Summary:
Purpose: Methotrexate (MTX) is an antimetabolite widely used in therapy with two distinct uses. At a low dose it is given as an immunomodulatory agent for the treatment of rheumatoid arthritis (DMARD) or other autoimmune disorders, while at a high dose as an anti-cancer treatment. However, the administration of MTX at low as well as at high dose has been associated with several adverse effects, which in several cases limit its use, among which are hepatic and renal dysfunction. Because MTX has a significant contribution and effectiveness in the treatment of the aforementioned diseases, it is important to study and find substances that will be able to reduce these side effects and make MTX treatment safer. This role could possibly be played by a flavonol isolated from the plant Ampelopsis grossedentata, family Vitaceae, dihydromyricetin (DHM). In the present work, we studied the potential hepatoprotective and nephroprotective effects of DHM in MTX-treated mice, and investigated the mechanism by which these protective effects are achieved.
Methods: MTX was administered intraperitoneally (IP) to mice of three different strains (C57Bl6, BALBc, 129/Sv), in order to find the appropriate experimental model of hepatotoxicity of MTX by evaluating the liver function markers aspartate and alanine aminotransferase. Then, 129/Sv mice were randomized into 6 groups and received either MTX once or DHM orally for 5 days or the combination of MTX and DHM. At the end of the administrations, a blood sample was taken to determine indicators of liver and kidney damage and hematological toxicity in the plasma. Finally, animals were euthanized to obtain the liver and kidney, for histological evaluation and analysis of the underlying mechanisms of MTX toxicity and DHM protection.
Results: Elevated levels of transaminases, creatinine and protein carbonyls were found in the plasma of the animals, indicating possible liver damage and increased oxidative stress. Histological evaluation of the organs, however, showed no significant lesions. Regarding the underlying mechanism in the liver, MTX appears to induce damage through mechanisms of apoptosis and inflammation. DHM reduces inflammation by reducing the expression of TNF-α, p-STAT3 and p-NF-Kb, as well as iNOS. At the same time, DHM also intervenes in the apoptotic pathway, as it reverses the increased levels of the apoptotic protein BAX and the decreased levels of the anti-apoptotic protein Bcl-xL observed after administration of MTX. In the kidney, the deleterious effect of MTX is based more on inflammation, and less on oxidative stress and apoptosis. In the inflammation-related markers, DHM reduces the phosphorylation of STAT3 and NF-Kb as well as the expression of IL-6. Finally, DHM exhibits antioxidant properties, as it restores the levels of protein carbonyls that increase under the influence of MTX in both the liver and kidneys.
Conclusions: DHM has a protective effect against MTX-induced hepatotoxicity and nephrotoxicity mediated by reducing inflammation and apoptosis while also involving antioxidant effects.
Keywords:
methotrexate, dihydromyricetin, hepatotoxicity, nephrotoxicity, oxidative stress, inflammation, OATP1B1
