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Isolating Brown Adipocytes from Murine Interscapular Brown Adipose Tissue for Gene and Protein Expression Analysis
Published on: March 12, 2021
Microarray profiling of isolated abdominal subcutaneous adipocytes from obese vs non-obese Pima Indians: increased
1Obesity and Diabetes Clinical Research Section, Phoenix Epidemiology and Clinical Research Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Phoenix, AZ, USA.
Aims/Hypothesis:
Obesity increases the risk of developing major diseases such as diabetes and cardiovascular disease. Adipose tissue, particularly adipocytes, may play a major role in the development of obesity and its comorbidities. The aim of this study was to characterise, in adipocytes from obese people, the most differentially expressed genes that might be relevant to the development of obesity.
Methods:
We carried out microarray gene profiling of isolated abdominal subcutaneous adipocytes from 20 non-obese (BMI 25+/-3 kg/m2) and 19 obese (BMI 55+/-8 kg/m2) non-diabetic Pima Indians using Affymetrix HG-U95 GeneChip arrays. After data analyses, we measured the transcript levels of selected genes based on their biological functions and chromosomal positions using quantitative real-time PCR.
Results:
The most differentially expressed genes in adipocytes of obese individuals consisted of 433 upregulated and 244 downregulated genes. Of these, 410 genes could be classified into 20 functional Gene Ontology categories. The analyses indicated that the inflammation/immune response category was over-represented, and that most inflammation-related genes were upregulated in adipocytes of obese subjects. Quantitative real-time PCR confirmed the transcriptional upregulation of representative inflammation-related genes (CCL2 and CCL3) encoding the chemokines monocyte chemoattractant protein-1 and macrophage inflammatory protein 1alpha. The differential expression levels of eight positional candidate genes, including inflammation-related THY1 and C1QTNF5, were also confirmed. These genes are located on chromosome 11q22-q24, a region with linkage to obesity in the Pima Indians.
Conclusions/Interpretation:
This study provides evidence supporting the active role of mature adipocytes in obesity-related inflammation. It also provides potential candidate genes for susceptibility to obesity.
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