An optimised protocol for the investigation of insulin signalling in a human cell culture model of adipogenesis

ABSTRACT

While there is no standardized protocol for the differentiation of human adipocytes in culture, common themes exist in the use of supra-physiological glucose and hormone concentrations, and an absence of exogenous fatty acids. These factors can have detrimental effects on some aspects of adipogenesis and adipocyte function. Here, we present methods for modifying the adipogenic differentiation protocol to overcome impaired glucose uptake and insulin signalling in human adipose-derived stem cell lines derived from the stromal vascular fraction of abdominal and gluteal subcutaneous adipose tissue. By reducing the length of exposure to adipogenic hormones, in combination with a physiological glucose concentration (5 mM), and the provision of exogenous fatty acids (reflecting typical dietary fatty acids), we were able to restore early insulin signalling events and glucose uptake, which were impaired by extended use of hormones and a high glucose concentration, respectively. Furthermore, the addition of exogenous fatty acids greatly increased the storage of triglycerides and removed the artificial demand to synthesize all fatty acids by de novo lipogenesis. Thus, modifying the adipogenic cocktail can enhance functional aspects of human adipocytes in vitro and is an important variable to consider prior to in vitro investigations into adipocyte biology.

KEYWORDS:

• Adipogenic cocktail
• adipose stem cell
• adipocyte
• cell model
• insulin signalling
• glucose uptake

Acknowledgments

This research was funded by the National Institute for Health Research (NIHR) Oxford Biomedical Research Centre (BRC) and the British Heart Foundation [RG/17/1/32663]. JG and KP received funding from the Medical Research Council under the Oxford-MRC Doctoral Training Partnership.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Data availability statement

The data that support the findings of this study are available from the corresponding author, K.E.P., upon reasonable request.

Correction Statement

This article has been corrected with minor changes. These changes do not impact the academic content of the article.

Additional information

Funding

This work was supported by the British Heart Foundation [RG/17/1/32663].