Lansoprazole promotes cisplatin-induced acute kidney injury via enhancing tubular necroptosis

Acute kidney injury (AKI) is the main obstacle that limits the use of cisplatin in cancer treatment. Proton pump inhibitors (PPIs), the most commonly used class of medications for gastrointestinal complications in cancer patients, have been reported to cause adverse renal events. However, the effect of PPIs on cisplatin-induced AKI remains unclear. Herein, the effect and mechanism of lansoprazole (LPZ), one of the most frequently prescribed PPIs, on cisplatin-induced AKI were investigated in vivo and in vitro. C57BL/6 mice received a single intraperitoneal (i.p.) injection of cisplatin (18 mg/kg) to induce AKI, and LPZ (12.5 or 25 mg/kg) was administered 2 hours prior to cisplatin administration and then once daily for another 2 days via i.p. injection. The results showed that LPZ significantly aggravated the tubular damage and further increased the elevated levels of serum creatinine and blood urea nitrogen induced by cisplatin. However, LPZ did not enhance cisplatin-induced tubular apoptosis, as evidenced by a lack of significant change in mRNA and protein expression of Bax/Bcl-2 ratio and TUNEL staining. Notably, LPZ increased the number of necrotic renal tubular cells compared to that by cisplatin treatment alone, which was further confirmed by the elevated necroptosis-associated protein expression of RIPK1, p-RIPK3 and p-MLKL. Furthermore, LPZ deteriorated cisplatin-induced inflammation, as revealed by the increased mRNA expression of pro-inflammatory factors including, NLRP3, IL-1β, TNF-α and caspase 1, as well as neutrophil infiltration. Consistently, in in vitro study, LPZ increased HK-2 cell death and enhanced inflammation, compared with cisplatin treatment alone. Collectively, our results demonstrate that LPZ aggravates cisplatin-induced AKI, and necroptosis may be involved in the exacerbation of kidney damage.     

Artesunate relieves acute kidney injury through inhibiting macrophagic Mincle-mediated necroptosis and inflammation to tubular epithelial cell

Artesunate is a widely used derivative of artemisinin for malaria. Recent researches have shown that artesunate has a significant anti-inflammatory effect on many diseases. However, its effect on acute kidney injury with a significant inflammatory response is not clear. In this study, we established a cisplatin-induced AKI mouse model and a co-culture system of BMDM and tubular epithelial cells (mTEC) to verify the renoprotective and anti-inflammatory effects of artesunate on AKI, and explored the underlying mechanism. We found that artesunate strongly down-regulated the serum creatinine and BUN levels in AKI mice, reduced the necroptosis of tubular cells and down-regulated the expression of the tubular injury molecule Tim-1. On the other hand, artesunate strongly inhibited the mRNA expression of inflammatory cytokines (IL-1β, IL-6 and TNF-α), protein levels of inflammatory signals (iNOS and NF-κB) and necroptosis signals (RIPK1, RIPK3 and MLKL) in kidney of AKI mouse. Notably, the co-culture system proved that Mincle in macrophage can aggravate the inflammation and necroptosis of mTEC induced by LPS, and artesunate suppressed the expression of Mincle in macrophage of kidney in AKI mouse. Overexpression of Mincle in BMDM restored the damage and necroptosis inhibited by artesunate in mTEC, indicating Mincle in macrophage is the target of artesunate to protect tubule cells in AKI. Our findings demonstrated that artesunate can significantly improve renal function in AKI, which may be related to the inhibition of Mincle-mediated macrophage inflammation, thereby reducing the damage and necroptosis to tubular cells that provide new option for the treatment of AKI.     

Neutral ceramidase deficiency protects against cisplatin-induced acute kidney injury

Cisplatin is a commonly used chemotherapeutic for the treatment of many solid organ cancers; however, its effectiveness is limited by the development of acute kidney injury (AKI) in 30% of patients. AKI is driven by proximal tubule cell death, leading to rapid decline in renal function. It has previously been shown that sphingolipid metabolism plays a role in regulating many of the biological processes involved in cisplatin-induced AKI. For example, neutral ceramidase (nCDase) is an enzyme responsible for converting ceramide into sphingosine, which is then phosphorylated to become sphingosine-1-phosphate, and our lab previously demonstrated that nCDase knockout (nCDase?/?) in mouse embryonic fibroblasts led to resistance to nutrient and energy deprivation–induced cell death via upregulation of autophagic flux. In this study, we further characterized the role of nCDase in AKI by demonstrating that nCDase?/? mice are resistant to cisplatin-induced AKI. nCDase?/? mice display improved kidney function, reduced injury and structural damage, lower rates of apoptosis, and less ER stress compared to wild-type mice following cisplatin treatment. Although the mechanism of protection is still unknown, we propose that it could be mediated by increased autophagy, as chloroquine treatment resensitized nCDase?/? mice to AKI development. Taken together, we conclude that nCDase may represent a novel target to prevent cisplatin-induced nephrotoxicity.     

PSTPIP2 inhibits cisplatin-induced acute kidney injury by suppressing apoptosis of renal tubular epithelial cells

Cisplatin (CP) is an effective chemotherapeutic agent widely used in the treatment of various solid tumours. However, CP nephrotoxicity is an important limitation for CP use; currently, there is no method to ameliorate cisplatin-induced acute kidney injury (AKI). Recently, we identified a specific role of proline–serine–threonine phosphatase-interacting protein 2 (PSTPIP2) in cisplatin-induced AKI. PSTPIP2 was reported to play an important role in a variety of diseases. However, the functions of PSTPIP2 in experimental models of cisplatin-induced AKI have not been extensively studied. The present study demonstrated that cisplatin downregulated the expression of PSTPIP2 in the kidney tissue. Administration of AAV-PSTPIP2 or epithelial cell-specific overexpression of PSTPIP2 reduced cisplatin-induced kidney dysfunction and inhibited apoptosis of renal tubular epithelial cells. Small interfering RNA-based knockdown of PSTPIP2 expression abolished PSTPIP2 regulation of epithelial cell apoptosis in vitro. Histone acetylation may impact gene expression at the epigenetic level, and histone deacetylase (HDAC) inhibitors were reported to prevent cisplatin-induced nephrotoxicity. The UCSC database was used to predict that acetylation of histone H3 at lysine 27 (H3K27ac) induces binding to the PSTPIP2 promoter, and this prediction was validated by a ChIP assay. Interestingly, an HDAC-specific inhibitor (TSA) was sufficient to potently upregulate PSTPIP2 in epithelial cells. Histone acetylation-mediated silencing of PSTPIP2 may contribute to cisplatin nephrotoxicity. PSTPIP2 may serve as a potential therapeutic target in the prevention of cisplatin nephrotoxicity.Subject terms: Cancer, Cancer prevention     

ERK-mediated suppression of cilia in cisplatin-induced tubular cell apoptosis and acute kidney injury

In kidneys, each tubular epithelial cell contains a primary cilium that protrudes from the apical surface. Ciliary dysfunction was recently linked to acute kidney injury (AKI) following renal ischemia–reperfusion. Whether ciliary regulation is a general pathogenic mechanism in AKI remains unclear. Moreover, the ciliary change during AKI and its underlying mechanism are largely unknown. Here we examined the change of primary cilium and its role in tubular cell apoptosis and AKI induced by cisplatin, a chemotherapy agent with notable nephrotoxicity. In cultured human proximal tubular HK-2 epithelial cells, cilia became shorter during cisplatin treatment, followed by apoptosis. Knockdown of Kif3a or Polaris (cilia maintenance proteins) reduced cilia and increased apoptosis during cisplatin treatment. We further subcloned HK-2 cells and found that the clones with shorter cilia were more sensitive to cisplatin-induced apoptosis. Mechanistically, cilia-suppressed cells showed hyperphosphorylation or activation of ERK. Inhibition of ERK by U0126 preserved cilia during cisplatin treatment and protected against apoptosis in HK-2 cells. In C57BL/6 mice, U0126 prevented the loss of cilia from proximal tubules during cisplatin treatment and protected against AKI. U0126 up-regulated Polaris, but not Kif3a, in kidney tissues. It is suggested that ciliary regulation by ERK plays a role in cisplatin-induced tubular apoptosis and AKI.     

Progranulin protects against cerebral ischemia-reperfusion (I/R) injury by inhibiting necroptosis and oxidative stress

Ischemic stroke is a leading cause of mortality and disability worldwide. Nevertheless, its molecular mechanisms have not yet been adequately illustrated. Progranulin (PGRN) is a secreted glycoprotein with pleiotropic functions. In the present study, we found that PGRN expression was markedly reduced in mice after stroke onset through middle cerebral artery occlusion (MCAO). We also showed that necroptosis was a mechanism underlying cerebral I/R injury. Importantly, PGRN knockdown in vivo significantly promoted the infarction volume and neurological deficits scores in mice after MCAO surgery. Necroptosis induced by MCAO was further accelerated by PGRN knockdown, as evidenced by the promoted expression of phosphorylated receptor-interacting protein (RIP) 1 kinase (RIPK1), RIPK3 and mixed lineage kinase domain-like (MLKL), which was accompanied with increased expression of cleaved Caspase-8 and Caspase-3. However, PGRN over-expression was neuroprotective. Additionally, PGRN-regulated ischemic stroke was related to ROS accumulation that MCAO-mice with PGRN knockdown exhibited severe oxidative stress, as proved by the aggravated malondialdehyde (MDA) and lipid peroxidation (LPO) contents, and the decreased superoxide dismutase (SOD) activity. However, PGRN over-expression in mice with cerebral ischemia showed anti-oxidative effects. Finally, PGRN was found to attenuate oxidative damage partly via its regulatory effects on necroptosis. Therefore, promoting PGRN expression could reduced cerebral I/R-induced brain injury by suppressing neroptosis and associated reactive oxygen species (ROS) production. These data elucidated that PGRN might provide an effective therapeutic treatment for ischemic stroke.     

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

BackgroundAcute 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.ObjectiveThis study aimed to evaluate the renoprotective effects of gypenoside XLIX (Gyp XLIX) in AKI.MethodsThe 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.ResultsIn 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.ConclusionGyp XLIX may serve as a potential therapeutic agent for AKI treatment via IGFBP7/ IGF1R-dependent mechanisms.     

VEGF-modified human embryonic mesenchymal stem cell implantation enhances protection against cisplatin-induced acute kidney injury

The implantation of mesenchymal stem cells (MSC) has been reported as a new technique to restore renal tubular structure and improve renal function in acute kidney injury (AKI). Vascular endothelial growth factor (VEGF) plays an important role in the renoprotective function of MSC. Whether upregulation of VEGF by a combination of MSC and VEGF gene transfer could enhance the protective effect of MSC in AKI is not clear. We investigated the effects of VEGF-modified human embryonic MSC (VEGF-hMSC) in healing cisplatin-injured renal tubular epithelial cells (TCMK-1) with a coculture system. We found that TCMK-1 viability declined 3 days after cisplatin pretreatment and that coculture with VEGF-hMSC enhanced cell protection via mitogenic and antiapoptotic actions. In addition, administration of VEGF-hMSC in a nude mouse model of cisplatin-induced kidney injury offered better protective effects on renal function, tubular structure, and survival as represented by increased cell proliferation, decreased cellular apoptosis, and improved peritubular capillary density. These data suggest that VEGF-modified hMSC implantation could provide advanced benefits in the protection against AKI by increasing antiapoptosis effects and improving microcirculation and cell proliferation.     

Irisin protects against vascular calcification by activating autophagy and inhibiting NLRP3-mediated vascular smooth muscle cell pyroptosis in chronic kidney disease

Irisin protects the cardiovascular system against vascular diseases. However, its role in chronic kidney disease (CKD) -associated vascular calcification (VC) and the underlying mechanisms remain unclear. In the present study, we investigated the potential link among Irisin, pyroptosis, and VC under CKD conditions. During mouse vascular smooth muscle cell (VSMC) calcification induced by β-glycerophosphate (β-GP), the pyroptosis level was increased, as evidenced by the upregulated expression of pyroptosis-related proteins (cleaved CASP1, GSDMD-N, and IL1B) and pyroptotic cell death (increased numbers of PI-positive cells and LDH release). Reducing the pyroptosis levels by a CASP1 inhibitor remarkably decreased calcium deposition in β-GP-treated VSMCs. Further experiments revealed that the pyroptosis pathway was activated by excessive reactive oxygen species (ROS) production and subsequent NLR family pyrin domain containing 3 (NLRP3) inflammasome activation in calcified VSMCs. Importantly, Irisin effectively inhibited β-GP-induced calcium deposition in VSMCs in vitro and in mice aortic rings ex vivo. Overexpression of Nlrp3 attenuated the suppressive effect of Irisin on VSMC calcification. In addition, Irisin could induce autophagy and restore autophagic flux in calcified VSMCs. Adding the autophagy inhibitor 3-methyladenine or chloroquine attenuated the inhibitory effect of Irisin on β-GP-induced ROS production, NLRP3 inflammasome activation, pyroptosis, and calcification in VSMCs. Finally, our in vivo study showed that Irisin treatment promoted autophagy, downregulated ROS level and thereby suppressed pyroptosis and medial calcification in aortic tissues of adenine-induced CKD mice. Together, our findings for the first time demonstrated that Irisin protected against VC via inducing autophagy and inhibiting VSMC pyroptosis in CKD, and Irisin might serve as an effective therapeutic agent for CKD-associated VC.Subject terms: Calcification, Chronic kidney disease     

Histone methyltransferase MLL1 drives renal tubular cell apoptosis by p53-dependent repression of E-cadherin during cisplatin-induced acute kidney injury

Mixed lineage leukemia 1 (MLL1) is a histone H3 lysine 4 (H3K4) methyltransferase that interacts with WD repeat domain 5 (WDR5) to regulate cell survival, proliferation, and senescence. The role of MLL1 in the pathogenesis of acute kidney injury (AKI) is unknown. In this study, we demonstrate that MLL1, WDR5, and trimethylated H3K4 (H3K4me3) were upregulated in renal tubular cells of cisplatin-induced AKI in mice, along with increased phosphorylation of p53 and decreased expression of E-cadherin. Administration of MM102, a selective MLL1/WDR5 complex inhibitor, improved renal function and attenuated tubular injury and apoptosis, while repressing MLL1, WDR5, and H3K4me3, dephosphorylating p53 and preserving E-cadherin. In cultured mouse renal proximal tubular cells (RPTCs) exposed to cisplatin, treatment with MM102 or transfection with siRNAs for either MLL1 or WDR5 also inhibited apoptosis and p53 phosphorylation while preserving E-cadherin expression; p53 inhibition with Pifithrin-α lowered cisplatin-induced apoptosis without affecting expression of MLL1, WDR5, and H3K4me3. Interestingly, silencing of E-cadherin offset MM102’s cytoprotective effects, but had no effect on p53 phosphorylation. These findings suggest that MLL1/WDR5 activates p53, which, in turn, represses E-cadherin, leading to apoptosis during cisplatin-induced AKI. Further studies showed that MM102 effectively inhibited cisplatin-triggered DNA damage response (DDR), as indicated by dephosphorylation of ataxia telangiectasia mutated (ATM) and ATM and Rad-3 related (ATR) proteins, dephosphorylation of checkpoint kinase 1 and 2 (Chk1 and Chk2); depression of γ-H2AX; and restrained cell cycle arrest, as evidenced by decreased expression of p21 and phospho-histone H3 at serine 10 in vitro and in vivo. Overall, we identify MLL1 as a novel DDR regulator that drives cisplatin-induced RPTC apoptosis and AKI by modulating the MLL1/WDR5-/ATR/ATM-Chk-p53-E-cadherin axis. Targeting the MLL1/WDR5 complex may have a therapeutic potential for the treatment of AKI.Subject terms: Acute kidney injury, Methylation     

Autologous cell therapy for cisplatin-induced acute kidney injury by using non-expanded adipose tissue-derived cells

Abstract Background aims. Recent studies have demonstrated that cultured mesenchymal stromal cells derived from adipose tissue are useful for regenerative cell therapy. The stromal vascular fraction (SVF) can be obtained readily without culturing and may be clinically applicable. We investigated the therapeutic effects of SVF and used it in the treatment of acute kidney injury (AKI). Methods. Liposuction aspirates were obtained from healthy donors who had provided written informed consent. We harvested the SVF and determined the growth factor secretion and anti-apoptotic ability with conditioned medium. To investigate the effect of SVF on AKI, cisplatin was injected into rats and SVF was administrated into the subcupsula of the kidney. Results. Both human and rat SVF cells secreted vascular endothelial growth factor-A (VEGF) and hepatocyte growth factor (HGF). Human SVF-conditioned media had an anti-apoptotic effect, which was inhibited by anti-HGF antibody (Ab) but not by anti-VEGF Ab. In vivo, SVF significantly ameliorated renal function, attenuated tubular damage and increased the cortical blood flow speed. In the SVF-treated group, VEGF levels in the cortex and HGF levels in both the cortex and medulla, especially tubules in the medulla, were significantly higher. Immunostaining revealed that SVF cells expressing VEGF and HGF and remained in the subcapsule on day 14. Conclusions. The present study demonstrates that a subcapsular injection of non-expanded SVF cells ameliorates rat AKI, and that the mechanism probably involves secretion of renoprotective molecules. Administration of human SVF may be clinically applicable and useful as a novel autologous cell therapy against kidney diseases.     

MFGE8 protects against CCl4-induced liver injury by reducing apoptosis and promoting proliferation of hepatocytes

Milk fat globule-EGF factor 8 (MFGE8) has been reported to play various roles in acute injury and inflammation response. However, the role of MFGE8 in liver injury is poorly investigated. The present research was designed to clarify the expression and function of MFGE8 in carbon tetrachloride (CCl₄)-induced liver injury. Using serum cytokine arrays, we selected a promising cytokine MFGE8 as the candidate in the process of hepatitis-fibrosis-hepatocellular carcinoma (HCC) progression, based on the elevated expression in both hepatic fibrosis and HCC models. We validated the increased expression of MFGE8 in liver tissues and serum samples of acute and chronic CCl₄-induced mice. Immunohistochemistry staining of mouse liver tissues indicated that elevated MFGE8 expression was mainly derived from the injured hepatocytes. In addition, MFGE8 expression in the supernatant of primary hepatocytes was accumulated with prolongation of culture time, and CCl₄ treatment further increased the expression of MFGE8. Moreover, a strong correlation between serum MFGE8 expression and liver transaminase activities suggested that MFGE8 may be a novel candidate in liver injury. Intriguingly, mice pretreated with MFGE8 were protected from CCl₄-induced liver injury through antiapoptosis role in the early stage and proproliferation role in the late stage. MFGE8 reduced apoptosis by inhibiting the activation of IRE1α/ASK1/JNK pathway and promoted proliferation by phosphorylation of ERK and AKT. Moreover, serum MFGE8 expression was increased in hepatitis patients while decreased in liver cirrhosis patients. All the results suggest MFGE8 as a novel marker and promising therapeutic agent of liver injury.     

Upregulation of lncRNA NONRATG019935.2 suppresses the p53-mediated apoptosis of renal tubular epithelial cells in septic acute kidney injury

Although increasing evidence has confirmed that the apoptosis of renal tubular epithelial cells (RTECs) is a crucial contributor to the onset and development of septic acute kidney injury (AKI), the pathological mechanism by which RTEC apoptosis is upregulated during septic AKI is not entirely clear. In this study, a rat model of septic AKI was induced by a cecal ligation puncture procedure or lipopolysaccharide (LPS) injection. Four differentially expressed long noncoding RNAs (DE-Lncs) in the rat model of septic AKI were determined using RNA-sequencing and verified by qRT-PCR. Among the four DE-Lncs, the expression level of lncRNA NONRATG019935.2 (9935) exhibited the most significant reduction in both septic AKI rats and LPS-treated NRK-52E cells (a rat RTEC line). The overexpression of 9935 suppressed cell apoptosis and p53 protein level in LPS-treated NRK-52E cells, and retarded septic AKI development in the rat model of septic AKI. Mechanistically, 9935 decreased the human antigen R (HuR)-mediated Tp53 mRNA stability by limiting the combination of HuR and the 3′UTR region of Tp53 mRNA in RTECs. The overexpression of HuR abrogated the inhibitory effect of pcDNA-9935 on the LPS-induced apoptosis of NRK-52E and rat primary RTECs. In conclusion, 9935 exerts its role in septic AKI by suppressing the p53-mediated apoptosis of RTECs, and this essential role of 9935 relies on its destructive effect on HuR-mediated Tp53 mRNA stability.Subject terms: Cell biology, Molecular biology     

Resveratrol protects astrocytes against traumatic brain injury through inhibiting apoptotic and autophagic cell death

Traumatic brain injury (TBI) is often caused by accidents that damage the brain. TBI can induce glutamate excitotoxicity and lead to neuronal and glial cell death. In this study, we investigated the mechanism of cell death during the secondary damage caused by TBI in vivo and in vitro, as well as the protective effect of resveratrol (RV). Here we report that glycogen synthase kinase-3β (GSK-3β) activation and microtubule-associated protein light chain 3 processing were induced in rat brains exposed to TBI. In the in vitro TBI model, apoptotic and autophagic cell death were induced through glutamate-mediated GSK-3β activation in normal CTX TNA2 astrocytes. The GSK-3β inhibitor SB216763 or transfection of GSK-3β small-interfering RNA increases cell survival. By contrast, overexpression of GSK-3β enhanced glutamate excitotoxicity. Administration of RV reduced cell death in CTX TNA2 astrocytes by suppressing reactive oxygen species (ROS)-mediated GSK-3β activation, the mechanism by which RV also exerted protective effects in vivo. Mitochondrial damages, including the opening of mitochondrial permeability transition pore (MPTP) and mitochondrial depolarization, were induced by glutamate through the ROS/GSK-3β pathway. Moreover, cyclosporine A, an MPTP inhibitor, suppressed mitochondrial damage and the percentages of cells undergoing autophagy and apoptosis and thereby increased cell survival. Taken together, our results demonstrated that cell death occurring after TBI is induced through the ROS/GSK-3β/mitochondria signaling pathway and that administration of RV can increase cell survival by suppressing GSK-3β-mediated autophagy and apoptosis. Therefore, the results indicated that resveratrol may serve as a potential therapeutic agent in the treatment of TBI.     

CGRP is essential for protection against alveolar epithelial cell necroptosis by activating the AMPK/L-OPA1 signaling pathway during acute lung injury

Alveolar epithelial cell (AEC) necroptosis is critical to disrupt the alveolar barrier and provoke acute lung injury (ALI). Here, we define calcitonin gene-related peptide (CGRP), the most abundant endogenous neuropeptide in the lung, as a novel modulator of AEC necroptosis in lipopolysaccharide (LPS)-induced ALI. Upon LPS-induced ALI, overexpression of Cgrp significantly mitigates the inflammatory response, alleviates lung tissue damage, and decreases AEC necroptosis. Similarly, CGRP alleviated AEC necroptosis under the LPS challenge in vitro. Previously, we identified that long optic atrophy 1 (L-OPA1) deficiency mediates mitochondrial fragmentation, leading to AEC necroptosis. In this study, we discovered that CGRP positively regulated mitochondrial fusion through stabilizing L-OPA1. Mechanistically, we elucidate that CGRP activates AMP-activated protein kinase (AMPK). Furthermore, the blockade of AMPK compromised the protective effect of CGRP against AEC necroptosis following the LPS challenge. Our study suggests that CRGP-mediated activation of the AMPK/L-OPA1 axis may have potent therapeutic benefits for patients with ALI or other diseases with necroptosis.     

Matrix metalloproteinase-10 protects against acute kidney injury by augmenting epidermal growth factor receptor signaling

Matrix metalloproteinase-10 (MMP-10) is a zinc-dependent endopeptidase involved in regulating a wide range of biologic processes, such as apoptosis, cell proliferation, and tissue remodeling. However, the role of MMP-10 in the pathogenesis of acute kidney injury (AKI) is unknown. In this study, we show that MMP-10 was upregulated in the kidneys and predominantly localized in the tubular epithelium in various models of AKI induced by ischemia/reperfusion (IR) or cisplatin. Overexpression of exogenous MMP-10 ameliorated AKI, manifested by decreased serum creatinine, blood urea nitrogen, tubular injury and apoptosis, and increased tubular regeneration. Conversely, knockdown of endogenous MMP-10 expression aggravated kidney injury. Interestingly, alleviation of AKI by MMP-10 in vivo was associated with the activation of epidermal growth factor receptor (EGFR) and its downstream AKT and extracellular signal-regulated kinase-1 and 2 (ERK1/2) signaling. Blockade of EGFR signaling by erlotinib abolished the MMP-10-mediated renal protection after AKI. In vitro, MMP-10 potentiated EGFR activation and protected kidney tubular cells against apoptosis induced by hypoxia/reoxygenation or cisplatin. MMP-10 was colocalized with heparin-binding EGF-like growth factor (HB-EGF) in vivo and activated it by a process of proteolytical cleavage in vitro. These studies identify HB-EGF as a previously unrecognized substrate of MMP-10. Our findings also underscore that MMP-10 can protect against AKI by augmenting EGFR signaling, leading to promotion of tubular cell survival and proliferation after injury.Subject terms: Apoptosis, Cell growth     

Astaxanthin protects against renal fibrosis through inhibiting myofibroblast activation and promoting CD8+ T cell recruitment

BackgroundRenal fibrosis is a common pathological hallmark of chronic kidney disease, and no effective treatment is clinically available to manage its progression. Astaxanthin was recently found to be anti-fibrotic, but its effect on renal fibrosis remains unclear.MethodsC57BL/6J mice were subjected to unilateral ureteral obstruction and intragastrically administered astaxanthin. Histopathology and immunohistochemistry were performed to evaluate renal fibrosis. Flow cytometry was used to examine lymphocyte accumulation in the fibrotic kidneys. Western blotting, real-time qPCR, and immunofluorescence were performed to cover the underlying mechanism concerning astaxanthin treatment during renal fibrosis.ResultsOral administration of astaxanthin effectively alleviates renal fibrosis in mice. In vitro, astaxanthin inhibited fibroblast activation by modulating Smad2, Akt and STAT3 pathways and suppressed epithelial-to-mesenchymal transition in renal tubular epithelial cells through Smad2, snail, and β-catenin. Moreover, astaxanthin significantly induced the rapid accumulation of CD8⁺ T cells in fibrotic kidneys, which was accompanied by elevated expression of IFN-γ. Accordingly, the depletion of CD8⁺ T cells strongly diminished the protective effect of astaxanthin. Further investigation showed that astaxanthin increased the population of CD8⁺ T cells by upregulating the expression of CCL5 in macrophages.ConclusionsThese findings highlight the beneficial effect of astaxanthin on fibroblast activation, epithelial-to-mesenchymal transition, and CD8⁺ T cell recruitment during renal fibrosis.General significanceThese data indicate that astaxanthin could serve as a therapeutic strategy to treat renal fibrotic conditions.     

Hydrogen sulfide protects against acetaminophen-induced acute liver injury by inhibiting apoptosis via the JNK/MAPK signaling pathway

Acetaminophen (APAP) is a widely used over-the-counter analgesic and antipyretic. It can cause hepatotoxicity. Recent studies demonstrated that hydrogen sulfide (H₂S) exhibits cell protection in several cell types. This study was designed to investigate whether H ₂S ameliorated APAP-induced acute liver injury and to elucidate its mechanisms. In this study, we analyzed the detailed biological and molecular processes of APAP-induced hepatotoxicity using a bioinformatics analysis, which showed that apoptosis and the c-Jun N-terminal kinase (JNK)/mitogen-activated protein kinase pathway were confirmed to play critical roles in these processes. We further investigated the protective effects of H ₂S on APAP-induced hepatotoxicity. In vivo, we observed that the exogenous supplement of H ₂S ameliorated APAP-induced liver injury. Cystathionine-β-synthase (CBS) and cystathionine-γ-lyase (CSE) systems were the endogenous pathway of H ₂S. The expression of CBS/CSE was decreased in APAP-treated mice, while H ₂S could significantly restore it. In addition, APAP-induced JNK activation was inhibited by H ₂S in vivo. In vitro, H ₂S abolished the active effects of APAP on caspase3, Bax, and Bcl-2 expressions as well as JNK phosphorylation in hepatocytes. It was found through flow cytometry that the amount of APAP-induced apoptotic hepatocytes was decreased in the presence of H ₂S. In conclusion, our results suggested that H ₂S attenuated APAP-induced apoptosis in hepatocytes through JNK/MAPK siganaling pathway.     

Ghrelin protects against cobalt chloride-induced hypoxic injury in cardiac H9c2 cells by inhibiting oxidative stress and inducing autophagy

Ghrelin is a multifunctional peptide that actively protects against cardiovascular ischemic diseases, but the underlying mechanisms are unclear. We used CoCl₂ to mimic hypoxic conditions in cardiac H9c2 cells in order to study the mechanism by which ghrelin protects cardiac myocytes against hypoxic injury by regulating the content of intracellular ROS and autophagy levels. Cell apoptosis and necrosis were evaluated by the flow cytometry assay, Hoechst staining, and LDH activity. Cell viability was detected by the WST-1 assay; ROS levels were assessed using DCFH2-DA; and Nox1, catalase and Mn-SOD were assayed by real-time PCR and activity assays. LC3II was measured by Western blot analysis. We observed that CoCl₂ induced apoptosis and death of H9c2 cells in a dose- and time-dependent manner. This was characterized by an increase in cell apoptosis, LDH activity, ROS content, Nox1 expression, and autophagy levels and a decrease in cell viability, catalase, and Mn-SOD activities. Ghrelin treatment significantly attenuated CoCl₂-induced hypoxic injury by decreasing cell apoptosis, LDH activity, ROS content, and Nox1 expression and increasing cell viability, autophagy levels, catalase, and Mn-SOD mRNA levels and activities. Further experiments revealed that inhibiting autophagy using 3-MA or AMPK pathway with compound C almost abrogated the induction of ghrelin in autophagy. This was associated with a decrease in cell viability and an increase in LDH activity. Our results indicate that ghrelin protected cardiac myocytes against CoCl₂-induced hypoxic injury by decreasing Nox1 expression, increasing the expression and activity of endogenous antioxidant enzymes, and inducing protective autophagy in an AMPK-dependent manner.