Canine obesity and inflammation: a study of microRNAs involved in the expression of macrophages in adipose tissue

Author(s)

Paola Caroline da Silva Nunes

• Document type: Master's Dissertation
• Press: Pirassununga.
• Institution: Universidade de São Paulo (USP). Faculdade de Zootecnica e Engenharia de Alimentos (FZE/BT)
• Defense date: 2022-06-22

Examining board members

Deise Carla Almeida Leite Dellova; Márcia de Oliveira Sampaio Gomes; Trícia Maria Ferreira de Sousa Oliveira

• Advisor: Deise Carla Almeida Leite Dellova

Abstract

Adipose tissue is a metabolic and endocrine organ, and its adipocytes have the ability to synthesize and secrete extracellular vesicles (EVs), thus allowing intercellular communication. MicroRNAs (miRNAs), present in adipose tissue and circulating EVs, are able to regulate gene expression. This study aimed to evaluate the small EVs in the serum of obese dogs, in addition to: 1) investigating the influence of weight loss on the size and concentration of these vesicles; 2) to evaluate the expression of three target miRNAs in small EVs from obese female dogs; and 3) to analyze markers of inflammation and a possible relationship between inflammatory status, serum EVs and target miRNAs expressed in serum EVs and adipose tissue of obese female dogs. The second chapter demonstrated that small EVs were isolated from whole blood, by means of serial centrifugations and ultracentrifugations, using the NanoSight equipment to determine the size and concentration of these small EVs present in the serum of overweight and obese dogs, before and after the weight loss program. The reduction in body weight and fat accumulation in these dogs was accompanied by a decrease in the size and concentration of small EVs, thus suggesting a reduction in the secretory activity of the dogs\' adipose tissue after 4 months of the weight loss program. In the third chapter, the size and concentration of serum EVs in obese female dogs was evaluated, as well as the miRNAs (miR-132, miR-26b and miR-155) contained in these EVs. The miRNAs were extracted from the small EVs and analyzed by the real-time PCR technique. A principal component analysis showed that a group of obese female dogs (obese with alteration) clustered together with higher levels of small EVs and serum triglyceride concentrations. These females had higher serum triglycerides, higher concentration of small EVs and higher expression of miR-132 and miR-26b, suggesting that with the expansion of adipose tissue, caused by obesity, greater production and release of EVs is occurring, which is probably influencing intercellular communication due to the action of the expression of miRNAs contained in these small EVs, as they seem to be involved in lipid metabolism and adipogenesis. Finally, in the fourth chapter, miR-26b, miR-155 and miR-132 and their target genes (MYCBP, SOCS1 and FOXO3) were evaluated with macrophage infiltrate and inflammation in the subcutaneous adipose tissue of obese female dogs, compared to control females. MiRNAs were extracted from small EVs and adipose tissue and analyzed by real-time PCR. The analysis of gene expression was also performed using the real-time PCR technique. No differences were observed regarding markers of inflammation, macrophage infiltration in subcutaneous adipose tissue, concentration and size of serum EVs, miRNA expression in EVs and subcutaneous adipose tissue, and expression of miRNA target genes in subcutaneous adipose tissue. However, we observed that the obese female dogs presented hypertrophy of the subcutaneous adipose tissue, revealing an alteration in the homeostasis of this tissue. No evidence was found to prove that the inflammatory process was installed and that is probably why we did not find changes in miRNAs and target genes evaluated, as well as the number of macrophages infiltrated in the subcutaneous adipose tissue. In short, these results contribute to the field of research on canine obesity, aiming to elucidate the role of small EVs and their miRNAs in this disease. (AU)

• FAPESP's process: 18/26547-0 - Canine obesity and inflammation: a study of microRNAs involved in the expression of macrophages in adipose tissue
• Grantee: Paola Caroline da Silva Nunes
• Support Opportunities: Scholarships in Brazil - Master