Novel method to characterize primary cultures of leiomyoma and myometrium with the use of confirmatory biomarker gene arrays

Objective: To develop a rapid biomarker method for confirming that leiomyoma and myometrium primary cultures maintain the molecular phenotype of the progenitor tissues. Design: Confirmation of primary cultures from leiomyoma and myometrium tissues. Setting: University hospital. Patient(s): Women undergoing hysterectomy for symptomatic leiomyomas. Intervention(s): Primary cell cultures, reverse-transcriptase polymerase chain reaction (RT-PCR), microarray, real time RT-PCR, and immunofluorescence. Main Outcome Measure: Relative messenger RNA and protein expression in leiomyoma and myometrial cell cultures. Result(s): We developed primary cell cultures from human leiomyoma and patient-matched myometrium obtained from hysterectomy specimens. In the primary cultures, we confirmed the presence of smooth muscle-specific α-actin as well as filamentous actin. Based on microarray analysis, we expected and confirmed, in the progenitor tissue and derived primary cultures, an overexpression of versican (8.31 fold ± 2.2 SEM and 4.3 fold ± 1.01 SEM, respectively), transforming growth factor beta-3 (5.66 fold ± 0.82 SEM and 4.92 fold ± 0.58 SEM, respectively), and cytochrome P450-26A1 (6.76 fold ± 0.80 SEM and 6.17 fold ± 2.02 SEM, respectively), and an underexpression of dermatopontin (-5.6 fold ± 1.82 SEM and -3.41 ± 1.20 SEM, respectively). Conclusion(s): Primary cell cultures offer a reliable in vitro model system for leiomyoma disease, if confirmed. Analysis of a gene array that distinguishes between myometrium and leiomyoma molecular phenotypes offers a rapid and reliable confirmation method, and provides confidence that in vitro findings resemble in vivo disease.