Tinospora crispa extract inhibits MMP-13 and migration of head and neck squamous cell carcinoma cell lines

Objective: To investigate the effect of Tinospora crispa (T. crispa) extract on matrix metalloproteinase-13 (MMP-13) expression and cell migration. Methods: The cytotoxicity of T. crispa extract was examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay on head and neck squamous cell carcinoma (HNSCC) cell lines. The effect on expression of MMP-13 was analysed by RT-PCR and ELISA. The migration was assessed by wound healing assay. Results: MMP-13 mRNA was highly expressed in the metastatic human HNSCC cell lines, HN22 and HSC-3. T. crispa extract at a concentration of 100.0 μg/mL caused about 50% reduction of cell survival. T. crispa extract at a non-toxic concentration of 12.5, 25.0 and 50.0 μg/mL significantly suppressed MMP-13 mRNA expression and secreted MMP-13 in both HN22 and HSC-3. The expression of tissue inhibitors of metalloproteinase-2 (TIMP-2) by HSC-3 cells was attenuated by 25.0 and 50.0 μg/mL of T. crispa extract. Addition of the extract to cells in a wound healing assay showed inhibition of cell migration by HN22 cells. Conclusions: These data suggest that T. crispa could be considered as a potential therapeutic drug to prevent metastasis of HNSCC

Defect in serine 46 phosphorylation of p53 contributes to acquisition of p53 resistance in oral squamous cell carcinoma cells

To investigate whether dysregulation of p53 phosphorylation confers tumor resistance to p53, we analysed the effects of wild-type p53 on oral squamous cell carcinoma(SCC) cell lines carrying various mutations of p53. Introduction of exogenous p53 neither induced apoptosis nor suppressed colony formation in HSC-3 cells lacking any detectable p53 and HSC-4 cells expressing mutant p53R248Q protein. Consistently, exogenous p53 did not induce proapoptotic p53-target genes in these p53-resistant cells. We found that phosphorylation of exogenous p53 on serine 46 (Ser46) was severely impaired in HSC-3 but not HSC-4 cells. A mutant mimicking Ser46-phosphorylation (p53S46D) enhanced proapoptotic Noxa promoter activity, and overcame the resistance to p53-mediated apoptosis and growth suppression in HSC-3 cells. Conversely, a mutant defective for Ser46-phosphorylation(p53S46A) failed to suppress the growth of p53-sensitive HSC-2 cells. In contrast to HSC-3 cells, p53S46D had no effect on HSC-4 cells, and inhibition of endogenous p53R248Q by siRNA restored p53- mediated apoptosis in HSC-4 cells, indicating a dominant- negative effect of p53R248Q protein on wild-type p53 function. These results demonstrate that the defect in Ser46 phosphorylation accounts for the p53 resistance of HSC-3 cells, and provide evidence for a mechanism underlying the acquisition of p53 resistance in oral SCC

Novel Epigenetic Modulation Chitosan-Based Scaffold as a Promising Bone Regenerative Material

Bone tissue engineering is a complicated field requiring concerted participation of cells, scaffolds, and osteoactive molecules to replace damaged bone. This study synthesized a chitosan-based (CS) scaffold incorporated with trichostatin A (TSA), an epigenetic modifier molecule, to achieve promising bone regeneration potential. The scaffolds with various biphasic calcium phosphate (BCP) proportions: 0%, 10%, 20%, and 40% were fabricated. The addition of BCP improved the scaffolds’ mechanical properties and delayed the degradation rate, whereas 20% BCP scaffold matched the appropriate scaffold requirements. The proper concentration of TSA was also validated. Our developed scaffold released TSA and sustained them for up to three days. The scaffold with 800 nM of TSA showed excellent biocompatibility and induced robust osteoblast-related gene expression in the primary human periodontal ligament cells (hPDLCs). To evaluate in vivo bone regeneration potential, the scaffolds were implanted in the mice calvarial defect model. The excellent bone regeneration ability was further demonstrated in the micro-CT and histology sections compared to both negative control and commercial bone graft product. New bone formed in the CS/BCP/TSA group revealed a trabeculae-liked characteristic of the mature bone as early as six weeks. The CS/BCP/TSA scaffold is an up-and-coming candidate for the bone tissue engineering scaffold