Dangua Fang(丹瓜方)regulating tricarboxylic acid cycle and respiratory chain and its mechanism in diabetic rats

OBJECTIVE:To investigate the influence and possible targets of Dangua Fang(丹瓜方)on tricarboxylic acid(TCA)cycle and respiratory chain to enrich the prescription’s mechanism of effective intervention on glycolipid metabolic diseases such as type 2 diabetes.METHODS:After interventional rats were fed with high glucose and high fat diet ad libitum for 4 weeks,intraperitoneally injected streptozotocin to induce diabetic model.According to blood glucose level,28 diabetic rats were selected and continued to be fed with high glucose and high fat diet,were stratified by body weight,and divided randomly by blood glucose into Model group(was given sterile water by gastric perfusion and injected aquae pro injection intraperitoneally),Dangua group[Dangua liquor(丹瓜方液)20.5 g·kg^(-1)·d^(-1) by perfusion and aquae pro injection intraperitoneally],Inhibitor group[sterile water by perfusion and nicotinamide phosphoribosyl transferase(Nampt)specific blocker GEN-6171.25 mg/kg intraperitoneally],DanInhit group(Dangua liquor and GEN-617 synchronously).Control group were continuously fed with ordinary diet.The intervention was last for 10 weeks.Body weight(BW),liver index(LI),glycosylated hemoglobin(HbA1c),TC,TG,free fatty acids(FFA),creatinine(Cr),and A-ketoglutarate(α-KG),Iso-citric acid(ICA),oxaloacetic acid(OAA)were tested.The cytochrome C oxidase(COX)and Succinate dehydrogenase(SDH)were evaluated by Colorimetry;Nampt protein,Adenosine triphosphate(ATP)synthase(ATPs),Nicotinamide adenine dinucleotide(NAD^(+))and its reduced(NADH)in liver were measured by enzyme linked immunosorbent assay.The expressions of Nampt and mitochondrialnadhdehydrogenase-1(mt-ND1)gene in liver was assessed by real-time polymerase chain reaction.Hepatic tissue staining was also completed.RESULTS:The levels of BW,ICA,α-KG and Nampt-mRNA in the Model group are lower than that in the Normal group(P<0.05),conversely,liver weight,LI,TC,HbA1c,SDH and ATPs,mt-ND1-mRNA,and Nampt protein in the Model group are higher(P<0.01,P<0.05).Compared with Model group,the levels of ICA,Nampt-mRNA and Nampt in Dangua group are significantly increased,and FFA obviously raised(P<0.01 and P<0.05);liver weight,BW,SDH are obviously lower,and HbA1c decreased significantly(P<0.01,P<0.05).TG,FFA and Nampt protein increased in the DanInhit group,TC,TG,BW obviously increased in the Inhibitor group,but SDH is decreased in both the two groups(P<0.05,P<0.01).Compared with Dangua group,DanInhib group has the lower levels of ICA,mt-ND1-mRNA,Nampt-mRNA,and the higher level of BW,LI and HbA1c.In the Inhibitor group,ICA and Nampt protein decreased,BW and LI,HbA1c and TG increased(P<0.01 or P<0.05).Tissue staining display that,in the model group there is obvious pathologic changes ie:fibrosis,steatosis and inflammatory cell infiltration.Lesions in the Dangua group are mild,and those of Inhibitor group are more obvious than the Model group,and DanInhit group is intermediately affected compared to Dangua group and Inhibitor group.CONCLUSION:Dangua Fang increases the metabolic flux of TCA cycle and optimizes respiratory chain function by up-regulating Nampt expression.

Effect of burn injury on relative anaplerosis and gluconeogenesis in rats by ^(13)C magnetic resonance spectrum

To introduce a safe and specific appr oach of 13 C magnetic resonance spectrum ( 13 C MRS) spectroscopy and investigate the alterations in hepatic anabolism. Methods: Relative anaplerotic, pyruvate recycling and gluconeog enic fluxes were measured by 13 C MRS isotopomer ana lysis of blood glucose from rats with 40% body surface area burn injury, and fro m rat s exposed to sham injury. A short chain fatty acid, [U 13 C] propionate which was avidly extracted by the liver, was infused intravenously to deliver 13 C into the citric acid cycle. Proton decoupled 13 C MRS of deproteinized plasma or extracts of the freeze clamped liver were used to determine the distribution of 13 C in blood or hepatic glucose.Results: There was no difference in the multiplets detected in the glucose carbon 2 anomer from blood or liver after 45 or 60 minutes of the i nfusion of the propionate, indicating that steady state isotopic conditions wer e achiev ed. Gluconeogenesis relative to citric acid cycle flux was not altered by burn i njury; in both sham and burn groups the rate of glucose production was about equ al to flux through citrate synthase. In the sham group of animals, the rate of e ntry of carbon skeletons into the citric acid cycle was about 4 times than that in the burn group. Similarly, flux through pyruvate kinase (again relative to ci trate synthase) was significantly increased after the burn injury. Conclusions: Since results from analysis of the blood glucose a re the same as that of the hepatic glucose, 13 C dis tribution in the glucose and hepatic metabolism can be assessed based on the 13 C MRS analysis of the blood glucose.