Renal Ischemia-reperfusion Model Research Articles

Pinocembrin alleviates renal ischemia-reperfusion injury/unilateral ureteral obstruction (UUO)-generated renal fibrosis by targeting the CYP1B1/ROS/MAPK axis.

In our research, we constructed models of renal ischemia-reperfusion (I/R)-exposed acute kidney injury (AKI) and unilateral ureteral obstruction (UUO)-stimulated renal fibrosis (RF) in C57BL/6 mice and HK-2 cells. We firstly authenticated that oral pinocembrin (PIN) administration obviously mitigated tissue damage and renal dysfunction induced by I/R injury, and PIN attenuated UUO-caused RF, as confirmed by the reduced expression of fibrotic markers as well as hematoxylin-eosin (H&E), Sirius red, immunohistochemistry, and Masson staining. Meanwhile, the beneficial role of PIN was again demonstrated in HK-2 cells with hypoxia-reoxygenation (H/R) or transforming growth factor beta-1 (TGF-β1) treatment. Importantly, the "ingredient-target-pathway-disease" network was established through bioinformatics analysis and molecular docking, which showed that PIN may target cytochrome P450 1B1 (CYP1B1) and modulate the mitogen-activated protein kinase (MAPK) pathway to exert its impact during injury. Furthermore, experiments confirmed that PIN usage remarkably constrained CYP1B1 expression, reactive oxygen species (ROS) production, MAPK-pathway-associated inflammation, or apoptosis during I/R injury or UUO exposure. PIN also ameliorated the elevated protein phosphorylation of MAPK pathway components [p38, extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase 1 (JNK ERK and JNK)], which validated the PIN-induced inhibition of the MAPK signaling pathway in renal I/R or UUO injury. Moreover, the AAV9 (adeno-associated virus 9)-packed CYP1B1 or pcDNA-CYP1B1 overexpression plasmid was utilized to treat C57BL/6 mice or HK-2 cells to overexpress CYP1B1, respectively. Notably, CYP1B1 overexpression considerably abolished PIN's restriction impact on ROS generation and MAPK pathway activation. In conclusion, via bioinformatics analysis, molecular docking, animal model, and cellular experiments, we proved that PIN alleviates renal I/R injury/UUO-generated renal fibrosis through regulating the CYP1B1/ROS/MAPK axis.

Involvement of Mineralocorticoid Receptor Activation by HMGB1 and RAGE in the Development of Acute Kidney Injury.

Although acute kidney injury (AKI) is associated with an increased risk of chronic kidney disease (CKD), the underlying mechanisms remain unclear. High mobility group box-1 (HMGB1), one of the ligands for the receptor for advanced glycation end products (RAGE), is elevated in patients with AKI. We recently demonstrated that the mineralocorticoid receptor (MR) is activated by the RAGE/Rac1 pathway, contributing to chronic renal damage in hypertensive mice. Therefore, this study investigated the role of the HMGB1/RAGE/MR pathway in AKI and progression to CKD. We performed a mouse model of renal ischemia-reperfusion (I/R) with or without MR antagonist (MRA). In vitro experiments were conducted using cultured endothelial cells to examine the interaction between the HMGB1/RAGE and Rac1/MR pathways. In renal I/R injury mice, renal MR activation was associated with elevated serum HMGB1, renal RAGE, and activated Rac1, all of which were suppressed by MRA. Renal I/R injury led to renal dysfunction, tubulointerstitial injury, and increased expressions of inflammation and fibrosis mediators, which were ameliorated by MRA. In vitro, RAGE-aptamer or MRA inhibited HMGB1-induced Rac1/MR activation and upregulation of MCP-1 and NF-κB expressions. 7 days following I/R injury, renal I/R injury mice developed CKD, whereas MRA prevented renal injury progression and decreased mortality rate. Furthermore, in case of MRA treatment even after I/R injury, attenuated renal dysfunction compared to untreated mice was also observed. Our findings suggest that HMGB1 may play a crucial role in AKI and CKD development by activating the Rac1/MR pathway through interactions with RAGE.

Omega 3 Fatty Acids Attenuate the Acute Kidney Injury to CKD Transition and Renal Fibrosis: Identification of Antifibrotic Metabolites.

AKI is an established risk factor for developing CKD. Recently, the renoprotective effect of omega-3 polyunsaturated fatty acids (ω3PUFAs) has attracted attention. This study aimed to evaluate the effect of ω3PUFAs on the transition of AKI to CKD, and to identify fatty acid active metabolites in renal tissue. Two mice models of AKI to CKD (7-week, male) and unilateral ureteral obstruction (UUO)-induced renal fibrosis (11-week, male) were fed linseed oil, rich in ω3PUFAs (Lin group), or with soybean oil, low in ω3PUFAs (Soy group). Renal fatty acids and metabolites composition in mice were measured by liquid chromatography-mass spectrometry. Rat renal fibroblast cells (NRK-49F cells) were used for in vitro study. At day 14 after 35 min bilateral renal ischemia reperfusion (IR), significant increase in survival was observed in the Lin group compared to the Soy group. Using the 30 min bilateral renal IR model (AKI to CKD model), the Lin group showed attenuated renal tissue damage and fibrosis. In addition, the antifibrotic effect of Lin group was also observed in UUO renal fibrosis model. In the two mice models, levels of eicosapentaenoic acid (EPA) and its metabolites were significantly elevated in renal tissue of mice fed with Lin. Cultured NRK-49F incubated with EPA and its metabolites 18-hydroxyeicosapentaenoic acid (18-HEPE), 17,18-epoxyeicosatetraenoic acid (17,18-EpETE) and 17,18-dihydroxyeicosatetraenoic acid (17,18-diHETE) displayed suppressed TGF-β1-stimulated α-smooth muscle actin protein expression. These effects were suppressed in the presence of an inhibitor of a cytochrome P450 involved in EPA metabolism. This observation suggests that the EPA metabolites have antifibrotic effects. ω3PUFAs prevents AKI to CKD and renal fibrosis. Moreover, the EPA metabolites 18-HEPE, 17,18-EpETE and 17,18-diHETE were found to have antifibrotic effects.

Ligustilide alleviates oxidative stress during renal ischemia-reperfusion injury through maintaining Sirt3-dependent mitochondrial homeostasis

BackgroundRenal ischemia-reperfusion (I/R) injury is an inevitable complication during renal transplantation and is closely related to patient prognosis. Mitochondrial damage induced oxidative stress is the core link of renal I/R injury. Ligustilide (LIG), a natural compound extracted from ligusticum chuanxiong hort and angelica sinensis, has exhibited the potential to protect mitochondrial function. However, whether LIG can ameliorate renal I/R injury requires further investigation. Delving deeper into the precise targets and mechanisms of LIG's effect on renal I/R injury is crucial. PurposeThis study aimed to elucidate the specific mechanism of LIG's protective effect on renal I/R injury. MethodsIn this study, an in vivo model of renal ischemia-reperfusion (I/R) injury was developed in mice, along with an in vitro model of hypoxia-reoxygenation (H/R) using human proximal renal tubular epithelial cells (HK-2). To assess the impact of LIG on renal injury, various methods were employed, including serum creatinine (Cr) and blood urea nitrogen (BUN) testing, hematoxylin and eosin (HE) staining, and immunohistochemistry (IHC) for kidney injury molecule-1 (KIM-1). The effects of LIG on oxidative stress were examined using fluorescent probes dihydroethidium (DHE) and dichlorodihydrofluorescein diacetate (DCFH-DA), TdT-mediated dUTP Nick-End Labeling (TUNEL) staining, and flow cytometry. Additionally, the influence of LIG on mitochondrial morphology and function was evaluated through transmission electron microscopy (TEM), Mito Tracker Red CMXRos staining, adenosine triphosphate (ATP) concentration assays, and JC-1 staining. The potential mechanism involving LIG and Sirt3 was explored by manipulating Sirt3 expression through cell transfection. ResultsThe results showed that LIG could provide protective function for mitochondria to alleviate oxidative stress induced by renal I/R. Further mechanistic studies indicated that LIG maintained mitochondrial homeostasis by targeting Sirt3. ConclusionOur findings demonstrated that LIG alleviated oxidative stress during renal I/R injury through maintaining Sirt3-dependent mitochondrial homeostasis. Overall, our data raised the possibility of LIG as a novel therapy for renal I/R injury.

The Effect of Interleukin-10 Immunotherapy on Renal Ischemia-Reperfusion Injury: A Systematic Review and Meta-Analysis of Preclinical Studies.

Renal ischemia-reperfusion is a common cause of acute kidney injury leading to significant morbidity and mortality. There are no effective treatments available in clinical practice. This meta-analysis aims to assess the effect of IL-10 immunotherapy on renal ischemia-reperfusion injury. Medline, Embase, Cochrane-library, Google Scholar and clinicaltrials.gov were searched up to 31 March 2023. Preclinical and clinical interventional studies investigating IL-10 immunotherapy for renal ischemia-reperfusion were eligible for inclusion. The primary endpoint was renal function (serum creatinine) following ischemia-reperfusion. The secondary endpoints included mitochondrial integrity, cellular proliferation, regulated cell death (TUNEL assay), expression of inflammatory cytokines (TNF-α, IL-6 and IL-1β), M1/M2 macrophage polarization, tissue integrity (tubular injury score), long-term kidney fibrosis (fibrotic area %) and adverse events (pulmonary toxicity, cardiotoxicity hepatotoxicity). The search returned 861 records. From these, 16 full texts were screened and subsequently, seven animal studies, corresponding to a population of 268 mice/rats, were included. Compared to the control treatment, IL-10 immunotherapy reduced serum creatinine more effectively within 24 h of administration (95% CI: -9.177, -5.601, I2 = 22.42%). IL-10 immunotherapy promoted mitochondrial integrity and cellular proliferation and reduced regulated cell death (95% CI: -11.000, -4.184, I2 = 74.94%). It decreased the expression of TNF-α, IL-6 and IL-1β, led to M2 polarization of the local macrophages, reduced tubular injury score (95% CI: -8.917, -5.755, I2 = 22.71%), and long-term kidney fibrosis (95% CI: -6.963, -3.438, I2 = 0%). No adverse outcomes were captured. In Conclusion, IL-10 immunotherapy safely improves outcomes in animal models of renal ischemia-reperfusion; the translational potential of IL-10 immunotherapy needs to be further investigated in clinical trials.

#829 Renoprotective effects of laxative linaclotide: inhibition of acute kidney injury and fibrosis in a rat model of renal ischemia-reperfusion

Abstract Background and Aims Acute kidney injury caused by renal ischemia/reperfusion (I/R) is a major clinical problem in many aspects of urology fields such as kidney transplantation and partial nephrectomy, but there is no effective treatment. Linaclotide, a guanylate cyclase C agonist clinically approved as a laxative, has recently been shown to be renoprotective in a chronic kidney disease (CKD) model. Therefore, the present study investigated the possibility that the renoprotective effects of linaclotide in renal I/R may extend beyond its gastrointestinal effects. Method The left renal pelvis of adult male Wistar rats was clamped for 45 minutes to induce renal I/R injury. Kidney tissue, colon, and blood were collected 24 hours and 14 days later. Linaclotide was administered for 14 days preoperatively and until postoperative tissue collection. Tissue sections were stained with standard histologic techniques such as HE, Sirius Red, F4/80, etc. As genes associated with I/R, TGF-β and related fibrosis markers α-SMA, MMP2, TIMP1 and inflammatory cytokines TNF, IL-1β and IL-6 were analyzed by quantitative PCR analysis. Results Serum creatinine levels decreased significantly after linaclotide administration, indicating improved renal function. Histologic examination showed less tubular damage and less collagen deposition on Sirius Red staining. Further analysis showed decreased expression of TGF-β and related fibrosis markers α-SMA, MMP2 and TIMP1, suggesting downregulation of the fibrotic TGF-β pathway by linaclotide. Furthermore, 1 day after I/R injury, linaclotide significantly reduced macrophage infiltration and suppressed key proinflammatory cytokines such as TNF, IL-1β, and IL-6. Conclusion These results suggest that linaclotide, with its established safety profile, could extend its benefits beyond gastrointestinal issues and potentially serve as a therapeutic intervention in many aspects of kidney surgeries. In addition, it could provide immediate and practical insights into the selection of laxatives for the management of patients with AKI or CKD, regardless of cause, and for those receiving dialysis or transplant therapy.

Role of cAMP/pCREB and GSK-3β/NF-κB p65 signaling pathways in the renoprotective effect of mirabegron against renal ischemia-reperfusion injury in rats

Acute kidney injury and other renal disorders are thought to be primarily caused by renal ischemia-reperfusion (RIR). Cyclic adenosine monophosphate (cAMP) has plenty of physiological pleiotropic effects and preserves tissue integrity and functions. This research aimed to examine the potential protective effects of the β3-adrenergic receptors agonist mirabegron in a rat model of RIR and its underlying mechanisms. Male rats enrolled in this work were given an oral dose of 30 mg/kg mirabegron for two days before surgical induction of RIR. Renal levels of kidney injury molecule-1 (KIM-1), monocyte chemoattractant protein-1 (MCP-1), tumor necrosis factor-alpha (TNF-α), Interleukin-10 (IL-10), cAMP, cAMP-responsive element binding protein (pCREB), and glycogen synthase kinase-3 beta (GSK-3β) were assessed along with blood urea nitrogen and serum creatinine. Additionally, caspase-3 and nuclear factor-kappa B (NF-κB) p65 were explored by immunohistochemical analysis. Renal specimens were inspected for histopathological changes. RIR led to renal tissue damage with elevated blood urea nitrogen and serum creatinine levels. The renal KIM-1, MCP-1, TNF-α, and GSK-3β were significantly increased, while IL-10, cAMP, and pCREB levels were reduced. Moreover, upregulation of caspase-3 and NF-κB p65 protein expression was seen in RIR rats. Mirabegron significantly reduced kidney dysfunction, histological abnormalities, inflammation, and apoptosis in the rat renal tissues. Mechanistically, mirabegron mediated these effects via modulation of cAMP/pCREB and GSK-3β/NF-κB p65 signaling pathways. Mirabegron administration could protect renal tissue and maintain renal function against RIR.

Renoprotective effects of laxative linaclotide: Inhibition of acute kidney injury and fibrosis in a rat model of renal ischemia-reperfusion

Ischemia-reperfusion (I/R) leads to tissue damage in transplanted kidneys, resulting in acute kidney injury (AKI) and chronic graft dysfunction, which critically compromises transplant outcomes, such as graft loss. Linaclotide, a guanylate cyclase C agonist clinically approved as a laxative, has recently been identified to exhibit renoprotective effects in a chronic kidney disease (CKD) model. This study evaluates the therapeutic effects of linaclotide on AKI triggered by I/R in a rat model with an initial comparison with other laxatives. Here, we show that linaclotide administration resulted in substantial reduction in serum creatinine levels, reflective of enhanced renal function. Histological examination revealed diminished tubular damage, and Sirius Red staining confirmed less collagen deposition, collectively indicating preserved structural integrity and mitigation of fibrosis. Further analysis demonstrated lowered expression of TGF-β and associated fibrotic markers, α-SMA, MMP2, and TIMP1, implicating the downregulation of the fibrogenic TGF-β pathway by linaclotide. Furthermore, one day after I/R insult, linaclotide profoundly diminished macrophage infiltration and suppressed critical pro-inflammatory cytokines such as TNF, IL-1β, and IL-6, signifying its potential to disrupt initial inflammatory mechanisms integral to AKI pathology. These findings suggest that linaclotide, with its established safety profile, could extend its benefits beyond gastrointestinal issues and potentially serve as a therapeutic intervention for organ transplantation. Additionally, it could provide immediate and practical insights into selecting laxatives for managing patients with AKI or CKD, regardless of the cause, and for those receiving dialysis or transplant therapy.

Bilateral Renal Ischemia-Reperfusion Model for Acute Kidney Injury in Mice.

Acute kidney injury (AKI) is defined as a rapid decline in renal function, in which persistent kidney dysfunction gradually progresses to chronic kidney disease (CKD) due to the irreversible loss of nephrons and their maladaptive repair. In recent years, the incidence of AKI has been increasing concerning diverse etiologies, including volume depletion, sepsis, nephrotoxicity, muscle injury, and major trauma, in which ischemia-reperfusion injury (IRI)accounts for most episodes. Development of the IRI model in mice is induced by surgical clamping of the renal pedicles, which provides powerful and controllable tools for preclinical models of AKI. Importantly, the IRI model is deployed at different stages of the AKI development, especially in the processes of AKI to CKD. Despite the IRI model being widely practiced in many laboratories, a series of variables still influence the results of this model. Here, we describe the procedure of IRI model development to provide a repeatable and reliable method for researchers to explore the underlying pathogenesis in the development of AKI and the progression of AKI to CKD.

M6A methylase WTAP participates in renal ischemia-reperfusion injury by regulating FOXO1 expression

To investigate the expression of WTAP, a m6A methylase, in a mouse model of renal ischemia-reperfusion (I/R) injury and the effect of WTAP knockdown on biological behavior of renal tubular epithelial cells exposed to I/R injury. Sixteen C57BL/6 mice with renal I/R injury or sham operation (n=8) were examined for blood urea nitrogen (BUN) and creatinine (Scr) levels to assess renal function, and renal pathologies were observed with HE staining. The expressions of WTAP and FOXO1 proteins in the kidneys of the mice were detected using immunohistochemistry. Human renal tubular epithelial cells (HK-2) were transfected with si-WTAP or si-NC followed by hypoxia-reoxygenation (H/R) exposure, Protein and mRNA expression were assessed by Western blot and qRT-PCR, and changes and changes in cell viability and apoptosis were assessed using CCK8 assay and TUNEL staining, respectively; LDH release level and caspase-3 activity of the cells were measured using commercial assay kits. FOXO1 m6A modification sites were predicted using SRAMP website (http://www.cuilab.cn/sramp/), and the interaction between WTAP and FOXO1 mRNA was analyzed with RIP experiment; the level of FOXO1 modified by m6A was detected by MeRIP-qPCR. Compared with sham-operated mice, the mice with renal I/R injury showed significantly increased Scr and BUN levels (P < 0.001) and renal expressions of WTAP mRNA and protein (P < 0.001). In cultured HK-2 cells, H/R exposure significantly decreased the cell viability (P < 0.001) and increased cellular LDH release (P < 0.001) and expressions of WTAP mRNA and protein (P < 0.001). WTAP knockdown obviously reduced the cell damage induced by I/R injury and significantly decreased the mRNA and protein levels of FOXO1 in the cells (P < 0.001). RIP experiment confirmed WTAP binding to FOXO1 mRNA, and inhibition of WTAP expression significantly reduced FOXO1 m6A level in HK-2 cells (P < 0.001). WTAP expression is up-regulated in the kidneys of mice with renal I/R injury and in HK-2 cells with H/R exposure. Inhibition of WTAP alleviates H/R-induced apoptotic damage in HK-2 cells possibly by inhibiting FOXO1 expression.

Acrolein produced during acute kidney injury promotes tubular cell death

Acute kidney injury is an important global health concern as it is associated with high morbidity and mortality. Polyamines, essential for cell growth and proliferation, are known to inhibit cardiovascular disease. However, under conditions of cellular damage, toxic acrolein is produced from polyamines by the enzyme spermine oxidase (SMOX). We used a mouse renal ischemia-reperfusion model and human proximal tubule cells (HK-2) to investigate whether acrolein exacerbates acute kidney injury by renal tubular cell death. Acrolein visualized by acroleinRED was increased in ischemia-reperfusion kidneys, particularly in tubular cells. When HK-2 cells were cultured under 1% oxygen for 24 h, then switched to 21% oxygen for 24 h (hypoxia-reoxygenation), acrolein accumulated and SMOX mRNA and protein levels were increased. Acrolein induced cell death and fibrosis-related TGFB1 mRNA in HK-2 cells. Administration of the acrolein scavenger cysteamine suppressed the acrolein-induced upregulation of TGFB1 mRNA. Cysteamine also inhibited a decrease in the mitochondrial membrane potential observed by MitoTrackerCMXRos, and cell death induced by hypoxia-reoxygenation. The siRNA-mediated knockdown of SMOX also suppressed hypoxia-reoxygenation-induced acrolein accumulation and cell death. Our study suggests that acrolein exacerbates acute kidney injury by promoting tubular cell death during ischemia-reperfusion injury. Treatment to control the accumulation of acrolein might be an effective therapeutic option for renal ischemia-reperfusion injury.

PS-B08-6: ROLE OF THE HMGB-1/RAGE AXIS AND MINERALOCORTICOID RECEPTOR IN THE PATHOGENESIS OF ACUTE KIDNEY INJURY

Background: Endothelial dysfunction plays a central role in the pathogenesis of cardio-renal syndrome. High mobility group box-1 (HMGB-1) is a protein with various roles in different cellular compartments, and indirectly regulates the activity of transcription and DNA repair in the nucleus. On the other hand, during tissue damage, it is released into the extracellular environment as damage-associated molecular patterns (DAMPs). HMGB-1 is elevated in the pathogenesis of acute kidney injury (AKI) and be involved in endothelial dysfunction through binding to toll like receptor (TLR) and receptor for advanced glycation end products (RAGE). In addition, we recently demonstrated that RAGE-mediated Rac1 activated mineralocorticoid receptor (MR) and subsequently resulted in podocytes damage. In the present study, we hypothesized that crosstalk between HMGB-1/ RAGE and Rac1-MR pathways could contribute to endothelial dysfunction in AKI. Methods: In the present study, we performed a renal ischemia-reperfusion model (I/R group) on C57BL/6J mice to evaluate the expression of Rac1-MR pathway and organ damage. In addition, the I/R group was pretreated with MR blocker (esaxerenone) and evaluated in the same way. In vitro study, we investigated whether HMGB-1 could activate Rac1-MR axis and thus induce endothelial injury in cultured human umbilical vein endothelial cells (HUVECs) by assessing expression levels of genes for MCP-1 and NF-κB with or without administration of RAGE aptamer or MR blocker. Results: In the I/R group, MR activation assessed by translocation into the nucleus and enhanced RAGE expression were observed along with HMGB-1 and RAGE elevation, and pretreatment with MR blocker suppressed the elevation of serum creatinine levels as well as tubular necrosis. Further, the loss of peritubular capillaries observed in I/R kidney was markedly suppressed by MR blocker. In vitro study, HMGB-1 supplementation significantly increased RAGE expression, GTP-bound Rac1 and enhanced MR translocation into the nucleus in HUVECs as well. We also found that HMGB-1 upregulated MCP-1 and NF-κB, all of which were significantly blocked by pretreatment of RAGE aptamer and MR blocker. Conclusion: These results suggest that there may be a close relationship between HMGB-1/ RAGE axis and Rac1/ MR activation, thus contributing endothelial injury. Using RAGE aptamer or MR blocker could be novel therapeutic strategies against endothelial dysfunction in patients with AKI.

Deficiency of mindin reduces renal injury after ischemia reperfusion

BackgroundAcute renal injury (AKI) secondary to ischemia reperfusion (IR) injury continues to be a significant perioperative problem and there is no effective treatment. Mindin belongs to the mindin/F-spondin family and involves in inflammation, proliferation, and cell apoptosis. Previous studies have explored the biological functions of mindin in liver and brain ischemic injury, but its role in AKI is unknown.MethodTo investigate whether mindin has a pathogenic role, mindin knockout (KO) and wild-type (WT) mice were used to establish renal IR model. After 30 min of ischemia and 24 h of reperfusion, renal histology, serum creatinine, and inflammatory response were examined to assess kidney injury. In vitro, proinflammatory factors and inflammatory signaling pathways were measured in mindin overexpression or knockdown and vector cells after hypoxia/reoxygenation (HR).ResultsFollowing IR, the kidney mindin level was increased in WT mice and deletion of mindin provided significant protection for mice against IR-induced renal injury as manifested by attenuated the elevation of serum creatinine and blood urea nitrogen along with less severity for histological alterations. Mindin deficiency significantly suppressed inflammatory cell infiltration, TNF-α and MCP-1 production following renal IR injury. Mechanistic studies revealed that mindin deficiency inhibits TLR4/JNK/NF-κB signaling activation. In vitro, the expression levels of TNF-α and MCP-1 were increased in mindin overexpression cells compared with vector cells following HR. Moreover, TLR4/JNK/NF-κB signaling activation was elevated in the mindin overexpression cells in response to HR stimulation while mindin knockdown inhibited the activation of TLR4/JNK/ NF-κB signaling after HR in vitro. Further study showed that mindin protein interacted directly with TLR4 protein. And more, mindin protein was confirmed to be expressed massively in renal tubule tissues of human hydronephrosis patients.ConclusionThese data demonstrate that mindin is a critical modulator of renal IR injury through regulating inflammatory responses. TLR4/JNK/NF-κB signaling most likely mediates the biological function of mindin in this model of renal ischemia.

GATA2 promotes oxidative stress to aggravate renal ischemia-reperfusion injury by up-regulating Redd1

Renal ischemia-reperfusion injury (RIRI) is a common pathophysiological process, and it is also an important cause of acute renal failure. Therefore, finding an effective therapeutic target for RIRI is extremely urgent. In our study, we constructed hypoxia-reoxygenation (HR) model in vitro and a renal ischemia-reperfusion (IR) model in vivo. Elevated levels of serum creatinine (Cr), blood urea nitrogen (BUN) tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and malondialdehyde (MDA) along with the decreased levels of superoxide dismutase (SOD) and glutathione (GSH) proved that kidney function was damaged after IR, and pathological changes of renal tissues were observed using HE staining and TUNEL staining. The protein of Redd1 expression level was detected to be upregulated after IR by western blot (WB). However, transfection of short hairpin RNA of Redd1 (sh-Redd1) alleviated the HR injury on LLC-PK1 cells, as evidenced by increased cell viability, proliferation and decreased cell apoptosis; additionally, the accumulation of ROS was inhibited. Sh-Redd1 also alleviated IR injury in the mouse model. Subsequently, GATA2 was proved to be upregulated in IR and HR models and was the transcription factor of Redd1. Knockdown of GATA2 efficiently mitigated the oxidative stress induced damages in vivo and in vitro, while these mitigations were reversed by transfection of Redd1 overexpression plasmid. In conclusion, our study clarified the possible underlying mechanism of protecting RIRI.

Potential of Sodium-Glucose Cotransporter Inhibitors as Agents for Prevention of Ischemic and Reperfusion Kidney Injury

Sodium-glucose cotransporter 2 inhibitors are a relatively new class of glucose-lowering agents with significant advantages over other groups due to their good safety profile and protective effects on the cardiovascular system and kidneys. Today, the question of their ability to prevent and reduce the severity of acute kidney injury remains relevant. The primary objective of the study is to investigate the Potential of Sodium-Glucose Cotransporter Inhibitors as Agents for the Prevention of Ischemic and Reperfusion Kidney Injury. To gratify that objective, the experiment was performed on 80 Wistar line male rats. Acute kidney injury was simulated by reproducing a bilateral 40-minute renal ischemia-reperfusion. sodium-glucose cotransporter inhibitors were administered before surgery: dapagliflozin at doses of 0.5 mg/kg and 1 mg/kg, canagliflozin - 8.6 mg/kg and 25.7 mg/kg, empagliflozin - 1 mg/kg and 2 mg/kg. The renoprotective effects were evaluated after 72 hours based on the following parameters: serum creatinine and urea concentrations, glomerular filtration rate and fractional sodium excretion, as well as the level of renal microcirculation. Based on the results acquired, Preliminary administration of dapagliflozin, canagliflozin and empagliflozin led to a statistically significant decrease in the level of serum creatinine concentration and fractional sodium excretion, as well as an increase in the glomerular filtration rate, compared with the control group. The results demonstrated their dosedependent effect and ability to improve the parameters of renal microcirculation. Plus, the results of the study demonstrate a high dose-dependent renoprotective potential of sodium-glucose cotransporter 2 inhibitors (dapagliflozin, canagliflozin and empagliflozin) on a model of bilateral renal ischemia-reperfusion. The results of this study can greatly contribute to the respective field.

Intrarenal Arterial Transplantation of Dexmedetomidine Preconditioning Adipose Stem-Cell-Derived Microvesicles Confers Further Therapeutic Potential to Attenuate Renal Ischemia/Reperfusion Injury through miR-122-5p/Erythropoietin/Apoptosis Axis

Intravenous adipose mesenchymal stem cells (ADSCs) attenuate renal ischemia/reperfusion (IR) injury but with major drawbacks, including the lack of a specific homing effect after systemic infusion, cell trapping in the lung, and early cell death in the damaged microenvironment. We examined whether intrarenal arterial transplantation of dexmedetomidine (DEX) preconditioning ADSC-derived microvesicles (DEX-MVs) could promote further therapeutic potential to reduce renal IR injury. We evaluated the effect of DEX-MVs on NRK-52E cells migration, hypoxia/reoxygenation (H/R)-induced cell death, and reactive oxygen species (ROS) amount and renal IR model in rats. IR was established by bilateral 45 min ischemia followed by 4 h reperfusion. Intrarenal MVs or DEX-MVs were administered prior to ischemia. Renal oxidative stress, hemodynamics and function, western blot, immunohistochemistry, and tubular injury scores were determined. The miR-122-5p expression in kidneys was analyzed using microarrays and quantitative RT-PCR and its action target was predicted by TargetScan. DEX-MVs were more efficient than MVs to increase migration capability and to further decrease H/R-induced cell death and ROS level in NRK-52E cells. Consistently, DEX-MVs were better than MV in increasing CD44 expression, improving IR-depressed renal hemodynamics and renal erythropoietin expression, inhibiting IR-enhanced renal ROS level, tubular injury score, miR-122-5p expression, pNF-κB expression, Bax/caspase 3/poly(ADP-ribose) polymerase (PARP)-mediated apoptosis, blood urea nitrogen, and creatinine levels. The use of NRK-52E cells confirmed that miR-122-5p mimic via inhibiting erythropoietin expression exacerbated Bax-mediated apoptosis, whereas miR-122-5p inhibitor via upregulating erythropoietin and Bcl-2 expression reduced apoptosis. In summary, intrarenal arterial DEX-MV conferred further therapeutic potential to reduce renal IR injury through the miR-122-5p/erythropoietin/apoptosis axis.

Investigation of the effects of lipoic acid and dihydrolipoate on experimental renal ischemia-reperfusion model

Objective: Ischemic/reperfusion (I/R) causes tissue injury and the leading cause of acute kidney injury. In this study, we aimed to investigate the effects of the long and short-term usage of ALA and short-term DHLA on oxidative stress markers in the experimental renal ischemia-reperfusion model. Methods: Forty male rats (250 to 300 gr) were divided into 5 groups: control; I/R group; long-term ALA+IR group; short-term ALA+IR group; and short-term DHLA+IR group. Ischemia was carried out for 45 minutes followed by reperfusion for 4 hours. Thiobarbituric acid reactive sunstances (TBARM), catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px) activities in tissue samples and serum total antioxidant status (TAS) and total oxidative stress (TOS) assayed by the spectrophotometrically. Tissue samples were investigated by histopathological analyzes. Results: TBARM (Control: 0.38±0.05. I/R: 1.37±0.17, long-term ALA-treated group:1.025±0.15, short-term ALA-treated group: 0.68±0.09, short-term DHLA-treated group: 0.38±0.04 (nmol/mg protein); p<0,001) CAT (Control: 0.12±0.02, I/R: 0.04±0.008, long-term ALA-treated group: 0.07±0.01, short-term ALA-treated group:0.06±0.008, short-term DHLA-treated group: 0.08±0.01 (k/mg protein); p<0.001), GSH-Px (Control: 0.45±0.04, I/R: 0.21±0.028, long-term ALA-treated group: 0.37±0.05, short-term ALA-treated group :0.34±0.05, short-term DHLA-treated group: 0.37±0.04 (U/mg protein); p<0.001), and serum OSI levels (Control: 1.32±0.15, I/R: 3.08±0.44, long-term ALA-treated group: 1.775±0.21, short-term ALA-treated group: 1.85±0.37, short-term DHLA-treated group: 1.53±0.21 (arbitrary unit) ; p<0.001) were improved in the long and short-term ALA-treated group and short-term DHLA-treated group compared to the I/R group. These findings were more prominent in histopathological tissue samples in the DHLA-treated group. Conclusion: We consider that both long-term and short-term ALA applications have the potential for the treatment of renal I/R damage. Besides, DHLA is more effective than ALA.

Kidney-targeted irradiation triggers renal ischemic preconditioning in mice.

Renal ischemia-reperfusion (I/R) causes acute kidney injury (AKI). Ischemic preconditioning (IPC) attenuates I/R-associated AKI. Whole body irradiation induces renal IPC in mice. Still, the mechanisms remain largely unknown. Furthermore, the impact of kidney-centered irradiation on renal resistance against I/R has not been studied. Renal irradiation (8.5 Gy) was done in male 8- to 12-wk-old C57bl/6 mice using a small animal radiation therapy device. Left renal I/R was performed by clamping the renal pedicles for 30 min, with simultaneous right nephrectomy, at 7, 14, and 28 days postirradiation. The renal reperfusion lasted 48 h. Following I/R, blood urea nitrogen (BUN) and serum creatinine (SCr) levels were lower in preirradiated mice compared with controls; so was the histological Jablonski score of AKI. The metabolomics signature of renal I/R was attenuated in preirradiated mice. The numbers of proliferating cell nuclear antigen (PCNA)-, cluster of differentiation molecule 11b (CD11b)-, and cell surface glycoprotein F4/80-positive cells in the renal parenchyma post-I/R were reduced in preirradiated versus control groups. Such IPC was significantly observed as early as day 14 postirradiation. RNA sequencing showed an upregulation of angiogenesis- and stress response-related signaling pathways in irradiated nonischemic kidneys on day 28. Qualitative RT-PCR confirmed the increased expression of vascular endothelial growth factor (VEGF), activin receptor-like kinase 5 (ALK5), heme oxygenase-1 (HO1), platelet endothelial cell adhesion molecule-1 (PECAM1), NADPH oxidase 2 (NOX2), and heat shock proteins 70 and 27 (HSP70 and HSP27, respectively) in irradiated kidneys compared with controls. In addition, irradiated kidneys showed an increased CD31-positive vascular area compared with controls. A 14-day gavage of irradiated mice with the antiangiogenic drug sunitinib before I/R abrogated the irradiation-induced IPC at both functional and structural levels. Our observations suggest that kidney-centered irradiation activates proangiogenic pathways and induces IPC, with preserved renal function and attenuated inflammation post-I/R.NEW & NOTEWORTHY This study based on a mouse model of renal ischemia-reperfusion (I/R) aimed to 1) test whether and how irradiation strictly centered on the kidney protects against the I/R injury and 2) determine the shortest efficient delay of kidney irradiation to achieve such nephroprotection. Kidney irradiation increased the vascular surface in the renal parenchyma and conferred resistance against renal I/R damage, which highlights novel putative strategies in the field of ischemic acute kidney injury.

Renal Injuries after Cardiac Arrest: A Morphological Ultrastructural Study.

Background: This study aims to investigate the probable lesions and injuries induced in the renal tissue after a cardiac arrest. The renal ischemia–reperfusion model in cardiac arrest describes the effects of ischemia in the kidneys, alongside a whole-body ischemia–reperfusion injury. This protocol excludes ischemic conditions caused by surgical vascular manipulation, venous injury or venous congestion. Methods: For the experimental study, 24 swine were subjected to cardiac arrest. Seven minutes later, the cardiopulmonary resuscitation technique was performed for 5 min. Afterwards, advanced life support was provided. The resuscitated swine consisted one group and the non-resuscitated the other. Tissue samples were obtained from both groups for light and electron microscopy evaluation. Results: Tissue lesions were observed in the tubules, parallel to destruction of the microvilli, reduction in the basal membrane invaginations, enlarged mitochondria, cellular vacuolization, cellular apoptosis and disorganization. In addition, fusion of the podocytes, destruction of the Bowman’s capsule parietal epithelium and abnormal peripheral urinary space was observed. The damage appeared more extensive in the non-resuscitated swine group. Conclusions: Acute kidney injury is not the leading cause of death after cardiac arrest. However, evidence suggests that the kidney damage after a cardiac arrest should be highly considered in the prognosis of the patients’ health outcome.

Urolithin A alleviates acute kidney injury induced by renal ischemia reperfusion through the p62-Keap1-Nrf2 signaling pathway.

Acute kidney injury (AKI) induced by renal ischemia reperfusion (RIR) is typically observed in renal surgeries and is a leading cause of renal failure. However, there is still an unmet medical need currently in terms of clinical treatments. Herein, we report the effect of Urolithin A (UA) in a mouse RIR model, wherein we demonstrated its underlying mechanism both in vitro and in vivo. The expression levels of p62 and Keap1 significantly decreased, while that of nuclear Nrf2 increased in vitro in a hypoxia cell model after UA treatment. Furthermore, the apoptosis of tubular cells was attenuated and the reactive oxygen species (ROS) levels were reduced in the kidneys in a mouse RIR model after UA administration. In this study, we demonstrated that UA can alleviate oxidative stress and promote autophagy by activating the p62-Keap1-Nrf2 signaling pathway, which could protect the kidneys from ischemia reperfusion injury.