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Editor's Note: this Article has been retracted; the Retraction Note is available at https://doi.org/10.1038/s41598-023-32033-5

PURPOSE OF REVIEW Diabetic kidney disease (DKD) is a significant health concern. Innovative strategies to prevent or limit the progression of DKD are urgently needed due to the limitation of existing treatments. KCa3.1, a potassium channel, is involved in a range of biological processes from cell survival to cell death. This review summarizes the current knowledge on the pathophysiological functions of the KCa3.1 channel, specifically its involvement in maintaining mitochondrial function. More specifically, the therapeutic potential of targeting KCa3.1 in DKD is systematically discussed in the review. RECENT FINDINGS Mitochondrial dysfunction contributes to the development and progression of DKD. Accumulating evidence indicates that KCa3.1 dysregulation plays a crucial role in mitochondrial dysfunction, in addition to driving cellular activation, proliferation and inflammation. Recent studies demonstrate that KCa3.1 deficiency improves diabetes-induced mitochondrial dysfunction in DKD, which is attributed to modulation of mitochondrial quality control through mitigating the altered mitochondrial dynamics and restoring abnormal BNIP3-mediated mitophagy. SUMMARY Based on its role in fibrosis, inflammation and mitochondrial dysfunction, pharmacological inhibition of KCa3.1 may offer a promising alternative for the treatment of DKD. Due to its safety profile in humans, the repurposing of senicapoc has the potential to expedite an urgently needed new drug in DKD.

Faecal microbiota transplantation (FMT) has attracted increasing attention as an intervention in many clinical conditions, including autoimmune, enteroendocrine, gastroenterological, and neurological diseases. For years, FMT has been an effective second-line treatment for Clostridium difficile infection (CDI) with beneficial outcomes. FMT is also promising in improving bowel diseases, such as ulcerative colitis (UC). Pre-clinical and clinical studies suggest that this microbiota-based intervention may influence the development and progression of chronic kidney disease (CKD) via modifying a dysregulated gut–kidney axis. Despite the high morbidity and mortality due to CKD, there are limited options for treatment until end-stage kidney disease occurs, which results in death, dialysis, or kidney transplantation. This imposes a significant financial and health burden on the individual, their families and careers, and the health system. Recent studies have suggested that strategies to reverse gut dysbiosis using FMT are a promising therapy in CKD. This review summarises the preclinical and clinical evidence and postulates the potential therapeutic effect of FMT in the management of CKD.

Chronic kidney disease (CKD) is rising in global prevalence and has become a worldwide public health problem, with adverse outcomes of kidney failure, cardiovascular disease, and premature death. However, current treatments are limited to slowing rather than reversing disease progression or restoring functional nephrons. Hence, innovative strategies aimed at kidney tissue recovery hold promise for CKD therapy. Mesenchymal stem cells (MSCs) are commonly used for regenerative therapy due to their potential for proliferation, differentiation, and immunomodulation. Accumulating evidence suggests that the therapeutic effects of MSCs are largely mediated by paracrine secretion of extracellular vesicles (EVs), predominantly exosomes. MSC-derived exosomes (MSC-Exos) replicate the functions of their originator MSCs via delivery of various genetic and protein cargos to target cells. More recently, MSC-Exos have also been utilized as natural carriers for targeted drug delivery. Therapeutics can be effectively incorporated into exosomes and then delivered to diseased tissue. Thus, MSC-Exos have emerged as a promising cell-free therapy in CKD. In this paper, we describe the characteristics of MSC-Exos and summarize their therapeutic efficacy in preclinical animal models of CKD. We also discuss the potential challenges and strategies in the use of MSC-Exos-based therapies for CKD in the future.

Chronic kidney disease (CKD) is a growing global public health problem. The implementation of evidence-based clinical practices only defers the development of kidney failure. Death, transplantation, or dialysis are the consequences of kidney failure, resulting in a significant burden on the health system. Hence, innovative therapeutic strategies are urgently needed due to the limitations of current interventions. Photobiomodulation (PBM), a form of non-thermal light therapy, effectively mitigates mitochondrial dysfunction, reactive oxidative stress, inflammation, and gut microbiota dysbiosis, all of which are inherent in CKD. Preliminary studies suggest the benefits of PBM in multiple diseases, including CKD. Hence, this review will provide a concise summary of the underlying action mechanisms of PBM and its potential therapeutic effects on CKD. Based on the findings, PBM may represent a novel, non-invasive and non-pharmacological therapy for CKD, although more studies are necessary before PBM can be widely recommended.

It is well-known that all progressive chronic kidney disease (CKD) is pathologically characterized by tubulointerstitial fibrosis process. Multiple studies have shown the critical role of inflammation and fibrosis in the development of CKD. Hence strategies that target inflammatory and fibrotic signaling pathways may provide promising opportunities to protect against renal fibrosis. Metformin has been used as the first-line glucose-lowering agent to treat patients with type 2 diabetes mellitus (T2DM) for over 50 years. Accumulating evidence suggests the potential for additional therapeutic applications of metformin, including mitigation of renal fibrosis. In this study, the anti-fibrotic effects of metformin independent of its glucose-lowering mechanism were examined in an adenine -induced mouse model of CKD. Expressions of inflammatory markers MCP-1, F4/80 and ICAM, fibrotic markers type IV collagen and fibronectin, and the cytokine TGF-β1 were increased in adenine-induced CKD when compared to control groups and significantly attenuated by metformin treatment. Moreover, treatment with metformin inhibited the phosphorylation of Smad3, ERK1/2, and P38 and was associated with activation of the AMP-activated protein kinase (AMPK) in the kidneys of adenine-treated mice. These results indicate that metformin attenuates adenine-induced renal fibrosis through inhibition of TGF-β1 signaling pathways and activation of AMPK, independent of its glucose-lowering action.

The G protein-coupled CXC chemokine receptor 4 (CXCR4) is a candidate therapeutic target for tissue fibrosis. A fully human single-domain antibody-like scaffold i-body AD-114-PA600 (AD-114) with specific high binding affinity to CXCR4 has been developed. To define its renoprotective role, AD-114 was administrated in a mouse model of renal fibrosis induced by folic acid (FA). Increased extracellular matrix (ECM) accumulation, macrophage infiltration, inflammatory response, TGF-β1 expression, and fibroblast activation were observed in kidneys of mice with FA-induced nephropathy. These markers were normalized or partially reversed by AD-114 treatment. In vitro studies demonstrated AD-114 blocked TGF-β1-induced upregulated expression of ECM, matrix metalloproteinase-2, and downstream p38 mitogen-activated protein kinase (p38 MAPK) and PI3K/AKT/mTOR signaling pathways in a renal proximal tubular cell line. Additionally, these renoprotective effects were validated in a second model of unilateral ureteral obstruction using a second generation of AD-114 (Fc-fused AD-114, also named AD-214). Collectively, these results suggest a renoprotective role of AD-114 as it inhibited the chemotactic function of CXCR4 as well as blocked CXCR4 downstream p38 MAPK and PI3K/AKT/mTOR signaling, which establish a therapeutic strategy for AD-114 targeting CXCR4 to limit renal fibrosis.

Receptor-interacting protein kinase-3 (RIPK3) is a multifunctional regulator of cell death and inflammation. RIPK3 controls cellular signalling through the formation of the domain-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome, which is recognised to mediate renal fibrogenesis. The role of RIPK3 in diabetic kidney disease (DKD) induced renal fibrosis has not been previously determined. To define the action of RIPK3 in the development of diabetic kidney disease, wild-type (WT), RIPK3 -/- and endothelium-derived nitric oxide synthase (eNOS)-/- mice were induced to develop diabetes mellitus using multiple low doses of streptozotocin and maintained for 24 weeks. RIPK3 activity and NLRP3 expression were upregulated and fibrotic responses were increased in the kidney cortex of WT mice with established diabetic nephropathy compared to control mice. Consistently, mRNA expression of inflammasome components, as well as transforming growth factor beta 1 (TGFβ1), α smooth muscle actin (α-SMA) and collagen deposition were increased in diabetic kidneys of WT mice compared to control mice. However, these markers were normalised or significantly reversed in kidneys of diabetic RIPK3 -/- mice. Renoprotection was also observed using the RIPK3 inhibitor dabrafenib in eNOS-/- diabetic mice as demonstrated by reduced collagen deposition and myofibroblast activation. These results suggest that RIPK3 is associated with the development of renal fibrosis in DKD due to the activation of the NLRP3 inflammasome. Inhibition of RIPK3 results in renoprotection. Thus, RIPK3 may be a potential target for therapeutic intervention in patients with diabetic kidney disease.

Renal fibrosis is common to all forms of progressive kidney disease. However, current therapies to limit renal fibrosis are largely ineffective. Phosphorylation of receptor-interacting serine/threonine-protein kinase (RIPK) 3 has been recently suggested to be a key regulator of the pyrin domain containing 3 (NLRP3) inflammasome, which provides new insights into mechanisms of chronic kidney disease (CKD). However, the specific effect of RIPK3 on renal cortical fibrosis has not been fully understood. To study the function of RIPK3, both genetic ablation and pharmacological inhibition of RIPK3 (dabrafenib) were used in the study. Our studies identify that RIPK3 promotes renal fibrosis via the activation of the NLRP3 inflammasome in a mouse model of folic acid-induced nephropathy. Both interventional strategies decreased the renal fibrotic response, and beneficial effects converged on the NLRP3 inflammasome. This study demonstrates a role for RIPK3 as the mediator of renal fibrosis via the upregulation of inflammasome activation. Dabrafenib, as an inhibitor of RIPK3, may be an effective treatment to limit the progression of the tubulointerstitial fibrosis.

We present the preparation of 11 nm polyacrylamide-stabilized polystyrene latex particles for conjugation to a microRNA model by surfactant-free RAFT emulsion polymerization. Our synthetic strategy involved the preparation of amphiphilic polyacrylamide-block-polystyrene copolymers, which were able to self-assemble into polymeric micelles and "grow" into polystyrene latex particles. The surface of these sterically stabilized particles was postmodified with a disulfide-bearing linker for the attachment of the microRNA model, which can be released from the latex particles under reducing conditions. These nanoparticles offer the advantage of ease of preparation via a scaleable process, and the versatility of their synthesis makes them adaptable to a range of applications.

Diabetic kidney disease (DKD) is a life‐limiting condition characterized by progressive and irreversible loss of renal function. Currently, the estimated glomerular filtration rate (eGFR) and albuminuria are used as key markers to define DKD. However, they may not accurately indicate the degree of renal dysfunction and injury. Current therapeutic approaches for DKD, including attainment of blood pressure goals, optimal control of blood glucose and lipid levels, and the use of agents to block the renin‐angiotensin‐aldosterone system (RAAS) can only slow the progression of DKD. Hence, early diagnosis and innovative strategies are needed to both prevent and treat DKD. In recent years, a novel class of noncoding RNA, microRNAs (miRNAs) are reported to be involved in all biological processes, including cellular proliferation, apoptosis, and differentiation. miRNAs are small noncoding RNAs that regulate gene expression by posttranscriptional and epigenetic mechanisms. They are found to be in virtually all body fluids and used successfully as biomarkers for various diseases. Urinary miRNAs correlate with clinical and histologic parameters in DKD and differential urinary miRNA expression patterns have been reported. Kidney fibrosis is the common end stage of various CKD including DKD. Transforming growth factor‐β(TGF‐β) is regarded as the master regulator of kidney fibrosis, which is likely at least in part through regulating miRNA expression. miRNA are widely involved in the progression of DKD via many molecular mechanisms. In this review, the involvement of miRNA in fibrosis, inflammation, hypertrophy, autophagy, endoplasmic reticulum (ER) stress, oxidative stress, insulin resistance, and podocyte injury will be discussed, as these mechanisms are believed to offer new therapeutic targets that can be exploited to develop important treatments for DKD over the next decade.

Hyperglycemia induces all isoforms of transforming growth factor β (TGFβ), which in turn play key roles in inflammation and fibrosis that characterize diabetic nephropathy. Sphingosine 1-phosphate (S1P) is a signaling sphingolipid, derived from sphingosine by the action of sphingosine kinase (SK). S1P mediates many biological processes, which mimic TGFβ signaling. To determine the role of SK1 and S1P in inducing fibrosis and inflammation, and the interaction with TGFβ-1, 2 and 3 signalling in diabetic nephropathy, human proximal tubular cells (HK2 cells) were exposed to normal (5 mmol/L) or high (30 mmol/L) glucose or TGFβ-1, -2, -3 ± an SK inhibitor (SKI-II) or SK1 siRNA. Control and diabetic wild type (WT) and SK1(-/-) mice were studied. Fibrotic and inflammatory markers, and relevant downstream signalling pathways were assessed. SK1 mRNA and protein expression was increased in HK2 cells exposed to high glucose or TGFβ1,-2,-3. All TGFβ isoforms induced fibronectin, collagen IV and macrophage chemoattractant protein 1 (MCP1), which were reversed by both SKI-II and SK1 siRNA. Exposure to S1P increased phospho-p44/42 expression, AP-1 binding and NFkB phosphorylation. WT diabetic mice exhibited increased renal cortical S1P, fibronectin, collagen IV and MCP1 mRNA and protein expression compared to SK1(-/-) diabetic mice. In summary, this study demonstrates that inhibiting the formation of S1P reduces tubulointerstitial renal inflammation and fibrosis in diabetic nephropathy.

Purpose of review Hypertension and hyperglycaemia are major risk factors that result in chronic kidney disease (CKD). Achievement of blood pressure goals, optimal control of blood glucose levels and the use of agents to block the renin-angiotensin-aldosterone system slow the progression of CKD. However, not all patients are benefited by these interventions and novel strategies to arrest or reverse the pathological processes inherent in CKD are needed. The therapeutic potential of targeting KCa3.1 in CKD will be discussed in this review. Recent findings Blockade of KCa3.1 ameliorates activation of renal fibroblasts in diabetic mice by inhibiting the transforming growth factor-β1/small mothers against decapentaplegic pathway. A concomitant reduction in nuclear factor-κB activation in human proximal tubular cells under diabetic conditions has been observed. Advanced glycosylated endproducts induce both protein expression and current density of KCa3.1, which, in turn, mediates migration and proliferation of vascular smooth muscle cells via Ca²⁺-dependent signalling pathways. Summary Studies have clearly demonstrated a causal role of chronic hyperglycaemia and hypertension in the development of CKD. However, a large proportion of patients develop end-stage kidney disease despite strict glycaemic control and the attainment of recommended blood pressure goals. Therefore, it is essential to identify and validate novel targets to reduce the development and progression of CKD. Recent findings demonstrate that genetic deletion or pharmacologic inhibition of KCa3.1 significantly reduces the development of diabetic nephropathy in animal models. However, the consequences of blockade of KCa3.1 in preventing and treating established diabetic nephropathy in humans warrants further study.

Progressive tubulointerstitial fibrosis has been recognized as a common pathological process that leads to the progression of all chronic kidney disease (CKD). Innovative strategies are needed to both prevent and treat CKD. Inflammatory and fibrotic signaling pathways play central roles in the progression of CKD regardless of aetiology. Hence, targeting inflammatory and fibrotic responses holds promise to limit renal fibrosis. Metformin has been the most prescribed glucose-lowering medicine worldwide, and its potential for many other therapeutic applications is also being explored intensively. Increasing evidence indicates metformin may limit renal fibrosis. However, the exact mechanisms whereby metformin limits renal injury are not fully understood. The anti-fibrotic effects of metformin, independent of improved glycaemic control was examined in a folic acid-induced mouse model of nephropathy for 14 days. Human proximal tubular cells (HK2 cells) exposed to TGF-β1 were used in in vitro models to examine mechanistic pathways. Folic acid induced nephropathy was associated with the overexpression of inflammatory markers MCP-1, F4/80, type IV collagen, fibronectin and TGF-β1 compared to control groups, which were partially attenuated by metformin treatment. In vitro studies confirmed that metformin inhibited TGF-β1 induced inflammatory and fibrotic responses through Smad3, ERK1/2, and P38 pathways in human renal proximal tubular cells. These results suggest that metoformin attenuates folic acid-induced renal interstitial fibrogenesis through TGF-β1 signaling pathways.

There is an urgent need to identify novel interventions for mitigating the progression of diabetic nephropathy. Diabetic nephropathy is characterized by progressive renal fibrosis, in which tubulointerstitial fibrosis has been shown to be the final common pathway of all forms of chronic progressive renal disease, including diabetic nephropathy. Therefore targeting the possible mechanisms that drive this process may provide novel therapeutics which allow the prevention and potentially retardation of the functional decline in diabetic nephropathy. Recently, the Ca2+-activated K+ channel KCa3.1 (KCa3.1) has been suggested as a potential therapeutic target for nephropathy, based on its ability to regulate Ca2+ entry into cells and modulate Ca2+-signalling processes. In the present review, we focus on the physiological role of KCa3.1 in those cells involved in the tubulointerstitial fibrosis, including proximal tubular cells, fibroblasts, inflammatory cells (T-cells and macrophages) and endothelial cells. Collectively these studies support further investigation into KCa3.1 as a therapeutic target in diabetic nephropathy.

Thioredoxin-interacting protein (TXNIP) expression is ubiquitous and is induced by a variety of cellular stresses, including high intracellular glucose. TXNIP is associated with activation of oxidative stress and tubulointerstitial fibrosis in diabetic nephropathy. Autophagy is a major pathway that delivers damaged proteins and organelles to lysosomes to maintain cellular homeostasis. This study aimed to investigate the dysregulation of autophagy and the regulation of TXNIP on autophagy in renal proximal tubular cells (PTCs) under diabetic conditions. The formation of autophagosomes was measured using transmission electron microscopy, and LC3-II, and the effectiveness of autophagic clearance was determined by p62 expression in diabetic kidney and in human PTCs exposed to high glucose (HG). The results collectively demonstrated increased expression of TXNIP, LC3/LC3-II and p62 in renal tubular cells of mice with diabetic nephropathy and in cultured human PTCs exposed to HG (30 mM/l) for 48 h compared with control. The formation of autophagic vacuoles was increased in HG-induced cells. Furthermore, silencing of TXNIP by siRNA transfection reduced autophagic vacuoles and the expression of LC3-II and p62 in human PTCs exposed to HG compared with control and partially reversed the accumulation of LC3-II and p62 induced by bafilomycin A1 (50 nM/l), a pharmacological inhibitor of autophagy which blocks the fusion of autophagosomes with lysosomes and impairs the degradation of LC3-II and p62. Collectively, these results suggest that hyperglycemia leads to dysfunction of autophagy in renal tubular cells and decreases autophagic clearance. HG-induced overexpression of TXNIP may contribute to the dysfunction of tubular autophagy in diabetes.

Background Fibroblast activation plays a critical role in diabetic nephropathy (DN). The Ca2+-activated K+ channel KCa3.1 mediates cellular proliferation of many cell types including fibroblasts. KCa3.1 has been reported to be a potential molecular target for pharmacological intervention in a diverse array of clinical conditions. However, the role of KCa3.1 in the activation of myofibroblasts in DN is unknown. These studies assessed the effect of KCa3.1 blockade on renal injury in experimental diabetes. Methods As TGF-β1 plays a central role in the activation of fibroblasts to myofibroblasts in renal interstitial fibrosis, human primary renal interstitial fibroblasts were incubated with TGF-β1+/- the selective inhibitor of KCa3.1, TRAM34, for 48 h. Two streptozotocin-induced diabetic mouse models were used in this study: wild-type KCa3.1+/+ and KCa3.1-/- mice, and secondly eNOS-/- mice treated with or without a selective inhibitor of KCa3.1 (TRAM34). Then, markers of fibroblast activation and fibrosis were determined. Results Blockade of KCa3.1 inhibited the upregulation of type I collagen, fibronectin, α-smooth muscle actin, vimentin and fibroblast-specific protein-1 in renal fibroblasts exposed to TGF-β1 and in kidneys from diabetic mice. TRAM34 reduced TGF-β1-induced phosphorylation of Smad2/3 and ERK1/2 but not P38 and JNK MAPK in interstitial fibroblasts. Conclusions These results suggest that blockade of KCa3.1 attenuates diabetic renal interstitial fibrogenesis through inhibiting activation of fibroblasts and phosphorylation of Smad2/3 and ERK1/2. Therefore, therapeutic interventions to prevent or ameliorate DN through targeted inhibition of KCa3.1 deserve further consideration.

The Ca2+-activated K+ channel KCa3.1 mediates cellular signaling processes associated with dysfunction of vasculature. However, the role of KCa3.1 in diabetic nephropathy is unknown. We sought to assess whether KCa3.1 mediates the development of renal fibrosis in two animal models of diabetic nephropathy. Wild-type and KCa3.1−/− mice, and secondly eNOS−/− mice, had diabetes induced with streptozotocin and then were treated with/without a selective inhibitor of KCa3.1 (TRAM34). Our results show that the albumin-to-creatinine ratio significantly decreased in diabetic KCa3.1−/− mice compared with diabetic wild-type mice and in diabetic eNOS−/− mice treated with TRAM34 compared with diabetic mice. The expression of monocyte chemoattractant protein-1 (MCP-1), intercellular adhesion molecule 1 (ICAM1), F4/80, plasminogen activator inhibitor type 1 (PAI-1), and type III and IV collagen significantly decreased (P < 0.01) in kidneys of diabetic KCa3.1−/− mice compared with diabetic wild-type mice. Similarly, TRAM34 reduced the expression of the inflammatory and fibrotic markers described above in diabetic eNOS−/− mice. Furthermore, blocking the KCa3.1 channel in both animal models led to a reduction of transforming growth factor-β1 (TGF-β1) and TGF-β1 type II receptor (TβRII) and phosphorylation of Smad2/3. Our results provide evidence that KCa3.1 mediates renal fibrosis in diabetic nephropathy through the TGF-β1/Smad signaling pathway. Blockade of KCa3.1 may be a novel target for therapeutic intervention in patients with diabetic nephropathy.

MicroRNAs (miRNAs) comprise of a novel class of endogenous small noncoding RNAs that frequently downregulate the expression of target genes. Recent reports suggest that miRNA-200b prevents epithelial-to-mesenchymal transition (EMT) in cancer cells by targeting the E-box binding transcription factors Zinc finger E-box-binding homeobox 1 (ZEB1) and Zinc finger E-box-binding homeobox 2 (ZEB2). About 35% of active fibroblasts are derived from EMT which is central to the development of progressive renal fibrosis. Hence, this study was designed to assess the effect of miRNA-200b on transforming growth factor-β (TGF-β1)-induced fibrotic responses in renal tubular cells. Morphologically, human kidney-2 cells transfected with miRNA-200b retained their epithelial cell characteristics when exposed to TGF-β1. miRNA-200b significantly increased E-cadherin ( P < 0.001) and reduced fibronectin mRNA and protein expression (both P < 0.01) independent of phospho-Smad2/3 and phospho-p38 and p42/44 signaling. Increased E-cadherin expression was associated with decreased expression of ZEB1 and ZEB2 and repression of fibronectin was mediated through direct targeting of the fibronectin mRNA, demonstrated using pMIR luciferase reporter assay and site-directed mutagenesis. These results suggest that miRNA-200b suppresses TGF-β1-induced EMT via inhibition of ZEB1 and ZEB2 and the extracellular matrix protein fibronectin by directing targeting of its 3′UTR mRNA, independent of pathways directly involved in TGF-β1 signaling.

Hyperglycemia upregulates thioredoxin interacting protein (TXNIP) expression, which in turn induces ROS production, inflammatory and fibrotic responses in the diabetic kidney. Dysregulation of autophagy contributes to the development of diabetic nephropathy. However, the interaction of TXNIP with autophagy/mitophagy in diabetic nephropathy is unknown. In this study, streptozotocin-induced diabetic rats were given TXNIP DNAzyme or scrambled DNAzyme for 12 weeks respectively. Fibrotic markers, mitochondrial function and mitochondrial reactive oxygen species (mtROS) were assessed in kidneys. Tubular autophagy and mitophagy were determined in kidneys from both human and rats with diabetic nephropathy. TXNIP and autophagic signaling molecules were examined. TXNIP DNAzyme dramatically attenuated extracellular matrix deposition in the diabetic kidneys compared to the control DNAzyme. Accumulation of autophagosomes and reduced autophagic clearance were shown in tubular cells of human diabetic compared to non-diabetic kidneys, which was reversed by TXNIP DNAzyme. High glucose induced mitochondrial dysfunction and mtROS production and inhibited mitophagy in proximal tubular cells, which was reversed by TXNIP siRNA. TXNIP inhibition suppressed diabetes-induced BNIP3 expression and activation of the mTOR signaling pathway. Collectively, hyperglycemia-induced TXNIP contributes to the dysregulation of tubular autophagy and mitophagy in diabetic nephropathy through activation of the mTOR signaling pathway.

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related mortality worldwide. However, there is a shortage of suitable diagnostic markers for early stages of NSCLC and therapeutic targets are limited. Right open reading frame (Rio) kinase 2 (RIOK2) and Nin one binding (NOB1) protein are important accessory factors in ribosome assembly and are highly expressed in malignant tumours; moreover, they interact with each other. However, the RIOK2 expression profile and its clinical significance as well as NOB1’s mechanism in NSCLC remain unknown. In this study, NSCLC cell lines and 15 NSCLC tumour tissues (paired with adjacent normal lung tissues) were collected for a real-time quantitative PCR (RT-qPCR) analysis. In addition, 153 NSCLC cases and 27 normal lung tissues were used in an immunohistochemical analysis to evaluate the RIOK2 and NOB1 expression profiles, their clinicopathological factors in NSCLC and their correlations with prognoses. RIOK2 and NOB1 were highly expressed in NSCLC cells and tissues and their expression profiles were significantly associated with the Tumour Node Metastasis (TNM) clinical stage, lymph node metastasis and differentiation. RIOK2 expression was correlated with NOB1. The results suggested that simultaneously determining the expression of RIOK2 and NOB1 will improve the diagnostic rate in early stages of NSCLC. Moreover, RIOK2 and NOB1 might be potential targets for NSCLC therapy.

Autophagy is emerging as an important pathway in many diseases including diabetic nephropathy. It is acknowledged that oxidative stress plays a critical role in autophagy dysfunction and diabetic nephropathy and KCa3.1 blockade ameliorates diabetic renal fibrosis through inhibiting TGF-β1 signaling pathway. To identify the role of KCa3.1 in dysfunctional tubular autophagy in diabetic nephropathy, human proximal tubular cells (HK2) transfected with scrambled or KCa3.1 siRNAs were exposed to TGF-β1 for 48 h, then autophagosome formation, the autophagy marker LC3, signaling molecules PI3K, Akt and mTOR and oxidative stress marker nitrotyrosine were examined respectively. In vivo, LC3, nitrotyrosine and phosphorylated mTOR were examined in kidneys of diabetic KCa3.1+/+ and KCa3.1−/− mice. The results demonstrated that TGF-β1 increased the formation of autophagic vacuoles, LC3 expression and phosphorylation of PI3K, Akt and mTOR in scrambled siRNA transfected HK2 cells compared to control cells, which was reversed in KCa3.1 siRNA transfected HK2 cells. In vivo, expression of LC3 and nitrotyrosine and phosphorylation of mTOR were significantly increased in kidneys of diabetic KCa3.1+/+ mice compared to non-diabetic mice, which were attenuated in kidneys of diabetic KCa3.1−/− mice. These results suggest that KCa3.1 activation contributes to dysfunctional tubular autophagy in diabetic nephropathy through PI3K/Akt/mTOR signaling pathways.

Aimed at the support problem of the deep large-section tunnel excavated in high-stress condition with fractured surrounding rock, the different characteristics of the stresses and displacements of the surrounding rock of the tunnel with different section supports and lateral pressure coefficients were analyzed using numerical simulation and theoretical calculation, so was the effect of the height of the double-layered I-section beam on the bearing capacity. Arched with bottom arch superimposed and fully-closed I-section beam suitable for large-section tunnel excavated in high-stress condition with fractured surrounding rock was developed. This support applies fully-closed and horse-shoe shaped structure with I-section beam overlapped and welded and its bearing capacity is 2.5 times higher than frequently-used I-section beam. Underground field testing showed that the deformation of the surrounding rock could be effectively controlled when the secondary arched with bottom arch superimposed and fully-closed I-section beam of high bearing capacity was used.

The aim of the present research was to investigate clinicopathologic correlations of immunohistochemically- demonstrated axin (axis inhibition) and β-catenin expression in primary hepatocellular carcinomas (HCCs), in comparison with paraneoplastic, cirrhotic and normal liver tissues. Variation in Axin expression across groups were significant (P0.01), correlating with alpha fetoprotein (AFP), HBsAg, cancer plugs in the portal vein, and clinical stage of HCCs(P0.05); however, there were no links with sex, age, and tumour size (P0.05). Differences in cell membrane β-catenin expression were also statistically significant (P0.01), again correlated with AFP, HBsAg, cancer plugs in the portal vein, and clinical stage in HCCs (P0.05) but not with sex, age, and tumour size (P0.05). Axin expression levels in tissues with reduced membrane β-catenin were low (P0.05), also being low with nuclear β-catenin expression (P0.05). Axin and β-catenin may play an important role in the genesis and progression of HCC via the Wnt signal transmission pathway. Simultaneous determination of axin, β-catenin, AFP, and HBsAg may be useful for early diagnosis, and metastatic and clinical staging of HCCs.

Hyperglycemia and hypoxia have independent and convergent roles in the development of renal disease. Transforming growth factor-β1(TGF-β1) is a key cytokine promoting the production of extracellular matrix proteins. The cationic-independent mannose 6-phosphate receptor (CI-M6PR) is a membrane protein that binds M6P-containing proteins. A key role is to activate latent TGF-β1. PXS25, a novel CI-MPR inhibitor, has antifibrotic properties in skin fibroblasts, but its role in renal fibrosis is unclear. The aim was to study the role of PXS25 in matrix protein production under high glucose ± hypoxic conditions in human proximal tubule (HK-2) cells. HK-2 cells were exposed to high glucose (30 mM) ± 100 μM PXS25 in both normoxic (20% O2) and hypoxic (1% O2) conditions for 72 h. Cellular fibronectin, collagen IV, and matrix metalloproteinase-2 (MMP-2) and MMP-9 were assessed. Total and active TGF-β1were measured by ELISA. High glucose and hypoxia independently induced TGF-β1production. Active TGF-β1, but not total TGF-β1was reduced with concurrent PXS25 in the presence of high glucose, but not in hyperglycemia+hypoxia conditions. Hyperglycemia induced fibronectin and collagen IV production ( P < 0.05), as did hypoxia, but only hyperglycemia-induced increases in matrix proteins were suppressed by concurrent PXS25 exposure. High glucose induced MMP-2 and -9 in normoxic and hypoxic conditions, which was not modified in the presence of PXS25. High glucose and hypoxia can independently induce endogenous active TGF-β1production in human proximal tubular cells. PXS25 inhibits conversion of high glucose-induced release of active TGF-β1, only in the absence of hypoxia.

Macrophage inflammatory protein-3 alpha (MIP-3α) is known to be upregulated early in the development of diabetic nephropathy (DN). However, the transcriptional regulation of MIP-3α is unknown. We previously demonstrated that the transcription factors KLF6 and PPAR-γ play key roles in regulating renal fibrotic and inflammatory responses to factors inherent in diabetes mellitus. Hence we determined the role of these transcription factors in regulating MIP-3α expression. HK-2 cells and STZ-induced diabetic rats were used. siRNAs, over-expressing constructs and CHIP promoter binding assays were used to determine the role of KLF6 and PPAR-γ in MIP-3α transcriptional regulation. KLF6 overexpression increased MIP-3α which was inhibited by concurrent exposure to PPAR-γ agonists. PPAR-γ agonists attenuated high glucose-induced MIP-3α secretion. Furthermore, MIP-3α secretion was up-regulated in PPAR-γ silenced cells, suggesting both KLF6 and PPAR-γ antagonistically regulate high glucose-induced MIP-3α secretion. The CHIP promoter binding assay confirmed that PPAR-γ binds to the MIP-3α promoter and negatively regulates MIP-3α expression. PPAR-γ agonists increased the binding activity of the PPAR-γ-MIP-3α promoter. In contrast, promoter binding activity decreased in KLF6 over-expressing cells. PPAR-γ decreased in KLF6 over-expressing cells and increased in KLF6 silenced cells, while PPAR-γ siRNA had no effect on KLF6 expression, suggesting that KLF6 acted upstream of PPAR-γ in the regulation of MIP-3α. In diabetic rats, renal MIP-3α and the macrophage marker ED-1 expression increased, which was inhibited by exposure to PPAR-γ agonists. The recognition of MIP-3α as a significant pathogenic mediator in diabetic nephropathy reaffirms the increasingly recognized role of inflammation in the progression of DN. Targeting pro-inflammatory chemokine MIP-3α and its signaling pathways will provide novel strategy to treat diabetic kidney disease.

The centrality of the transcriptional regulator Snail in epithelial-to-mesenchymal transformation (EMT), known to occur in models of diabetic nephropathy, has not been established. Transforming growth factor beta-1 (TGFbeta1) is induced in diabetic nephropathy and induces both Snail and EMT. Hypoxia inducible factors (HIFs) are known to induce Snail, independent of TGFbeta1. Notch induction is integral to Snail induction and EMT in tumour cells, but its role in the kidney is unknown. The present study was undertaken to determine the upstream regulators of Snail in the kidney in high glucose and hypoxic conditions. HK-2 cells were cultured in normoxic, hypoxic, high glucose and combined hypoxic/high glucose conditions. The expression of HIF1alpha, NotchIC, Snail, Lysyl oxidase-like 2 (Loxl2), and Hairy and Enhancer Split-1 (Hes1) were measured. We found that hypoxia increased HIF1alpha expression; however, concurrent exposure to high glucose blunted this effect. A similar pattern was observed in Lox12 expression, suggesting that Loxl2 was downstream of HIF1alpha, which was confirmed using siRNA techniques. Snail was upregulated by hypoxia and high glucose and in combination the effect was additive, suggesting independent upstream activation pathways by the two stimuli. Hes1 was upregulated by high glucose and to a lesser extent by hypoxia, but the effect of the combined stimuli was no greater than that observed with high glucose alone. NotchIC was downregulated by both hypoxia and high glucose, and in combination the effect was additive. Therefore, this study suggests that hypoxia and high glucose induce Snail expression through distinct pathways, independent of Notch signalling.

We demonstrated recently that thioredoxin-interacting protein (Txnip) and the transcription factor Krüppel-like factor 6 (KLF6) were up-regulated in both in vivo and in vitro models of diabetic nephropathy, thus promoting renal injury. Conversely, peroxisome proliferator-activated receptor-gamma (PPAR-gamma) agonists have been shown to be renoprotective. Hence, this study was undertaken to determine whether Txnip expression is regulated by the transcription factors KLF6 and PPAR-gamma. By using siRNAs and overexpressing constructs, the role of KLF6 and PPAR-gamma in Txnip transcriptional regulation was determined in human kidney proximal tubule cells and in streptozocin-induced diabetes mellitus in Sprague-Dawley rats, in vitro and in vivo models of diabetic nephropathy, respectively. KLF6 overexpression increased Txnip expression and promoter activity, which was inhibited by concurrent exposure to PPAR-gamma agonists. In contrast, reduced expression of KLF6 by siRNA or exposure to PPAR-gamma agonists attenuated high glucose-induced Txnip expression and promoter activity. KLF6-Txnip promoter binding was decreased in KLF6-silenced cells, whereas PPAR-gamma agonists increased PPAR-gamma-Txnip promoter binding. Indeed, silencing of KLF6 increased PPAR-gamma expression, suggesting endogenous regulation of PPAR-gamma expression by KLF6. Moreover, renal KLF6 and Txnip expression increased in rats with diabetes mellitus and was inhibited by PPAR-gamma agonist treatment; however, KLF6 expression did not change in HK-2 cells exposed to PPAR-gamma agonists. Hence, Txnip expression and promoter activity are mediated via divergent effects of KLF6 and PPAR-gamma transcriptional regulation.

BACKGROUND Nuclear factor-kappa B (NF-kappaB) is believed to be involved in the pathogenesis of various inflammatory diseases, including oral lichen planus (OLP). The objective of the present study was to investigate the possible relationship between NF-kappaB activation and expression of tumor necrosis factor-alpha (TNF-alpha) in OLP and their expression pattern in relation to several clinical features. METHODS Thirty OLP cases were divided into atrophic-erosive form (14 cases) and reticular form (16 cases) according to their clinical manifestations. The expression of NF-kappaB p65 and TNF-alpha of both two groups were investigated by immunohistochemical staining, and the percentage of positive cells was calculated in each case. Biopsies of 10 normal oral mucosa (NOM) also underwent the same procedure as controls. RESULTS Nuclear factor-kappa B p65 nuclear staining was found in nuclei of basal and suprabasal epithelial keratinocytes in OLP, however, no positive staining was found in NOM. Positive TNF-alpha staining was detected in cytoplasm of basal epithelial keratinocytes in OLP, and only scattered staining was detected in NOM. Expression of NF-kappaB p65 and TNF-alpha were significantly different with respect to clinical forms and lesion sites (P 0.05) in 30 OLP cases. NF-kappaB nuclear staining positively correlated (r = 0.676, P < 0.01) with TNF-alpha overexpression in OLP. CONCLUSIONS Nuclear factor-kappa B activation and its correlation with overexpression of TNF-alpha may play an important role in pathogenesis of OLP. There might be a positive regulatory loop between NF-kappaB and TNF-alpha, which may contribute to inflammation in OLP; NF-kappaB may also protect epithelial keratinocytes from excessive apoptosis.

Transforming growth factor-beta(1) (TGFbeta(1)) is recognized as both a fibrogenic and inflammatory cytokine and plays a critical role in the kidney pathophysiology. The dysregulation of TGFbeta(1) has been linked with the development of diabetic nephropathy. Connective tissue growth factor (CTGF) is a fibrogenic cytokine and is recognized as a downstream mediator of TGFbeta(1) in kidney fibrosis. TGFbeta(1) is involved in immunomodulation and fibrosis in the kidney. However, CTGF plays a more specific role in the fibrogenic pathways in the kidney proximal tubule cells. Moreover, CTGF facilitates TGFbeta(1) signaling and promotes renal fibrosis. This suggests CTGF could be a potential target for kidney fibrosis. Long-term inhibition and targeting TGFbeta(1) directly is problematic, therefore, a more fruitful direction targeting diabetic nephropathy may involve the development of therapeutic strategies specifically targeting CTGF.

Hyperglycemia is a causative factor in the pathogenesis of diabetic nephropathy. Here, we demonstrate the transcriptional profiles of the human proximal tubule cell line (HK-2 cells) exposed to high glucose using cDNA microarray analysis. Thioredoxin-interacting protein (Txnip) was the gene most significantly increased among 10 strongly up-regulated and 15 down-regulated genes. Txnip, heat shock proteins 70 and 90, chemokine (C-C motif) ligand 20, and matrix metalloproteinase-7 were chosen for verification of gene expression. Real-time reverse transcriptase-polymerase chain reaction confirmed the mRNA expression levels of these five genes, consistent with microarray analysis. The increased protein expression of Txnip, CCL20, and MMP7 were also verified by Western blotting and enzyme-linked immunosorbent assay. Increased expression of Txnip and of nitrotyrosine, as a marker of oxidative stress, were confirmed in vivo in diabetic Ren-2 rats. Subsequent studies focused on the dependence of Txnip expression on up-regulation of transforming growth factor (TGF)-beta1 under high-glucose conditions. Overexpression of Txnip and up-regulation of Txnip promoter activity were observed in cells in which the TGF-beta1 gene was silenced in HK-2 cells using short interfering RNA technology. High glucose further increased both Txnip expression and its promoter activity in TGF-beta1 silenced cells compared with wild-type cells exposed to high glucose, suggesting that high glucose induced Txnip through a TGF-beta1-indepen-dent pathway.

AIMS: Smad7 is an inhibitory Smad that regulates transforming growth factor-beta (TGF-beta) signaling. Connective tissue growth factor (CTGF) is recognized as a potent downstream mediator of the fibrogenic effects of TGF-beta1. SMAD binding sites have been identified in both TGF-beta and CTGF promoters. The effect of CTGF on Smad7 expression and its role in the regulation of Smad7 induced by TGF-beta1 in renal tubular cells is unknown. METHODS: Human model of proximal tubular cells (HK-2 cells) was used and confirmed using a diabetic rat model. RT-PCR was performed to measure Smad7, TGF-beta1 and Smad2 and ELISA was performed to measure active TGF-beta1. CTGF or TGF-beta1 was silenced in HK-2 cells using siRNA methodology. RESULTS: TGF-beta1 induced Smad7 in a time-dependent manner, peaking at 30 min (P

Background Epidermal growth factor (EGF) has been shown to play a role in the nephromegaly and enhanced sodium reabsorption observed in diabetic nephropathy. This is recognized to be dependent on activation of serine threonine glucocorticoid kinase-1 (SGK-1). However, the roles of EGF and SGK-1 in renal fibrogenesis observed under high glucose conditions have not been established. Methods Primary cultures of human cortical fibroblasts (CFs) were used as the model in which to study the dependent and independent effects of high glucose, EGF and SGK-1 on the expression of the extracellular matrix protein (ECM) fibronectin. Wild type CFs expressing SGK-1, or cells in which SGK-1 was effectively silenced using siRNA methodology, were exposed to normal (5 mM) or high (25 mM) glucose, or EGF (10 ng/ml) for 48 hr and fibronectin assessed. The role of the EGF-receptor and its relationship to SGK-1 signaling was studied using concurrent treatment with PKI166, a specific inhibitor of EGF-receptor. Results Exposure of CF to high glucose and EGF increased phosphorylated EGF-R, SGK-1, and fibronectin expression in wild-type cells. Inhibition of the EGF-R reduced SGK-1 and fibronectin expression in control, and following exposure to EGF and high glucose conditions. In cells in which SGK-1 was silenced, fibronectin was reduced and there was no significant increase in pEGF-R, suggesting that SGK-1 is downstream of the EGF-R and negatively inhibits EGF-R activation. Conclusion These results suggest that high glucose induced fibronectin expression is mediated through the EGF-R and downstream expression of SGK-1.

Early detection of oral precancerous and malignant lesions is still a diagnostic challenge for most of clinicians, and ideal adjuncts for detection of these lesions are currently unavailable. Our preliminary study has indicated that rose bengal (RB) staining might have the potency as a diagnostic aid; however, its clinical significance and reliability in hospital-based population are still not clear. In the present study, we investigated the efficacy of RB staining in detection of oral precancerous and malignant lesions. RB staining was conducted in 132 patients, and staining results were determined by a 4-graded shade guide, which had been quantitatively measured in the 1976 CIEL*a*b* space by instrumental colorimetry. Histological examination was performed in 128 of 132 patients after RB staining. The sensitivity and specificity to detect epithelial dysplasia (DP) and oral squamous cell carcinoma were 93.9 and 73.7%, respectively. The positive and negative likelihood ratios were 3.570 and 0.082, respectively. Moreover, RB staining seemed promising to detect DP in oral leukoplakia, lichen planus and leukokeratosis. In this study, 5 of 6 DP or oral squamous cell carcinoma were identified by RB staining before histological examination. In conclusion, RB staining may be a valuable diagnostic test in detection of oral precancerous and malignant lesions.

Summary Kruppel-like factor 4 (KLF4) is known to mitigate inflammation in several cell types. Using human proximal tubule cells, the present study aimed to investigate the role of KLF4 in regulating transforming growth factor (TGF)-β1 induced inflammatory and fibrotic responses. Human kidney proximal tubular cells were exposed to high glucose, or TGF-β1 and KLF4 expressions were determined. Cells were then transfected with empty vector or KLF4 and exposed to 2-ng/mL TGF-β1 for up to 72 h. Inflammatory proteins (macrophage migration inhibitory factor and monocyte chemoattractant protein-1) and pro-fibrotic proteins (fibronectin and collagen IV) were measured after 72 h by enzyme-linked immunosorbent assay and western blot, respectively. To determine the relevance to in vivo models of chronic kidney disease, KLF4 protein expression in streptozotocin-induced diabetic mice was determined. Kruppel-like factor 4 messenger RNA (mRNA) levels were significantly reduced in high glucose-treated human kidney proximal tubular cells. High glucose increased TGF-β1 mRNA expression, which significantly increased migration inhibitory factor and monocyte chemoattractant protein-1 protein secretion. Transforming growth factor-β1 significantly increased fibronectin and collagen IV protein expression. The overexpression of KLF4 significantly reduced TGF-β–mediated increases in migration inhibitory factor and monocyte chemoattractant protein-1 but had no effect on TGF-β–mediated fibronectin and collagen IV mRNA and protein expression. The levels of KLF4 mRNA were significantly reduced in the diabetic kidney, and diabetic animals had a significant reduction in renal tubular expression of KLF4 proteins. This data suggest that KLF4 reduces inflammation induced by TGF-β1, suggesting a therapeutic role for KLF4 in diabetic nephropathy.

Transforming growth factor-β1(TGF-β1) is not only an important fibrogenic but also immunomodulatory cytokine in the human kidney. We have recently demonstrated that TGF-β1induces interleukin-8 (IL-8), macrophage chemoattractant protein-1 (MCP-1), and fibronectin production in renal proximal tubular (HK-2) cells. However, the unique dependence of IL-8, MCP-1, and fibronectin on TGF-β1expression is unknown. The TGF-β1gene was effectively silenced in HK-2 cells using small-interference (si) RNA. Basal secretion of IL-8 and MCP-1 decreased (both P < 0.05) but, paradoxically, fibronectin increased ( P < 0.05) in TGF-β1-silenced cells compared with cells transfected with nonspecific siRNA. Significant increases were observed in mRNA for the TGF-β2( P < 0.05), TGF-β3( P < 0.05) isoforms and pSmad2 ( P < 0.05), which were reflected in protein expression. Concurrent exposure to pan-specific TGF-β antibody reversed the observed increase in fibronectin expression, suggesting that TGF-β2and TGF-β3isoforms mediate the increased fibronectin expression in TGF-β1-silenced cells. An increase in the DNA binding activity of activator protein-1 (AP-1; P < 0.05) was also observed in TGF-β1-silenced cells. In contrast, nuclear factor-κB (NF-κB) DNA binding activity was significantly decreased ( P < 0.0005). These studies demonstrate that TGF-β1is a key regulator of IL-8 and MCP-1, whereas fibronectin expression is regulated by a complex interaction between the TGF-β isoforms in the HK-2 proximal tubular cell line. Decreased expression of TGF-β1reduces chemokine production in association with reduced NF-κB DNA binding activity, suggesting that immunomodulatory pathways in the kidney are specifically dependent on TGF-β1. Conversely, decreased expression of TGF-β1results in increased TGF-β2, TGF-β3, AP-1, and pSmad2 that potentially mediates the observed increase in fibronectin.

Krüppel-like factor 4 (KLF4; also known as gut-enriched Krüppel-like factor or GKLF) is known to exhibit checkpoint function during the G1/S and G2/M transitions of the cell cycle. The mechanism by which KLF4 exerts these effects is not fully established. Here we investigated the expression profile of KLF4 in an inducible system over a time course of 24 h. Using oligonucleotide microarrays, we determined that the fold changes relative to control in expression levels of KLF4 exhibited a time-dependent increase from 3- to 20-fold between 4 and 24 h following KLF4 induction. During this period and among a group of 473 cell cycle regulatory genes examined, 96 were positively correlated and 86 were negatively correlated to KLF4's expression profile. Examples of upregulated cell cycle genes include those encoding tumor suppressors such as MCC and FHIT, and cell cycle inhibitors such as CHES1 and CHEK1. Examples of downregulated genes include those that promote the cell cycle including several cyclins and those required for DNA replication. Unexpectedly, several groups of genes involved in macromolecular synthesis, including protein biosynthesis, transcription, and cholesterol biosynthesis, were also significantly inhibited by KLF4. Thus, KLF4 exerts a global inhibitory effect on macromolecular biosynthesis that is beyond its established role as a cell cycle inhibitor.

Renal cortical fibroblasts have key roles in mediating intercellular communication with neighboring/infiltrating cells and extracellular matrix (ECM) and maintenance of renal tissue architecture. They express a variety of cytokines, chemokines, growth factors and cell adhesion molecules, playing an active role in paracrine and autocrine interactions and regulating both fibrogenesis and the interstitial inflammatory response. They additionally have an endocrine function in the production of epoetin. Tubulointerstitial fibrosis, the common pathological consequence of renal injury, is characterized by the accumulation of extracellular matrix largely due to excessive production in parallel with reduced degradation, and activated fibroblasts characterized by a myofibroblastic phenotype. Fibroblasts in the kidney may derive from resident fibroblasts, from the circulating fibroblast population or from haemopoetic progenitor or stromal cells derived from the bone marrow. Cells exhibiting a myofibroblastic phenotype may derive from these sources and from tubular cells undergoing epithelial to mesenchymal transformation in response to renal injury. The number of interstitial myofibroblasts correlates closely with tubulointerstitial fibrosis and progressive renal failure. Hence inhibiting myofibroblast formation may be an effective strategy in attenuating the development of renal failure in kidney disease of diverse etiology.

Thiazolidinediones are synthetic agonists of peroxisome proliferator-activated receptors that have shown promise inin vitroand animal models of nephropathy. Encouraging early-stage data have also been generated in patients with diabetic nephropathy. Carol Pollock and colleagues outline what we know and what we need to find out before thiazolidinediones such as pioglitazone and rosiglitazone can fulfill their clinical potential. Up to a third of people with diabetes mellitus suffer end-stage renal failure due to diabetic nephropathy. Strategies to delay progression of diabetic nephropathy—including glycemic and blood pressure control, modification of the renin–angiotensin system and management of lipid levels with statins—have been effective, but development of new strategies is essential if the ever-increasing burden of this disease is to be minimized. Thiazolidinediones (TZDs) are a family of compounds used as oral hypoglycemic agents in patients with type 2 diabetes mellitus. The therapeutic effects of TZDs are largely a function of their activity as ligands of peroxisome proliferator-activated receptor gamma (PPARγ), a transcription factor that has a central role in adipogenesis and insulin sensitization. In vitro animal and clinical studies have shown that TZDs ameliorate symptoms and pathogenic mechanisms of diabetic and nondiabetic nephropathy, including proteinuria, excessive deposition of glomerular matrix, cellular proliferation, inflammation and fibrosis. Many of these favorable effects occur under both normal and high-glucose conditions. The mechanisms responsible probably involve both PPARγ-dependent and PPARγ-independent pathways. So, TZDs and other agonists of PPARγ offer promise for treatment of diabetic nephropathy; however, before their putative renoprotective effects can be translated into clinical practice, the complex mechanisms of PPARγ activity and regulation will need to be investigated further.

Peroxisome proliferator-activated receptor-γ (PPARγ) are ligand-activated transcription factors that regulate cell growth, inflammation, lipid metabolism, and insulin sensitivity. We recently demonstrated that PPARγ agonists limit high glucose-induced inflammation in a model of proximal tubular cells (PTC; Panchapakesan U, Pollock CA, and Chen XM. Am J Physiol Renal Physiol 287: F528–F534, 2004). However, the role of PPARγ in the excess extracellular matrix production is largely unknown. We evaluated the effect of 24- to 48-h 8 μM l-805645 or 10 μM pioglitazone on 25 mM d-glucose-induced markers of fibrosis in HK-2 cells. High d-glucose induced nuclear binding of activator protein-1 (AP-1) to 140.8 ± 10.9% ( P < 0.05), which was attenuated with L-805645 and pioglitazone to 82.3 ± 14.4 ( P < 0.01 vs. high d-glucose) and 99.3 ± 12.2% ( P < 0.05 vs. high d-glucose), respectively. High d-glucose increased total production of transforming growth factor (TGF)-β1139.6 ± 6.5% ( P < 0.05), which was reversed with L-805645 and pioglitazone to 68.73 ± 5.7 ( P < 0.01 vs. high d-glucose) and 112 ± 13.6% ( P < 0.05 vs. high d-glucose). L-805645 and pioglitazone reduced high d-glucose-induced fibronectin from 156.0 ± 24.9 ( P < 0.05) to 81.9 ± 16.0 and 57.4 ± 12.7%, respectively (both P < 0.01 vs. high d-glucose). Collagen IV was not induced by high d-glucose. L-805645 and pioglitazone suppressed collagen IV to 68.0 ± 14.5 ( P < 0.05) and 46.5 ± 11.6% ( P < 0.01) vs. high d-glucose, respectively. High d-glucose increased the nuclear binding of NF-κB to 167 ± 22.4% ( P < 0.05), which was not modified with PPARγ agonists. In conclusion, PPARγ agonists exert antifibrotic effects in human PTC in high glucose by attenuating the increase in AP-1, TGF-β1, and the downstream production of the extracellular matrix protein fibronectin.

Krüppel-like factor 4 (KLF4) is a zinc-finger-containing transcription factor, the expression of which is enriched in the postmitotic cells of the intestinal epithelium. KLF4 is a target gene of the tumor suppressor adenomatous polyposis coli (APC). We sought to determine the role of KLF4 in suppressing the tumorigenecity of RKO colon cancer cells, which do not express KLF4. We utilized an established system in RKO cells, in which an inducible promoter controls expression of KLF4. Four independent assays were used to assess the effects of KLF4 induction on tumor cells. We find that KLF4 overexpression reduces colony formation, cell migration and invasion, and in vivo tumorigenecity. The mechanism of action of KLF4 does not involve apoptosis. These findings, along with our previous findings that KLF4 induces G1/S arrest, suggest that KLF4 is a cell cycle checkpoint protein that can reduce tumorigenecity of colon cancer cells.

Krüppel-like factor 4 (KLF4) is an epithelially enriched, zinc finger-containing transcription factor, the expression of which is associated with growth arrest. Constitutive expression of KLF4 inhibits G1/S transition of the cell cycle but the manner by which it accomplishes this effect is unclear. To better understand the biochemical function of KLF4, we identified its target genes using cDNA microarray analysis in an established human cell line containing inducible KLF4. RNA extracted from induced and control cells were hybridized differentially to microarray chips containing 9600 human cDNAs. In all, 84 genes with significantly increased expression and 107 genes with significantly reduced expression due to KLF4 induction were identified. The affected genes are sorted to several clusters on the basis of functional relatedness. A major cluster belongs to genes involved in cell-cycle control. Within this cluster, many up-regulated genes are inhibitors of the cell cycle and down-regulated genes are promoters of the cell cycle. Another up-regulated gene cluster includes nine keratin genes, of which seven are located in a specific region on chromosome 12. The results indicate that KLF4 is involved in the control of cell proliferation and does so by eliciting changes in expression of numerous cell-cycle regulatory genes in a concerted manner. Furthermore, KLF4 regulates expression of a group of epithelial-specific keratin genes in a manner consistent with a potential locus control region function.

This paper presents a class of hybrid one-step methods that are obtained by using Cramer's rule and rational approximation to function exp(q). The algorithms fall into the catalogue of implicit formula, which involves sth order derivative and s+1 free parameters. The order of the algorithms satisfies s+1≤p≤2s+2. The stability of the methods is also studied, necessary and sufficient conditions for A-stability and L-stability are given. In addition, some examples are also given to demonstrate the method presented.

With China’s economy changing into the new normal, the business environment faced by commercial banks has undergone major changes. The real economy is facing many difficulties. The rising rate of non-performing loans has become a business problem which commercial banks have to face. Therefore, in this kind of condition, The debt-for-equity swap seems to be a good recipe for commercial banks to regain their confidence in management. However, the author thinks that the debt-for-equity swap will bring many risks to commercial banks: Financial risk, Liquidity risk, Cash Payment risk and Moral risk. Therefore, if the debt-for-equity swap will be carried out massively, it is not necessarily a good thing for the commercial Banks. So we need to take a cold look at the debt-for-equity swap.

Objectives: To identify and characterize lung cancer associated protein N35 and attempt to learn the prospective possibility of the clinical application of the protein N35. Methods: Immunoprecipitation, immunoblotting, differential centrifigation and subcellular assay, immunohistochemistry, Nglycanase digestion, mitotic cell immunoflourescence and multiple methods of affinity chromatography have been used with the monoclonal antibody N-35 to detect the distribution of the protein N35 among the various cancer cell lines and normal human tissue, the relationship between the protein N35 and glycoprotein, the location of the subcellular structure and chromosomal domain of the protein N35,the most effective way of purification of tumor associated protein N35. Results: The protein N35 is a glycoprotein, distributes to the human lung cancer cell line GLC-82, human cervical cancer cell line Hela, human hepatic cancer cell line HepG-2 and human breast cancer cell line PMC with different relative molecular mass(Mr), but no expression of the protein ingredient in normal human fresh tissue; concentrates at the nuclei significantly ,much more than at the mitochondrail and membrane, locates at the centriole of the chromosomal domain. Conclusions: The lung cancer associated protein N35 might be expressed only by the cancer cells and related with the proliferation of cancer cells as a role of tumor cell growth regulator.