Background: Atherosclerosis is a major pathological basis of cardiovascular disease and is influenced by both genetic and environmental factors. Although genome-wide association studies have identified the LAMB2 p.E987K variant as a susceptibility locus for atherosclerosis, its functional role remains unclear. Methods: In this study, a rabbit model carrying the homologous LAMB2 p.E991K mutation was generated using the CRISPR/Cas9-SpG system and subjected to a high-fat diet to induce atherosclerosis. Results: LAMB2 mutant rabbits exhibited increased body weight and significant lipid metabolic abnormalities. Oil Red O staining demonstrated enhanced lipid accumulation and larger atherosclerotic plaques in the aorta. In addition, α-SMA expression was reduced, whereas CD4 and MCP-1 expression was elevated, suggesting vascular smooth muscle cell loss and altered immune responses. Laminin β2 (LAMB2) expression was markedly decreased and accompanied by basement membrane disruption. Furthermore, activation of the NLRP3 inflammasome was observed in mutant rabbits. Conclusions: These findings demonstrate that the LAMB2 p.E991K mutation promotes HFD-induced atherosclerosis by impairing basement membrane integrity, enhancing inflammation, and disrupting lipid metabolism, highlighting LAMB2 as a genetic modifier of atherosclerosis susceptibility.