Mukwevho E, Kohn TA, Lang D, Nyatia E, Smith J, Ojuka EO. Caffeine induces hyperacetylation of histones at the MEF2 site on the Glut4 promoter and increases MEF2A binding to the site via a CaMK-dependent mechanism. Am J Physiol Endocrinol Metab 294: E582-E588, 2008. First published January 15, 2008 doi:10.1152/ajpendo.00312.2007.-This study was conducted to explore the mechanism by which caffeine increases GLUT4 expression in C2C12 myotubes. Myoblasts were differentiated in DMEM containing 2% horse serum for 13 days and the resultant myotubes exposed to 10 mM caffeine in the presence or absence of 25 M KN93 or 10 mM dantrolene for 2 h. After the treatment, cells were kept in serum-free medium and harvested between 0 and 6 h later, depending on the assay. Chromatin immunoprecipitation (ChIP) assays revealed that caffeine treatment caused hyperacetylation of histone H3 at the myocyte enhancer factor 2 (MEF2) site on the Glut4 promoter (P Ͻ 0.05) and increased the amount of MEF2A that was bound to this site ϳ2.2-fold (P Ͻ 0.05) 4 h posttreatment compared with controls. These increases were accompanied by an ϳ1.8-fold rise (P Ͻ 0.05 vs. control) in GLUT4 mRNA content at 6 h post-caffeine treatment. Both immunoblot and immunocytochemical analyses showed reduced nuclear content of histone deacetylase-5 in caffeinetreated myotubes compared with controls at 0 -2 h posttreatment. Inclusion of 10 mM dantrolene in the medium to prevent the increase in cytosolic Ca 2ϩ , or 25 M KN93 to inhibit Ca 2ϩ /calmodulindependent protein kinase (CaMK II), attenuated all the above caffeine-induced changes. These data indicate that caffeine increases GLUT4 expression by acetylating the MEF2 site to increase MEF2A binding via a mechanism that involves CaMK II. myocyte enhancer factor 2; glucose transporter 4; histone deacetylase; chromatin immunoprecipitation assay; Ca 2ϩ /calmodulin-dependent protein kinase II; histone H3 REGULAR EXERCISE INCREASES THE CONTENT of the GLUT4 glucose transporter that increases glucose disposal and protects against type II diabetes (8, 28). There is mounting evidence that the increase in cytosolic Ca 2ϩ

Activation of calmodulin dependent protein kinase (CaMK)II by exercise is beneficial in controlling membrane lipids associated with type 2 diabetes and obesity. Regulation of lipid metabolism is crucial in the improvement of type 2 diabetes and obesity associated symptoms. The role of CaMKII in membrane associated lipid metabolism was the focus of this study. Five to six weeks old male Wistar rats were used in this study. GC×GC-TOFMS technique was used to determine the levels of polyunsaturated fatty acids (linoleic acid, arachidonic acid and 11,14-eicosadienoic acid). Carnitine palmitoyltransferase (Cpt-1) and acetyl-CoA carboxylase (Acc-1) genes expression were assessed using quantitative real time PCR (qPCR). From the results, CaMKII activation by exercise increased the levels of arachidonic acid and 11,14-eicosadienoic acid while a decrease in the level of linolenic acid was observed in the skeletal muscle. The results indicated that exercise-induced CaMKII activation increased CPT-1 expression and decreased ACC-1 expression in rat skeletal muscle. All the observed increases with activation of CaMKII by exercise were aborted when KN93, an inhibitor of CaMKII was injected in exercising rats. This study demonstrated that CaMKII activation by exercise regulated lipid metabolism. This study suggests that CaMKII can be a vital target of therapeutic approach in the management of diseases such as type 2 diabetes and obesity that have increased to epidemic proportions recently.

No abstract

There is an increased prevalence of nonalcoholic steatohepatitis ( NASH ) in adolescents. The suckling period is developmentally plastic, affecting later health outcomes. We investigated whether neonatal administration of curcumin would provide protection against the development of NASH later in adolescence in rats fed a high‐fructose diet. From postnatal day ( PN ) 6 to PN 21, the pups ( N = 128) were allocated to four groups and orally gavaged daily with either 0.5% dimethyl sulfoxide solution (vehicle control), curcumin (500 mg·kg −1 ), fructose (20%, w/v) or curcumin and fructose combined. All the pups were weaned and half the rats in each group had tap water, whereas the other received fructose (20%) as their drinking fluid ad libitum for 6 weeks. The rats’ liver NASH scores, lipid content, and RNA gene expression ratios of AMPK α and TNF α were determined. Hepatic lipid content was similar across the treatment groups in the males ( P > 0.05, ANOVA ). In the females, the hepatic lipid content in the treatment groups ranged from 2.7 to 4.3%. The livers of male and female rats that had fructose either as neonates and/or postweaning had significantly marked inflammation ( P = 0.0112, Kruskal–Wallis) and fibrosis ( P < 0.0001, ANOVA ) which were attenuated by curcumin. The hepatic gene expression ratios for AMPK α in both sexes were significantly downregulated ( P < 0.0001, ANOVA ), whereas the expression ratios of TNF α were significantly upregulated ( P < 0.0001) in rats fed a high‐fructose diet pre and/or postweaning compared to the other groups. Neonatal curcumin administration is a potential natural pharmacological candidate for the prevention of NASH .

Obesity is the accumulation of excess body fat and the hallmark of type II diabetes mellitus, characterized by hyperglycemia. Glycemic control is very critical to reduce long-term vascular complications resulting from the progressive nature of hyperglycemia. In previous studies, thermally reduced graphene oxide (rGO)-based hydrogel biocomposites were prepared and in vitro drug release studies confirmed their potential as a biodegradable-targeted drug delivery system. Thus, the in vitro biological evaluation of these rGO-based hydrogels was investigated. The hydrogels were encapsulated with chloroquine diphosphate (CQ) and proguanil (P) drugs to investigate potential of combination therapy. The non-toxic nature of the hydrogels was investigated by the use of the MTT assay against 3T3-L1 and c2c12 cell lines. 3T3-L1 pre-adipocytes were grown, differentiated and treated with the drug-encapsulated hydrogels to detect the effect on the adipose tissue cells by quantitative real-time polymerase chain reaction (qPCR) identifying gene expression levels by utilizing gene markers specific for diabetes and obesity: cpt-1, glut-4, acc1, pgc-1, mef2a and nrf-1 with comparison to positive control metformin. The cytoxicity studies confirmed non-toxic nature of the hydrogels; identified dosage of drugs encapsulated were effective within investigated treatment time and qPCR revealed an upregulation of CPT-1, GLUT-4, PGC-1, MEF2A and NRF-1 marker genes, but a downregulation of ACC-1 marker gene. The results from the expression of investigated genes suggest the anti-obesity potential of drugs released from the hydrogels. There were identified positive effects employing combination therapy, but further studies are required to ascertain the actual effect of the drugs in combination, by further varying the ratios of drugs (instead of the presented 1:1 ratio) employed. Statistically, the results from the individual drug release treatments were not significantly different from positive control metformin treatments, but the combination therapy investigation showed more promise.