Indole-3-Aldehyde Delays Abdominal Aortic Aneurysm Progression By Modulating Phenotypic Switching Of Vascular Smooth Muscle Cells

Objective: Abdominal aortic aneurysms(AAA)is one of the most severe diseases in the field of vascular surgery.Not only does it have an adverse impact on patients’ health but imposes substantial financial burden on their families as well as society.Despite the development of new techniques,the option for the treatment of AAA is still limited.Particularly,there is a lack of effective pharmacological treatment.The complexity of AAA’s pathogenesis poses further challenges to medication development.Recent studies have demonstrated that the intestinal microbiota and its metabolites played a significant role in the treatment of cardiovascular diseases.However,it is unclear how the microbiota and its metabolites involve and regulate the process of AAA pathogenesis.Therefore,this study aimed to investigate the regulatory role of the tryptophan metabolite indole-3-aldehyde(3-IAld)derived from the intestinal microbiota in the development and progression of AAA.Methods:(1)Blood and tissue samples were collected from patients with AAA and healthy participants.The difference in the concentration of 3-IAld in serum between AAA and Shame groups was then analyzed,using high performance liquid chromatography(HPLC).Immunofluorescence was used to detect differences in protein expression of alpha-smooth muscle actin(α-SMA),smooth muscle protein 22 alpha(SM22α),and osteopontin(OPN)in human aneurysm tissues and healthy controls.The difference in expression levels of aryl hydrocarbon receptor(AHR)in arterial wall tissues between the two groups were detected by quantitative real-time polymerase chain reaction(q RT-PCR)and western blotting,respectively.(2)The AAA mouse model was built using the elastase perfusion method,and50 mice were randomly allocated to five groups with 10 mice in each group: sham-operated group(Sham,saline perfusion),AAA group(AAA,elastase perfusion),low-dose 3-IAld treatment group(3-IAld50,elastase perfusion,50mg/kg.d gavage),high-dose 3-IAld treatment group(3-IAld150,elastase infusion,150mg/kg.d gavage),and the 3-IAld high concentration treatment group + AHR antagonist group(3-IAld150 + CH-22319,elastase infusion,150mg/kg.d gavage + 10mg/kg.d intraperitoneal injection).These mice were euthanized two weeks after the operation.Serum and abdominal aortic tissues were immediately collected for subsequent experiments.HPLC was used to detect the difference in the concentration of indole-3-aldehyde in the serum of the AAA,high-dose 3-IAld treatment group,and sham-operated mice.The specimens were also subjected to hematoxylin-eosin(HE)staining,elastic fiber(EVG)staining,and Masson staining to observe the histomorphology of vessel walls in different groups.q RT-PCR and immunofluorescence were used to detect differences in the expression of α-SMA,SM22α,and OPN in the arterial walls of different groups.Immunohistochemistry was used to detect differences in the expression of matrix metalloproteinase-2(MMP-2)and MMP-9 in the arterial walls of different groups.q RT-PCR and immunofluorescence were used to detect differences in the expression of AHR in the arterial walls of different groups.Results:(1)In comparison to the healthy population,AAA patients showed a reduction in the concentration of 3-IAld in their serum.In addition,there was a decrease in the expression of α-SMA and SM22α in tissues,as well as the expression of the AHR in the arterial wall.By contrast,an increased expression of OPN was observed in the aneurysm tissues of patients.(2)Animal experimental studies showed that mice with AAA had lower serum levels of 3-IAld than the sham group,and the high-dose 3-IAld treatment group had higher serum levels of 3-IAld than the AAA group.Administration of exogenous 3-IAld in mice with the AAA model significantly reduced the expansion of aneurysm diameter and improved the degradation of elastic fibers in the arterial wall.Moreover,the treatment inhibited the phenotype of VSMCs in the vessel wall of mice with AAA.In the 3-IAld administration group,the expression of AHR significantly increased compared to the AAA group,and the most significant increase was observed in the 3-IAld150 group.The addition of CH-22319,an AHR antagonist,delayed the therapeutic effect of 3-IAld on AAA,further supporting the notion that 3-IAld delayed the progression of abdominal aortic aneurysm through AHR regulation of VSMCs phenotype transition.Conclusions: The expression and concentration of 3-IAld and its receptor AHR in the serum of AAA patients decreased,which was negatively correlated with the aortic diameter,accompanied by phenotype conversion of VSMCs in aortic wall aneurysm specimens of AAA patients.In addition,3-IAld was found to inhibit the progression of AAA mouse models constructed by elastase infusion.This maybe indicated the mechanism that 3-IAld regulated the phenotypic transformation of VSMCs via specific binding of AHR.