Determination of estrogen receptor messenger ribonucleic acid (mRNA) and cytochrome P450 aromatase mRNA levels in adipocytes and adipose stromal cells by competitive polymerase chain reaction amplification.
Clinical evidence suggests that sex hormones affect adipose tissue metabolism and deposition. To investigate the biosynthesis and possible action of estrogen in adipose tissue, we report the use of competitive, specific polymerase chain reaction amplifications to determine levels of estrogen receptor (ER) messenger RNA (mRNA) and cytochrome P450 aromatase mRNA in adipocytes and adipose stromal cells. This extremely sensitive technique uses coamplification of a homologous animal species complementary RNA to control for differences in amplification efficiencies. The DNA amplification products are identified by Southern hybridization with species-specific radiolabeled oligonucleotide probes. Abdominal adipose tissue obtained from female patients during elective abdominoplasty was separated by collagenase digestion and centrifugation into floating adipocytes and pelleted adipose stromal cells. Our results demonstrate higher ER mRNA levels in adipocytes compared to adipose stromal cells, whereas cytochrome P450 aromatase mRNA levels are higher in adipose stromal cells compared to adipocytes. The finding of ER mRNA in adipose tissue suggests the presence of the ER in adipose tissue. In addition the inverse correlation of ER mRNA and cytochrome P450 aromatase mRNA levels in adipocytes and adipose stromal cells suggests a paracrine relationship whereby estrogen produced by adipose stromal cells affects adjacent adipocytes.
Price
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Greenville Hospital System/Clemson University, Biomedical Research Cooperative, South Carolina 29605.
O'Brien
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D017667:Adipocytes / Q000378:metabolism*; D000273:Adipose Tissue / Q000166:cytology*; D000818:Animals; D001141:Aromatase / Q000096:biosynthesis* / Q000235:genetics; D001483:Base Sequence; D015152:Blotting, Northern; D017931:DNA Primers / Q000737:chemistry; D005260:Female; D006801:Humans; D051379:Mice; D008875:Middle Aged; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes / Q000737:chemistry; D009841:Oligonucleotides; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D011960:Receptors, Estrogen / Q000096:biosynthesis* / Q000235:genetics; D017154:Stromal Cells / Q000378:metabolism*; D014158:Transcription, Genetic
D017931:DNA Primers; D015345:Oligonucleotide Probes; D009841:Oligonucleotides; D012333:RNA, Messenger; D011960:Receptors, Estrogen; D001141:Aromatase