O-95: Human leiomyomas minimize retinoic acid exposure by altering transport and metabolism gene expression

Abstract

Uterine leiomyomas may represent an alteration in myometrial cell differentiation. The leiomyocyte displays a phenotypic pattern of disrupted and excessive extracellular matrix production not seen in the normal myocyte. Retinoic acid plays a central role in cellular differentiation, and alterations in retinoic acid exposure could alter the cellular phenotype from a myocyte to a leiomyocyte. In this study, we used human leiomyomata and patient-matched myometrium to determine whether there were alterations in retinoic acid exposure in uterine leiomyomata. Laboratory study. After IRB approval, we obtained fibroid:myometrium pairs from five patients undergoing abdominal hysterectomy. Methodology included microarray analysis, real time reverse transcriptase-polymerase chain reaction (RT-PCR), western blot analysis, immunohistochemistry, and high pressure liquid chromatography (HPLC). Statistical evaluation performed using the Wilcoxon Rank-Sum test, with significance defined as α<0.05. Microarray analysis demonstrated under-expression of multiple steps in the production of retinoic acid, and real time RT-PCR confirmed decreased mRNA levels of specific components in the retinoic acid production process, including cellular retinol binding protein (-2.62 +/- 0.04 fold), cellular retinoic acid binding protein (-2.42 +/- 0.20 fold), alcohol dehydrogenase (-3.97 +/- 0.03 fold) and aldehyde dehydrogenase (-3.10 +/- 0.07 fold). In addition, there was over-expression of cP450 retinoic acid metabolizing protein (+5.40 +/- 0.53 fold), which can decrease the concentration of available retinoic acid. All findings were statistically significant. Western blot analysis and immunohistochemistry confirmed that mRNA alterations translated to altered protein expression. In order to confirm that retinoic acid concentrations were altered in human leiomyomata, we determined in situ concentrations of retinoic acid, as well as concentrations after spiking the tissue with retinoic acid. These HPLC studies demonstrated decreased retinoic acid concentrations in leiomyoma relative to patient-matched myometrium, and more rapid metabolism of exogenously added retinoic acid (both for all trans retinoic acid and 9-cis-retinoic acid). Molecular alterations in the retinoic acid pathway of human leiomyomas result in decreased exposure to retinoic acid. Minimizing retinoic acid exposure diminishes the differentiation influence of retinoic acid, potentially influencing the altered differentiation pattern of the leiomyocyte. Furthermore, the distinct alterations in retinoic acid metabolism may provide insight into the origins of human leiomyoma development, and form the basis for novel therapeutic interventions for symptomatic leiomyoma disease.