Shin-ichi Usui

In Vivo klotho Gene Transfer Ameliorates Angiotensin II-Induced Renal Damage

The klotho gene, originally identified by insertional mutagenesis in mice, suppresses the expression of multiple aging-associated phenotypes. This gene is predominantly expressed in the kidney. Recent studies have shown that expression of renal klotho gene is regulated in animal models of metabolic diseases and in humans with chronic renal failure. However, little is known about the mechanisms and the physiological relevance of the regulation of the expression of the klotho gene in the kidney in some diseased conditions. In the present study, we first investigated the role of angiotensin II in the regulation of renal klotho gene expression. Long-term infusion of angiotensin II downregulated renal klotho gene expression at both the mRNA and protein levels. This angiotensin II-induced renal klotho downregulation was an angiotensin type 1 receptor-dependent but pressor-independent event. Adenovirus harboring mouse klotho gene (ad-klotho, 3.3×1010 plaque forming units) was also intravenously administered immediately before starting angiotensin II infusion in some rats. This resulted in a robust induction of Klotho protein in the liver at day 4, which was still detectable 14 days after the gene transfer. Ad-klotho gene transfer, but not ad-lacZ gene transfer, caused an improvement of creatinine clearance, decrease in urinary protein excretion, and amelioration of histologically demonstrated tubulointerstitial damage induced by angiotensin II administration. Our data suggest that downregulation of the renal klotho gene may have an aggravative role in the development of renal damage induced by angiotensin II, and that induction of the klotho gene may have therapeutic possibilities in treating angiotensin II-induced end organ damage.

Iron overload augments angiotensin II-induced cardiac fibrosis and promotes neointima formation.

Background—Abnormal iron deposition may cause oxidant-induced damage in various organs. We have previously reported that continuous administration of angiotensin II to rats results in an overt iron deposition in the renal tubular epithelial cells, which may have a role in angiotensin II–induced renal damage. In the present study, we investigated the role of iron in the development of cardiac injury induced by angiotensin II. Methods and Results—Angiotensin II was continuously infused to rats at a dose of 0.7 mg/kg per day for 7 consecutive days. No iron deposits were observed in the hearts of untreated rats, whereas iron deposition was seen in the cells in the subepicardial and granulation regions after angiotensin II infusion. Concomitant administration of deferoxamine, an iron chelator, significantly reduced the extent of cardiac fibrosis, which suggests that iron deposition aggravates the cardiac fibrosis induced by angiotensin II. Iron overload caused by the administration of iron-dextran resulted in an augmentation of cardiac fibrosis and the generation of neointimal cells in the coronary artery in angiotensin II–infused rats. By contrast, neointima was not formed in the cardiac vessels in norepinephrine-infused rats with iron overload. Conclusions—Cardiac iron deposition may be involved in the development of cardiac fibrosis induced by angiotensin II. In addition, iron overload may enhance the formation of neointima under conditions of increased circulating angiotensin II but not catecholamines.

Abnormal Iron Deposition in Renal Cells in the Rat with Chronic Angiotensin II Administration

Acute experimental iron loading causes iron to accumulate in the renal tissue. The accumulation of iron may play a role in enhancing oxidant-induced tubular injury by producing increased amounts of reactive oxygen species. From findings in cells from heme oxygenase-1 (HO-1)-deficient mice, HO-1 is postulated to prevent abnormal intracellular iron accumulation. Recently, it has been reported that HO-1 is induced in the renal tubular epithelial cells, in which iron is deposited after iron loading, and that this HO-1 induction may be involved in ameliorating iron-induced renal toxicity. We previously showed that chronic administration of angiotensin II to rats induces HO-1 expression in the tubular epithelial cells. These observations led us to investigate whether there is a link between iron deposition and HO-1 induction in renal tubular cells in rats undergoing angiotensin II infusion. In the present study, rats were given angiotensin II for continuously 7 days. Prussian blue staining revealed the distinct deposits of iron in the proximal tubular epithelial cells after angiotensin II administration. Electron microscopy demonstrated that iron particles were present in the lysosomes of these cells. Histologic and immunohistochemical analyses showed that stainable iron and immunoreactive ferritin and HO-1 were colocalized in the tubular epithelial cells. Treatment of angiotensin II-infused rats with an iron chelator, deferoxamine, blocked the abnormal iron deposition in kidneys and also the induced expression of HO-1 and ferritin expression. Furthermore, deferoxamine treatment suppressed the angiotensin II-induced increase in the urinary excretion of protein and N-acetyl-β-d-glucosaminidase, a marker of tubular injury; however, deferoxamine did not affect the angiotensin II-induced decrease in glomerular filtration rate. These results suggest that angiotensin II causes renal injury, in part, by inducing the deposition of iron in the kidney.