Single-cell profiling reveals the origin of phenotypic variability in adipogenesis.
Phenotypic heterogeneity in a clonal cell population is a well-observed but poorly understood phenomenon. Here, a single-cell approach is employed to investigate non-mutative causes of phenotypic heterogeneity during the differentiation of 3T3-L1 cells into fat cells. Using coherent anti-Stokes Raman scattering microscopy and flow cytometry, adipogenic gene expression, insulin signaling, and glucose import are visualized simultaneously with lipid droplet accumulation in single cells. Expression of adipogenic genes PPARgamma, C/EBPalpha, aP2, LP2 suggests a commitment to fat cell differentiation in all cells. However, the lack of lipid droplet in many differentiating cells suggests adipogenic gene expression is insufficient for lipid droplet formation. Instead, cell-to-cell variability in lipid droplet formation is dependent on the cascade responses of an insulin signaling pathway which includes insulin sensitivity, kinase activity, glucose import, expression of an insulin degradation enzyme, and insulin degradation rate. Increased and prolonged insulin stimulation promotes lipid droplet accumulation in all differentiating cells. Single-cell profiling reveals the kinetics of an insulin signaling cascade as the origin of phenotypic variability in drug-inducible adipogenesis.
Le
Thuc T
TT
Weldon School of Biomedical Engineering, Purdue University, West Lafayette, Indiana, United States of America.
Cheng
Ji-Xin
JX
D041721:3T3-L1 Cells / Q000166:cytology / Q000502:physiology*; D050156:Adipogenesis / Q000502:physiology*; D000818:Animals; D002135:Calcium-Binding Proteins; D002454:Cell Differentiation / Q000502:physiology*; D005947:Glucose / Q000378:metabolism; D007328:Insulin / Q000378:metabolism; D007339:Insulysin / Q000235:genetics / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D050356:Lipid Metabolism; D051379:Mice; D008853:Microscopy / Q000295:instrumentation / Q000379:methods; D015388:Organelles / Q000378:metabolism / Q000648:ultrastructure; D047495:PPAR gamma / Q000235:genetics / Q000378:metabolism; D010641:Phenotype; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology
D002135:Calcium-Binding Proteins; D005947:Glucose; D007328:Insulin; D007339:Insulysin; D036341:Intercellular Signaling Peptides and Proteins; D047495:PPAR gamma; D019869:Phosphatidylinositol 3-Kinases; C512925:Dlk1 protein, mouse