High-calorie diet exacerbates prostate neoplasia in mice with haploinsufficiency of Pten tumor suppressor gene.

OBJECTIVE: Association between prostate cancer and obesity remains controversial. Allelic deletions of PTEN, a tumor suppressor gene, are common in prostate cancer in men. Monoallelic Pten deletion in mice causes low prostatic intraepithelial neoplasia (mPIN). This study tested the effect of a hypercaloric diet on prostate cancer in Pten (+/-) mice. METHODS: 1-month old mice were fed a high-calorie diet deriving 45% calories from fat for 3 and 6 months before prostate was analyzed histologically and biochemically for mPIN progression. Because Pten (+/-) mice are protected against diet-induced insulin resistance, we tested the role of insulin on cell growth in RWPE-1 normal human prostatic epithelial cells with siRNA knockdown of PTEN. RESULTS: In addition to activating PI3 kinase/Akt and Ras/MAPkinase pathways, high-calorie diet causes neoplastic progression, angiogenesis, inflammation and epithelial-mesenchymal transition. It also elevates the expression of fatty acid synthase (FAS), a lipogenic gene commonly elevated in progressive cancer. SiRNA-mediated downregulation of PTEN demonstrates increased cell growth and motility, and soft agar clonicity in addition to elevation in FAS in response to insulin in RWPE-1 normal human prostatic cells. Downregulating FAS in addition to PTEN, blunted the proliferative effect of insulin (and IL-6) in RWPE-1 cells. CONCLUSION: High-calorie diet promotes prostate cancer progression in the genetically susceptible Pten haploinsufficient mouse while preserving insulin sensitivity. This appears to be partly due to increased inflammatory response to high-caloric intake in addition to increased ability of insulin to promote lipogenesis.

Mitochondrial impairment in the five-sixth nephrectomy model of chronic renal failure: proteomic approach.

BACKGROUND: Kidney injuries provoke considerable adjustment of renal physiology, metabolism, and architecture to nephron loss. Despite remarkable regenerative capacity of the renal tissue, these adaptations often lead to tubular atrophy, interstial and glomerular scaring, and development of chronic kidney disease. The therapeutic strategies for prevention of the transition from acute kidney damage to a chronic condition are limited. The purpose of this study was to elucidate large-scale alterations of the renal cortex proteome in partially nephrecromized rats at an early stage of chronic kidney disease. METHODS: Sprague-Dawley 5/6 nephrectomized rats and sham-operated controls were sacrificed at day 28 post-surgery. To identify proteins with notable alteration of expression we applied a 2D-proteomics approach followed by mass-spectrometry. Altered expression of identified and related proteins was validated by Western blotting and immunohistochemistry. RESULTS: Proteins with increased levels of expression after partial nephrectomy were albumin and vimentin. Proteins with decreased expression were metabolic or mitochondrial. Western blotting analysis showed that the renal cortex of nephrectomized rats expressed decreased amount (by ~50%) of proteins from the inner mitochondrial compartment - the beta-oxidation enzyme MCAD, the structural protein GRP-75, and the oxidative phosphorylation protein COXIV. Mitochondrial DNA copy number was decreased by 30% in the cortex of PNx rats. In contrast, the levels of an outer mitochondrial membrane protein, VDAC1, remained unchanged in remnant kidneys. Mitochondrial biogenesis was not altered after renal mass ablation as was indicated by unchanged levels of PPARγ and PGC1α proteins. Autophagy related protein Beclin 1 was up-regulated in remnant kidneys, however the level of LC3-II protein was unchanged. BNIP3 protein, which can initiate both mitochondrial autophagy and cell death, was up-regulated considerably in kidneys of nephrecomized rats. CONCLUSIONS: The results of the study demonstrated that notable alterations in the renal cortex of 5/6 nephrectomized rats were associated with mitochondrial damage, however mitochondrial biogenesis and autophagy for replacement of damaged mitochondria were not stimulated. Accumulation of dysfunctional mitochondria after 5/6 nephrectomy may cause multiple adjustments in biosynthetic pathways, energy production, ROS signaling, and activation of pro-cell death regulatory pathways thus contributing to the development of chronic kidney disease.

Peroxisome proliferator-activated receptor δ agonist, HPP593, prevents renal necrosis under chronic ischemia.

The Goldblatt's 2 kidney 1 clip (2K1C) rat animal model of renovascular hypertension is characterized by ischemic nephropathy of the clipped kidney. 2K1C rats were treated with a specific peroxisome proliferator-activated receptor δ (PPARδ) agonist, HPP593. Clipped kidneys from untreated rats developed tubular and glomerular necrosis and massive interstitial, periglomerular and perivascular fibrosis. HPP593 kidneys did not exhibit any histochemical features of necrosis; fibrotic lesions were present only in perivascular areas. Necrosis in the untreated clipped kidneys was associated with an increased oxidative stress, up regulation and mitochondrial translocation of the pro-death protein BNIP3 specifically in tubules. In the kidneys of HPP593-treated rats oxidative stress was attenuated and BNIP3 protein decreased notably in the mitochondrial fraction when compared to untreated animals. In untreated clipped kidneys, mitochondria were dysfunctional as revealed by perturbations in the levels of MCAD, COXIV, TFAM, and Parkin proteins and AMPK activation, while in HPP593-treated rats these proteins remained at the physiological levels. Nuclear amounts of oxidative stress-responsive proteins, NRF1 and NRF2 were below physiological levels in treated kidneys. Mitochondrial biogenesis and autophagy were inhibited similarly in both treated and untreated 2K1C kidneys as indicated by a decrease in PGC1-α and deficiency of the autophagy-essential proteins LC3-II and ATG5. However, HPP593 treatment resulted in increased accumulation of p62 protein, an autophagic substrate and an enhancer of NRF2 activity. Therefore, inhibition of BNIP3 activation by the preservation of mitochondrial function and control of oxidative stress by PPARδ is the most likely mechanism to account for the prevention of necrotic death in the kidney under conditions of persistent ischemia.

The cardiotonic steroid hormone marinobufagenin induces renal fibrosis: implication of epithelial-to-mesenchymal transition.

We recently demonstrated that the cardiotonic steroid marinobufagenin (MBG) induced fibrosis in rat hearts through direct stimulation of collagen I secretion by cardiac fibroblasts. This stimulation was also responsible for the cardiac fibrosis seen in experimental renal failure. In this study, the effect of MBG on the development of renal fibrosis in rats was investigated. Four weeks of MBG infusion triggered mild periglomerular and peritubular fibrosis in the cortex and the appearance of fibrotic scars in the corticomedullary junction of the kidney. MBG also significantly increased the protein levels and nuclear localization of the transcription factor Snail in the tubular epithelia. It is known that activation of Snail is associated with epithelial-to-mesenchymal transition (EMT) during renal fibrosis. To examine whether MBG alone can trigger EMT, we used the porcine proximal tubular cell line LLC-PK1. MBG (100 nM) caused LLC-PK1 cells grown to confluence to acquire a fibroblast-like shape and have an invasive motility. The expressions of the mesenchymal proteins collagen I, fibronectin, and vimentin were increased twofold. However, the total level of E-cadherin remained unchanged. These alterations in LLC-PK1 cells in the presence of MBG were accompanied by elevated expression and nuclear translocation of Snail. During the time course of EMT, MBG did not have measurable inhibitory effects on the ion pumping activity of its natural ligand, Na(+)-K(+)-ATPase. Our data suggest that the MBG may be an important factor in inducing EMT and, through this mechanism, elevated levels of MBG in chronic renal failure may play a role in the progressive fibrosis.