RPL13A and EEF1A1 Are Suitable Reference Genes for qPCR during Adipocyte Differentiation of Vascular Stromal Cells from Patients with Different BMI and HOMA-IR.
Real-time or quantitative PCR (qPCR) is a useful technique that requires reliable reference genes for data normalization in gene expression analysis. Adipogenesis is among the biological processes suitable for this technique. The selection of adequate reference genes is essential for qPCR gene expression analysis of human Vascular Stromal Cells (hVSCs) during their differentiation into adipocytes. To the best of our knowledge, there are no studies validating reference genes for the analyses of visceral and subcutaneous adipose tissue hVSCs from subjects with different Body Mass Index (BMI) and Homeostatic Model Assessment of Insulin Resistance (HOMA-IR) index. The present study was undertaken to analyze this question. We first analyzed the stability of expression of five potential reference genes: CYC, GAPDH, RPL13A, EEF1A1, and 18S ribosomal RNA, during in vitro adipogenic differentiation, in samples from these types of patients. The expression of RPL13A and EEF1A1 was not affected by differentiation, thus being these genes the most stable candidates, while CYC, GAPDH, and 18S were not suitable for this sort of analysis. This work highlights that RPL13A and EEF1A1 are good candidates as reference genes for qPCR analysis of hVSCs differentiation into adipocytes from subjects with different BMI and HOMA-IR.
IBIMA, Universidad de Málaga, Campus Teatinos s/n, 29010, Málaga, España.
Adriana-Mariel
AM
Gentile
IBIMA, Universidad de Málaga, Campus Teatinos s/n, 29010, Málaga, España.
Said
S
Lhamyani
CIBER Pathophysiology of obesity and nutrition CB06/03, Carlos III Health Institute, Malaga, 29010, Spain, Laboratory of Biomedical Research, Virgen de la Victoria Clinical University Hospital, Málaga, 29010, Spain.
Leticia
L
Coín-Aragüez
CIBER Pathophysiology of obesity and nutrition CB06/03, Carlos III Health Institute, Malaga, 29010, Spain, Laboratory of Biomedical Research, Virgen de la Victoria Clinical University Hospital, Málaga, 29010, Spain.
Wilfredo
W
Oliva-Olivera
Biomedical Sciences Program, Health Sciences Department, College of Arts and Sciences, Qatar University, P.O. Box 2713, Doha, Qatar.
Hatem
H
Zayed
Unidad de Gestión Clínica de Alergia Intercentros, Hospital Universitario Virgen Macarena, Sevilla, Spain.
Antonio
A
Vega-Rioja
Unidad de Gestión Clínica de Alergia Intercentros, Hospital Universitario Virgen Macarena, Sevilla, Spain.
Javier
J
Monteseirin
Unidad de Gestion Clínica Intercentros de Endocrinología y Nutrición, Instituto de Investigación Biomédica de Málaga (IBIMA), Hospital Regional Universitario de Málaga/Universidad de Málaga, 29009, Málaga, Spain.|Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Málaga, Spain.
Silvana-Yanina
SY
Romero-Zerbo
CIBER Pathophysiology of obesity and nutrition CB06/03, Carlos III Health Institute, Malaga, 29010, Spain, Laboratory of Biomedical Research, Virgen de la Victoria Clinical University Hospital, Málaga, 29010, Spain.|Endocrinology Service, Virgen de la Victoria Clinical University Hospital, Malaga, 29010, Spain.
Francisco-José
FJ
Tinahones
Unidad de Gestion Clínica Intercentros de Endocrinología y Nutrición, Instituto de Investigación Biomédica de Málaga (IBIMA), Hospital Regional Universitario de Málaga/Universidad de Málaga, 29009, Málaga, Spain.|Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Málaga, Spain.
Francisco-Javier
FJ
Bermúdez-Silva
CIBER Pathophysiology of obesity and nutrition CB06/03, Carlos III Health Institute, Malaga, 29010, Spain, Laboratory of Biomedical Research, Virgen de la Victoria Clinical University Hospital, Málaga, 29010, Spain.
Rajaa
R
El Bekay
D017667:Adipocytes / Q000166:cytology / Q000378:metabolism*; D050156:Adipogenesis / Q000235:genetics; D000328:Adult; D000368:Aged; D015992:Body Mass Index; D002454:Cell Differentiation / Q000235:genetics*; D002478:Cells, Cultured; D020869:Gene Expression Profiling / Q000379:methods / Q000592:standards*; D006801:Humans; D007333:Insulin Resistance; D008875:Middle Aged; D020648:Peptide Elongation Factor 1 / Q000235:genetics*; D012015:Reference Standards; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods / Q000592:standards; D012269:Ribosomal Proteins / Q000235:genetics*; D017154:Stromal Cells / Q000166:cytology / Q000378:metabolism*; D055815:Young Adult
C094814:EEF1A1 protein, human; D020648:Peptide Elongation Factor 1; C402487:RPL13A protein, human; D012269:Ribosomal Proteins