Proof-of-concept for CRISPR/Cas9 gene editing in human preadipocytes: Deletion of FKBP5 and PPARG and effects on adipocyte differentiation and metabolism

• Published in: Scientific reports Vol. 10; no. 1; p. 10565
• Main Authors: Kamble, Prasad G., Hetty, Susanne, Vranic, Milica, Almby, Kristina, Castillejo-López, Casimiro, Abalo, Xesús M., Pereira, Maria J., Eriksson, Jan W.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 29-06-2020; Nature Publishing Group
• Subjects: 631/1647; 631/443; Adipocytes; Adipocytes - metabolism; Adipogenesis; Adipogenesis - genetics; Adipose tissue; Adipose Tissue - metabolism; Adult; Aged; Animal models; Cell culture; Cell Differentiation - genetics; Clustered Regularly Interspaced Short Palindromic Repeats - genetics; CRISPR; CRISPR-Cas Systems - genetics; Diabetes mellitus (non-insulin dependent); Electroporation; Female; Gene deletion; Gene Editing - methods; Gene Knockout Techniques - methods; Genes; Genome editing; Genomes; Glucocorticoid receptors; Glucocorticoids; Glucose metabolism; Humanities and Social Sciences; Humans; Insulin; Metabolism; Middle Aged; multidisciplinary; Peroxisome proliferator-activated receptors; Phenotypes; PPAR gamma - genetics; Preadipocytes; Proof of Concept Study; Science; Science (multidisciplinary); Tacrolimus Binding Proteins - genetics; Transcription
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-020-67293-y

CRISPR/Cas9 has revolutionized the genome-editing field. So far, successful application in human adipose tissue has not been convincingly shown. We present a method for gene knockout using electroporation in preadipocytes from human adipose tissue that achieved at least 90% efficiency without any need for selection of edited cells or clonal isolation. We knocked out the
FKBP5
and
PPARG
genes in preadipocytes and studied the resulting phenotypes. PPARG knockout prevented differentiation into adipocytes. Conversely, deletion of FKBP51, the protein coded by the
FKBP5
gene, did not affect adipogenesis. Instead, it markedly modulated glucocorticoid effects on adipocyte glucose metabolism and, furthermore, we show some evidence of altered transcriptional activity of glucocorticoid receptors. This has potential implications for the development of insulin resistance and type 2 diabetes. The reported method is simple, easy to adapt, and enables the use of human primary preadipocytes instead of animal adipose cell models to assess the role of key genes and their products in adipose tissue development, metabolism and pathobiology.