Ontogenetic profile of the expression of thyroid hormone receptors in rat and human corpora cavernosa of the penis

Abstract

Introduction: In the last few years, various studies have underlined a correlation between thyroid function and male sexual function, hypothesizing a direct action of thyroid hormones on the penis.

Aim: To study the spatiotemporal distribution of mRNA for the thyroid hormone nuclear receptors (TR) alpha1, alpha2 and beta in the penis and smooth muscle cells (SMCs) of the corpora cavernosa of rats and humans during development.

Methods: We used several molecular biology techniques to study the TR expression in whole tissues or primary cultures from human and rodent penile tissues of different ages.

Main outcome measure: We measured our data by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) amplification, Northern blot and immunohistochemistry.

Results: We found that TRalpha1 and TRalpha2 are both expressed in the penis and in SMCs during ontogenesis without development-dependent changes. However, in the rodent model, TRbeta shows an increase from 3 to 6 days post natum (dpn) to 20 dpn, remaining high in adulthood. The same expression profile was observed in humans. While the expression of TRbeta is strictly regulated by development, TRalpha1 is the principal isoform present in corpora cavernosa, suggesting its importance in SMC function. These results have been confirmed by immunohistochemistry localization in SMCs and endothelial cells of the corpora cavernosa.

Conclusions: The presence of TRs in the penis provides the biological basis for the direct action of thyroid hormones on this organ. Given this evidence, physicians would be advised to investigate sexual function in men with thyroid disorders.

Figure 1

RT–PCR analysis of TR_α1, TR_α2, and TR_β expression in penises from rats of different ages. The PCR products are derived from total RNA from rat penises of 4, 20, 60, and 300 dpn rats using specific primers for TR_s and GAPDH. GAPDH mRNA amplification was performed to verify the integrity of the extracted total RNA. Densitometric evaluation of TR_s mRNA expression over the GAPDH housekeeping gene was obtained from band intensity of RT-PCR products. The densitometric analysis of TR_α1, TR_α2, and TR_β from three independent experiments ± SD (*P < 0.05) is shown.

Figure 2

Expression of mRNA from various TR isoforms in rat penis during development and adulthood. Representative Northern blot analysis of 20 µg of total RNA extracted from rat penises of 3–6, 20, and 60 dpn animals. Samples were separated in denaturing gel, blotted on nylon filter, and hybridized with the cDNA fragments corresponding to the indicated TR_s or GAPDH. Densitometric evaluation of TR_s mRNA expression over the GAPDH housekeeping gene was obtained from band intensity. The densitometric analysis of TR_α1, TR_α2, and TR_β from three independent experiments ± SD (*P < 0.05) is shown.

Figure 3

Localization of TR_α1 obtained by immunohistochemistry experiment of histological sections of adult (60 dpn) rat penis. A and C represent the negative control. B and D are stained with the anti-TR_α1 antibody (original magnification: 20×).

Figure 4

RT–PCR analysis of TR_α1, TR_α2, and TR_β expression in corpus cavernosum SMCs from rats of different ages. The products are derived from total RNA from rat corpus cavernosum SMCs from 3 (CC3) and 20 (CC20) dpn rats using specific primers for various TR_s and GAPDH. GAPDH mRNA amplification was performed to verify the integrity of the extracted total RNA. Densitometric evaluation of TR_s mRNA expression over the GAPDH housekeeping gene was obtained from band intensity of RT-PCR products. The histograms are calculated after normalization on GAPDH and on the basis of the amount of volume of PCR amplification loaded in the gel (1/5 of volume for TR_α1 and TR_α2, and 4/5 for the TR_β isoform). The densitometric analysis of TR_α1, TR_α2, and TR_β from three independent experiments ± SD (*P < 0.05) is shown.

Figure 5

(A) Expression of mRNA from various TR isoforms in adult human corpora cavernosa. Representative Northern blot analysis of 20 µg of total RNA extracted from the penis of a 65-year-old man. Samples were separated in denaturing gel, blotted on nylon filter, and hybridized with the cDNA fragments corresponding to the indicated TRs or GAPDH probe to assess their integrity and concentration. (B) RT–PCR analysis of TR_α1, TR_α2, and TR_β expression in human corpus cavernosum SMCs. The products are derived from total RNA from human fetal (hfPSMC) and adult (hAdutCC) corpus cavernosum SMCs using specific primers for various TR_s and GAPDH. GAPDH mRNA amplification was performed to verify the integrity of the extracted total RNA. Densitometric evaluation of TR_s mRNA expression over the GAPDH housekeeping gene was obtained from band intensity of RT-PCR products. The histograms are calculated on the basis of the amount of volume of PCR amplification loaded in the gel (1/5 of volume for TR_α1 and TR_α2, and 4/5 for the TR_β isoform). The densitometric analysis of TR_α1, TR_α2, and TR_β from three independent experiments ± SD (*P < 0.05) is shown.