Abstract 82: The Second X Chromosome Contributes to Stroke Sensitivity in the Absence of Activational Hormonal Effects

Abstract

Introduction: Stroke is a sexually dimorphic disease. Stroke sensitivity in the elderly is altered by the decline of estrogen levels in females. Our previous studies suggested a X chromosomal effect on stroke sensitivity in aged mice; however, these mice were gonadal intact and the activational effect of hormones cannot be ruled out. In this study, we utilized the XY* mice model in which mice of four genotypes were produced (XX, XO, XY, and XXY) to dissociate the effect of X chromosome from that of Y. To rule out the activational effect of hormones, we gonadectomized all mice at 3 weeks old, and let them survive for 6-8 weeks to achieve “surgical post-menopause”. We hypothesized that the second X chromosome mediates the stroke sensitivity in the absence of activational sex hormones. Two X escapee genes ( Kdm6A and Kdm5C ) that escape from the X chromosome inactivation and involved in microglial activation, were also examined to explore the potential mechanisms. Methods: “Surgical postmenopausal” XY* mice were subjected to a 60-min middle cerebral artery occlusion (MCAO) (n=6-8/gp for stroke and = 4-5/gp for sham). Infarct volumes and behavior deficits were quantified 3 days after MCAO. Plasma inflammatory cytokine (TNFα, IL-1β, IL-4, iNOS, TGF-β1 and IL-10) levels were determined by ELISA. KDM6A and KDM5C expression in microglia was examined by IHC. Results: XX mice had significantly larger infarct size than XO mice in striatum (55.02±12.38% vs. 36.06±9.25%, P=0.0387) and the ipsilateral hemisphere (45.33±10.77% vs. 26.55±6.92%, p=0.0413); the same pattern was seen in XXY vs. XY group in striatum (58.47±14.26% vs. 34.22±15.24%, P=0.0027). Neurological deficit scores were higher in XX vs. XO mice. Mice with two copies of the X chromosome had significantly higher plasma levels of pro-inflammatory cytokines (TNFα, IL-1β) than the mice with one X. Significantly higher levels of KDM6A or KDM5C in Iba-1 positive microglial cells were also found in mice with two vs. one copy of X. Conclusion: The second X chromosome contributes to stroke sensitivity in the absence of activational sex hormones. Our future goal is to use Kdm6A and Kdm5C microglial conditional knockout mice to explore the molecular mechanisms underlying chromosomal factors in stroke sensitivity.