Bioassessment of the inflammatory response of macrophages to collagen-chitosan scaffold blended with Aloe vera

The in vitro macrophage response on Aloe vera-blended collagen-chitosan (COL-CS-AV) scaffold was studied using Murine Raw 264 macrophage cells. The cells were cultured on 1-cm² scaffolds in a standard condition; at a predefined period, the cultured constructs were subjected to various experimental studies. The cytocompatibility of the constructs was assessed by dead/live examination by acridine orange/ethidium bromide (AO/EB) fluorescent dyes, and the live cells were quantified by MTT assay. The morphology of macrophages was determined using scanning electron microscopy (SEM). The cellular apoptosis and necrosis were examined by flow cytometer using annexin V FITC conjugate and propidium iodide (PI) fluorescent probes. The cellular total antioxidant level was estimated from the cell lysate of macrophages cultured on the scaffold construct. Matrix metalloproteinase-9 (MMP-9) activity was assessed by the gelatinolytic property of cell lysate by zymography technique. The expression of pro-inflammatory markers TNFα and IL-6 was studied by western blot analysis using its specific antibody. The COL-CS-AV scaffold showed promising results over the collagen-chitosan. Overall, the increased biological property with less activation of macrophage response by downregulating the TNFα and IL-6 was observed in the Aloe vera-blended scaffold construct. In support of this, the active form of MMP-9 disappeared in earlier days than without Aloe vera-blended scaffold. Through these encouraging results, we conclude that the COL-CS-AV scaffold may be used as a better solution for tissue engineering applications.