Effects of obesity on adipose stromal/stem cell immunomodulation and mitochondrial respiration capacity
Adnan, Amna (2021)
2021
Master's Programme in Biomedical Technology
Lääketieteen ja terveysteknologian tiedekunta - Faculty of Medicine and Health Technology
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Hyväksymispäivämäärä
2021-05-12
Julkaisun pysyvä osoite on
https://urn.fi/URN:NBN:fi:tuni-202104233368
https://urn.fi/URN:NBN:fi:tuni-202104233368
Tiivistelmä
Background and aims: Adipose tissue (AT) derived adipose stem/stromal cells (ASC) are multipotent cells. They not only possess excellent proliferation and differentiation capacity but are also known for their low immunogenicity and the ability for immunomodulation. Obesity is be-coming increasingly a worldwide health concern causing low-grade chronic inflammation in AT. Macrophages are known to keep the tissue homeostasis. Obesity-induced inflammation not only disturbs the tissue-resident macrophage ratio but also affects the ASC functions. Metabolic dysfunction of AT has been previously studied but the effect of obesity-related inflammation on ASC functions is still not clearly known. This study aimed to investigate the effect of obesity on ASC immunomodulatory capacity, especially macrophage polarization and cytokine secretion. Also, the aim was to study the effect of donor weight and in-vitro inflammatory environment on the mitochondrial respiratory capacity of ASC.
Materials and methods: ASC were isolated from human AT samples that were obtained from two donors undergoing bariatric surgery, before and after weight loss. Monocyte derived macrophages were polarized from frozen PBMCs (peripheral blood mono nuclear cells) into pro-inflammatory (M1), anti-inflammatory (M2), and regulatory macrophages (Mreg). These three types of macrophages were cultured in mono and co-culture with ASC collected from two different donors before and after weight loss. Macrophage polarization was studied with light and fluorescent microscopy, immunophenotype was analyzed with flow cytometry analysis and the amount of cytokine secreted in mono and co-cultures with ASC. Metabolic capacity of ASC collected from lean and obese donors was measured by quantifying the mitochondrial respiration capacity of these cells at two different time points.
Results: M1, M2 and Mreg macrophages with characteristic morphology, phenotype, and cytokine secretion were polarized from frozen PBMCs. M1 macrophages showed high expression of CD86, CD11c and HLA-DR, M2 showed high expression of CD206, CD163 and CD86 while Mreg macrophages showed decreased expression of above-mentioned markers. The mono and co-cultures of M1 and Mreg macrophages secreted pro-inflammatory cytokines. The co-cultures with obese derived ASC secreted more pro-inflammatory cytokines, particularly IL-6, IL-12p70 and MCP-1. Moreover, the cellular respiratory capacity of ASC increased after the weight loss.
Conclusion: Our study showed that different macrophage types can be polarized from frozen PBMCs in vitro. It was studied that Mreg macrophages have both regulatory and pro-inflammatory nature. Also, it was observed that secretion of proinflammatory cytokines particularly IL-6, IL-12p70 and MCP-1 was significantly high in ASC obtained from obese donors com-pared with M1 monocultures and the immunosuppressive capacity of ASC was independent of donor weight. Moreover, it was observed that ASC after weight loss have increased cellular respiration capacity and inflammatory environment rapidly increases the cellular respiration capacity of ASC in vitro. Although this study showed some insight of ASC immunomodulation, additional co-cultures of macrophages with higher number of ASC obtained from obese and lean donors should be carried out with optimized study design that will provide further insight of obesity induced inflammation and its effects on immunomodulatory functions of ASC.
Materials and methods: ASC were isolated from human AT samples that were obtained from two donors undergoing bariatric surgery, before and after weight loss. Monocyte derived macrophages were polarized from frozen PBMCs (peripheral blood mono nuclear cells) into pro-inflammatory (M1), anti-inflammatory (M2), and regulatory macrophages (Mreg). These three types of macrophages were cultured in mono and co-culture with ASC collected from two different donors before and after weight loss. Macrophage polarization was studied with light and fluorescent microscopy, immunophenotype was analyzed with flow cytometry analysis and the amount of cytokine secreted in mono and co-cultures with ASC. Metabolic capacity of ASC collected from lean and obese donors was measured by quantifying the mitochondrial respiration capacity of these cells at two different time points.
Results: M1, M2 and Mreg macrophages with characteristic morphology, phenotype, and cytokine secretion were polarized from frozen PBMCs. M1 macrophages showed high expression of CD86, CD11c and HLA-DR, M2 showed high expression of CD206, CD163 and CD86 while Mreg macrophages showed decreased expression of above-mentioned markers. The mono and co-cultures of M1 and Mreg macrophages secreted pro-inflammatory cytokines. The co-cultures with obese derived ASC secreted more pro-inflammatory cytokines, particularly IL-6, IL-12p70 and MCP-1. Moreover, the cellular respiratory capacity of ASC increased after the weight loss.
Conclusion: Our study showed that different macrophage types can be polarized from frozen PBMCs in vitro. It was studied that Mreg macrophages have both regulatory and pro-inflammatory nature. Also, it was observed that secretion of proinflammatory cytokines particularly IL-6, IL-12p70 and MCP-1 was significantly high in ASC obtained from obese donors com-pared with M1 monocultures and the immunosuppressive capacity of ASC was independent of donor weight. Moreover, it was observed that ASC after weight loss have increased cellular respiration capacity and inflammatory environment rapidly increases the cellular respiration capacity of ASC in vitro. Although this study showed some insight of ASC immunomodulation, additional co-cultures of macrophages with higher number of ASC obtained from obese and lean donors should be carried out with optimized study design that will provide further insight of obesity induced inflammation and its effects on immunomodulatory functions of ASC.