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While many large-scale risk factors for calcific aortic valve disease (CAVD) have been identified, the molecular etiology and subsequent pathogenesis of CAVD have yet to be fully understood. Specifically, it is unclear what biological phenomena underlie the significantly higher occurrence of CAVD in the male population. We hypothesized the existence of intrinsic, cellular-scale differences between male and female valvular interstitial cells (VICs) that contribute to male sex being a risk factor for CAVD. Differences in gene expression profiles between healthy male and female porcine VICs were investigated via microarray analysis. Mean expression values of each probe set in the male samples were compared to the female samples, and biological processes were analyzed for overrepresentation using Gene Ontology term enrichment analysis. There were 183 genes identified as significantly (fold change>2; P<0.05) different in male versus female aortic valve leaflets. Within this significant gene list there were 298 overrepresented biological processes, several of which are relevant to pathways identified in CAVD pathogenesis. In particular, pathway analysis indicated that cellular proliferation, apoptosis, migration, ossification, angiogenesis, inflammation, and extracellular matrix reorganization were all significantly represented in the data set. These gene expression findings also translated into functional differences in VIC behavior in the  in vitro  environment, as sex-related differences in proliferation and apoptosis were confirmed in VIC populations cultured  in vitro . These data suggest that a sex-related propensity for CAVD exists on the cellular level in healthy subjects, a phenomenon that could have significant clinical implications. These findings also strongly support discontinuing the use of mixed-sex VIC cultures, thereby changing the current standard in the field.

Sex-Related Differences in Gene Expression by Porcine Aortic Valvular Interstitial Cells