Citation
- Authors: Picarda E. et al.
- Year: 2022
- Journal: Sci Adv 8 eabm7012
- Applications: in vitro / DNA / jetPRIME
- Cell type: HEK-293T
Description: Human embryonic kidney Fibroblast
Known as: HEK293T, 293T
Method
Lentivirus production
One day before transfection, 293 T cells were seeded on collagen-coated 10-cm cell culture dish in DMEM with l-glutamine, 10% FBS, and 1% pen/strep, at a density allowing for 80% confluency after 24 hours. Cells
were transduced using a jetPRIME transfection reagent (Polyplus) according to the manufacturer’s protocol. Briefly, the lentiviral vector containing the expression cassette was cotransfected with the envelope expressing plasmid pCI-VSVG and the lentiviral packaging plasmid pCMVR8.74 for 4 hours. Then, the transfection medium was replaced with fresh DMEM with l-glutamine, 10% FBS, and 1% pen/strep. Forty-eight hours later, the viral supernatant was collected, filtered through 0.45-um mesh to remove cellular debris, and stored at −80°C.
Abstract
The immune checkpoint B7-H3 (CD276) is a member of the B7 family that has been studied in the tumor microenvironment and immunotherapy, but its potential role in metabolism remains largely unknown. Here, we show that B7-H3 is highly expressed in mouse and human adipose tissue at steady state, with the highest levels in adipocyte progenitor cells. B7-H3 is rapidly down-regulated upon the initiation of adipocyte differentiation. Combined RNA sequencing and metabolic studies reveal that B7-H3 stimulates glycolytic and mitochondrial activity of adipocyte progenitors. Loss of B7-H3 in progenitors results in impaired oxidative metabolism program and increased lipid accumulation in derived adipocytes. Consistent with these observations, mice knocked out for B7-H3 develop spontaneous obesity, metabolic dysfunction, and adipose tissue inflammation. Our results reveal an unexpected metabolic role for B7-H3 in adipose tissue and open potential new avenues for the treatment of metabolic diseases by targeting the B7-H3 pathway.