Quercetin enhances decidualization through AKT-ERK-p53 signaling and supports a role for senescence in endometriosis.

BACKGROUND: Patients with endometriosis suffer with chronic pelvic pain and infertility, and from the lack of pharmacologic therapies that consistently halt disease progression. Differences in the endometrium of patients with endometriosis vs. unaffected controls are well-documented. Specifically, shed endometrial tissues (delivered to the pelvic cavity via retrograde menstruation) reveal that a subset of stromal cells exhibiting pro-inflammatory, pro-fibrotic, and pro-senescence-like phenotypes is enhanced in endometriosis patients compared to controls. Additionally, cultured biopsy-derived endometrial stromal cells from endometriosis patients exhibit impaired decidualization, a defined differentiation process required for human embryo implantation and pregnancy. Quercetin, a senolytic agent, shows therapeutic potential for pulmonary fibrosis, a disorder attributed to senescent pulmonary fibroblasts. In rodent models of endometriosis, quercetin shows promise, and quercetin improves decidualization in vitro. However, the exact mechanisms are not completely understood. Therefore, we investigated the effects of quercetin on menstrual effluent-derived endometrial stromal cells from endometriosis patients and unaffected controls to define the signaling pathways underlying quercetin's effects on endometrial stromal cells. METHODS: Menstrual effluent-derived endometrial stromal cells were collected and cultured from unaffected controls and endometriosis patients and then, low passage cells were treated with quercetin (25 µM) under basal or standard decidualization conditions. Decidualization responses were analyzed by measuring the production of IGFBP1 and PRL. Also, the effects of quercetin on intracellular cAMP levels and cellular oxidative stress responses were measured. Phosphokinase arrays, western blotting, and flow cytometry methods were performed to define the effects of quercetin on various signaling pathways and the potential mechanistic roles of quercetin. RESULTS: Quercetin significantly promotes decidualization of control- and endometriosis-endometrial stromal cells. Quercetin substantially reduces the phosphorylation of multiple signaling molecules in the AKT and ERK1/2 pathways, while enhancing the phosphorylation of p53 and total p53 levels. Furthermore, p53 inhibition blocks decidualization while p53 activation promotes decidualization. Finally, we provide evidence that quercetin increases apoptosis of endometrial stromal cells with a senescent-like phenotype. CONCLUSIONS: These data provide insight into the mechanisms of action of quercetin on endometrial stromal cells and warrant future clinical trials to test quercetin and other senolytics for treating endometriosis.

Magnesium deficiency with high calcium-to-magnesium ratio promotes a metastatic phenotype in the CT26 colon cancer cell line.

Magnesium (Mg) plays important roles in maintaining genomic stability and cellular redox. Mg also serves as nature's physiological calcium (Ca) channel antagonist, controlling intracellular Ca entry. Because Ca is the most important second messenger, its intracellular concentration is tightly regulated. Excess intracellular Ca can activate aberrant signaling pathways leading to the acquisition of pathological characteristics and cell injury. Several epidemiological studies have linked Mg deficiency (MgD) and increased Ca:Mg ratios with higher incidences of colon cancer and increased mortality. While it is estimated that less than 50% of the US population consumes the recommended daily allowance for Mg, Ca supplementation is widespread. Therefore, we studied the effect of MgD, with variable Ca:Mg ratios on cellular oxidative stress, cell migration, calpain activity, and associated signaling pathways using the CT26 colon cancer cell line. MgD (with Ca:Mg ratios >1) elevated intracellular Ca levels, calpain activity and TRPM7 expression, as well as oxidative stress and cell migration, consistent with observed degradation of full-length E-cadherin, ß-catenin, and N-terminal FAK. MgD was accompanied by enhanced degradation of IκBα and the transactivation domain containing the C-terminus of NF-κB p65 (RelA). MgD-exposed CT26 cells exhibited increased p53 degradation and aneuploidy, markers of genomic instability. By contrast, these pathological changes were not observed when CT26 were cultured under MgD conditions where the Ca:Mg ratio was kept at 1. Together, these data support that exposure of colon cancer cells to MgD with physiological Ca concentrations (or increasing Ca:Mg ratios) leads to the acquisition of a more aggressive, metastatic phenotype.

Activation of the cholinergic anti-inflammatory pathway by GTS-21 attenuates cisplatin-induced acute kidney injury in mice.

Acute kidney injury (AKI) is the most common side effect of cisplatin, a widely used chemotherapy drug. Although AKI occurs in up to one third of cancer patients receiving cisplatin, effective renal protective strategies are lacking. Cisplatin targets renal proximal tubular epithelial cells leading to inflammation, reactive oxygen species, tubular cell injury, and eventually cell death. The cholinergic anti-inflammatory pathway is a vagus nerve-mediated reflex that suppresses inflammation via α7 nicotinic acetylcholine receptors (α7nAChRs). Our previous studies demonstrated the renoprotective and anti-inflammatory effects of cholinergic agonists, including GTS-21. Therefore, we examined the effect of GTS-21 on cisplatin-induced AKI. Male C57BL/6 mice received either saline or GTS-21 (4mg/kg, i.p.) twice daily for 4 days before cisplatin and treatment continued through euthanasia; 3 days post-cisplatin mice were euthanized and analyzed for markers of renal injury. GTS-21 significantly reduced cisplatin-induced renal dysfunction and injury (p<0.05). GTS-21 significantly attenuated renal Ptgs2/COX-2 mRNA and IL-6, IL-1ß, and CXCL1 protein expression, as well as neutrophil infiltration after cisplatin. GTS-21 blunted cisplatin-induced renal ERK1/2 activation, as well as renal ATP depletion and apoptosis (p<0.05). GTS-21 suppressed the expression of CTR1, a cisplatin influx transporter and enhanced the expression of cisplatin efflux transporters MRP2, MRP4, and MRP6 (p<0.05). Using breast, colon, and lung cancer cell lines we showed that GTS-21 did not inhibit cisplatin's tumor cell killing activity. GTS-21 protects against cisplatin-AKI by attenuating renal inflammation, ATP depletion and apoptosis, as well as by decreasing renal cisplatin influx and increasing efflux, without impairing cisplatin-mediated tumor cell killing. Our results support further exploring the cholinergic anti-inflammatory pathway for preventing cisplatin-induced AKI.

Magnesium improves cisplatin-mediated tumor killing while protecting against cisplatin-induced nephrotoxicity.

Approximately 30% of all cancer patients treated with cisplatin, a widely used broad-spectrum chemotherapeutic agent, experience acute kidney injury (AKI). Almost all patients receiving cisplatin have magnesium (Mg) losses, which are proposed to aggravate AKI. Currently, there are no methods to successfully treat or prevent cisplatin-AKI. Whereas Mg supplementation has been shown to reduce AKI in experimental models and several small clinical trials, the effects of Mg status on tumor outcomes in immunocompetent tumor-bearing mice and humans have not been investigated. The purpose of this study was to further examine the effects of Mg deficiency (±Mg supplementation) on cisplatin-mediated AKI and tumor killing in immunocompetent mice bearing CT26 colon tumors. Using a model where cisplatin alone (20 mg/kg cumulative dose) produced minimal kidney injury, Mg deficiency significantly worsened cisplatin-mediated AKI, as determined by biochemical markers (blood urea nitrogen and plasma creatinine) and histological renal changes, as well as markers of renal oxidative stress, inflammation, and apoptosis. By contrast, Mg supplementation blocked cisplatin-induced kidney injury. Using LLC-PK1 renal epithelial cells, we observed that Mg deficiency or inhibition of Mg uptake significantly enhanced cisplatin-induced cytotoxicity, whereas Mg supplementation protected against cytotoxicity. However, neither Mg deficiency nor inhibition of Mg uptake impaired cisplatin-mediated killing of CT26 tumor cells in vitro. Mg deficiency was associated with significantly larger CT26 tumors in BALB/c mice when compared with normal-fed control mice, and Mg deficiency significantly reduced cisplatin-mediated tumor killing in vivo. Finally, Mg supplementation did not compromise cisplatin's anti-tumor efficacy in vivo.

Magnesium protects against cisplatin-induced acute kidney injury without compromising cisplatin-mediated killing of an ovarian tumor xenograft in mice.

Cisplatin, a commonly used chemotherapeutic for ovarian and other cancers, leads to hypomagnesemia in most patients and causes acute kidney injury (AKI) in 25-30% of patients. Previously, we showed that magnesium deficiency worsens cisplatin-induced AKI and magnesium replacement during cisplatin treatment protects against cisplatin-mediated AKI in non-tumor-bearing mice (Solanki MH, Chatterjee PK, Gupta M, Xue X, Plagov A, Metz MH, Mintz R, Singhal PC, Metz CN. Am J Physiol Renal Physiol 307: F369-F384, 2014). This study investigates the role of magnesium in cisplatin-induced AKI using a human ovarian tumor (A2780) xenograft model in mice and the effect of magnesium status on tumor growth and the chemotherapeutic efficacy of cisplatin in vivo. Tumor progression was unaffected by magnesium status in saline-treated mice. Cisplatin treatment reduced tumor growth in all mice, irrespective of magnesium status. In fact, cisplatin-treated magnesium-supplemented mice had reduced tumor growth after 3 wk compared with cisplatin-treated controls. While magnesium status did not interfere with tumor killing by cisplatin, it significantly affected renal function following cisplatin. Cisplatin-induced AKI was enhanced by magnesium deficiency, as evidenced by increased blood urea nitrogen, creatinine, and other markers of renal damage. This was accompanied by reduced renal mRNA expression of the cisplatin efflux transporter Abcc6. These effects were significantly reversed by magnesium replacement. On the contrary, magnesium status did not affect the mRNA expression of cisplatin uptake or efflux transporters by the tumors in vivo. Finally, magnesium deficiency enhanced platinum accumulation in the kidneys and renal epithelial cells, but not in the A2780 tumor cells. These findings demonstrate the renoprotective role of magnesium during cisplatin AKI, without compromising the chemotherapeutic efficacy of cisplatin in an ovarian tumor-bearing mouse model.

Omega-3 polyunsaturated fatty acids enhance cytokine production and oxidative stress in a mouse model of preterm labor.

OBJECTIVE: Omega-3 polyunsaturated fatty acid (ω-3 PUFA) supplementation during pregnancy remains controversial. We sought to examine the effects of ω-3 PUFA on inflammation and oxidative stress in vitro and in vivo using a model of preterm labor. METHODS: In vivo. Female Swiss Webster mice were fed a normal diet or a 5% fish oil (FO) diet for 3 weeks then mated with normal-fed males. On gestational day 15, dams were injected with either saline (n=10 per group) or lipopolysaccharide (LPS, intrauterine) (n=10 per group). Maternal plasma, amniotic fluid, placentas, and uteri were collected 4 h later and assessed for cytokines; maternal plasma and amniotic fluids were analyzed for oxidative stress. In vitro. RAW264.7 mouse macrophage-like cells were treated with either: vehicle, H2O2, docosahexaenoic acid (DHA), or eicosapentaenoic acid (EPA) (0, 0.1-100 µM) and analyzed for oxidative stress. RESULTS: In vivo. Administration of the 5% FO diet enhanced LPS-induced cytokines in the placenta (P<0.05-0.01) and increased tumor necrosis factor-α in the uterus (P<0.05) and amniotic fluid (P<0.01) when compared to LPS-treated normal-fed animals. Maternal plasma obtained from FO-fed dams showed higher LPS-induced oxidative stress than control-fed animals (P<0.035). However, no differences in oxidative stress were observed in the amniotic fluid. In vitro. Treatment of macrophage-like cells with ω-3 PUFA significantly and dose-dependently increased oxidative stress (P<0.001-0.0001). CONCLUSIONS: Supplementation with FO for prior to and during pregnancy significantly increased LPS-induced inflammation in the amniotic fluid, uterus, and placenta and significantly increased maternal systemic oxidative stress in vivo. Likewise, DHA and EPA induced oxidative stress in macrophage-like cells in vitro.

Magnesium protects against cisplatin-induced acute kidney injury by regulating platinum accumulation.

Despite its success as a potent antineoplastic agent, ∼25% of patients receiving cisplatin experience acute kidney injury (AKI) and must discontinue therapy. Impaired magnesium homeostasis has been linked to cisplatin-mediated AKI, and because magnesium deficiency is widespread, we examined the effect of magnesium deficiency and replacement on cisplatin-induced AKI in physiologically relevant older female mice. Magnesium deficiency significantly increased cisplatin-associated weight loss and markers of renal damage (plasma blood urea nitrogen and creatinine), histological changes, inflammation, and renal cell apoptosis and modulated signaling pathways (e.g., ERK1/2, p53, and STAT3). Conversely, these damaging effects were reversed by magnesium. Magnesium deficiency alone significantly induced basal and cisplatin-mediated oxidative stress, whereas magnesium replacement attenuated these effects. Similar results were observed using cisplatin-treated LLC-PK1 renal epithelial cells exposed to various magnesium concentrations. Magnesium deficiency significantly amplified renal platinum accumulation, whereas magnesium replacement blocked the augmented platinum accumulation after magnesium deficiency. Increased renal platinum accumulation during magnesium deficiency was accompanied by reduced renal efflux transporter expression, which was reversed by magnesium replacement. These findings demonstrate the role of magnesium in regulating cisplatin-induced AKI by enhancing oxidative stress and thus promoting cisplatin-mediated damage. Additional in vitro experiments using ovarian, breast, and lung cancer cell lines showed that magnesium supplementation did not compromise cisplatin's chemotherapeutic efficacy. Finally, because no consistently successful therapy to prevent or treat cisplatin-mediated AKI is available for humans, these results support developing more conservative magnesium replacement guidelines for reducing cisplatin-induced AKI in cancer patients at risk for magnesium deficiency.

Maternal metformin treatment decreases fetal inflammation in a rat model of obesity and metabolic syndrome.

OBJECTIVE: Obesity and metabolic syndrome are associated with systemic inflammation and increased perinatal morbidity. Metformin improves metabolic and inflammatory biomarkers in nonpregnant adults. Using in vivo and in vitro models, we examined the effect of metformin on maternal and fetal inflammation. STUDY DESIGN: Female Wistar rats (6-7 weeks old) were fed a normal diet (NORM) or a high-fat/high-sugar diet (HCAL) for 5-6 weeks to induce obesity/metabolic syndrome. After mating with NORM-fed male rats, one-half of the HCAL-fed female rats received metformin (300 mg/kg, by mouth daily). All dams continued their respective diets until gestational day 19, at which time maternal and fetal outcomes were assessed. Maternal and fetal plasma and placentas were analyzed for metabolic and inflammatory markers. Cultured human placental JAR cells were pretreated with vehicle or metformin (10 µmol/L-2.5 mmol/L) before tumor necrosis factor α (TNF-α; 50 ng/mL), and supernatants were assayed for interleukin-6 (IL-6). RESULTS: HCAL rats gained more prepregnancy weight than NORM rats (P = .03), had higher levels of plasma insulin and leptin, and exhibited dyslipidemia (P < .05). Fetuses that were exposed to the HCAL diet had elevated plasma IL-6, TNF-α, and chemokine (C-C motif) ligand 2 levels (P < .05) and enhanced placental TNF-α levels (P < .05). Maternal metformin did not impact maternal markers but significantly decreased diet-induced TNF-α and chemokine (C-C motif) ligand 2 in the fetal plasma. Finally, metformin dose-dependently reduced TNF-α-induced IL-6 and IκBα levels in cultured placental JAR cells. CONCLUSION: Diet induced-obesity/metabolic syndrome during pregnancy significantly enhanced fetal and placental cytokine production; maternal metformin reduced fetal cytokine levels. Similarly, metformin treatment of a placental cell line suppressed TNF-α-induced IL-6 levels by NFκB inhibitor.

Maternal magnesium supplementation reduces intrauterine growth restriction and suppresses inflammation in a rat model.

OBJECTIVE: Intrauterine growth restriction (IUGR) is associated with increased inflammatory responses. We sought to investigate whether magnesium (Mg) attenuates inflammation and IUGR in a rat model. STUDY DESIGN: Pregnant Wistar rats (12 weeks, gestational day 18) were randomly assigned to 1 of 4 groups: normal diet with bilateral uterine artery ligation (BL) (n = 6) or sham surgery (SH) (n = 5); and Mg chloride (MgCl2) 1% (wt/vol) in the drinking water throughout gestation + BL (MgBL) (n = 6) or SH (MgSH) (n = 5). Dams were euthanized 24 hours postsurgery (gestational day 19). Maternal plasma, fetal plasma (pooled), individual amniotic fluid (AF) samples, and placentas (PL) were collected and assessed from live fetal pups only (BL, n = 36; SH, n = 20; MgBL, n = 20; MgSH, n = 20). All samples were analyzed for cytokines/chemokines (interleukin [IL]-6, IL-1ß, chemokine [C-X-C motif] ligand 1 [CXCL1], chemokine [C-C motif] ligand 2 [CCL2], and tumor necrosis factor [TNF-α] sensitivity <3 pg/mL) using a multiplex platform. Data were analyzed using Mann Whitney, analysis of variance, and Fisher exact tests. RESULTS: The incidence of IUGR (pup weight <10th percentile of SH) in the MgBL group was significantly lower (31%) than the BL group (86.3%) (relative risk, 0.36; 95% confidence interval, 0.2-0.6; P < .0001). BL significantly increased AF levels of IL-6, IL-1ß, TNF-α (P < .05), and CCL2 (P < .001) vs SH and PL levels of IL-6, IL-1ß, CCL2 and CXCL1 (P < .001), and TNF-α (P < .05) vs SH. Maternal MgCl2 supplementation significantly decreased IL-1ß, TNF-α, and CCL2 levels in AF and IL-1ß in PL tissues of MgBL vs BL rats (P < .0001). CONCLUSION: Maternal oral MgCl2 supplementation reduced BL-induced IUGR by 64% and suppressed cytokine/chemokine levels in the AF and PL.

Magnesium sulfate ameliorates maternal and fetal inflammation in a rat model of maternal infection.

OBJECTIVE: Magnesium sulfate is proposed to have neuroprotective effects in the offspring. We examined the effects of maternal magnesium sulfate administration on maternal and fetal inflammatory responses in a rat model of maternal infection. STUDY DESIGN: Pregnant rats were injected with saline, Gram-negative bacterial endotoxin lipopolysaccharide or lipopolysaccharide with magnesium sulfate (pre- and/or after lipopolysaccharide) to mimic infection. Maternal blood, amniotic fluid, fetal blood, and fetal brains were collected 4 hours after lipopolysaccharide and assayed for tumor necrosis factor, interleukin-6, monocyte chemoattractant protein-1, and growth-related oncogene-KC. In addition, the effect of magnesium sulfate on cytokine production by an astrocytoma cell line was assessed. RESULTS: Lipopolysaccharide administration induced tumor necrosis factor, interleukin-6, monocyte chemoattractant protein-1, and growth-related oncogene-KC expression in maternal and fetal compartments. Maternal magnesium sulfate treatment significantly attenuated lipopolysaccharide-induced multiple proinflammatory mediator levels in maternal and fetal compartments. CONCLUSION: Antenatal magnesium sulfate administration significantly ameliorated maternal, fetal, and gestational tissue-associated inflammatory responses in an experimental model of maternal infection.

Progesterone suppresses the fetal inflammatory response ex vivo.

OBJECTIVE: Progesterone supplementation has been shown to be efficacious in preventing preterm birth. We sought to investigate the effects of progesterone on fetal inflammatory responses. STUDY DESIGN: Fetal mononuclear cells were isolated from umbilical cord blood and exposed to vehicle or progesterone (P4) for 1 hour prior to lipopolysaccharide (LPS) stimulation. Supernatants were assayed for tumor necrosis factor-alpha. Similar experiments were performed using cyclic adenosine monophosphate (cAMP) and progesterone modulators. The effect of P4 treatment on intracellular cAMP levels was also determined. RESULTS: LPS treatment led to a significant increase in cytokine production by fetal mononuclear cells. Despite the lack of detectable nuclear progesterone receptors, P4 suppressed this inflammatory response. R5020 (progesterone agonist), forskolin (cAMP inducer), and dibutyryl cAMP (cAMP agonist) all achieved immunosuppression. The cAMP antagonist, Rp-cAMP, blocked the inhibitory effect of progesterone. P4 significantly increased intracellular cAMP levels. CONCLUSION: Progesterone rapidly suppresses the fetal inflammatory response, possibly via nongenomic activation of the cAMP cascade.

Cholinergic neural signals to the spleen down-regulate leukocyte trafficking via CD11b.

The cholinergic anti-inflammatory pathway is a physiological mechanism that inhibits cytokine production and diminishes tissue injury during inflammation. Recent studies demonstrate that cholinergic signaling reduces adhesion molecule expression and chemokine production by endothelial cells and suppresses leukocyte migration during inflammation. It is unclear how vagus nerve stimulation regulates leukocyte trafficking because the vagus nerve does not innervate endothelial cells. Using mouse models of leukocyte trafficking, we show that the spleen, which is a major point of control for cholinergic modulation of cytokine production, is essential for vagus nerve-mediated regulation of neutrophil activation and migration. Administration of nicotine, a pharmacologic agonist of the cholinergic anti-inflammatory pathway, significantly reduces levels of CD11b, a beta(2)-integrin involved in cell adhesion and leukocyte chemotaxis, on the surface of neutrophils in a dose-dependent manner and this function requires the spleen. Similarly, vagus nerve stimulation significantly attenuates neutrophil surface CD11b levels only in the presence of an intact and innervated spleen. Further mechanistic studies reveal that nicotine suppresses F-actin polymerization, the rate-limiting step for CD11b surface expression. These studies demonstrate that modulation of leukocyte trafficking via cholinergic signaling to the spleen is a specific, centralized neural pathway positioned to suppress the excessive accumulation of neutrophils at inflammatory sites. Activating this mechanism may have important therapeutic potential for preventing tissue injury during inflammation.

A pilot study to examine the effects of smoking cessation on serum markers of inflammation in women at risk for cardiovascular disease.

BACKGROUND: The links among smoking, inflammation, and cardiovascular disease (CVD) are well established. Several studies have demonstrated that quitting smoking reverses the risk of coronary heart disease within 5 to 10 years. However, the immediate effects of quitting smoking on inflammatory biomarkers associated with CVD risk have not been well described. METHODS: In this pilot study, we examined a panel of circulating inflammatory biomarkers associated with CVD in "at-risk" women during the smoking cessation program. Forty-six women enrolled in a smoking cessation program consented to attend four study visits over 6 to 7 weeks. Health/medical information and blood were collected at each visit. Circulating levels of C-reactive protein (CRP), tumor necrosis factor (TNF), interleukin (IL)-6, soluble TNF receptor (sTNFR)-I, sTNFR-II, and soluble vascular cell adhesion molecule (sVCAM)-1 were measured, and changes between baseline levels (visit 1, while smoking) and visits 2 through 4 were determined. RESULTS: Significant reductions in circulating levels of TNF, sTNFR-I, sTNFR-II, and sVCAM-1 were observed among participants over the course of the smoking cessation program. Serum levels of both IL-6 and CRP declined during the smoking cessation program; the changes were not statistically significant, however. CONCLUSIONS: These findings suggest there are rapid consequences of smoking cessation on inflammatory biomarkers in women at risk for CVD. Additional, larger studies including diverse smokers desiring to quit are required to confirm changes in "measurable milestones" that could serve as motivating factors to assist smokers to quit.

Nicotinic acetylcholine receptor expression and regulation in the rat kidney after ischemia-reperfusion injury.

The cholinergic anti-inflammatory pathway is a mechanism whereby local inflammation is modulated by the brain via the vagus nerve and nicotinic acetylcholine receptors (nAChRs). The nAChR family are ligand-gated ion channels that consist of many different subtypes formed by the specific assembly of five polypeptide subunits including alpha1-10, beta1-4, gamma, delta, and epsilon. The alpha7 receptor (alpha7nAChR) mediates the anti-inflammatory effects of cholinergic stimulation. We recently demonstrated that cholinergic agonists attenuate renal ischemia-reperfusion (I/R) injury in rats. We also showed that tubular epithelial cells express functional nAChRs in vitro. The current studies report the expression, localization, and regulation of the alpha7nAChR in the rat kidney after I/R injury. We also examined, in this model, potential interactions between cholinergic stimulation and the STAT3 pathway, a key signaling cascade that has been linked to alpha7nAChR activation. RT-PCR and immunohistochemistry showed constitutive expression of many nAChR subunits. Immunohistochemistry localized basal alpha7nAChR expression to the endothelium of cortical peritubular capillaries, and its distribution was upregulated after I/R injury. Western blotting also showed an increase in alpha7nAChR subunit protein after renal I/R injury. Interestingly, pretreatment with nicotine, which improves the outcome after renal I/R injury, reduced the alpha7nAChR protein after I/R injury. Finally, we found that I/R injury stimulated the STAT3 pathway, whereas pretreatment with nicotine downregulated its activation. These results suggest that the alpha7nAChR plays an important role in the pathophysiology of renal I/R injury.