The Role And Mechanism Of Nampt In The Onset And Development Of Atherosclerosis

Atherosclerosis is the leading cause of various cardiovascular diseases such as stroke,coronary heart disease,peripheral vascular disease and so on.With the continuous improvement of living standards,the incidence of atherosclerosis rises rapidly.The induction of atherosclerosis is lipid metabolism disorders.The plaques are formed by the accumulation of diseased vascular cells and apoptotic cells.Once the rupture of plaque will cause blood vessel embolism,inducing stroke and endangering human life and health.Nicotinamide phosphoribosyltransferase(NAMPT)is a fat factor secreted by adipocytes.Nampt plays an important role in both intracellular and extracellular activity.Namatin is a rate-limiting enzyme synthesized by nicotinamide adenine dinucleotide(NAD),which converts the substrate nicotinamide(NAM)into product nicotinamide mononucleotide(NMN).NMN generates NAD under catalytic conditions.NAD,known as coenzyme I,is an essential coenzyme involved in the redox reaction of the mitochondrial respiratory chain.The biological function of NAD is extremely important and NAD may be able to determine the fate of cells directly.Our group previously demonstrated the neuroprotective effect of Nampt after cerebral ischemia.However,the exact role of Nampt in the pathogenesis of ischemic stroke remains unclear.Therefore,we investigate the effect of Nampt on the development and progression of atherosclerosis by using NMN in vitro and Apo E-/-NamptTg mice.We found that Nampt could promote the development of atherosclerosis.In this study,we further studied the role of TNF-α in the development of atherosclerosis.The mechanism of the role of Nampt in atherosclerosis is discussed,which may provide a new target for the treatment of atherosclerosis.Methods: 1.Eight male Apo E-/-mice were devided into two groups with western high fat diet at the same time.Two months later,the experimental group drank NMN water,and the dosage of NMN was 300 mg/kg/day.The control group drank normal water at the same time.The effect of NMN on the thickness and size of plaque were observed by oil red O staining and HE staining.F4 / 80 immunohistochemical staining was used to observe the macrophage infiltration of aortic root plaque,and the effect of NMN supplementation on plaque stability was determined.TNF-α immunohistochemical staining was used to determine the effect of NMN on the expression of inflammatoryfactors.2.We established Apo E-/-;NamptTg mice as experimental group,and Apo E-/-mice as control.After 4 months western high fat diet,the effect of Nampt overexpression on aortic lipid deposition was observed by oil red O staining.Oil red O staining and HE staining were used to observe the effect of Nampt overexpression on the thickness and size of atherosclerotic plaque.The effects of Nampt overexpression on the stability of atherosclerotic plaques were observed by F4/80 immunohistochemistry.We use masson trichrome stainingand sirius red staining to observe the effect of Nampt on atherosclerotic plaque fibrous cap and collagen.TNF-α and IL-1β were immunohistochemically stained to observe the effect of Nampt overexpression on the expression of inflammatory cytokines in atherosclerotic plaques.ICAM-1 and VCAM-1 were used to observe the effect of Nampt overexpression on the level of chemokines in plaques.TUNEL was used to observe the effect of Nampt overexpression on the number of apoptotic cells in plaques.In addition,CD47 immunohistochemical staining of frozen sections of aorta root was carried out to explore the mechanism of Nampt on atherosclerosis.3.The experimental group were given TNF-α by intraperitoneal injection at a doseage of 5 mg/kg/day for one months,and Control group were given PBS by intraperitoneal injection at the same time.Oil red O staining was used to observe the effect of Nampt overexpression on the thickness and size of atherosclerotic plaque.The effects of Nampt overexpression on the stability of atherosclerotic plaques were observed by F4/80 immunohistochemistry andα-SMA immunohistochemical staining.CD47 immunohistochemical staining and TUNEL cell apoptosis detection were used to further explore the mechanism of Nampt in the development of atherosclerosis.Results: 1.Compared with the control group,the mouse in NMN group had more lipid deposition on the aorta,larger plaque area,higher degree of macrophage infiltration,poor stability of plaque,and elevated level of TNF-α.NMN promotes the formation of atherosclerotic plaques,inflammatory infiltration and plaque instability.2.The results of O staining showed that compared with Apo E-/-group,the mouse in Apo E-/-;NamptTg group had more lipid deposition.The overexpression of Nampt increased plaque area and thickness,macrophage infiltration,necrosis of the center,type III collagen,plaque instability,the level of inflammatory factors(TNF-alpha,IL-1 beta)and chemokine(ICAM-1,VCAM-1)and the number of cells apoptosis in plaque.3.Overexpression of Nampt promotes the occurrence and development of atherosclerosis along with the increase of CD47 expression.The inhibition of TNF-α may not affect blood lipid and plaque area.The level of F4/80 and α-SMA decreased and plaque stability was improved when the TNF-α was suppressed.In addition,the inhibition of TNF-α may cause the lower level of CD47 and suppression of plaque cell apoptosis death.Conclusion: 1.The supplementation of NMN can promote the development of atherosclerosis.2.Nampt overexpression can lead to the development of atherosclerosis,and the atherosclerosis deterioration may be related to the up-regulation of CD47.3.Inhibition of TNF-α can delay the process of atherosclerosis.Nampt may promote the development of atherosclerosis through the secretion of TNF-α which can induce the up-regulation of CD47.