Fat's all, folks: culturing and manipulating peri-prostatic adipocytes to probe impacts on prostate cancer biology

Obesity, officially recognised as a global epidemic by the World Health Organization, will soon overtake smoking as the largest preventable risk factor for cancer. By 2035, more than half the world's population is expected to be overweight or obese with a significant increase in obesity-related health expenditures. However, despite the increase in prevalence and the overall lower life expectancy associated with obesity, mechanisms underpinning obesity-driven diseases are not well understood. Adipocytes pose many challenges for in vitro culture due to their poor cell-to-surface attachment and low viability. Their large size and high lipid content can also present methodological challenges for downstream experiments. Several mouse and human-derived primary pre-adipocyte cell lines have been established over the years. However, they show limited renewal capacity and they cannot be cultured long term in vitro. Commercial cell lines available, which can be cultured long term, fail to represent organ-specific adipocyte heterogeneity. Adipose tissue from different organs and fat depots can show significant heterogeneity in terms of metabolism and overall secretome and extracellular matrix production. The prostate, for example, is surrounded by peri-prostatic adipose tissue (PPAT), the volume of which is associated with an increased risk of lethal prostate cancer and a reduced therapy response. Here, we outline a protocol for ex vivo culture of fresh PPAT and non-prostatic adipose tissue (NPAT), which reflects donor- and depot-specific characteristics. Ex vivo culture of PPAT/NPAT explants maintains cell-cell interactions and preserves local tissue architecture within adipose tissue. We have also described establishment of immortalised, patient PPAT-derived pre-adipocytes and patient-matched NPAT pre-adipocytes that can be in vitro differentiated into mature adipocytes. The protocols outlined here could be readily adapted to other organ-specific fat depots, such as mammary/bone marrow adipose tissue, and to tissues of non-human origin.