Ameliorative effect of white tea from 50-year-old tree of Camellia sinensis L. (Theaceae) on kidney damage in diabetic mice via SIRT1/AMPK pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Diabetic kidney damage (DKD) is one of the most common complications of diabetes, which is known as a chronic inflammatory kidney disease caused by persistent hyperglycemia. White tea was originally used as a folk medicine to treat measles in ancient China. What arouses our interest is that there is a traditional method to treat diabetes with white tea taken from over 30-year-old tree of Camellia sinensis L. However, there are few reports on the renal protection of white tea. AIM OF THE STUDY: This present study was designed to study the potential protective effects of white tea (WT) and old tree white tea (OTWT) on high-fat-diet (HFD) combined with streptozotocin (STZ)-induced type 2 diabetic mice to explore the possible mechanism of WT/OTWT against DKD. MATERIALS AND METHODS: C57BL/6 mice were randomly divided into four groups: NC, T2D, WT (400 mg/kg·b.w, p.o.), OTWT (400 mg/kg·b.w, p.o.). Diabetes was established in all groups except NC group, by six weeks of HFD feeding combined with STZ (50 mg/kg, i.p.) for 3 times, treatments were administered for six weeks and then all the animals were decapitated; kidney tissues and blood samples were collected for the further analysis, including: levels of insulin, lipid metabolism (TG, TC, HDL, LDL, FFA), antioxidative enzymes (catalase (CAT), super oxide dismutase (SOD), glutathione peroxidase (GPx)), blood urea nitrogen (BUN) and creatine, inflammatory cytokines (TNF-α, IL-1ß, COX-2, iNOS, MCP-1), advanced glycation end products (AGE), receptor of AGE (RAGE), Nrf2, AMPK, SIRT1, and PGC-1α. H&E, PAS and Masson staining were performed to examine the histopathological alterations of the kidneys. RESULTS: Our data showed that WT and OTWT reversed the abnormal serum lipids (TG, TC, HDL, LDL, FFA) in T2D mice, upregulated antioxidative enzymes levels (CAT, SOD, GPx) and inhibit the excessive production of proinflammatory mediators (including MCP-1, TNF-α, IL1ß, COX-2 and iNOS) by varying degrees, and OTWT was more effective. In histopathology, OTWT could significantly alleviate the accumulation of renal AGE in T2D mice, thereby improving the structural changes of the kidneys, such as glomerular hypertrophy, glomerular basement membrane thickening and kidney FIbrosis. CONCLUSIONS: Both WT and OTWT could alleviate the diabetic changes in T2D mice via hypoglycemic, hypolipidemic, anti-oxidative and anti-inflammatory effects, while OTWT was more evident. OTWT could prominently alleviate the accumulation of AGE in the kidneys of T2D mice, thereby ameliorating the renal oxidative stress and inflammatory damage, which was associated with the activation of SIRT1/AMPK pathway.

A different perspective on the filtration barrier after kidney stone formation: An immunohistochemical and biochemical study.

The aim of this study is to investigate whether the filtration barrier is affected by experimental kidney stone formation. Thirty-two rats divided into 4 equally groups (n = 8) at random. Group I control; Group II 1% ethylene glycol; Group III 1% Ethylene glycol + 0.25% Ammonium chloride; Group IV 1% Ethylene glycol + 0.5% Ammonium chloride group. Tissues applied hematoxylin-eosin, periodic-acid-Schiff, Pizzolato's staining. Immunohistochemically stained with integrin α3ß1, type IV collagen, laminin, nephrin, CD2-associated protein (CD2AP) and podocin to show the filtration barrier structure. The TUNEL method was used for apoptosis. The amount of calcium, magnesium, creatinine and uric acid in urine and blood samples, also urine microprotein determined. Stones were formed in all experimental groups. Urine calcium, creatinine, uric acid levels decreased, magnesium levels were not changed. No statistically significant change was observed in blood serum results and TUNEL analysis. Immunohistochemical results showed an increase in nephrin, podocin, CD2AP, laminin and a decrease in integrin α3ß1 and type IV collagen. Consequently, there is an increase in the expression densities of the proteins incorporated in the structure to prevent loss of functionality in the cellular part supporting the structure against a weakening of the basement membrane structure in the glomerular structure in which urine is filtered.

Rivaroxaban, a Direct Factor Xa Inhibitor, Ameliorates Hypertensive Renal Damage Through Inhibition of the Inflammatory Response Mediated by Protease-Activated Receptor Pathway.

Background An enhanced renin-angiotensin system causes hypertensive renal damage. Factor Xa not only functions in the coagulation cascade but also activates intracellular signaling through protease-activated receptors ( PAR ). We investigated the effects of rivaroxaban, a factor Xa inhibitor, on hypertensive renal damage in hypertensive mice overexpressing renin (Ren-TG). Methods and Results The 12- to 16-week-old Ren-TG and wild-type mice were orally administered with or without 6 or 12 mg/kg of rivaroxaban for 1 or 4 months. Plasma factor Xa was significantly increased in the Ren-TG compared with the wild-type mice and was reduced by 12 mg/kg of rivaroxaban ( P<0.05). Urinary albumin excretion (UAE) was higher in the nontreated 8-month-old Ren-TG than in the wild-type mice (69.6±29 versus 20.1±8.2 µg/day; P<0.01). Treatment with 12 mg/kg of rivaroxaban for 4 months decreased the UAE to 38.1±13.2 µg/day ( P<0.01). Moreover, rivaroxaban treatment attenuated histologic changes of glomerular hypertrophy, mesangial matrix expansion, effacement of the podocyte foot process, and thickened glomerular basement membrane in the Ren-TG. The renal expression of PAR -2 was increased in the Ren-TG, but was inhibited with rivaroxaban treatment. In vitro study using the human podocytes showed that the expressions of PAR -2 and inflammatory genes and nuclear factor--κB activation were induced by angiotensin II stimulation, but were inhibited by rivaroxaban. PAR -2 knockdown by small interfering RNA also attenuated the PAR -2-related inflammatory gene expressions. Conclusions These findings indicate that rivaroxaban exerts protective effects against angiotensin II-induced renal damage, partly through inhibition of the PAR -2 signaling-mediated inflammatory response.

Amelioration of Diabetic Nephropathy Using a Retinoic Acid Receptor ß2 Agonist.

Vitamin A (VA) and its derivatives, known as retinoids, play critical roles in renal development through retinoic acid receptor ß2 (RARß2). Disruptions in VA signaling pathways are associated with the onset of diabetic nephropathy (DN). Despite the known role of RARß2 in renal development, the effects of selective agonists for RARß2 in a high-fat diet (HFD) model of DN are unknown. Here we examined whether AC261066 (AC261), a highly selective agonist for RARß2, exhibited therapeutic effects in a HFD model of DN in C57BL/6 mice. Twelve weeks of AC261 administration to HFD-fed mice was well tolerated with no observable side effects. Compared with HFD-fed mice, HFD + AC261-treated mice had improved glycemic control and reductions in proteinuria and urine albumin-to-creatinine ratio. Several cellular hallmarks of DN were mitigated in HFD + AC261-treated mice, including reductions in tubule lipid droplets, podocyte (POD) effacement, endothelial cell collapse, mesangial expansion, and glomerular basement membrane thickening. Mesangial and tubule interstitial expression of the myofibroblast markers α-smooth muscle actin (α-SMA) and type IV collagen (Col-IV) was lower in HFD + AC261-treated mice compared with HFD alone. Ultrastructural and immunohistochemistry analyses showed that, compared with HFD-fed mice, HFD + AC261-treated mice showed preservation of POD foot process and slit-diaphragm morphology, an increase in the levels of slit-diagram protein podocin, and the transcription factor Wilms tumor-suppressor gene 1 in PODs. Given the need for novel DN therapies, our results warrant further studies of the therapeutic properties of AC261 in DN.

Targeted inhibition of Axl receptor tyrosine kinase ameliorates anti-GBM-induced lupus-like nephritis.

Glomerulonephritis (GN) is a typical lesion in autoantibody and immune complex disorders, including SLE. Because the Gas6/Axl pathway has been implicated in the pathogenesis of many types of GN, targeting this pathway might ameliorate GN. Consequently, we have studied the efficacy and mechanism of R428, a potent selective Axl inhibitor, in the prevention of experimental anti-GBM nephritis. Axl upregulation was investigated with Sp1/3 siRNA in the SV40-transformed mesangial cells. For Axl inhibition, a daily dose of R428 (125 mg/kg) or vehicle was administered orally. GN was induced with anti-GBM sera. Renal disease development was followed by serial blood urine nitrogen (BUN) determinations and by evaluation of kidney histology at the time of sacrifice. Axl-associated signaling proteins were analyzed by Western blotting and inflammatory cytokine secretion was analyzed by Proteome array. SiRNA data revealed the transcription factor Sp1 to be an important regulator of mesangial Axl expression. Anti-GBM serum induced severe nephritis with azotemia, protein casts and necrotic cell death. R428 treatment diminished renal Axl expression and improved kidney function, with significantly decreased BUN and glomerular proliferation. R428 treatment inhibited Axl and significantly decreased Akt phosphorylation and renal inflammatory cytokine and chemokine expression; similar effects were observed in anti-GBM antiserum-treated Axl-KO mice. These studies support a role for Axl inhibition in glomerulonephritis.

Actin dynamics at focal adhesions: a common endpoint and putative therapeutic target for proteinuric kidney diseases.

Proteinuria encompasses diverse causes including both genetic diseases and acquired forms such as diabetic and hypertensive nephropathy. The basis of proteinuria is a disturbance in size selectivity of the glomerular filtration barrier, which largely depends on the podocyte: a terminally differentiated epithelial cell type covering the outer surface of the glomerulus. Compromised podocyte structure is one of the earliest signs of glomerular injury. The phenotype of diverse animal models and podocyte cell culture firmly established the essential role of the actin cytoskeleton in maintaining functional podocyte structure. Podocyte foot processes, actin-based membrane extensions, contain 2 molecularly distinct "hubs" that control actin dynamics: a slit diaphragm and focal adhesions. Although loss of foot processes encompasses disassembly of slit diaphragm multiprotein complexes, as long as cells are attached to the glomerular basement membrane, focal adhesions will be the sites in which stress due to filtration flow is counteracted by forces generated by the actin network in foot processes. Numerous studies within last 20 years have identified actin binding and regulatory proteins as well as integrins as essential components of signaling and actin dynamics at focal adhesions in podocytes, suggesting that some of them may become novel, druggable targets for proteinuric kidney diseases. Here we review evidence supporting the idea that current treatments for chronic kidney diseases beneficially and directly target the podocyte actin cytoskeleton associated with focal adhesions and suggest that therapeutic reagents that target the focal adhesion-regulated actin cytoskeleton in foot processes have potential to modernize treatments for chronic kidney diseases.

Tubular basement membrane immune complex deposition is associated with activity and progression of lupus nephritis: a large multicenter Chinese study.

Tubulointerstitial injury is found frequently in lupus nephritis. Immune complex deposits can occur in the tubular basement membranes (TBMs), although its significance in lupus nephritis patients remains unclear. This study assessed the clinical and prognostic features of lupus nephritis patients with TBM deposits in a large Chinese multicenter cohort. Complete data were collected from 195 patients with renal biopsy-proven lupus nephritis diagnosed in the Peking University First Hospital as the discovery cohort. A total of 102 lupus nephritis patients were enrolled from another four centers as the validation cohort. The status of TBM deposits was retrospectively assessed using electron microscopy, and the associations of the deposits with clinical data, pathological characteristics and renal outcomes were further analyzed. The percentage of positive TBM deposits was nearly 30% in the lupus nephritis patients. Using immuno-gold labeling, we found that 10/10 patients were positive for IgG, 7/10 were C3d positive, 6/10 were C1q positive, and 1/10 were C4d positive. Patients with TBM deposits presented with more active features, including a higher SLEDAI score (SLE Disease Activity Index) ( p < 0.001), higher serum creatinine level ( p = 0.001) and lower serum C3 level ( p < 0.001). These patients also presented with higher scores for most renal pathological indices, including the total activity indices score ( p < 0.001) and total chronicity indices score ( p = 0.001). TBM deposits affected renal outcomes in the univariate Cox hazards regression analysis (HR = 4.2, 95% CI = 1.3-14.3, p = 0.02). In conclusion, TBM deposits were common in lupus nephritis patients and correlated closely with the clinical disease activity and renal outcome.

Hyperoside pre-treatment prevents glomerular basement membrane damage in diabetic nephropathy by inhibiting podocyte heparanase expression.

Glomerular basement membrane (GBM) damage plays a pivotal role in pathogenesis of albuminuria in diabetic nephropathy (DN). Heparan sulfate (HS) degradation induced by podocyte heparanase is the major cause of GBM thickening and abnormal perm-selectivity. In the present study, we aimed to examine the prophylactic effect of hyperoside on proteinuria development and GBM damage in DN mouse model and the cultured mouse podocytes. Pre-treatment with hyperoside (30 mg/kg/d) for four weeks could significantly decrease albuminuria, prevent GBM damage and oxidative stress in diabetes mellitus (DM) mice. Immunofluorescence staining, Real time PCR and Western blot analysis showed that decreased HS contents and increased heparanase expression in DN mice were also significantly improved by hyperoside pre-treatment. Meanwhile, transmission electron microscope imaging showed that hyperoside significantly alleviated GBM thickening in DN mice. In addition, hyperoside pre-treatment inhibited the increased heparanase gene (HPR1) promoter activity and heparanase expression induced by high glucose or reactive oxidative species (ROS) in cultured podocytes. Our data suggested that hyperoside has a prophylactic effect on proteinuria development and GBM damage in DM mice by decreasing podocyte heparanase expression.

Rituximab for Treatment of Membranoproliferative Glomerulonephritis and C3 Glomerulopathies.

Membranoproliferative glomerulonephritis (MPGN) is a histological pattern of injury resulting from predominantly subendothelial and mesangial deposition of immunoglobulins or complement factors with subsequent inflammation and proliferation particularly of the glomerular basement membrane. Recent classification of MPGN is based on pathogenesis dividing MPGN into immunoglobulin-associated MPGN and complement-mediated C3 glomerulonephritis (C3GN) and dense deposit disease (DDD). Current guidelines suggest treatment with steroids, cytotoxic agents with or without plasmapheresis only for subjects with progressive disease, that is, nephrotic range proteinuria and decline of renal function. Rituximab, a chimeric B-cell depleting anti-CD20 antibody, has emerged in the last decade as a treatment option for patients with primary glomerular diseases such as minimal change disease, focal-segmental glomerulosclerosis, or idiopathic membranous nephropathy. However, data on the use of rituximab in MPGN, C3GN, and DDD are limited to case reports and retrospective case series. Patients with immunoglobulin-associated and idiopathic MPGN who were treated with rituximab showed partial and complete responses in the majorities of cases. However, rituximab was not effective in few cases of C3GN and DDD. Despite promising results in immunoglobulin-associated and idiopathic MPGN, current evidence on this treatment remains weak, and controlled and prospective data are urgently needed.

Neutrophil Protease Cleavage of Von Willebrand Factor in Glomeruli - An Anti-thrombotic Mechanism in the Kidney.

Adequate cleavage of von Willebrand factor (VWF) prevents formation of thrombi. ADAMTS13 is the main VWF-cleaving protease and its deficiency results in development of thrombotic microangiopathy. Besides ADAMTS13 other proteases may also possess VWF-cleaving activity, but their physiological importance in preventing thrombus formation is unknown. This study investigated if, and which, proteases could cleave VWF in the glomerulus. The content of the glomerular basement membrane (GBM) was studied as a reflection of processes occurring in the subendothelial glomerular space. VWF was incubated with human GBMs and VWF cleavage was assessed by multimer structure analysis, immunoblotting and mass spectrometry. VWF was cleaved into the smallest multimers by the GBM, which contained ADAMTS13 as well as neutrophil proteases, elastase, proteinase 3 (PR3), cathepsin-G and matrix-metalloproteinase 9. The most potent components of the GBM capable of VWF cleavage were in the serine protease or metalloprotease category, but not ADAMTS13. Neutralization of neutrophil serine proteases inhibited GBM-mediated VWF-cleaving activity, demonstrating a marked contribution of elastase and/or PR3. VWF-platelet strings formed on the surface of primary glomerular endothelial cells, in a perfusion system, were cleaved by both elastase and the GBM, a process blocked by elastase inhibitor. Ultramorphological studies of the human kidney demonstrated neutrophils releasing elastase into the GBM. Neutrophil proteases may contribute to VWF cleavage within the subendothelium, adjacent to the GBM, and thus regulate thrombus size. This anti-thrombotic mechanism would protect the normal kidney during inflammation and could also explain why most patients with ADAMTS13 deficiency do not develop severe kidney failure.

Effect of mycophenolic acid in experimental, nontransplant glomerular diseases: new mechanisms beyond immune cells.

Mycophenolic acid (MPA) was introduced into clinical practice as immunosuppressive drug therapy to prevent allograft rejection. Since then, its clinical application has widened. Our goal was to review the lessons learned from experimental nontransplant glomerular disease models on the mechanisms of MPA therapy. T and B lymphocytes are preferentially dependent on de novo purine synthesis. By inhibiting the rate-limiting enzyme of de novo purine synthesis, MPA depletes the pool of deoxyguanosine triphosphate (dGTP) and inhibits proliferation of these immune cells. Furthermore, MPA can also induce apoptosis of immune cells and is known to inhibit synthesis of fucose- and mannose-containing membrane glycoproteins altering the surface expression and binding ability of adhesion molecules. However, MPA exerts a direct effect also on nonimmune cells. Mesangial cells are partially dependent on de novo purine biosynthesis and are thus susceptible to MPA treatment. Additionally, MPA can inhibit apoptosis in podocytes and seems to be beneficial in preserving the expression of nephrin and podocin, and by attenuation of urokinase receptor expression leads to decreased foot-process effacement. In summary, our manuscript sheds light on the molecular mechanisms underlying the antiproteinuric effect of MPA. Overall, MPA is an excellent treatment option in many immunologic glomerulopathies because it possesses immunosuppressive properties, has a remarkable effect on nonimmune cells and counteracts the proliferation of mesangial cells, expansion of mesangial matrix, and foot-process effacement of podocytes combined with a low systemic toxicity.

Neuropilin1 regulates glomerular function and basement membrane composition through pericytes in the mouse kidney.

Neuropilin1 (Nrp1) is a co-receptor best known to regulate the development of endothelial cells and is a target of anticancer therapies. However, its role in other vascular cells including pericytes is emergent. The kidney is an organ with high pericyte density and cancer patients develop severe proteinuria following administration of NRP1B-neutralizing antibody combined with bevacizumab. Therefore, we investigated whether Nrp1 regulates glomerular capillary integrity after completion of renal development using two mouse models; tamoxifen-inducible NG2Cre to delete Nrp1 specifically in pericytes and administration of Nrp1-neutralizing antibodies. Specific Nrp1 deletion in pericytes did not affect pericyte number but mutant mice developed hematuria with glomerular basement membrane defects. Despite foot process effacement, albuminuria was absent and expression of podocyte proteins remained unchanged upon Nrp1 deletion. Additionally, these mice displayed dilation of the afferent arteriole and glomerular capillaries leading to glomerular hyperfiltration. Nidogen-1 mRNA was downregulated and collagen4α3 mRNA was upregulated with no significant effect on the expression of other basement membrane genes in the mutant mice. These features were phenocopied by treating wild-type mice with Nrp1-neutralizing antibodies. Thus, our results reveal a postdevelopmental role of Nrp1 in renal pericytes as an important regulator of glomerular basement membrane integrity. Furthermore, our study offers novel mechanistic insights into renal side effects of Nrp1 targeting cancer therapies.

Restoration of Mimecan Expression by Grape Seed Procyanidin B2 Through Regulation of Nuclear Factor-kappaB in Mice With Diabetic Nephropathy.

Grape seed procyanidin B2 (GSPB2) exerts a variety of potent protective pharmacological effects on diabetic complications. The renal protective effects of GSPB2 and the target protein mimecan regulated by GSPB2, discovered in a previous quantitative proteomic analysis, were assessed in mice with diabetic nephropathy Twenty-four db/db mice were divided into 2 groups of the vehicle-treated and GSPB2-treated (30 mg/kg/d) diabetic groups. All animals were observed for 10 weeks. Treatment with GSPB2 resulted in an improvement in body weight increase and serum levels of triglyceride, total cholesterol, advanced glycation end products, and urinary albumin excretion in comparison with the vehicle-treated diabetic mice (P < .05), although these levels were still higher than those in the control group. Treatment with GSPB2 significantly reduced the extent of glomerular basement membranes thickening, mesangial expansion, and glomerular area as well. Mimecan protein expressions in diabetes mellitus were decreased approximately by 28% when compared with those in the control group (P < .05), and restored remarkably after GSPB2 treatment (P < .05). The expression of nuclear factor-κB (NF-κB) p65 in nuclear extracts, markedly higher in the diabetic mice than in the controls, was significantly suppressed by GSPB2. The findings of this study revealed that mimecan might become a new therapeutic target in the future and indicated that GSPB2 had beneficial effects not only on oxidative stress, but also on renal fibrosis, particularly in the diabetic kidney.

Lipoprotein X Causes Renal Disease in LCAT Deficiency.

Human familial lecithin:cholesterol acyltransferase (LCAT) deficiency (FLD) is characterized by low HDL, accumulation of an abnormal cholesterol-rich multilamellar particle called lipoprotein-X (LpX) in plasma, and renal disease. The aim of our study was to determine if LpX is nephrotoxic and to gain insight into the pathogenesis of FLD renal disease. We administered a synthetic LpX, nearly identical to endogenous LpX in its physical, chemical and biologic characteristics, to wild-type and Lcat-/- mice. Our in vitro and in vivo studies demonstrated an apoA-I and LCAT-dependent pathway for LpX conversion to HDL-like particles, which likely mediates normal plasma clearance of LpX. Plasma clearance of exogenous LpX was markedly delayed in Lcat-/- mice, which have low HDL, but only minimal amounts of endogenous LpX and do not spontaneously develop renal disease. Chronically administered exogenous LpX deposited in all renal glomerular cellular and matrical compartments of Lcat-/- mice, and induced proteinuria and nephrotoxic gene changes, as well as all of the hallmarks of FLD renal disease as assessed by histological, TEM, and SEM analyses. Extensive in vivo EM studies revealed LpX uptake by macropinocytosis into mouse glomerular endothelial cells, podocytes, and mesangial cells and delivery to lysosomes where it was degraded. Endocytosed LpX appeared to be degraded by both human podocyte and mesangial cell lysosomal PLA2 and induced podocyte secretion of pro-inflammatory IL-6 in vitro and renal Cxl10 expression in Lcat-/- mice. In conclusion, LpX is a nephrotoxic particle that in the absence of Lcat induces all of the histological and functional hallmarks of FLD and hence may serve as a biomarker for monitoring recombinant LCAT therapy. In addition, our studies suggest that LpX-induced loss of endothelial barrier function and release of cytokines by renal glomerular cells likely plays a role in the initiation and progression of FLD nephrosis.

Short- and Long-Term Tracking of Anionic Ultrasmall Nanoparticles in Kidney.

While biodistribution of nanoparticles (NPs) has been widely studied at the organ level, relatively little is known about their disposition in organs at the cellular level, especially after long-term exposure. The kidney is regarded as the key organ for the clearance of ultrasmall NPs (<5.5 nm). However, recent studies indicate that NPs in this size range could accumulate in the kidney for extended times without urinary excretion. Using negatively charged quantum dots (QDs) (∼3.7 nm) as a model system, we examined the suborgan disposition of anionic ultrasmall NPs in the kidney at the cellular level after intravenous injection by multiphoton microscopy coupled with fluorescence lifetime imaging. Most of the NPs were initially distributed in the peritubular capillaries or glomerular arterioles after injection, whereas they passed through the fenestrated glomerular endothelium and were gradually taken up by mesangial cells up to 30 days after injection. Only trace amounts of anionic QDs could be detected in the urine, which could be attributed to the barrier of the anionic glomerular basement membrane preventing filtration of anionic QDs. In contrast, cationic QDs of similar size (∼5.67 nm) were found to be readily excreted into urine. This study thus highlights the importance of surface charge in determining renal clearance of ultrasmall NPs. It provides a framework for characterizing and predicting the subcellular disposition in organs and long-term targeting of other NPs, with a physiologically based kinetic model being subsequently developed to describe the suborgan kinetics of anionic ultrasmall NPs.

Erythropoietin protects the tubular basement membrane by promoting the bone marrow to release extracellular vesicles containing tPA-targeting miR-144.

Renal fibrosis is an inevitable outcome of chronic kidney disease (CKD). Erythropoietin (EPO) has been recently reported to be able to mitigate renal fibrosis. The mechanism underlying the protective effect of EPO, however, remains elusive. In the present study, employing a mouse model of renal tubulointerstitial fibrosis induced by unilateral ureteral obstruction (UUO), we demonstrated that EPO markedly reduced the disruption of the tubular basement membrane (TBM) through attenuating the activation of tissue plasminogen activator (tPA) and matrix metalloproteinase 9 (MMP9), the major matrix proteolytic network in the obstructed kidney. Instead of acting directly on tPA in the kidney, EPO strongly increased the level of circulating microRNA (miR)-144, which was delivered to the injured renal fibroblasts via extracellular vesicles (EVs) to target the tPA 3'-untranslated region and suppress tPA expression. The protective effect of EPO on mouse TBM was inhibited by miR-144 antagomir. Furthermore, in vitro results confirmed that EPO could stimulate bone marrow-derived Sca-1(+)CD44(+)CD11b(-)CD19(-) cells to secrete miR-144-containing EVs, which markedly suppressed tPA expression, as well as metalloproteinase 9 (MMP9) level and activity, in cultured renal fibroblasts. In conclusion, our study provides the first evidence that EPO protects mouse renal TBM through promoting bone marrow cells to generate and secrete miR-144, which, in turn, is efficiently delivered into the mouse kidney via EVs to inhibit the activation of the tPA/MMP9-mediated proteolytic network. This finding thus suggests that EPO, a hormone widely used to treat anemia in CKD, is a potential therapeutic strategy for renal fibrosis.

Efficacy and safety of tacrolimus vs. cyclophosphamide for idiopathic membranous nephropathy: A meta-analysis of Chinese adults.

The efficacy and safety of tacrolimus (TAC) and cyclophosphamide (CTX) in the treatment of idiopathic membranous nephropathy (IMN) were compared in Chinese adult patients using a meta- analysis of the available literatures. Randomized controlled clinical trials (RCTs) of the treatment of primary IMN with TAC or CTX combined with corticosteroids in the English databases PubMed, Embase and Cochrane, as well as Chinese databases, were searched. Qualified studies were subjected to quality assessment and meta-analysis. A total of 8 RCTs, including 359 Chinese patients, were included in the meta-analysis. The complete remission rate and overall remission rate in the TAC treatment group after 6 months of treatment were higher than those in the CTX treatment group. No significant difference in remission rate was found after 12 months of treatment. There was no significant difference in the adverse reaction between the two groups at the 6th or 12th months. TAC-based treatment was associated with a faster response than CTX at the 6th month, but there was no significant difference between the two groups at 12th month in Chinese adults. Further study is needed to evaluate the long-term efficacy and safety of this treatment regimen.

PF-1355, a mechanism-based myeloperoxidase inhibitor, prevents immune complex vasculitis and anti-glomerular basement membrane glomerulonephritis.

Small vessel vasculitis is a life-threatening condition and patients typically present with renal and pulmonary injury. Disease pathogenesis is associated with neutrophil accumulation, activation, and oxidative damage, the latter being driven in large part by myeloperoxidase (MPO), which generates hypochlorous acid among other oxidants. MPO has been associated with vasculitis, disseminated vascular inflammation typically involving pulmonary and renal microvasculature and often resulting in critical consequences. MPO contributes to vascular injury by 1) catabolizing nitric oxide, impairing vasomotor function; 2) causing oxidative damage to lipoproteins and endothelial cells, leading to atherosclerosis; and 3) stimulating formation of neutrophil extracellular traps, resulting in vessel occlusion and thrombosis. Here we report a selective 2-thiouracil mechanism-based MPO inhibitor (PF-1355 [2-(6-(2,5-dimethoxyphenyl)-4-oxo-2-thioxo-3,4-dihydropyrimidin-1(2H)-yl)acetamide) and demonstrate that MPO is a critical mediator of vasculitis in mouse disease models. A pharmacokinetic/pharmacodynamic response model of PF-1355 exposure in relation with MPO activity was derived from mouse peritonitis. The contribution of MPO activity to vasculitis was then examined in an immune complex model of pulmonary disease. Oral administration of PF-1355 reduced plasma MPO activity, vascular edema, neutrophil recruitment, and elevated circulating cytokines. In a model of anti-glomerular basement membrane disease, formerly known as Goodpasture disease, albuminuria and chronic renal dysfunction were completely suppressed by PF-1355 treatment. This study shows that MPO activity is critical in driving immune complex vasculitis and provides confidence in testing the hypothesis that MPO inhibition will provide benefit in treating human vasculitic diseases.

Notoginsenoside R1 ameliorates podocyte adhesion under diabetic condition through α3ß1 integrin upregulation in vitro and in vivo.

BACKGROUND: Decreased expression of α3ß1 integrin may contribute to reduction in podocyte adhesion to glomerular basement membrane (GBM), which represents a novel early mechanism leading to diabetic kidney disease (DKD). Here, we examined the protective effects of Notoginsenoside R1 (NR1) on podocyte adhesion and α3ß1 integrin expression under diabetic condition in vitro and in vivo. METHODS: Conditionally immortalized mouse podocytes were exposed to high glucose (HG) with 10 and 100µg /ml of NR1 for 24 h. Podocyte adhesion, albuminuria, oxidative markers, renal histopathology, podocyte number per glomerular volume, integrin-linked kinase (ILK) activity and α3ß1 integrin expression were measured in vitro and in vivo. RESULTS: HG decreased podocyte adhesive capacity and α3ß1 integrin expression, the main podocyte anchoring dimer to the GBM. However, NR1 ameliorated impaired podocyte adhesive capacity and partially restored α3ß1 integrin protein and mRNA expression. These in vitro observations were confirmed in vivo. In streptozotocin(STZ)-induced diabetic rats, treatment with NR1 (5 and 10 mg· kg(-1)· d(-1)) for 12 weeks partially restored the number of podocytes per glomerular volume and glomerular α3ß1 integrin expression, as well as ameliorated albuminuria, histopathology and oxidative stress. NR1 also inhibited glomerular ILK activity in diabetic rats. CONCLUSION: NR1, a novel antioxidant, ameliorated glucose-induced impaired podocyte adhesive capacity and subsequent podocyte depopulation partly through α3ß1 integrin upregulation. These findings might provide a potential new therapeutic option for the treatment of DKD.

Laminin α2-mediated focal adhesion kinase activation triggers Alport glomerular pathogenesis.

It has been known for some time that laminins containing α1 and α2 chains, which are normally restricted to the mesangial matrix, accumulate in the glomerular basement membranes (GBM) of Alport mice, dogs, and humans. We show that laminins containing the α2 chain, but not those containing the α1 chain activates focal adhesion kinase (FAK) on glomerular podocytes in vitro and in vivo. CD151-null mice, which have weakened podocyte adhesion to the GBM rendering these mice more susceptible to biomechanical strain in the glomerulus, also show progressive accumulation of α2 laminins in the GBM, and podocyte FAK activation. Analysis of glomerular mRNA from both models demonstrates significant induction of MMP-9, MMP-10, MMP-12, MMPs linked to GBM destruction in Alport disease models, as well as the pro-inflammatory cytokine IL-6. SiRNA knockdown of FAK in cultured podocytes significantly reduced expression of MMP-9, MMP-10 and IL-6, but not MMP-12. Treatment of Alport mice with TAE226, a small molecule inhibitor of FAK activation, ameliorated fibrosis and glomerulosclerosis, significantly reduced proteinuria and blood urea nitrogen levels, and partially restored GBM ultrastructure. Glomerular expression of MMP-9, MMP-10 and MMP-12 mRNAs was significantly reduced in TAE226 treated animals. Collectively, this work identifies laminin α2-mediated FAK activation in podocytes as an important early event in Alport glomerular pathogenesis and suggests that FAK inhibitors, if safe formulations can be developed, might be employed as a novel therapeutic approach for treating Alport renal disease in its early stages.