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Quantitative Analysis of Growth Hormone (GH) Pre-mRNA Expression in Cultured Rat Anterior Pituitary Cells by an Intron-Specific and Competitive PCR Method

First Department of Internal Medicine, Kagawa Medical University, 1750-1 Ikenobe, Miki-cho, Kita-gun, Kagawa, 761-07, Japan

Address all correspondence and requests for reprints to: Dr. M. Sato, Kagawa Medical University, First Department of Internal Medicine, 1750-1 Ikenobe, Miki-cho, Kagawa, Kita-gun 761-07, Japan.

We have developed a novel method of quantifying growth hormone (GH) pre-mRNA expression in anterior pituitary cells. DNA-free total RNA extracted from cultured rat anterior pituitary cells was reverse transcribed (RT) to cDNA, and RT products were subsequently quantitated by competitive PCR using intron-specific primers of rat GH gene. After 6-h of incubation in treated cells, dexamethasone (Dex) and triiodo-L-thyronine (T3) significantly increased GH pre-mRNA levels (3.2- and 2.2-fold compared to nontreated cells, respectively). However, Northern blot analysis did not detect significant changes in GH mRNA levels. After 24-h incubation with Dex and T3, significant increases in GH mRNA levels were detected on Northern blots, but GH pre-mRNA levels did not differ between treated and non-treated cells. These findings suggest that both Dex and T3 treatments rapidly increase GH pre-mRNA levels in normal somatotropes. This method has high sensitivity and widespread application to the analysis of pre-mRNAs of target genes.