Biochemical and functional analysis of corticotropin releasing factor purified from an aqueous extract of human placenta used as wound healer
Human placental extract constitutes of innumerable therapeutically important components mostly used in wound healing arising from the skin and burn injuries. However, there is still some bioactive present in the placental extracts yet to be characterized to better under the complex process of wound healing mediated by the placental extract. In this study, the presence of corticotropin releasing factor (CRF) in an aqueous extract of human placenta was detected and quantified by dot blot and CRF-ELISA immunoassay kit respectively. Subsequently, it was purified by immuno-affinity chromatography and quantified as 0.45 ± 0.05 μg of CRF per ml of placental extract where its molecular weight found to be 4.78 kDa by MALDI-TOF. To study functional analysis of CRF, an in vitro WI-38 lung fibroblast cell scratch wound model was used which indicated proliferation, motility of cells after treatment with purified CRF. Moreover, reduction in apoptosis rate of cells during closure of wound was observed from microscopy studies and FACS analysis. Also, Antalarmin, an antagonist of CRF type 1 receptor inhibited the wound closure potency of the purified component. Faster healing of wound with an elevation of IL-6 and TGF-β during early stages of repair by placental CRF was observed on excision rat model. The process of healing was accompanied by the decrease in the level of TNF-α and IFN-γ.