Investigação de alterações bioquímicas, moleculares e comportamentais relacionadas à patogênese da hipermetioninemia: estudos in vitro e in vivo

Abstract

Hypermethioninemia is an inherited metabolic disorder characterized by high levels of methionine (Met) and its metabolites such as methionine sulfoxide (MetO). The symptoms and clinical signs remain poorly understood, besides being varied and nonspecific, making it difficult to diagnose and select the most appropriate therapy. Thus, this study aimed to investigate the effect of high concentrations of Met and MetO on behavioral, cellular and molecular changes in experimental models in vitro and in vivo. In the in vitro protocol, the cytotoxic effects of exposure of cortical astrocytes to Met (1 and 2 mM), MetO (0.5 mM) and the combination of Met (1 and 2 mM) and MetO (0.5 mM) . For the in vivo model, an acute and a chronic protocol were used in which the Wistar rats were divided into four groups: I (control); II (Met 0.2-0.4 g/kg); III (MetO 0.05 - 0.1 g/kg) and IV (association of Met + MetO). After 1 h, 3 h and/or 21 days, behavioral, serum biochemical, neurochemical and thromboregulatory parameters were evaluated in brain, liver, kidney, blood and platelets. First, it was observed that chronic administration of Met and/or MetO altered serum biochemical parameters such as glucose, triglycerides, total cholesterol and urea levels, besides increased ALA-D enzyme activity and induced oxidative stress in liver and kidney. In the acute and chronic protocols Met and/or MetO reduced the activity of the enzymes NTPDase, 5'-nucleotidase and adenosine deaminase in platelets and serum. In addition, there was an increase in the production of reactive oxygen species and lipid peroxidation, reduction of the levels of non-enzymatic antioxidant compounds, reduction of superoxide dismutase activity, catalase and increase in glutathione S-transferase activity in platelets and serum of rats submitted to acute and chronic protocols. In the cerebral cortex, acute administration of Met and/or MetO induced oxidative stress, decreased cell viability, caused DNA damage and cell death by apoptosis, increased the activity of caspases 3 and 9 and reduced membrane potential. In the chronic protocol, Met and/or MetO induced impairment in spatial and short-term memory without altering the animals' locomotion. In the cerebral cortex, hippocampus and striatum Met and/or MetO induced oxidative stress and increased acetylcholinesterase (AChE) activity. Additionally, in the hippocampus there was a reduction of Na+, K+-ATPase activity, levels of neurotrophic factor derived from the brain and number of neurons in the CA3 region and dentate gyrus. In the astrocytes, it was observed that Met and/or MetO did not change cell viability and proliferation, but altered the morphology of these cells, induced oxidative stress, increased AChE activity and reduced Na+, K+- ATPase activity. In addition, in the astrocytes treated with Met and/or MetO there was a reduction in NTPDase activity using ATP as substrate and 5'-nucleotidase. On the other hand, there was an increase in NTPDase activity using ADP as substrate. The findings of the present study demonstrated that hypermethioninemia is able to alter several cellular and molecular mechanisms inducing important alterations in the homeostasis of different tissues and cells, impairing behavioral and physiological aspects. Therefore, these findings may be very important to aid in the diagnosis and future studies of possible therapeutic targets, as antioxidants, for patients with this pathology.