Dixon D.,Molecular Pathogenesis Group |
Dixon D.,Bureau of Health of Wujin District |
Di X.,U.S. National Institutes of Health |
Andrews D.M.K.,U.S. National Institutes of Health |
And 7 more authors.
Experimental and Molecular Medicine | Year: 2012
Previously, we found that high doses of genistein show an inhibitory effect on uterine leiomyoma (UtLM) cell proliferation. In this study, using microarray analysis and Ingenuity Pathways AnalysisTM, we identified genes (up- or down-regulated, ≥ 1.5 fold, P ≤ 0.001), functions and signaling pathways that were altered following treatment with an inhibitory concentration of genistein (50 μg/ml) in UtLM cells. Downregulation of TGF-β signaling pathway genes, activin A, activin B, Smad3, TGF-β2 and genes related to cell cycle regulation, with the exception of the upregulation of the CDK inhibitor P15, were identified and validated by real-time RT-PCR studies. Western blot analysis further demonstrated decreased protein expression of activin A and Smad3 in genistein-treated UtLM cells. Moreover, we found that activin A stimulated the growth of UtLM cells, and the inhibitory effect of genis-tein was partially abrogated in the presence of activin A. Overexpression of activin A and Smad3 were found in tissue samples of leiomyoma compared to matched myometrium, supporting the contribution of activin A and Smad3 in promoting the growth of UtLM cells. Taken together, these results suggest that down-regulation of activin A and Smad3, both members of the TGF-β pathway, may offer a mechanistic explanation for the inhibitory effect of a high-dose of genistein on UtLM cells, and might be potential therapeutic targets for treatment of clinical cases of uterine leiomyomas. Copyright © 2012 by the Korean Society for Biochemistry and Molecular Biology.
Mozzachio K.,Mozzachio Veterinary Services |
Moore A.B.,Molecular Pathogenesis Group |
Kissling G.E.,U.S. National Institutes of Health |
Dixon D.,Molecular Pathogenesis Group
Toxicologic Pathology | Year: 2016
Uterine leiomyomas in miniature pet pigs occur similarly to those in women with regard to frequency, age, parity, and cycling. Clinical signs, gross, and histologic features of the porcine tumors closely resemble uterine leiomyomas (fibroids) in women. Although fibroids are hormonally responsive in women, the roles of estrogen and progesterone have not been fully elucidated. In this study, immunohistochemistry was used to assess the expression of the steroid hormone receptors, estrogen receptor alpha (ER-α), estrogen receptor beta (ER-β) and progesterone receptor (PR), and cell proliferation markers, proliferating cell nuclear antigen (PCNA) and Ki-67 in tumor and matched myometrial tissues sampled from miniature pigs. A €quickscore€ method was used to determine receptor expression and labeling indices were calculated for the markers. ER-α/β and PR were localized to the nuclei of smooth muscle cells in both tissues. PR expression was intense and diffuse throughout all tissues, with correlation between tumors and matched myometria. Conversely, ER-α expression was variable between the myometrial and tumor tissues, as well as between animals. ER-β expression was low. PCNA and Ki-67 were localized to the nucleus and expression varied among tumors; however, normal tissues were overall negative. These findings support further investigation into the use of the miniature pig as a model of fibroids in women. © Society of Toxicologic Pathology.
Gao X.,Nanjing Medical University |
Gao X.,Molecular Pathogenesis Group |
Wang Q.,Nanjing Medical University |
Wang J.,Nanjing Medical University |
And 6 more authors.
Archives of Toxicology | Year: 2012
Exposure to fenvalerate was demonstrated to be toxic to the male reproductive system. Our previous data revealed that intracellular calcium plays an important role in regulating the above toxicity, through actions on both T-type calcium channels and endoplasmic reticulum calcium signals. The present study explored the effects of fenvalerate on the expression of calmodulin in mouse testis and GC-2spd(ts) cells, and its association with fenvalerateinduced male reproductive toxicity. Male mice were subjected to different doses (3.71, 18.56, 37.12, 92.81 mg/kg bw) of fenvalerate or vehicle control for 4 weeks. Expression of calmodulin was determined by real-time polymerase chain reaction (PCR) and Western blot analysis in mouse testis. Similar approaches were utilized in GC-2spd(ts) cells cultured with 5 μM fenvalerate at different time points. In the in vivo study, all mice survived through the entire 4 weeks. Administration of fenvalerate resulted in a dose-dependent reduction in testis weight/body weight, sperm motility, and increased head abnormality rate. By histological staining, mice treated with fenvalerate at higher doses showed dilated seminiferous tubules and disturbed arrangement of spermatogenic cells. Meanwhile, both mRNA and protein expression of calmodulin were significantly increased in the testes of mice exposed to fenvalerate compared to control mice. Moreover, in the in vitro study, 5 μM fenvalerate significantly increased the expression of calmodulin at the mRNA and protein levels in GC-2spd(ts) cells after 8 h of incubation and sustained these levels for at least 24 h. Collectively, these data suggested that enhanced expression of calmodulin correlates with male reproductive damage induced by fenvalerate. © Springer-Verlag 2012.
PubMed | Molecular Pathogenesis Group
Type: Journal Article | Journal: Environmental health perspectives | Year: 2015
It has been proposed that cadmium (Cd) is an environmental metalloestrogen and that its action is mediated via the estrogen receptor (ER). Cd mimics the effects of estrogen in the rat uterus, and blood Cd concentrations positively correlate with ER levels in uteri of women with fibroids.In the present study we explored whether Cd could stimulate proliferation of estrogen-responsive human uterine leiomyoma (ht-UtLM) cells and uterine smooth muscle cells (ht-UtSMCs) through classical interactions with ER and ER, or by nongenomic mechanisms.We used estrogen response element (ERE) reporters, phosphorylated receptor tyrosine kinase arrays, Western blot analysis, estrogen binding, and cell proliferation assays to evaluate the effects of Cd on ht-UtLM cells and ht-UtSMCs.Cd stimulated growth of both cell types at lower concentrations and inhibited growth at higher concentrations ( 50 M). Cd did not significantly bind to ER or ER, nor did it show transactivation in both cell types transiently transfected with ERE reporter genes. However, in both cells types, Cd (0.1 M and 10 M) activated p44/42 MAPK (ERK1/2), and a MAPK inhibitor (PD98059) abrogated Cd-induced cell proliferation. Cd in ht-UtLM cells, but not in ht-UtSMCs, activated the growth factor receptors EGFR, HGFR, and VEGF-R1 upstream of MAPK. Additional studies in ht-UtLM cells showed that AG1478, an EGFR inhibitor, abolished Cd-induced phosphorylation of EGFR and MAPK.Our results show that low concentrations of Cd stimulated cell proliferation in estrogen-responsive uterine cells by nongenomic activation of MAPK, but not through classical ER-mediated pathways.