Chronic kidney disease with comorbid cardiac dysfunction exacerbates cardiac and renal damage

To address the pathophysiological mechanisms underlying chronic kidney disease with comorbid cardiac dysfunction, we investigated renal and cardiac, functional and structural damage when myocardial infarction (MI) was applied in the setting of kidney injury (induced by 5/6 nephrectomy—STNx). STNx or Sham surgery was induced in male Sprague–Dawley rats with MI or Sham surgery performed 4 weeks later. Rats were maintained for a further 8 weeks. Rats (n = 36) were randomized into four groups: Sham+Sham, Sham+MI, STNx+Sham and STNx+MI. Increased renal tubulointerstitial fibrosis (P < 0.01) and kidney injury molecule‐1 expression (P < 0.01) was observed in STNx+MI compared to STNx+Sham animals, while there were no further reductions in renal function. Heart weight was increased in STNx+MI compared to STNx+Sham or Sham+MI animals (P < 0.05), despite no difference in blood pressure. STNx+MI rats demonstrated greater cardiomyocyte cross‐sectional area and increased cardiac interstitial fibrosis compared to either STNx+Sham (P < 0.01) or Sham+MI (P < 0.01) animals which was accompanied by an increase in diastolic dysfunction. These changes were associated with increases in ANP, cTGF and collagen I gene expression and phospho‐p38 MAPK and phospho‐p44/42 MAPK protein expression in the left ventricle. Addition of MI accelerated STNx‐induced structural damage but failed to significantly exacerbate renal dysfunction. These findings highlight the bidirectional response in this model known to occur in cardiorenal syndrome (CRS) and provide a useful model for examining potential therapies for CRS.     

The effects of mesenchymal stem cell mitochondrial transplantation on doxorubicin‐mediated nephrotoxicity in rats

The effect of dysfunctional mitochondria in several cell pathologies has been reported in renal diseases, including diabetic nephropathy and acute kidney injury. Previous studies have reported that mitochondrial transplantation provided surprising results in myocardial and liver ischemia, as well as in Parkinson's disease. We aimed to investigate the beneficial effects of isolated mitochondria transplantation from mesenchymal stem cells (MSCs) in vivo, to mitigate renal damage that arises from doxorubicin‐mediated nephrotoxicity and its action mechanism. In this study, a kidney model of doxorubicin‐mediated nephrotoxicity was used and isolated mitochondria from MSCs were transferred to the renal cortex of rats. The findings showed that the rate of isolated mitochondria from MSCs maintains sufficient membrane integrity, and was associated with a beneficial renal therapeutic effect. Following doxorubicin‐mediated renal injury, isolated mitochondria or vehicle infused into the renal cortex and rats were monitored for five days. This study found that mitochondrial transplantation decreased cellular oxidative stress and promoted regeneration of tubular cells after renal injury (P < .001, P = .009). Moreover, mitochondrial transplantation reduced protein accumulation of tubular cells and reversed renal deficits (P = .01, P < .001). Mitochondrial transplantation increased Bcl‐2 levels, and caspase‐3 levels decreased in injured renal cells (P < .015, P < .001). Our results provide a direct link between mitochondria dysfunction and doxorubicin‐mediated nephrotoxicity and suggest a therapeutic effect of transferring isolated mitochondria obtained from MSCs against renal injury. To our knowledge, this study is the first study in the literature that showed good therapeutic effects of mitochondrial transplantation in a nephrotoxicity model, which is under‐researched.     

Nodakenin alleviated obstructive nephropathy through blunting Snail1 induced fibrosis

Tubulointerstitial fibrosis plays an important role in end‐stage renal failure, and there are only limited therapeutic options available to preserve organ function. In the present study, we identified that nodakenin, a coumarin isolated from the roots of Angelicae gigas, functions effectively against unilateral ureteral obstruction (UUO)‐induced fibrosis via down‐regulating Snail1 expression. We established UUO‐induced renal fibrosis in mice and then administered with nodakenin orally ata a dose of 1 and 10 mg/kg. The in‐vivo results indicated that nodakenin protected obstructive nephropathy through its anti‐inflammatory and anti‐fibrotic properties. Nodakenin prevented the infiltration of inflammatory cells, alleviated the levels of pro‐inflammatory cytokines, reduced the polarization of macrophages and down‐regulating the aberrant deposition of extracellular matrix at the site of injury. Of note, nodakenin dramatically impeded Smad3, NF‐κB p65 phosphorylation and Snail1 expression. In line with in vivo studies, nodakenin suppressed the expression of Snail1, Smad3 phosphorylation and fibrogenesis in TGF‐β1‐treated renal epithelial cells in‐vitro. Furthermore, we found that the effect of nodaknin against fibrosis was reversed in Snail1 overexpressing cells, whereas nodakenin could not further reduce expression of fibrogenesis in Snail1 silenced cells, suggesting that nodaknein may function through a Snail1‐dependent manner. Collectively, this study reveal a critical role of nodakenin in the cure of renal fibrosis.     

Knockout of receptor for advanced glycation end‐products attenuates age‐related renal lesions

Pro‐aging effects of endogenous advanced glycation end‐products (AGEs) have been reported, and there is increasing interest in the pro‐inflammatory and ‐fibrotic effects of their binding to RAGE (the main AGE receptor). The role of dietary AGEs in aging remains ill‐defined, but the predominantly renal accumulation of dietary carboxymethyllysine (CML) suggests the kidneys may be particularly affected. We studied the impact of RAGE invalidation and a CML‐enriched diet on renal aging. Two‐month‐old male, wild‐type (WT) and RAGE^?/? C57Bl/6 mice were fed a control or a CML‐enriched diet (200 μg CML/g_food) for 18 months. Compared to controls, we observed higher CML levels in the kidneys of both CML WT and CML RAGE^?/? mice, with a predominantly tubular localization. The CML‐rich diet had no significant impact on the studied renal parameters, whereby only a trend to worsening glomerular sclerosis was detected. Irrespective of diet, RAGE^?/? mice were significantly protected against nephrosclerosis lesions (hyalinosis, tubular atrophy, fibrosis and glomerular sclerosis) and renal senile apolipoprotein A‐II (ApoA‐II) amyloidosis (p < 0.001). A positive linear correlation between sclerosis score and ApoA‐II amyloidosis score (r = 0.92) was observed. Compared with old WT mice, old RAGE^?/? mice exhibited lower expression of inflammation markers and activation of AKT, and greater expression of Sod2 and SIRT1. Overall, nephrosclerosis lesions and senile amyloidosis were significantly reduced in RAGE^?/? mice, indicating a protective effect of RAGE deletion with respect to renal aging. This could be due to reduced inflammation and oxidative stress in RAGE^?/? mice, suggesting RAGE is an important receptor in so‐called inflamm‐aging.     

Kidney regeneration through nephron neogenesis in medaka

Although renal regeneration is limited to repair of the proximal tubule in mammals, some bony fish are capable of renal regeneration through nephron neogenesis in the event of renal injury. We previously reported that nephron development in the medaka mesonephros is characterized by four histologically distinct stages, generally referred to as condensed mesenchyme, nephrogenic body, relatively small nephron, and the mature nephron. Developing nephrons are positive for wt1 expression during the first three of these stages. In the present study, we examined the regenerative response to renal injury, artificially induced by the administration of sublethal amounts of gentamicin in adult medaka. Similar to previous reports in other animals, the renal tubular epithelium and the glomerulus of the medaka kidney exhibited severe damage after exposure to this agent. However, kidneys showed substantial recovery after gentamicin administration, and a significant number of developing nephrons appeared 14 days after gentamicin administration (P < 0.01). Similarly, the expression of wt1 in developing nephrons also indicated the early stages of nephrogenesis. These findings show that medaka has the ability to regenerate kidney through nephron neogenesis during adulthood and that wt1 is a suitable marker for detecting nephrogenesis.     

Persistent activation of autophagy in kidney tubular cells promotes renal interstitial fibrosis during unilateral ureteral obstruction

Renal fibrosis is the final, common pathway of end-stage renal disease. Whether and how autophagy contributes to renal fibrosis remains unclear. Here we first detected persistent autophagy in kidney proximal tubules in the renal fibrosis model of unilateral ureteral obstruction (UUO) in mice. UUO-associated fibrosis was suppressed by pharmacological inhibitors of autophagy and also by kidney proximal tubule-specific knockout of autophagy-related 7 (PT-Atg7 KO). Consistently, proliferation and activation of fibroblasts, as indicated by the expression of ACTA2/α-smooth muscle actin and VIM (vimentin), was inhibited in PT-Atg7 KO mice, so was the accumulation of extracellular matrix components including FN1 (fibronectin 1) and collagen fibrils. Tubular atrophy, apoptosis, nephron loss, and interstitial macrophage infiltration were all inhibited in these mice. Moreover, these mice showed a specific suppression of the expression of a profibrotic factor FGF2 (fibroblast growth factor 2). In vitro, TGFB1 (transforming growth factor β 1) induced autophagy, apoptosis, and FN1 accumulation in primary proximal tubular cells. Inhibition of autophagy suppressed FN1 accumulation and apoptosis, while enhancement of autophagy increased TGFB1-induced-cell death. These results suggest that persistent activation of autophagy in kidney proximal tubules promotes renal interstitial fibrosis during UUO. The profibrotic function of autophagy is related to the regulation on tubular cell death, interstitial inflammation, and the production of profibrotic factors.     

Atg5-mediated autophagy deficiency in proximal tubules promotes cell cycle G2/M arrest and renal fibrosis

Macroautophagy/autophagy protects against cellular stress. Renal sublethal injury-triggered tubular epithelial cell cycle arrest at G₂/M is associated with interstitial fibrosis. However, the role of autophagy in renal fibrosis is elusive. Here, we hypothesized that autophagy activity in tubular epithelial cells is pivotal for inhibition of cell cycle G₂/M arrest and subsequent fibrogenic response. In both renal epithelial cells stimulated by angiotensin II (AGT II) and the murine kidney after unilateral ureteral obstruction (UUO), we observed that occurrence of autophagy preceded increased production of COL1 (collagen, type I). Pharmacological enhancement of autophagy by rapamycin suppressed COL1 accumulation and renal fibrosis. In contrast, genetic ablation of autophagy by proximal tubular epithelial cell-specific deletion of Atg5, with reduction of the LC3-II protein level and degradation of SQSTM1/p62, showed marked cell cycle arrest at the G₂/M phase, robust COL1 deposition, and severe interstitial fibrosis in a UUO model, as compared with wild-type mice. In vitro, AGT II exposure triggered autophagy preferentially in the G₁/S phase, and increased COL1 expression in the G₂/M phase in renal epithelial cells. Stimulation of Atg5-deficient primary proximal tubular cells with AGT II also resulted in elevated G₂/M arrest and COL1 production. Pharmacological or genetic inhibition of autophagy increased AGT II-mediated G₂/M arrest. Enhanced expression of ATG5, but not the autophagy-deficient ATG5 mutant K130R, rescued the G₂/M arrest, suggesting the regulation of cell cycle progression by ATG5 is autophagy dependent. In conclusion, Atg5-mediated autophagy in proximal epithelial cells is a critical host-defense mechanism that prevents renal fibrosis by blocking G₂/M arrest.     

Perinatally administered losartan augments renal ACE2 expression but not cardiac or renal Mas receptor in spontaneously hypertensive rats

Since the identification of the alternative angiotensin converting enzyme (ACE)2/Ang‐(1‐7)/Mas receptor axis, renin‐angiotensin system (RAS) is a new complex target for a pharmacological intervention. We investigated the expression of RAS components in the heart and kidney during the development of hypertension and its perinatal treatment with losartan in young spontaneously hypertensive rats (SHR). Expressions of RAS genes were studied by the RT‐PCR in the left ventricle and kidney of rats: normotensive Wistar, untreated SHR, SHR treated with losartan since perinatal period until week 9 of age (20 mg/kg/day) and SHR treated with losartan only until week 4 of age and discontinued until week 9. In the hypertrophied left ventricle of SHR, cardiac expressions of Ace and Mas were decreased while those of AT1 receptor (Agtr1a) and Ace2 were unchanged. Continuous losartan administration reduced LV weight (0.43 ± 0.02; P < 0.05 versus SHR) but did not influence altered cardiac RAS expression. Increased blood pressure in SHR (149 ± 2 in SHR versus 109 ± 2 mmHg in Wistar; P < 0.05) was associated with a lower renal expressions of renin, Agtr1a and Mas and with an increase in ACE2. Continuous losartan administration lowered blood pressure to control levels (105 ± 3 mmHg; P < 0.05 versus SHR), however, only renal renin and ACE2 were significantly up‐regulated (for both P < 0.05 versus SHR). Conclusively, prevention of hypertension and LV hypertrophy development by losartan was unrelated to cardiac or renal expression of Mas. Increased renal Ace2, and its further increase by losartan suggests the influence of locally generated Ang‐(1‐7) in organ response to the developing hypertension in SHRs.     

Acute treatment with relaxin protects the kidney against ischaemia/reperfusion injury

Although recent preclinical and clinical studies have demonstrated that recombinant human relaxin (rhRLX) may have important therapeutic potential in acute heart failure and chronic kidney diseases, the effects of acute rhRLX administration against renal ischaemia/reperfusion (I/R) injury have never been investigated. Using a rat model of 1‐hr bilateral renal artery occlusion followed by 6‐hr reperfusion, we investigated the effects of rhRLX (5 μg/Kg i.v.) given both at the beginning and after 3 hrs of reperfusion. Acute rhRLX administration attenuated the functional renal injury (increase in serum urea and creatinine), glomerular dysfunction (decrease in creatinine clearance) and tubular dysfunction (increase in urinary excretion of N‐acetyl‐β‐glucosaminidase) evoked by renal I/R. These beneficial effects were accompanied by a significant reduction in local lipid peroxidation, free radical‐induced DNA damage and increase in the expression/activity of the endogenous antioxidant enzymes Mn‐ and CuZn‐superoxide dismutases (SOD). Furthermore, rhRLX administration attenuated the increase in leucocyte activation, as suggested by inhibition of myeloperoxidase activity, intercellular‐adhesion‐molecule‐1 expression, interleukin (IL)‐1β, IL‐18 and tumour necrosis factor‐α production as well as increase in IL‐10 production. Interestingly, the reduced oxidative stress status and neutrophil activation here reported were associated with rhRLX‐induced activation of endothelial nitric oxide synthase and up‐regulation of inducible nitric oxide synthase, possibly secondary to activation of Akt and the extracellular signal‐regulated protein kinase (ERK) 1/2, respectively. Thus, we report herein that rhRLX protects the kidney against I/R injury by a mechanism that involves changes in nitric oxide signalling pathway.     

Inactivation of fatty acid amide hydrolase protects against ischemic reperfusion injury-induced renal fibrogenesis

Although cannabinoid receptors (CB) are recognized as targets for renal fibrosis, the roles of endogenous cannabinoid anandamide (AEA) and its primary hydrolytic enzyme, fatty acid amide hydrolase (FAAH), in renal fibrogenesis remain unclear. The present study used a mouse model of post-ischemia-reperfusion renal injury (PIR) to test the hypothesis that FAAH participates in the renal fibrogenesis. Our results demonstrated that PIR showed upregulated expression of FAAH in renal proximal tubules, accompanied with decreased AEA levels in kidneys. Faah knockout mice recovered the reduced AEA levels and ameliorated PIR-triggered increases in blood urea nitrogen, plasma creatinine as well as renal profibrogenic markers and injuries. Correspondingly, a selective FAAH inhibitor, PF-04457845, inhibited the transforming growth factor-beta 1 (TGF-β1)–induced profibrogenic markers in human proximal tubular cell line (HK-2 cells) and mouse primary cultured tubular cells. Knockdown of FAAH by siRNA in HK-2 cells had similar effects as PF-04457845. Tubular cells isolated from Faah^?/? mice further validated the protection against TGF-β1–induced damages. The CB 1 or CB2 receptor antagonist and exogenous FAAH metabolite arachidonic acid failed to reverse the protective effects of FAAH inactivation in HK-2 cells. However, a substrate-selective inhibitor of AEA-cyclooxygenase-2 (COX-2) pathway significantly suppressed the anti-profibrogenic actions of FAAH inhibition. Further, the AEA-COX-2 metabolite, prostamide E2 exerted anti-fibrogenesis effect. These findings suggest that FAAH activation and the consequent reduction of AEA contribute to the renal fibrogenesis, and that FAAH inhibition protects against fibrogenesis in renal cells independently of CB receptors via the AEA-COX-2 pathway by the recovery of reduced AEA.     

Cannabinoid receptor 2 plays a central role in renal tubular mitochondrial dysfunction and kidney ageing

Kidney is one of the most important organs in maintaining the normal life activities. With the high abundance of mitochondria, renal tubular cell plays the vital role in functioning in the reabsorption and secretion of kidney. Reports have shown that mitochondrial dysfunction is of great importance to renal tubular cell senescence and subsequent kidney ageing. However, the underlying mechanisms are not elucidated. Cannabinoid receptor 2 is one of the two receptors responsible for the activation of endocannabinoid system. CB2 is primarily upregulated in renal tubular cells in chronic kidney diseases and mediates fibrogenesis. However, the role of CB2 in tubular mitochondrial dysfunction and kidney ageing has not been clarified. In this study, we found that CB2 was upregulated in kidneys in 24-month-old mice and d -galactose (d -gal)-induced accelerated ageing mice, accompanied by the decrease in mitochondrial mass. Furthermore, gene deletion of CB2 in d -gal-treated mice could greatly inhibit the activation of β-catenin signalling and restore the mitochondrial integrity and Adenosine triphosphate (ATP) production. In CB2 knockout mice, renal tubular cell senescence and kidney fibrosis were also significantly inhibited. CB2 overexpression or activation by the agonist AM1241 could sufficiently induce the decrease in PGC-1α and a variety of mitochondria-related proteins and trigger cellular senescence in cultured human renal proximal tubular cells. CB2-activated mitochondrial dysfunction and cellular senescence could be blocked by ICG-001, a blocker for β-catenin signalling. These results show CB2 plays a central role in renal tubular mitochondrial dysfunction and kidney ageing. The intrinsic mechanism may be related to its activation in β-catenin signalling.     

Curcumin alleviates acute kidney injury in a dry‐heat environment by reducing oxidative stress and inflammation in a rat model

Curcumin exhibits anti‐inflammatory and antioxidant activities. We investigated the protective effects of curcumin in a renal injury rat model under dry‐heat conditions. We divided Sprague‐Dawley rats into four groups: dry‐heat 0‐ (normal temperature control group), 50‐, 100‐, and 150‐minute groups. Each group was divided into five subgroups (n = 10): normal saline (NS), sodium carboxymethylcellulose (CMCNa), and curcumin pretreated low, medium, and high‐dose (50, 100, and 200 mg/kg, respectively) groups. Compared to the normal temperature group, serum creatinine, blood urea nitrogen, urinary kidney injury molecule‐1, and neutrophil gelatinase‐associated load changes in lipoprotein (NGAL) levels were significantly increased in the dry‐heat environment group (P < .05); inducible nitric oxide synthase (iNOS) and cyclooxygenase‐2 (COX‐2) expression and malondialdehyde (MDA) and related inflammatory factor levels were increased in the kidney tissue. Superoxide dismutase (SOD) and catalase (CAT) levels were decreased. However, following all curcumin pretreatment, the serum levels of kidney injury indicators and NGAL were decreased in the urine compared to those in the NS and CMCNa groups (P < .05), whereas renal SOD and CAT activities were increased and MDA was decreased (P < .05). Renal tissues of the 150‐minute group showed obvious pathological changes. Compared to the NS group, pathological changes in the renal tissues of the 100‐ and 200‐mg/kg curcumin groups were significantly reduced. Furthermore, iNOS and COX‐2 expression and inflammatory factor levels were decreased after curcumin treatment. Curcumin exerted renoprotective effects that were likely mediated by its antioxidant and anti‐inflammatory effects in a dry‐heat environment rat model.     

Up-regulation of the human-specific CHRFAM7A gene protects against renal fibrosis in mice with obstructive nephropathy

Renal fibrosis is a major factor in the progression of chronic kidney diseases. Obstructive nephropathy is a common cause of renal fibrosis, which is also accompanied by inflammation. To explore the effect of human-specific CHRFAM7A expression, an inflammation-related gene, on renal fibrosis during obstructive nephropathy, we studied CHRFAM7A transgenic mice and wild type mice that underwent unilateral ureteral obstruction (UUO) injury. Transgenic overexpression of CHRFAM7A gene inhibited UUO-induced renal fibrosis, which was demonstrated by decreased fibrotic gene expression and collagen deposition. Furthermore, kidneys from transgenic mice had reduced TGF-β1 and Smad2/3 expression following UUO compared with those from wild type mice with UUO. In addition, the overexpression of CHRFAM7A decreased release of inflammatory cytokines in the kidneys of UUO-injured mice. In vitro, the overexpression of CHRFAM7A inhibited TGF-β1-induced increase in expression of fibrosis-related genes in human renal tubular epithelial cells (HK-2 cells). Additionally, up-regulated expression of CHRFAM7A in HK-2 cells decreased TGF-β1-induced epithelial-mesenchymal transition (EMT) and inhibited activation f TGF-β1/Smad2/3 signalling pathways. Collectively, our findings demonstrate that overexpression of the human-specific CHRFAM7A gene can reduce UUO-induced renal fibrosis by inhibiting TGF-β1/Smad2/3 signalling pathway to reduce inflammatory reactions and EMT of renal tubular epithelial cells.     

HU210-Induced Downregulation in Cannabinoid CB1 Receptor Binding Strongly Correlates with Body Weight Loss in the Adult Rat

In vitro autoradiography was used to examine changes in cannabinoid CB1 receptors (targeted with [³H] CP55,940) in rats treated with the potent cannabinoid agonist HU210. Animals were administered with HU210 (25, 50, 100 μg/kg) for 4 or 14 days or received a single 100 μg/kg injection of HU210 and sacrificed 24 h later. The acute dose resulted in a decrease in binding in the caudate putamen and hippocampus. A dose dependent, region-specific reduction (P < 0.0001) in [³H] CP55,940 binding was seen in all brain regions examined after 4 and 14 days treatment. A decrease in body weight was recorded during the first 4 days of treatment but after this animals began to gain weight. Correlations (0.865 < r < 0.659, P < 0.0001) between body weight on day four and CB1 receptor binding were found in all brain regions examined suggesting that downregulation of CB1 receptors may contribute to the induction of tolerance to body weight loss induced by HU210.     

Nicotinamide reduces renal interstitial fibrosis by suppressing tubular injury and inflammation

Renal interstitial fibrosis is a common pathological feature in progressive kidney diseases currently lacking effective treatment. Nicotinamide (NAM), a member of water‐soluble vitamin B family, was recently suggested to have a therapeutic potential for acute kidney injury (AKI) in mice and humans. The effect of NAM on chronic kidney pathologies, including renal fibrosis, is unknown. Here we have tested the effects of NAM on renal interstitial fibrosis using in vivo and in vitro models. In vivo, unilateral urethral obstruction (UUO) induced renal interstitial fibrosis as indicated Masson trichrome staining and expression of pro‐fibrotic proteins, which was inhibited by NAM. In UUO, NAM suppressed tubular atrophy and apoptosis. In addition, NAM suppressed UUO‐associated T cell and macrophage infiltration and induction of pro‐inflammatory cytokines, such as TNF‐α and IL‐1β. In cultured mouse proximal tubule cells, NAM blocked TGF–β‐induced expression of fibrotic proteins, while it marginally suppressed the morphological changes induced by TGF‐β. NAM also suppressed the expression of pro‐inflammatory cytokines (eg MCP‐1 and IL‐1β) during TGF‐β treatment of these cells. Collectively, the results demonstrate an anti‐fibrotic effect of NAM in kidneys, which may involve the suppression of tubular injury and inflammation.     

Protective effect of heme oxygenase induction in ethinylestradiol‐induced cholestasis

Estrogen‐induced cholestasis is characterized by impaired hepatic uptake and biliary bile acids secretion because of changes in hepatocyte transporter expression. The induction of heme oxygenase‐1 (HMOX1), the inducible isozyme in heme catabolism, is mediated via the Bach1/Nrf2 pathway, and protects livers from toxic, oxidative and inflammatory insults. However, its role in cholestasis remains unknown. Here, we investigated the effects of HMOX1 induction by heme on ethinylestradiol‐induced cholestasis and possible underlying mechanisms. Wistar rats were given ethinylestradiol (5 mg/kg s.c.) for 5 days. HMOX1 was induced by heme (15 μmol/kg i.p.) 24 hrs prior to ethinylestradiol. Serum cholestatic markers, hepatocyte and renal membrane transporter expression, and biliary and urinary bile acids excretion were quantified. Ethinylestradiol significantly increased cholestatic markers (P ≤ 0.01), decreased biliary bile acid excretion (39%, P = 0.01), down‐regulated hepatocyte transporters (Ntcp/Oatp1b2/Oatp1a4/Mrp2, P ≤ 0.05), and up‐regulated Mrp3 (348%, P ≤ 0.05). Heme pre‐treatment normalized cholestatic markers, increased biliary bile acid excretion (167%, P ≤ 0.05) and up‐regulated hepatocyte transporter expression. Moreover, heme induced Mrp3 expression in control (319%, P ≤ 0.05) and ethinylestradiol‐treated rats (512%, P ≤ 0.05). In primary rat hepatocytes, Nrf2 silencing completely abolished heme‐induced Mrp3 expression. Additionally, heme significantly increased urinary bile acid clearance via up‐regulation (Mrp2/Mrp4) or down‐regulation (Mrp3) of renal transporters (P ≤ 0.05). We conclude that HMOX1 induction by heme increases hepatocyte transporter expression, subsequently stimulating bile flow in cholestasis. Also, heme stimulates hepatic Mrp3 expression via a Nrf2‐dependent mechanism. Bile acids transported by Mrp3 to the plasma are highly cleared into the urine, resulting in normal plasma bile acid levels. Thus, HMOX1 induction may be a potential therapeutic strategy for the treatment of ethinylestradiol‐induced cholestasis.     

Deletion of Smad3 protects against C-reactive protein-induced renal fibrosis and inflammation in obstructive nephropathy

Introduction and Aims: Elevated plasma levels of C-reactive protein (CRP) are closely associated with progressive renal injury in patients with chronic kidney disease (CKD). Here, we tested a hypothesis that CRP may promote renal fibrosis and inflammation via a TGF-β/Smad3-dependent mechanism.Methods: Role and mechanisms of TGF-β/Smad3 in CRP-induced renal fibrosis and inflammation were examined in a mouse model of unilateral ureteral obstruction (UUO) induced in CRP Tg/Smad3 KO mice and in a rat tubular epithelial cell line in which Smad3 gene is stably knocked down (S3KD-NRK52E).Results: We found that mice overexpressing the human CRP gene were largely promoted renal inflammation and fibrosis as evidenced by increasing IL-1β, TNF-α, MCP-1 expression, F4/80⁺ macrophages infiltration, and marked accumulation of α-smooth muscle actin (α-SMA), collagen I and fibronectin in the UUO kidney, which were blunted when Smad3 gene was deleted in CRPtg-Smad3KO. Mechanistically, we found that the protection of renal inflammation and fibrosis in the UUO kidney of CRPtg-Smad3KO mice was associated with the inactivation of CD32-NF-κB and TGF-β/Smad3 signaling.Conclusion: In conclusion, Smad3 deficiency protects against CRP-mediated renal inflammation and fibrosis in the UUO kidney by inactivating CD32-NF-κB and TGF-β/Smad3 signaling.     

Tubular cell-derived exosomal miR-150-5p contributes to renal fibrosis following unilateral ischemia-reperfusion injury by activating fibroblast in vitro and in vivo

Unilateral ischemia reperfusion injury (UIRI) with longer ischemia time is associated with an increased risk of acute renal injury and chronic kidney disease. Exosomes can transport lipid, protein, mRNA, and miRNA to corresponding target cells and mediate intercellular information exchange. In this study, we aimed to investigate whether exosome-derived miRNA mediates epithelial-mesenchymal cell communication relevant to renal fibrosis after UIRI. The secretion of exosomes increased remarkably in the kidney after UIRI and in rat renal tubular epithelium cells (NRK-52E) after hypoxia treatment. The inhibition of exosome secretion by Rab27a knockout or GW4869 treatment ameliorates renal fibrosis following UIRI in vivo. Purified exosomes from NRK-52E cells after hypoxia treatment could activate rat kidney fibroblasts (NRK-49F). The inhibition of exosome secretion in hypoxic NRK-52E cells through Rab27a knockdown or GW4869 treatment abolished NRK-49F cell activation. Interestingly, exosomal miRNA array analysis revealed that miR-150-5p expression was increased after hypoxia compared with the control group. The inhibition of exosomal miR-150-5p abolished the ability of hypoxic NRK-52E cells to promote NRK-49F cell activation in vitro, injections of miR-150-5p enriched exosomes from hypoxic NRK-52E cells aggravated renal fibrosis following UIRI, and renal fibrosis after UIRI was alleviated by miR-150-5p-deficient exosome in vivo. Furthermore, tubular cell-derived exosomal miR-150-5p could negatively regulate the expression of suppressor of cytokine signaling 1 to activate fibroblast. Thus, our results suggest that the blockade of exosomal miR-150-5p mediated tubular epithelial cell-fibroblast communication may provide a novel therapeutic target to prevents UIRI progression to renal fibrosis.