Transiently proliferating perivascular microglia harbor M1 type and precede cerebrovascular changes in a chronic hypertension model.

BACKGROUND: Microglia play crucial roles in the maintenance of brain homeostasis. Activated microglia show a biphasic influence, promoting beneficial repair and causing harmful damage via M2 and M1 microglia, respectively. It is well-known that microglia are initially activated to the M2 state and subsequently switch to the M1 state, called M2-to-M1 class switching in acute ischemic models. However, the activation process of microglia in chronic and sporadic hypertension remains poorly understood. We aimed to clarify the process using a chronic hypertension model, the deoxycorticosterone acetate (DOCA)-salt-treated Wistar rats. METHODS: After unilateral nephrectomy, the rats were randomly divided into DOCA-salt, placebo, and control groups. DOCA-salt rats received a weekly subcutaneous injection of DOCA (40 mg/kg) and were continuously provided with 1% NaCl in drinking water. Placebo rats received a weekly subcutaneous injection of vehicle and were provided with tap water. Control rats received no administration of DOCA or NaCl. To investigate the temporal expression profiles of M1- and M2-specific markers for microglia, the animals were subjected to the immunohistochemical and biochemical studies after 2, 3, or 4 weeks DOCA-salt treatment. RESULTS: Hypertension occurred after 2 weeks of DOCA and salt administration, when round-shaped microglia with slightly shortened processes were observed juxtaposed to the vessels, although the histopathological findings were normal. After 3 weeks of DOCA and salt administration, M1-state perivascular and parenchyma microglia significantly increased, when local histopathological findings began to be observed but cerebrovascular destruction did not occur. On the other hand, M2-state microglia were never observed around the vessels at this period. Interestingly, prior to M1 activation, about 55% of perivascular microglia transiently expressed Ki-67, one of the cell proliferation markers. CONCLUSIONS: We concluded that the resting perivascular microglia directly switched to the pro-inflammatory M1 state via a transient proliferative state in DOCA-salt rats. Our results suggest that the activation machinery of microglia in chronic hypertension differs from acute ischemic models. Proliferative microglia are possible initial key players in the development of hypertension-induced cerebral vessel damage. Fine-tuning of microglia proliferation and activation could constitute an innovative therapeutic strategy to prevent its development.

Up-regulation of NOX1/NADPH oxidase following drug-induced myocardial injury promotes cardiac dysfunction and fibrosis.

Cardiac fibrosis is a common feature in failing heart and therapeutic strategy to halt the progression of fibrosis is highly needed. We here report on NOX1, a non-phagocytic isoform of superoxide-producing NADPH oxidase, which promotes cardiac fibrosis in a drug-induced myocardial injury model. A single-dose administration of doxorubicin (DOX) elicited cardiac dysfunction accompanied by increased production of reactive oxygen species and marked elevation of NOX1 mRNA in the heart. In mice deficient in Nox1 (Nox1-/Y), cardiac functions were well retained and overall survival was significantly improved. However, increased level of serum creatine kinase was equivalent to that of wild-type mice (Nox1+/Y). At 4 days after DOX treatment, severe cardiac fibrosis accompanied by increased hydroxyproline content and activation of matrix metalloproteinase-9 was demonstrated in Nox1+/Y, but it was significantly attenuated in Nox1-/Y. When H9c2 cardiomyocytes were exposed to their homogenate, a dose-dependent increase in NOX1 mRNA was observed. Up-regulation of NOX1 mRNA in H9c2 co-incubated with their homogenate was abolished in the presence of TAK242, a TLR4 inhibitor. When isolated cardiac fibroblasts were exposed to H9c2 homogenates, increased proliferation and up-regulation of collagen 3a1 mRNA were demonstrated. These changes were significantly attenuated in cardiac fibroblasts exposed to homogenates from H9c2 harboring disrupted Nox1. These findings suggest that up-regulation of NOX1 following cellular damage promotes cardiac dysfunction and fibrosis by aggravating the pro-fibrotic response of cardiac fibroblasts. Modulation of the NOX1/NADPH oxidase signaling pathway may be a novel therapeutic strategy for preventing heart failure after myocardial injury.

Pitavastatin suppresses hyperglycaemia-induced podocyte injury via bone morphogenetic protein-7 preservation.

Podocytes form the essential components of the glomerular filtration barrier and play a critical role in diabetic nephropathy. Recent evidence suggests that HMG-CoA reductase inhibitors (statins) exert renoprotective effects. We investigated whether pitavastatin directly suppresses hyperglycaemia-induced podocyte injury using cultured podocytes and, if so, the mechanism of the beneficial effects. Cultured podocytes were exposed to media containing normal (NG; 5 mmol/L) or high (HG; 25 mmol/L) glucose for 1 week. HG increased the lethal injury of podocytes and disruption of F-actin fibers, and reduced the mRNA expression of novel podocyte markers, synaptopodin and Wilms tumor-1 (WT-1), in association with decreased bone morphogenetic protein-7 (BMP-7) expression. Pitavastatin (100 nmol/L) reduced podocyte injury and restored the mRNA expression of synaptopodin and WT1; however, these protective effects were abolished by BMP-7 siRNA. Additionally, pitavastatin suppressed HG-induced Rho kinase activation, as assessed by the phosphorylation level of myosin phosphatase targeting subunit 1 (MYTP1), and C3 exotoxin, a Rho inhibitor, mimicked the effect of pitavastatin on BMP-7 preservation. Pitavastatin attenuates hyperglycaemia-induced podocyte injury via Rho-Rho kinase-dependent BMP-7 preservation.

Pitavastatin Exhibits Protective Effects on Podocytes Accompanied by BMP-7 Up-Regulation and Rho Suppression.

BACKGROUND/AIMS: Podocytes injury is involved in the development of diabetic nephropathy. This study was designed to confirm the reno- and podocyte-protective effects of pitavastatin in diabetic rats and clarify its mechanisms. METHODS: Wistar rats were divided into 4 treatment groups: control, streptozotocin (STZ; 55 mg/kg)-induced diabetes, STZ with pitavastatin (10 mg/kg/day), and STZ with tempol (1 mmol/l). RESULTS: STZ-induced diabetic rats exhibited increases in urinary protein excretion and plasma creatinine, and a decrease in creatinine clearance. Pitavastatin significantly improved these parameters without reducing cholesterol levels, whereas tempol did not. The treatment with STZ-enhanced renal fibrosis, mesangial proliferation, transforming growth factor (TGF)-ß, MCP-1 and suppressed Rho in association with decrement of bone morphogenetic protein (BMP)-7 expression in renal cortex. Moreover, STZ decreased podocyte related factors, podocin and nephrin, and BMP-7 in podocytes. Pitavastatin significantly ameliorated all these indices. On the other hand, improvement by tempol was found only in TGF-ß, MCP-1 and histological changes. CONCLUSION: Pitavastatin exhibited reno- and podocyte-protective effects accompanied by BMP-7 preservation and Rho suppression.

Short-Term Caloric Restriction Suppresses Cardiac Oxidative Stress and Hypertrophy Caused by Chronic Pressure Overload.

BACKGROUND: Caloric restriction (CR) prevents senescent changes, in which reactive oxygen species (ROS) have a critical role. Left ventricular (LV) hypertrophy is a risk factor for cardiovascular diseases. We examined whether CR alters cardiac redox state and hypertrophy from chronic pressure overload. METHODS AND RESULTS: Male c57BL6 mice were subjected to ascending aortic constriction (AAC) with ad libitum caloric intake (AL + AAC group) or 40% restricted caloric intake (CR + AAC group). CR was initiated 2 weeks before AAC and was continued for 4 weeks. Two weeks after constriction, AAC increased LV wall thickness, impaired transmitral flow velocity, and augmented myocyte hypertrophy and fibrosis, in association with enhancement of BNP and collagen III expressions in the AL + AAC group. In the AL + AAC group, oxidative stress in cardiac tissue and mitochondria were enhanced, and NADPH oxidase activity and mitochondrial ROS production were elevated. These changes were significantly attenuated in the CR + AAC group. Additionally, in antioxidant systems, myocardial glutathione peroxidase and superoxide dismutase activities were enhanced in the CR + AAC group. CONCLUSIONS: Chronic pressure overload increased cardiac oxidative damage, in association with cardiac hypertrophy and fibrosis. Short-term CR suppressed oxidative stress and improved cardiac function, suggesting that short-term CR could be a useful strategy to prevent pressure overload-induced cardiac injury.

Telmisartan protects against vascular dysfunction with peroxisome proliferator-activated receptor-γ activation in hypertensive 5/6 nephrectomized rats.

BACKGROUND/AIMS: Telmisartan and losartan, angiotensin II type 1 (AT1) receptor antagonists, are used to manage hypertension. We previously reported that telmisartan, a partial agonist of peroxisome proliferator-activated receptor-γ (PPAR-γ), exhibited stronger vasoprotection than the same dose of losartan in normotensive chronic kidney disease (CKD) rats. We investigated whether telmisartan could inhibit vascular dysfunction in hypertensive CKD rats, via both AT1 receptor blockade and PPAR-γ activation, more effectively than losartan, which decreased blood pressure to a similar extent as telmisartan. METHODS: Two or three branches of the left renal artery were ligated and the right kidney was removed to make hypertensive CKD rats. Telmisartan (5 mg/kg), losartan (10 mg/kg) or telmisartan plus the PPAR-γ antagonist GW9662 was administered. RESULTS: Blood pressure was increased in CKD rats. Telmisartan and losartan decreased blood pressure to the same levels. Impaired endothelium-dependent vasodilation, hyperplasia and decreased phospho-eNOS (Ser(1177)) expression in CKD rat aortas were improved by telmisartan. The aortic infiltration by macrophages and expression of osteopontin were enhanced in CKD rats and suppressed by telmisartan. GW9662 partly canceled the normalization of vascular dysfunction. While losartan attenuated vascular changes, the extent of this attenuation was greater in the telmisartan-treated group. CONCLUSION: Telmisartan exhibited vasoprotection via PPAR-γ agonistic properties in hypertensive CKD rats.