Microbiota control acute arterial inflammation and neointimal hyperplasia development after arterial injury

Abstract

BackgroundThe microbiome has a functional role in a number of inflammatory processes and disease states. While neointimal hyperplasia development has been linked to inflammation, a direct role of the microbiota in neointimal hyperplasia has not yet been established. Germ-free (GF) mice are an invaluable model for studying causative links between commensal organisms and the host. We hypothesized that GF mice would exhibit altered neointimal hyperplasia following carotid ligation compared to conventionally raised (CONV-R) mice.MethodsTwenty-week-old male C57BL/6 GF mice underwent left carotid ligation under sterile conditions. Maintenance of sterility was assessed by cultivation and 16S rRNA qPCR of stool. Neointimal hyperplasia was assessed by morphometric and histologic analysis of arterial sections after 28 days. Local arterial cell proliferation and inflammation was assessed by immunofluorescence for Ki67 and inflammatory cell markers at five days. Systemic inflammation was assessed by multiplex immunoassays of serum. CONV-R mice treated in the same manner served as the control cohort. GF and CONV-R mice were compared using standard statistical methods.ResultsAll GF mice remained sterile during the entire study period. Twenty-eight days after carotid ligation, CONV-R mice had significantly more neointimal hyperplasia development compared to GF mice, as assessed by intima area, media area, intima+media area, and intima area/(intima+media) area. The collagen content of the neointimal lesions appeared qualitatively similar on Masson’s trichrome staining. There was significantly reduced Ki67 immunoreactivity in the media and adventitia of GF carotid arteries 5 days after ligation. GF mice also had increased arterial infiltration of anti-inflammatory M2 macrophages compared to CONV-R mouse arteries and a reduced proportion of mature neutrophils. GF mice had significantly reduced serum IFN-γ-inducible protein (IP)-10 and MIP-2 5 days after carotid ligation, suggesting a reduced systemic inflammatory response.ConclusionsGF mice have attenuated neointimal hyperplasia development compared to CONV-R mice, which is likely related to altered kinetics of wound healing and acute inflammation. Recognizing the role of commensals in the regulation of arterial remodeling will provide a deeper understanding of the pathophysiology of restenosis and support strategies to treat or reduce restenosis risk by manipulating microbiota.

Animal Models of Neointimal Hyperplasia and Restenosis

“…An influence of the microbiome on the development of atherosclerosis and restenosis has also been suggested in recent studies. A comparison of aseptic and conventionally raised mice showed that the latter had more advanced neointimal formation, which was probably related to changes in the kinetics of acute inflammation and wound healing ( 33 ). Intestinal dysbiosis also appeared to contribute to the development of atherosclerosis by modulating inflammatory events caused by differences in the production of microbial metabolites ( 34 , 35 ).…”

Germ-free housing conditions do not affect aortic root and aortic arch lesion size of late atherosclerotic low-density lipoprotein receptor-deficient mice

“…During the last decade, an increasing number of studies has provided a wealth of association-based [1][2][3] and causal evidence, 1,[4][5][6][7] linking the microbiota and specific bacterial community members [8][9][10] to the development of atherosclerotic lesions and cardiovascular disease (CVD) (for overview see Table 1). 11 The gut microbiota has been recognized as an environmental factor that influences endothelial cell functions and contributes to vascular inflammatory phenotypes, [12][13][14] fostering arterial thrombosis through various prothrombotic mechanisms. [15][16][17][18] While experiments with germ-free (GF) apolipoprotein E (Apoe)deficient mouse models at chow diet conditions have repeatedly shown a protective role of the microbiota in atherogenesis, 4,6 it remains controversial how HFD and different feeding regimens affect atherosclerotic lesion development in mouse atherosclerosis models under GF housing conditions.…”

Dysbiosis of Gut Microbiota and Metabolite Phenylacetylglutamine in Coronary Artery Disease Patients With Stent Stenosis

“…Accumulating evidence has revealed the association between gut dysbiosis and arterial remodeling. Compared to conventional mice, germ-free mice show attenuated neointimal hyperplasia development and reduced local inflammation in arteries after carotid ligation (21). Concomitantly, CAD patients with instent hyperplasia and stenosis showed significant alteration in gut microbiota diversity and composition compared to CAD patients with in-stent patency.…”