Activation of the estrogen receptor contributes to the progression of pulmonary lymphangioleiomyomatosis via matrix metalloproteinase-induced cell invasiveness

Context: The role of estrogens in the pathogenesis of lymphangioleiomyomatosis (LAM), an aggressive and destructive, eventually fatal lung disease of women, is poorly understood. Objective: The study was conducted to test the hypothesis that the lung disease in LAM is estrogen mediated and to determine whether estrogens contribute to the invasiveness of LAM. Design: In vitro cell culture of spindle-shaped LAM cells(LAMD-SM) were isolated and propagated from affected lungs. Estrogen receptor (ER)-alpha and ER beta analyses were conducted by RT-PCR. ER alpha and ER beta, tissue inhibitor of metalloproteinase-2, and matrix metalloproteinases (MMP)-2 had Western blot analysis for protein assessment. Activity assays were performed for MT1-MMP, MMP-2, and tissue inhibitor of metalloproteinase-2. Assessment of MMP-2 promoter function was done via transfection assays. Cell invasion chambers were used to determine and quantitate cell invasiveness. Setting: The study was conducted at an academic medical center. Patients: Tissue and cells were obtained from patients as outlined in approved institution review board protocol (97/007). Intervention: LAMD-SM cells were treated with a specific MMP-2 antibody or a nonspecific inhibitor, doxycycline. Main Outcome Measures: Activity of MMP-2 and invasiveness of LAMD-SM cells were measured. Results: LAMD-SM cells express functional ERs (ER alpha and ER beta), which undergo rapid intracellular turnover in their unbound state. 17 beta-Estradiol (E-2) enhances the transcriptional ER activity. E-2-induced ER activation increases synthesisandactivity of MMP-2 through posttranscriptional mechanisms in LAMD-SM. The E-2/ER-mediated increase of MMP-2 promotes LAMD-SM invasiveness, in assays in vitro, which can be inhibited by specific antibodies against MMP-2 or doxycycline, an inhibitor of MMPs. Conclusion: The invasion and destruction of lung parenchyma in LAM is, at least partially, an estrogen-MMP-driven process, which has major implications for therapeutic interventions.