The Inhibitory Effect Of SSO On Experimental Abdominal Aortic Aneurysm In Mice

BackgroundAbdominal aortic aneurysm(AAA)is a common arterial dilatation disease in vascular surgery and has become an important health risk in the elderly.The prevalence of abdominal aortic aneurysm in elderly men over 65 years of age can reach 8%,and as the lumen diameter of abdominal aortic aneurysm increases with age,the risk of rupture also increases,and in case of rupture,the mortality rate of patients can be over 80%.Currently,surgery is the only effective treatment route to prevent and treat ruptured abdominal aortic aneurysms.Surgical intervention is considered for patients with a maximum aneurysm diameter of more than 5.5 cm and 5 cm for men and women,respectively.In contrast,for smaller diameter abdominal aortic aneurysms,there are no drugs that can effectively inhibit the progression of the aneurysm,and regular follow-up and periodic monitoring of the change in aneurysm diameter by imaging are currently the main preventive interventions.Therefore,the search for clinical agents suitable for the treatment of abdominal aortic aneurysms is of crucial importance.The formation of abdominal aortic aneurysms is a complex pathophysiologic process.The conventional view is that atherosclerosis is the most common cause of AAA.Chronic infiltrative inflammation of the duct wall and macrophage activation are the main features of atherosclerosis and the basis for abdominal aortic aneurysm formation.It has been shown that the most striking pathological feature of abdominal aortic aneurysm development is the increased infiltration of macrophages and lymphocytes.These inflammatory cells also secrete matrix metalloproteinases(MMPs)and various inflammatory factors such as tumor necrosis factor-α,interleukin-1β,and interleukin-6.The local inflammatory response promotes the degradation of elastic fibers and apoptosis of vascular smooth muscle cells in the arterial vessel wall,resulting in reduced vessel wall compliance and a decrease in The lumen is continuously dilated under the impact of blood flow,which eventually leads to rupture of abdominal aortic aneurysm.CD36,also known as scavenger receptor B2,is a transmembrane protein pattern receptor expressed on the surface of a variety of cells and can bind to a variety of ligands,thereby mediating different biological processes such as innate immunity,atherosclerosis and inflammation.Macrophages internalize oxidized low-density lipoprotein(ox-LDL)with the help of their surface CD36 receptor,thereby confining it to the vascular intima.Prolonged uptake of ox-LDL by macrophages leads to intracellular accumulation of lipids and the formation of foam cells,which is the key initiation process of atherosclerosis.Thus,CD36 plays a crucial role in lipid metabolism,especially in the formation of atherosclerosis.Given the close association between abdominal aortic aneurysm and atherosclerosis and the important role of CD36 in the whole pathological process of atherosclerosis,as well as the fact that some scholars have found significantly higher expression of CD36 in the aneurysmal tissues of AAA patients compared with normal human aortic tissues in recent years,we speculate that CD36 may be related to the development of abdominal aortic aneurysm.Sulfosuccinimidyl oleate(SSO)is a specific inhibitor of the CD36 receptor.SSO inhibits the uptake of oxidized LDL by macrophages through specific inhibition of FAT/CD36mediated cellular fatty acid uptake,and has been shown to improve neuroinflammation after diabetic cardiometabolic disorders and cerebral ischemic stroke.Some ameliorative effects were observed.However,the role of SSO in abdominal aortic aneurysms has not been investigated.In this study,we will examine the expression level of CD36 in patients with abdominal aortic aneurysm and mouse abdominal aortic aneurysm model,and further investigate the effect of CD36 inhibitor SSO on mouse abdominal aortic aneurysm model to clarify the potential relationship between CD36 and in abdominal aortic aneurysm.Objective1.To study the expression of CD36 in the serum and tissues of the aneurysm wall of patients with abdominal aortic aneurysm.2.To study the expression of CD36 in serum and aneurysm wall tissues of a mouse abdominal aortic aneurysm model.3.To clarify the potential relationship between CD36 and AAA,to study the effect of SSO inhibitor on mouse abdominal aortic aneurysm model,and then to provide effective experimental data to support the treatment of abdominal aortic aneurysm.Methods1.Ten blood samples were obtained from patients with an abdominal aortic aneurysm diagnosed by computed tomography(CT)at Shandong Provincial Hospital between May 2021 and December 2021,along with 8 blood samples from similarly aged healthy volunteers.We collected eight pathologic tissue samples from the abdominal aortic aneurysm wall collected from patients undergoing repair of an open abdominal aortic aneurysm at the Department of Vascular Surgery at Shandong Provincial Hospital,as well as eight agematched normal tissue samples of the abdominal aortic vessel wall obtained from multiple organ donors.CD36 expression in human serum was determined using ELISA;mRNA and protein expression levels of CD36 in the aneurysmal wall tissues were determined by qRTPCR,Western blotting,immunohistochemical staining and other experimental techniques,respectively.2.Angiotensin Ⅱ-induced ApoE-/-mice were used to establish a mouse abdominal aortic aneurysm model.The mice were randomly divided into three groups,Sham,AAA and AAA+SSO.Micro-osmotic pumps were implanted subcutaneously in the posterior neck of the mice,and saline was pumped into the Sham group,while Ang II was pumped into the AAA and AAA+SSO groups for 28 days of induction.Meanwhile,mice in the AAA+SSO group were given SSO by intraperitoneal injection for intervention,and dosing was started immediately after the micro-osmotic pump implantation procedure for 28 days.mice in the Sham and AAA groups were injected intraperitoneally with the same volume of co-solvent.The change in the maximum diameter of the abdominal aorta of the mice during the modeling process was measured using ultrasound detection.After 28 days of drug administration,the aneurysmal segments of the mice were taken and sections were prepared for HE and EVG staining to clarify the morphological changes of the abdominal aortic aneurysms from macroscopic to microscopic levels,and were compared and analyzed to observe the basic morphological differences between the different groups and to determine whether the modeling was successful.The serum of each group of mice was collected for ELISA assay to determine their CD36 content.The mRNA and protein expression of CD36 in the aneurysmal wall of each group were compared using qRT-PCR,Western blotting and immunohistochemical staining.p65,p-p65,MMP-2,MMP-9 were measured in the abdominal aorta of each group by WB test.qRT-PCR was performed to detect IL-1β in the abdominal aorta of each group.The mRNA expression levels of IL-1β,IL-6,TNF-α,MMP-2,MMP-9 and NF-κB were measured by qRT-PCR in the abdominal aortic tissues of each group of mice.In addition,the expression of α-SMA,CD68,CD31 and B220 in the abdominal aorta of each group of mice was analyzed by immunohistochemical staining to evaluate the extent of vascular smooth muscle loss and inflammatory cell infiltration in each group of mice.Results1.CD36 expression was upregulated in the aneurysm wall tissues of patients with abdominal aortic aneurysms,and there was no significant difference in serum CD36 levels between patients with abdominal aortic aneurysms and normal subjects.2.CD36 expression was increased in mice with abdominal aortic aneurysm wall tissues.There was no significant difference in serum CD36 levels between groups of mice.3.SSO reduced the incidence of abdominal aortic aneurysm in experimental mice and effectively reduced the maximum diameter of the dilated abdominal aorta.4.SSO significantly inhibited the loss of smooth muscle cells and degradation of elastic fibers in the aortic aneurysm wall of experimental mice,and effectively improved the histopathological damage of the aortic aneurysm wall in mice.5.SSO attenuated the inflammatory response and neovascularization of the aortic aneurysm wall in experimental mice.6.SSO inhibited the expression of CD36,p65 and p-p65 in the abdominal aortic aneurysm tissues of experimental mice.ConclusionIn this study,we demonstrated for the first time the ability of SSO to inhibit the formation and development of abdominal aortic aneurysms in experimental mice.SSO as a CD36-specific inhibitor significantly reduced the expression level of CD36 in mouse abdominal aortic tissues,while SSO significantly inhibited the degradation of elastic fibers in the wall of mouse abdominal aortic aneurysms,reduced the loss of smooth muscle cells in the wall,and attenuated inflammatory cell infiltration and neovascularization in the wall.SSO could significantly inhibit the degradation of elastic fibers in the wall of mouse abdominal aortic aneurysms,reduce the loss of smooth muscle cells in the wall,reduce the inflammatory infiltration and neovascularization of the wall,and thus limit the occurrence and development of abdominal aortic aneurysms.And the inhibitory effect of SSO on AAA progression in mice may be related to the regulation of NF-κB pathway.SSO may be a potential therapeutic approach to inhibit the progression of abdominal aortic aneurysms.