Susceptibility to Infectious Diseases in obEsity: an endocRine trAnslational socioLogic Evaluation, "SIDERALE"

The current project aims at investigating 1) the susceptibility to infectious disease in obese patients, the role of tailored dietetic schemes of Mediterranean diet (MD), and melatonin supplementation on infective events; 2) association of the oral and/or gut microbiota signature with bacterial or viral infections in the above described obese cohorts; 3) the mutual relationship between biomedical values, environmental conditions and lifestyle habits through structured questionnaires and therefore the possible impact of this study on the National Health System (NHS).

The role of melatonin as chronobiologic influencer of circadian metabolic processes and as anti-obesogenic and weight-reducing effector comes to the fore in these last decades. Moreover, melatonin is also known to possess anti-inflammatory and antioxidant properties that may ameliorate the condition of low-grade chronic inflammation (metaflammation) observed in patients with obesity. Nevertheless, although there has been a tremendous interest in the role of the clock genes in regulating metabolic processes, relatively less effort has been expended examining how the regulation of circadian rhythms and metabolic inputs can affect the susceptibility to infections. To address this question, Unit 1 will enroll patients with essential obesity (20 subjects, BMI :30-35 Kg/m2) and with genetic forms of obesity (20 subjects) of both genders, aged 18-65yrs, to the main proof of concept pilot study.

At baseline (T0), patients will be subjected to anthropometric determinations, including body weight and height for BMI calculation, body composition by mean of DXA Scan, blood, serum and urine collection, oral cavity swab and stool sampling. Patients will undergo a session of behavioural dietary counselling by which patients will be educated to adhere to a hypocaloric MD for the first 12w (T1) and to add to the MD the supplementation with 1mg/die of melatonin for other 12w (T2). The same patients will be visited and subjected to the anthropometric determinations and biological samples at T0, T1, T2 of protocol. The endpoints for Unit 1 will be: 1.to investigate the impact of tailored hypocaloric MD and hypocaloric MD plus melatonin (supplement formulation, 1mg/die before sleep) on the number of events - i.e. flu- or flulike syndromes, skin, respiratory, digestive, urinary infections - per patient of the 4 groups, recording the number of infectious events during the clinical examinations in six months and 1 year follow up and detect the SARS-CoV-2, Epstein-Barr, Cytomegalovirus, Varicella and Measles IgG and IgM, hepatitis C and hepatitis B core antibodies and Quantiferon TB Gold (QFT-GIT). The obtained clinical results on infections events from the SIDERALE study will be used to formulate hypotheses for a prospective study and for the formulation of a specific validate questionnaire.

2.to assess the impact of a tailored hypocaloric MD and hypocaloric MD plus melatonin (supplement formulation, 1mg/die before sleep) on anthropometric determinations, glucose and lipid metabolism, thyroid and adrenal axis, leptin, adiponectin, IGFBP2, sirtuin 1 (SIRT1), ESR, high-sensitivity CRP, fibrinogen and inflammatory cytokines (TNF, IL-6, IL-1Beta, IL-18) and melatonin that will be evaluated at T0, T1, T2.

3.to assess the impact of a tailored hypocaloric MD and hypocaloric MD plus melatonin (supplement formulation, 1mg/die before sleep) on the entrainment of circadian clock misalignment, evaluating clock gene expression by RT-qPCR in peripheral blood mononuclear cells (PBMC) and concomitantly, on the regulation of Toll like-receptors (TLRs) and suppressor cytokine signaling 3 (SOCS3), known to play a crucial role in eliciting immunity and improving the well-known susceptibility to infections in patients with obesity, all evaluated by RT-qPCR in PBMC of all patients cohorts at 8 am, 12 pm and 12 am.

4.to quantify the impact of our intervention on the NHS in terms of decrease of antibiotic and antiviral treatment and ER (emergency room) admissions in six months and 1 year of follow up.

Moreover, an in vitro analysis will be performed on 3T3-L1 cells (mouse pre-adipocytes). 3T3-L1 differentiated in adipocytes will be synchronized and treated with melatonin in presence of TLRs agonists (i.e.

lipopolysaccharide, Pam3CSK, ssRNA40, imiquimod). Evaluation of TLRs signalling pathway will be performed by western blot. Evaluation of inflammatory cytokines (TNF, IL-6, IL-1, IL-18) and chemokines (MCP-1, CXCL5) will be performed by CLIA and/or ELISA and/or western blot.

• Change in body weight (kg) after hypocaloric MD and hypocaloric MD plus melatonin in patients with obesity and lipodystrophy. [Срок оценки: Change from baseline at 3-6-12 months]
• BMI modification after hypocaloric MD and hypocaloric MD plus melatonin in patients with obesity and lipodystrophy: body weight (kg) will be combined with height (m) to obtain BMI (kg/m^2) [Срок оценки: Change from baseline at 3-6-12 months]
• Change in waist and hip circumference after hypocaloric MD and hypocaloric MD plus melatonin in patients with obesity and lipodystrophy. [Срок оценки: Change from baseline at 3-6-12 months]
• Change in body composition assessed by Bioelectrical impedance analysis (BIA) after hypocaloric MD and hypocaloric MD plus melatonin in patients with obesity and lipodystrophy . [Срок оценки: Change from baseline at 3-6-12 months]
• Change in body composition assessed by DXA scan after hypocaloric MD and hypocaloric MD plus melatonin in patients with obesity and lipodystrophy. [Срок оценки: Change from baseline at 3-6-12 months]
• Change in glucose profile assessed by blood glucose levels after hypocaloric MD and hypocaloric MD plus melatonin in patients with obesity and lipodystrophy. [Срок оценки: Change from baseline at 3-6-12 months]
• Change in glucose profile assessed by blood glycated hemoglobin (HbA1c) levels after hypocaloric MD and hypocaloric MD plus melatonin in patients with obesity and lipodystrophy. [Срок оценки: Change from baseline at 3-6-12 months]
• Change in insulin profile assessed by blood insulin levels after hypocaloric MD and hypocaloric MD plus melatonin in patients with obesity and lipodystrophy. [Срок оценки: Change from baseline at 3-6-12 months]
• Change in insulin resistance assessed by homeostasis model assessment - insulin resistance (HOMA-IR) Index after hypocaloric MD and hypocaloric MD plus in patients with obesity and lipodystrophy. [Срок оценки: Change from baseline at 3-6-12 months]
• Change in lipid profile assessed by blood total cholesterol levels after hypocaloric MD and hypocaloric MD plus melatonin in patients with obesity and lipodystrophy. [Срок оценки: Change from baseline at 3-6-12 months]
• Change in lipid profile assessed by blood LDL cholesterol levels after hypocaloric MD and hypocaloric MD plus in patients with obesity and lipodystrophy. [Срок оценки: Change from baseline at 3-6-12 months]
• Change in lipid profile assessed by blood HDL cholesterol levels after hypocaloric MD and hypocaloric MD plus melatonin in patients with obesity and lipodystrophy. [Срок оценки: Change from baseline at 3-6-12 months]