SCIENTIFIC REPORT RESULTS
We included in the project study a total of 517 patients newly diagnosed with type 2 diabetes (ND-T2D) of which 367 were completely regarding clinical tests (parameters determined). Of these 170 were reevaluated at 12 months after inclusion in the project. For the control group we included 130 people without diabetes. Patients reevaluated after 12 months of lifestyle improvement were investigated according to the protocol at baseline as follows:
1. Clinical Module: historical data, general clinical examination
2. Psycho-nutritional evaluation for food behavioral profiling (a) and motion behavioral profiling (b)
3. Detailed anthropometric study: BMI (body mass index), body fat distribution based on physical methods (skin fold, perimeter) or laboratory (performance devices to identify the different compartments of fat based on bio-impedance)
4. The study of basal metabolism and respiratory quotient (by indirect calorimetry)
5. Fasting blood was collected for necessary biological samples and determining clinical, biochemical and oxidative stress parameters according to project proposal, as follows: isolation of peripheral mononuclear cells (PBMC), determining the intensity of respiratory burst (RB) by chemiluminescence, the determination of non-protein erythrocyte glutathione by spectrophotometric method and determination of MCP-1 (monocyte chemoattractand protein-1) by ELISA (kit EIA 4857, DRG Instruments, Germany).
6. Biochemical Profile for glucidic, lipidic, liver and kidney profile, including HbA1c (HPLC), fructozamine measurements (kit FRZ SPIN REACT REF1001158) and free fatty acids (AGL, MBS268756)
7. Determination of insulin, C-peptide, proinsulin (for the investigation of pancreatic beta-cell function) and also leptin, adiponectin and CTRP3 (for investigating the function of adipocyte) was performed for all subjects using commercially available ELISA kits according to the manufacturer's instructions, as follows: EIA-2935 (insulin), EIA-1560 (proinsulin), EIA-1293 (C-peptide), EIA 2395 (leptin), EIA-4177 (adiponectin), all from DRG Instruments, Germany and respectively kit Adipogen AG-45A-0042PP-KI01 for CTRP3.
8. Determination of IL-6 (EIA-4640), TNF-α (EIA-4641), ICAM (EIA 4781) and Reactive C Protein (CRP, EIA 3954) DRG Instruments, Germany (as inflammation markers), GLP-1 (kit MBS760336) and Pref-1
9. Determination of Total Oxidant Capacity (TOS, PerOX TOS/TOC KC5100, Immun Diagnostik) and Total Antioxidant Capacity (TAS, ImAmOx TAS/TAC KC 5200)
10. Determination of PON2 lactonase activity from PBMNC cells.
11. Determination of some antioxidant enzymes activity as Gluthation Peroxidase (GPx, kit CGP1-1KT, Sigma-Aldrich Co. LLC., St Louis, USA), Superoxide Dismutase (SOD, kit 19160-1KT-F, Sigma-Aldrich Co. LLC., St Louis, USA) and Paraoxonase 1 (PON1, arylesterase activity) were done for the patients we had also the samples after 12 months lifestyle changes.
Depending on the amount of funding in 2012-2016 we did some investigation and statistical analysis for some parameters on smaller datasets. For instance:
- we studied the association between paraoxonase 2, anthropometric markers and metabolic syndrome. The article with title "Negative association between paraoxonase 2, anthropometric markers and metabolic syndrome" was published in 2015 in Open Life Science 10, pp. 365-371 (impact factor 0.710)
- we study the associations of two inflammatory markers (IL-6 and TNF-alpha) with different metabolic variables in type 2 diabetic patients on 138 patients out of 517. The article with title "Changes in the serum proinflammatory cytokines in patients with elevated HOMA-IR and type 2 diabetes mellitus" was published in 2015 in Farmacia, vol.63 (1), pp.132-139 (Impact Factor 1.251)
- the results obtained from analysis of resting metabolic rate (RMR) were published on one hand in a book chapter included in Rainer Days Collection "Anthropology and Environment" named "The relationship between resting metabolic rate and anthropometric markers in patients with type 2 diabetes newly diagnosed" (vol. 5 Ed. Niculescu, pp. 152-159; PDF ISBN: 978-973-748-859-6) and on the other hand in the article entitled "Gender influence on resting metabolic rate and adipocytokines levels in newly diagnosed type 2 diabetic patients with metabolic syndrome" was published in 2014 in Rom. J. Diabetes Nutr. Metab. Dis. 21 (3): 193-202; (CNCSIS B+).
We also aimed to study the role of PON2 in modulation of synthesis and secretion of insulin in beta-pancreatic cell lines (PANC-1, INS-1E). With our model of in vitro islets, we showed that intracellular insulin accumulation was lower at higher glucose concentrations (glucotoxicity) and this was associated with insulin secretion and PON2 changes in all cell lines studied. The effect was amplified by adding oleic acid in a combination with palmitic acid (lipotoxicity).
Based on data and activities submitted, members of this research study group have published or presented following works:
- the project was presented in 2012, 2013, 2014, 2015 and 2016 at the Congress of the Romanian Medical Association (Bucharest, Romania) and also at the Congress of the Romanian Society of Diabetes, Nutrition and Metabolic Diseases (Sibiu, Romania)
- the project results were used by publishing 14 articles, 8 of which in ISI journals. In addition, three articles were submitted for publication in ISI journals in 2016 and two manuscripts are in preparation
- we presented in summary form of poster or oral presentations 33 national/international conferences in the field, including 26 posters and 7 oral presentations.
- a book chapter included in Rainer Days Collection "Anthropology and Environment" named "The relationship between resting metabolic rate and anthropometric markers in patients with type 2 diabetes newly diagnosed" (vol. 5 Ed. Niculescu, pp. 152-159; PDF ISBN: 978-973-748-859-6)
Following the concept that prevention is preferable to treatment, taking into account the results achieved in the present project during the 5 years of implementation, we have designed a method to predict the occurrence of diabetes, objective and accurate enough to motivate adoption of conduct relating to lifestyle or administration of plant extract. We are convinced that without a good prediction cannot conceive of any prevention program. In turn, the prediction mechanism requires a good knowledge of the pathogensis and identification by a validated prediction method.
To achieve this, our project results represented the basis of one dedicated software called "Diabetes prediction prototype", which is one of the products of our project. Briefly, the software includes the following sections: a) Input of variables (for example blood glucose values over a period of time); b) Processes k steps (represents the period of time for which the prediction is made); c) Step by step (gives the current calculations and the current state); d) Probability values of the last vector (a graphical view of the probabilities for high blood glucose in the future); e) Prediction of behavior (a graphical depiction of the daily probabilities for a specified time frame); f) Vector components - probability plot.
This software will be further validated with longer duration of follow-up patients, to follow (at least 3 years).
A second software "PromKappa" was used in this project to allow the employment of a class of genes associated with diabetes for the characterization of three diabetes phenotypes: T1D, T2D and IDM.
All project results were presented to the "One-Day Symposium" organized to the Romanian Academy on 25th April 2016 and were grouped in a book entitled „A GOOD PREVENTION NEEDS A GOOD PREDICTION", Editura ILEX, Bucharest, 2016.