Dihydromyricetin Modulates Nrf2 and NF-κB Crosstalk to Alleviate Methotrexate-Induced Lung Toxicity.

BACKGROUND: Methotrexate (MTX) is an effective anticancer, anti-inflammatory, and immunomodulatory agent. However, it induces a serious pneumonitis that leads to irreversible fibrotic lung damage. This study addresses the protective role of the natural flavonoid dihydromyricetin (DHM) against MTX-induced pneumonitis via modulation of Nrf2/NF-κB signaling crosstalk. METHODS: Male Wistar rats were divided into 4 groups: control, which received the vehicle; MTX, which received a single MTX (40 mg/kg, i.p) at day 9 of the experiment; (MTX + DHM), which received oral DHM (300 mg/kg) for 14 days and methotrexate (40 mg/kg, i.p) on the 9th day; and DHM, which received DHM (300 mg/kg, p.o) for 14 days. RESULTS: Lung histopathological examination and scoring showed a decline in MTX-induced alveolar epithelial damage and decreased inflammatory cell infiltration by DHM treatment. Further, DHM significantly alleviated the oxidative stress by decreasing MDA while increasing GSH and SOD antioxidant levels. Additionally, DHM suppressed the pulmonary inflammation and fibrosis through decreasing levels of NF-κB, IL-1ß, and TGF-ß1 while promoting the expression of Nrf2, a positive regulator of antioxidant genes, and its downstream modulator, HO-1. CONCLUSION: This study identified DHM as a promising therapeutic target against MTX-induced pneumonitis via activation of Nrf2 antioxidant signaling while suppressing the NF-κB mediated inflammatory pathways.

Role of EGF/ERK1/2/HO-1 axis in mediating methotrexate induced testicular damage in rats and the ameliorative effect of xanthine oxidase inhibitors.

Objectives: Methotrexate (MTX) is commonly used in the management of several malignancies and autoimmune disorders; however, testicular damage is one of the most detrimental effects of MTX administration. The current the protective effect of xanthine oxidase inhibitors either purine analogue; allopurinol (ALL) or non-purine analogue; febuxostat (FEB) in testicular injury induced by MTX in rats.Materials and methods: Thirty-two rats were randomly allocated to four groups; control (received vehicles), MTX (received single dose, 20 mg/kg, i.p.), MTX + ALL (received MTX plus ALL) and MTX + FEB (received MTX plus ALL). ALL and FEB were administered orally at 100- and 10 mg/kg, respectively for 15 days. Total and free testosterone were measured in serum. In addition, total antioxidant capacity (TAC), epidermal growth factor (EGF), malondialdehyde (MDA), tumor necrosis factor-α (TNF-α), extracellular signal-regulating kinase1/2 (ERK1/2), and total nitrite/nitrate (NOx) end products were measured in testicular tissues. At the same time, immunoexpression of HO-1in testicular tissue was measured. Histopathological examination was done.Results: ALL and FEB increased total and free serum testosterone. Both drugs showed a significant reduction in testicular MDA, NOx, TNF-α levels with an increase in TAC, EGF, and ERK1/2 levels in testicular tissue. Furthermore, both drugs enhanced HO-1 immunoexpression in testicular tissue. All these findings were parallel to the preservation of normal testicular architecture in rats treated with ALL and FEB.Conclusion: All and FEB were equally protective against testicular damage induced by MTX through anti-inflammatory, anti-apoptotic, and antioxidant actions. Their effects might be through activation of the EGF/ERK1/2/HO-1 pathway.

Dihydromyricetin alleviates methotrexate-induced hepatotoxicity via suppressing the TLR4/NF-κB pathway and NLRP3 inflammasome/caspase 1 axis.

The anticancer drug methotrexate (MTX) is known to cause hepatotoxicity as a possibly fatal adverse effect that hinders its clinical application. Although the natural flavonoid, dihydromyricetin (DHM), has antioxidant and anti-inflammatory effects; its role against MTX-induced hepatotoxicity has not been explored yet. For this, rats were administrated DHM orally for two weeks at a dose of 300 mg/kg per day, with or without a single i.p. injection of 40 mg/kg MTX on the 9th day of the experiment. MTX caused deterioration in liver structure and function, depicted by an increase in liver enzymes; ALT and AST. Moreover, MTX induced oxidative stress, shown by increasing malondialdehyde and decreasing reduced glutathione and total antioxidant capacity, initiated the inflammatory response via upregulated expression of Toll-like receptor 4 (TLR4) and its downstream transcription factor, nuclear factor-kappa B (NF-κB p65). Consequent to TLR4 signaling cascade, Nucleotide-binding oligomerization domain (NOD)-like receptor family pyrin domain containing 3 (NLRP3) inflammosome was activated and caused caspase 1 mediated transformation of proinflammatory cytokines interleukin 1ß (IL-1ß) and interleukin 18 (IL-18) into their active forms. Interestingly, administering DHM with MTX improved liver structure and function, as well as significantly decreased all oxidative stress and inflammatory signaling. Collectively, DHM possesses antioxidant and anti-inflammatory properties that can ameliorate MTX-induced hepatotoxicity, through down-regulation of liver TLR4/NF-κB and therefore prohibit activation of NLRP3/caspase 1 pathway.

Dihydromyricetin protects against high glucose-induced endothelial dysfunction: Role of HIF-1α/ROR2/NF-κB.

OBJECTIVES: Dihydromyricetin (DHM), a natural flavonoid isolated from vine tea with anti-inflammatory activity was evaluated for its ability to prevent vascular endothelial dysfunction caused by hyperglycaemia. METHODS: Vasoconstrictor (phenylephrine-PE) and vasodilator (acetylcholine-ACh) responses were monitored for female rat aorta rings maintained in a bioassay organ bath for 3 h at 37 °C in either low (LG: 10 mM) or high (HG: 40 mM, to mimic hyperglycaemia) glucose-Krebs buffer in the absence or presence of 50 µM DHM. Tissues recovered from the organ bath at 3 h were fixed and analyzed for morphological changes and their expression of eNOS, iNOS, HIF-1α, GLUT1, ROR2 tyrosine kinase, NF-κB, TNF-α, Bax, Bcl2, caspase-3, and forindices of increased oxidative stress. KEY FINDINGS: HG-incubated tissues showed increased PE-stimulated contractile response and decreased ACh-mediated endothelial vasodilation. DHM prevented both of these changes. Besides, HG incubation increased the immunoreactivity to iNOS, HIF-1α, GLUT1, ROR2, NF-κB, TNF-α, Bax, and active caspase-3, and decreased the expression of eNOS and Bcl2. Hyperglycaemia-like conditions also increased the indices of oxidative/nitrosative stress. These HG-induced changes, which were accompanied by an increase in tissue adventitial thickness and inflammatory cell infiltration, were all prevented by DHM. CONCLUSION: Our data demonstrate an anti-inflammatory protective action of DHM to preserve vascular function in the setting of hyperglycaemia.

Angiotensin aldosterone inhibitors improve survival and ameliorate kidney injury induced by sepsis through suppression of inflammation and apoptosis.

Sepsis is an extensive life-threatening illness that occurs due to an abnormal host response that extends through the initial storm of inflammation and oxidative stress and terminates at the late stage of immunosuppression. Among global intensive care units, sepsis-induced acute kidney injury is reported with high mortality rate. The purpose of this study was to evaluate the protective effect of the renin angiotensin aldosterone system (RAAS) inhibition on sepsis outcomes. Cecal ligation and puncture (CLP) procedure was applied for sepsis induction. The experimental design constituted of five groups of rats: sham, CLP-nontreated and CLP-treated with ramipril (10 mg/kg, p.o.), losartan (20 mg/kg, i.p.) and spironolactone (25 mg/kg, p.o.). Twenty-four hours after surgery, rats were euthanized for blood and tissue samples, which were used for assessment of serum inflammatory markers, and oxidative stress parameters, as well as to kidney function parameters. The tissue samples were used for histological and caspase-3 assessment. A survival study was conducted using another set of animals. Our results showed that the different RAAS inhibitors showed protective effects evidenced by enhanced overall survival following sepsis (80% in ramipril and spironolactone-treated and 60% in losartan-treated vs. 10% in the septic group), in addition to improved renal function parameters and reduction of oxidative stress and inflammation. The timely use of RAAS inhibitors during sepsis might represent a new therapeutic approach in septic patient.

Antifibrotic effects of bezafibrate and pioglitazone against thioacetamide-induced liver fibrosis in albino rats.

Activation of hepatic stellate cells is a central event in hepatic fibrogenesis that offers multiple potential sites for therapeutic interventions. Peroxisome proliferator-activated receptors are implicated in liver fibrosis. We aimed to evaluate the effect of bezafibrate and pioglitazone on a thioacetamide (TAA) rat model of liver fibrosis and to clarify the possible underlying mechanisms. Rats received intraperitoneal injections of TAA for 6 weeks. Daily oral treatments with bezafibrate or pioglitazone were started with the first day of TAA intoxication. Serum liver function tests, hepatic malondialdehyde (MDA), total nitrite and nitrate (NOx), superoxide dismutase, and hepatic histopathology were assessed to evaluate hepatic damage. Alpha smooth muscle actin (α-SMA) and tissue inhibitor metalloproteinase-1 (TIMP-1) and caspase-3 were also assessed. The TAA group experienced significant deterioration of liver functions, increased oxidative stress, and increased liver tissue NOx. Administration of bezafibrate or pioglitazone resulted in significant improvement of all liver functions and reduced oxidative stress in hepatic tissues. Only administration of bezafibrate significantly reduced NOx levels. Liver tissues from the TAA-treated group showed disrupted normal architecture. Administration of bezafibrate or pioglitazone attenuated this picture. Stronger α-SMA expression was detected in the TAA group. Treatment with bezafibrate or pioglitazone decreased the α-SMA expression.

Toxicological Assessment of Venlafaxine: Acute and Subchronic Toxicity Study in Rats.

Antidepressants are the most commonly prescribed drugs for psychiatric treatment, and venlafaxine (VEN) is one of the most popular options. Venlafaxine is a nontricyclic dual-acting serotonin-norepinephrine reuptake inhibitor. Although an increased incidence of acute toxicity and addiction has been reported, controlled studies examining its toxic effects on different organs are still lacking. This study investigated the possible toxic effects of VEN on the liver, kidney, and gastric tissues. Three groups of rats were administered saline, a single LD50 dose (350 mg/kg), or 100 mg/kg VEN daily, followed by increases in the dose of 50 mg/kg every 10 days for 30 days (about 10 times the therapeutic doses). The following parameters of liver and kidney injury were then assayed: alanine aminotransferase, aspartate aminotransferase, γ-glutamyl transferase, prothrombin time, partial thromboplastin time, blood urea nitrogen, and serum creatinine. A histopathological examination was then conducted. Both acute and subchronic administration of VEN produced multiple clinical manifestations in the experimental animals, including seizures, coma, and even death. Moreover, the liver and renal function tests indicated injury in these tissues. Furthermore, the histopathological examination showed signs of organ toxicity after both acute and chronic VEN exposure. This study has shown that VEN has harmful effects on the liver, kidney, and stomach in either a single high dose (LD50) or repeated exposure to 10 times the therapeutic doses. As a result, strategies to increase awareness of these effects among physicians and the public are needed because this drug may be addictive.

Effect of histone deacetylase inhibitor on epithelial-mesenchymal transition of liver fibrosis.

Liver fibrosis is an excessively reversible wound healing process and the fibrotic disorder is the activation of hepatic stellate cell that requires extensive alterations in gene expression. As reversible deacetylation of histone proteins modulate gene expression, we examined the effect of valproic acid (VPA) as selective histone deacetylase inhibitor on CCl-4 induced liver fibrosis. Thirty rats were divided into three equal groups; control group, fibrotic group and VPA-treated group. The rats were sacrificed after 6 weeks of liver fibrosis induction. The histopathological effect on liver tissue was examined. The expression of α-SMA and Smad-4 mRNA and serum levels of TGF-ß1, alanine aminotransferase, and aspartate aminotransferase were determined. Treatment of rats with VPA attenuated carbon tetrachloride-induced liver fibrosis. Moreover, α-SMA and Smad-4 expression was repressed under VPA treatment and both serum TGF-ß1 and liver enzymes were significantly decreased. The histone deacetylase inhibitor-1 VPA inhibits the epithelial-mesenchymal transition and affects hepatic stellate cell activation during liver fibrosis through downregulation of Smad4 and α-SMA expression which may serve as a promising agent in liver fibrosis treatment. © 2018 IUBMB Life, 70(6):511-518, 2018.

Protective effect of C-peptide on experimentally induced diabetic nephropathy and the possible link between C-peptide and nitric oxide.

Diabetic nephropathy one of the major microvascular diabetic complications. Besides hyperglycemia, other factors contribute to the development of diabetic complications as the proinsulin connecting peptide, C-peptide. We described the role of C-peptide replacement therapy on experimentally induced diabetic nephropathy, and its potential mechanisms of action by studying the role of nitric oxide (NO) as a mediator of C-peptide effects by in vivo modulating its production by NG-nitro-l-arginine methyl ester (L-NAME). Renal injury markers measured were serum urea, creatinine, tumor necrosis factor alpha, and angiotensin II, and malondialdehyde, total antioxidant, Bcl-2, and NO in renal tissue. In conclusion, diabetic induction resulted in islet degenerations and decreased insulin secretion with its metabolic consequences and subsequent renal complications. C-Peptide deficiencies in diabetes might have contributed to the metabolic and renal error, since C-peptide treatment to the diabetic rats completely corrected these errors. The beneficial effects of C-peptide are partially antagonized by L-NAME coadministration, indicating that NO partially mediates C-peptide effects.

Tramadol Induced Adrenal Insufficiency: Histological, Immunohistochemical, Ultrastructural, and Biochemical Genetic Experimental Study.

Tramadol is a synthetic, centrally acting analgesic. It is the most consumed narcotic drug that is prescribed in the world. Tramadol abuse has dramatically increased in Egypt. Long term use of tramadol can induce endocrinopathy. So, the aim of this study was to analyze the adrenal insufficiency induced by long term use of tramadol in experimental animals and also to assess its withdrawal effects through histopathological and biochemical genetic study. Forty male albino rats were used in this study. The rats were divided into 4 groups (control group, tramadol-treated group, and withdrawal groups). Tramadol was given to albino rats at a dose of 80 mg/kg body weight for 3 months and after withdrawal periods (7-15 days) rats were sacrificed. Long term use of tramadol induced severe histopathological changes in adrenal glands. Tramadol decreased the levels of serum cortisol and DHEAS hormones. In addition, it increased the level of adrenal MDA and decreased the genetic expression of glutathione peroxidase and thioredoxin reductase in adrenal gland tissues. All these changes started to return to normal after withdrawal of tramadol. Thus, it was confirmed that long term use of tramadol can induce severe adrenal insufficiency.

Possible Protective Effect of Diacerein on Doxorubicin-Induced Nephrotoxicity in Rats.

Nephrotoxicity is one of the limiting factors for using doxorubicin (DOX). Interleukin 1 has major role in DOX-induced nephrotoxicity, so we investigated the effect of interleukin 1 receptor antagonist diacerein (DIA) on DOX-induced nephrotoxicity. DIA (25 and 50 mg/kg/day) was administered orally to rats for 15 days, in the presence or absence of nephrotoxicity induced by a single intraperitoneal injection of DOX (15 mg/kg) at the 11th day. We measured levels of serum urea, creatinine, renal reduced glutathione (GSH), malondialdehyde (MDA), total nitrites (NO x ), catalase, and superoxide dismutase (SOD). In addition, caspase-3, tumor necrosis factor alpha (TNFα), nuclear factor kappa B (NFκB) expressions, and renal histopathology were assessed. Our results showed that DOX-induced nephrotoxicity was ameliorated or reduced by both doses of DIA, but diacerein high dose (DHD) showed more improvement than diacerein low dose (DLD). This protective effect was manifested by significant improvement in all measured parameters compared to DOX treated group by using DHD. DLD showed significant improvement of creatinine, MDA, NO x , GSH, histopathology, and immunohistochemical parameters compared to DOX treated group.