Umed V. Ramani

Anti-obesity potential of Clerodendron glandulosum.Coleb leaf aqueous extract.

ETHNOPHARMACOLOGICAL RELEVANCE Clerodendron glandulosum.Coleb leaf aqueous extract (CG) is traditionally used by people of North-East India to alleviate symptoms of diabetes, obesity and hypertension. Previous study from our laboratory have documented anti-diabetic and anti-hypertensive properties of CG extract but, till date there are no pharmacological studies available on its anti-obesity potential. This inventory investigates the underlining molecular mechanism/s of CG induced regulation of in vivo HFD induced obesity and in vitro adipocyte differentiation. AIM To evaluate effects of CG extract on (i) expression of genes regulating visceral adiposity and (ii) in vitro adipocyte differentiation and LEP release. MATERIALS AND METHODS Body weight, lee index, plasma lipids and LEP, mRNA expression of PPARγ-2, SREBP1c, FAS, CPT-1 and LEP in epididymal adipose tissue of control and experimental groups were evaluated. Also, potential of CG extract on in vitro adipocyte differentiation and LEP release was assessed. RESULTS Supplementation of CG extract to HFD fed mice significantly prevented HFD induced increment in bodyweight, lee index, plasma lipids and LEP, visceral adiposity and adipocyte hypertrophy. Also, CG extract supplementation resulted in down regulation of PPARγ-2, SREBP1c, FAS and LEP expression along with up-regulation of CPT-1 in epididymal adipose tissue compared to HFD fed mice. In vitro study recorded significant anti-adipogenic effect of CG extract that resulted in decreased adipogenesis, TG accumulation, LEP release, G3PDH activity along with higher glycerol release without significantly altering viability of 3T3L1 pre-adipocytes. CONCLUSIONS Clerodendron glandulosum.Coleb extract prevents adipocyte differentiation and visceral adiposity by down regulation of PPARγ-2 related genes and Lep expression thus validating its traditional therapeutic use in controlling obesity.

Myostatin gene silencing by RNA interference in chicken embryo fibroblast cells.

Myostatin (MSTN), a member of transforming growth factor-β (TGF-β) superfamily, is a negative regulator of the skeletal muscle growth, and suppresses the proliferation and differentiation of myoblast cells. Dysfunction of MSTN gene either by natural mutation or genetic manipulation (knockout or knockdown) has been reported to interrupt its proper function and to increase the muscle mass in many mammalian species. RNA interference (RNAi) mediated by small interfering RNAs (siRNAs) or short hairpin RNAs (shRNAs) has become a powerful tool for gene knockdown studies. In the present study transient silencing of MSTN gene in chicken embryo fibroblast cells was evaluated using five different shRNA expression constructs. We report here up to 68% silencing of myostatin mRNA using these shRNA constructs in transiently transfected fibroblasts (p<0.05). This was, however, associated with induction of interferon responsive genes (OAS1, IFN-β) (3.7-64 folds; p<0.05). Further work on stable expression of antimyostatin shRNA with minimum interferon induction will be of immense value to increase the muscle mass in the transgenic animals.

Sida rhomboidea. Roxb Leaf Extract Down-Regulates Expression of PPARγ2 and Leptin Genes in High Fat Diet Fed C57BL/6J Mice and Retards in Vitro 3T3L1 Pre-Adipocyte Differentiation

Sida rhomboidea. Roxb leaf extract (SRLE) is being used by the populace of North-East India to alleviate symptoms of diabetes and obesity. We have previously reported its hypolipidemic and anti-diabetic properties. In this study, we report the effect of SRLE on (i) in vivo modulation of genes controlling high fat diet (HFD) induced obesity and (ii) in vitro 3T3L1 pre-adipocyte differentiation and leptin release. Supplementation with SRLE significantly prevented HFD induced increment in bodyweight, plasma lipids and leptin, visceral adiposity and adipocyte hypertrophy. Also, SRLE supplementation reduced food intake, down regulated PPARγ2, SREBP1c, FAS and LEP expressions and up-regulated CPT-1 in epididymal adipose tissue compared to obese mice. In vitro adipogenesis of 3T3L1 pre-adipocytes was significantly retarded in the presence of SRLE extract. Also decreased triglyceride accumulation, leptin release and glyceraldehyde-3-Phosphate dehydrogenase activity along with higher glycerol release without significant alteration of viability of 3T3L1 pre-adipocytes, was recorded. Our findings suggest that prevention of HFD induced visceral adiposity is primarily by down regulation of PPARγ2 and leptin gene expression coupled with attenuation of food intake in C57BL/6J mice. SRLE induced prevention of pre-adipocytes differentiation, and leptin release further substantiated these findings and scientifically validates the potential application of SRLE as a therapeutic agent against obesity.

A modified enrichment protocol for adult caprine skeletal muscle stem cell

To establish an adequate model to study the proliferation and differentiation of adult caprine skeletal muscle in response to bioactive compounds, a pool of satellite cells (SC) was derived from the rectus abdominis muscle of adult goat. Skeletal muscle contains a population of adult stem cells, named as satellite cells that reside beneath the basal lamina of skeletal muscle fiber and other populations of cells. These SC are multipotent stem cells, since cells cultured in the presence of specific cell lineage inducing cocktails can differentiate into several types of mesenchymal lineage, such as osteocytes and adipocytes. In the present study, we have developed a modified protocol for isolating satellite cells (>90%) and examined their myogenic and contractile properties in vitro.

A preliminary sketch of horn cancer transcriptome in Indian zebu cattle

Horn cancer, a type of squamous cell carcinoma, in zebu cattle is an expensive affair in Indian agriculture sector, which accounts for 83.34% of total tumors found. In general, cancer tissue confirms considerably different expression patterns when compared to a normal stage. This includes not only up/down regulation, but also, the aberrant gene expression, the presence of different non-coding RNAs (ncRNAs), pseudogenes expression and genes involved in unusual pathways. We employed Roche 454 next generation sequencing platform to sequence Bos indicus cancerous and normal horn tissue transcripts. This resulted into a total of 909,345 high-confidence deep sequencing reads and detected a range of unusual transcriptional events including tumor associated genes. We also validated expression of two of the four tested genes in five other similar tissue samples by RT-qPCR. Further, seven cancer specific non-coding transcripts were accessed and a few of them have been suggested as cancer specific markers. This study for the first time provides primary transcriptome sketch of Bos indicus horn cancer tissue, and also demonstrates the suitability of the 454 sequencer for transcriptome analysis, which supports the concept of varied gene expression in cancerous condition.