Rujun Gong

A novel STAT3 inhibitor, S3I-201, attenuates renal interstitial fibroblast activation and interstitial fibrosis in obstructive nephropathy.

Accumulation of both interstitial myofibroblasts and excessive production of extracellular matrix proteins is a common pathway contributing to chronic kidney disease. In a number of tissues, activation of STAT3 (signal transducer and activator of transcription 3) increases expression of multiple profibrotic genes. Here, we examined the effect of a STAT3 inhibitor, S3I-201, on activation of renal interstitial fibroblasts and progression of renal fibrosis. Treatment of cultured rat renal interstitial fibroblasts with S3I-201 inhibited their activation, as evidenced by dose- and time-dependent blockade of alpha-smooth muscle actin and fibronectin expression. In a mouse model of renal interstitial fibrosis induced by unilateral ureteral obstruction, STAT3 was activated, and administration of S3I-201 attenuated both this activation and extracellular matrix protein deposition following injury. S3I-201 reduced infiltration of the injured kidney by inflammatory cells and suppressed the injury-induced expression of fibronectin, alpha-smooth muscle actin, and collagen type-1 proteins, as well as the expression of multiple cytokines. Furthermore, S3I-201 inhibited proliferation and induced apoptosis preferentially in renal interstitial fibroblasts of the obstructed kidney. Thus, our results suggest that increased STAT3 activity mediates activation of renal interstitial fibroblasts and the progression of renal fibrosis. Inhibition of STAT3 signaling with S3I-201 may hold therapeutic potential for fibrotic kidney diseases.

Hepatocyte Growth Factor Modulates Matrix Metalloproteinases and Plasminogen Activator/Plasmin Proteolytic Pathways in Progressive Renal Interstitial Fibrosis

Evidence suggests that hepatocyte growth factor (HGF) ameliorates renal fibrosis in animal models of chronic renal disease by promoting extracellular matrix catabolism. This study examined the molecular mechanisms of HGF-induced alterations in matrix degradation both in vitro and in vivo. In vitro, HGF increased the collagen catabolizing activity of human proximal tubular epithelial cells (HKC) that were treated with TGF-beta1. Increased collagen catabolism was associated with enhanced activity of both matrix metalloproteinases (MMP) and plasminogen activators (PA)/plasmin proteolytic pathways. HGF abrogated TGF-beta1-induced production of the profibrotic tissue inhibitor of metalloproteinase-2 (TIMP-2) and plasminogen activator inhibitor-1 (PAI-1). In addition, HGF induced the production of MMP-9. In vivo, continuous infusion of HGF in the rat remnant kidney model ameliorated renal fibrosis and tubulointerstitial collagen deposition. This was associated with increased tubular expression of MMP-9, enhanced in situ gelatinolytic activity, partially restored plasmin activity and decreased expression of TIMP-2 and PAI-1 in tubular cells, and upregulation of renal TIMP-3 expression. Conversely, blocking of endogenous HGF by an anti-HGF neutralizing antibody increased renal fibrosis and interstitial collagen. This was accompanied by decreased tubular expression of MMP-9, less in situ proteolytic activity, and elevated expression of TIMP-2 and PAI-1 in tubular cells. Collectively, these findings demonstrate that HGF ameliorates renal fibrosis by enhancing extracellular matrix catabolism via both MMP and the PA/plasmin proteolytic pathways.

Genetic or Pharmacologic Blockade of EGFR Inhibits Renal Fibrosis

Although enhanced activation of the EGF receptor (EGFR) associates with the development and progression of renal fibrosis, the mechanisms linking these observations are not completely understood. Here, after unilateral ureteral obstruction (UUO), wild-type mice exhibited sustained EGFR phosphorylation in the kidney and developed renal fibrosis that was more severe than the renal fibrosis observed in waved-2 mice, which have reduced EGFR tyrosine kinase activity. Waved-2 mice also showed fewer renal tubular cells arrested at G2/M, reduced expression of α-smooth muscle actin (α-SMA), downregulation of multiple genes encoding profibrogenic cytokines, including TGF-β1, and dephosphorylation of Smad3, STAT3, and ERK1/2. Administration of the specific EGFR inhibitor gefitinib recapitulated this phenotype in wild-type mice after UUO. Furthermore, inactivation of either EGFR or STAT3 reduced UUO-induced expression of lipocalin-2, a molecule associated with the pathogenesis of CKD. In cultured renal interstitial fibroblasts, inhibition of EGFR also abrogated TGF-β1- or serum-induced phosphorylation of EGFR, STAT3, ERK1/2, and Smad3 as well as expression of α-SMA and extracelluar matrix proteins. Taken together, these data suggest that EGFR may mediate renal fibrogenesis by promoting transition of renal epithelial cells to a profibrotic phenotype, increased production of inflammatory factors, and activation of renal interstitial fibroblasts. Inhibition of EGFR may have therapeutic potential for fibrotic kidney disease.

Anti-Inflammatory Effect of Hepatocyte Growth Factor in Chronic Kidney Disease: Targeting the Inflamed Vascular Endothelium

Recent studies show that hepatocyte growth factor (HGF) has potent anti-inflammatory effects in multiple animal models of disease in various organ systems, including the kidney, suggesting that HGF may suppress a common proinflammatory process. The aim of this study was to examine the molecular mechanism of HGFs anti-inflammatory actions in a model of chronic kidney disease. Beginning 2 wk after subtotal nephrectomy, rats received a continuous infusion of recombinant HGF, neutralization of endogenous HGF by daily injection of an anti-HGF antibody, or preimmune IgG for an additional 2 wk. The effects on inflammation and injury were examined. HGF administration ameliorated whereas neutralizing endogenous HGF worsened renal inflammation in remnant kidneys. This was accompanied by parallel alterations in endothelial activation and inflammation, marked respectively by de novo E-selectin expression in renal vascular endothelium and leukocyte adhesion to endothelium. In vitro, HGF abrogated monocyte adhesion to TNF-alpha-activated endothelial monolayers and suppressed endothelial expression of E-selectin, which depended on NF-kappaB signaling. In addition, HGF suppressed NF-kappaB reporter gene activity that was induced by TNF-alpha and counteracted TNF-alpha-elicited NF-kappaB interaction with kappaB elements at the E-selectin gene level. Dissection of the NF-kappaB signaling cascade revealed that suppression of NF-kappaB depended on HGFs inhibitory action on NF-kappaB and IkappaB phosphorylation and IkappaB degradation. In vivo, continuous infusion of exogenous HGF markedly diminished sequestration of circulating fluorescence-labeled macrophages in the remnant kidney, mimicking the action of an E-selectin blocking antibody. These findings suggest that HGF has potent and direct anti-inflammatory effects on the basis of suppression of NF-kappaB activation and downstream endothelial inflammation.

Candesartan suppresses chronic renal inflammation by a novel antioxidant action independent of AT1R blockade.

Reactive oxygen species are thought to be critical inducers of renal inflammation and destruction. We examined the effects of candesartan, a highly selective angiotensin II type I receptor (AT1R) blocker, on renal inflammation and oxidative stress. Candesartan suppressed TNF-induced chemokine expression and NFkappaB activation independent of AT1R blockade in cultured renal tubular epithelial cells. This receptor blocker decreased reactive oxygen generation elicited by either TNF or the pro-oxidant hydrogen peroxide and reinstated redox homeostasis, suggesting a direct antioxidant effect. This result was unique to candesartan among several angiotensin II receptor blockers and occurred in cells lacking the AT1R. A dose 5 times the standard therapeutic dose lessened renal inflammation and suppressed tubular NFkappaB activation in spontaneously hypertensive rats. An ultrahigh dose (15 times standard) produced an even greater beneficial effect. Angiotensin II infusion did not cause any hemodynamic changes at either candesartan dose denoting a complete blockade of systemic and renal AT1R. There was, however, a dose-dependent improvement of renal redox homeostasis. Our study suggests that candesartan suppresses redox-sensitive NFkappaB-mediated renal inflammation by a direct antioxidant effect independent of AT1R blockade.

Sustained activation of EGFR triggers renal fibrogenesis after acute kidney injury.

Severe acute kidney injury (AKI) is frequently accompanied by maladaptive repair and renal fibrogenesis; however, the molecular mechanisms that mediate these acute and chronic consequences of AKI remain poorly understood. In this study, we examined the role of epidermal growth factor receptor (EGFR) in these processes using waved-2 (Wa-2) mice, which have reduced EGFR activity, and their wild-type (WT) littermates after renal ischemia. Renal EGFR phosphorylation was induced within 2 days after ischemia, increased over time, and remained elevated at 28 days in WT mice, but this was diminished in Wa-2 mice. At the early stage of postischemia (2 days), Wa-2 mice developed more severe acute renal tubular damage with less reparative responses as indicated by enhanced tubular cell apoptosis, and reduced dedifferentiation and proliferation as compared to WT animals. At the late stage of postischemia (28 days), Wa-2 mice exhibited a less severe renal interstitial fibrosis as shown by reduced activation/proliferation of renal myofibroblasts and decreased deposition of extracellular matrix proteins. EGFR activation also contributed to cell cycle arrest at the G2/M phase, a cellular event associated with production of profibrogenetic factors, in the injured kidney. Collectively, these results indicate that severe AKI results in sustained activation of EGFR, which is required for reparative response of renal tubular cells initially, but eventually leads to fibrogenesis.

Sex Differences in Estrogen Receptor Gene Polymorphism and Its Association with Lupus Nephritis in Chinese

Background/Aims: Lupus nephritis (LN) is a clinical heterogeneous autoimmune disease and genetic factors contribute to the development of LN. One of the most striking characteristics in LN is the high prevalence among childbearing women, as well as that its clinical manifestation differs in women and men, suggesting the role of sex hormones in its pathogenesis. Methods: ThePvuII and XbaI restriction fragment length polymorphism (RFLP) of estrogen receptor (ER) gene were analyzed in 245 biopsy-proven LN patients (58 males and 187 females) and 172 normal controls (101 males and 71 females) by PCR-RFLP. The clinical and pathological features of 49 male and 152 female LN patients with different genotypes were analyzed. Results: It was found that genotype PpXx, ppxx and Ppxx were three major genotypes of ER gene in both of lupus patients and control groups. The distribution of ER gene polymorphism was quite different in lupus patients of different genders. The frequency of the PpXx genotype in male LN patients was significantly higher than both the gender matched normal controls (p < 0.05) and the female LN patients (p < 0.05), while no difference was shown in the frequency of PpXx genotype between female LN patients and gender matched controls. Interestingly, skin rashes and arthritis were found more common in the patients with PpXx genotype. The frequency of hematological abnormalities and hypertension were higher in patients with ppxx genotype (p < 0.05), while capillary thrombi and glomerular sclerosis were more frequently complicated in the patients with ppxx genotype. In addition, the renal vasculitis and interstitial injury were more frequent in those with Ppxx genotype (p < 0.01). Conclusion: The distribution of ER gene polymorphism in LN patients is distinct with different gender. The PpXxgenotype of ER gene may be associated with the susceptibility of SLE in male. ER gene polymorphism is probably one of the genetic factors contributing to the development of clinical heterogeneity and sexually dimorphic manifestations of LN.

The renaissance of corticotropin therapy in proteinuric nephropathies

Refractory nephrotic syndrome continues to be a therapeutic challenge despite advances in immunosuppression and blockade of the renin–angiotensin–aldosterone cascade. Adrenocorticotropic hormone (ACTH), a pituitary neuroimmunoendocrine polypeptide, was widely used in the 1950s as an effective therapy for childhood nephrotic syndrome, but has since been replaced by synthetic glucocorticoid analogues. In addition to controlling steroidogenesis, ACTH also acts as an important physiological agonist of the melanocortin system. Clinical and experimental evidence now suggests that ACTH has antiproteinuric, lipid-lowering and renoprotective properties, which are not fully explained by its steroidogenic effects. ACTH therapy is effective in inducing remission of nephrotic syndrome in patients with a variety of proteinuric nephropathies, even those resistant to steroids and other immunosuppressants. This Perspectives article describes the biophysiology of ACTH, with an emphasis on its melanocortin actions, particularly in renal parenchymal cells, which could potentially explain the therapeutic effects of ACTH in nephrotic glomerulopathies.

Hepatocyte growth factor suppresses acute renal inflammation by inhibition of endothelial E-selectin.

Vascular endothelial activation, marked by de novo expression of E-selectin, is an early and essential event in the process of leukocyte extravasation and inflammation. Evidence suggests that hepatocyte growth factor (HGF) ameliorates inflammation in animal models of renal disease, implying that HGF might inhibit specific components of the inflammatory response. This study examined the effect of HGF on endothelial E-selectin expression in acute inflammation induced by tumor necrosis factor (TNF)-α. In vitro, HGF suppressed TNF-α-induced cell surface expression of E-selectin in human umbilical vein endothelial cells (HUVEC) and inhibited E-selectin mediated monocytic adhesion to endothelial monolayers. HGF activated phosphatidylinositol 3-kinase (PI3K)–Akt that in turn inhibited its downstream transducer glycogen synthase kinase (GSK)3. Blockade of the PI3K–Akt pathway with specific inhibitors abrogated HGF induced inhibitory phosphorylation of GSK3 and suppression of E-selectin. In addition, selective inhibition of GSK3 activity by lithium suppressed TNF-α-induced E-selectin expression and monocytic adhesion, reminiscent of the action of HGF. Moreover, ectopic expression of an uninhibitable mutant GSK3β, in which the regulatory serine-9 is replaced by alanine, abolished HGFs suppressive effect on endothelial E-selectin. In vivo, administration of exogenous HGF reduced endothelial expression of E-selectin induced by bolus injection of TNF-α. This was associated with less sequestration of circulating fluorescence-labeled macrophages in the kidney. These findings suggest that HGF ameliorates acute renal inflammation in part by downregulating E-selectin mediated macrophage adhesion to the inflamed endothelium.

Inhibition of glycogen synthase kinase 3β prevents NSAID- induced acute kidney injury

Clinical use of non-steroidal anti-inflammatory drugs (NSAIDs) like diclofenac (DCLF) is limited by multiple adverse effects, including renal toxicity leading to acute kidney injury. In mice with DCLF-induced nephrotoxicity TDZD-8, a selective glycogen synthase kinase (GSK)3β inhibitor, improved acute kidney dysfunction, ameliorated tubular necrosis and apoptosis associated with induced cortical expression of cyclooxygenase-2 (COX-2) and prostaglandin E2. This renoprotective effect was blunted but still largely preserved in COX-2 null mice, suggesting that other GSK3β targets beyond COX-2 functioned in renal protection. Indeed, TDZD-8 diminished the mitochondrial permeability transition in DCLF-injured kidneys. In vitro, GSK3β inhibition reinstated viability and suppressed necrosis and apoptosis in DCLF-stimulated tubular epithelial cells. DCLF elicited oxidative stress, enhanced the activity of the redox-sensitive GSK3β and promoted a mitochondrial permeability transition by interacting with cyclophilin D, a key component of the mitochondrial permeability transition pore. TDZD-8 blocked GSK3β activity, prevented GSK3β mediated cyclophilin D phosphorylation and the ensuing mitochondrial permeability transition, concomitant with normalization of intracellular ATP. Conversely, ectopic expression of a constitutively active GSK3β abolished the effects of TDZD-8. Hence, inhibition of GSK3β ameliorates NSAID-induced acute kidney injury by induction of renal cortical COX-2 and direct inhibition of the mitochondrial permeability transition.