Cassia Abbreviata Enhances Glucose Uptake and Glucose Transporter 4 Translocation in C2C12 Mouse Skeletal Muscle Cells.

Abstract

Background: This study aim at assessing C. abbreviata aqueous extracts for its potential to exhibit anti-diabetic activity in skeletal muscle cells. In addition to the toxicological and glucose absorption studies, the action of C. abbreviata extracts on some major genes involved in the insulin signaling pathway was established.

Methods: The in vitro cytotoxic effects C. abbreviata was evaluated on muscle cells using the MTT assay and the in vitro glucose uptake assay conducted using a modified glucose oxidase method described by Van de Venter et al. (2008). The amount of GLUT-4 on cell surfaces was estimated quantitatively using the flow cytometry technique. Real time quantitative PCR (RT-qPCR) was used to determine the expression of GLUT-4, IRS-1, PI3 K, Akt1, Akt2, PPAR-γ.

Results: Cytotoxicity tests revealed that all extracts tested at various concentrations were non-toxic (LC₅₀ > 5000). Aqueous extracts of leaves, bark and seeds resulted in a dose-dependent increase in glucose absorption by cells, after 1 h, 3 h and 6 h incubation period. Extracts of all three plant parts had the best effect after 3 h incubation, with the leaf extract showing the best activity across time (Glucose uptake of 29%, 56% and 42% higher than untreated control cells after treatment with 1 mg/ml extract at 1 h, 3 h and 6 h, respectively). All extracts, with the exception 500 µg/ml seed extract, induced a two-fold increase in GLUT-4 translocation while marginally inducing GLUT-10 translocation in the muscle cells. The indirect immunofluorescence confirmed that GLUT-4 translocation indeed occurred. There was an increased expression of GLUT-4, IRS1 and PI3 K in cells treated with insulin and bark extract as determined by the RT-qPCR.

Conclusion: The study reveals that glucose uptake involves GLUT-4 translocation through a mechanism that is likely to involve the upstream effectors of the PI3-K/Akt pathway.