HOXC10, HOXC11, HOXD10, and HOXD11 Expression Is Not Modulated by Sex Steroids in Primary Human Endometrial Stromal Cells
To test whether HOXC10, HOXC11, HOXD10, and HOXD11 expression is regulated by sex steroids, the signal corresponding to each HOX transcript was measured in primary human stromal cells after treatment with estrogen or progesterone. Endometrial samples from proliferative phase of the menstrual cycle were used as a source of primary cultures of stromal cells. Cells were grown to confluence in charcoal-stripped, phenol red-free media and serum-starved for 24 h before a 24-h treatment with physiologic concentrations of 17-p estradiol, progesterone, or both. RNA was extracted and used for RT-PCR. HOXC10, HOXC11, HOXD10, and HOXD11 mRNA was idetified in the endometrial stromal cells (Fig. 4).
However, treatment with sex steroids did not significantly alter the level of transcript of any of these four genes (Fig. 3). HOXA10 transcripts are increased with sex-steroid treatment as previously described and shown as a positive control. Each experiment was repeated five times. Average densitometry readings were used to quantify and normalize the results; 17-p estradiol, progesterone, or both combined did not significantly alter HOXC10, HOXC11, HOXD10, and HOXD11 expression in human stromal cells (data not shown).
FIG. 3. Densitometric analysis of HOXC10, HOXC11, HOXD10, and HOXD11 signal in 10 samples normalized to G3PDH. In the secretory phase, expression of (A) HOXC10, (B) HOXC11, and (D) HOXD11 decreased to 4%, and (C) HOXD10 decreased to 25% of the proliferative phase expression (P < 0.001, P < 0.001, P < 0.02, P < 0.01, respectively). Values represent the mean ± SEM (n = 10). Each experiment was repeated three times.
FIG. 4. HOXC10, HOXC11, HOXD10, and HOXD11 expression in stromal cells in response to sex steroids as demonstrated by RT-PCR. Cells that were demonstrated to be estrogen- and progesterone receptor-positive expressed (A) HOXC10, (B) HOXC11, (C) HOXD10, and (D) HOXD11. Treatment with 1 X 10_8 M 17p-estradiol (E), 1 X 10_6 M progesterone (P), or both hormones (E + P) resulted in no statistically significant difference in HOXC10, HOXC11, HOXD10, and HOXD11 signal in between treatment groups. Lane L: DNA ladder (100 bp). The response of HOXA10 to E + P is shown as a control. Each experiment was repeated five times.