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NAV3, a Tumor Suppressor Gene, Is Decreased in Uterine Leiomyoma Tissue and Cells.

ABSTRACT: NAV 3 is a tumor suppressor of unknown function in leiomyomas. The objective of this study is to assess NAV3 expression and its potential role in human uterine leiomyomas. NAV3 protein expression was examined in patient leiomyoma and patient-matched myometrial tissue samples by Western blot and immunohistochemistry. NAV3 mRNA and protein expression was assessed in leuprolide acetate- and cetrorelix-treated cell line leiomyoma samples. RNAseq analysis of placebo-treated leiomyoma compared with myometrium demonstrated the presence of transcripts encoding for several neuronal proteins. For NAV3, RNA sequence analysis demonstrated decreased expression in leiomyoma as compared with myometrium (0.86?±?0.03 fold). Presence of NAV3 mRNA was also decreased in leiomyoma surgical samples (0.43 fold ±?0.05, p?=?0.026) compared with patient-matched myometrium. Confirmatory qRT-PCR results on immortalized leiomyoma and myometrial cell lines similarly demonstrated a decrease in expression of NAV3 in leiomyomas (0.28?±?0.02, p?=?0.00075). Immunohistochemical analysis demonstrated a significant decrease in NAV 3 protein in leiomyomas (H-score 154.7?±?6.2) as compared with myometrium (H-score; 312.5?±?14.7, p?

SUBMITTER: Aly JM

PROVIDER: S-EPMC7539815 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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NAV3, a Tumor Suppressor Gene, Is Decreased in Uterine Leiomyoma Tissue and Cells.

Aly Jasmine M JM Lewis Terrence D TD Parikh Toral T Britten Joy J Malik Minnie M Catherino William H WH

Reproductive sciences (Thousand Oaks, Calif.) 20200101 3

NAV 3 is a tumor suppressor of unknown function in leiomyomas. The objective of this study is to assess NAV3 expression and its potential role in human uterine leiomyomas. NAV3 protein expression was examined in patient leiomyoma and patient-matched myometrial tissue samples by Western blot and immunohistochemistry. NAV3 mRNA and protein expression was assessed in leuprolide acetate- and cetrorelix-treated cell line leiomyoma samples. RNAseq analysis of placebo-treated leiomyoma compared with my ...[more]

PMID: 32046415

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Project description:BACKGROUND:Uterine leiomyomas, the most common tumors of the female reproductive system, are characterized by excessive deposition of disordered stiff extracellular matrix and fundamental alteration in the mechanical signaling pathways. Specifically, these alterations affect the normal dynamic state of responsiveness to mechanical cues in the extracellular environment. These mechanical cues are converted through integrins, cell membrane receptors, to biochemical signals including cytoskeletal signaling pathways to maintain mechanical homeostasis. Leiomyoma cells overexpress β1 integrin and other downstream mechanical signaling proteins. We previously reported that simvastatin, an antihyperlipidemic drug, has antileiomyoma effects through cellular, animal model, and epidemiologic studies. OBJECTIVE:This study aimed to examine the hypothesis that simvastatin might influence altered mechanotransduction in leiomyoma cells. STUDY DESIGN:This is a laboratory-based experimental study. Primary leiomyoma cells were isolated from 5 patients who underwent hysterectomy at the Department of Gynecology and Obstetrics of the Johns Hopkins University Hospital. Primary and immortalized human uterine leiomyoma cells were treated with simvastatin at increasing concentrations (0.001, 0.01, 0.1, and 1 μM, or control) for 48 hours. Protein and mRNA levels of β1 integrin and extracellular matrix components involved in mechanical signaling were quantified by quantitative real-time polymerase chain reaction, western blotting, and immunofluorescence. In addition, we examined the effect of simvastatin on the activity of Ras homolog family member A using pull-down assay and gel contraction. RESULTS:We found that simvastatin significantly reduced the protein expression of β1 integrin by 44% and type I collagen by 60% compared with untreated leiomyoma cells. Simvastatin-treated cells reduced phosphorylation of focal adhesion kinase down to 26%-60% of control, whereas it increased total focal adhesion kinase protein expression. Using a Ras homolog family member A pull-down activation assay, we observed reduced levels of active Ras homolog family member A in simvastatin-treated cells by 45%-85% compared with control. Consistent with impaired Ras homolog family member A activation, simvastatin treatment reduced tumor gel contraction where gel area was 122%-153% larger than control. Furthermore, simvastatin treatment led to reduced levels of mechanical signaling proteins involved in β1 integrin downstream signaling, such as A-kinase anchor protein 13, Rho-associated protein kinase 1, myosin light-chain kinase, and cyclin D1. CONCLUSION:The results of this study suggest a possible therapeutic role of simvastatin in restoring the altered state of mechanotransduction signaling in leiomyoma. Collectively, these findings are aligned with previous epidemiologic studies and other reports and support the need for clinical trials.

Project description:BackgroundUterine leiomyoma is the most common benign tumor in reproductive-age women. Each leiomyoma is thought to be a benign monoclonal tumor arising from a single transformed myometrial smooth muscle cell; however, it is not known what leiomyoma cell type is responsible for tumor growth. Thus, we tested the hypothesis that a distinct stem/reservoir cell-enriched population, designated as the leiomyoma-derived side population (LMSP), is responsible for cell proliferation and tumor growth.Principal findingsLMSP comprised approximately 1% of all leiomyoma and 2% of all myometrium-derived cells. All LMSP and leiomyoma-derived main population (LMMP) but none of the side or main population cells isolated from adjacent myometrium carried a mediator complex subunit 12 mutation, a genetic marker of neoplastic transformation. Messenger RNA levels for estrogen receptor-α, progesterone receptor and smooth muscle cell markers were barely detectable and significantly lower in the LMSP compared with the LMMP. LMSP alone did not attach or survive in monolayer culture in the presence or absence of estradiol and progestin, whereas LMMP readily grew under these conditions. LMSP did attach and survive when directly mixed with unsorted myometrial cells in monolayer culture. After resorting and reculturing, LMSP gained full potential of proliferation. Intriguingly, xenografts comprised of LMSP and unsorted myometrial smooth muscle cells grew into relatively large tumors (3.67 ± 1.07 mm(3)), whereas xenografts comprised of LMMP and unsorted myometrial smooth muscle cells produced smaller tumors (0.54 ± 0.20 mm(3), p<0.05, n = 10 paired patient samples). LMSP xenografts displayed significantly higher proliferative activity compared with LMMP xenografts (p<0.05).ConclusionsOur data suggest that LMSP, which have stem/reservoir cell characteristics, are necessary for in vivo growth of leiomyoma xenograft tumors. Lower estrogen and progesterone receptor levels in LMSP suggests an indirect paracrine effect of steroid hormones on stem cells via the mature neighboring cells.

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NAV3, a Tumor Suppressor Gene, Is Decreased in Uterine Leiomyoma Tissue and Cells.

Publications

NAV3, a Tumor Suppressor Gene, Is Decreased in Uterine Leiomyoma Tissue and Cells.

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