Kidney Fibrosis Research Articles

Nephropathy II Decoction Attenuates Renal Fibrosis via Regulating TLR4 and Gut Microbiota Along the Gut-Kidney Axis.

Nephropathy II Decoction (NED) is a widely used Chinese medicinal formulation for managing chronic kidney disease (CKD). Despite its extensive application, the precise mechanisms underlying its therapeutic effects remain poorly understood. This study aims to elucidate the role of NED in attenuating renal fibrosis and to explore its impact on the gut-kidney axis. The principal constituents of NED were analyzed using ultra-performance LC-tandem mass spectrometry (UPLC-MS/MS). A bilateral renal ischemia-reperfusion injury (bIRI) model was employed to induce fibrosis. RT-qPCR was utilized to assess the expression of mRNA related to the toll-like receptor 4 (TLR4) and myeloid differentiation factor 88 (MyD88) and nuclear factor-κB (NF-κB) signaling pathway. Western blotting analysis was performed to identify changes in renal fibrosis markers, TLR4/MyD88/NF-κB pathway proteins, and the colon proteins ZO-1 and Occludin-1. Serum levels of uremic toxins were quantified using enzyme-linked immunosorbent assay (ELISA), and 16S ribosomal RNA (rRNA) gene sequencing was conducted to explore changes in the gut microbiome of the mice. Our study demonstrated that mice in the NED group exhibited reduced serum creatinine, blood urea nitrogen, and urinary protein levels, alongside improvements in kidney damage and a decrease in renal fibrosis markers. In the bIRI group, TLR4/MyD88/NF-κB protein and mRNA levels, as well as intestinal tight junction proteins and enterogenic uremic toxins, were significantly reduced. NED treatment reversed these changes and modified the gut microbiota. Furthermore, fecal microbial transplantation (FMT) alleviated kidney damage and fibrosis in bIRI mice. In summary, NED ameliorates kidney injury and fibrosis by modulating the gut microbiota and may further attenuate fibrosis through the inhibition of TLR4 expression, thereby influencing the gut-kidney axis.

Inhibition of SIRT1/HSF1 pathway contributes to doxorubicin-induced nephrotoxicity in ovarian tumor-bearing mice.

Doxorubicin (DOX) is a common drug used in chemotherapy to treat for advanced ovarian cancer, but it can cause organ damage, particularly to the kidneys. This study aimed to investigate whether the SIRT1/HSF1 pathway is associated with DOX-induced nephrotoxicity. Bioinformatics analysis was performed using single-cell RNA sequencing (scRNA-seq) data from DOX-treated kidneys to investigate the potential mechanism of DOX-induced renal damage. To explore the role of HSF1 in DOX-induced nephrotoxicity, the lentivirus HSF1 (Lv-HSF1) was injected after tumor implantation, followed by DOX administration. DOX prevented ovarian tumor growth but caused renal injury in mice, as evidenced by elevated UACR, increased blood BUN levels, and abnormalities in kidney structure and fibrosis. Bioinformatic analysis revealed fewer podocytes in the kidneys of DOX-exposed mice than in those of control mice, which was further confirmed by examining renal tissue and murine podocyte cells. Gene set enrichment analysis revealed significant enrichment of HSF1-dependent transactivation and HSF1 activation pathways specifically within podocytes obtained from DOX-treated mice, which was also validated in renal tissue samples. Furthermore, HSF1A attenuated DOX-induced podocyte injury in vitro. Lv-HSF1-targeted podocytes mitigate DOX-induced podocyte injury in vivo. Notably, SIRT1 expression was significantly downregulated in both kidney tissues and podocytes subjected to DOX treatment. The observed damage to podocytes induced by DOX may be attributed to an increase in HSF1 acetylation facilitated through the downregulation of SIRT1, a process that can be counteracted by the administration of the SIRT1 agonist RSV. Collectively, these findings demonstrated that suppression of the SIRT1/HSF1 signaling pathway contributes to DOX-mediated nephrotoxicity in mice bearing ovarian tumors.

Krüppel-Like Factor 6 Induces RNA Polymerase II Subunit RPB1 to Promote Kidney Injury.

Initial proximal tubule cell injury and dedifferentiation contribute to acute kidney injury (AKI), and persistent dedifferentiation drives fibrosis and chronic kidney disease (CKD). Proximal tubule-specific knockdown of zinc-finger transcription factor Krüppel-like factor 6 (Klf6) attenuates the AKI to CKD transition. Our aim was to study the early transcriptional mechanisms by which KLF6 induction exacerbates proximal tubular injury and eventual fibrosis. Aristolochic acid I-treated wild-type and KLF6 overexpression mice underwent single nucleus (sn)RNA-seq and snATAC-seq (acute phase) and assessment of kidney function, injury, and fibrosis (remodeling phase). POLR2A was knocked down in human kidney cells, and cell number, gene expression, differentiation, DNA damage, and cell cycle assessed. Kidney sections from fibrotic mouse models and human CKD secondary to aristolochic acid and diabetes were assessed for RPB1 expression. snRNA-seq identified an injured proximal tubule cluster with high expression of Klf6 and RNA polymerase II subunit a (Polr2a) encoding RPB1. After injury, RPB1-positive cells accumulated and were associated with dedifferentiated proximal tubules. POLR2A knockdown in injured cells increased cell death, but reduced inflammatory and fibrotic gene expression, dedifferentiation, DNA damage, and G2/M cell cycle arrest, with a transcriptional switch from long genes to short genes. snATAC-seq demonstrated an open chromatin region in Polr2a intron 1 in injured proximal tubule cells, containing a KLF6 binding site. Knockdown of KLF6 reduced POLR2A induction, whilst proximal tubule-specific KLF6 further increased Polr2a levels after injury. Mice with tubule-specific KLF6 induction had more RPB1-positive proximal tubules and more injury post-AKI. Human kidney samples with DNA-damage induced CKD and diabetic kidney disease also had high POLR2A/RPB1 expression in dedifferentiated proximal tubule cells. Prolonged high expression of RPB1 is associated with dedifferentiated proximal tubule cells. Mice overexpressing KLF6 had higher expression of RPB1 and worse kidney injury after DNA damage.

CCN2 Activates Cellular Senescence Leading to Kidney Fibrosis in Folic Acid-Induced Experimental Nephropathy

Cellular communication network factor 2 (CCN2, also known as CTGF) is a complex protein that regulates numerous cellular functions. This biomolecule exhibits dual functions, depending on the context, and can act as a matricellular protein or as a growth factor. CCN2 is an established marker of fibrosis and a well-known mediator of kidney damage, involved in the regulation of inflammation, extracellular matrix remodeling, cell death, and activation of tubular epithelial cell (TECs) senescence. In response to kidney damage, cellular senescence mechanisms are activated, linked to regeneration failure and progression to fibrosis. Our preclinical studies using a total conditional CCN2 knockout mouse demonstrate that CCN2 plays a significant role in the development of a senescence phenotype after exposure to a nephrotoxic agent. CCN2 induces cell growth arrest in TECs, both in the early phase and in the chronic phase of folic acid nephropathy (FAN), associated with cell-death/necroinflammation and fibrosis, respectively. Renal CCN2 overexpression was found to be linked to excessive collagen accumulation in tubulointerstitial areas, microvascular rarefaction, and a decline in renal function, which were observed three weeks following the initial injury. All these findings were markedly diminished in conditional CCN2 knockout mice. In the FAN model, injured senescent TECs are associated with microvascular rarefaction, and both were modulated by CCN2. In primary cultured endothelial cells, as previously described in TECs, CCN2 directly induced senescence. The findings collectively demonstrate the complexity of CCN2, highlight the pivotal role of cellular senescence as an important mechanism in renal injury, and underscore the critical function of this biomolecule in kidney damage progression.

Therapeutic potential of voltage-dependent potassium channel subtype 1.3 blockade in alleviating macrophage-related renal inflammation and fibrogenesis

Macrophage polarization and infiltration are notable characteristics of kidney injury and fibrosis. Although voltage-dependent potassium channel subtype (Kv) 1.3 is involved in macrophage-induced inflammation, its precise mechanism has not been elucidated. Therefore, this study aimed to explore the role of Kv1.3 in renal injury and macrophage polarization. Herein, mouse models of kidney injury were established through unilateral ureteral obstruction (UUO) and ischemia-reperfusion injury (IRI). For intervention, a selective Kv1.3 blocker, margatoxin (MgTx), was administered intraperitoneally. Blood and kidney samples were collected on days 3 and 7 following UUO surgery to evaluate renal Kv1.3 expression, kidney injury, macrophage polarization changes, cytokine levels, phosphorylation of extracellular signal-regulated kinase (ERK) and nuclear factor kappa-light-chain-enhancer of activated B (NF-κB). Kidney samples were also collected 24 h after IRI to assess kidney injury and evaluate renal Kv1.3 expression, as well as the phosphorylation levels of ERK and NF-κB. Histological analysis of MgTx-treated UUO and IRI mice revealed that Kv1.3 inhibition markedly alleviated renal damage induced by UUO and IRI, substantially reducing the levels of myofibroblast markers, specifically α-smooth muscle actin and transforming growth factor-β1. In UUO mice, Kv1.3 expression and proportions of monocyte-derived cells in peripheral blood and M1 macrophages notably increased but reversed after MgTx treatment, indicating diminished macrophage infiltration. Additionally, MgTx treatment downregulated various M1-related proinflammatory markers, including tumor necrosis factor-α, inducible nitric oxide synthase, and interleukin (IL)-1β, and upregulated M2-associated markers such as IL-10, arginase-1, and CD206. Moreover, Kv1.3 overexpression in THP-1 cells upregulated M1 macrophage markers and proinflammatory cytokines, enhanced their migratory ability. This indicates an increased polarization towards the M1 phenotype, which correlates with impaired renal tubular epithelial cells. Notably, Kv1.3 upregulation both in vivo and in vitro led to increased phosphorylation of ERK and NF-κB, possibly promoting M1 macrophage polarization. This study establishes Kv1.3 as a pivotal regulator of renal fibrosis and macrophage polarization, showing that its inhibition leads to reduced infiltration and migration of M1 macrophages, mitigation of renal injury via suppression of ERK/NF-κB signaling. Altogether, these findings suggest the potential of Kv1.3 as a promising therapeutic target for renal fibrosis.

The VEGF decoy receptor soluble Fms-like tyrosine kinase 1 binds to macrophages

BackgroundSoluble Fms-like Tyrosine kinase-1 (sFLT1) is a native inhibitor of VEGF, best known for its antiangiogenic effects in preeclampsia. sFLT1 also reduces chronic inflammation and promotes tissue repair. In experimental diabetic nephropathy, we previously found that sFLT1 ameliorates kidney fibrosis and reduces the infiltration of macrophages. How sFLT1 regulates inflammation is still incompletely understood. Based on the direct association of sFLT1 with various cell types, we here studied whether sFLT1 interacts with macrophages to modulate inflammation.MethodsUsing various macrophage cell lines, sFLT1 cell surface binding was detected with flow cytometry. Enzyme studies, mass spectrometry and RNAseq were employed to identify potential sFLT1 cell surface interactors and effects of sFLT1 on macrophage signaling.ResultsSoluble FLT1 binds to primary macrophages, THP-1 and RAW264.7 macrophages in vitro. Alternative activation with IL-4 increases sFLT1 binding in THP-1 macrophages, whereas proinflammatory activation with IFN-γ and LPS decreases binding. Binding of sFLT1 depends on heparan sulphates, and colocalizes with the membrane heparin sulfate proteoglycan neuropilin-1. Incubation with sFLT1 reduces the gene expression of chemokine receptors.ConclusionOur results show that sFLT1, while typically associated with angiogenesis, also directly interacts with macrophages. Alternative activation of macrophages by IL-4 strongly increases binding of sFLT1 to the cell surface membrane, possibly via the VEGF co-receptor neuropilin-1. Considering sFLT1’s anti-inflammatory effects in animal studies, our findings indicate a novel function for sFLT1 to directly control anti-inflammatory macrophage function.

Uncoupling protein 1-mediated protective effects of β3-adrenergic receptor agonist on kidney fibrosis via promoting adipose tissue browning in diabetic mice.

Uncoupling protein 1-mediated protective effects of β3-adrenergic receptor agonist on kidney fibrosis via promoting adipose tissue browning in diabetic mice.

A bioprinted and scalable model of human tubulo-interstitial kidney fibrosis.

Chronic kidney disease (CKD) affects more than 10% of the global population. As kidney function negatively correlates with the presence of interstitial fibrosis, the development of new anti-fibrotic therapies holds promise to stabilize functional decline in CKD patients. The goal of the study was to generate a scalable bioprinted 3-dimensional kidney tubulo-interstitial disease model of kidney fibrosis. We have generated novel human PDGFRβ+ pericytes, CD10+ epithelial and CD31+ endothelial cell lines and compared their transcriptomic signature to their in vivo counterpart using bulk RNA sequencing in comparison to human kidney single cell RNA-sequencing datasets. This comparison indicated that the novel cell lines still expressed kidney cell specific genes and shared many features with their native cell-state. PDGFRβ+ pericytes showed three-lineage differentiation capacity and differentiated towards myofibroblasts following TGFβ treatment. We utilized a fibrinogen/gelatin-based hydrogel as bioink and confirmed a good survival rate of all cell types within the bioink after printing. We then combined all three cells in a bioprinted model using separately printed compartments for tubule epithelium, and interstitial endothelium and pericytes. We confirmed that this 3D printed model allows to recapitulate key disease driving epithelial-mesenchymal crosstalk mechanisms of kidney fibrosis since injury of epithelial cells prior to bioprinting resulted in myofibroblast differentiation and fibrosis driven by pericytes after bioprinting. The bioprinted model was also scalable up to a 96-well format.

Molecular mechanisms of andrographolide-induced kidney injury and senescence via SIRT3 inhibition.

Molecular mechanisms of andrographolide-induced kidney injury and senescence via SIRT3 inhibition.

Low-dose bisphenol A plus arsenite: Continuous or intermittent exposures in Sprague-Dawley rats; Effects on kidney oxidative stress, DNA damage, ferroptosis, and fibrosis

Low-dose bisphenol A plus arsenite: Continuous or intermittent exposures in Sprague-Dawley rats; Effects on kidney oxidative stress, DNA damage, ferroptosis, and fibrosis

ASK1 limits kidney glucose reabsorption, growth, and mid-late proximal tubule KIM-1 induction when diabetes and Western diet are combined with SGLT2 inhibition.

Selonsertib is an apoptosis signal-regulating kinase 1 inhibitor (ASK1i) that attenuated the decline in creatinine-based estimated GFR in humans with type 2 diabetes and kidney disease but increased the rate of acute kidney injury. This study explored the individual and combined kidney effects of selonsertib and the antihyperglycemic sodium-glucose cotransporter 2 inhibitor (SGLT2i) dapagliflozin in Western diet-fed male Akita mice, a murine model of early type 1 diabetes mellitus showing signs of systemic but no kidney inflammation. ASK1i reduced elevated plasma levels of proinflammatory cytokines/chemokines (IL-6, MCP1/CCL2, KC/CXCL1, and IP-10/CXCL10) without significantly changing hyperglycemia, glomerular hyperfiltration, and albuminuria or affecting the blood glucose and glomerular hyperfiltration-lowering effect of SGLT2i. A potential sign of tubular stress, SGLT2i modestly upregulated kidney cortex transcription of proinflammatory and profibrotic genes and distal tubule injury marker Ngal. Adding ASK1i to SGLT2i lowered the transcription of many of these genes, including Ngal. However, ASK1i enhanced kidney glucose reabsorption independent of SGLT2i, and combined ASK1i + SGLT2i increased kidney weight by 30%. This was associated with and positively correlated with the upregulation of the tubular stress/injury marker KIM-1, primarily in the mid-to-late proximal tubule. Combined ASK1i + SGLT2i increased the tubular injury score but not signs of kidney inflammation or fibrosis beyond a robust increase in kidney mRNA expression of Il6, Ccl2 (Mcp1), and Timp1, associated with increased plasma IL-6 levels. The data support the hypothesis that housekeeping functions of ASK1 limit glucose reabsorption and the associated growth and cellular stress induced in the mid-to-late proximal tubule by combining hyperglycemia and Western diet with SGLT2 inhibition.NEW & NOTEWORTHY Selonsertib is an apoptosis signal-regulating kinase 1 (ASK1) inhibitor that attenuated creatinine-based eGFR decline in humans with type 2 diabetes and kidney disease but increased acute kidney injury rates. Here, we report evidence in a murine model of early type 1 diabetes mellitus that housekeeping functions of ASK1 limit glucose reabsorption and the associated growth and cellular stress induced in the mid-to-late proximal tubule by combining hyperglycemia and Western diet with SGLT2 inhibition.

Morin Hydrate Improves Kidney Functions in DEHP-Intoxicated Mice via NF-kB/TNFα/Oxidative Stress/Apoptosis Pathway.

Di (2-ethylhexyl) phthalate (DEHP) is a plasticiser used in plastic products; and it dissolves easily and leaks into the environment. Due to its lipophilic nature, DEHP accumulates in organisms and can bioaccumulate through food chains. The natural flavonoid, like morin hydrate, possesses various pharmacological properties, including anti-inflammatory, antioxidant, and free radical scavenging. The purpose of this study was to investigate the effect of morin hydrate (MH) on kidney function of DEHP-treated mice. To investigate the underlying processes of the proposed objective, DEHP (500 mg/kg) and DEHP, along with MH at doses of 10 and 100 mg/kg, were administered to Swiss albino mice for 14 days. Our results showed that MH treatment improved kidney function by decreasing creatinine and urea levels in DEHP-intoxicated mice. Furthermore, the MH also alleviates DEHP-induced kidney fibrosis and kidney histoarchitecture. DEHP-mediated oxidative stress and stimulated apoptosis in the kidney were also mitigated by MH treatment. The elevated expression of NF-kB/TNF-α by the DEHP treatment was also down-regulated by the MH treatment. In addition, the abundance of HSP70 increased in the kidneys after DEHP treatment, and MH treatment also decreased the abundance of HSP70 in the kidneys. In conclusion, DEHP treatment caused kidney toxicity in mice, and MH mitigates the kidney functions via modulating NF-kB/TNF-α/oxidative stress/apoptosis pathway. Our findings provide the new findings that MH protects the kidneys from DEHP intoxication, highlighting the potential of MH as a protective treatment for DEHP toxicity and offering hope for future research and treatments.

Albiflorin inhibits inflammation to improve liver fibrosis by targeting the CXCL12/CXCR4 axis in mice

Liver fibrosis is a common pathological feature of chronic hepatic injury that currently lacks effective therapeutic interventions. Albiflorin (ALB), a pinane-type monoterpene derived from Paeonia lactiflora Pall, has notable anti-inflammatory and hepatoprotective effects. However, the potential role of ALB against liver fibrosis is largely unknown. In this study, we discovered that ALB significantly inhibited CCl4-induced liver fibrosis in mice. This was evidenced by improvements in liver and kidney function indexes, fibrosis indicators, and histopathological findings. In vitro studies also showed that ALB inhibited TGF-β1-induced LX-2 cell activation and reduced the expression of α-SMA and collagen I. Additionally, we found that ALB mitigates inflammation and ameliorates liver fibrosis by targeting the CXCL12/CXCR4 axis, as confirmed using the CXCR4 inhibitor AMD3100 in CCl4-treated mice. Notably, combining ALB with metformin (MET) enhanced the inhibition of liver fibrosis progression. These findings highlight that ALB exerts anti-liver fibrosis effects by targeting the CXCL12/CXCR4 axis, underscoring its potential as a standalone treatment or as an adjuvant therapy.

Mineralocorticoid receptor antagonists promote renal immunosenescence.

Chronic kidney disease (CKD) is often associated with chronic inflammation, influenced by the activation of mineralocorticoid receptors (MR). This review focuses on changes in immune cells and explores the important role that MR antagonists (MRAs), especially the new nonsteroidal MRA, finerenone, play in alleviating renal and cardiac injury by affecting the transformation of stimulated immune cells. We found that MR can promote the transformation of macrophages to M1 pro-inflammatory phenotype through IL-14 receptor and mitogen-activated protein kinase (MAPK)-JNK. MR also activates helper T cells and reduces the generation of regulatory T cells by promoting the interaction between nuclear factor and activator protein-1, increasing the secretion of IL-2 and IL-18, increasing the expression of CD38 and CD69, especially the IL-17/IL-23 axis. The above immune system changes jointly mediate inflammation leading to kidney damage and fibrosis. In addition, we propose that the NLRP3 inflammasome is associated with macrophage imbalance. Preclinical studies indicate that finerenone effectively reduces inflammation and prevents structural kidney damage without significant systemic blood pressure changes. These data will provide some ideas for further research in the field of immune mechanisms in future, and drug research targeting specific targets and channels may also become a new type of diagnostic and treatment measure.

Unilateral Ureteral Obstruction Model for Investigating Kidney Interstitial Fibrosis.

Kidney fibrosis is the final pathological outcome of progressive chronic kidney disease (CKD). The unilateral ureteral obstruction (UUO) model is widely used to elucidate the molecular and cellular mechanisms underlying kidney interstitial fibrosis and to identify potential therapeutic targets. This model is established in mice through surgical ligation of a unilateral ureter, a procedure that is relatively straightforward and easy to perform. However, the UUO mouse model is known to exhibit significant variability and inconsistency, influenced by factors such as mouse strain, age, sex, anesthesia type, duration of surgery, body temperature during the procedure, the operator's surgical skills, feeding conditions, and the overall health status of the mice. Variations in surgical techniques, suture placement, and the duration of obstruction contribute to the variability in outcomes. Additionally, inconsistent sampling of obstructed kidneys further increases variability in the assessment of kidney fibrosis. This study outlines the process of developing the UUO mouse model and evaluating interstitial fibrosis, discusses the technical challenges contributing to the model's unpredictability, and proposes potential solutions. These insights aim to establish a more standardized and universally applicable approach for investigating kidney fibrosis.

Single-cell analysis of proximal tubular cells with different DNA content reveals functional heterogeneity in the acute kidney injury to chronic kidney disease transition.

Single-cell analysis of proximal tubular cells with different DNA content reveals functional heterogeneity in the acute kidney injury to chronic kidney disease transition.

Chymase Inhibition Attenuates Kidney Fibrosis in a Chronic Mouse Model of Renal Ischemia-Reperfusion Injury.

Although various factors contribute to the transition from acute kidney injury (AKI) to chronic kidney disease (CKD), no clinically effective pharmacological treatment has been established. We investigated whether chymase inhibition is effective in preventing renal fibrosis, a key process in the transition from AKI to CKD. Male BALB/c mice were subjected to unilateral ischemia-reperfusion (I/R) injury, and TY-51469, a chymase-specific inhibitor, was administered intraperitoneally at a dose of 10 mg/kg/day for 6 weeks. The 45 min ischemic period followed by 6 weeks of reperfusion resulted in severe renal atrophy. Renal fibrosis was particularly pronounced in the transition region between the cortex and medulla in placebo-treated mice. The expression of mouse mast cell protease 4 (MMCP-4, a mouse chymase) mRNA, the number of chymase-positive mast cells, and fibrosis-related factors, such as transforming growth factor (TGF)-β1 and collagen I, were all significantly increased in I/R-injured kidneys. However, treatment with TY-51469 significantly suppressed fibrosis formation, along with the inhibition of renal chymase and TGF-β1 expression. These findings suggest that chymase inhibition may be a potential therapeutic strategy for preventing the transition from AKI to CKD by reducing fibrosis.

Engineering IL-10 and rapamycin to bind collagen leads to improved anti fibrotic efficacy in lung and kidney fibrosis

Fibrotic diseases are involved in 45% of deaths in the United States. In particular, fibrosis of the kidney and lung are major public health concerns due to their high prevalence and lack of existing treatment options. Here, we harness the pathophysiological features of fibrotic diseases, namely leaky vasculature and aberrant extracellular matrix (ECM) protein deposition (i.e. collagen), to target an anti-fibrotic biologic and a small molecule drug to disease sites of fibrosis, thus improving the therapeutic potential of both the biologic and small molecule in mouse models of both lung and kidney fibrosis. First, we identify and validate two collagen-targeting drug delivery systems that preferentially accumulate in fibrotic organs: von Willebrand Factor’s A3 domain (VWF-A3) and decorin-derived collagen-binding peptide-conjugated micelles (CBP-micelles). We then engineer and recombinantly express novel candidate biologic therapies based on the anti-inflammatory cytokine IL-10: A3-IL-10 and A3-Serum Albumin-IL-10 (A3-SA-IL-10). Simultaneously, we stably encapsulate the potential anti-fibrotic water-insoluble drug, rapamycin, in CBP-micelles. We show that these novel formulations of therapeutics bind to collagen in vitro and that their efficacy in mouse models of lung and kidney fibrosis is improved, compared to free, untargeted drugs. Our results demonstrate that collagen-targeted anti-fibrotic drugs may be next generation therapies of high clinical potential.

SGLT2 inhibitor empagliflozin ameliorates tubulointerstitial fibrosis in DKD by downregulating renal tubular PKM2

Background and objectiveSodium-glucose cotransporter 2 (SGLT2) inhibitors have been shown to prevent the progression of diabetic kidney disease (DKD). However, their impact on renal fibrosis remains largely uninvestigated. This study aimed to explore the effect of SGLT2 inhibitor empagliflozin on renal fibrosis in DKD patients and DKD models, and the molecular mechanisms involved.MethodsKidney samples of DKD patients and DKD models were used in this study. DKD mouse models included STZ-treated CD-1 mice and HFD-fed C57BL/6 mice were all treated with empagliflozin for 6 to 12 weeks. Kidney pathological changes were analysed and fibrotic factors were detected. HK-2 cells were treated with normal glucose (NG), high glucose (HG), or HG with empagliflozin. RNA sequencing was employed to identify the differentially expressed genes. Epithelial–mesenchymal transition (EMT) markers were detected. Binding of transcription factor and target gene was determined using a dual-luciferase reporter assay.ResultsEmpagliflozin significantly ameliorated kidney fibrosis in DKD patients and DKD models. This was evidenced by tubulointerstitial fibrosis reduction observed through PAS and Masson staining, along with fibrotic factors downregulation. RNA sequencing and the subsequent in vitro and in vivo validation identified PKM2 as the most significantly upregulated glycolytic enzyme in DKD patients and models. Empagliflozin downregulated PKM2 and alleviated EMT and renal fibrosis. Importantly, empagliflozin improves fibrosis by downregulating PKM2. The downregulation of PKM2 by empagliflozin was achieved by inhibiting the binding of estrogen-related receptor α at the promoter.ConclusionsEmpagliflozin ameliorates kidney fibrosis via downregulating PKM2 in DKD.

Development and validation of multi-center serum creatinine-based models for noninvasive prediction of kidney fibrosis in chronic kidney disease.

Kidney fibrosis is a key pathological feature in the progression of chronic kidney disease (CKD), traditionally diagnosed through invasive kidney biopsy. This study aimed to develop and validate a noninvasive, multi-center predictive model incorporating machine learning (ML) for assessing kidney fibrosis severity using biochemical markers. This multi-center retrospective study included 598 patients with kidney fibrosis from four hospitals. A training cohort of 360 patients from Shanghai Tongji Hospital was used to develop a predictive nomogram and ML model, with fibrosis severity classified as mild or moderate-to-severe based on Banff scores. Logistic regression identified key predictors, which were incorporated into a nomogram and ML model. An external validation cohort of 238 patients from three additional hospitals was used for model evaluation. Serum creatinine (Scr), estimated glomerular filtration rate (eGFR), parathyroid hormone (PTH), brain natriuretic peptide (BNP), and sex were identified as independent predictors of kidney fibrosis severity. The nomogram demonstrated superior discriminative ability in the training cohort (AUC: 0.89, 95% CI: 0.85-0.92) compared to eGFR (AUC: 0.83, 95% CI: 0.78-0.87) and Scr (AUC: 0.87, 95% CI: 0.83-0.91). Among ML models, the Random Forest (RF) model achieved the highest AUC (0.98). In external validation, the nomogram and RF models maintained robust performance with AUCs of 0.86 and 0.79, respectively. This study presents a validated, noninvasive, multi-center Scr-based machine learning model for assessing kidney fibrosis severity in CKD. The integration of a clinical nomogram and ML approach offers a novel, practical alternative to biopsy for dynamic fibrosis evaluation.