Poster Presentation ESA-SRB-ANZBMS 2024 in conjunction with ENSA

Flow cytometric isolation of adrenocortical cells using CD36L1 expression (#505)

Brigette M Clarke 1 2 3 , Svjetlana Kireta 1 4 , Patrick T Coates 1 4 , David J Torpy 1 2 , Plinio R Hurtado 1 4
  1. University of Adelaide , Adelaide, South Australia, Australia
  2. Endocrine and Metabolic Unit, Royal Adelaide Hospital, Adelaide, South Australia, Australia
  3. Endocrine and Diabetes Services, The Queen Elizabeth Hospital, Woodville South, SA, Australia
  4. Central and Northern Adelaide Renal and Transplantation Service, Royal Adelaide Hospital, Adelaide, South Australia, Australia

Flow cytometry permits the rapid analysis of the characteristics of large numbers of individual cells within suspension. Flow cytometric studies of adrenocortical cells have been very limited and there is no established cell surface marker for the isolation of a pure population of primary adrenocortical cells. The scavenger receptor class B member 1 (SCARB1/CD36L1) is a cell surface receptor with high affinity to high-density lipoproteins, involved in the uptake and delivery of cholesterol ester to cells. Within the human adrenal CD36L1 contributes to cholesterol transportation into steroidogenic cells for steroid hormone production (1). The primary adrenal cell isolate comprises a heterogenous cell population including fibroblasts, hematopoietic, vascular endothelial, medullary and adrenocortical cells. The aim of this experiment was to confirm whether CD36L1 expression could be used to isolate adrenocortical cells from this mixed cell population.

Primary adrenal cells isolated from human adrenal glands were prepared and stained with Fixable Viability stain 780, anti-human CD45, CD34 and CD36L-1 before undergoing flow cytometric cell sorting. The primary cell population was gated to sort viable singlets into CD45+, CD45-/CD34+ and CD45-/CD36L1+ populations. Sorted CD36L1+ cells were evaluated by assessment of ACTH-responsiveness in cell culture and comparison of adrenal specific gene expression as measured by real-time RT-PCR to unsorted primary adrenocortical cells.   

CD36L1+ cells demonstrate ACTH responsiveness in cell culture with stimulated cortisol production equivalent to the unsorted primary cell isolate. Expression of SF1, CYP11B1, CYP11B2 & CYP17A1 by CD36L1+ cells was similar to or increased when compared to the unsorted primary adrenal cell isolate.

CD36L1 expression can be used for flow cytometric isolation of a pure adrenocortical cell population. This finding is important as a step towards further characterisation studies of primary adrenocortical cells by flow cytometry with the view to establishing specific surface markers for zonal identification and isolation.

  1. 1. Azhar S, Reaven E. Scavenger receptor class BI and selective cholesteryl ester uptake: partners in the regulation of steroidogenesis. Mol Cell Endocrinol. 2002;195(1-2):1-26.