Supplementary Material for: Cyclic Mechanical Stretch Induced Smooth Muscle Cell Changes in Cerebral Aneurysm Progress by Reducing Collagen Type IV and Collagen Type VI Levels

Background/Aims: Cerebral aneurysm growth is characterized by continuous structural weakness of local smooth muscle cells, though the mechanism is unclear. In this study, we examine protein changes in cerebral aneurysm and human brain vascular smooth muscle cells after cyclic mechanical stretch. We further explore the relationship between the smooth muscle cell changes and reductions in the levels of collagen types IV and VI. Methods: Saccular cerebral aneurysms (n=10) were collected, and temporal artery samples were used as controls. Quantitative proteomics were analyzed and histopathological changes were examined. Smooth muscle cells were cultured in a flexible silicone chamber and subjected to 15% cyclic mechanical stretch. The effect of stretch on the cell viability, function, gene and protein expression were further studied for the understanding the molecular mechanism of aneurysm development. Results: Proteomics analysis revealed 92 proteins with increased expression and 88 proteins with decreased expression compared to the controls (p<0.05). KEGG pathway analysis showed that the change in focal adhesion and extracellular matrix-receptor interaction, suggesting the involvement of collagen type IV and VI. The aneurysm tissue exhibited fewer smooth muscle cells and lower levels of collagen type IV and VI. Human brain vascular smooth muscle cell culture showed spindle-like cells and obvious smooth muscle cell layer. Cell proteomics analysis showed that decreased expression of 118 proteins and increased expression of 32 proteins in smooth muscle cells after cyclic mechanical stretch. KEGG pathway analysis indicated that focal adhesion and ECM-receptor interaction were involved. After cyclic mechanical stretch, collagen type IV and IV expression were decreased. Moreover, the stretch induced MMP-1 and MMP-3 expression elevation. Conclusion: We demonstrated that collagen type IV and VI were decreased in cerebral aneurysms and continuous cyclic mechanical stretch induced smooth muscle cell changes. Smooth muscle cell protection provides an additional therapeutic option to prevent the growth of cerebral aneurysms.