Transcriptome analysis in rat kidneys: importance of genes involved in programmed hypertension

Abstract

Suboptimal conditions in pregnancy can elicit long-term effects on the health of offspring. The most common outcome is programmed hypertension. We examined whether there are common genes and pathways in the kidney are responsible for generating programmed hypertension among three different models using next generation RNA sequencing (RNA-Seq) technology. Pregnant Sprague-Dawley rats received dexamethasone (DEX, 0.1 mg/kg) from gestational day 16 to 22, 60% high-fructose (HF) diet, or NG-nitro-l-arginine-methyester (l-NAME, 60 mg/kg/day) to conduct DEX, HF, or l-NAME model respectively. All three models elicited programmed hypertension in adult male offspring. We observed five shared genes (Bcl6, Dmrtc1c, Egr1, Inmt, and Olr1668) among three different models. The identified differential genes (DEGs) that are related to regulation of blood pressure included Aqp2, Ptgs1, Eph2x, Hba-a2, Apln, Guca2b, Hmox1, and Npy. RNA-Seq identified genes in arachidonic acid metabolism are potentially gatekeeper genes contributing to programmed hypertension. In addition, HF and DEX increased expression and activity of soluble epoxide hydrolase (Ephx2 gene encoding protein). Conclusively, the DEGs in arachidonic acid metabolism are potentially gatekeeper genes in programmed hypertension. The roles of DEGs identified by the RNA-Seq in this study deserve further clarification, to develop the potential interventions in the prevention of programmed hypertension.

Figure 1

Effects of dexamethasone (DEX), high-fructose intake (HF), and N^G-nitro-l-arginine-methyester (l-NAME) on systolic blood pressure in male adult offspring. n = 6 per group. * p < 0.05 vs. control.

Figure 2

(A) Venn diagram depicting unique and shared (overlapping circles) sets of DEGs between high-fructose intake (HF, black circle), dexamethasone treatment (DEX, white circle), and l-NAME treatment (grey circle); (B) Confirmatory analysis of kidney gene expression by qPCR in the l-NAME model. Individual gene expression was determined by qPCR and expressed as fold change in log base 2 vs. control and graphed alongside fold change derived from RNA-Seq analysis.

Figure 3

(A) Effects of HF on gene expression of arachidonic acid metabolism pathway in the kidney. Confirmatory analysis of kidney gene expression by qPCR in HF vs. control. Individual gene expression was determined by qPCR and expressed as fold change in log base 2 vs. control and graphed alongside fold change derived from RNA-Seq analysis; (B) Representative Western blots showing soluble epoxide hydrolase (SEH) protein (62 kDa) in offspring kidneys at 12 weeks of age. Relative abundance of renal sEH is quantified; (C) Effect of HF on renal 14,15-dihydroxyeicosatrienoic acid level; (D) Effects of DEX on gene expression of arachidonic acid metabolism pathway in the kidney. Confirmatory analysis of kidney gene expression by qPCR in DEX vs. control; (E) Representative Western blots showing sEH protein in offspring kidneys at 16 weeks of age; (F) Effect of DEX on renal 14,15-dihydroxyeicosatrienoic acid level. * p < 0.05 vs. control.

Figure 3

(A) Effects of HF on gene expression of arachidonic acid metabolism pathway in the kidney. Confirmatory analysis of kidney gene expression by qPCR in HF vs. control. Individual gene expression was determined by qPCR and expressed as fold change in log base 2 vs. control and graphed alongside fold change derived from RNA-Seq analysis; (B) Representative Western blots showing soluble epoxide hydrolase (SEH) protein (62 kDa) in offspring kidneys at 12 weeks of age. Relative abundance of renal sEH is quantified; (C) Effect of HF on renal 14,15-dihydroxyeicosatrienoic acid level; (D) Effects of DEX on gene expression of arachidonic acid metabolism pathway in the kidney. Confirmatory analysis of kidney gene expression by qPCR in DEX vs. control; (E) Representative Western blots showing sEH protein in offspring kidneys at 16 weeks of age; (F) Effect of DEX on renal 14,15-dihydroxyeicosatrienoic acid level. * p < 0.05 vs. control.