New evidences on the regulation of SF-1 expression by POD1/TCF21 in adrenocortical tumor cells

OBJECTIVES: Transcription Factor 21 represses steroidogenic factor 1, a nuclear receptor required for gonadal development, sex determination and the regulation of adrenogonadal steroidogenesis. The aim of this study was to investigate whether silencing or overexpression of the gene Transcription Factor 21 could modulate the gene and protein expression of steroidogenic factor 1 in adrenocortical tumors. METHODS: We analyzed the gene expression of steroidogenic factor 1 using qPCR after silencing endogenous Transcription Factor 21 in pediatric adrenal adenoma-T7 cells through small interfering RNA. In addition, using overexpression of Transcription Factor 21 in human adrenocortical carcinoma cells, we analyzed the protein expression of steroidogenic factor 1 using Western blotting. RESULTS: Transcription Factor 21 knockdown increased the mRNA expression of steroidogenic factor 1 by 5.97-fold in pediatric adrenal adenoma-T7 cells. Additionally, Transcription Factor 21 overexpression inhibited the protein expression of steroidogenic factor 1 by 0.41-fold and 0.64-fold in two different adult adrenocortical carcinoma cell cultures, H295R and T36, respectively. CONCLUSIONS: Transcription Factor 21 is downregulated in adrenocortical carcinoma cells. Taken together, these findings support the hypothesis that Transcription Factor 21 is a regulator of steroidogenic factor 1 and is a tumor suppressor gene in pediatric and adult adrenocortical tumors.

POD-1/Tcf21 overexpression reduces endogenous SF-1 and StAR expression in rat adrenal cells

During gonad and adrenal development, the POD-1/capsulin/TCF21transcription factor negatively regulates SF-1/NR5A1expression, with higher SF-1 levels being associated with increased adrenal cell proliferation and tumorigenesis. In adrenocortical tumor cells, POD-1 binds to the SF-1 E-box promoter region, decreasing SF-1 expression. However, the modulation of SF-1 expression by POD-1 has not previously been described in normal adrenal cells. Here, we analyzed the basal expression of Pod-1 and Sf-1 in primary cultures of glomerulosa (G) and fasciculata/reticularis (F/R) cells isolated from male Sprague-Dawley rats, and investigated whether POD-1 overexpression modulates the expression of endogenous Sf-1 and its target genes in these cells. POD-1 overexpression, following the transfection of pCMVMycPod-1, significantly decreased the endogenous levels of Sf-1 mRNA and protein in F/R cells, but not in G cells, and also decreased the expression of the SF-1 target StAR in F/R cells. In G cells overexpressing POD-1, no modulation of the expression of SF-1 targets, StAR and CYP11B2, was observed. Our data showing that G and F/R cells respond differently to ectopic POD-1 expression emphasize the functional differences between the outer and inner zones of the adrenal cortex, and support the hypothesis that SF-1 is regulated by POD-1/Tcf21 in normal adrenocortical cells lacking the alterations in cellular physiology found in tumor cells.

Differential expression of upstream stimulatory factor (USF) 2 variants in eutopic endometria from women with endometriosis: estradiol regulation

BACKGROUND: Endometriosis, pro-inflammatory and invasive benign disease estrogen dependent, abnormally express in endometria the enzyme P450Arom, positively regulated by steroid factor-1 (SF-1). Our objective was to study the nuclear protein contents of upstream stimulating factor 2 (USF2a and USF2b), a positive regulator of SF-1, throughout the menstrual cycle in eutopic endometria from women with and without (control) endometriosis and the involvement of nuclear estrogen receptors (ER) and G-coupled protein estrogen receptor (GPER)-1. RESULTS: Upstream stimulating factor 2 protein contents were higher in mid (USF2b) and late (USF2a and USF2b) secretory phase in eutopic endometria from endometriosis than control (p < 0.05). In isolated control epithelial cells incubated with E2 and PGE2, to resemble the endometriosis condition, the data showed: (a) significant increase of USF2a and USF2b nuclear protein contents when treated with E2, PPT (specific agonist for ERa) or G1 (specific agonist for GPER1); (b) no increase in USF2 binding to SF-1 E-Box/DNA consensus sequence in E2-treated cells; (c) USF2 variants protein contents were not modified by PGE2; (d) SF-1 nuclear protein content was significantly higher than basal when treated with PGE2, E2 or G1, stimulation unaffected by ICI (nuclear ER antagonist); and (e) increased (p < 0.05) cytosolic protein contents of P450Arom when treated with PGE2, E2, PPT or G1 compared to basal, effect that was additive with E2 + PGE2 together. Nevertheless, in endometriosis cells, the high USF2, SF-1 and P450Arom protein contents in basal condition were unmodified. CONCLUSION: These data strongly suggest that USF2 variants and P450Arom are regulated by E2 through ERa and GPER1, whereas SF-1 through GPER1, visualized by the response of the cells obtained from control endometria, being unaffected the endogenously stimulated cells from endometriosis origin. The lack of E2 stimulation on USF2/SF-1 E-Box/DNA-sequence binding and the absence of PGE2 effect on USF2 variants opposite to the strong induction that they exert on SF1 and P450 proteins suggest different mechanisms and indirect regulations. The sustained USF2 variants protein expression during the secretory phase in eutopic endometria from women with endometriosis may participate in the pathophysiology of this disease strongly associated with infertility and its characteristic endometrial invasion to ectopic sites in the pelvic cavity.

Expression of TM4SF1 and its effect on proliferation, migration and invasion in pancreatic cancer cells / 中华胰腺病杂志

Objective To determine the expression of TM4SF1 mRNA in 5 human pancreatic cancer cell lines,and investigate its effect on the proliferation,migration and invasion of pancreatic cancer cells.Methods The expression of TM4SF1 mRNA in MPanc96,MiaPaCa-2,PANC1,AsPC-1,HPAC cells was determined by qRT-PCR,and the results were compared with that of human pancreatic ductal epithelial (HPDE) cells.RNA interference method was used to transiently transfect siRNA targeting at TM4SF1 and negative control siRNA into MPanc96,MiaPaCa-2 cells.The proliferation of cells were measured by MTS method,and migration and invasion of cells were determined by Transwell.Results The expression levels of TM4SF1 mRNA in pancreatic cancer cell lines MPanc96,MiaPaCa-2,PANC1,AsPC-1 and HPAC were 1.205 ± 0.073,1.096 ± 0.260,1.382 ± 0.075,1.374 ± 0.363 and 0.744 ± 0.096,which were significantly highly than that in HPDE (0.020 ± 0.003,P ＜ 0.01).Compared with cells transfected with negative control siRNA,the proliferation of MPanc96 and MiaPaCa-2 cells transfected with siRNA targeting at TM4SF1 was not significantly changed,but the migration abilitiy was decreased by (62.5 ± 7.6) ％ and (72.8 ± 4.0) ％,and invasion abilitiy was decreased by (69.5 ± 5.7) ％ and (78.6 ± 6.3) ％.Conclusions TM4SF1 is highly expressed in pancreatic cancer cells and appears to promote the migration and invasion abilities of the cancer cells.

SF-1 gene mutation and gonad development / 国际儿科学杂志

Orphan nuclear receptor steroidogenic factor 1 (SF-1) is a member of the nuclear receptor superfamily. It regulates the expression of multiple genes involved in adrenal and gonadal development,steroidogenesis,and reproductive function. Twenty five different mutations have been described and their clinical features are various. Detection of SF-1 gene may be contribute to the etiological diagnosis for 46, XY disorder of sex development patients.

Influence of fluoride on the expression TM9SF1 mRNA and Ras mRNA of human osteoblasts / 中国地方病学杂志

Objective To detect the influence of fluoride on the expression TM9SF1 mRNA and Ras mRNA of osteoblasts. Methods The third generation of primary cultured osteoblasts were exposed to a series concentrations of 0,2.5,5.0, 10.0,20.0 mg/L fluoride for 10 days. The influence of different doses of fluorine on the expression of TM9SF1 mRNA and Ras mRNA of osteoblasts cultured in vitro was investigated by SYBR Green I methods. Results The osteoblasts of the control group and the 2.5 mg/L group were in the shape of long spindle, triangle or irregular polygon and had processes, and the cytoplasm was translucent, adjacent cells affixed to each other under light microscope. Those of the 20.0 mg/L group shaped as long spindle or irregular polygon, and some vacuolization and granular materials appeared in cytoplasm. The number of the cells decreased and the volume increased significantly. After exposed to fluoride for 10 days, osteoblasts of 2.5 mg/L group morphologically proliferated. There were statistical siguificances between each groups of TM9SF1 mRNA in human osteoblasts(F = 322.82, P < 0.01). The highest in the 2.5 mg/L group(9326.0 ± 115.97), the expression of TM9SF1 mRNA decreased along with the increasing dose of fluorine. There were statistical significances between 5.0, 10.0,20.0 mg/L groups(6495.0 ± 323.9, 4387.5 ± 545.2, 5962.5 ± 536.7) and control group(9221.0 ± 107.5, all P< 0.01). There was a statistical significance between each groups of Ras mRNA in human osteoblasts(F = 703.28, P < 0.01). The highest in the control group, the expression of Ras mRNA decreased along with the increasing of dose of fluorine. There were statistical significance between 2.5, 5.0, 10.0, 20.0 mg/L groups(6144.5 ± 270.82,5603.5 ± 88.39,3181.0 ± 159.81,4067.5 ± 37.4) and control group(6571.0 ± 196.58). Conclusion The influence on TM9SFI mRNA and Ras mRNA expression in osteoblasts correlates with the dose of fluorine.

Immunosuppressive effects of human semen on macrophage of mouse / 中国免疫学杂志

This paper reports the functional changes of macrophage under the effect of SF1, the first fraction of human semen separated by Sephadex G-100. Two kinds of Balb/c mouse macrophage, i. e. peri toneal macrophage (PEC-M?) and the bone marrow derived macrophage (BM-M?), were treated by SF1 in vivo (i.p. ) and in vitro. It has been proved that SF1 is a potent immunosuppressive fraction; (1) As compared with the control, the phagocytosis of SF1 treated macrophage to SRBC coated with antibody declines significantly in terms of phagocytosis rate and index (p