Enhanced Expression of the Female-Biased Gene Gsta2 in Male Proximal Tubule Cells Improves Renal Resilience to Ischemia-Reperfusion Injury.

Abstract

Elevated <i>Gsta2</i> expression in female proximal tubules associates with female resilience to ischemia-reperfusion injury. Transgenic expression of <i>Gsta2</i> in male proximal tubule cells directed by human <i>HNF4A</i> enhancers ameliorated ischemia-reperfusion injury outcomes. Ectopic <i>Gsta2</i> attenuated protein peroxidation and double-strand DNA breakage, reduced fibrosis, and improved proximal tubule repair. Genetic sex is an important determinant of kidney injury and repair, with female kidneys typically exhibiting greater resilience to AKI. Among the sexually dimorphic genes in mouse proximal tubule cells, <i>Gsta2</i>, encoding an NFE2 like BZIP transcription factor 2–regulated antioxidant enzyme, is strongly enriched in females. Here, we hypothesized that augmenting <i>Gsta2</i> expression in male proximal tubule cells will enhance resistance to ischemia-reperfusion injury (IRI). To enable proximal tubule cell–specific expression of transgenes, we mapped and verified enhancer regions directing proximal tubule expression of human <i>HNF4A.</i> A synthetic HNF4A enhancer cassette driving <i>Gsta2</i> was introduced into a safe harbor locus in transgenic mice, thereby enhancing the expression of <i>Gsta2</i> in male mice. After unilateral nephrectomy, transgenic and wild-type male mice were subjected to IRI. Post-IRI outcomes were assessed by examining kidney function, histologic injury, and fibrotic progression for up to 28 days postinjury. Enhancing <i>Gsta2</i> expression in male proximal tubule cells led to significantly higher glomerular filtration rates and attenuated fibrotic remodeling after IRI. Early-phase transcriptional analyses 4 hours postinjury showed reduced expression of immediate early genes (<i>Jun</i>, <i>Fos</i>, <i>Egr1</i>), suggesting a reduced stress response, diminished DNA double-strand DNA breaks (gamma-H2AX, the phosphorylated form of the histone variant H2AX incorporation into chromatin), and lower protein peroxidation. Later-stage transgenic kidneys exhibited a reduction in fibrosis-associated transcripts (<i>Acta2</i>, <i>Col1a1</i>, <i>Col3a1</i>) and markers of failed proximal tubule cells repair (<i>Havcr1</i>, <i>Vcam1</i>, <i>Ccl2</i>). Ectopic expression of <i>Gsta2</i> in male proximal tubule cells reduced oxidative injury, injury-associated fibrosis, and maladaptive stress signaling after IRI.