gx50对与皮肤衰老相关nfκb通路调节作用-regulatory effect of gx50 on nf κ b pathway related to skin age.docxVIP

EMSA (electrophoretic mobility shift assay) experiment to test the nuclear transcription fator-κB DNA-binding activity.We carry out Western Blot to detect the plasma protein of skin tissue according to the standard of phosphorylation degree of IκB which restrain the activity of nuclear transcription fator-κB.At the cellular level, we conduct the primary culture of mice epidemis and identify purity by immunofluorescence staining choosing anti-PCK (Mouse Anti-Pan Cytokeratin) as the epidemal cell’s surface markers. We treat the cells with GX50 and LPS and blank control and extract membrane protein, nucleoprotein and plasma protein and measure their concentration by ultraviolet specrophotometer. It is found that GX50 can effectively activate α7 nicotin acetylcholine receptor through drug design method.We detect the ability of GX50 combine to α7 nicotin acetylcholine receptor of epidemis membrane with the application of Octet RED biological interactions analyzer.We carried out EMSA experiment to test the nuclear transcription fator-κB activity of the epidemal cells. We carry out Western Blot to detect the phosphorylation degree of IκB.Results (1)The results of the hemoatoxylin and eosin staining show that epidermis is flatter on the left than the right and the size and the shape of the basal cells is more regular and extracellular matrix arranged closely;Dermal has more follicle and more dense and more melanocytes and less melanin deposition on the left than the right; Borders of Epidermal and dermal are more obviously and the connection beween the epidermis and the dermis is more firmly on the left than the right.Immunohistochemistry of collagen protein expression in the skin of the frozen sections show that the density of collagen protein on the left is higher than that on theright（P＜0.05）.The results of EMSA of the skin tissues show that the volume of the nuclear transcription fator-κB shuffling into the nuclear in the GX50-treated skin sample was lessthan