The incidence and severity of acute kidney injury is increased in patients with diabetes and with aging. However, the mechanisms involved have not been clearly established. The present study examined the effects of aging and diabetes on the severity of renal ischemia-reperfusion (IR) injury in Sprague-Dawley (SD) and type 2 diabetic (T2DN) rats. T2DN rats develop diabetes at 3 mo of age and progressive proteinuria and diabetic nephropathy as they age from 6 to 18 mo. Plasma creatinine levels after bilateral IR were significantly higher (3.4 ± 0.1 mg/dl) in 18-mo-old elderly T2DN rats than in middle-aged (12 mo) T2DN rats with less severe diabetic nephropathy or young (3 mo) and elderly (18 mo) control SD rats (1.5 ± 0.2, 1.8 ± 0.1, and 1.7 ± 0.1 mg/dl, respectively). Elderly T2DN rats exhibited a greater fall in medullary blood flow 2 h following renal IR and a more severe and prolonged decline in glomerular filtration rate than middle-aged T2DN and young or elderly SD rats. The basal expression of the adhesion molecules ICAM-1 and E-selectin and the number of infiltrating immune cells was higher in the kidney of elderly T2DN than age-matched SD rats or young and middle-aged T2DN rats before renal IR. These results indicate that elderly T2DN rats with diabetic nephropathy are more susceptible to renal IR injury than diabetic animals with mild injury or age-matched control animals. This is associated with increased expression of ICAM-1, E-selectin and immune cell infiltration, renal medullary vasocongestion, and more prolonged renal medullary ischemia.

Background: Sirtuin 3 (SIRT3) has a crucial role in the cardiovascular diseases. Our previous study revealed that SIRT3 knockout (SIRT3KO) promoted cardiac pericyte–fibroblast transition. In this study, we investigated the involvement of pericyte and iron in angiotensin II (Ang-II)-mediated renal fibrosis in the SIRT3KO mice. Methods and Results: NG2-DsRed mice and NG2-DsRed-SIRT3 knockout (SIRT3KO) mice were infused with saline or Ang-II (1000 ng/kg/min) for 4 weeks. Renal fibrosis, iron content and reactive oxygen species (ROS) were measured. Masson’s trichrome staining showed that SIRT3KO enhanced Ang-II-induced renal fibrosis. Immunostaining showed that Ang-II treatment increased the number of NG2-DsRed+ cells in the kidney, and SIRT3KO further enhanced NG2-DsRed+ cells. Moreover, SIRT3KO promoted pericyte differentiation into fibroblasts as evidenced by co-staining NG2-DsRed/FSP-1. Furthermore, DsRed/FSP-1+ and DsRed/transforming growth factor-β1 (TGF-β1)+ fibroblasts were elevated by SIRT3KO after Ang-II infusion. Ang-II-induced collagen I and TGF-β1 expression was also enhanced in the SIRT3KO mice. SIRT3KO significantly exacerbated Ang-II-induced iron accumulation. This was accompanied by an increase in acetyl-p53, HO-1 and FPN expression. Further, SIRT3KO sensitized Ang-II-induced upregulation of p47phox and gp91phox together with increased ROS formation in the kidney. Conclusion: Our study suggests that SIRT3 deficiency sensitized Ang-II-induced renal fibrosis by the mechanisms involved in promoting differentiation of pericytes into fibroblasts, exacerbating iron overload and accelerating NADPH oxidase-derived ROS formation.

BACKGROUND: Coronary microvascular dysfunction (CMD) has been shown to contribute to cardiac hypertrophy and heart failure (HF) with preserved ejection fraction. At this point, there are no proven treatments for CMD. METHODS: We have shown that histone acetylation may play a critical role in the regulation of CMD. By using a mouse model that replaces lysine with arginine at residues K98, K117, K161, and K162R of p53 (p53 ^4KR ), preventing acetylation at these sites, we test the hypothesis that acetylation-deficient p53 ^4KR could improve CMD and prevent the progression of hypertensive cardiac hypertrophy and HF. Wild-type and p53 ^4KR mice were subjected to pressure overload by transverse aortic constriction to induce cardiac hypertrophy and HF. RESULTS: Echocardiography measurements revealed improved cardiac function together with a reduction of apoptosis and fibrosis in p53 ^4KR mice. Importantly, myocardial capillary density and coronary flow reserve were significantly improved in p53 ^4KR mice. Moreover, p53 ^4KR upregulated the expression of cardiac glycolytic enzymes and Gluts (glucose transporters), as well as the level of fructose-2,6-biphosphate; increased PFK-1 (phosphofructokinase 1) activity; and attenuated cardiac hypertrophy. These changes were accompanied by increased expression of HIF-1α (hypoxia-inducible factor-1α) and proangiogenic growth factors. Additionally, the levels of SERCA-2 were significantly upregulated in sham p53 ^4KR mice, as well as in p53 ^4KR mice after transverse aortic constriction. In vitro, p53 ^4KR significantly improved endothelial cell glycolytic function and mitochondrial respiration and enhanced endothelial cell proliferation and angiogenesis. Similarly, acetylation-deficient p53 ^4KR significantly improved coronary flow reserve and rescued cardiac dysfunction in SIRT3 (sirtuin 3) knockout mice. CONCLUSIONS: Our data reveal the importance of p53 acetylation in coronary microvascular function, cardiac function, and remodeling and may provide a promising approach to improve hypertension-induced CMD and to prevent the transition of cardiac hypertrophy to HF.

Background: Heart failure with preserved ejection fraction (HFpEF) is characterized by a diastolic dysfunction and is highly prevalent in aged women. Our study showed that ablation of endothelial Sirtuin 3 (SIRT3) led to diastolic dysfunction in male mice. However, the sex-specific role of endothelial SIRT3 deficiency on blood pressure and diastolic function in female mice remains to be investigated. Methods and Results: In this study, we demonstrate that the ablation of endothelial SIRT3 in females elevated blood pressure as compared with control female mice. Diastolic function measurement also showed that the isovolumic relaxation time (IVRT) and myocardial performance index (MPI) were significantly increased, whereas the E’ velocity/A’ velocity (E’/A’) ratio was reduced in the endothelial-specific SIRT3 knockout (SIRT3 ECKO) female mice. To further investigate the regulatory role of endothelial SIRT3 on blood pressure and diastolic dysfunction in metabolic stress, SIRT3 ECKO female mice were fed a normal diet and high-fat diet (HFD) for 20 weeks. The knockout of endothelial SIRT3 resulted in an increased blood pressure in female mice fed with an HFD. Intriguingly, SIRT3 ECKO female mice + HFD exhibited impaired coronary flow reserve (CFR) and more severe diastolic dysfunction as evidenced by an elevated IVRT as compared with control female mice + HFD. In addition, female SIRT3 ECKO mice had higher blood pressure and diastolic dysfunction as compared to male SIRT3 ECKO mice. Moreover, female SIRT3 ECKO mice + HFD had an impaired CFR and diastolic dysfunction as compared to male SIRT3 ECKO mice + HFD. Conclusions: These results implicate a sex-specific role of endothelial SIRT3 in regulating blood pressure and diastolic function in mice. Deficiency of endothelial SIRT3 may be responsible for a diastolic dysfunction in aged female.

Angiotensin II (ANG II) is the key contributor to renal fibrosis and injury. The present study investigated the role of endothelium prolyl hydroxylase 2 (PHD2) in ANG II-mediated renal fibrosis and injury. In vitro, endothelial cells (ECs) were isolated from PHD2^f/f control [wild-type (WT)] mice or PHD2 EC knockout (PHD2^ECKO) mice. In vivo, WT and PHD2^ECKO mice were infused with ANG II (1,000 ng·kg⁻¹·min⁻¹) for 28 days. Renal fibrosis, reactive oxygen species (ROS), and iron contents were measured. Knockout of PHD2 resulted in a significant increase in the expression of hypoxia-inducible factor (HIF)-1α and HIF-2α in ECs. Intriguingly, knockout of PHD2 significantly reduced expression of the ANG II type 1 receptor (AT₁R) in ECs. WT mice infused with ANG II caused increases in renal fibrosis, ROS formation, and iron contents. ANG II treatment led to a downregulation of PHD1 expression and upregulation of HIF-1α and HIF-2α in the renal cortex and medulla. Knockout of PHD2 in EC blunted ANG II-induced downregulation of PHD1 expression. Furthermore, knockout of PHD2 in ECs attenuated ANG II-induced expression of HIF-1α, HIF-2α, transforming growth factor-β1, p47 ^phox, gp91 ^phox, heme oxygenase-1, and ferroportin. This was accompanied by a significant suppression of renal fibrosis, ROS formation, and iron accumulation. In summary, knockout of endothelial PHD2 suppressed the expression of AT₁R in ECs and blunted ANG II-induced downregulation of PHD1 and upregulation of HIF-α in the kidney. Our study, for the first time, demonstrates a necessary role of endothelial PHD2 in ANG II-mediated renal fibrosis and injury.

Endothelial glycolytic metabolism plays an important role in the process of angiogenesis. TP53‐induced glycolysis and apoptosis regulator (TIGAR) is a significant mediator of cellular energy homeostasis. However, the role of TIGAR in endothelial metabolism, angiogenesis, and coronary flow reserve (CFR) has not been studied. The present study investigated whether knockout (KO) of TIGAR improves endothelial glycolytic function and angiogenesis. In vitro, aortic endothelial cells (ECs) from TIGAR KO mice exhibited increased expression of 6‐phosphofructo‐2‐kinase/fructose‐2,6‐bisphosphatase isoform‐3 (PFKFB3) and increased glycolytic function. These were accompanied by increased mitochondrial basal/maximal respiration and ATP production. Furthermore, knockout of TIGAR in ECs enhanced endothelial proliferation, migration, and tube formation. Knockout of TIGAR also significantly increased aortic sprouting ex vivo. In vivo, knockout of TIGAR increased the expression of proangiogenic factor, angiopoietin‐1 (Ang‐1) in mouse hearts. Knockout of TIGAR also significantly increased coronary capillary density with enhanced CFR in these hearts. Furthermore, TIGAR KO mice subjected to pressure overload (PO), a common model to study angiogenesis and cardiac hypertrophy, exhibited elevated expression of Ang‐1, VEGF, and PFKFB3 than that of the wild‐type (WT) mice. WT mice subjected to PO exhibited a significant reduction of coronary capillary density and impaired CFR, but TIGAR KO mice did not. In addition, knockout of TIGAR blunted TAC‐induced cardiac hypertrophy and dysfunction seen in the WT mice. In conclusion, knockout of TIGAR improves endothelial angiogenetic capabilities by enhancing the endothelial glycolytic function, mitochondrial respiration, and proangiogenic signaling, which leads to increased coronary capillary density and vascular function and protects against chronic stress.