Withaferin A promotes proliferation and migration of brain endothelial cells

Purpose: To investigate the effect of withaferin A (WFA) on the proliferation and migration of brain endothelial cells. Methods: BALB-5023 mouse microvascular cells were treated with a range of withaferin A (WFA) concentrations from 10 to 100 ng/mL. Dojindo’s CCK-8 cell proliferation kit was used for the analysis of cell proliferation. Transwell cell culture inserts were used to determine the migration potential of WFAtreated endothelial cells. Absorbance was measured at 450 nm on an enzyme-linked immunosorbent (ELISA) reader. Results: The results revealed a significant increase in the proliferation and migration of endothelial cells following treatment with a low concentration (30 ng/mL) of WFA compared with the higher concentration (> 10 ng/mL). The effect was further enhanced when WFA was used in combination with soluble Fas ligand (sFasL). Autocrine signaling of vascular endothelial growth factor (VEGF) by endothelial cells was significantly increased following treatment with WFA or in combination with sFasL. WFA increased the expression of Fas on endothelial cells, suggesting the involvement of sFasL in the proliferation and migration of brain endothelial cells. Conclusion: Thus, WFA promotes the proliferation and migration of endothelial cells through increase in the expression of Fas and secretion of VEGF.

Development of hydroxyapatite-based nanomaterials to enhance biological response of osteoblast cells for clinical application

Purpose: To develop a novel chitosan/gelatin-hydroxyapatite (CGHaP) microspheres for evaluating the biological response of pre-osteoblast cells. Methods: The microsphere was prepared by water-in-oil emulsion method. Cell proliferation was studied using AlamarBlue colorimetric assay and DAPI staining while alkaline phosphatase assay was carried out by colorimetric assay method. Chitosan microspheres as well as chitosan-hydroxyapatite microspheres was prepared and tested for biological response from MC3T3-E1 cell line. Results: The results showed that CGHaP promotes MC3T3-E1 cell proliferation and spread on the surface of microspheres. The cells were clustered with more actin filaments and well-linked with neighbouring cells or adjacent cells when cultured in CGHaP microspheres whereas fewer cells were spread on chitosan (CH) microspheres. CGHaP microspheres significantly (p < 0.05) promoted cell attachment, proliferation and extracellular matrix mineralization. CGHaP microspheres presented significantly (p < 0.02) higher calcium deposition (0.5 ng) than CH microspheres (0.28 ng). Specifically, CGHaP microspheres exhibited high ALP activity (8 units; 2-fold) compared to CH with 3 units, after 7 days of incubation. The results suggest that CGHaP possesses a great ability to facilitate bone ingrowth formation and possibility of good osteointegration in vivo. Conclusion: The nanomaterial enhances the proliferation of pre-osteoblast cells in tissue engineering microspheres. The outcome of this study may have a major impact on the development of novel nanomaterials for bone tissue engineering.