Potential of Modulating Aldosterone Signaling and Mineralocorticoid Receptor with microRNAs to Attenuate Diabetic Kidney Disease.

Diabetic Kidney Disease (DKD) is a significant complication of diabetes and primary cause of end-stage renal disease globally. The exact mechanisms underlying DKD remain poorly understood, but multiple factors, including the renin-angiotensin-aldosterone system (RAAS), play a key role in its progression. Aldosterone, a mineralocorticoid steroid hormone, is one of the key components of RAAS and a potential mediator of renal damage and inflammation in DKD. miRNAs, small noncoding RNA molecules, have attracted interest due to their regulatory roles in numerous biological processes. These processes include aldosterone signaling and mineralocorticoid receptor (MR) expression. Numerous miRNAs have been recognized as crucial regulators of aldosterone signaling and MR expression. These miRNAs affect different aspects of the RAAS pathway and subsequent molecular processes, which impact sodium balance, ion transport, and fibrosis regulation. This review investigates the regulatory roles of particular miRNAs in modulating aldosterone signaling and MR activation, focusing on their impact on kidney injury, inflammation, and fibrosis. Understanding the complex interaction between miRNAs and the RAAS could lead to a new strategy to target aldosterone signaling and MR activation using miRNAs. This highlights the potential of miRNA-based interventions for DKD, with the aim of enhancing kidney outcomes in individuals with diabetes.

YB-1: the Jekyll and Hyde of kidney disease?

Y-box-binding protein 1 is a well-described and important regulator of gene transcription, which is linked to various pathologic conditions, including inflammation and fibrosis of the kidney. The identification of a novel and protective crosstalk pathway between podocytes and tubular cells in the kidney with Y-box-binding protein 1 acting as a paracrine messenger sheds new light and provides novel opportunities for renoprotection.

Glucagon-like peptide-1 receptor signaling modifies the extent of diabetic kidney disease through dampening the receptor for advanced glycation end products-induced inflammation.

Glucagon like peptide-1 (GLP-1) is a hormone produced and released by cells of the gastrointestinal tract following meal ingestion. GLP-1 receptor agonists (GLP-1RA) exhibit kidney-protective actions through poorly understood mechanisms. Here we interrogated whether the receptor for advanced glycation end products (RAGE) plays a role in mediating the actions of GLP-1 on inflammation and diabetic kidney disease. Mice with deletion of the GLP-1 receptor displayed an abnormal kidney phenotype that was accelerated by diabetes and improved with co-deletion of RAGE in vivo. Activation of the GLP-1 receptor pathway with liraglutide, an anti-diabetic treatment, downregulated kidney RAGE, reduced the expansion of bone marrow myeloid progenitors, promoted M2-like macrophage polarization and lessened markers of kidney damage in diabetic mice. Single cell transcriptomics revealed that liraglutide induced distinct transcriptional changes in kidney endothelial, proximal tubular, podocyte and macrophage cells, which were dominated by pathways involved in nutrient transport and utilization, redox sensing and the resolution of inflammation. The kidney-protective action of liraglutide was corroborated in a non-diabetic model of chronic kidney disease, the subtotal nephrectomised rat. Thus, our findings identify a novel glucose-independent kidney-protective action of GLP-1-based therapies in diabetic kidney disease and provide a valuable resource for exploring the cell-specific kidney transcriptional response ensuing from pharmacological GLP-1R agonism.

Ameliorating diabetes-associated atherosclerosis and diabetic nephropathy through modulation of soluble guanylate cyclase.

Diabetes mellitus (DM) is an independent risk factor for micro- and macrovascular complications such as nephropathy and atherosclerosis respectively, which are the major causes of premature morbidity and mortality in Type 1 and Type 2 diabetic patients. Endothelial dysfunction is the critical first step of vascular disease and is characterized by reduced bioavailability of the essential endothelial vasodilator, nitric oxide (NO), coupled with an elevation in inflammation and oxidative stress. A novel pathway to bolster NO activity is to upregulate soluble guanylate cyclase (sGC), an enzyme responsible for mediating the protective actions of NO. Two classes of sGC modulators exist, activators and stimulators, with differing sensitivity to oxidative stress. In this study, we investigated the therapeutic effects of the sGC stimulator BAY 41-2272 (Bay 41) and the sGC activator BAY 60-2770 (Bay 60) on endpoints of atherosclerosis and renal disease as well as inflammation and oxidative stress in diabetic Apolipoprotein E knockout (ApoE-/-) mice. We hypothesized that under oxidative conditions known to accompany diabetes, sGC activation might be more efficacious than sGC stimulation in limiting diabetic vascular complications. We demonstrate that Bay 60 not only significantly decreased nitrotyrosine staining (P < 0.01) and F4/80 positive cells by 75% (P < 0.05), but it also significantly reduced total plaque area (P < 0.05) and improved endothelial function (P < 0.01). Our data suggest an important anti-atherogenic role for Bay 60 accompanied by reduced oxidative stress and inflammation under diabetic settings. Treatment with the stimulator Bay 41, on the other hand, had minimal effects or caused no changes with respect to cardiovascular or renal pathology. In the kidneys, treatment with Bay 60 significantly lessened urinary albuminuria, mesangial expansion and nitrotyrosine staining under diabetic conditions. In summary, our head-to-head comparator is the first preclinical study to show that a sGC activator is more efficacious than a sGC stimulator for the treatment of diabetes-associated vascular and renal complications.

Pro-resolving lipid mediators: regulators of inflammation, metabolism and kidney function.

Obesity, diabetes mellitus, hypertension and cardiovascular disease are risk factors for chronic kidney disease (CKD) and kidney failure. Chronic, low-grade inflammation is recognized as a major pathogenic mechanism that underlies the association between CKD and obesity, impaired glucose tolerance, insulin resistance and diabetes, through interaction between resident and/or circulating immune cells with parenchymal cells. Thus, considerable interest exists in approaches that target inflammation as a strategy to manage CKD. The initial phase of the inflammatory response to injury or metabolic dysfunction reflects the release of pro-inflammatory mediators including peptides, lipids and cytokines, and the recruitment of leukocytes. In self-limiting inflammation, the evolving inflammatory response is coupled to distinct processes that promote the resolution of inflammation and restore homeostasis. The discovery of endogenously generated lipid mediators - specialized pro-resolving lipid mediators and branched fatty acid esters of hydroxy fatty acids - which promote the resolution of inflammation and attenuate the microvascular and macrovascular complications of obesity and diabetes mellitus highlights novel opportunities for potential therapeutic intervention through the targeting of pro-resolution, rather than anti-inflammatory pathways.

Asymmetric Synthesis and Biological Screening of Quinoxaline-Containing Synthetic Lipoxin A4 Mimetics (QNX-sLXms).

Failure to resolve inflammation underlies many prevalent pathologies. Recent insights have identified lipid mediators, typified by lipoxins (LXs), as drivers of inflammation resolution, suggesting potential therapeutic benefit. We report the asymmetric preparation of novel quinoxaline-containing synthetic-LXA4-mimetics (QNX-sLXms). Eight novel compounds were screened for their impact on inflammatory responses. Structure-activity relationship (SAR) studies showed that (R)-6 (also referred to as AT-02-CT) was the most efficacious and potent anti-inflammatory compound of those tested. (R)-6 significantly attenuated lipopolysaccharide (LPS)- and tumor-necrosis-factor-α (TNF-α)-induced NF-κB activity in monocytes and vascular smooth muscle cells. The molecular target of (R)-6 was investigated. (R)-6 activated the endogenous LX receptor formyl peptide receptor 2 (ALX/FPR2). The anti-inflammatory properties of (R)-6 were further investigated in vivo in murine models of acute inflammation. Consistent with in vitro observations, (R)-6 attenuated inflammatory responses. These results support the therapeutic potential of the lead QNX-sLXm (R)-6 in the context of novel inflammatory regulators.

Targeted deletion of nicotinamide adenine dinucleotide phosphate oxidase 4 from proximal tubules is dispensable for diabetic kidney disease development.

BACKGROUND: The nicotinamide adenine dinucleotide phosphate oxidase isoform 4 (Nox4) mediates reactive oxygen species (ROS) production and renal fibrosis in diabetic kidney disease (DKD) at the level of the podocyte. However, the mitochondrial localization of Nox4 and its role as a mitochondrial bioenergetic sensor has recently been reported. Whether Nox4 drives pathology in DKD within the proximal tubular compartment, which is densely packed with mitochondria, is not yet known. METHODS: We generated a proximal tubular-specific Nox4 knockout mouse model by breeding Nox4flox/flox mice with mice expressing Cre recombinase under the control of the sodium-glucose cotransporter-2 promoter. Subsets of Nox4ptKO mice and their Nox4flox/flox littermates were injected with streptozotocin (STZ) to induce diabetes. Mice were followed for 20 weeks and renal injury was assessed. RESULTS: Genetic ablation of proximal tubular Nox4 (Nox4ptKO) resulted in no change in renal function and histology. Nox4ptKO mice and Nox4flox/flox littermates injected with STZ exhibited the hallmarks of DKD, including hyperfiltration, albuminuria, renal fibrosis and glomerulosclerosis. Surprisingly, diabetes-induced renal injury was not improved in Nox4ptKO STZ mice compared with Nox4flox/flox STZ mice. Although diabetes conferred ROS overproduction and increased the mitochondrial oxygen consumption rate, proximal tubular deletion of Nox4 did not normalize oxidative stress or mitochondrial bioenergetics. CONCLUSIONS: Taken together, these results demonstrate that genetic deletion of Nox4 from the proximal tubules does not influence DKD development, indicating that Nox4 localization within this highly energetic compartment is dispensable for chronic kidney disease pathogenesis in the setting of diabetes.

Potential Targeting of Renal Fibrosis in Diabetic Kidney Disease Using MicroRNAs.

Diabetic kidney disease (DKD) is a major health problem and one of the leading causes of end-stage renal disease worldwide. Despite recent advances, there exists an urgent need for the development of new treatments for DKD. DKD is characterized by the excessive synthesis and deposition of extracellular matrix proteins in glomeruli and the tubulointerstitium, ultimately leading to glomerulosclerosis as well as interstitial fibrosis. Renal fibrosis is the final common pathway at the histological level leading to an end-stage renal failure. In fact, activation of the nuclear factor erythroid 2-related factor 2 pathway by bardoxolone methyl and inhibition of transforming growth factor beta signaling by pirfenidone have been assumed to be effective therapeutic targets for DKD, and various basic and clinical studies are currently ongoing. MicroRNAs (miRNAs) are endogenously produced small RNA molecules of 18-22 nucleotides in length, which act as posttranscriptional repressors of gene expression. Studies have demonstrated that several miRNAs contribute to renal fibrosis. In this review, we outline the potential of using miRNAs as an antifibrosis treatment strategy and discuss their clinical application in DKD.

Therapeutic Potential of Lipoxin A4 in Chronic Inflammation: Focus on Cardiometabolic Disease.

Several studies have shown that failure to resolve inflammation may contribute to the progression of many chronic inflammatory disorders. It has been suggested targeting the resolution of inflammation might be a novel therapeutic approach for chronic inflammatory diseases, including inflammatory bowel disease, diabetic complications, and cardiometabolic disease. Lipoxins [LXs] are a class of endogenously generated mediators that promote the resolution of inflammation. Biological actions of LXs include inhibition of neutrophil infiltration, promotion of macrophage polarization, increase of macrophage efferocytosis, and restoration of tissue homeostasis. Recently, several studies have demonstrated that LXs and synthetic analogues protect tissues from acute and chronic inflammation. The mechanism includes down-regulation of pro-inflammatory cytokines and chemokines (e.g., interleukin-1ß and tumor necrosis factor-α), inhibition of the activation of the master pro-inflammatory pathway (e.g., nuclear factor κ-light-chain-enhancer of activated B cells pathway) and increased release of the pro-resolving cytokines (e.g., interleukin-10). Three generations of LXs analogues are well described in the literature, and more recently a fourth generation has been generated that appears to show enhanced potency. In this review, we will briefly discuss the potential therapeutic opportunity provided by lipoxin A4 as a novel approach to treat chronic inflammatory disorders, focusing on cardiometabolic disease and the current drug development in this area.

Specialized pro-resolving mediators in diabetes: novel therapeutic strategies.

Diabetes mellitus (DM) is an important metabolic disorder characterized by persistent hyperglycemia resulting from inadequate production and secretion of insulin, impaired insulin action, or a combination of both. Genetic disorders and insulin receptor disorders, environmental factors, lifestyle choices and toxins are key factors that contribute to DM. While it is often referred to as a metabolic disorder, modern lifestyle choices and nutrient excess induce a state of systemic chronic inflammation that results in the increased production and secretion of inflammatory cytokines that contribute to DM. It is chronic hyperglycemia and the low-grade chronic-inflammation that underlies the development of microvascular and macrovascular complications leading to damage in a number of tissues and organs, including eyes, vasculature, heart, nerves, and kidneys. Improvements in the management of risk factors have been beneficial, including focus on intensified glycemic control, but most current approaches only slow disease progression. Even with recent studies employing SGLT2 inhibitors demonstrating protection against cardiovascular and kidney diseases, kidney function continues to decline in people with established diabetic kidney disease (DKD). Despite the many advances and a greatly improved understanding of the pathobiology of diabetes and its complications, there remains a major unmet need for more effective therapeutics to prevent and reverse the chronic complications of diabetes. More recently, there has been growing interest in the use of specialised pro-resolving mediators (SPMs) as an exciting therapeutic strategy to target diabetes and the chronic complications of diabetes.

Lipoxins Protect Against Inflammation in Diabetes-Associated Atherosclerosis.

Increasing evidence points to the fact that defects in the resolution of inflammatory pathways predisposes individuals to the development of chronic inflammatory diseases, including diabetic complications such as accelerated atherosclerosis. The resolution of inflammation is dynamically regulated by the production of endogenous modulators of inflammation, including lipoxin A4 (LXA4). Here, we explored the therapeutic potential of LXA4 and a synthetic LX analog (Benzo-LXA4) to modulate diabetic complications in the streptozotocin-induced diabetic ApoE-/- mouse and in human carotid plaque tissue ex vivo. The development of diabetes-induced aortic plaques and inflammatory responses of aortic tissue, including the expression of vcam-1, mcp-1, il-6, and il-1ß, was significantly attenuated by both LXA4 and Benzo-LXA4 in diabetic ApoE-/- mice. Importantly, in mice with established atherosclerosis, treatment with LXs for a 6-week period, initiated 10 weeks after diabetes onset, led to a significant reduction in aortic arch plaque development (19.22 ± 2.01% [diabetic]; 12.67 ± 1.68% [diabetic + LXA4]; 13.19 ± 1.97% [diabetic + Benzo-LXA4]). Secretome profiling of human carotid plaque explants treated with LXs indicated changes to proinflammatory cytokine release, including tumor necrosis factor-α and interleukin-1ß. LXs also inhibited platelet-derived growth factor-stimulated vascular smooth muscle cell proliferation and transmigration and endothelial cell inflammation. These data suggest that LXs may have therapeutic potential in the context of diabetes-associated vascular complications.

Lipoxins Regulate the Early Growth Response-1 Network and Reverse Diabetic Kidney Disease.

Background The failure of spontaneous resolution underlies chronic inflammatory conditions, including microvascular complications of diabetes such as diabetic kidney disease. The identification of endogenously generated molecules that promote the physiologic resolution of inflammation suggests that these bioactions may have therapeutic potential in the context of chronic inflammation. Lipoxins (LXs) are lipid mediators that promote the resolution of inflammation.Methods We investigated the potential of LXA4 and a synthetic LX analog (Benzo-LXA4) as therapeutics in a murine model of diabetic kidney disease, ApoE-/- mice treated with streptozotocin.Results Intraperitoneal injection of LXs attenuated the development of diabetes-induced albuminuria, mesangial expansion, and collagen deposition. Notably, LXs administered 10 weeks after disease onset also attenuated established kidney disease, with evidence of preserved kidney function. Kidney transcriptome profiling defined a diabetic signature (725 genes; false discovery rate P≤0.05). Comparison of this murine gene signature with that of human diabetic kidney disease identified shared renal proinflammatory/profibrotic signals (TNF-α, IL-1ß, NF-κB). In diabetic mice, we identified 20 and 51 transcripts regulated by LXA4 and Benzo-LXA4, respectively, and pathway analysis identified established (TGF-ß1, PDGF, TNF-α, NF-κB) and novel (early growth response-1 [EGR-1]) networks activated in diabetes and regulated by LXs. In cultured human renal epithelial cells, treatment with LXs attenuated TNF-α-driven Egr-1 activation, and Egr-1 depletion prevented cellular responses to TGF-ß1 and TNF-αConclusions These data demonstrate that LXs can reverse established diabetic complications and support a therapeutic paradigm to promote the resolution of inflammation.

RAGE Deletion Confers Renoprotection by Reducing Responsiveness to Transforming Growth Factor-ß and Increasing Resistance to Apoptosis.

Signaling via the receptor of advanced glycation end products (RAGE)-though complex and not fully elucidated in the setting of diabetes-is considered a key injurious pathway in the development of diabetic nephropathy (DN). We report here that RAGE deletion resulted in increased expression of fibrotic markers (collagen I and IV, fibronectin) and the inflammatory marker MCP-1 in primary mouse mesangial cells (MCs) and in kidney cortex. RNA sequencing analysis in MCs from RAGE-/- and wild-type mice confirmed these observations. Nevertheless, despite these gene expression changes, decreased responsiveness to transforming growth factor-ß was identified in RAGE-/- mice. Furthermore, RAGE deletion conferred a more proliferative phenotype in MCs and reduced susceptibility to staurosporine-induced apoptosis. RAGE restoration experiments in RAGE-/- MCs largely reversed these gene expression changes, resulting in reduced expression of fibrotic and inflammatory markers. This study highlights that protection against DN in RAGE knockout mice is likely to be due in part to the decreased responsiveness to growth factor stimulation and an antiapoptotic phenotype in MCs. Furthermore, it extends our understanding of the role of RAGE in the progression of DN, as RAGE seems to play a key role in modulating the sensitivity of the kidney to injurious stimuli such as prosclerotic cytokines.

Perinatal exposure to high dietary advanced glycation end products in transgenic NOD8.3 mice leads to pancreatic beta cell dysfunction.

The contribution of environmental factors to pancreatic islet damage in type 1 diabetes remains poorly understood. In this study, we crossed mice susceptible to type 1 diabetes, where parental male (CD8+ T cells specific for IGRP206-214; NOD8.3) and female (NOD/ShiLt) mice were randomized to a diet either low or high in AGE content and maintained on this diet throughout pregnancy and lactation. After weaning, NOD8.3+ female offspring were identified and maintained on the same parental feeding regimen for until day 28 of life. A low AGE diet, from conception to early postnatal life, decreased circulating AGE concentrations in the female offspring when compared to a high AGE diet. Insulin, proinsulin and glucagon secretion were greater in islets isolated from offspring in the low AGE diet group, which was akin to age matched non-diabetic C57BL/6 mice. Pancreatic islet expression of Ins2 gene was also higher in offspring from the low AGE diet group. Islet expression of glucagon, AGEs and the AGE receptor RAGE, were each reduced in low AGE fed offspring. Islet immune cell infiltration was also decreased in offspring exposed to a low AGE diet. Within pancreatic lymph nodes and spleen, the proportions of CD4+ and CD8+ T cells did not differ between groups. There were no significant changes in body weight, fasting glucose or glycemic hormones. This study demonstrates that reducing exposure to dietary AGEs throughout gestation, lactation and early postnatal life may benefit pancreatic islet secretion and immune infiltration in the type 1 diabetic susceptible mouse strain, NOD8.3.

The Use of Targeted Next Generation Sequencing to Explore Candidate Regulators of TGF-ß1's Impact on Kidney Cells.

Aims/Hypothesis: Transforming growth factor-beta (TGF-ß1) plays an important regulatory role in the progression of chronic kidney failure. Further, damage to kidney glomerular mesangial cells is central to the progression of diabetic nephropathy. The aim of this study was to explore the genetic associations between mRNA, microRNA, and epigenetics in mesangial cells in response to TGF-ß1. Methods: The regulatory effects of TGF-ß1 on mesangial cells were investigated at different molecular levels by treating mesangial cells with TGF-ß1 for 3 days followed by genome-wide miRNA, RNA, DNA methylation, and H3K27me3 expression profiling using next generation sequencing (NGS). Results: Our results provide the first comprehensive, computationally integrated report of RNA-Seq, miRNA-Seq, and epigenomic analyses across all genetic variations, confirming the occurrence of DNA methylation and H3K27me3 in response to TGF-ß1. Our findings show that the expression of KLF7 and Gja4 are involved in TGF-ß1 regulated DNA methylation. Our data also provide evidence of the association between epigenetic changes and the expression of genes closely related to TGF-ß1 regulation. Conclusion: This study has advanced our current knowledge of mechanisms that contribute to the expression of TGF-ß1-regulated genes involved in the pathogenesis of kidney disease. The molecular underpinnings of TGF-ß1 stimulation of kidney cells was determined, thereby providing a robust platform for further target exploration.

Increased liver AGEs induce hepatic injury mediated through an OST48 pathway.

The protein oligosaccharyltransferase-48 (OST48) is integral to protein N-glycosylation in the endoplasmic reticulum (ER) but is also postulated to act as a membrane localised clearance receptor for advanced glycation end-products (AGE). Hepatic ER stress and AGE accumulation are each implicated in liver injury. Hence the objective of this study was to increase the expression of OST48 and examine the effects on hepatic function and structure. Groups of 8 week old male mice (n = 10-12/group) over-expressing the gene for OST48, dolichyl-diphosphooligosaccharide-protein glycosyltransferase (DDOST+/-), were followed for 24 weeks, while randomised to diets either low or high in AGE content. By week 24 of the study, either increasing OST48 expression or consumption of high AGE diet impaired liver function and modestly increased hepatic fibrosis, but their combination significantly exacerbated liver injury in the absence of steatosis. DDOST+/- mice had increased both portal delivery and accumulation of hepatic AGEs leading to central adiposity, insulin secretory defects, shifted fuel usage to fatty and ketoacids, as well as hepatic glycogen accumulation causing hepatomegaly along with hepatic ER and oxidative stress. This study revealed a novel role of the OST48 and AGE axis in hepatic injury through ER stress, changes in fuel utilisation and glucose intolerance.

Protective Effect of let-7 miRNA Family in Regulating Inflammation in Diabetes-Associated Atherosclerosis.

The let-7 miRNA family plays a key role in modulating inflammatory responses. Vascular smooth muscle cell (SMC) proliferation and endothelial cell (EC) dysfunction are critical in the pathogenesis of atherosclerosis, including in the setting of diabetes. Here we report that let-7 levels are decreased in diabetic human carotid plaques and in a model of diabetes-associated atherosclerosis, the diabetic ApoE-/- mouse. In vitro platelet-derived growth factor (PDGF)- and tumor necrosis factor-α (TNF-α)-induced vascular SMC and EC activation was associated with reduced let-7 miRNA expression via Lin28b, a negative regulator of let-7 biogenesis. Ectopic overexpression of let-7 in SMCs inhibited inflammatory responses including proliferation, migration, monocyte adhesion, and nuclear factor-κB activation. The therapeutic potential of restoring let-7 levels using a let-7 mimic was tested: in vitro in SMCs using an endogenous anti-inflammatory lipid (lipoxin A4), ex vivo in murine aortas, and in vivo via tail vein injection in a 24-h murine model. Furthermore, we delivered let-7 mimic to human carotid plaque ex vivo and observed significant changes to the secretome in response to let-7 therapy. Restoration of let-7 expression could provide a new target for an anti-inflammatory approach in diabetic vascular disease.

miR-21 promotes renal fibrosis in diabetic nephropathy by targeting PTEN and SMAD7.

The cytokine transforming growth factor (TGF)-ß1 plays a central role in diabetic nephropathy (DN) with data implicating the miRNA (miR) miR-21 as a key modulator of its prosclerotic actions. In the present study, we demonstrate data indicating that miR-21 up-regulation positively correlates with the severity of fibrosis and rate of decline in renal function in human DN. Furthermore, concomitant analyses of various models of fibrotic renal disease and experimental DN, confirm tubular miR-21 up-regulation. The fibrotic changes associated with increased miR-21 levels are proposed to include the regulation of TGF-ß1-mediated mothers against decapentaplegic homolog 3 (SMAD3)- and phosphoinositide 3-kinase (PI3K)-dependent signalling pathways via co-ordinated repression of mothers against decapentaplegic homolog 7 (SMAD7) and phosphatase and tensin homologue (PTEN) respectively. This represents a previously uncharacterized interaction axis between miR-21 and PTEN-SMAD7. Targeting of these proteins by miR-21 resulted in de-repression of the respective pathways as reflected by increases in SMAD3 and V-Akt murine thymoma viral oncogene homolog 1 (AKT) phosphorylation. Many of the changes typically induced by TGF-ß1, including phosphorylation of signalling mediators, were further enhanced by miR-21. Collectively, these data present a unified model for a key role for miR-21 in the regulation of renal tubular extracellular matrix (ECM) synthesis and accumulation and provide important insights into the molecular pathways implicated in the progression of DN.

Study of microRNA in diabetic nephropathy: isolation, quantification and biological function.

In recent years, several studies have reported dysregulation of microRNA expression in disease with a growing interest focussed on targeting microRNAs as a novel therapy for human disease. This is especially true in diabetic nephropathy where the expression of several microRNAs is dysregulated, contributing to the increased expression and accumulation of extracellular matrix proteins and increased pro-fibrotic signalling, ultimately resulting in renal fibrosis. The development of various techniques and microRNA reagents has enabled work to progress very rapidly in this area. In the present article, the authors describe the methods they have used that have enabled them to contribute to our current understanding of the role of microRNAs in diabetic nephropathy.