Selective progesterone receptor modulator asoprisnil down-regulates collagen synthesis in cultured human uterine leiomyoma cells through up-regulating extracellular matrix metalloproteinase inducer.

BACKGROUND: A recent clinical trial demonstrated that selective progesterone receptor modulator asoprisnil is effective in reducing uterine leiomyoma volume. We investigated the effects of asoprisnil in vitro on the expression of the extracellular matrix (ECM)-remodeling enzymes and collagens in cultured leiomyoma and matching normal myometrial cells. METHODS: The expression of extracellular matrix metalloproteinase inducer (EMMPRIN), matrix metalloproteinases (MMPs), tissue inhibitors of MMP (TIMPs) and collagens were assessed by western blot analysis. RESULTS: Untreated cultured leiomyoma cells had significantly lower EMMPRIN (P < 0.05), MMP-1 (P < 0.05) and membrane type 1-MMP (MT1-MMP) (P < 0.01) protein contents, but significantly higher TIMP-1 (P < 0.05), TIMP-2 (P < 0.01), type I (P < 0.05) and type III (P < 0.01) collagen protein contents compared with untreated cultured myometrial cells. Treatment with asoprisnil at concentrations > or =10(-7) M for 48 h significantly (P < 0.05) increased EMMPRIN, MMP-1 and MT1-MMP protein contents, and decreased TIMP-1 (P < 0.05), TIMP-2 (P < 0.01), type I (P < 0.01) and type III (P < 0.05 at 10(-7) M; P < 0.01 at 10(-6) M) collagen protein contents in cultured leiomyoma cells compared with control cultures. However, asoprisnil treatment did not affect the protein contents of ECM-remodeling enzymes and collagens in cultured myometrial cells. CONCLUSIONS: These results suggest that asoprisnil may reduce collagen deposit in the ECM of cultured leiomyoma cells through decreasing collagen synthesis and enhancing the expression of EMMPRIN, MMPs and TIMPs without comparable effects on cultured myometrial cells.

A novel selective progesterone receptor modulator asoprisnil activates tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated signaling pathway in cultured human uterine leiomyoma cells in the absence of comparable effects on myometrial cells.

CONTEXT: We previously demonstrated that asoprisnil, a selective progesterone receptor modulator, induces apoptosis of cultured uterine leiomyoma cells. This study was conducted to evaluate whether asoprisnil activates TNF-related apoptosis-inducing ligand (TRAIL)-mediated apoptotic pathway in cultured uterine leiomyoma and matching myometrial cells. OBJECTIVE AND METHODS: After subculture in phenol red-free DMEM supplemented with 10% fetal bovine serum for 120 h, cultured cells were stepped down to serum-free conditions for 24 h in the absence or presence of graded concentrations of asoprisnil. The levels of TRAIL signaling molecules and cellular inhibitors of apoptosis protein were assessed by Western blot analysis. RESULTS: TRAIL contents in untreated cultured leiomyoma cells were significantly (P < 0.01) lower compared with those in untreated cultured myometrial cells. There was no difference in death receptor (DR)4 and DR5 contents between the two types of cells. Asoprisnil treatment significantly (P < 0.05) increased TRAIL, DR4, and DR5 contents in cultured leiomyoma cells in a dose-dependent manner with a cleavage of caspase-8, -7, and -3, and decreased X-linked chromosome-linked inhibitor of apoptosis protein contents. In cultured myometrial cells, however, asoprisnil treatment did not affect either TRAIL signaling molecule or cellular inhibitors of apoptosis protein contents. The concomitant treatment with 100 ng/ml P4 significantly (P < 0.05) reversed asoprisnil-induced increase in DR4 and cleaved poly(adenosine 5'-diphosphate-ribose) polymerase contents in cultured leiomyoma cells. CONCLUSIONS: These results suggest that asoprisnil induces apoptosis of cultured leiomyoma cells by activating TRAIL-mediated apoptotic pathway and down-regulating X-linked chromosome-linked inhibitor of apoptosis protein levels in the absence of comparable effects on myometrial cells.

Comparative effects of SPRM asoprisnil (J867) on proliferation, apoptosis, and the expression of growth factors in cultured uterine leiomyoma cells and normal myometrial cells.

Progesterone plays a pivotal role in controlling uterine leiomyoma growth. The authors review studies they conducted to evaluate the comparative effects of asoprisnil on proliferation, apoptosis, and growth factor expression in cultured leiomyoma and normal myometrial cells. Treatment with asoprisnil decreased the proliferating cell nuclear antigen-positive rate and the number of viable cells and increased the terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick end labeling- positive rate in cultured leiomyoma cells in a dose-dependent manner ( P < .05). Similarly, asoprisnil decreased Bcl-2 expression and increased cleaved caspase-3 and cleaved poly(adenosine 5'-diphosphate-ribose) polymerase in leiomyoma cells but not in normal myometrial cells. Similarly, asoprisnil decreased epidermal growth factor (EGF), insulin-like growth factor-I (IGF-I), and transforming growth factor (TGF) beta mRNA and protein expression, as well as EGF receptor, IGF-IR alpha, and TGF RII protein expression in leiomyoma cells but not in cultured normal myometrial cells. These results suggest that asoprisnil selectively inhibits proliferation by downregulating the growth factors and their receptor expression and induces apoptosis in leiomyoma cells without affecting proliferation and apoptosis in normal myometrial cells.

A novel selective progesterone receptor modulator asoprisnil (J867) down-regulates the expression of EGF, IGF-I, TGFbeta3 and their receptors in cultured uterine leiomyoma cells.

BACKGROUND: This study was conducted to evaluate the effects of a novel selective progesterone receptor modulator (SPRM) asoprisnil on the expression of growth factors and their receptors and on growth factor-induced proliferation of cultured uterine leiomyoma and matching myometrial cells. METHODS: The expression of epidermal growth factor (EGF), insulin-like growth factor-I (IGF-I) and transforming growth factor (TGFbeta3) was assessed by immunocytochemistry and semi-quantitative RT-PCR. The expression of phosphorylated EGF receptor (p-EGFR), IGF-I receptor alpha subunit (IGF-IRalpha) and phosphorylated TGFbeta receptor type II (p-TGFbeta RII) was assessed by Western blot analysis. Cell proliferation was assessed by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assay. RESULTS: Treatment with 10(-7) M asoprisnil decreased EGF, IGF-I and TGFbeta3 mRNA and protein expression as well as p-EGFR, IGF-IRalpha and p-TGFbeta RII protein expression in leiomyoma cells cultured for 72 h. EGF (100 ng/ml), IGF-I (100 ng/ml) and TGFbeta3 (10 ng/ml) increased the number of viable leiomyoma cells cultured for 72 h, whereas the concomitant treatment with 10(-7) M asoprisnil antagonized the growth factor-induced increase in leiomyoma cell proliferation. In cultured myometrial cells, however, asoprisnil affected neither the growth factor and their receptor expression nor the cell proliferation. CONCLUSION: Asoprisnil inhibits the expression of EGF, IGF-I, TGFbeta3 and their receptors in cultured leiomyoma cells without affecting their expressions in myometrial cells.

A novel selective progesterone receptor modulator asoprisnil (J867) inhibits proliferation and induces apoptosis in cultured human uterine leiomyoma cells in the absence of comparable effects on myometrial cells.

CONTEXT: Asoprisnil, a selective progesterone (P4) receptor (PR) modulator (SPRM) with mixed P4 agonist/antagonist activities, reduces uterine leiomyoma volume in a dose-dependent manner in the presence of follicular phase estrogen concentrations. The evidence from clinical studies suggests that asoprisnil may directly target the uterine leiomyomata. OBJECTIVE AND METHODS: The present study evaluated the effects of asoprisnil on cell proliferation, the expression of apoptosis-related proteins, and apoptosis in cultured human uterine leiomyoma cells and matched normal myometrial cells. PR-A and PR-B expression in the two types of cells was comparatively evaluated. Cell proliferation, proliferating cell nuclear antigen (PCNA)-positive rate, and TUNEL-positive rate were assessed by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assay, immunocytochemistry, and terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick end labeling (TUNEL) assay, respectively. The expression of apoptosis-related proteins and PR was assessed by Western blot analysis. RESULTS: Compared with untreated cultures, asoprisnil decreased the number of viable cultured cells, the PCNA-positive rate, and PCNA protein expression in cultured leiomyoma cells. Asoprisnil increased the TUNEL-positive rate, cleaved caspase-3, and cleaved poly(adenosine 5'-diphosphate-ribose) polymerase expression and decreased Bcl-2 protein expression in cultured leiomyoma cells. These effects were dose and time dependent. In cultured myometrial cells, however, asoprisnil did not affect cell proliferation and apoptosis. PR-B expression was elevated in cultured leiomyoma cells compared with cultured myometrial cells, whereas no differences in PR-A expression were noted between the two cell types. CONCLUSIONS: These results show that asoprisnil inhibits proliferation and induces apoptosis in cultured uterine leiomyoma cells in the absence of comparable effects on cultured normal myometrial cells, suggesting a cell type-specific effect.