Isoliquiritigenin pretreatment regulates ER stress and attenuates cisplatin-induced nephrotoxicity in male Wistar rats.

Cisplatin (CP) is a chemotherapeutic drug used to treat solid tumors. However, studies have revealed its nephrotoxic effect. Oxidative stress, endoplasmic reticulum (ER) stress, and mitochondrial dysfunction are involved in CP-induced renal damage. Thus, preconditioning (hormetic effect) of ER stress is a strategy to prevent CP-induced renal damage. On the other hand, isoliquiritigenin (IsoLQ) is recognized as a flavonoid with antioxidant properties and an inducer of ER stress. Therefore, we evaluated the ER stress-inducing capacity of IsoLQ and its possible protective effect against CP-induced nephrotoxicity in adult male Wistar rats. The findings reflected that IsoLQ pretreatment might decrease renal damage by reducing plasma creatinine and blood urea nitrogen levels in animals with CP-induced nephrotoxicity. These may be associated with IsoLQ activating ER stress and unfolded protein response (UPR). We found increased messenger RNA levels of the ER stress marker glucose-related protein 78 kDa (GRP78). In addition, we also found that pretreatment with IsoLQ reduced the levels of CCAAT/enhancer-binding protein-homologous protein (CHOP) and X-box-binding protein 1 (XBP1) in the renal cortex, reflecting that IsoLQ can regulate the UPR and activation of the apoptotic pathway. Moreover, this preconditioning with IsoLQ of ER stress had oxidative stress-regulatory effects, as it restored the activity of glutathione peroxidase and glutathione reductase enzymes. Finally, IsoLQ modifies the protein expression of mitofusin 2 (Mfn-2) and voltage-dependent anion channel (VDAC). In conclusion, these data suggest that IsoLQ pretreatment has a nephroprotective effect; it could functionally regulate the ER and mitochondria and reduce CP-induced renal damage by attenuating hormesis-mediated ER stress.

Therapeutic Effect of Curcumin on 5/6Nx Hypertriglyceridemia: Association with the Improvement of Renal Mitochondrial ß-Oxidation and Lipid Metabolism in Kidney and Liver.

Chronic kidney disease (CKD) prevalence is constantly increasing, and dyslipidemia in this disease is characteristic, favoring cardiovascular events. However, the mechanisms of CKD dyslipidemia are not fully understood. The use of curcumin (CUR) in CKD models such as 5/6 nephrectomy (5/6Nx) has shown multiple beneficial effects, so it has been proposed to correct dyslipidemia without side effects. This work aimed to characterize CUR's potential therapeutic effect on dyslipidemia and alterations in lipid metabolism and mitochondrial ß-oxidation in the liver and kidney in 5/6Nx. Male Wistar rats were subjected to 5/6Nx and progressed by 4 weeks; meanwhile, CUR (120 mg/kg) was administered for weeks 5 to 8. Our results showed that CUR reversed the increase in liver and kidney damage and hypertriglyceridemia induced by 5/6Nx. CUR also reversed mitochondrial membrane depolarization and ß-oxidation disorders in the kidney and the increased lipid uptake and the high levels of proteins involved in fatty acid synthesis in the liver and kidney. CUR also decreased lipogenesis and increased mitochondrial biogenesis markers in the liver. Therefore, we concluded that the therapeutic effect of curcumin on 5/6Nx hypertriglyceridemia is associated with the restoration of renal mitochondrial ß-oxidation and the reduction in lipid synthesis and uptake in the kidneys and liver.

A ketogenic diet attenuates acute and chronic ischemic kidney injury and reduces markers of oxidative stress and inflammation.

BACKGROUND: Ischemic kidney injury is a common clinical condition resulting from transient interruption of the kidney's normal blood flow, leading to oxidative stress, inflammation, and kidney dysfunction. The ketogenic diet (KD), a low-carbohydrate, high-fat diet that stimulates endogenous ketone body production, has potent antioxidant and anti-inflammatory effects in distinct tissues and might thus protect the kidney against ischemia and reperfusion (IR) injury. MAIN METHODS: Male Wistar rats were fed a KD or a control diet (CD) for three days before analyzing metabolic parameters or testing nephroprotection. We used two different models of kidney IR injury and conducted biochemical, histological, and Western blot analyses at 24 h and two weeks after surgery. KEY FINDINGS: Acute KD feeding caused protein acetylation, liver AMPK activation, and increased resistance to IR-induced kidney injury. At 24 h after IR, rats on KD presented reduced tubular damage and improved kidney functioning compared to rats fed with a CD. KD attenuated oxidative damage (protein nitration, 4-HNE adducts, and 8-OHdG), increased antioxidant defenses (GPx and SOD activity), and reduced inflammatory intermediates (IL6, TNFα, MCP1), p50 NF-κB expression, and cellular infiltration. Also, KD prevented interstitial fibrosis development at two weeks, up-regulation of HSP70, and chronic Klotho deficiency. SIGNIFICANCE: Our findings demonstrate for the first time that short-term KD increases tolerance to experimental kidney ischemia, opening the opportunity for future therapeutic exploration of a dietary preconditioning strategy to convey kidney protection in the clinic.

Unilateral Ureteral Obstruction for 28 Days in Rats Is Not Associated with Changes in Cardiac Function or Alterations in Mitochondrial Function.

Our work evaluated cardiac function and mitochondrial bioenergetics parameters in hearts from male Wistar rats subjected to the UUO model during 28 days of progression. We measured markers of kidney damage and inflammation in plasma and renal fibrosis by histological analysis and Western blot. Cardiac function was evaluated by echocardiography and proteins involved in cardiac damage by Western blot. Oxygen consumption and transmembrane potential were monitored in cardiac mitochondria using high-resolution respirometry. We also determined the activity of ATP synthase and antioxidant enzymes such as glutathione peroxidase, glutathione reductase, and catalase. Our results show that, although renal dysfunction is established in animals subjected to ureteral obstruction, cardiac function is maintained along with mitochondrial function and antioxidant enzymes activity after 28 days of injury evolution. Our results suggest that renocardiac syndrome might develop but belatedly in obstruction-induced renal damage, opening the opportunity for treatment to prevent this condition.

Anti-Inflammatory Effect of Allicin Associated with Fibrosis in Pulmonary Arterial Hypertension.

Pulmonary arterial hypertension (PAH) is characterized by pulmonary vascular remodeling. Recent evidence supports that inflammation plays a key role in triggering and maintaining pulmonary vascular remodeling. Recent studies have shown that garlic extract has protective effects in PAH, but the precise role of allicin, a compound derived from garlic, is unknown. Thus, we used allicin to evaluate its effects on inflammation and fibrosis in PAH. Male Wistar rats were divided into three groups: control (CON), monocrotaline (60 mg/kg) (MCT), and MCT plus allicin (16 mg/kg/oral gavage) (MCT + A). Right ventricle (RV) hypertrophy and pulmonary arterial medial wall thickness were determined. IL-1ß, IL-6, TNF-α, NFκB p65, Iκß, TGF-ß, and α-SMA were determined by Western blot analysis. In addition, TNF-α and TGF-ß were determined by immunohistochemistry, and miR-21-5p and mRNA expressions of Cd68, Bmpr2, and Smad5 were determined by RT-qPCR. Results: Allicin prevented increases in vessel wall thickness due to TNF-α, IL-6, IL-1ß, and Cd68 in the lung. In addition, TGF-ß, α-SMA, and fibrosis were lower in the MCT + A group compared with the MCT group. In the RV, allicin prevented increases in TNF-α, IL-6, and TGF-ß. These observations suggest that, through the modulation of proinflammatory and profibrotic markers in the lung and heart, allicin delays the progression of PAH.

Progressive Reduction in Mitochondrial Mass Is Triggered by Alterations in Mitochondrial Biogenesis and Dynamics in Chronic Kidney Disease Induced by 5/6 Nephrectomy.

The five-sixth nephrectomy (5/6Nx) model is widely used to study the mechanisms involved in chronic kidney disease (CKD) progression. Mitochondrial impairment is a critical mechanism that favors CKD progression. However, until now, there are no temporal studies of the change in mitochondrial biogenesis and dynamics that allow determining the role of these processes in mitochondrial impairment and renal damage progression in the 5/6Nx model. In this work, we determined the changes in mitochondrial biogenesis and dynamics markers in remnant renal mass from days 2 to 28 after 5/6Nx. Our results show a progressive reduction in mitochondrial biogenesis triggered by reducing two principal regulators of mitochondrial protein expression, the peroxisome proliferator-activated receptor-gamma coactivator 1-alpha and the peroxisome proliferator-activated receptor alpha. Furthermore, the reduction in mitochondrial biogenesis proteins strongly correlates with the increase in renal damage markers. Additionally, we found a slow and gradual change in mitochondrial dynamics from fusion to fission, favoring mitochondrial fragmentation at later stages after 5/6Nx. Together, our results suggest that 5/6Nx induces the progressive reduction in mitochondrial mass over time via the decrease in mitochondrial biogenesis factors and a slow shift from mitochondrial fission to fusion; both mechanisms favor CKD progression in the remnant renal mass.

Effects of Allicin on Pathophysiological Mechanisms during the Progression of Nephropathy Associated to Diabetes.

This study aimed to assess the impact of allicin on the course of diabetic nephropathy. Study groups included control, diabetes, and diabetes-treated rats. Allicin treatment (16 mg/kg day/p.o.) started after 1 month of diabetes onset and was administered for 30 days. In the diabetes group, the systolic blood pressure (SBP) increased, also, the oxidative stress and hypoxia in the kidney cortex were evidenced by alterations in the total antioxidant capacity as well as the expression of nuclear factor (erythroid-derived 2)-like 2/Kelch ECH associating protein 1 (Nrf2/Keap1), hypoxia-inducible factor 1-alpha (HIF-1α), vascular endothelial growth factor (VEGF), erythropoietin (Epo) and its receptor (Epo-R). Moreover, diabetes increased nephrin, and kidney injury molecule-1 (KIM-1) expression that correlated with mesangial matrix, the fibrosis index and with the expression of connective tissue growth factor (CTGF), transforming growth factor-ß1 (TGF-ß1), and α-smooth muscle actin (α-SMA). The insulin levels and glucose transporter protein type-4 (GLUT4) expression were decreased; otherwise, insulin receptor substrates 1 and 2 (IRS-1 and IRS-2) expression was increased. Allicin increased Nrf2 expression and decreased SBP, Keap1, HIF-1α, and VEGF expression. Concurrently, nephrin, KIM-1, the mesangial matrix, fibrosis index, and the fibrotic proteins were decreased. Additionally, allicin decreased hyperglycemia, improved insulin levels, and prevented changes in (GLUT4) and IRSs expression induced by diabetes. In conclusion, our results demonstrate that allicin has the potential to help in the treatment of diabetic nephropathy. The cellular mechanisms underlying its effects mainly rely on the regulation of antioxidant, antifibrotic, and antidiabetic mechanisms, which can contribute towards delay in the progression of renal disease.

Temporal Alterations in Mitochondrial ß-Oxidation and Oxidative Stress Aggravate Chronic Kidney Disease Development in 5/6 Nephrectomy Induced Renal Damage.

Five-sixths nephrectomy (5/6Nx) model is widely used for studying the mechanisms involved in chronic kidney disease (CKD) progression, a kidney pathology that has increased dramatically in recent years. Mitochondrial impairment is a key mechanism that aggravates CKD progression; however, the information on mitochondrial bioenergetics and redox alterations along a time course in a 5/6Nx model is still limited and in some cases contradictory. Therefore, we performed for the first time a time-course study of mitochondrial alterations by high-resolution respirometry in the 5/6Nx model. Our results show a decrease in mitochondrial ß-oxidation at early times, as well as a permanent impairment in adenosine triphosphate (ATP) production in CI-linked respiration, a permanent oxidative state in mitochondria and decoupling of these organelles. These pathological alterations are linked to the early decrease in complex I and ATP synthase activities and to the further decrease in complex III activity. Therefore, our results may suggest that mitochondrial bioenergetics impairment is an early event in renal damage, whose persistence in time aggravates CKD development in the 5/6Nx model.

Protection against renal ischemia and reperfusion injury by short-term time-restricted feeding involves the mitochondrial unfolded protein response.

Food restriction improves metabolic health and increases resistance to stress in experimental animals. However, most studies have focused on long-term dietary restriction protocols consisting of several weeks or months of limited food ingestion. Here it was investigated the impact of 2-h time-restricted feeding (TRF) for one week on stress resistance in a rat model of kidney injury induced by ischemia and reperfusion (IR). At baseline, TRF reduced blood glucose, increased ß-hydroxybutyrate and improved body composition in male Wistar rats. Importantly, implementing the one-week TRF schedule before ischemia significantly improved renal function, suppressed tubular injury, prevented the activation of extracellular signal-regulated kinase 1/2 (ERK1/2) and inhibited the development of interstitial fibrosis. These benefits were related to increased antioxidant protection, reduction in dynamin-related protein 1 (DRP1)-mediated mitochondrial fragmentation and modulation of the mitochondrial unfolded protein response (UPRmt). Specifically, preoperative TRF boosted the activation of the UPRmt in the acute phase after renal IR while promoted its resolution at the stage of fibrosis. Our study indicates that dietary preconditioning by short-term TRF improves the outcome of renal IR injury, and suggests that an optimal intervention that promotes kidney protection may not necessarily require adherence to restrictive diets for prolonged periods of time.

Chronic impairment of mitochondrial bioenergetics and ß-oxidation promotes experimental AKI-to-CKD transition induced by folic acid.

Recent studies suggest that mitochondrial bioenergetics and oxidative stress alterations may be common mechanisms involved in the progression of renal damage. However, the evolution of the mitochondrial alterations over time and the possible effects that their prevention could have in the progression of renal damage are not clear. Folic acid (FA)-induced kidney damage is a widely used experimental model to induce acute kidney injury (AKI), which can evolve to chronic kidney disease (CKD). Therefore, it has been extensively applied to study the mechanisms involved in AKI-to-CKD transition. We previously demonstrated that one day after FA administration, N-acetyl-cysteine (NAC) pre-administration prevented the development of AKI induced by FA. Such therapeutic effect was related to mitochondrial preservation. In the present study, we characterized the temporal course of mitochondrial bioenergetics and redox state alterations along the progression of renal damage induced by FA. Mitochondrial function was studied at different time points and showed a sustained impairment in oxidative phosphorylation capacity and a decrease in ß-oxidation, decoupling, mitochondrial membrane potential depolarization and a pro-oxidative state, attributed to the reduction in activity of complexes I and III and mitochondrial cristae effacement, thus favoring the transition from AKI to CKD. Furthermore, the mitochondrial protection by NAC administration before AKI prevented not only the long-term deterioration of mitochondrial function at the chronic stage, but also CKD development. Taken together, our results support the idea that the prevention of mitochondrial dysfunction during an AKI event can be a useful strategy to prevent the transition to CKD.

Ketone bodies, stress response, and redox homeostasis.

The ketone body ß-hydroxybutyrate is no longer viewed simply as a metabolic intermediate, as it regulates a broad range of physiological processes at cellular and systemic levels. Particularly, ß-hydroxybutyrate functions as a stress response molecule and orchestrates an antioxidant defense program to maintain redox homeostasis in response to environmental and metabolic challenges, such as ischemia. This property of ß-hydroxybutyrate might be key for the beneficial effect of calorie restriction on stress response and disease processes.

Mechanisms of Fasting-Mediated Protection against Renal Injury and Fibrosis Development after Ischemic Acute Kidney Injury.

Ischemia-reperfusion injury of the kidney may lead to renal fibrosis through a combination of several mechanisms. We recently demonstrated that fasting protects the rat kidney against oxidative stress and mitochondrial dysfunction in early acute kidney injury, and also against fibrosis development. Here we show that preoperative fasting preserves redox status and mitochondrial homeostasis at the chronic phase of damage after severe ischemia. Also, the protective effect of fasting coincides with the suppression of inflammation and endoplasmic reticulum stress, as well as the down-regulation of the mechanistic target of rapamycin (mTOR) and extracellular signal-regulated kinase 1/2 (ERK1/2) signaling pathways in the fibrotic kidney. Our results demonstrate that fasting targets multiple pathophysiological mechanisms to prevent renal fibrosis and damage that results after renal ischemia-reperfusion injury.

Fasting reduces oxidative stress, mitochondrial dysfunction and fibrosis induced by renal ischemia-reperfusion injury.

Food deprivation protects against ischemia-reperfusion (IR) injury through unknown mechanisms. In an experimental rat model of acute IR injury, we found that preoperative fasting for 3 days protects rats from tubular damage and renal functional decline by increasing antioxidant protection independently of the NF-E2-related factor 2 (Nrf2), and by maintaining mitochondrial morphology and function. In addition, further analysis revealed that fasting protects against tubulointerstitial fibrosis. In summary, our results point out to fasting as a robust nutritional intervention to limit oxidative stress and mitochondrial dysfunction in early acute kidney injury and also to promote long-term protection against fibrosis.

ß-Hydroxybutyrate: A signaling metabolite in starvation response?

Ketone bodies ß-hydroxybutyrate (BHB) and acetoacetate are important metabolic substrates for energy production during prolonged fasting. However, BHB also has signaling functions. Through several metabolic pathways or processes, BHB modulates nutrient utilization and energy expenditure. These findings suggest that BHB is not solely a metabolic intermediate, but also acts as a signal to regulate metabolism and maintain energy homeostasis during nutrient deprivation. We briefly summarize the metabolism and emerging physiological functions of ketone bodies and highlight the potential role for BHB as a signaling molecule in starvation response.