The Relationship Between The Inactivation Of Cysteine 674 At SERCA2 And Abdominal Aortic Aneurysm And The Underlying Mechanisms Involved

Abdominal aortic aneurysm is a tumor-like expansion of the abdominal aorta,which has a high mortality rate after rupture and seriously threatens to the health of human.It is extremely important to clarify the pathogenesis of abdominal aortic aneurysm for the prevention and treatment.Sarcoplasmic/endoplasmic reticulum calcium ATPase（SERCA）2 is the main subtype of SERCA in blood vessels.SERCA2transports cytosolic Ca²⁺to the sarcoplasmic reticulum and endoplasmic reticulum through the hydrolysis of ATP,and maintains intracellular Ca²⁺homeostasis.The sulfhydryl group of cysteine located at position 674（C674）of SERCA2 is an important glutathionylation site for the vasodilator nitric oxide（NO）.NO increases the activity of SERCA2 through glutathione in this site.The irreversible oxidation of C674 is significantly increased in a series of pathological conditions such as diabetes and atherosclerosis.Angiotensin II（Ang II）is a common inducer for abdominal aortic aneurysm.We found that Ang II significantly increased the irreversible oxidation of C674 in abdominal aortic and abdominal aortic aneurysm,presumably the irreversible oxidation of C674 was involved in the formation of abdominal aortic aneurysms under pathological conditions.Therefore,we constructed SERCA2 C674S knock-in mice（SKI）in C57BL/6J background to simulate the irreversible oxidation of C674 and backcrossed them into low-density lipoprotein receptor gene deficient（LDLR-/-）background to explore the relationship between the irreversible oxidation of cysteine674 at SERCA2 and abdominal aortic aneurysm and the underlying mechanisms involved.The aim is to provide a new theoretical basis and intervention target for the abdominal aortic aneurysm’s prevention and treatment.We performed Ang II perfusion in SKI/LDLR-/-mice and their littermate control LDLR-/-mice to induce abdominal aortic aneurysm.Compared with LDLR-/-mice,the survival rate of SKI/LDLR-/-mice was lower and the incidence of abdominal aortic aneurysm was higher.The average length and volume of the abdominal aortic aneurysm were increased significantly.Most of abdominal aortic aneurysms in SKI/LDLR-/-mice were associated with thrombosis.The histological results showed that in the abdominal aortic aneurysm from SKI/LDLR-/-mice,the elastic fiber was destroyed more severely,and the collagen deposition was more slightly compared with the ones from LDLR-/-mice.In aorta and aortic smooth muscle cells in C57BL/6J background,we examined the expression of proteins associated with abdominal aortic aneurysm and inflammatory response related factors in SKI mice and control mice.We found that the irreversible oxidation of C674 significantly up-regulated the protein expression of collagen I（COL I）,collagen III（COL III）,matrix metalloproteinase 2（MMP2）,osteopontin（OPN）,phospho-p65 nuclear factor kappa B（p-p65 NF-κB）,vascular cell adhesion molecule1（VCAM1）and intercellular cell adhesion molecule1（ICAM1）.In addition,the irreversible oxidation of C674 led to a sustained increase in the concentration of intracellular Ca²⁺,which activated the calcineurin-nuclear factor of activated T cells（CaN-NFAT）signaling pathway,and mediated the up-regulation of the above proteins in SKI smooth muscle cells.Our results showed that the irreversible oxidation of SERCA2 C674 caused by the susceptibility to pathological factors in abdominal aortic aneurysms aggravated Ang II induced abdominal aortic aneurysm,and we speculated that it was related to activation of CaN-NFAT signaling pathway in aortic smooth muscle cells.The activation of CaN-NFAT signaling pathway increased the expression of abdominal aortic aneurysm related proteins and inflammatory response related factors,which caused the imbalance of ECM synthesis and degradation,and inflammatory response,and consequently promoted the formation of abdominal aortic aneurysms.