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Digital archive of theses discussed at the University of Pisa

 

Thesis etd-10142021-115838


Thesis type
Tesi di specializzazione (5 anni)
Author
MENGOZZI, ALESSANDRO
URN
etd-10142021-115838
Thesis title
Targeting SIRT1 rescues age- and obesity-induced human microvascular dysfunction
Department
MEDICINA CLINICA E SPERIMENTALE
Course of study
MEDICINA INTERNA
Supervisors
relatore Prof. Virdis, Agostino
Keywords
  • ageing
  • endothelial function
  • mitochondria
  • obesity
  • SIRT1
Graduation session start date
03/11/2021
Availability
Withheld
Release date
03/11/2024
Summary
Background.
Age- and metabolism-related microvascular endothelial dysfunction is the first step of the healthy-to-disease phenotype transition. In vitro and in vivo evidence suggests a key role of deacetylase silent information regulator two-1 (SIRT1) in mediating the benefits of caloric restriction, likely through a modulation of mitochondria environment. However, its potential contribution in preserving cardiometabolic health in human is yet to be explored. The present study aims to investigate the role of SIRT1 in age- and obesity-related microvascular dysfunction in humans.

Material and methods.
Patients with obesity (n=48) and control subjects (n=47) undergoing elective laparoscopic surgery were recruited and stratified based on their age (above or below 40 years) in four groups: young patients with obesity (YO), young control (YC), old patients with obesity (OO) and control (OC). Small resistance arteries were extracted from adipose tissue and mounted on a pressurised micromyograph. Media-lumen ratio (M/L) and media cross-sectional area (MCSA) were assessed as functional parameters. Endothelial function was measured as percentage relaxation to acetylcholine, alone and after incubation with L-NAME to accurately assess contribution of NO. The same curves were repeated after incubation with a SIRT1 agonist (SRT1720) and a mitochondria reactive oxygen species (mtROS) scavenger (MitoTEMPO). Vascular production of mtROS and nitric oxide availability were assessed by confocal microscopy by the means of MitoSOX and DAF-FM. Gene expression of SIRT1 and mitochondrial proteins was assessed by qPCR directly on the vessel wall. ChIP assay was employed to investigate SIRT1-dependent epigenetic regulation of mitochondrial proteins.

Results.
M/L showed a direct relationship with age, and the slope of this relationship was higher in the obese population. Endothelial function was progressively reduced across the four groups, with a YC-OC-YO-OO pattern. The same pattern was observed for NO availability, while mtROS followed was increased along the same groups. Patients with obesity showed lower expression of SIRT1 and mitochondrial antioxidant proteins (FOXO3 and SOD2), and a higher expression of pro-oxidant and aging mitochondria proteins p66Shc and Arginase II. Endothelial function of OO, YO and OC groups was rescued by SRT1720 and MitoTEMPO in a similar manner. Consistently, vessel of OO group incubated with SRT1720 showed a reduced mtROS, an increased nitric oxide availability and an improved response to mitochondrial swelling assay, superimposable to the one of YC group. These effects were explained by SIRT1-dependent chromatin changes leading to reduced p66Shc expression and upregulation of proteins involved in mitochondria respiratory chain (SIRT3, ATP6, CytB, ND2, ND5).

Conclusions.
SIRT1 is reduced in the vascular wall of small resistance arteries of patients with obesity and with older age. This is reflected by a reduced NO and an increased mtROS, which in turn translate into an impaired endothelial function. Therapeutic modulation of SIRT1 directly on the small vessels restores obesity and age-related endothelial dysfunction. It exerts an epigenetic control on several mitochondrial pathways influencing mtROS production, NO availability, and the expression of proteins of the mitochondria respiratory chain. Early targeting of SIRT1 might represent a crucial strategy to prevent or revert the microvascular dysfunction related to age and obesity.
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