Evaluation of miRNAs regulation of BDNF and IGF1 genes in T2DM insulin resistance in experimental models: bioinformatics based approach / Avaliação da regulação por miRNAs dos genes BDNF e IGF1 na resistência a insulina no diabetes do tipo 2 em modelos experimentais: in silico

microRNAs (miRNAs) are recognized as diabetes mellitus type 2 (T2DM) biomarkers useful for disease metabolism comprehension and have great potential as therapeutics targets. BDNF and IGF1 increased expression are highly involved in the benefits of insulin and glucose paths, however, they are down-regulated in insulin resistance conditions, while their expression increase is correlated to the improvement of glucose and insulin metabolism. Studies suggest the microRNA regulation of these genes in several different contexts, providing a novel investigation approach for comprehending T2DM metabolism and revealing potential therapeutic targets. In the present study, we investigate in different animal models (human, rat, and mouse) miRNAs that target BDNF and IGF1 in skeletal muscle tissue with T2DM physiological conditions. Bioinformatics tools and databases were used to miRNA prediction, molecular homology, experimental validation of interactions, expression in the studied physiological condition, and network interaction. The findings showed three miRNAs candidates for IGF1(miR-29a, miR-29b, and miR-29c) and one for BDNF (miR-206). The experimental evaluations and the search for the expression in skeletal muscle from T2DM subjects confirmed the predicted interaction between miRNA-mRNA for miR-29b and miR-206 through human, rat, and mouse models. This interaction was reaffirmed in multiple network analyses. In conclusion, our results show the regulation relationship between miR-29b and miR-206 with the investigated genes, in several tissues, suggesting an inhibition pattern. Nevertheless, these data show a large number of possible interaction physiological processes, for future biotechnological prospects.

Os microRNAs (miRNAs) são reconhecidos como biomarcadores do diabetes mellitus tipo 2 (DM2), úteis para a compreensão do metabolismo da doença, e possuem grande potencial como alvos terapêuticos. O aumento da expressão de BDNF e IGF1 está altamente envolvido nos benefícios as vias de insulina e glicose, porém, são regulados negativamente em condições de resistência à insulina, enquanto seu aumento de expressão está correlacionado com a melhora do metabolismo da glicose e da insulina. Estudos sugerem a regulação desses genes por microRNA em vários contextos diferentes, proporcionando uma nova abordagem de investigação para compreender o metabolismo do DM2 e revelar potenciais alvos terapêuticos. No presente estudo, investigamos em diferentes modelos animais (humanos, ratos e camundongos) miRNAs que têm como alvo BDNF e IGF1 em tecido muscular esquelético com condições fisiológicas de DM2. As análises foram realizadas utilizando ferramentas de bioinformática e bancos de dados para predição de miRNA, homologia molecular, validação experimental de interações, expressão na condição fisiológica estudada e interação em rede. Os resultados mostraram três candidatos a miRNAs para IGF1 (miR-29a, miR-29b e miR-29c) e um para BDNF (miR-206). As avaliações experimentais e a busca pela expressão no músculo esquelético de indivíduos com DM2 confirmaram a interação prevista entre miRNA-mRNA para miR-29b e miR-206 através de modelos humanos, ratos e camundongos. Essa interação foi reafirmada em múltiplas análises de rede. Em conclusão, nossos resultados mostram a relação de regulação entre miR-29b e miR-206 com os genes investigados, em diversos tecidos, sugerindo um padrão de inibição. Contudo, esses dados mostram um grande número de possíveis processos fisiológicos de interação para perspectivas biotecnológicas.

Evaluation of miRNAs regulation of BDNF and IGF1 genes in T2DM insulin resistance in experimental models: bioinformatics based approach.

microRNAs (miRNAs) are recognized as diabetes mellitus type 2 (T2DM) biomarkers useful for disease metabolism comprehension and have great potential as therapeutics targets. BDNF and IGF1 increased expression are highly involved in the benefits of insulin and glucose paths, however, they are down-regulated in insulin resistance conditions, while their expression increase is correlated to the improvement of glucose and insulin metabolism. Studies suggest the microRNA regulation of these genes in several different contexts, providing a novel investigation approach for comprehending T2DM metabolism and revealing potential therapeutic targets. In the present study, we investigate in different animal models (human, rat, and mouse) miRNAs that target BDNF and IGF1 in skeletal muscle tissue with T2DM physiological conditions. Bioinformatics tools and databases were used to miRNA prediction, molecular homology, experimental validation of interactions, expression in the studied physiological condition, and network interaction. The findings showed three miRNAs candidates for IGF1(miR-29a, miR-29b, and miR-29c) and one for BDNF (miR-206). The experimental evaluations and the search for the expression in skeletal muscle from T2DM subjects confirmed the predicted interaction between miRNA-mRNA for miR-29b and miR-206 through human, rat, and mouse models. This interaction was reaffirmed in multiple network analyses. In conclusion, our results show the regulation relationship between miR-29b and miR-206 with the investigated genes, in several tissues, suggesting an inhibition pattern. Nevertheless, these data show a large number of possible interaction physiological processes, for future biotechnological prospects.

Evaluation of Hypoglycemic Therapy Through Physical Exercise in n5STZ-Induced Diabetes Rats.

BACKGROUND: Diabetes mellitus is a syndrome with multiple etiologies involving insulin, in which there is a lack of production and/or loss of sensitivity to this hormone resulting in insulin resistance. Treatment and control of this disease requires changes in diet, use of medication, and lifestyle, such as physical activity. These modifications may compromise quality-of-life if there is no proper guidance for the treatment or alert to possible complications caused by the disease. METHODS: This study aimed to evaluate biochemical and hematological parameters, and to assess brain derived neurotrophic factor levels in diabetic Wistar rats submitted to chronic physical exercise. RESULTS: The results demonstrated an increase in plasma concentration of brain-derived neurotrophic factor (BDNF) in association with hyperglycemia reduction in diabetic animals. DISCUSSION: The results obtained suggest that there is a regulation of glucose homeostasis between peripheral tissues and the central nervous system. Exercise-induced BDNF also improved levels of glycemia, body weight, and dyslipidemia. In hematological evaluation, BDNF increase was positively correlated with an improvement in leukocyte parameters. Electrophoresis analyses demonstrated a reduction in levels of pro-inflammatory proteins, lipoprotein fractions, and albumin preservation in diabetic animals trained with elevated concentration of plasma BDNF. CONCLUSION: In conclusion, this study demonstrated that chronic exercise was able to elevate BDNF levels in plasma, which resulted directly in positive hypoglycemic activity in diabetic animals and a reduction of the metabolic syndrome associated with diabetes mellitus.

Two Kunitz-type inhibitors with activity against trypsin and papain from Pithecellobium dumosum seeds: purification, characterization, and activity towards pest insect digestive enzyme.

Two trypsin inhibitors (called PdKI-3.1 and PdKI-3.2) were purified from the seeds of the Pithecellobium dumosum tree. Inhibitors were obtained by TCA precipitation, affinity chromatography on Trypsin-Sepharose and reversed-phase-HPLC. SDS-PAGE analysis with or without reducing agent showed that they are a single polypeptide chain, and MALDI-TOF analysis determined molecular masses of 19696.96 and 19696.36 Da, respectively. The N-terminal sequence of both inhibitors showed strong identity to the Kunitz family trypsin inhibitors. They were stable over a wide pH (2-9) and temperature (37 to 100 degrees C) range. These inhibitors reduced over 84% of trypsin activity with inhibition constant (Ki) of 4.20 x 10(-8) and 2.88 x 10(-8) M, and also moderately inhibited papain activity, a cysteine proteinase. PdKI-3.1 and PdKI-3.2 mainly inhibited digestive enzymes from Plodia interpunctella, Zabrotes subfasciatus and Ceratitis capitata guts. Results show that both inhibitors are members of the Kunitz-inhibitor family and that they affect the digestive enzyme larvae of diverse orders, indicating a potential insect antifeedant.

Identification of a Kunitz-type proteinase inhibitor from Pithecellobium dumosum seeds with insecticidal properties and double activity.

A trypsin inhibitor, PdKI, was purified from Pithecellobium dumosum seeds by TCA precipitation, trypsin-sepharose chromatography, and reversed-phase-HPLC. PdKI was purified 217.6-fold and recovered 4.7%. SDS-PAGE showed that PdKI is a single polypeptide chain of 18.9 kDa and 19.7 kDa by MALDI-TOF. The inhibition on trypsin was stable in the pH range 2-10 and at a temperature of 50 degrees C. The Ki values were 3.56 x 10(-8)and 7.61 x 10(-7) M with competitive and noncompetitive inhibition mechanisms for trypsin and papain, respectively. The N-terminal sequence identified with members of Kunitz-type inhibitors from the Mimosoideae and Caesalpinoideae subfamilies. PdKI was effective against digestive proteinase from Zabrotes subfasciatus, Ceratitis capitata, Plodia interpunctella, Alabama argillaceae, and Callosobruchus maculatus, with 69, 66, 44, 38, and 29% inhibition, respectively. Results support that PdKI is a member of the Kunitz inhibitor family and its insecticidal properties indicate a potent insect antifeedant.