenized products. The results of identification would provide a basis for screening out more active andrographolide derivatives.
Animals ; Chromatography, High Pressure Liquid ; Diterpenes ; chemistry ; metabolism ; Drugs, Chinese Herbal ; chemistry ; metabolism ; Microsomes, Liver ; chemistry ; metabolism ; Molecular Structure ; Rats ; Tandem Mass Spectrometry

Objective To investigate the clinical efficacy of acupuncture at Huatuo jiaji points plus point Shixuan bloodletting in treating cervical spondylotic radiculopathy.Methods One hundred patients with cervical spondylotic radiculopathy were randomly allocated to treatment and control groups, 50 cases each. The treatment group received acupuncture at Huatuo jiaji points plus point Shixuan bloodletting and the control group, acupuncture at Huatuo jiaji points alone. Plasma viscosity was measured in the two groups before and after treatment. The clinical therapeutic effects were compared between the two groups.Results There was a statistically significant pre-/post-treatment difference in plasma viscosity in the two groups (P<0.05). There was a statistically significant post-treatment difference in plasma viscosity between the treatment and control groups (P<0.05). The total efficacy rate was 94.0% in the treatment group and 84.0% in the control group; there was a statistically significant difference between the two groups (P<0.05).Conclusion Acupuncture at Huatuo jiaji points plus point Shixuan bloodletting is an effective way to treat cervical spondylotic radiculopathy.

AIM: To investigate the expression of survivin in pancreas in the streptozotocin-induced diabetic mice. METHODS: Low dose of streptozotocin was used to induce diabetes mellitus in BALB/c mice. Body weight and blood glucose concentrations were examined at 1, 2, 3 and 4 weeks after the streptozotocin injection. Expression of survivin mRNA was detected by real-time FQ-PCR. RESULTS: Survivin was expressed in the pancreas of normal BALB/c mice. Low dose of streptozotocin provoked hyperglycaemia with increased survivin expression in the pancreas, but blood glucose concentration and expression of survivin was not significantly changed in control group. CONCLUSION: Survivin is expressed in the pancreas of normal BALB/c mice. Streptozotocin increases survivin expression in the pancreas, which may be related with islets regeneration.