Single Cell Characterization of Adipose Tissue in the Setting of HIV and Aging

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Project Summary of Supplement: Both the process of aging and HIV-infection are associated with pro-inflammatory environments that are characterized by elevated levels of cytokines and inflammatory mediators. Stimulation of innate immune receptors by both pathogen-associated molecular patterns and damage associated molecular patterns likely is contributing to the systemic inflammation seen with advancing age, and HIV-infection. There is increasing evidence that adipose tissue-derived inflammation likely contributes to this pro-inflammatory environment. Both obesity and redistribution of adipose tissue occur with the process of aging and HIV-infection and are strongly linked to metabolic syndrome. However, the mechanisms whereby adipose tissue contributes to the development of metabolic syndrome are not fully understood. Adipose tissue is a cellular community that includes adipocytes, endothelial cells, pre-adipocytes, and innate immune cells such as macrophages, dendritic cells, and monocytes. The purpose of this proposal is to characterize both immune and non- immune cell populations within adipose tissue using single cell RNA sequencing, in the context of aging, and HIV-infection in order to understand how they contribute to systemic inflammation. In Aim 1, fat pad biopsies will be collected from HIV-positive, and HIV-negative adults in the following age groups (21- 40) and (≥ 60 years). Multicolor flow cytometry and single cell RNA seq will be used to characterize both immune (macrophages, dendritic cells, monocytes) and non-immune cell populations within adipose tissue by evaluation of innate immune pathways, such as the NLRP3 inflammasome, cytokines (IL-1, IL-18, IL-6, TNF- , IL-10 etc.), and transcriptomics of specific cells to evaluate cellular signaling networks and characterize new cell populations. Aim 2 seeks to characterize protein expression in the context of adipose tissue architecture by using CyTOF imaging mass cytometry. The supernatant of adipose tissue cell cultures will also be collected and used to perform proteomic analysis of the secretome of adipose tissue cells in order to better understand the inflammatory output of these cells. Clinical characteristics (e.g., BMI) and co-morbidities (e.g., diabetes) will be evaluated in conjunction with the experimental data. Our findings will allow us to better understand the underlying innate immune mechanisms within specific cell populations that contribute to adipose tissue derived inflammation and ultimately affect the pro-inflammatory environment that is seen in the setting of aging, and HIV-infection. 2