SARS-CoV-2 N Protein Induces Acute Kidney Injury via Smad3-Dependent G1 Cell Cycle Arrest Mechanism.

COVID-19 is infected by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and can cause severe multiple organ injury and death. Kidney is one of major target organs of COVID-19 and acute kidney injury (AKI) is common in critically ill COVID-19 patients. However, mechanisms through which COVID-19 causes AKI remain largely unknown and treatment remains unspecific and ineffective. Here, the authors report that normal kidney-specifically overexpressing SARS-CoV-2 N develops AKI, which worsens in mice under ischemic condition. Mechanistically, it is uncovered that SARS-CoV-2 N-induced AKI is Smad3-dependent as SARS-CoV-2 N protein can interact with Smad3 and enhance TGF-ß/Smad3 signaling to cause tubular epithelial cell death and AKI via the G1 cell cycle arrest mechanism. This is further confirmed in Smad3 knockout mice and cells in which deletion of Smad3 protects against SARS-CoV-2 N protein-induced cell death and AKI in vivo and in vitro. Most significantly, it is also found that targeting Smad3 with a Smad3 pharmacological inhibitor is able to inhibit SARS-CoV-2 N-induced AKI. In conclusion, the authors identify that SARS-CoV-2 N protein is a key mediator for AKI and induces AKI via the Smad3-dependent G1 cell cycle arrest mechanism. Targeting Smad3 may represent as a novel therapy for COVID-19-asscoaited AKI.

Relationship between the status of phospholipase A2 receptor and prognosis of idiopathic membranous nephropathy.

AIM: Serum levels of phospholipase A2 receptor antibody (PLA2R; SAb) and glomerular deposits of PLA2R antigen (GAg) have been detected in patients with idiopathic membranous nephropathy (IMN). However, the correlation between these immunologic factors and their associations with the status and prognosis of IMN remain uncertain. METHODS: Fifty-one patients with biopsy-proven IMN diagnosed between March of 2015 and December of 2016 were enrolled in this study. All the patients were followed until March of 2017.We used enzyme-linked immunosorbent assay and immunofluorescence to measure the SAb and GAg, respectively. RESULTS: The positive rate of GAg was significantly higher than SAb in patients with IMN (88.24 vs 66.77%, P = 0.017). Compared with SAb- patients, SAb+ patients had a higher baseline proteinuria (6.21 vs 3.40 g/24 h), lower serum albumin (22.49 ± 6.59 vs 29.09 ± 7.40 g/L) and poorer renal function (88.96 ± 21.17 vs 107.25 ± 20.04 mL/min per 1.73 m2 ), as well as a higher renal IgG4 level (P < 0.05). A comparison of SAb+/GAg+ and SAb-/GAg+ tissues yielded similar results (P < 0.01). Regarding prognosis, SAb- patients had a higher rate of complete remission after immunosuppressive treatment than SAb+ patients (P = 0.042). CONCLUSION: The disease status and prognosis correlated more closely with the SAb than with the GAg in our cohort of patients with IMN. Furthermore, SAb+ patients had more severe clinical symptoms and a worse prognosis, which was probably associated with increased IgG4 deposition.

Petchiether A attenuates obstructive nephropathy by suppressing TGF-ß/Smad3 and NF-κB signalling.

Obstructive nephropathy is the end result of a variety of diseases that block drainage from the kidney(s). Transforming growth factor-ß1 (TGF-ß1)/Smad3-driven renal fibrosis is the common pathogenesis of obstructive nephropathy. In this study, we identified petchiether A (petA), a novel small-molecule meroterpenoid from Ganoderma, as a potential inhibitor of TGF-ß1-induced Smad3 phosphorylation. The obstructive nephropathy was induced by unilateral ureteral obstruction (UUO) in mice. Mice received an intraperitoneal injection of petA/vehicle before and after UUO or sham operation. An in vivo study revealed that petA protected against renal inflammation and fibrosis by reducing the infiltration of macrophages, inhibiting the expression of proinflammatory cytokines (interleukin-1ß and tumour necrosis factor-α) and reducing extracellular matrix deposition (α-smooth muscle actin, collagen I and fibronectin) in the obstructed kidney of UUO mice; these changes were associated with suppression of Smad3 and NF-κB p65 phosphorylation. Petchiether A inhibited Smad3 phosphorylation in vitro and down-regulated the expression of the fibrotic marker collagen I in TGF-ß1-treated renal epithelial cells. Further, we found that petA dose-dependently suppressed Smad3-responsive promoter activity, indicating that petA inhibits gene expression downstream of the TGF-ß/Smad3 signalling pathway. In conclusion, our findings suggest that petA protects against renal inflammation and fibrosis by selectively inhibiting TGF-ß/Smad3 signalling.

The baseline levels and risk factors for high-sensitive C-reactive protein in Chinese healthy population.

BACKGROUND: Recent studies show that C-reactive protein (CRP) is not only a biomarker but also a pathogenic mediator contributing to the development of inflammation and ageing-related diseases. However, serum levels of CRP in the healthy ageing population remained unclear, which was investigated in the present study. METHODS: Serum levels of high sensitive C-reactive protein (hs-CRP), glucose (Glu), triglyceride (TG), cholesterol (CHOL), high-density lipoprotein cholesterol (HDL-c), low-density lipoprotein cholesterol (LDL-c), superoxide dismutase (SOD), serum creatinine (SCr), serum uric acid (SUA) were measured in 6060healthy subjects (3672 male and 2388 female, mean age:45.9 years) who received routine physical examination at Sun Yat-sen Memorial Hospital, Guangzhou, China. RESULTS: In total of 6060 healthy people, serum levels of hs-CRP were significantly increased with ageing (P < 0.05), particularly in those with age over 45-year-old (1.31[0.69-2.75] vs 1.05[0.53-2.16]mg/L, P < 0.001). Interestingly, levels of serum hs-CRP were significantly higher in male than female population (1.24[0.65-2.57] vs 1.07[0.53-2.29]mg/L, P < 0.001). Correlation analysis also revealed that serum levels of hs-CRP positively correlated with age and SUA, but inversely correlated with serum levels of HDL-c and SOD (all P < 0.05). CONCLUSIONS: Baseline levels of serum hs-CRP are increased with ageing and are significantly higher in male than female healthy population. In addition, elevated serum levels of hs-CRP are also associated with increased SUA but decreased HDL-c and SOD. Thus, serum levels of hs-CRP may be an indicator associated with ageing in healthy Chinese population.

Novel lncRNA Erbb4-IR Promotes Diabetic Kidney Injury in db/db Mice by Targeting miR-29b.

Transforming growth factor-ß/Smad signaling plays an important role in diabetic nephropathy. The current study identified a novel Smad3-dependent long noncoding RNA (lncRNA) Erbb4-IR in the development of type 2 diabetic nephropathy (T2DN) in db/db mice. We found that Erbb4-IR was highly expressed in T2DN of db/db mice and specifically induced by advanced glycosylation end products (AGEs) via a Smad3-dependent mechanism. The functional role of Erbb4-IR in T2DN was revealed by kidney-specific silencing of Erbb4-IR to protect against the development of T2DN, such as elevated microalbuminuria, serum creatinine, and progressive renal fibrosis in db/db mice, and to block AGE-induced collagen I and IV expression in mouse mesangial cells (mMCs) and mouse tubular epithelial cells (mTECs). Mechanistically, we identified that the Erbb4-IR-microRNA (miR)-29b axis was a key mechanism of T2DN because Erbb4-IR was able to bind the 3' untranslated region of miR-29b genomic sequence to suppress miR-29b expression at transcriptional level. In contrast, silencing of renal Erbb4-IR increased miR-29b and therefore protected the kidney from progressive renal injury in db/db mice and prevented mTECs and mMCs from AGE-induced loss of miR-29b and fibrotic response in vitro. Collectively, we identify that Erbb4-IR is a Smad3-dependent lncRNA that promotes renal fibrosis in T2DN by suppressing miR-29b. Targeting Erbb4-IR may represent a novel therapy for T2DN.

Smad7 protects against acute kidney injury by rescuing tubular epithelial cells from the G1 cell cycle arrest.

Smad7 plays a protective role in chronic kidney disease; however, its role in acute kidney injury (AKI) remains unexplored. Here, we report that Smad7 protects against AKI by rescuing the G1 cell cycle arrest of tubular epithelial cells (TECs) in ischemia/reperfusion-induced AKI in mice in which Smad7 gene is disrupted or restored locally into the kidney. In Smad7 gene knockout (KO) mice, more severe renal impairment including higher levels of serum creatinine and massive tubular necrosis was developed at 48 h after AKI. In contrast, restored renal Smad7 gene locally into the kidney of Smad7 KO mice protected against AKI by promoting TEC proliferation identified by PCNA+ and BrdU+ cells. Mechanistic studies revealed that worsen AKI in Smad7 KO mice was associated with a marked activation of TGF-ß/Smad3-p21/p27 signaling and a loss of CDK2/cyclin E activities, thereby impairing TEC regeneration at the G1 cell cycle arrest. In contrast, restored Smad7 locally into the kidneys of Smad7 KO mice protected TECs from the G1 cell cycle arrest and promoted TEC G1/S transition via a CDK2/cyclin E-dependent mechanism. In conclusion, Smad7 plays a protective role in AKI. Blockade of TGF-ß/Smad3-p21/p27-induced G1 cell cycle arrest may be a key mechanism by which Smad7 treatment inhibits AKI. Thus, Smad7 may be a novel therapeutic agent for AKI.

C-reactive protein promotes acute kidney injury via Smad3-dependent inhibition of CDK2/cyclin E.

Acute kidney injury (AKI) is exacerbated in C-reactive protein transgenic mice but alleviated in Smad3 knockout mice. Here we used C-reactive protein transgenic/Smad3 wild-type and C-reactive protein transgenic/Smad3 knockout mice to investigate the signaling mechanisms by which C-reactive protein promotes AKI. Serum creatinine was elevated, and the extent of tubular epithelial cell necrosis following ischemia/reperfusion-induced AKI was greater in C-reactive protein transgenics but was blunted when Smad3 was deleted. Exacerbation of AKI in C-reactive protein transgenics was associated with increased TGF-ß/Smad3 signaling and expression of the cyclin kinase inhibitor p27, but decreased phosphorylated CDK2 and expression of cyclin E. Concomitantly, tubular epithelial cell proliferation was arrested at the G1 phase in C-reactive protein transgenics with fewer cells entering the S-phase cell cycle as evidenced by fewer bromodeoxyuridine-positive cells. In contrast, the protection from AKI in C-reactive protein transgenic/Smad3 knockout mice was associated with decreased expression of p27 and promotion of CDK2/cyclin E-dependent G1/S transition of tubular epithelial cells. In vitro studies using tubular epithelial cells showed that C-reactive protein activates Smad3 via both TGF-ß-dependent and ERK/MAPK cross talk mechanisms, Smad3 bound directly to p27, and blockade of Smad3 or the Fc receptor CD32 prevented C-reactive protein-induced p27-dependent G1 cell cycle arrest. In vivo, treatment of C-reactive protein transgenics with a Smad3 inhibitor largely improved AKI outcomes. Thus, C-reactive protein may promote AKI by impairing tubular epithelial cell regeneration via the CD32-Smad3-p27-driven inhibition of the CDK2/cyclin E complex. Targeting Smad3 may offer a new treatment approach for AKI.

C-Reactive Protein Promotes Diabetic Kidney Disease in db/db Mice via the CD32b-Smad3-mTOR signaling Pathway.

C-reactive protein (CRP) is associated with progressive diabetic nephropathy in patients with type-2 diabetes (T2DN). However, role of CRP in T2DN remains unclear. We report here that CRP is pathogenic in T2DN in db/db mice that express human CRP (CRPtg-db/db). Compared to the littermate db/db mice, CRPtg-db/db developed more severe T2DN, showing higher levels of fasting blood glucose and microalbuminuria and more progressive renal inflammation and fibrosis. Enhanced T2DN in CRPtg-db/db mice were associated with over-activation of CRP-CD32b, NF-κB, TGF-ß/Smad3, and mTOR signaling. Further studies in vitro defined that CRP activated Smad3 directly at 15 mins via the CD32b- ERK/p38 MAP kinase crosstalk pathway and indirectly at 24 hours through a TGF-ß1-dependent mechanism. Importantly, CRP also activated mTOR signaling at 30 mins via a Smad3-dependent mechanism as Smad3 bound mTOR physically and CRP-induced mTOR signaling was abolished by a neutralizing CD32b antibody and a specific Smad3 inhibitor. Finally, we also found that CRP induced renal fibrosis through a CD32b-Smad3-mTOR pathway because blocking mTOR signaling with rapamycin inhibited CRP-induced CTGF and collagen I expression. Thus, CRP is pathogenic in T2DN. CRP may promote CD32b- NF-κB signaling to mediate renal inflammation; whereas, CRP may enhance renal fibrosis in T2DN via CD32b-Smad3-mTOR signaling.

miRNA-29b improves bone healing in mouse fracture model.

A number of miRNAs regulates bone remodeling and their levels in circulation were associated with bone fracture, however no miRNAs have yet been shown to improve fracture healing directly. This study aimed to investigate the effect of miR-29b-3p on mice femoral fracture healing through site-specific delivery with microbubble-ultrasound system. miR-29b-3p promoted osteogenesis of mouse bone marrow-derived mesenchymal stem cells as indicated with quantitative real-time polymerase chain reaction (qPCR) and Alizarin red S staining. Animal study showed that single injection of miR-29b-3p at week 2 post fracture improved healing outcome as indicated by significant decrease of callus width and area with radiographic analysis without causing significant weight loss. Static bone histomorphometry analysis showed that miR-29b-3p increased bone volume fraction (BV/TV), and micro-computed tomography (micro-CT) measurement showed increased BV/TV of high density bone and bone mineral density (BMD) of the callus. 3 point bending mechanical test showed improved relative stiffness. However, repeated injection of miR-29b-3p at weeks 2 and 3 did not result in additive therapeutic outcome, and caused increased total tissue volume and reduced BMD of the callus. This is the first report showing significant therapeutic effect of miR-29b-3p on femoral fracture healing through site-specific delivery with microbubble-ultrasound system. Further studies are warranted to investigate the underlying mechanisms and to refine the treatment protocol.

Targeting c-fms kinase attenuates chronic aristolochic acid nephropathy in mice.

Aristolochic acid nephropathy (AAN) is a progressive kidney disease caused by some Chinese herbal medicines, but treatment remains ineffective. Macrophage accumulation is an early feature in human and experimental AAN; however, the role of macrophages in chronic AAN is unknown. We report here that targeting macrophages with fms-I, a selective inhibitor of the tyrosine kinase activity of the macrophage colony-stimulating factor receptor, suppressed disease progression in a mouse model of chronic AAN. Treatment with fms-I (10mg/kg/BID) from day 0 to 28 (prevention study) or from day 14 to 28 (intervention study) substantially inhibited macrophage accumulation and significantly improved renal dysfunction including a reduction in proteinuria and tubular damage. Progressive interstitial fibrosis (myofibroblast accumulation and collagen deposition) and renal inflammation (increased expression of MCP-1, MIF, and TNF-α) were also attenuated by fms-I treatment. These protective effects involved inhibition of TGF-ß/Smad3 and NF-kB signaling. In conclusion, the present study establishes that macrophages are key inflammatory cells that exacerbates progressive tubulointerstitial damage in chronic AAN via mechanisms associated with TGF-ß/Smad3-mediated renal fibrosis and NF-κB-driven renal inflammation. Targeting macrophages via a c-fms kinase inhibitor may represent a novel therapy for chronic AAN.

Identification of Genes Associated with Smad3-dependent Renal Injury by RNA-seq-based Transcriptome Analysis.

Transforming growth factor-ß/Smad3 signaling plays a critical role in the process of chronic kidney disease (CKD), but targeting Smad3 systematically may cause autoimmune disease by impairing immunity. In this study, we used whole-transcriptome RNA-sequencing to identify the differential gene expression profile, gene ontology, pathways, and alternative splicing related to TGF-ß/Smad3 in CKD. To explore common dysregulation of genes associated with Smad3-dependent renal injury, kidney tissues of Smad3 wild-type and knockout mice with immune (anti-glomerular basement membrane glomerulonephritis) and non-immune (obstructive nephropathy)-mediated CKD were used for RNA-sequencing analysis. Totally 1922 differentially expressed genes (DEGs) were commonly found in these CKD models. The up-regulated genes are inflammatory and immune response associated, while decreased genes are material or electron transportation and metabolism related. Only 9 common DEGs were found to be Smad3-dependent in two models, including 6 immunoglobulin genes (Ighg1, Ighg2c, Igkv12-41, Ighv14-3, Ighv5-6 and Ighg2b) and 3 metabolic genes (Ugt2b37, Slc22a19, and Mfsd2a). Our results identify transcriptomes associated with renal injury may represent a common mechanism for the pathogenesis of CKD and reveal novel Smad3 associated transcriptomes in the development of CKD.

MicroRNAs in renal fibrosis.

MicroRNAs (miRNAs) are endogenous short non-coding RNAs that regulate most of important cellular processes by inhibiting gene expression through the post-transcriptional repression of their target mRNAs. In kidneys, miRNAs have been associated in renal development, homeostasis, and physiological functions. Results from clinical and experimental animal studies demonstrate that miRNAs play essential roles in the pathogenesis of various renal diseases. Chronic kidney diseases (CKD) is characterized by renal fibrosis. Transforming growth factor beta (TGF-ß) is recognized as a major mediator of renal fibrosis because it is able to stimulate the accumulation of extracellular matrix (ECM) proteins to impair normal kidney function. Recently, emerging evidence demonstrate the relationship between TGF-ß signaling and miRNAs expression during renal diseases. TGF-ß regulates expression of several microRNAs, such as miR-21, miR-192, miR-200, miR-433, and miR-29. MiR-21, miR-192, and miR-433 which are positively induced by TGF-ß signaling play a pathological role in kidney diseases. In contrast, members in both miR-29 and miR-200 families which are inhibited by TGF-ß signaling protect kidneys from renal fibrosis by suppressing the deposition of ECM and preventing epithelial-to-mesenchymal transition, respectively. Clinically, the presence of miRNAs in blood and urine has been examined to be early biomarkers for detecting renal diseases. From experimental animal studies of CKD, targeting microRNAs also provides evidence about therapeutic potential of miRNAs during renal diseases. Now, it comes to the stage to examine the exact mechanisms of miRNAs during the initiation and progression of renal diseases. Therefore, determining the function of miRNAs in renal fibrosis may facilitate the development of both early diagnosis and treatment of renal diseases.

Long Noncoding RNA Arid2-IR Is a Novel Therapeutic Target for Renal Inflammation.

Increasing evidence shows that microRNAs play an important role in kidney disease. However, functions of long noncoding RNAs (lncRNAs) in kidney diseases remain undefined. We have previously shown that TGF-ß1 plays a diverse role in renal inflammation and fibrosis and Smad3 is a key mediator in this process. In this study, we used RNA-sequencing to identify lncRNAs related to renal inflammation and fibrosis in obstructive nephropathy induced in Smad3 wild-type and knockout mice. We found that Arid2-IR was a Smad3-associated lncRNA as a Smad3 binding site was found in the promoter region of Arid2-IR and deletion of Smad3 abolished upregulation of Arid2-IR in the diseased kidney. In vitro knockdown of Arid2-IR from tubular epithelial cells produced no effect on TGF-ß-induced Smad3 signaling and fibrosis but inhibited interleukin-1ß-stimulated NF-κB-dependent inflammatory response. In contrast, overexpression of Arid2-IR promoted interleukin-1ß-induced NF-κB signaling and inflammatory cytokine expression without alteration of TGF-ß1-induced fibrotic response. Furthermore, treatment of obstructed kidney with Arid2-IR shRNA blunted NF-κB-driven renal inflammation without effect on TGF-ß/Smad3-mediated renal fibrosis. Thus, Arid2-IR is a novel lncRNA that functions to promote NF-κB-dependent renal inflammation. Blockade of Arid2-IR may represent a novel and specific therapy for renal inflammatory disease.

Smad7 inhibits AngII-mediated hypertensive nephropathy in a mouse model of hypertension.

The TGFß (transforming growth factor ß)/SMAD and NF-κB (nuclear factor κB) signalling pathways play a key role in hypertensive nephropathy. The present study examined whether targeting these pathways by SMAD7, a downstream inhibitor of both pathways, blocks AngII (angiotensin II)-induced hypertensive kidney disease in mice. A doxycycline-inducible SMAD7-expressing plasmid was delivered into the kidney by a non-invasive ultrasound-microbubble technique before and after AngII infusion. Results showed that pre-treatment with SMAD7 prevented AngII-induced progressive renal injury by inhibiting an increase in proteinuria and serum creatinine while improving the glomerular filtration rate. Similarly, treatment with SMAD7 in the established hypertensive nephropathy at day 14 after AngII infusion halted the progressive renal injury. These preventive and therapeutic effects of SMAD7 on hypertensive kidney injury were associated with inhibition of AngII-induced up-regulation of SMURF2 (SMAD-specific E3 ubiquitin protein ligase 2) and Sp1 (specificity protein 1), blockade of TGFß/Smad3-mediated renal fibrosis and suppression of NF-κB-driven renal inflammation. Moreover, overexpression of SMAD7 also prevented AngII-induced loss of renal miR-29b, an miRNA with an inhibitory role in both TGFß/Smad3 and NF-κB pathways. In conclusion, SMAD7 may be a therapeutic agent for AngII-mediated hypertensive nephropathy. Inhibition of the Sp1/SMAD3/NF-κB/miR-29b regulatory network may be a mechanism by which SMAD7 inhibits hypertensive nephropathy.

MicroRNAs in Diabetic Kidney Disease.

Rapid growth of diabetes and diabetic kidney disease exerts a great burden on society. Owing to the lack of effective treatments for diabetic kidney disease, treatment relies on drugs that either reduces its progression or involve renal replacement therapies, such as dialysis and kidney transplantation. It is urgent to search for biomarkers for early diagnosis and effective therapy. The discovery of microRNAs had lead to a new era of post-transcriptional regulators of gene expression. Studies from cells, experimental animal models and patients under diabetic conditions demonstrate that expression patterns of microRNAs are altered during the progression of diabetic kidney disease. Functional studies indicate that the ability of microRNAs to bind 3' untranslated region of messenger RNA not only shows their capability to regulate expression of target genes, but also their therapeutic potential to diabetic kidney disease. The presence of microRNAs in plasma, serum, and urine has been shown to be possible biomarkers in diabetic kidney disease. Therefore, identification of the pathogenic role of microRNAs possesses an important clinical impact in terms of prevention and treatment of progression in diabetic kidney disease because it allows us to design novel and specific therapies and diagnostic tools for diabetic kidney disease.

MicroRNA-29b inhibits diabetic nephropathy in db/db mice.

Inflammation and its consequent fibrosis are two main features of diabetic nephropathy (DN), but target therapy on these processes for DN remains yet ineffective. We report here that miR-29b is a novel therapeutic agent capable of inhibiting progressive renal inflammation and fibrosis in type 2 diabetes in db/db mice. Under diabetic conditions, miR-29b was largely downregulated in response to advanced glycation end (AGE) product, which was associated with upregulation of collagen matrix in mesangial cells via the transforming growth factor-ß (TGF-ß)/Smad3-dependent mechanism. These pathological changes were reversed by overexpressing miR-29b, but enhanced by knocking-down miR-29b. Similarly, loss of renal miR-29b was associated with progressive diabetic kidney injury, including microalbuminuria, renal fibrosis, and inflammation. Restored renal miR-29b by the ultrasound-based gene therapy was capable of attenuating diabetic kidney disease. Further studies revealed that inhibition of Sp1 expression, TGF-ß/Smad3-dependent renal fibrosis, NF-κB-driven renal inflammation, and T-bet/Th1-mediated immune response may be mechanisms associated with miR-29b treatment in db/db mice. In conclusion, miR-29b may play a protective role in diabetic kidney disease and may have therapeutic potential for diabetic kidney complication.

C-reactive protein promotes acute kidney injury by impairing G1/S-dependent tubular epithelium cell regeneration.

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 FcγRII (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.

Identification of novel long noncoding RNAs associated with TGF-ß/Smad3-mediated renal inflammation and fibrosis by RNA sequencing.

We have previously shown that transforming growth factor-ß/Smad3-dependent miRNAs play a critical role in renal inflammation and fibrosis. However, off-target effects of miRNAs limit their therapeutic application. Recently, emerging roles of long noncoding RNAs (lncRNAs) in diseases have been recognized. In this study, we used high-throughput RNA sequencing to identify the Smad3-dependent lncRNAs related to renal inflammation and fibrosis in Smad3 knockout mouse models of unilateral ureteral obstructive nephropathy and immunologically induced anti-glomerular basement membrane glomerulonephritis. Compared with wild-type mice, 151 lncRNAs in the unilateral ureteral obstructive nephropathy kidney and 413 lncRNAs in kidneys with anti-glomerular basement membrane glomerulonephritis were significantly altered in Smad3 knockout mice. Among them, 21 common lncRNAs were up-regulated in wild-type, but down-regulated in Smad3 knockout, kidneys in both disease models in which progressive renal inflammation and fibrosis were abolished when the Smad3 gene was deleted or suppressed. Real-time PCR confirmed these findings and revealed the functional link between Smad3-dependent lncRNAs np_5318/np_17856 and progressive kidney injury. Results demonstrate that the identification and characterization of functional lncRNAs associated with kidney disease may represent a promising research direction into renal disorder and may lead to the development of new lncRNA therapies for kidney diseases.