An Adipocyte Cell Culture Model to Study the Impact of Protein and Micro-RNA Modulation on Adipocyte Function
- Resource Type
- Authors
- Jager, Jennifer; Gaudfrin, Mélanie; Gilleron, Jérôme; Cormont, Mireille; Tanti, Jean-François; Tanti, Jean‐François
- Source
- Journal of visualized experiments : JoVE
Journal of visualized experiments : JoVE, JoVE, 2021, ⟨10.3791/61925⟩
- Subject
- General Chemical Engineering
Cell Culture Techniques
General Biochemistry, Genetics and Molecular Biology
chemistry.chemical_compound
Mice
Insulin resistance
Adipocyte
3T3-L1 Cells
microRNA
medicine
Adipocytes
Animals
Humans
[SDV.MHEP.EM] Life Sciences [q-bio]/Human health and pathology/Endocrinology and metabolism
cell culture
biology
General Immunology and Microbiology
Chemistry
General Neuroscience
micro-RNA (miR)
RNA
Cell Differentiation
Transfection
small interfering RNA (si-RNA)
[SDV.MHEP.EM]Life Sciences [q-bio]/Human health and pathology/Endocrinology and metabolism
medicine.disease
Cell biology
Insulin receptor
MicroRNAs
transfection
Diabetes Mellitus, Type 2
Cell culture
Lipogenesis
biology.protein
- Language
- ISSN
- 1940-087X
Alteration of adipocyte function contributes to the pathogenesis of metabolic diseases including Type 2 diabetes and insulin resistance. This highlights the need to better understand the molecular mechanism involved in adipocyte dysfunction to develop new therapies against obesity-related diseases. Modulating the expression of proteins and micro-RNAs in adipocytes remains highly challenging. This paper describes a protocol to differentiate murine fibroblasts into mature adipocytes and to modulate the expression of proteins and micro-RNAs in mature adipocytes through reverse-transfection using small-interfering RNA (siRNA) and micro-RNA mimicking (miR mimic) oligonucleotides. This reverse-transfection protocol involves the incubation of the transfection reagent and the oligonucleotides to form a complex in the cell culture plate to which the mature adipocytes are added. The adipocytes are then allowed to reattach to the adherent plate surface in the presence of the oligonucleotides/transfection reagent complex. Functional analyses such as the study of insulin signaling, glucose uptake, lipogenesis, and lipolysis can be performed on the transfected 3T3-L1 mature adipocytes to study the impact of protein or micro-RNA manipulation on adipocyte function.