Effect of growth hormone on fatty acid synthase gene expression in porcine adipose tissue cultures
We describe an efficient in vitro assay to test growth hormone effects on mRNA levels and fatty acid synthase (FAS, EC. 2.3.1.85) activity. Swine adipose tissue explants were long-term cultured in medium containing growth hormone and FAS mRNA levels and enzyme activity were measured. We quantified F...
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| Format: | Article |
| Language: | English |
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Sociedade Brasileira de Genética
2006-01-01
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| Series: | Genetics and Molecular Biology |
| Subjects: | |
| Online Access: | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1415-47572006000100032 |
| _version_ | 1818154210045198336 |
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| author | Andrea A.F.B.V. José Lúcia. E. Alvares Eduardo F. Delgado Luiz L. Coutinho Dante P.D. Lanna |
| author_facet | Andrea A.F.B.V. José Lúcia. E. Alvares Eduardo F. Delgado Luiz L. Coutinho Dante P.D. Lanna |
| author_sort | Andrea A.F.B.V. José |
| collection | DOAJ |
| description | We describe an efficient in vitro assay to test growth hormone effects on mRNA levels and fatty acid synthase (FAS, EC. 2.3.1.85) activity. Swine adipose tissue explants were long-term cultured in medium containing growth hormone and FAS mRNA levels and enzyme activity were measured. We quantified FAS transcripts by competitive reverse transcriptase PCR (RT-PCR) using total RNA from cultured adipose tissue explants and RT-PCR standard-curves were constructed using a cloned 307 bp segment of native FAS cDNA and a shorter fragment from which a 64 bp (competitor, 243 bp) internal sequence had been deleted. A known amount of competitor was added to each PCR as an internal control and µ-actin transcripts were also measured to correct for differences in total RNA extraction and reverse transcription efficiency. In cultures with added growth hormone FAS mRNA levels decreased 70% (p < 0.01) and FAS enzyme activity decreased 22% (p < 0.05). These in vitro growth hormone effects were consistent with those observed in vivo, showing that in vitro adipose tissue culture combined with RT-PCR is a useful and accurate tool for studying growth hormone modulation of adipose tissue metabolism. This technique allowed the diagnosis of lower levels of FAS mRNA in the presence of growth hormone and these low levels were associated with decreased FAS activity in the adipose tissue explants. |
| first_indexed | 2024-12-11T14:22:53Z |
| format | Article |
| id | doaj.art-74f7bda3181040e9ab02e472b5bc0601 |
| institution | Directory Open Access Journal |
| issn | 1415-4757 1678-4685 |
| language | English |
| last_indexed | 2024-12-11T14:22:53Z |
| publishDate | 2006-01-01 |
| publisher | Sociedade Brasileira de Genética |
| record_format | Article |
| series | Genetics and Molecular Biology |
| spelling | doaj.art-74f7bda3181040e9ab02e472b5bc06012022-12-22T01:02:50ZengSociedade Brasileira de GenéticaGenetics and Molecular Biology1415-47571678-46852006-01-0129118018610.1590/S1415-47572006000100032Effect of growth hormone on fatty acid synthase gene expression in porcine adipose tissue culturesAndrea A.F.B.V. JoséLúcia. E. AlvaresEduardo F. DelgadoLuiz L. CoutinhoDante P.D. LannaWe describe an efficient in vitro assay to test growth hormone effects on mRNA levels and fatty acid synthase (FAS, EC. 2.3.1.85) activity. Swine adipose tissue explants were long-term cultured in medium containing growth hormone and FAS mRNA levels and enzyme activity were measured. We quantified FAS transcripts by competitive reverse transcriptase PCR (RT-PCR) using total RNA from cultured adipose tissue explants and RT-PCR standard-curves were constructed using a cloned 307 bp segment of native FAS cDNA and a shorter fragment from which a 64 bp (competitor, 243 bp) internal sequence had been deleted. A known amount of competitor was added to each PCR as an internal control and µ-actin transcripts were also measured to correct for differences in total RNA extraction and reverse transcription efficiency. In cultures with added growth hormone FAS mRNA levels decreased 70% (p < 0.01) and FAS enzyme activity decreased 22% (p < 0.05). These in vitro growth hormone effects were consistent with those observed in vivo, showing that in vitro adipose tissue culture combined with RT-PCR is a useful and accurate tool for studying growth hormone modulation of adipose tissue metabolism. This technique allowed the diagnosis of lower levels of FAS mRNA in the presence of growth hormone and these low levels were associated with decreased FAS activity in the adipose tissue explants.http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1415-47572006000100032mRNA quantificationenzyme activityswinecultures of adipose tissue explantscompetitive RT-PCR |
| spellingShingle | Andrea A.F.B.V. José Lúcia. E. Alvares Eduardo F. Delgado Luiz L. Coutinho Dante P.D. Lanna Effect of growth hormone on fatty acid synthase gene expression in porcine adipose tissue cultures Genetics and Molecular Biology mRNA quantification enzyme activity swine cultures of adipose tissue explants competitive RT-PCR |
| title | Effect of growth hormone on fatty acid synthase gene expression in porcine adipose tissue cultures |
| title_full | Effect of growth hormone on fatty acid synthase gene expression in porcine adipose tissue cultures |
| title_fullStr | Effect of growth hormone on fatty acid synthase gene expression in porcine adipose tissue cultures |
| title_full_unstemmed | Effect of growth hormone on fatty acid synthase gene expression in porcine adipose tissue cultures |
| title_short | Effect of growth hormone on fatty acid synthase gene expression in porcine adipose tissue cultures |
| title_sort | effect of growth hormone on fatty acid synthase gene expression in porcine adipose tissue cultures |
| topic | mRNA quantification enzyme activity swine cultures of adipose tissue explants competitive RT-PCR |
| url | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1415-47572006000100032 |
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