Pooja Narain Adtani1, Malathi Narasimhan1, Alan M Punnoose2, Himavanth Reddy Kambalachenu3. 1. Department of Oral Pathology and Microbiology, Faculty of Dental Sciences, Sri Ramachandra University and Research Institute, Chennai, India. 2. Center for Regenerative Medicine and Stem Cell Research, Central Research Facility, Sri Ramachandra University and Research Institute, Chennai, India. 3. Department of Biomedical Sciences, Sri Ramachandra University and Research Institute, Chennai, India.
Abstract
AIM: The aim of the present study was to investigate the in vitro antifibrogenic effects of Centella asiatica Linn (CA) and its bioactive triterpene aglycone asiatic acid (AA) on arecoline-induced fibrosis in primary human buccal fibroblasts (HBF). METHODS: An ethanolic extract of CA was prepared, and AA was purchased commercially. High-performance thin-layer chromatography (HPTLC) was performed to quantify AA in the CA extract; colorimetric assay (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) was performed to determine an half-maximal inhibitory concentration. HBF were cultured and stimulated with arecoline. The inhibitory effects of CA and AA at different concentrations were assessed using gene-expression studies on fibrosis-related markers: transforming growth factor-β1, collagen 1 type 2, and collagen 3 type 1. The stimulatory effect of arecoline and the inhibitory effect of AA on fibroblast morphology and extracellular matrix were assessed qualitatively using Masson trichrome stain. RESULTS: The HPTLC analysis determined 1.2% AA per 100 g of CA extract. Arecoline produced a concentration-dependent increase in the fibrotic markers, treatment with CA significantly downregulated fibrotic markers at higher concentrations, and AA downregulated at lower concentrations. Arecoline altered fibroblast morphology and stained strongly positive for collagen, and AA treatment regained fibroblast morphology with faint collagen staining. CONCLUSION: CA and AA can be used as antifibrotic agents.
AIM: The aim of the present study was to investigate the in vitro antifibrogenic effects of Centella asiatica Linn (CA) and its bioactive triterpeneaglyconeasiatic acid (AA) on arecoline-induced fibrosis in primary human buccal fibroblasts (HBF). METHODS: An ethanolic extract of CA was prepared, and AA was purchased commercially. High-performance thin-layer chromatography (HPTLC) was performed to quantify AA in the CA extract; colorimetric assay (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) was performed to determine an half-maximal inhibitory concentration. HBF were cultured and stimulated with arecoline. The inhibitory effects of CA and AA at different concentrations were assessed using gene-expression studies on fibrosis-related markers: transforming growth factor-β1, collagen 1 type 2, and collagen 3 type 1. The stimulatory effect of arecoline and the inhibitory effect of AA on fibroblast morphology and extracellular matrix were assessed qualitatively using Masson trichrome stain. RESULTS: The HPTLC analysis determined 1.2% AA per 100 g of CA extract. Arecoline produced a concentration-dependent increase in the fibrotic markers, treatment with CA significantly downregulated fibrotic markers at higher concentrations, and AA downregulated at lower concentrations. Arecoline altered fibroblast morphology and stained strongly positive for collagen, and AA treatment regained fibroblast morphology with faint collagen staining. CONCLUSION: CA and AA can be used as antifibrotic agents.