The Role of Metformin in Controlling Oxidative Stress in Muscle of Diabetic Rats

Metformin can act in muscle, inhibiting the complex I of the electron transport chain and decreasing mitochondrial reactive oxygen species. Our hypothesis is that the inhibition of complex I can minimize damage oxidative in muscles of hypoinsulinemic rats. The present study investigated the effects of insulin and/or metformin treatment on oxidative stress levels in the gastrocnemius muscle of diabetic rats. Rats were rendered diabetic (D) with an injection of streptozotocin and were submitted to treatment with insulin (D+I), metformin (D+M), or insulin plus metformin (D+I+M) for 7 days. The body weight, glycemic control, and insulin resistance were evaluated. Then, oxidative stress levels, glutathione antioxidant defense system, and antioxidant status were analyzed in the gastrocnemius muscle of hypoinsulinemic rats. The body weight decreased in D+M compared to ND rats. D+I and D+I+M rats decreased the glycemia and D+I+M rats increased the insulin sensitivity compared to D rats. D+I+M reduced the oxidative stress levels and the activity of catalase and superoxide dismutase in skeletal muscle when compared to D+I rats. In conclusion, our results reveal that dual therapy with metformin and insulin promotes more benefits to oxidative stress control in muscle of hypoinsulinemic rats than insulinotherapy alone

Identification of salivary and urinary biomarkers for the development of sustainable platforms for the diagnosis and monitoring of diabetes mellitus

The diagnosis and monitoring of diabetes mellitus (DM) by blood is highly accurate, however it is an invasive procedure, it causes discomfort to patients and has a high cost for both the health system and patients. The development of alternatives non-invasive platforms with other biological samples, which are sustainable, with high sensitivity and reduced cost, can collaborate to diabetes surveillance. In this thesis, potential salivary and urinary biomarkers were investigated as alternatives for the diagnosis and monitoring of DM divided in 3 studies. Firstly, an analysis was performed with the Fourie transform infrared spectroscopy (FTIR) tool to evaluate the urinary components of non-diabetic, diabetic and diabetic rats treated with insulin. In another study, the FTIR platform was used in association with the classification of the support vector machine (SVM) for discriminate non-diabetic subjects and type 2 diabetic patients using saliva. Besides that, a systematic review and meta-analysis was carried out to assess the correlation between the salivary alpha-2-macroglobulin (A2MG) protein with glycemia and glycated hemoglobin (HbA1c) to systematically review the effectiveness of this protein to determine the levels of glycemia and HbA1c in type 2 diabetic patients. Briefly, Here we outline the potential of the FTIR platform as an alternative tool without reagent, non-invasive and highly sensitive for screening and monitoring diabetic patients using a reduced volume and minimal sample preparation (saliva or urine). Besides, the data show a strong correlation between HbA1C and salivary A2MG that should be proved in large cohort of patients to monitoring diabetes using saliva.FAPEMIG - Fundação de Amparo a Pesquisa do Estado de Minas GeraisTese (Doutorado)O diagnóstico e o monitoramento do diabetes mellitus (DM) pelo sangue é altamente preciso, entretanto é um procedimento invasivo, gera desconforto aos pacientes e possui um elevado custo tanto para o sistema de saúde como para os pacientes. O desenvolvimento de plataformas não invasivas alternativas com outras amostras biológicas de forma sustentável, com alta sensibilidade e custo reduzido podem colaborar com a vigilância de DM. Nesta tese, foram investigados possíveis biomarcadores salivares e urinários como potenciais alternativas para o diagnóstico e monitoramento do DM divididos em três estudos. Inicialmente, foi realizada análise com a ferramenta de espectroscopia de infravermelho com transformada de Fourier (FTIR) para avaliar os componentes urinários de ratos não diabéticos, diabéticos e diabéticos tratados com insulina. Em outro estudo, uma plataforma de FTIR foi utilizada em associação com a classificação de máquina de vetor de suporte para discriminação de sujeitos não-diabéticos e pacientes com DM tipo 2 por meio da análise de saliva. Além disso, foi realizada uma revisão sistemática e metanálise que avaliou a correlação entre a proteína alfa-2-macroglobulina (A2MG) salivar com a glicemia e hemoglobina glicada (HbA1c) para revisar sistematicamente a eficácia desta proteína para determinar os níveis de glicemia e HbA1c de pacientes com DM tipo 2. Em resumo, podemos concluir que a plataforma FTIR apresenta grande potencial para diagnóstico e monitoramento de pacientes diabéticos de forma não invasiva, sem utilização de reagente, e altamente sensível utilizando um baixo volume com preparação mínima de amostra (saliva ou urina). Além disto, os dados apresentam uma significativa correlação entre HbA1C e A2MG salivar que necessitam ser provados em estudos com grandes coortes para vigilância do diabetes pela saliva.2022-11-1

Salivary diagnosis of diabetes mellitus using ATR-FTIR-spectroscopy

Introduction: Monitoring of blood glucose is an invasive, painful and costly procedure in diabetes. Consequently, the search for more a cost-effective (reagent-free), non-invasive and specific diabetes diagnostic method is of great interest. Objective: This research investigated the application of ATR-FTIR spectroscopy as an alternative for the diagnosis of DM by quantitative salivary spectrum analysis. Material and methods: Wistar rats were divided in non-diabetic (ND), diabetic (D) and diabetic 6U-treated of insulin (D6U). DM was induced by an intraperitoneal injection (60 mg/kg) of streptozotocin (STZ). The animals were submitted to 28 days of diabetes, and on the 21st day, the treatment was started for 7 days with insulin or vehicle solution according to the group. Body weight, food and water intake, glycemia, urinary volume and urine concentration of urine were evaluated during the experiment. The salivary profile was analyzed by ATR-FTIR spectroscopy and the vibrational modes were evaluated for diagnostic ability by ROC curve. Results: Thirteen vibrational modes of saliva spectra of ND, D and D6U were identified, and of these, four vibrational modes were pre-validated as potential biomarkers for diagnosis by the ROC curve, with a significant correlation with glycemia. Compared to the ND, 1377 cm-1, 1255 cm-1, 628 cm-1 and 616 cm-1 bands of D rats gave a sensitivity and specificity of 100% (p<0.001). In addition, the spectral biomarkers 1255 cm-1 and 628 cm-1 demonstrated a high correlation with glycemia (R2 of 0.84 and 0.8595, respectively). Conclusion: Altogether, 1255 cm-1 and 628 cm-1 spectral salivary biomarkers may provide a novel robust alternative for diabetes diagnostics.CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível SuperiorCNPq - Conselho Nacional de Desenvolvimento Científico e TecnológicoFAPEMIG - Fundação de Amparo a Pesquisa do Estado de Minas GeraisDissertação (Mestrado)Introdução: Atualmente o diagnóstico do diabetes é realizado por procedimento invasivo, dolorido e de custo alto. Consequentemente a busca por um método diagnóstico mais barato (sem utilização de reagentes), não-invasivo e específico ao diabetes é de grande interesse. Objetivo: Esta pesquisa investigou a aplicação da espectroscopia de reflexão total atenuada no infravermelho com transformada de Fourier (ATR-FTIR) em amostras de saliva como alternativa para o diagnóstico de DM. Materiais e métodos: Ratos Wistar foram divididos em: não-diabético (ND), diabético (D) e diabético tratado com 6U de insulina (D6U). O DM foi induzido por uma injeção intraperitoneal (60 mg / kg) de estreptozotocina (STZ). Vinte e um dias após a indução do diabetes foi iniciado o tratamento, que ocorreu durante 7 dias, com insulina ou veículo. O peso corporal, a ingestão alimentar e hídrica, glicemia, volume urinário e concentração de glicose na urina foram avaliados durante o experimento. O perfil salivar foi analisado por espectroscopia ATR-FTIR e os modos vibracionais foram avaliados quanto à capacidade diagnóstica pela curva ROC. Resultados: Foram identificados treze modos vibracionais dos espectros da saliva de animais ND, D e D6U, e destes, 4 modos vibracionais foram pré-validados como potenciais biomarcadores para diagnóstico pela curva ROC e significativa correlação com a glicemia. Em comparação com o espectro ND, os modos vibracionais 1377 cm-1, 1255 cm-1, 628 cm-1 e 616 cm-1 de animais D apresentaram uma sensibilidade e especificidade de 100% (p <0.001). Além disso, os biomarcadores espectrais 1255 cm-1 e 628 cm-1 demonstraram uma alta correlação com a glicemia (r de 0,84 e 0,8595, respectivamente). Conclusão: Dessa forma, os espectros salivares 1255 cm-1 e 628 cm-1 podem ser utilizados como uma nova alternativa para o diagnóstico de diabetes

Monitoring glucose levels in urine using FTIR spectroscopy combined with univariate and multivariate statistical methods.

The development of novel platforms for non-invasive continuous glucose monitoring applied in the screening and monitoring of diabetes is crucial to improve diabetes surveillance systems. Attenuated total reflection-Fourier transform infrared (ATR-FTIR) spectroscopy of urine can be an alternative as a sustainable, label-free, fast, non-invasive, and highly sensitive analysis to detect changes in urine promoted by diabetes and insulin treatment. In this study, we used ATR-FTIR to evaluate the urinary components of non-diabetic (ND), diabetic (D), and diabetic insulin-treated (D + I) rats. As expected, insulin treatment was capable to revert changes in glycemia, 24-h urine collection volume, urine creatinine, urea, and glucose excretion promoted by diabetes. Several differences in the urine spectra of ND, D, and D + I were observed, with urea, creatinine, and glucose analytes being related to these changes. Principal components analysis (PCA) scores plots allowed for the discrimination of ND and D + I from D with an accuracy of ∼ 99 %. The PCA loadings associated with PC1 confirmed the importance of urea and glucose vibrational modes for this discrimination. Univariate analysis of second derivative spectra showed a high correlation (r: 0.865, p < 0.0001) between the height of 1074 cm-1 vibrational mode with urinary glucose concentration. In order to estimate the amount of glucose present in the infrared spectra from urine, multivariate curve resolution-alternating least square (MCR-ALS) was applied and a higher predicted concentration of glucose in the urine was observed with a correlation of 78.9 % compared to urinary glucose concentration assessed using enzyme assays. In summary, ATR-FTIR combined with univariate and multivariate chemometric analyses provides an innovative, non-invasive, and sustainable approach to diabetes surveillance

Adaptogenic potential of royal jelly in liver of rats exposed to chronic stress.

Restraint and cold stress increase both corticosterone and glycemia, which lead to oxidative damages in hepatic tissue. This study assessed the effect of royal jelly (RJ) supplementation on the corticosterone level, glycemia, plasma enzymes and hepatic antioxidant system in restraint and cold stressed rats. Wistar rats were allocated into no-stress, stress, no-stress supplemented with RJ and stress supplemented with RJ groups. Initially, RJ (200mg/Kg) was administered for fourteen days and stressed groups were submitted to chronic stress from the seventh day. The results showed that RJ supplementation decreases corticosterone levels and improves glycemia control after stress induction. RJ supplementation also decreased the body weight, AST, ALP and GGT. Moreover, RJ improved total antioxidant capacity, SOD activity and reduced GSH, GR and lipoperoxidation in the liver. Thus, RJ supplementation reestablished the corticosterone levels and the hepatic antioxidant system in stressed rats, indicating an adaptogenic and hepatoprotective potential of RJ

Salivary SARS-CoV-2 RNA for diagnosis of COVID-19 patients: A systematic review and meta-analysis of diagnostic accuracy

Accurate, self-collected, and non-invasive diagnostics are critical to perform mass-screening diagnostic tests for COVID-19. This systematic review with meta-analysis evaluated the accuracy, sensitivity, and specificity of salivary diagnostics for COVID-19 based on SARS-CoV-2 RNA compared with the current reference tests using a nasopharyngeal swab (NPS) and/or oropharyngeal swab (OPS). An electronic search was performed in seven databases to find COVID-19 diagnostic studies simultaneously using saliva and NPS/OPS tests to detect SARS-CoV-2 by RT-PCR. The search resulted in 10,902 records, of which 44 studies were considered eligible. The total sample consisted of 14,043 participants from 21 countries. The accuracy, specificity, and sensitivity for saliva compared with the NPS/OPS was 94.3 % (95 %CI = 92.1;95.9), 96.4 % (95 %CI = 96.1;96.7), and 89.2 % (95 %CI = 85.5;92.0), respectively. Besides, the sensitivity of NPS/OPS was 90.3 % (95 %CI = 86.4;93.2) and saliva was 86.4 % (95 %CI = 82.1;89.8) compared to the combination of saliva and NPS/OPS as the gold standard. Based on low to moderate certainty level these findings suggest a similarity in SARS-CoV-2 RNA detection between NPS/OPS swabs and saliva, and the association of both testing approaches as a reference standard can increase by 3.6 % the SARS-CoV-2 detection compared with NPS/OPS alone. This study supports saliva as an attractive alternative for diagnostic platforms to provide a non-invasive detection of SARS-CoV-2

Salivary Detection of Zika Virus Infection Using ATR-FTIR Spectroscopy Coupled with Machine Learning Algorithms and Univariate Analysis: A Proof-of-Concept Animal Study

Zika virus (ZIKV) diagnosis is currently performed through an invasive, painful, and costly procedure using molecular biology. Consequently, the search for a non-invasive, more cost-effective, reagent-free, and sustainable method for ZIKV diagnosis is of great relevance. It is critical to prepare a global strategy for the next ZIKV outbreak given its devastating consequences, particularly in pregnant women. Attenuated total reflection–Fourier transform infrared (ATR-FTIR) spectroscopy has been used to discriminate systemic diseases using saliva; however, the salivary diagnostic application in viral diseases is unknown. To test this hypothesis, we intradermally challenged interferon-gamma gene knockout C57/BL6 mice with ZIKV (50 µL,105 FFU, n = 7) or vehicle (50 µL, n = 8). Saliva samples were collected on day three (due to the peak of viremia) and the spleen was also harvested. Changes in the salivary spectral profile were analyzed by Student’s t test (p −1 as a potential candidate to discriminate ZIKV and control salivary samples. Three PCs explained 93.2% of the cumulative variance in PCA analysis and the spectrochemical analysis with LDA achieved an accuracy of 93.3%, with a specificity of 87.5% and sensitivity of 100%. The LDA-SVM analysis showed 100% discrimination between both classes. Our results suggest that ATR-FTIR applied to saliva might have high accuracy in ZIKV diagnosis with potential as a non-invasive and cost-effective diagnostic tool

Biomarkers of chronic stress in liver tissue of rats stressed by restraint and cold.

<p>Plasma corticosterone level after seven days of stress-induction (A). No stress (NS), No Stress supplemented with Royal Jelly (NSRJ), Stress (S) and Stress supplemented with Royal Jelly (SRJ). Blood glucose level before and after the last stress induction (B). No stress (NS), No Stress supplemented with Royal Jelly (NSRJ), Stress group before the last stress session (Sb), Stress group after the last stress session (Sa), Stress group supplemented with RJ before the last stress session (SRJb), Stress group supplemented with RJ after the last stress session (SRJa). Pearson correlation of mean values of corticosterone levels and blood glucose after the last stress-induction (C). Values are expressed as means ± SEM. *p < 0.05 vs NS, # p < 0.05 vs S, & p < 0.05 vs Sb, § p < 0.05 vs Sa (One-way ANOVA followed by Tukey test). Outliers were detected by performing Grubb’s test using an online GraphPad outlier calculator (<a href="http://graphpad.com/quickcalcs/Grubbs1.cfm" target="_blank">http://graphpad.com/quickcalcs/Grubbs1.cfm</a>).</p

The effect of the stress-induction by restraint and cold and supplementation of royal jelly on body weight, water intake, food intake and hepatic enzymes activities in plasma.

<p>The effect of the stress-induction by restraint and cold and supplementation of royal jelly on body weight, water intake, food intake and hepatic enzymes activities in plasma.</p

Superoxide dismutase activity in liver tissue of rats stressed by immobilization and cold.

<p>No stress (NS), No Stress supplemented with Royal Jelly (NSRJ), Stress (S) and Stress supplemented with Royal Jelly (SRJ). Values are expressed as mean±SEM, * p < 0.05 vs. NS, # p < 0.05 vs. S (One-way ANOVA followed by Tukey test). Outliers were detected by performing Grubb’s test using an online GraphPad outlier calculator (<a href="http://graphpad.com/quickcalcs/Grubbs1.cfm" target="_blank">http://graphpad.com/quickcalcs/Grubbs1.cfm</a>).</p