Costunolide Inhibits Chronic Kidney Disease Development by Attenuating IKKß/NF-κB Pathway.

Background: Chronic kidney disease (CKD) is a significant worldwide health concern that leads to high mortality rates. The bioactive substance costunolide (CTD) has demonstrated several pharmacological effects and holds promise as a CKD treatment. This study aims to investigate the impact of CTD on CKD and delve into its mechanisms of action. Methods: Unilateral ureteral obstruction (UUO) methods and renal fibrosis mice models were created. Various concentrations of CTD were injected into UUO mice models to investigate the therapeutic effects of CTD on renal fibrosis of mice. Then, renal morphology, pathological changes, and the expression of genes related to fibrosis, inflammation and ferroptosis were analysed. RNA sequencing was utilized to identify the main biological processes and pathways involved in renal injury. Finally, both overexpression and inhibition of IKKß were studied to examine their respective effects on fibrosis and inflammation in both in vitro and in vivo models. Results: CTD treatment was found to significantly alleviate fibrosis, inflammation and ferroptosis in UUO-induced renal fibrosis mice models. The results of RNA sequencing suggested that the IKKß acted as key regulatory factor in renal injury and the expression of IKKß was increased in vitro and in vivo renal fibrosis model. Functionally, down-regulated IKKß expression inhibits ferroptosis, inflammatory cytokine production and collagen deposition. Conversely, IKKß overexpression exacerbates progressive renal fibrosis. Mechanistically, CTD alleviated renal fibrosis and inflammation by inhibiting the expression of IKKß and attenuating IKKß/NF-κB pathway. Conclusion: This study demonstrates that CTD could mitigate renal fibrosis, ferroptosis and inflammation in CKD by modulating the IKKß/NF-κB pathway, which indicates targeting IKKß has an enormous potential for treating CKD.

TREM2 deficiency aggravates renal injury by promoting macrophage apoptosis and polarization via the JAK-STAT pathway in mice.

The triggering receptor expressed on myeloid cells 2 (TREM2) is an immune receptor that affects cellular phenotypes by modulating phagocytosis and metabolism, promoting cell survival, and counteracting inflammation. Its role in renal injury, in particular, unilateral ureteral obstruction (UUO) or ischemia-reperfusion injury (IRI)-induced renal injury remains unclear. In our study, WT and Trem2-/- mice were employed to evaluate the role of TREM2 in renal macrophage infiltration and tissue injury after UUO. Bone marrow-derived macrophages (BMDM) from both mouse genotypes were cultured and polarized for in vitro experiments. Next, the effects of TREM2 on renal injury and macrophage polarization in IRI mice were also explored. We found that TREM2 expression was upregulated in the obstructed kidneys. TREM2 deficiency exacerbated renal inflammation and fibrosis 3 and 7 days after UUO, in association with reduced macrophage infiltration. Trem2-/- BMDM exhibited increased apoptosis and poorer survival compared with WT BMDM. Meanwhile, TREM2 deficiency augmented M1 and M2 polarization after UUO. Consistent with the in vivo observations, TREM2 deficiency led to increased polarization of BMDM towards the M1 proinflammatory phenotype. Mechanistically, TREM2 deficiency promoted M1 and M2 polarization via the JAK-STAT pathway in the presence of TGF-ß1, thereby affecting cell survival by regulating mTOR signaling. Furthermore, cyclocreatine supplementation alleviated cell death caused by TREM2 deficiency. Additionally, we found that TREM2 deficiency promoted renal injury, fibrosis, and macrophage polarization in IRI mice. The current data suggest that TREM2 deficiency aggravates renal injury by promoting macrophage apoptosis and polarization via the JAK-STAT pathway. These findings have implications for the role of TREM2 in the regulation of renal injury that justify further evaluation.

Irisin ameliorates UUO-induced renal interstitial fibrosis through TGF-ß1/periostin/MMP-2 signaling pathway.

Renal fibrosis is the most common pathway in progressive kidney diseases. The unilateral ureteral obstruction (UUO) model is used to induce progressive renal fibrosis. We evaluated the effects of irisin on renal interstitial fibrosis in UUO mice. The GSE121190, GSE36496, GSE42303, and GSE96101 datasets were downloaded from the Gene Expression Omnibus (GEO) database. In total, 656 differentially expressed genes (DEGs) were identified in normal and UUO mouse renal samples. Periostin and matrix metalloproteinase-2 (MMP-2) were selected to evaluate the effect of irisin on renal fibrosis in UUO mice. In UUO mice, irisin ameliorated renal function, decreased the expression of periostin and MMP-2, and attenuated epithelial-mesenchymal transition and extracellular matrix deposition in renal tissues. In HK-2 cells, irisin treatment markedly attenuated TGF-ß1-induced expression of periostin and MMP-2. Irisin treatment also inhibited TGF-ß1-induced epithelial-mesenchymal transition, extracellular matrix formation, and inflammatory responses. These protective effects of irisin were abolished by the overexpression of periostin and MMP-2. In summary, irisin treatment can improve UUO-induced renal interstitial fibrosis through the TGF-ß1/periostin/MMP-2 signaling pathway, suggesting that irisin may be used for the treatment of renal interstitial fibrosis.

MicroRNA-10 Family Promotes Renal Fibrosis through the VASH-1/Smad3 Pathway.

Renal fibrosis (RF) stands as a pivotal pathological process in the advanced stages of chronic kidney disease (CKD), and impeding its progression is paramount for delaying the advancement of CKD. The miR-10 family, inclusive of miR-10a and miR-10b, has been implicated in the development of various fibrotic diseases. Nevertheless, the precise role of miR-10 in the development of RF remains enigmatic. In this study, we utilized both an in vivo model involving unilateral ureteral obstruction (UUO) in mice and an in vitro model employing TGF-ß1 stimulation in HK-2 cells to unravel the mechanism underlying the involvement of miR-10a/b in RF. The findings revealed heightened expression of miR-10a and miR-10b in the kidneys of UUO mice, accompanied by a substantial increase in p-Smad3 and renal fibrosis-related proteins. Conversely, the deletion of these two genes led to a notable reduction in p-Smad3 levels and the alleviation of RF in mouse kidneys. In the in vitro model of TGF-ß1-stimulated HK-2 cells, the co-overexpression of miR-10a and miR-10b fostered the phosphorylation of Smad3 and RF, while the inhibition of miR-10a and miR-10b resulted in a decrease in p-Smad3 levels and RF. Further research revealed that miR-10a and miR-10b, through binding to the 3'UTR region of Vasohibin-1 (VASH-1), suppressed the expression of VASH-1, thereby promoting the elevation of p-Smad3 and exacerbating the progression of RF. The miR-10 family may play a pivotal role in RF.

Bulk and single-cell transcriptome profiling identify potential cellular targets of the long noncoding RNA Gas5 in renal fibrosis.

The long noncoding RNA growth arrest-specific 5 (lncRNA Gas5) is implicated in various kidney diseases. In this study, we investigated the lncRNA Gas5 expression profile and its critical role as a potential biomarker in the progression of chronic kidney disease. Subsequently, we assessed the effect of lncRNA Gas5 deletion on renal fibrosis induced by unilateral ureteral obstruction (UUO). The results indicated that loss of lncRNA Gas5 exacerbates UUO-induced renal injury and extracellular matrix deposition. Notably, the deletion of lncRNA Gas5 had a similar effect on control mice. The fibrogenic phenotype observed in mice lacking lncRNA Gas5 correlates with peroxisome proliferator-activated receptor (PPAR) signaling pathway activation and aberrant cytokine and chemokine reprogramming. Single-cell RNA sequencing analysis revealed key transcriptomic features of fibroblasts after Gas5 deletion, revealing heterogeneous cellular states suggestive of a propensity for renal fibrosis. Our findings indicate that lncRNA Gas5 regulates the differentiation and activation of immune cells and the transcription of key genes in the PPAR signaling pathway. These data offer novel insights into the involvement of lncRNA Gas5 in renal fibrosis, potentially paving the way for innovative diagnostic and therapeutic targets.

Huoxue Jiedu Huayu recipe inhibits macrophage-secreted vascular endothelial growth factor-a on angiogenesis and alleviates renal fibrosis in the contralateral kidneys of unilateral ureteral obstruction rats.

OBJECTIVE：To elucidate the mechanism by which Huoxue Jiedu Huayu recipe (, HJHR) regulates angiogenesis in the contralateral kidney of unilateral ureteral obstruction (UUO) rats and the mechanism by which it reduces of renal fibrosis. METHODS: Male Wistar rats were randomly divided into 4 groups: the sham group, UUO group (180 d of left ureter ligation), UUO plus eplerenone (EPL) group, and UUO plus HJHR group. After 180 d of oral drug administration, blood and contralateral kidneys were collected for analysis. Angiogenesis- and fibrosis-related indexes were detected. RESULTS: HJHR and EPL improved structural damage and renal interstitial fibrosis in the contralateral kidney and reduced the protein expression levels of α-smooth muscle actin (α-SMA), vimentin and collagen I. Moreover, these treatments could reduce the expression of vascular endothelial growth factor-A (VEGFA) by inhibiting the infiltration of macrophages. Furthermore, HJHR and EPL significantly reduced the expression of CD34 and CD105 by downregulating VEGFA production, which inhibited angiogenesis. Finally, the coexpressions of CD34, CD105 and α-SMA were decreased in the HJHR and EPL groups, indicating that endothelial-to-mesenchymal transition was inhibited. CONCLUSIONS: These findings confirm that HJHR alleviates contralateral renal fibrosis by inhibiting VEGFA-induced angiogenesis, encourage the use of HJHR against renal interstitial fibrosis and provide a theoretical basis for the clinical management of patients with CKD.

Eplerenone reduces lymphangiogenesis in the contralateral kidneys of UUO rats.

Inflammation and fibrosis often occur in the kidney after acute injury, resulting in chronic kidney disease and consequent renal failure. Recent studies have indicated that lymphangiogenesis can drive renal inflammation and fibrosis in injured kidneys. However, whether and how this pathogenesis affects the contralateral kidney remain largely unknown. In our study, we uncovered a mechanism by which the contralateral kidney responded to injury. We found that the activation of mineralocorticoid receptors and the increase in vascular endothelial growth factor C in the contralateral kidney after unilateral ureteral obstruction could promote lymphangiogenesis. Furthermore, mineralocorticoid receptor activation in lymphatic endothelial cells resulted in the secretion of myofibroblast markers, thereby contributing to renal fibrosis. We observed that this process could be attenuated by administering the mineralocorticoid receptor blocker eplerenone, which, prevented the development of fibrotic injury in the contralateral kidneys of rats with unilateral ureteral obstruction. These findings offer valuable insights into the intricate mechanisms underlying kidney injury and may have implications for the development of therapeutic strategies to mitigate renal fibrosis in the context of kidney disease.

Early growth response protein 2 promotes partial epithelial-mesenchymal transition by phosphorylating Smad3 during renal fibrosis.

Chronic kidney disease (CKD) is a serious health problem worldwide, which ultimately leads to end-stage renal disease (ESRD). Renal fibrosis is the common pathway and major pathological manifestation for various CKD proceeding to ESRD. However, the underlying mechanisms and effective therapies are still ambiguous. Early growth response 2 (EGR2) is reportedly involved in organ formation and cell differentiation. To determine the role of EGR2 in renal fibrosis, we respectively confirmed the increased expression of EGR2 in kidney specimens from both CKD patients and mice with location in proximal tubules. Genetic deletion of EGR2 attenuated obstructive nephropathy while EGR2 overexpression further promoted renal fibrosis in mice subjected to unilateral ureteral obstruction (UUO) due to extracellular matrix (ECM) deposition mediating by partial epithelial-mesenchymal transition (EMT) as well as imbalance between matrix metalloproteinases (MMPs) and tissue inhibitor of MMPs (TIMPs). We found that EGR2 played a critical role in Smad3 phosphorylation, and inhibition of EGR2 reduced partial EMT leading to blockade of ECM accumulation in cultured human kidney 2 cells (HK2) treated with transforming growth factor ß1 (TGF-ß1). In addition, the transcription co-stimulator signal transducer and activator of transcription 3 (STAT3) phosphorylation was confirmed to regulate the transcription level of EGR2 in TGF-ß1-induced HK2 cells. In conclusion, this study demonstrated that EGR2 played a pathogenic role in renal fibrosis by a p-STAT3-EGR2-p-Smad3 axis. Thus, targeting EGR2 could be a promising strategy for CKD treatment.

Atg5 deficiency in macrophages protects against kidney fibrosis via the CCR6-CCL20 axis.

BACKGROUND: Autophagy is a lysosome-dependent degradation pathway that regulates macrophage activation, differentiation, and polarization. Autophagy related 5 (Atg5) is a key protein involved in phagocytic membrane elongation in autophagic vesicles that forms a complex with Atg12 and Atg16L1. Alterations in Atg5 are related to both acute and chronic kidney diseases in experimental models. However, the role of macrophage-expressed Atg5 in acute kidney injury remains unclear. METHODS: Using a myeloid cell-specific Atg5 knockout (MΦ atg5-/-) mouse, we established renal ischemia/reperfusion and unilateral ureteral obstruction models to evaluate the role of macrophage Atg5 in renal macrophage migration and fibrosis. RESULTS: Based on changes in the serum urea nitrogen and creatinine levels, Atg5 deletion had a minimal effect on renal function in the early stages after mild injury; however, MΦ atg5-/- mice had reduced renal fibrosis and reduced macrophage recruitment after 4 weeks of ischemia/reperfusion injury and 2 weeks of unilateral ureteral obstruction injury. Atg5 deficiency impaired the CCL20-CCR6 axis after severe ischemic kidneys. Chemotactic responses of bone marrow-derived monocytes (BMDMs) from MΦ atg5-/- mice to CCL20 were significantly attenuated compared with those of wild-type BMDMs, and this might be caused by the inhibition of PI3K, AKT, and ERK1/2 activation. CONCLUSIONS: Our data indicate that Atg5 deficiency decreased macrophage migration by impairing the CCL20-CCR6 axis and inhibited M2 polarization, thereby improving kidney fibrosis.

ELABELA-derived peptide ELA13 attenuates kidney fibrosis by inhibiting the Smad and ERK signaling pathways. / ELABELA衍生肽ELA13通过抑制Smad和ERK信号通路减轻肾纤维化.

Kidney fibrosis is an inevitable result of various chronic kidney diseases (CKDs) and significantly contributes to end-stage renal failure. Currently, there is no specific treatment available for renal fibrosis. ELA13 (amino acid sequence: RRCMPLHSRVPFP) is a conserved region of ELABELA in all vertebrates; however, its biological activity has been very little studied. In the present study, we evaluated the therapeutic effect of ELA13 on transforming growth factor-ß1 (TGF-ß1)-treated NRK-52E cells and unilateral ureteral occlusion (UUO) mice. Our results demonstrated that ELA13 could improve renal function by reducing creatinine and urea nitrogen content in serum, and reduce the expression of fibrosis biomarkers confirmed by Masson staining, immunohistochemistry, real-time polymerase chain reaction (RT-PCR), and western blot. Inflammation biomarkers were increased after UUO and decreased by administration of ELA13. Furthermore, we found that the levels of essential molecules in the mothers against decapentaplegic (Smad) and extracellular signal-regulated kinase (ERK) pathways were reduced by ELA13 treatment in vivo and in vitro. In conclusion, ELA13 protected against kidney fibrosis through inhibiting the Smad and ERK signaling pathways and could thus be a promising candidate for anti-renal fibrosis treatment.

Icariin alleviates renal inflammation and tubulointerstitial fibrosis via Nrf2-mediated attenuation of mitochondrial damage.

Tubulointerstitial fibrosis is an inevitable consequence of all progressive chronic kidney disease (CKD) and contributes to a substantial health burden worldwide. Icariin, an active flavonoid glycoside obtained from Epimedium species, exerts potential antifibrotic effect. The study aimed to explore the protective effects of icariin against tubulointerstitial fibrosis in unilateral ureteral obstruction (UUO)-induced CKD mice and TGF-ß1-treated HK-2 cells, and furthermore, to elucidate the underlying mechanisms. The results demonstrated that icariin significantly improved renal function, alleviated tubular injuries, and reduced fibrotic lesions in UUO mice. Furthermore, icariin suppressed renal inflammation, reduced oxidative stress as evidenced by elevated superoxide dismutase activity and decreased malondialdehyde level. Additionally, TOMM20 immunofluorescence staining and transmission electron microscope revealed that mitochondrial mass and morphology of tubular epithelial cells in UUO mice was restored by icariin. In HK-2 cells treated with TGF-ß1, icariin markedly decreased profibrotic proteins expression, inhibited inflammatory factors, and protected mitochondria along with preserving mitochondrial morphology, reducing reactive oxygen species (ROS) and mitochondrial ROS (mtROS) overproduction, and preserving membrane potential. Further investigations demonstrated that icariin could activate nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) pathway both in vivo and in vitro, whereas inhibition of Nrf2 by ML385 counteracted the protective effects of icariin on TGF-ß1-induced HK-2 cells. In conclusion, icariin protects against renal inflammation and tubulointerstitial fibrosis at least partly through Nrf2-mediated attenuation of mitochondrial dysfunction, which suggests that icariin could be developed as a promising therapeutic candidate for the treatment of CKD.

FTO-mediated m6A mRNA demethylation aggravates renal fibrosis by targeting RUNX1 and further enhancing PI3K/AKT pathway.

Chronic kidney disease (CKD) is a global health burden, with ineffective therapies leading to increasing morbidity and mortality. Renal interstitial fibrosis is a common pathway in advanced CKD, resulting in kidney function and structure deterioration. In this study, we investigate the role of FTO-mediated N6-methyladenosine (m6A) and its downstream targets in the pathogenesis of renal fibrosis. M6A modification, a prevalent mRNA internal modification, has been implicated in various organ fibrosis processes. We use a mouse model of unilateral ureteral obstruction (UUO) as an in vivo model and treated tubular epithelial cells (TECs) with transforming growth factor (TGF)-ß1 as in vitro models. Our findings revealed increased FTO expression in UUO mouse model and TGF-ß1-treated TECs. By modulating FTO expression through FTO heterozygous mutation mice (FTO+/- ) in vivo and small interfering RNA (siRNA) in vitro, we observed attenuation of UUO and TGF-ß1-induced epithelial-mesenchymal transition (EMT), as evidenced by decreased fibronectin and N-cadherin accumulation and increased E-cadherin levels. Silencing FTO significantly improved UUO and TGF-ß1-induced inflammation, apoptosis, and inhibition of autophagy. Further transcriptomic assays identified RUNX1 as a downstream candidate target of FTO. Inhibiting FTO was shown to counteract UUO/TGF-ß1-induced RUNX1 elevation in vivo and in vitro. We demonstrated that FTO signaling contributes to the elevation of RUNX1 by demethylating RUNX1 mRNA and improving its stability. Finally, we revealed that the PI3K/AKT pathway may be activated downstream of the FTO/RUNX1 axis in the pathogenesis of renal fibrosis. In conclusion, identifying small-molecule compounds that target this axis could offer promising therapeutic strategies for treating renal fibrosis.

Hypermethylation leads to the loss of HOXA5, resulting in JAG1 expression and NOTCH signaling contributing to kidney fibrosis.

Epigenetic regulations, including DNA methylation, are critical to the development and progression of kidney fibrosis, but the underlying mechanisms remain elusive. Here, we show that fibrosis of the mouse kidney was associated with the induction of DNA methyltransferases and increases in global DNA methylation and was alleviated by the DNA methyltransferase inhibitor 5-Aza-2'-deoxycytidine (5-Aza). Genome-wide analysis demonstrated the hypermethylation of 94 genes in mouse unilateral ureteral obstruction kidneys, which was markedly reduced by 5-Aza. Among these genes, Hoxa5 was hypermethylated at its gene promoter, and this hypermethylation was associated with reduced HOXA5 expression in fibrotic mouse kidneys after ureteral obstruction or unilateral ischemia-reperfusion injury. 5-Aza prevented Hoxa5 hypermethylation, restored HOXA5 expression, and suppressed kidney fibrosis. Downregulation of HOXA5 was verified in human kidney biopsies from patients with chronic kidney disease and correlated with the increased kidney fibrosis and DNA methylation. Kidney fibrosis was aggravated by conditional knockout of Hoxa5 and alleviated by conditional knockin of Hoxa5 in kidney proximal tubules of mice. Mechanistically, we found that HOXA5 repressed Jag1 transcription by directly binding to its gene promoter, resulting in the suppression of JAG1-NOTCH signaling during kidney fibrosis. Thus, our results indicate that loss of HOXA5 via DNA methylation contributes to fibrogenesis in kidney diseases by inducing JAG1 and consequent activation of the NOTCH signaling pathway.

Impaired TFEB-mediated autophagy-lysosome fusion promotes tubular cell cycle G2/M arrest and renal fibrosis by suppressing ATP6V0C expression and interacting with SNAREs.

Increasing evidence suggests that autophagy plays a major role during renal fibrosis. Transcription factor EB (TFEB) is a critical regulator of autophagy- and lysosome-related gene transcription. However, the pathophysiological roles of TFEB in renal fibrosis and fine-tuned mechanisms by which TFEB regulates fibrosis remain largely unknown. Here, we found that TFEB was downregulated in unilateral ureteral obstruction (UUO)-induced human and mouse fibrotic kidneys, and kidney-specific TFEB overexpression using recombinant AAV serotype 9 (rAAV9)-TFEB in UUO mice alleviated renal fibrosis pathogenesis. Mechanically, we found that TFEB's prevention of extracellular matrix (ECM) deposition depended on autophagic flux integrity and its subsequent blockade of G2/M arrest in tubular cells, rather than the autophagosome synthesis. In addition, we together RNA-seq with CUT&Tag analysis to determine the TFEB targeted gene ATP6V0C, and revealed that TFEB was directly bound to the ATP6V0C promoter only at specific site to promote its expression through CUT&Run-qPCR and luciferase reporter assay. Interestingly, TFEB induced autophagic flux integrity, mainly dependent on scaffold protein ATP6V0C-mediated autophagosome-lysosome fusion by bridging with STX17 and VAMP8 (major SNARE complex) by co-immunoprecipitation analysis, rather than its mediated lysosomal acidification and degradation function. Moreover, we further investigated the underlying mechanism behind the low expression of TEFB in UUO-induced renal fibrosis, and clearly revealed that TFEB suppression in fibrotic kidney was due to DNMT3a-associated TFEB promoter hypermethylation by utilizing methylation specific PCR (MSP) and bisulfite-sequencing PCR (BSP), which could be effectively recovered by 5-Aza-2'-deoxycytidine (5A-za) to alleviate renal fibrosis pathogenesis. These findings reveal for the first time that impaired TFEB-mediated autophagosome-lysosome fusion disorder, tubular cell G2/M arrest and renal fibrosis appear to be sequentially linked in UUO-induced renal fibrosis and suggest that DNMT3a/TFEB/ATP6V0C may serve as potential therapeutic targets to prevent renal fibrosis.

Kangxianling formula attenuates renal fibrosis by regulating gut microbiota.

BACKGROUND: Renal fibrosis (RF) produced adverse effect on kidney function. Recently, intestinal dysbiosis is a key regulator that promotes the formation of renal fibrosis. This study will focus on exploring the protective mechanism of Kangxianling Formula (KXL) on renal fibrosis from the perspective of intestinal flora. METHODS: Unilateral Ureteral Obstruction (UUO) was used to construct rats' model with RF, and receive KXL formula intervention for 1 week. The renal function indicators were measured. Hematoxylin-eosin (HE), Masson and Sirus red staining were employed to detect the pathological changes of renal tissue in each group. The expression of α-SMA, Col-III, TGF-ß, FN, ZO-1, and Occuludin was detected by immunofluorescence and immunohistochemistry. Rat feces samples were collected and analyzed for species' diversity using high-throughput sequencing 16S rRNA. RESULTS: Rats in UUO groups displayed poor renal function as well as severe RF. The pro-fibrotic protein expression in renal tissues including α-SMA, Col-III, TGF-ß and FN was increased in UUO rats, while ZO-1 and Occuludin -1 expression was downregulated in colon tissues. The above changes were attenuated by KXL treatment. 16S rRNA sequencing results revealed that compared with the sham group, the increased abundance of pathogenic bacteria including Acinetobacter, Enterobacter and Proteobacteria and the decreased abundance of beneficial bacteria including Actinobacteriota, Bifidobacteriales, Prevotellaceae, and Lactobacillus were found in UUO group. After the administration of KXL, the growth of potential pathogenic bacteria was reduced and the abundance of beneficial bacteria was enhanced. CONCLUSION: KXL displays a therapeutical potential in protecting renal function and inhibiting RF, and its mechanism of action may be associated with regulating intestinal microbiota.

Integrative transcriptomic and proteomic profile revealed inhibition of oxidative phosphorylation and peroxisomes during renal interstitial fibrosis.

Effective therapies of chronic kidney disease (CKD) are lacking due to the unclear molecular pathogenesis. Previous single omics-studies have described potential molecular regulation mechanism of CKD only at the level of transcription or translation. Therefore, this study generated an integrated transcriptomic and proteomic profile to provide deep insights into the continuous transcription-translation process during CKD. The comprehensive datasets identified 14,948 transcripts and 6423 proteins, 233 up-regulated and 364 down-regulated common differentially expressed genes of transcriptome and proteome were selected to further combined bioinformatics analysis. The obtained results revealed reactive oxygen species (ROS) metabolism and antioxidant system due to imbalance of mitochondria and peroxisomes were significantly repressed in CKD. Overall, this study presents a valuable multi-omics analysis that sheds light on the molecular mechanisms underlying CKD. SIGNIFICANCE: Chronic kidney disease (CKD) is a progressive and irreversible condition that results in abnormal kidney function and structure, and is ranked 18th among the leading causes of death globally, leading to a significant societal burden. Hence, there is an urgent need for research to detect new, sensitive, and specific biomarkers. Omics-based studies offer great potential to identify underlying disease mechanisms, aid in clinical diagnosis, and develop novel treatment strategies for CKD. Previous studies have mainly focused on the regulation of gene expression or protein synthesis in CKD, thereby compelling us to conduct a meticulous analysis of transcriptomic and proteomic data from the UUO mouse model. Here, we have performed a unified analysis of CKD model by integrating transcriptomes and protein suites for the first time. Our study contributes to a deeper understanding of the pathogenesis of CKD and provides a basis for subsequent disease management and drug development.

Demethylzeylasteral protects against renal interstitial fibrosis by attenuating mitochondrial complex I-mediated oxidative stress.

ETHNOPHARMACOLOGICAL RELEVANCE: Renal interstitial fibrosis (RIF) is a main pathological process in chronic kidney disease (CKD). Demethylzeylasteral (DML), a major component of Tripterygium wilfordii Hook. f., has anti-renal fibrosis effects. However, its mechanism of action remains incompletely understood. AIM OF THE STUDY: The present study was designed to comprehensively examine the effects of DML on RIF and the underlying mechanisms. MATERIALS AND METHODS: Pathological experiments were performed to determine the therapeutic effect of DML on a mouse model of UUO-induced RIF. To determine the novel mechanisms underlying the therapeutic effects of DML against RIF, a comprehensive transcriptomics analysis was performed on renal tissues, which was further verified by a series of experiments. RESULTS: Pathological and immunohistochemical staining showed that DML inhibited UUO-induced renal damage and reduced the expression of fibrosis-related proteins in mice. Transcriptomic analysis revealed that the partial subunits of mitochondrial complex (MC) I and II may be targets by which DML protects against RIF. Furthermore, DML treatment reduced mitochondrial reactive oxygen species (ROS) levels, consequently promoting ATP production and mitigating oxidative stress-induced injury in mice and cells. Notably, this protective effect was attributed to the inhibition of MC I activity, suggesting a crucial role for this specific complex in mediating the therapeutic effects of DML against RIF. CONCLUSIONS: This study provides compelling evidence that DML may be used to treat RIF by effectively suppressing mitochondrial oxidative stress injury mediated by MC I. These findings offer valuable insights into the pharmacological mechanisms of DML and its potential clinical application for patients with CKD.

LncRNA Meg3 Aggravates Renal Fibrosis Caused by Unilateral Ureteral Obstruction in Rats by Activating the Hedgehog Pathway.

BACKGROUND: The hedgehog signaling pathway exerts vital functions in regulating epithelial-to-mesenchymal transition (EMT) in renal interstitial fibrosis (RIF). It was reported that lncRNA-maternally expressed gene 3 (lncRNA Meg3) can regulate hepatic fibrosis by regulating the expression of smoothened (Smo) in the hedgehog signaling pathway. However, the specific role of lncRNA Meg3 in renal fibrosis resulting from unilateral ureteral obstruction (UUO) by regulating the hedgehog signaling pathway has not been reported. Hence, this research aimed to expound the effects of lncRNA Meg3 on renal fibrosis induced by UUO in rats via the hedgehog pathway. METHODS: Peripheral blood was collected from patients with chronic kidney disease (CKD, CKD group) and healthy volunteers (Normal group) at the same period. In addition, 6-week-old male Sprague-Dawley (SD) rats were divided to Sham, UUO, UUO+shRNA Negative control (shNC), and UUO+sh-Meg3 groups, and their kidney tissues and serum were gathered. Next, quantitative real-time polymerase chain reaction (qRT-PCR) was employed for detecting the lncRNA Meg3 expression level in the serum of patients and renal tissue of rats; kits for testing levels of blood urea nitrogen (BUN), creatinine (Cr), hydroxyproline (HYP), and 24-hour urine protein (24-up) in rats of each group; hematoxylin and eosin (HE) staining and Masson staining for observing kidney tissue and renal fibrosis level in rats; western blot for measuring levels of collagen type III (Col III), α-Smooth muscle actin (α-SMA), fibronectin, E-cadherin, sonic hedgehog (Shh), patched (Ptch) protein, smoothened (Smo) protein and glioma-associated oncogene homolog 1 (Gli1) protein expression. RESULTS: LncRNA Meg3 was highly expressed in CKD patients and UUO rats (p < 0.01). In contrast to the UUO+shNC group, knocking down lncRNA Meg3 improved renal injury, relieved pathological renal lesions, and reduced kidney fibrosis and related protein levels. It inhibited the hedgehog pathway in kidney tissues of UUO rats (p < 0.05 and p < 0.01). CONCLUSIONS: LncRNA Meg3 can aggravate UUO-induced rat renal fibrosis by activating the hedgehog pathway.

Beclin-1-Derived Peptide MP1 Attenuates Renal Fibrosis by Inhibiting the Wnt/ß-Catenin Pathway.

Renal fibrosis is distinguished by the abnormal deposition of extracellular matrix and progressive loss of nephron function, with a lack of effective treatment options in clinical practice. In this study, we discovered that the Beclin-1-derived peptide MP1 significantly inhibits the abnormal expression of fibrosis and epithelial-mesenchymal transition-related markers, including α-smooth muscle actin, fibronectin, collagen I, matrix metallopeptidase 2, Snail1, and vimentin both in vitro and in vivo. H&E staining was employed to evaluate renal function, while serum creatinine (Scr) and blood urea nitrogen (BUN) were used as main indices to assess pathologic changes in the obstructed kidney. The results demonstrated that daily treatment with MP1 during the 14-day experiment significantly alleviated renal dysfunction and changes in Scr and BUN in mice with unilateral ureteral obstruction. Mechanistic research revealed that MP1 was found to have a significant inhibitory effect on the expression of crucial components involved in both the Wnt/ß-catenin and transforming growth factor (TGF)-ß/Smad pathways, including ß-catenin, C-Myc, cyclin D1, TGF-ß1, and p-Smad/Smad. However, MP1 exhibited no significant impact on either the LC3II/LC3I ratio or P62 levels. These findings indicate that MP1 improves renal physiologic function and mitigates the fibrosis progression by inhibiting the Wnt/ß-catenin pathway. Our study suggests that MP1 represents a promising and novel candidate drug precursor for the treatment of renal fibrosis. SIGNIFICANCE STATEMENT: This study indicated that the Beclin-1-derived peptide MP1 effectively mitigated renal fibrosis induced by unilateral ureteral obstruction through inhibiting the Wnt/ß-catenin pathway and transforming growth factor-ß/Smad pathway, thereby improving renal physiological function. Importantly, unlike other Beclin-1-derived peptides, MP1 exhibited no significant impact on autophagy in normal cells. MP1 represents a promising and novel candidate drug precursor for the treatment of renal fibrosis focusing on Beclin-1 derivatives and Wnt/ß-catenin pathway.

Activation of the YAP/KLF5 transcriptional cascade in renal tubular cells aggravates kidney injury.

The Hippo/YAP pathway plays a critical role in tissue homeostasis. Our previous work demonstrated that renal tubular YAP activation induced by double knockout (dKO) of the upstream Hippo kinases Mst1 and Mst2 promotes tubular injury and renal inflammation under basal conditions. However, the importance of tubular YAP activation remains to be established in injured kidneys in which many other injurious pathways are simultaneously activated. Here, we show that tubular YAP was already activated 6 h after unilateral ureteral obstruction (UUO). Tubular YAP deficiency greatly attenuated tubular cell overproliferation, tubular injury, and renal inflammation induced by UUO or cisplatin. YAP promoted the transcription of the transcription factor KLF5. Consistent with this, the elevated expression of KLF5 and its target genes in Mst1/2 dKO or UUO kidneys was blocked by ablation of Yap in tubular cells. Inhibition of KLF5 prevented tubular cell overproliferation, tubular injury, and renal inflammation in Mst1/2 dKO kidneys. Therefore, our results demonstrate that tubular YAP is a key player in kidney injury. YAP and KLF5 form a transcriptional cascade, where tubular YAP activation induced by kidney injury promotes KLF5 transcription. Activation of this cascade induces tubular cell overproliferation, tubular injury, and renal inflammation.