Kenji Tsuiji

Inhibitory effect of curcumin on uterine leiomyoma cell proliferation

Objective. Uterine leiomyomas are the most common gynaecological benign tumour and greatly affect reproductive health and wellbeing. They are the predominant indication for hysterectomy in premenopausal women. Curcumin, a well-known component of turmeric, has been reported to prevent various diseases such as cancer, diabetes and obesity. Previous study reported that curcumin represses the proliferation of several tumour cells. However, there has not been a precise characterisation of the curcumin-induced inhibition of uterine leiomyoma cells. In this study, we investigated the inhibitory effect of curcumin on leiomyoma cells proliferation. Study design. Eker rat-derived uterine leiomyoma cell lines (ELT-3 cells) were used. Cell proliferation was assessed by counting the number of cells and MTS assay. The activation of peroxisome proliferator-activated receptor-gamma (PPARγ) was evaluated by luciferase assay. Results. We found that curcumin significantly inhibited ELT-3 cell proliferation. PPARγ was expressed in ELT-3 cells and curcumin acted as a PPARγ ligand. This inhibitory effect of curcumin was attenuated by the treatment of cells with PPARγ antagonist. Conclusion. These experimental findings in vitro show that the inhibitory effect of curcumin on ELT-3 cell proliferation occurs through the activation of PPARγ. Curcumin may be useful as an alternative therapy for uterine leiomyoma.

Curcumin induces cross-regulation between autophagy and apoptosis in uterine leiomyosarcoma cells.

Objective Uterine leiomyosarcoma (LMS) has an unfavorable response to standard chemotherapy. A natural occurring compound, curcumin, has been shown to have inhibitory effects on cancers. We previously demonstrated that curcumin reduced uterine LMS cell proliferation by targeting the AKT-mTOR pathway and activating apoptosis. To further explore the anticancer effect of curcumin, we investigated the efficacy of curcumin on autophagy in LMS cells. Methods Cell proliferation in human uterine LMS cell lines, SKN and SK-UT-1, was assessed after exposure to rapamycin or curcumin. Autophagy was detected by Western blotting for light chain 3 and sequestosome 1 (SQSTM1/p62) expression. Apoptosis was confirmed by Western blotting for cleaved poly (ADP-ribose) polymerase (PARP). Results Both rapamycin and curcumin potently inhibited SKN and SK-UT-1 cell proliferation in a dose-dependent manner. Curcumin induced autophagy and apoptosis in SKN and SK-UT-1 cells, whereas rapamycin, a specific mTOR inhibitor, did not. Curcumin increased extracellular signal-regulated kinase 1/2 activity in both SKN and SK-UT-1 cells, whereas PD98059, an MEK1 inhibitor, inhibited both the extracellular signal-regulated kinase 1/2 pathway and curcumin-induced autophagy. Conclusions These experimental findings suggest that curcumin is a potent inhibitor of cell proliferation in uterine LMS and provide new insights about ongoing signaling events leading to the possible development of a new therapeutic agent.

Epigallocatechin-3-gallate potentiates curcumin’s ability to suppress uterine leiomyosarcoma cell growth and induce apoptosis

BackgroundUterine leiomyosarcoma (LMS) has an unfavorable response to standard chemotherapeutic regimens. Two natural occurring compounds, curcumin and epigallocatechin gallate (EGCG), are reported to have anti-cancer activity. We previously reported that curcumin reduced uterine LMS cell proliferation by targeting the AKT–mTOR pathway. However, challenges remain in overcoming curcumin’s low bioavailability.MethodsThe human LMS cell line SKN was used. The effect of EGCG, curcumin or their combination on cell growth was detected by MTS assay. Their effect on AKT, mTOR, and S6 was detected by Western blotting. The induction of apoptosis was determined by Western blotting using cleaved-PARP specific antibody, caspase-3 activity and TUNEL assay. Intracellular curcumin level was determined by a spectrophotometric method. Antibody against EGCG cell surface receptor, 67-kDa laminin receptor (67LR), was used to investigate the role of the receptor in curcumin’s increased potency by EGCG.ResultsIn this study, we showed that the combination of EGCG and curcumin significantly reduced SKN cell proliferation more than either drug alone. The combination inhibited AKT, mTOR, and S6 phosphorylation, and induced apoptosis at a much lower curcumin concentration than previously reported. EGCG enhanced the incorporation of curcumin. 67LR antibody partially rescued cell proliferation suppression by the combination treatment, but was not involved in the EGCG-enhanced intracellular incorporation of curcumin.ConclusionsEGCG significantly lowered the concentration of curcumin required to inhibit the AKT–mTOR pathway, reduce cell proliferation and induce apoptosis in uterine LMS cells by enhancing intracellular incorporation of curcumin, but the process was independent of 67LR.

The anti-diabetic drug metformin inhibits vascular endothelial growth factor expression via the mammalian target of rapamycin complex 1/hypoxia-inducible factor-1α signaling pathway in ELT-3 cells.

The aim of this study was to elucidate whether metformin can regulate the expression of vascular endothelial growth factor (VEGF) in rat-derived uterine leiomyoma cells (ELT-3 cells). In vitro studies were conducted using ELT-3 cells. Under normoxic conditions, metformin suppressed VEGF protein levels in the supernatant and cells in a dose-dependent manner. In hypoxia-mimicking conditions, VEGF and hypoxia-inducible factor-1α (HIF-1α) proteins were both highly expressed and were suppressed by the metformin treatment. Metformin did not affect HIF-1α mRNA levels, which indicated that its effects occurred at the post-translational level. Metformin inhibited mammalian target of rapamycin complex 1 (mTORC1) activity by phosphorylating the mTORC1 component raptor. This study revealed the anti-angiogenic activity of metformin in ELT-3 cells by suppressing the expression of VEGF via the mTORC1/HIF-1α pathway. These results indicate that metformin may represent an effective alternative in the future treatment of uterine leiomyomas.

The antidiabetic drug metformin inhibits uterine leiomyoma cell proliferation via an AMP-activated protein kinase signaling pathway

Uterine leiomyomas are the most common gynecological benign tumors and greatly affect reproductive health and wellbeing. Metformin is the most widely used antidiabetic drug in the world, and there is increasing evidence of a potential efficacy of this agent as an anticancer drug. In order to understand metformin’s anti-tumorigenic potential better, in this study, we investigated the inhibitory effect of metformin and expression of key targets of metformin cell signaling in leiomyoma cells. Cell proliferation was assessed after exposure to metformin. Apoptosis was assessed by western blotting for cleaved-PARP and TUNEL staining. The expressions of phosphorylated AMPK and phosphorylated S6 were determined by western blotting. Metformin potently inhibited ELT-3 cell proliferation in a dose-dependent manner. Western blotting analysis demonstrated that metformin induced phosphorylation of AMPK and the inhibitory effect was attenuated with AMPK inhibitor, compound C. In parallel, treatment with metformin decreased phosphorylation of S6 protein. These experimental findings show that metformin is a potent inhibitor of cell proliferation in leiomyoma cells. This effect is mediated by AMPK activation and subsequent inhibition of the mTOR pathway. Thus, this study provides a possible mechanism of the action of metformin in the inhibition of leiomyoma cell growth.

Antiproliferative effect of adiponectin on rat uterine leiomyoma ELT-3 cells

Objective. Although fibroids greatly affect reproductive health, the pathophysiology and epidemiology are not well known. Recently, we have reported the relationship between uterine leiomyoma and metabolic syndrome. Many studies have indicated that reductions in plasma adiponectin levels play major roles in the development of metabolic syndrome. In this study, we investigated the significant repressive effect of adiponectin on rat uterine leiomyoma ELT-3 cells proliferation. Study design. Expression of adiponectin receptor 1 and receptor 2 was evaluated by RT-PCR and Western blot analysis. Cell proliferation was assessed by the MTS assay and cell counting. Apoptosis was evaluated by Hoechst staining and cleaved poly (ADP-ribose) polymerase (PARP). Results. Adiponectin receptor 1 and receptor 2 were expressed in ELT-3 cells. Adiponectin repressed rat uterine leiomyoma ELT-3 cells cell proliferation without inducing apoptosis. Conclusion. The repression of adiponectin on leiomyoma cell proliferation in the rat may explain a crucial role of adiponectin in the association of metabolic syndrome with uterine leiomyoma.

Aldosterone stimulates the proliferation of uterine leiomyoma cells.

Objective. Although uterine leiomyomas are the most common gynaecological benign tumour and greatly affect reproductive health and well being, the pathophysiology and epidemiology of uterine leiomyomas are not well known. Elevated blood pressure has an independent, positive association with risk for clinically detected uterine leiomyomas. Aldosterone is a key biological peptide in the renin-angiotensin-aldosterone system that regulates blood pressure. In this study, we investigated the siginificant stimulatory effect of aldosterone on leiomyoma cells proliferation. Study design. This study investigated the potential role of aldosterone in the proliferation of ELT-3 leiomyoma cells. Results. Aldosterone-induced ELT-3 leiomyoma cell proliferation and the expression of mineralocorticoid receptor (MR) were confirmed. Pre-incubating the cells with the MR blockers spironolactone or eplerenone effectively repressed aldosterone-induced and angiotensin II (Ang II)-induced cell proliferation. Treatment of aldosterone increased the levels of Ang II type-1 receptor mRNA. Conclusion. These experimental findings in vitro show the presence of complex regulation of Ang II and aldosterone induced leiomyoma cell proliferation.