Neurotrimin: a novel neural regulator of uterine leiomyoma development

Prior studies have demonstrated that estrogen exposure increases the myometrial expression of neurotrimin (NTM/HNT-1), a member of the immunoglobulin IgLON family of cell adhesion molecules that regulate neurite outgrowth and cellular adhesion. We hypothesize that NTM will be increased in leiomyoma tissue as estrogen-rich environments promote its growth. Additionally, we hypothesize that treatment with the selective progesterone receptor modulator (SPRM), ulipristal acetate (UPA), reduces NTM expression. Molecular analysis in a two-dimensional immortalized human myometrial and leiomyoma cell lines. RNA sequence (RNAseq) analysis was performed on placebo and matched UPA (10mg or 20mg) treated patient leiomyoma and myometrium samples from a prospective, randomized, placebo-controlled clinical trial. Results were performed in triplicate with qRT-PCR and results reported as mean ± SEM. For western blot analysis, calculations were done using software from Bio-Rad. Data is presented as fold difference between leiomyoma and myometrium (L: M) relative density units that was corrected for internal control using COX IV. Immunohistochemistry was also performed. Alterations of NTM mRNA transcripts by RNAseq in human uterine leiomyoma specimen demonstrated increased expression of transcripts in leiomyoma as compared to myometrium (5.22 fold, p=0.0003) and a reduced expression of NTM following UPA treatment (0.75 fold,p=0.134), when UPA treated leiomyomma is compared to placebo. In untreated patient samples, qRT-PCR showed an increased expression of NTM in leiomyoma compared to myometrium (1.95±0.05 fold, p=.037). However, qRT-PCR results on immortalized leiomyoma and myometrial cell lines demonstrated a reduced expression of NTM in leiomyoma (0.13±0.02 fold, p=0.005). Western blot analysis in immortalized leiomyoma and myometrial cell lines demonstrated an up-regulation of NTM protein expression (2.4± 0.04 fold, p=0.03). Following UPA treatment of an immortalized cell line (10-7M UPA for 72 hours), there is reduced expression of NTM in treated versus untreated leiomyoma (0.67± 0.04 fold, p=.0.003) change in protein expression. A significant increase in intensity of staining in leiomyoma as compared to myometrium was seen following immunohistochemistry, supporting an increased cytoplasmic expression of NTM in leiomyoma. NTM, a neural cell adhesion molecule highly expressed within brain tissues, is identified for the first time in leiomyoma tissue. NTM is increased in leiomyoma as compared to myometrium in both cell line and patient tissue and similarly down-regulated following treatment with UPA. Prior studies have shown NTM to be affected by estrogen regulation. We show here that its expression is affected by progesterone modulation as well.