Hepatic-derived BMP9 is involved in hepatic fibrosis-induced kidney injury through inhibition of renal VEGFA.

Clinical observations suggest that acute kidney injury (AKI) occurs in approximately 20-50% of hospitalized cirrhotic patients, suggesting a link between the liver and kidney. Bone morphogenetic protein 9 (BMP9) is a protein produced primarily by the liver and can act on other tissues at circulating systemic levels. Previous studies have demonstrated that controlling abnormally elevated BMP9 in acute liver injury attenuates liver injury; however, reports on whether BMP9 plays a role in liver injury-induced AKI are lacking. By testing we found that liver injury in mice after bile duct ligation (BDL) was accompanied by a significant upregulation of the kidney injury marker kidney injury molecule (KIM-1). Interestingly, all these impairments were alleviated in the kidneys of hepatic BMP9 knockout (BMP9-KO) mice. Peritubular capillary injury is a key process leading to the progression of AKI, and previous studies have demonstrated that vascular endothelial growth factor A (VEGFA) plays a key role in maintaining the renal microvascular system. In animal experiments, we found that high levels of circulating BMP9 had an inhibitory effect on VEGFA expression, while renal tubular epithelial cell injury was effectively attenuated by VEGFA supplementation in the hypoxia-enriched-oxygen (H/R) constructs of the AKI cell model in both humans and mice. Overall, we found that elevated BMP9 in hepatic fibrosis can affect renal homeostasis by regulating VEGFA expression. Therefore, we believe that targeting BMP9 therapy may be a potential means to address the problem of clinical liver fibrosis combined with AKI.

Genetic and pharmacological inhibition of GRPR protects against acute kidney injury via attenuating renal inflammation and necroptosis.

Gastrin-releasing peptide (GRP) binds to its receptor (GRP receptor [GRPR]) to regulate multiple biological processes, but the function of GRP/GRPR axis in acute kidney injury (AKI) remains unknown. In the present study, GRPR is highly expressed by tubular epithelial cells (TECs) in patients or mice with AKI, while histone deacetylase 8 may lead to the transcriptional activation of GRPR. Functionally, we uncovered that GRPR was pathogenic in AKI, as genetic deletion of GRPR was able to protect mice from cisplatin- and ischemia-induced AKI. This was further confirmed by specifically deleting the GRPR gene from TECs in GRPRFlox/Flox//KspCre mice. Mechanistically, we uncovered that GRPR was able to interact with Toll-like receptor 4 to activate STAT1 that bound the promoter of MLKL and CCL2 to induce TEC necroptosis, necroinflammation, and macrophages recruitment. This was further confirmed by overexpressing STAT1 to restore renal injury in GRPRFlox/Flox/KspCre mice. Concurrently, STAT1 induced GRP synthesis to enforce the GRP/GRPR/STAT1 positive feedback loop. Importantly, targeting GRPR by lentivirus-packaged small hairpin RNA or by treatment with a novel GRPR antagonist RH-1402 was able to inhibit cisplatin-induced AKI. In conclusion, GRPR is pathogenic in AKI and mediates AKI via the STAT1-dependent mechanism. Thus, targeting GRPR may be a novel therapeutic strategy for AKI.

Critical Overview of Hepatic Factors That Link Non-Alcoholic Fatty Liver Disease and Acute Kidney Injury: Physiology and Therapeutic Implications.

Non-alcoholic fatty liver disease (NAFLD) is defined as a combination of a group of progressive diseases, presenting different structural features of the liver at different stages of the disease. According to epidemiological surveys, as living standards improve, the global prevalence of NAFLD increases. Acute kidney injury (AKI) is a class of clinical conditions characterized by a rapid decline in kidney function. NAFLD and AKI, as major public health diseases with high prevalence and mortality, respectively, worldwide, place a heavy burden on societal healthcare systems. Clinical observations of patients with NAFLD with AKI suggest a possible association between the two diseases. However, little is known about the pathogenic mechanisms linking NAFLD and AKI, and the combination of the diseases is poorly treated. Previous studies have revealed that liver-derived factors are transported to distal organs via circulation, such as the kidney, where they elicit specific effects. Of note, while NAFLD affects the expression of many hepatic factors, studies on the mechanisms whereby NAFLD mediates the generation of hepatic factors that lead to AKI are lacking. Considering the unique positioning of hepatic factors in coordinating systemic energy metabolism and maintaining energy homeostasis, we hypothesize that the effects of NAFLD are not only limited to the structural and functional changes in the liver but may also involve the entire body via the hepatic factors, e.g., playing an important role in the development of AKI. This raises the question of whether analogs of beneficial hepatic factors or inhibitors of detrimental hepatic factors could be used as a treatment for NAFLD-mediated and hepatic factor-driven AKI or other metabolic disorders. Accordingly, in this review, we describe the systemic effects of several types of hepatic factors, with a particular focus on the possible link between hepatic factors whose expression is altered under NAFLD and AKI. We also summarize the role of some key hepatic factors in metabolic control mechanisms and discuss their possible use as a preventive treatment for the progression of metabolic diseases.

Insulin-like growth factor binding protein 7 promotes acute kidney injury by alleviating poly ADP ribose polymerase 1 degradation.

The novel biomarker, insulin-like growth factor binding protein 7 (IGFBP7), is used clinically to predict different types of acute kidney injury (AKI) and has drawn significant attention as a urinary biomarker. However, as a secreted protein in the circulation of patients with AKI, it is unclear whether IGFBP7 acts as a key regulator in AKI progression, and if mechanisms underlying its upregulation still need to be determined. Here we found that IGFBP7 is highly expressed in the blood and urine of patients and mice with AKI, possibly via a c-Jun-dependent mechanism, and is positively correlated with kidney dysfunction. Global knockout of IGFBP7 ameliorated kidney dysfunction, inflammatory responses, and programmed cell death in murine models of cisplatin-, kidney ischemia/reperfusion-, and lipopolysaccharide-induced AKI. IGFBP7 mainly originated from kidney tubular epithelial cells. Conditional knockout of IGFBP7 from the kidney protected against AKI. By contrast, rescue of IGFBP7 expression in IGFBP7-knockout mice restored kidney damage and inflammation. IGFBP7 function was determined in vitro using recombinant IGFBP7 protein, IGFBP7 knockdown, or overexpression. Additionally, IGFBP7 was found to bind to poly [ADP-ribose] polymerase 1 (PARP1) and inhibit its degradation by antagonizing the E3 ubiquitin ligase ring finger protein 4 (RNF4). Thus, IGFBP7 in circulation acts as a biomarker and key mediator of AKI by inhibiting RNF4/PARP1-mediated tubular injury and inflammation. Hence, over-activation of the IGFBP7/PARP1 axis represents a promising target for AKI treatment.

Stratifin promotes renal dysfunction in ischemic and nephrotoxic AKI mouse models via enhancing RIPK3-mediated necroptosis.

Stratifin (SFN) is a member of the 14-3-3 family of highly conserved soluble acidic proteins, which regulates a variety of cellular activities such as cell cycle, cell growth and development, cell survival and death, and gene transcription. Acute kidney injury (AKI) is prevalent disorder characterized by inflammatory response, oxidative stress, and programmed cell death in renal tubular epithelial cells, but there is still a lack of effective therapeutic target for AKI. In this study, we investigated the role of SFN in AKI and the underlying mechanisms. We established ischemic and nephrotoxic AKI mouse models caused by ischemia-reperfusion (I/R) and cisplatin, respectively. We conducted proteomic and immunohistochemical analyses and found that SFN expression levels were significantly increased in AKI patients, cisplatin- or I/R-induced AKI mice. In cisplatin- or hypoxia/reoxygenation (H/R)-treated human proximal tubule epithelial cells (HK2), we showed that knockdown of SFN significantly reduced the expression of kidney injury marker Kim-1, attenuated programmed cell death and inflammatory response. Knockdown of SFN also significantly alleviated the decline of renal function and histological damage in cisplatin-caused AKI mice in vivo. We further revealed that SFN bound to RIPK3, a key signaling modulator in necroptosis, to induce necroptosis and the subsequent inflammation in cisplatin- or H/R-treated HK2 cells. Overexpression of SFN increased Kim-1 protein levels in cisplatin-treated MTEC cells, which was suppressed by RIPK3 knockout. Taken together, our results demonstrate that SFN that enhances cisplatin- or I/R-caused programmed cell death and inflammation via interacting with RIPK3 may serve as a promising therapeutic target for AKI treatment.

LncRNA MIR31HG is induced by tocilizumab and ameliorates rheumatoid arthritis fibroblast-like synoviocyte-mediated inflammation via miR-214-PTEN-AKT signaling pathway.

Fibroblast-like synoviocytes (FLS) obtained from the joint synovium of rheumatoid arthritis (RA) patients exhibit hyperplasia and aggressive inflammatory phenotypes. This study was designed to explore the anti-inflammatory mechanism of IL-6R inhibitor, tocilizumab, in FLS-mediated inflammation in RA from the perspective of non-coding RNAs (ncRNAs). To this end, we sorted primary FLS obtained from the synovium of patients with RA and cultured them in vitro. The cells were then treated with tocilizumab and subjected to lncRNA- and miRNA-seq to identify the ncRNAs regulated by tocilizumab treatment using bioinformatic analysis and experimental verification. Tocilizumab treatment enhanced the expression of lncRNA MIR31HG and reduced that of micoRNA-214 (miR-214). In addition, miR-214 activated the AKT signaling pathway by directly targeting MIR31HG and PTEN. In addition, the tocilizumab-MIR31HG-miR-214-PTEN-AKT axis regulated the proliferation, migration, and production of inflammatory molecules and matrix metalloproteinases (MMPs) in RA-FLS. Furthermore, co-culture experiments showed that this axis could inhibit the inflammatory phenotype of macrophages and protect chondrocytes. In summary, our study shows that tocilizumab suppresses RA-FLS inflammation by regulating the MIR31HG-miR-214-PTEN-AKT pathway, and presents new insights on RA pathogenesis and potential targets for RA therapy.

Eleutheroside B-loaded poly (lactic-co-glycolic acid) nanoparticles protect against renal fibrosis via Smad3-dependent mechanism.

Although renal fibrosis is a common complication of chronic kidney disease (CKD), effective options for its treatment are currently limited. In this study, we evaluated the renal protective effect and possible mechanism of eleutheroside B. In order to solve the allergic reactions, side effects, and low oral bioavailability of eleutheroside B, we successfully prepared PLGA (poly [lactic-co-glycolic acid])-eleutheroside B nanoparticles (NPs) with the diameter of about 128 nm. In vitro and in vivo results showed that eleutheroside B could inhibit expression levels of α-smooth muscle actin (α-SMA) and collagen I. Molecular docking results showed that eleutheroside B bound to Smad3 and significantly decreased the expression of phospho-Smad3 (p-Smad3). Silencing Smad3 reversed the fibrotic protective effect of eleutheroside B in HK2 cells. Furthermore, small animal imaging showed that NPs can selectively accumulate in the UUO kidneys of mice, and retention time reached as long as 7 days. In conclusion, our results suggested that eleutheroside B is a potential drug to protect renal fibrosis and PLGA-eleutheroside B NPs could facilitate specific targeted therapy for renal fibrosis.

Gypenoside XLIX loaded nanoparticles targeting therapy for renal fibrosis and its mechanism.

Renal fibrosis is the main pathological feature of the occurrence and development of chronic nephropathy. At present, there is no effective treatment, except for renal transplantation and dialysis. Previous studies have shown that nano-preparations can be used as a therapeutic tool to target organs. In this study, we studied the therapeutic effect and mechanism of Chinese medicine monomer Gypenoside (Gyp) XLIX on renal fibrosis and explored the targeting and therapeutic effects of polylactic acid-co-glycoside (PLGA)-Gyp XLIX nanoparticles in unilateral ureteral occlusion (UUO) kidney. Gyp XLIX and PLGA-Gyp XLIX nanoparticles were used to treat UUO mice and Human renal tubular epithelial (HK2) cells stimulated by transforming growth factor-ß (TGF-ß). Histopathological and molecular biological techniques were used to detect the expression of type I collagen and alpha-smooth muscle actin (α-SMA). To investigate the in vivo targeting of PLGA nanoparticles, they were loaded with 1,1-dioctadecyl-3,3,3,3-tetramethylindotricarbocyanine iodide and injected into UUO mice. We evaluated the effect of Gyp XLIX nanoparticles on TGF-ß/Smad3 pathway, a central driver for renal fibrosis in Smad-deficient HK2 cells. Fluorescence imaging showed that the PLGA nanoparticles around 120 nm could be targeted to the UUO kidney. Compared with Gyp XLIX, PLGA-Gyp XLIX nanoparticles could effectively inhibit renal fibrosis and reduce collagen deposition and reduce renal tubular necrosis. Gyp XLIX decreased the phosphorylation of Smad3, but could not further reduce the levels of type I collagen and α-SMA in Smad-deficient cells. This study opens a promising way for targeted drug treatment of renal fibrosis.

Gypenoside XLIX protects against acute kidney injury by suppressing IGFBP7/IGF1R-mediated programmed cell death and inflammation.

BACKGROUND: Acute kidney injury (AKI), characterised by excessive inflammatory cell recruitment and programmed cell death, has a high morbidity and mortality; however, effective and specific therapies for AKI are still lacking. OBJECTIVE: This study aimed to evaluate the renoprotective effects of gypenoside XLIX (Gyp XLIX) in AKI. METHODS: The protective effects of Gyp XLIX were tested in two AKI mouse models established using male C57BL/6 mice (aged 6-8 weeks) by a single intraperitoneal injection of cisplatin (20 mg/kg) or renal ischemia-reperfusion for 40 min. Gyp XLIX was administered intraperitoneally before cisplatin administration or renal ischemia-reperfusion. Renal function, tubular injury, renal inflammation and programmed cell death were evaluated. In addition, the renoprotective effects of Gyp XLIX were also evaluated in cisplatin- or hypoxia-treated tubular epithelial cells. The mechanisms underlying these effects were then explored using RNA sequencing. RESULTS: In vivo, Gyp XLIX substantially suppressed the increase in serum creatinine and blood urea nitrogen levels. Moreover, tubular damage was alleviated by Gyp XLIX as shown by periodic acid-Schiff staining, electron microscopy and molecular analysis of KIM-1. Consistently, we found that Gyp XLIX suppressed renal necroptosis though the RIPK1/RIPK3/MLKL pathway. The anti-inflammatory and antinecroptotic effects were further confirmed in vitro. Mechanistically, RNA sequencing showed that Gyp XLIX markedly suppressed the levels of IGF binding protein 7 (IGFBP7). Co-immunoprecipitation and western blot analysis further showed that Gyp XLIX reduced the binding of IGFBP7 to IGF1 receptor (IGF1R). Additionally, picropodophyllin, an inhibitor of IGF1R, abrogated the therapeutic effects of Gyp XLIX on cisplatin-induced renal cell injury; this finding indicated that Gyp XLIX may function by activating IGF1R-mediated downstream signalling Additionally, we also detected the metabolic distribution of Gyp XLIX after injection; Gyp XLIX had a high concentration in the kidney and exhibited a long retention time. These findings may shed light on the application of Gyp XLIX for AKI treatment clinically. CONCLUSION: Gyp XLIX may serve as a potential therapeutic agent for AKI treatment via IGFBP7/ IGF1R-dependent mechanisms.

Eleutheroside B Protects against Acute Kidney Injury by Activating IGF Pathway.

Acute kidney injury (AKI) is a common, complex, and severe clinical syndrome characterized by rapid decline in renal function, combined with tissue damage. Currently, the prevention and treatment of AKI are focused on symptomatic treatment, rather than treating the underlying causes. Therefore, there is no specific treatment to prevent renal injury except for renal dialysis. In this study, we used cisplatin-induced AKI mouse and human kidney-2 (HK-2) cell models to evaluate the renal protective effect of eleutheroside B, an active compound in traditional Chinese medicines. MTT assay was used to detect the effect of eleutheroside B on proliferation of human HK-2 cells in presence and in absence of cisplatin. Western blot and immunostaining were used to detect the protein level of kidney injury molecule-1 (KIM-1), cleaved caspase-3, receptor-interacting protein kinase (RIPK)-1, and RIPK-3. Real-time PCR was used to detect the mRNA levels of chemokines (like monocyte chemotactic protein 1, MCP-1) and pro-inflammatory cytokines including interleukin-6 (IL-6) and tumor necrosis factor (TNF-α). Flow cytometry assay was used to detect apoptosis of HK-2 cells. In vivo results showed that eleutheroside B reduced the increase in serum creatinine and blood urea nitrogen (BUN) levels in the AKI model. Periodic acid-Schiff staining and Western blot analysis of KIM-1 showed that eleutheroside B alleviated tubular cell injury. Further, eleutheroside B reduced macrophage infiltration and production of inflammatory cytokines, inhibited the activation of nuclear factor (NF)-κB, and inhibited apoptosis and programmed necrosis. The mechanism may be that eleutheroside B can activate the insulin-like growth factor (IGF) pathway and its downstream pathway by downregulating the expression of IGFBP-7, thus promoting cell proliferation. Therefore, our results suggest that eleutheroside B is a potential drug for AKI treatment.

Restoration of E-cadherin by PPBICA protects against cisplatin-induced acute kidney injury by attenuating inflammation and programmed cell death.

E-cadherin is a major component of tubular adherent proteins that maintain intercellular contacts and cell polarity in epithelial tissue. It is involved in pathological processes of renal cell carcinoma and fibrotic diseases via epithelial-mesenchymal transition. Although studies have shown E-cadherin is significantly downregulated in acute kidney injury (AKI), its function in AKI is unknown. Here, we evaluated cell damage and inflammation in cisplatin-stimulated tubular epithelial cell lines after disrupting E-cadherin and restoring it with PPBICA, a small molecule identified by high-throughput screening. We also determined the therapeutic potential of restoring E-cadherin in vivo. Results show cisplatin reduced E-cadherin expression both in mouse kidney and proximal tubular epithelial cell lines (mTECs). PPBICA restored E-cadherin levels, which increased cell viability while attenuating programmed cell death. This may be mediated via deactivation of the RIPK1/RIPK3 axis and decreased caspase3 cleavage. In addition, PPBICA suppressed inflammatory response in cisplatin-treated mTECs, which correlated with suppressed NF-κB phosphorylation and promoter activity. In contrast, disruption of E-cadherin promoted cell damage and inflammation. PPBICA failed to further attenuate kidney damage in E-cadherin knockdown cells, indicating that PPBICA protects against mTECs through E-cadherin restoration. We also found that peritoneal injection of PPBICA in mice prevented loss of renal function and tubular damage by suppressing NF-κB-driven renal inflammation and RIPK-regulated programmed cell death. This was driven by restoration of E-cadherin in cisplatin nephropathy. Additionally, PPBICA attenuated cisplatin-induced kidney damage in an established AKI model, indicating its therapeutic potential in the treatment of AKI. In conclusion, E-cadherin plays functional roles in tubule integrity, programmed cell death, and renal inflammation. Our results underscore the potential of E-cadherin restoration as a novel therapeutic strategy for AKI.

Structural effects on the catalytic activity of carbon-supported magnetite nanocomposites in heterogeneous Fenton-like reactions.

The catalytic reactivity of synthetic bare magnetite nanoparticles, activated carbon supported magnetite (AC-Mt), and graphene oxide supported magnetite (GO-Mt) for heterogeneous Fenton-like oxidation of methylene blue (MB) were compared, in order to investigate how the structural features of the support impact catalytic activity of the nanocomposites. The different effects of AC and GO on MB removal rate, hydroxyl radical (˙OH) production, iron leaching, and surface deactivation have been systematically studied. The rate constant of MB removal by AC-Mt was 0.1161 min-1, one order of magnitude larger than the value of bare magnetite nanoparticles (0.0566 min-1). The higher catalytic activity of AC-Mt might be attributed to the larger reactive surface area of well-dispersed magnetite for ˙OH production and the recharge of the magnetite surface by the AC support via Fe-O-C bonds. However, the removal rate of MB by GO-Mt was one order of magnitude slower than that of bare magnetite nanoparticles under the same experimental conditions, presumably due to the wrapping of GO around magnetite nanoparticles or extensive aggregation of GO-Mt composites. These findings revealed the significant influence of support structure on the catalytic activity of carbon-supported magnetite nanocomposites, which is important for the development of efficient magnetite-based catalysts for wastewater treatments.

Three-Dimensional Graphene Supported Bimetallic Nanocomposites with DNA Regulated-Flexibly Switchable Peroxidase-Like Activity.

A synergistic bimetallic enzyme mimetic catalyst, three-dimensional (3D) graphene/Fe3O4-AuNPs, was successfully fabricated which exhibited flexibly switchable peroxidase-like activity. Compared to the traditional 2D graphene-based monometallic composite, the introduced 3D structure, which was induced by the addition of glutamic acid, and bimetallic anchoring approach dramatically improved the catalytic activity, as well as the catalysis velocity and its affinity for substrate. Herein, Fe3O4NPs acted as supporters for AuNPs, which contributed to enhance the efficiency of electron transfer. On the basis of the measurement of Mott-Schottky plots of graphene and metal anchored hybrids, the catalysis mechanism was elucidated by the decrease of Fermi level resulted from the chemical doping behavior. Notably, the catalytic activity was able to be regulated by the adsorption and desorption of single-stranded DNA molecules, which laid a basis for its utilization in the construction of single-stranded DNA-based colorimetric biosensors. This strategy not only simplified the operation process including labeling, modification, and imprinting, but also protected the intrinsic affinity between the target and biological probe. Accordingly, based on the peroxidase-like activity and its controllability, our prepared nanohybrids was successfully adopted in the visualized and label-free sensing detections of glucose, sequence-specific DNA, mismatched nucleotides, and oxytetracycline.

A HPLC-MS/MS method for the quantitation of free, conjugated, and total HDND-7, a novel hesperetin derivative, in rat plasma and tissues: Application to the pharmacokinetic and tissue distribution study.

A sensitive and reliable HPLC-MS/MS method was developed and validated for the determination of free (unconjugated), glucuronidated, sulfated, and total (free and conjugated) HDND-7 in rat plasma and tissues. Plasma and tissues samples were treated prior to and after the enzyme hydrolysis. Chromatographic separation was achieved on a Phenomenex Luna C18 column (150 × 4.6mm, 3 µm), using isocratic mobile phase consisting of 0.1% formic acid-acetonitrile (50:50, v/v) at a flow rate of 300 µl/min. The detection was performed on a triple quadruple tandem mass spectrometer using positive electrospray ionization (ESI) source with a chromatographic run time of 5.0 min. The detection was operated by multiple reaction monitoring (MRM) of the transitions of m/z 429.3 → 223.9 for HDND-7 and 272.9 → 152.9 for naringenin (IS), respectively. This method was validated in terms of specificity, linearity, precision, accuracy, and stability. The calibration curves for plasma and tissues were linear over a wide concentration range of 0.02-40 µg/ml with a lower limit of quantification (LLOQ) of 0.02 µg/ml. Mean extraction recoveries in plasma and tissues ranged from 87.4 to 97.1% and from 54.2 to 70.5%, respectively. The intra- and inter-day precision values were below 15% and the accuracy was within ± 15%. The samples were stable under all the tested conditions. This method has been successfully applied to the pharmacokinetic study following oral doses of 25, 50 and 100mg/kg and intravenous dose of 25mg/kg, and tissue distribution study following oral dose of 50mg/kg.