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Abstract Previously, this study demonstrates the critical role of myeloid specific TLR4 in macrophage‐mediated progressive renal injury in anti‐glomerular basement membrane (anti‐GBM) crescentic glomerulonephritis (cGN); however, the underlying mechanism remains largely unknown. In this study, single‐cell RNA sequencing (scRNA‐seq), pseudotime trajectories reconstruction, and motif enrichment analysis are used, and macrophage diversity in anti‐GBM cGN under tight regulation of myeloid‐TLR4 is uncovered. Most significantly, a myeloid‐TLR4 deletion‐induced novel reparative macrophage phenotype (Nr4a1+Ear2+) with significant upregulated anti‐inflammatory and tissue repair‐related signaling is discovered, thereby suppressing the M1 proinflammatory responses in anti‐GBM cGN. This is further demonstrated in vitro that deletion of TLR4 from bone marrow‐derived macrophages (BMDMs) induces the Nr4a1/Ear2‐expressing anti‐inflammatory macrophages while blocking LPS‐stimulated M1 proinflammatory responses. Mechanistically, activation of the Nr4a1/Ear2‐axis is recognized as a key mechanism through which deletion of myeloid‐TLR4 promotes the anti‐inflammatory macrophage differentiation in vivo and in vitro. This is confirmed by specifically silencing macrophage Nr4a1 or Ear2 to reverse the anti‐inflammatory effects on TLR4 deficient BMDMs upon LPS stimulation. In conclusion, the findings decode a previously unidentified role for a myeloid‐TLR4 dependent Nr4a1/Ear2 negative feedback mechanism in macrophage‐mediated progressive renal injury, implying that activation of Nr4a1‐Ear2 axis can be a novel and effective immunotherapy for anti‐GBM cGN.

Abstract Cystic fibrosis transmembrane conductance regulator (CFTR), known as a cAMP-activated Cl− channel, is widely expressed at the apical membrane of epithelial cells in a wide variety of tissues. Of note, despite the abundant expression of CFTR in mammalian kidney, the role of CFTR in kidney disease development is unclear. Here, we report that CFTR expression is downregulated in the UUO (unilateral ureteral obstruction)-induced kidney fibrosis mouse model and human fibrotic kidneys. Dysfunction or downregulation of CFTR in renal epithelial cells leads to alteration of genes involved in Epithelial-Mesenchymal Transition (EMT) and kidney fibrosis. In addition, dysregulation of CFTR activates canonical Wnt/β-catenin signaling pathways, whereas the β-catenin inhibitor reverses the effects of CFTR downregulation on EMT marker. More interestingly, CFTR interacts with Dishevelled 2 (Dvl2), a key component of Wnt signaling, thereby suppressing the activation of β-catenin. Compared to wild type, deltaF508 mice with UUO treatment exhibit significantly higher β-catenin activity with aggregated kidney fibrogenesis, which is reduced by forced overexpression of CFTR. Taken together, our study reveals a novel mechanism by which CFTR regulates Wnt/β-catenin signaling pertinent to progression of kidney fibrosis and indicates a potential treatment target.

To expand the armamentarium of treatment for chronic kidney disease (CKD), we explored the utility of boosting endogenously synthesized N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP), which is augmented by inhibition of the angiotensin converting enzyme. Male BALB/c mice underwent unilateral ureteral ligation (UUO) or sham operation and received exogenously administered Ac-SDKP delivered via a subcutaneous osmotic minipump or Captopril treatment by oral gavage. Seven days after UUO, there were significant reductions in the expression of both collagen 1 and collagen 3 in kidneys treated with Ac-SDKP or Captopril, and there was a trend towards reductions in collagen IV, α-SMA, and MCP-1 versus control. However, no significant attenuation of interstitial injury or macrophage infiltration was observed. These findings are in contrary to observations in other models and underscore the fact that a longer treatment time frame may be required to yield anti-inflammatory effects in BALB/c mice treated with Ac-SDKP compared to untreated mice. Finding an effective treatment regimen for CKD requires fine-tuning of pharmacologic protocols.

ObjectiveIncreasing evidence shows that TGF-β1 is a key mediator in diabetic nephropathy (DN) and induces renal fibrosis positively by Smad3 but negatively by Smad7. However, treatment of DN by blocking the TGF-β/Smad pathway remains limited. The present study investigated the anti-fibrotic effect of a traditional Chinese medicine, Chaihuang-Yishen granule (CHYS), on DN.Research design and methodsProtective role of CHYS in DN was examined in an accelerated type 1 DN induced by streptozotocin in uninephrectomized Wistar rats. CHYS, at a dose of 0.56 g/kg body weight, was administered by a daily gastric gavage for 20 weeks and the therapeutic effect and potential mechanisms of CHYS on diabetic kidney injury were examined.ResultsTreatment with CHYS attenuated diabetic kidney injury by significantly inhibiting 24-h proteinuria and progressive renal fibrosis including glomerulosclerotic index, tubulointerstitial fibrosis index, and upregulation of extracellular matrix (collagen I, IV, and fibronectin), despite the same levels of blood glucose. Further studies revealed that inhibition of renal fibrosis in CHYS-treated diabetic rats was associated with inhibition of TGF-β1/Smad3 signaling as demonstrated by upregulation of Smad7 but downregulation of TGF-β1, TGF-β receptors, activation of Smad3, and expression of miRNA-21.ConclusionsCHYS may be a therapeutic agent for DN. CHYS attenuates DN by blocking TGF-β/Smad3-mediated renal fibrosis.

Smad7 has been shown to negatively regulate fibrosis and inflammation, but its role in angiotensin II (Ang II)-induced hypertensive cardiac remodeling remains unknown. Therefore, the present study investigated the role of Smad7 in hypertensive cardiopathy induced by angiotensin II infusion. Hypertensive cardiac disease was induced in Smad7 gene knockout (KO) and wild-type (WT) mice by subcutaneous infusion of Ang II (1.46 mg/kg/day) for 28 days. Although equal levels of high blood pressure were developed in both Smad7 KO and WT mice, Smad7 KO mice developed more severe cardiac injury as demonstrated by impairing cardiac function including a significant increase in left ventricular (LV) mass (P

Liver fibrosis is a major cause of liver failure, but treatment remains ineffective. In the present study, we investigated the mechanisms and anti-hepatofibrotic activities of asiatic acid (AA) in a rat model of liver fibrosis induced by carbon tetrachloride (CCl(4)) and in vitro in TGF-beta1-stimulated rat hepatic stellate cell line (HSC-T6). Treatment with AA significantly attenuated CCl(4)-induced liver fibrosis and functional impairment in a dosage-dependent manner, including blockade of the activation of HSC as determined by inhibiting de novo alpha smooth muscle actin (a-SMA) and collagen matrix expression, and an increase in ALT and AST (all p

Macrophages are a rich source of macrophage migration inhibitory factor (MIF). It is well established that macrophages and MIF play a pathogenic role in anti-glomerular basement membrane crescentic glomerulonephritis (anti-GBM CGN). However, whether macrophages mediate anti-GBM CGN via MIF-dependent mechanism remains unexplored, which was investigated in this study by specifically deleting MIF from macrophages in MIFf/f−lysM−cre mice. We found that compared to anti-GBM CGN induced in MIFf/f control mice, conditional ablation of MIF in macrophages significantly suppressed anti-GBM CGN by inhibiting glomerular crescent formation and reducing serum creatinine and proteinuria while improving creatine clearance. Mechanistically, selective MIF depletion in macrophages largely inhibited renal macrophage and T cell recruitment, promoted the polarization of macrophage from M1 towards M2 via the CD74/NF-κB/p38MAPK-dependent mechanism. Unexpectedly, selective depletion of macrophage MIF also significantly promoted Treg while inhibiting Th1 and Th17 immune responses. In summary, MIF produced by macrophages plays a pathogenic role in anti-GBM CGN. Targeting macrophage-derived MIF may represent a novel and promising therapeutic approach for the treatment of immune-mediated kidney diseases.

Transforming growth factor β (TGF-β)/Smad3 plays a vital role in hypertensive cardiac fibrosis. The long non-coding RNA (lncRNA) Erbb4-IR is a novel Smad3-dependent lncRNA that mediates kidney fibrosis. However, the role of Erbb4-IR in hypertensive heart disease remains unexplored and was investigated in the present study by ultrasound-microbubble-mediated silencing of cardiac Erbb4-IR in hypertensive mice induced by angiotensin II. We found that chronic angiotensin II infusion induced hypertension and upregulated cardiac Erbb4-IR, which was associated with cardiac dysfunction, including a decrease in left ventricle ejection fraction (LVEF) and LV fractional shortening (LVFS) and an increase in LV mass. Knockdown of cardiac Erbb4-IR by Erbb4-IR short hairpin RNA (shRNA) gene transfer effectively improved the angiotensin II-induced deterioration of cardiac function, although blood pressure was not altered. Furthermore, silencing cardiac Erbb4-IR also inhibited angiotensin II-induced progressive cardiac fibrosis, as evidenced by reduced collagen I and III, alpha-smooth muscle actin (α-SMA), and fibronectin accumulation. Mechanistically, improved hypertensive cardiac injury by specifically silencing cardiac Erbb4-IR was associated with increased myocardial Smad7 and miR-29b, revealing that Erbb4-IR may target Smad7 and miR-29b to mediate angiotensin II-induced hypertensive cardiac fibrosis. In conclusion, Erbb4-IR is pathogenic in angiotensin II (Ang II)-induced cardiac remodeling, and targeting Erbb4-IR may be a novel therapy for hypertensive cardiovascular diseases.

“Cytokine storm” is common in critically ill COVID-19 patients, however, mechanisms remain largely unknown. Here, we reported that overexpression of SARS-CoV-2 N protein in diabetic db/db mice significantly increased tubular death and the release of HMGB1, one of the damage-associated molecular patterns (DAMPs), to trigger M1 proinflammatory macrophage activation and production of IL-6, TNF-α, and MCP-1 via a Mincle-Syk/NF-κB-dependent mechanism. This was further confirmed in vitro that overexpression of SARS-CoV-2 N protein caused the release of HMGB1 from injured tubular cells under high AGE conditions, which resulted in M1 macrophage activation and production of proinflammatory cytokines via a Mincle-Syk/NF-κB-dependent mechanism. This was further evidenced by specifically silencing macrophage Mincle to block HMGB1-induced M1 macrophage activation and production of IL-6, TNF-α, and MCP-1 in vitro. Importantly, we also uncovered that treatment with quercetin largely improved SARS-CoV-2 N protein-induced AKI in db/db mice. Mechanistically, we found that quercetin treatment significantly inhibited the release of a DAMP molecule HMGB1 and inactivated M1 pro-inflammatory macrophage while promoting reparative M2 macrophage responses by suppressing Mincle-Syk/NF-κB signaling in vivo and in vitro. In conclusion, SARS-CoV-2 N protein-induced AKI in db/db mice is associated with Mincle-dependent M1 macrophage activation. Inhibition of this pathway may be a mechanism through which quercetin inhibits COVID-19-associated AKI.

TGF-β stimulated tumor-associated neutrophils (TANs) can exert pro-tumoral functions. Here the authors show that Smad3 activation in TANs is associated with an N2-like polarization state and poor outcome in patients with non-small cell lung carcinoma and that Smad3 targeting reprograms TANs to an antitumor state suppressing tumor growth in preclinical lung cancer models.

Kallistatin is a multiple functional serine protease inhibitor that protects against vascular injury, organ damage and tumor progression. Kallistatin treatment reduces inflammation and fibrosis in the progression of chronic kidney disease (CKD), but the molecular mechanisms underlying this protective process and whether kallistatin plays an endogenous role are incompletely understood. In the present study, we observed that renal kallistatin levels were significantly lower in patients with CKD. It was also positively correlated with estimated glomerular filtration rate (eGFR) and negatively correlated with serum creatinine level. Unilateral ureteral obstruction (UUO) in animals also led to down-regulation of kallistatin protein in the kidney, and depletion of endogenous kallistatin by antibody injection resulted in aggravated renal fibrosis, which was accompanied by enhanced Wnt/β-catenin activation. Conversely, overexpression of kallistatin attenuated renal inflammation, interstitial fibroblast activation and tubular injury in UUO mice. The protective effect of kallistatin was due to the suppression of TGF-β and β-catenin signaling pathways and subsequent inhibition of epithelial-to-mesenchymal transition (EMT) in cultured tubular cells. In addition, kallistatin could inhibit TGF-β-mediated fibroblast activation via modulation of Wnt4/β-catenin signaling pathway. Therefore, endogenous kallistatin protects against renal fibrosis by modulating Wnt/β-catenin-mediated EMT and fibroblast activation. Down-regulation of kallistatin in the progression of renal fibrosis underlies its potential as a valuable clinical biomarker and therapeutic target in CKD. [ABSTRACT FROM AUTHOR]

Unresolved inflammation can lead to tissue fibrosis and impaired organ function. Macrophage–myofibroblast transition (MMT) is one newly identified mechanism by which ongoing chronic inflammation causes progressive fibrosis in different forms of kidney disease. However, the mechanisms underlying MMT are still largely unknown. Here, we discovered a brain-specific homeobox/POU domain protein Pou4f1 (Brn3a) as a specific regulator of MMT. Interestingly, we found that Pou4f1 is highly expressed by macrophages undergoing MMT in sites of fibrosis in human and experimental kidney disease, identified by coexpression of the myofibroblast marker, α-SMA. Unexpectedly, Pou4f1 expression peaked in the early stage in renal fibrogenesis in vivo and during MMT of bone marrow-derived macrophages (BMDMs) in vitro. Mechanistically, chromatin immunoprecipitation (ChIP) assay identified that Pou4f1 is a Smad3 target and the key downstream regulator of MMT, while microarray analysis defined a Pou4f1-dependent fibrogenic gene network for promoting TGF-β1/Smad3-driven MMT in BMDMs at the transcriptional level. More importantly, using two mouse models of progressive renal interstitial fibrosis featuring the MMT process, we demonstrated that adoptive transfer of TGF-β1-stimulated BMDMs restored both MMT and renal fibrosis in macrophage-depleted mice, which was prevented by silencing Pou4f1 in transferred BMDMs. These findings establish a role for Pou4f1 in MMT and renal fibrosis and suggest that Pou4f1 may be a therapeutic target for chronic kidney disease with progressive renal fibrosis. [ABSTRACT FROM AUTHOR]

Transforming growth factor-β/Smad3 signaling plays an important role in diabetic nephropathy, but its underlying working mechanism remains largely unexplored. The current study uncovered the pathogenic role and underlying mechanism of a novel Smad3-dependent long noncoding RNA (lncRNA) (LRNA9884) in type 2 diabetic nephropathy (T2DN). We found that LRNA9884 was significantly upregulated in the diabetic kidney of db/db mice at the age of 8 weeks preceding the onset of microalbuminuria and was associated with the progression of diabetic renal injury. LRNA9884 was induced by advanced glycation end products and tightly regulated by Smad3, and its levels were significantly blunted in db/db mice and cells lacking Smad3. More importantly, kidney-specific silencing of LRNA9884 effectively attenuated diabetic kidney injury in db/db mice, as shown by the reduction of histological injury, albuminuria excretion, and serum creatinine. Mechanistically, we identified that LRNA9884 promoted renal inflammation-driven T2DN by triggering MCP-1 production at the transcriptional level, and its direct binding significantly enhanced the promoter activity of MCP-1. Thus, LRNA9884 is a novel Smad3-dependent lncRNA that is highly expressed in db/db mice associated with T2DN development. Targeting of LRNA9884 effectively blocked MCP-1-dependent renal inflammation, therefore suppressing the progressive diabetic renal injury in db/db mice. This study reveals that LRNA9884 may be a novel and precision therapeutic target for T2DN in the future. [ABSTRACT FROM AUTHOR]

It is well established that Smad3 is a key downstream effector of transforming growth factor-β (TGF-β) signaling in tissue fibrogenesis. We reported here that targetting Smad3 specifically with a Smad3 inhibitor SIS3 is able to prevent or halt the progression of renal fibrosis in a mouse model of unilateral ureteral obstructive nephropathy (UUO).We found that preventive treatment with SIS3 at the time of disease induction largely suppressed progressive renal fibrosis by inhibiting α-smooth muscle actin (α-SMA) + myofibroblast accumulation and extracellular matrix (collagen I (Col.I) and fibronectin (FN)) production. Importantly, we also found that treatment with SIS3 on established mouse model of UUO from day 4 after UUO nephropathy halted the progression of renal fibrosis. Mechanistically, the preventive and therapeutic effects of SIS3 on renal fibrosis were associated with the inactivation of Smad3 signaling and inhibition of TGF-β1 expression in the UUO kidney. In conclusion, results from the present study suggest that targetting Smad3 may be a specific and effective therapy for renal fibrosis. [ABSTRACT FROM AUTHOR]

Angiotensin-converting enzyme inhibitors (ACEi) are widely employed to deter the progression of chronic kidney disease (CKD). Besides controlling hypertension and reduction of intra-glomerular pressure, ACEi appear to have anti-fibrotic effects in the renal cortex. N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP), an endogenous tetrapeptide that is degraded by ACE, has also been shown to ameliorate the pro-fibrotic phenotype displayed in CKD in our recent study. Whether the anti-fibrotic properties of ACEi are mediated by Ac- SDKP has not been fully investigated. To delineate the role of Ac-SDKP in ACE blockade, 12-week-old male BALB/c mice underwent sham operation or unilateral ureteric obstruction (UUO). UUO mice were subjected to: i) vehicle; ii) captopril or iii) captopril in conjunction with S17092, a prolyl oligopeptidase inhibitor. After 7 days, mice were sacrificed and kidneys harvested for analyses. After UUO, there were heightened expressions of collagen I, collagen III, fibronectin and a-SMA associated with significant levels of tubulointerstitial injury on histological examination. Furthermore, p44/42 Mitogen-Activated Protein Kinase (MAPK) and Transforming Growth Factor Beta 1(TGF-ß1) signaling were up-regulated. These were significantly ameliorated by captopril treatment alone but unaffected by co-administration of captopril with S17092. Captopril treatment had resulted in elevated urinary Ac-SDKP levels, an effect which was eliminated by the co-administration with S17092. This study allowed the investigation of the renoprotective property of ACEi in the absence of Ac-SDKP and proved conclusively that Ac-SDKP is the prime anti-fibrotic mediator of captopril, acting via p44/42 MAPK and TGF-ß1 signaling pathways. Future research to expand CKD armamentarium should explore the utility of augmenting Ac-SDKP levels. [ABSTRACT FROM AUTHOR]

Tubular cell necrosis is a key histological feature of acute kidney injury (AKI). Necroptosis is a type of programed necrosis, which is executed by mixed lineage kinase domain-like protein (MLKL) upon its binding to the plasma membrane. Emerging evidence indicates that necroptosis plays a critical role in the development of AKI. However, it is unclearwhether renal tubular cells undergo necroptosis in vivo and how the necroptotic pathway is regulated during AKI. Repulsive guidance molecule (RGM)-b is a member of the RGM family. Our previous study demonstrated that RGMb is highly expressed in kidney tubular epithelial cells, but its biological role in the kidney has not been well characterized. In the present study, we found that RGMb reduced membrane-associated MLKL levels and inhibited necroptosis in cultured cells. During ischemia/reperfusion injury (IRI) or oxalate nephropathy, MLKL was induced to express on the apical membrane of proximal tubular (PT) cells. Specific knockout of Rgmb in tubular cells (Rgmb cKO) increased MLKL expression at the apical membrane of PT cells and induced more tubular cell death and more severe renal dysfunction compared with wild-type mice. Treatment with the necroptosis inhibitor Necrostatin-1 or GSK'963 reduced MLKL expression on the apical membrane of PT cells and ameliorated renal function impairment after IRI in both wild-type and Rgmb cKO mice. Taken together, our results suggest that proximal tubular cell necroptosis plays an important role in AKI, and that RGMb protects against AKI by inhibiting MLKL membrane association and necroptosis in proximal tubular cells. [ABSTRACT FROM AUTHOR]

Angiotensin-converting enzyme-2 (ACE2) is downregulated in hypertensive nephropathy. The present study investigated the mechanisms whereby loss of ACE2 promoted angiotensin II-induced hypertensive nephropathy in ACE2 gene knockout mice. We found that compared with wild-type animals, mice lacking ACE2 developed much more severe hypertensive nephropathy in response to chronic angiotensin II infusion, including higher levels of blood pressure, urinary protein excretion, serum creatinine, and progressive renal fibrosis and inflammation. Mechanistic studies revealed that worsening kidney injury in ACE2 knockout mice was associated with an increase in Smurf2 (Smad-specific E3 ubiquitin protein ligase 2), a decrease in renal Smad7, and marked activation of TGF-β (transforming growth factor β)/Smad3 and NF-κB (nuclear factor κ-light-chain-enhancer of activated B cells) signaling, suggesting that Smurf2-dependent Smad7 ubiquitin degradation may be a key mechanism whereby loss of ACE2 promotes angiotensin II-induced TGF-β/Smad3 and NF-κB-mediated hypertensive nephropathy. This was validated by restoring Smad7 locally in the kidneys of ACE2 knockout mice to block angiotensin II-induced TGF-β/Smad3-mediated renal fibrosis and NF-κB-driven renal inflammation. Moreover, we found that angiotensin II could induce microRNA-21 in the mouse kidney and in cultured mesangial cells via a Smad3-dependent mechanism, which was enhanced by deleting ACE2 but inhibited by overexpressing renal Smad7. In conclusion, loss of ACE2 promotes angiotensin II-induced renal injury by targeting Smad7 for degradation via a Smurf2-dependent mechanism. Overexpression of renal Smad7 protects against hypertensive nephropathy by inactivating angiotensin II-induced TGF-β/Smad3 and NF-κB pathways and by targeting the Smad3-dependent microRNA-21 axis. [ABSTRACT FROM AUTHOR]

CRP (C-reactive protein) is regarded as an inflammatory biomarker in AKI (acute kidney injury), but its exact role in AKI remains unclear. Thus we sought to investigate the role of CRP in AKI. Clinically, elevated serum CRP levels were found to associate closely with increased serum creatinine and urea levels (P < 0.01) in patients with AKI, which then fell after recovery from AKI. To determine the role of CRP in AKI, an ischaemia/reperfusion mouse model of AKI was developed using Tg (transgenic) mice that express human CRP. Compared with the WT (wild-type) mice, CRP Tg mice developed more severe renal injury at 24 h after ischaemia as determined by significantly increased serum creatinine and tubular necrosis. This was associated with an impaired TEC (tubular epithelium cell) regeneration as shown by an over 60% reduction in PCNA+ (proliferating-cell nuclear antigen) and BrdU+ (bromodeoxyuridine) TECs in CRP Tg mice with AKI. In vitro, the addition of CRP to a human TEC line (HK-2) also largely suppressed the proliferation of TECs. The functional role of CRP in AKI was demonstrated further by the blocking of CRP binding to the Fcγ RII (Fcγ receptor II) with a neutralizing anti-CD32 antibody, which restored TEC proliferation and prevented AKI in CRP Tg mice. Moreover, we found that impaired G1/S transition by suppression of the phosphorylation of CDK2 (cyclin-dependent kinase 2) and expression of cyclin E may be a key mechanism by which CRP inhibits TEC regeneration during the AKI repair process. In conclusion, CRP plays a pathogenic role in AKI by inhibiting G1/S-dependent TEC regeneration. The results of the present study suggest that targeting CRP signalling may offer a new therapeutic potential for AKI. [ABSTRACT FROM AUTHOR]

RPS19 (ribosomal protein S19), a component of the 40S small ribosomal subunit, has recently been identified to bind the pro-inflammatory cytokine macrophage MIF (migration inhibitory factor). In vitro experiments identify RPS19 as the first endogenous MIF inhibitor by blocking the binding of MIF to its receptor CD74 and MIF functions on monocyte adherence to endothelial cells. In the present study, we sought to establish whether recombinant RPS19 can exert anti-inflammatory effects in a mouse model of anti-GBM (glomerular basement membrane) GN (glomerulonephritis) in which MIF is known to play an important role. Accelerated anti-GBM GN was induced in C57BL/6J mice by immunization with sheep IgG followed 5 days later by administration of sheep anti-mouse GBM serum. Groups of eight mice were treated once daily by intraperitoneal injection with 6 mg of RPS19/kg of body weight or an irrelevant control protein (human secretoglobin 2A1), or received no treatment, from day 0 until being killed on day 10. Mice that received control or no treatment developed severe crescentic anti-GBM disease on day 10 with increased serum creatinine, declined creatinine clearance and increased proteinuria. These changes were associated with up-regulation of MIF and its receptor CD74 activation of ERK (extracellular-signal-regulated kinase) and NF-?B (nuclear factor ?B) signalling, prominent macrophage and T-cell infiltration, as well as up-regulation of Th1 [T-bet and IFN? (interferon ? )] and Th17 [STAT3 (signal transducer and activator of transcription 3) and IL (interleukin)-17A] as well as IL-1ß and TNFa (tumour necrosis factor a). In contrast, RPS19 treatment largely prevented the development of glomerular crescents and glomerular necrosis, and prevented renal dysfunction and proteinuria (all P<0.001). Of note, RPS19 blocked up-regulation of MIF and CD74 and inactivated ERK and NF-?B signalling, thereby inhibiting macrophage and T-cell infiltration, Th1 and Th17 responses and up-regulation of pro-inflammatory cytokines (all P<0.01). These results demonstrate that RPS19 is a potent anti-inflammatory agent, which appears to work primarily by inhibiting MIF signalling. [ABSTRACT FROM AUTHOR]

Background: IL-17A is a pro-inflammatory cytokine that plays important role in inflammatory disease pathology and tumor microenvironment. The aim of this study is to investigate the effect of IL-17A on the progression of hepatocellular carcinoma (HCC). Methodology and Principal Finding: Expression pattern of IL-17A in clinical HCC samples (n = 43) was determined by immunohistochemistry staining. Transcript levels of MMP2, MMP9 and IL-17A were measured in another 50 pairs (including tumor and related non-tumor tissues) HCC samples. Cell growth, focus formation, cell migration, invasion and western blot assays were used to characterize the functional and signaling mechanisms in IL-17A-treated HCC. Association study was used to identify clinical significance of IL-17A in HCC. Compared with paired non-tumor tissue, higher frequency of IL-17Apositive cells was detected in tumor tissues in HCCs with metastasis, and the frequency of IL-17A-positive cells was also significantly associated with poor prognosis of HCC (P = 0.01). Functional study found that IL-17A could promote HCC cell migration and invasion. Further molecular analysis also showed that IL-17A could upregulate MMP2 and MMP9 expression via NF-kB signaling activation. Conclusions: IL-17A could promote HCC metastasis by the upregulation of MMP2 and MMP9 expression via activating NF-kB signaling pathway. [ABSTRACT FROM AUTHOR]

Aristolochic acid nephropathy (AAN) has become a worldwide disease and is the most severe complication related to the use of traditional Chinese medicine. However, the pathogenic mechanisms of AAN remain unclear and therapies are limited. The present study tested the hypothesis that transforming growth factor (TGF)β/Smad3 may be a key pathway leading to chronic AAN. This was examined in vivo in Smad3 wild-type/knockout (WT/KO) mice and in vitro in tubular epithelial cells with knockdown of Smad2 or Smad3. Results revealed that chronic administration of aristolochic acid (AA) resulted in a severe AAN characterized by progressive renal dysfunction and tubulointerstitial fibrosis including epithelial-mesenchymal transition (EMT) in Smad3 WT mice, but not in Smad3 KO mice, suggesting a critical role for Smad3 in the development of AAN. This was further tested in vitro. We found that AA was able to activate Smad signaling to mediate EMT and renal fibrosis via both TGF-β-dependent and JNK/MAP kinase-dependent mechanisms because blockade of JNK and specific knockdown of Smad3, but not Smad2, were able to attenuate AA-stimulated collagen matrix expression and EMT. In conclusion, TGF-β/Smad3 may be an essential mediator for chronic AAN. Results from this study indicate that specific blockade of the TGF-β/Smad3 signaling pathway may have therapeutic potential for chronic AAN. [ABSTRACT FROM AUTHOR]

Connective tissue growth factor (CTGF) plays a critical role in angiotensin II (Ang II)-mediated hypertensive nephropathy. The present study investigated the mechanisms and specific roles of individual Smads in Ang II-induced CTGF and collagen I expression in tubular epithelial cells with deletion of transforming growth factor (TGF)-beta1, overexpression of Smad7, or knockdown of Smad2 or Smad3. We found that Ang II-induced tubular CTGF and collagen I mRNA and protein expressions were regulated positively by phosphorylated Smad2/3 but negatively by Smad7 because overexpression of Smad7-abolished Ang II-induced Smad2/3 phosphorylation and upregulation of CTGF and collagen I in vitro and in a rat model of remnant kidney disease. Additional studies revealed that, in addition to a late (24-hour) TGF-beta-dependent Smad2/3 activation, Ang II also induced a rapid activation of Smad2/3 at 15 minutes and expression of CTGF and collagen I in tubular epithelial cells lacking the TGF-beta gene, which was blocked by the addition of an Ang II type 1 receptor antagonist (losartan) and inhibitors to extracellular signal-regulated kinase 1/2 (PD98059) and p38 (SB203580) but not by inhibitors to Ang II type 2 receptor (PD123319) or c-Jun N-terminal kinase (SP600125), demonstrating a TGF-beta-independent, Ang II type 1 receptor-mediated extracellular signal-regulated kinase/p38 mitogen-activated protein kinase cross-talk pathway in Ang II-mediated CTGF and collagen I expression. Importantly, the ability of knockdown of Smad3, but not Smad2, to inhibit Ang II-induced CTGF and collagen I expression further revealed an essential role for Smad3 in Ang II-mediated renal fibrosis. In conclusion, Ang II induces tubular CTGF expression and renal fibrosis via the TGF-beta-dependent and -independent Smad3 signaling pathways, suggesting that targeting Smad3 may have therapeutic potential for hypertensive nephropathy. [ABSTRACT FROM AUTHOR]