Phd in MOLECULAR MEDICINE

Thesis title: Role of SIRT1 in the regulation of adipose tissue and metabolism: implications with the ketogenic diet.

Sirtuin1 (SIRT1) has a regulatory role mainly in metabolism and aging, extending health span through effects able to prevent metabolic derangements and degenerative diseases, including aging-related bone loss. Being a histone deacetylase using NAD as a cofactor, its activity increases with caloric restriction or starvation. Overnutrition reduce SIRT1, while transgenic mice for SIRT1 overexpression are protected from fatty liver, type 2 diabetes (T2D), age- induced cancer and osteoporosis. We demonstrated that circulating SIRT1 – as well as the tissue – is inversely correlated with fat amounts and coherently associated with the levels of the two well-known adipokines adiponectin and leptin, being predictive of visceral adipose tissue (VAT), specifically. In addition, circulating SIRT1 proved to be directly correlated with bone mineral density and was the best predictor of trabecular structure, reflecting the negative relationship with sclerostin – an osteoblast inhibitor – observed in murine studies. Ketogenesis is a metabolic process involved in controlling energy homeostasis by a fine epigenetic regulation implemented during fasting or intense exercise. Endogenous or exogenous ketosis delay aging by improving anti-inflammatory and mitochondrial activities, mnemonic and muscle performance and reduces metabolic diseases through the disposal of fatty acids. Being proposed as an alternative therapy option even during acute illness such as COVID-19, the ketogenic diet (KD) has been successfully introduced for the treatment of obesity and its main complications such as NAFLD and T2D. Recent advances in understanding the influence on health span and the shared outcomes between ketone bodies and SIRT1 suggest a joint work with the same beneficial intent, probably also via epigenetic modulation. The aim of our study was to understand whether a KD leads to the increase of SIRT1, assuming the role of SIRT1 as a downstream effector of the metabolic adaptations observed during ketosis, beyond caloric restriction and weight loss. We evaluated 20 C57BL/6J 8 weeks old male mice treated with isocaloric KD or standard diet of the same caloric intake for 4 weeks. Blood and urine were collected at baseline and after the sacrifice. SIRT1 was evaluated in serum, liver, and adipose tissue including VAT, subcutaneous adipose tissue (SAT) and brown adipose tissue (BAT). All organs were weighed dry and collected as follows: 1/2 in 10% Neutral Buffered Formalin Solution (for histology) and 1/2 dry at -80°C (for molecular analyses). After 4 weeks, despite the 1.7-fold higher caloric intake, KD-fed mice showed no significative changes in weight gain, VAT (g), SAT (g), glycemia (mg/dl) and TRG (mg/dl). Ketosis was efficaciously achieved in the treated group (p<0.001) and serum SIRT1 increased significantly compared to the controls under normal chow (p<0.05). Tissue SIRT1 expression increased both in VAT and SAT in mice under KD compared to normal chow group (p<0.05), while no differences were seen regarding liver and BAT. This study supports the hypothesis of a ketone body-mediated activation of SIRT1, specifically exerted in white adipose tissue, able to counteract the development of metabolic derangements. The possible cooperation between ketone bodies and SIRT1 could be of considerable interest in terms of promoting metabolic health.