羊脐带提取物的制备工艺及其抗炎功效探究OA

In this study,sheep umbilical cord extract(S-UCE)was obtained through a multi-step purification process involving enzymatic hydrolysis,homogenization,ultrasonication and filtration.The protein mass concentration of S-UCE,as determined by the bicinchoninic acid(BCA)assay,was(1.64±0.37)mg/mL.The effect of S-UCE on cell viability of human dermal fibroblasts(HDFs),human immortalized keratinocytes(HaCaT),and RAW 264.7 macrophages were evaluated using the MTT colorimetric assay.Results demonstrate that S-UCE at protein mass concentrations ranging from 0 to 400 μg/mL exhibit no cytotoxicity and promote cell proliferation by 50％-100％in the aforementioned cell lines.An inflammatory model was established using lipopolysaccharide(LPS)-stimulated RAW264.7 cells.Morphological changes were observed using microscope,while nitric oxide(NO)release in the culture supernatant was quantified using the Griess reagent method.Enzyme-linked immunosorbent assay(ELISA)was employed to measure the expression levels of pro-inflammatory cytokines,including tumor necrosis factor-alpha(TNF-α),interleukin(IL)-6,and IL-1 β.Furthermore,western blotting and immunofluorescence assays were performed to detect the protein expression of polarization markers CD86(M1 phenotype)and CD206(M2 phenotype)in RAW264.7 cells.The results indicate that S-UCE at protein mass concentrations of 50-200 μg/mL significantly attenuate LPS-induced morphological alterations in RAW264.7 cells,reduce NO production,and suppress the expression of IL-6,IL-1β,and TNF-α.Notably,S-UCE at 100 μg/mL exhibits the most potent anti-inflammatory effects,downregulating the relative protein expression of CD86 while upregulating CD206 expression.These findings demonstrate that the prepared S-UCE possesses favorable biosafety and anti-inflammatory properties,providing an experimental foundation for the development of novel animal-derived functional ingredients in cosmetic applications.