HXJN Activates ADAM17 To Regulate Platelet Surface Protein Through P38

Background and Objective: Thrombotic diseases are a series of diseases caused by blood clots formed on the surface of blood vessels or heart intima,resulting in the interruption of blood flow.The incidence rate is increasing year by year,and has become a major health problem worldwide.Platelets are non nucleated cells formed by the cytoplasmic shedding of mature bone marrow megakaryocytes.They are mainly involved in hemostasis and thrombosis,and also play an important role in pathophysiological processes such as inflammation and angiogenesis.HXJN is a traditional Chinese medicine compound preparation based on Buyang Huanwu Decoction.It is mainly used to treat atherosclerosis,coronary heart disease and other diseases caused by deficiency of Qi and blood and blood stasis.Previous studies have found that a variety of active components in HXJN can activate ADAM17(a disintegrin and metalloproteinase 17)in vascular endothelial cells to play anti-inflammatory and cytoprotective roles.Related studies have shown that platelet membrane receptor glycoprotein Ibα can produce extracellular fragment shedding under the action of ADAM17,which weakens its interaction with von Willebrand factor.The binding of v WF and other ligands is an important mechanism of platelet function regulation in vivo.The aim of this study was to investigate the effect of HXJN on the activation of tumor necrosis factor-α converting enzyme(TACE)in platelets and the role of ADAM17 in regulating platelet membrane receptor GPIbα signaling pathway,further clarification the key link and molecular mechanism of HXJN.It provides a new direction for the research of traditional Chinese medicine compound and monomer.Methods: DAMI cells(human megakaryocytic leukemia cells)were cultured in vitro or fresh human platelets were isolated.The quality of isolated platelets was evaluated by Giemsa staining of platelet rich plasma.After treated with different concentrations of HXJN aqueous extract,the content of GPIbα extracellular fragment(GC)in cell culture medium or platelet rich plasma(PRP)was detected by ELISA;Cell lysates were collected that is to were detect the expression levels of p-TACE / TACE,p-p38 and GPIbα by Western blot;The activity of ADAM17 was detected by fluorescence method.Results:(1)The isolated platelets were observed by Giemsa staining,and the physiological state and function of platelets were good,which could be further tested;(2)HXJN induced a significant increase of TACE activity in platelets;(3)HXJN could induce a rapid and significant increase of GC content of GPIbα extracellular fragment in DAMI cells culture supernatant or PRP;(4)The levels of p-TACE(thr735),p-p38 in HXJN induced cells increased significantly with time;(5)The increase of GC content induced by HXJN was significantly inhibited by p38 inhibitor.Conclusion: By activating p38 signal,HXJN induces phosphorylation of ADAM17 in platelets and platelet precursor cells and increases the activity of ADAM17 enzyme,which makes the extracellular ligand binding fragment of platelet membrane protein receptor GPIbα fall off,and then regulates the function of platelets.It will provide a new direction for the research of traditional Chinese medicine compound and monomer.