Aromatase expression in the human fetal osteoblastic cell line SV-HFO

A number of clinical studies have highlighted the importance of estrogen in bone growth and maintenance in men and postmenopausal women. In these instances, estrogen is synthesized locally within bone tissue by aromatase, encoded by the CYP19 gene. The mechanisms regulating aromatase expression in bone, however, are unclear. In this work we characterized the expression of aromatase activity and gene transcripts in the human fetal osteoblastic cell line, SV-HFO. Aromatase activity and gene transcript expression were stimulated by dexamethasone. Oncostatin M strongly stimulated aromatase expression in synergy with dexamethasone. These factors induced CYP19 transcripts that included the sequence of exon I.4 in their 5'UTR. Consistent with this, a reporter construct harboring the genomic sequence of the promoter region of exon I.4 (promoter I.4) was also activated by dexamethasone and oncostatin M. 5' deletion and mutation analysis revealed important roles for a glucocorticoid response element, an interferon gamma activating sequence and a putative binding site for Sp1. Transfection of exogenous glucocorticoid receptor, STAT3 or Sp1 increased promoter activity, indicating a potential role for these transcription factors in regulating aromatase expression in SV-HFO cells. These data suggest that the SV-HFO cell line is a valuable model with which to elucidate the mechanisms regulating local estrogen synthesis in osteoblasts.

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Forskolin and dexamethasone synergistically induce aromatase (CYP19) expression in the human osteoblastic cell line SV-HFO

Acta Endocrinologica ◽

10.1530/eje.1.01882 ◽

2005 ◽

Vol 152 (4) ◽

pp. 619-624 ◽

Cited By ~ 10

Author(s):

Masatada Watanabe ◽

Shuji Ohno ◽

Shizuo Nakajin

Keyword(s):

Oncostatin M ◽

Osteoblastic Cell ◽

Aromatase Activity ◽

Gene Transcript ◽

Rt Pcr ◽

Mineral Density ◽

Osteoblastic Cell Line ◽

Human Osteoblastic Cell ◽

Estrogen Concentration ◽

Cyp19 Expression

Objective: Recent progress supports the importance of local estrogen secretion in human bone tissues to increase and maintain bone mineral density. In a previous study, we reported that the expression of aromatase (CYP19) is dexamethasone (Dex)-dependent and oncostatin M (OSM) increases the expression synergistically with Dex. In the present study, we examined the effects of forskolin (FSK) as another potential synergistic factor. Results: Co-administration of 100 nM Dex and 10 μM FSK increased aromatase activity 4-fold compared with Dex alone. The results of reverse transcriptase (RT)-PCR suggest that the amount of CYP19 gene transcript was also up-regulated by FSK synergistically with Dex, and that promoter I.4, which is not activated by FSK alone, is activated by FSK synergistically with Dex. The results of RT-PCR also suggest that promoter II, which responds to FSK, was not activated even in the presence of FSK in SV-HFO. The promoter I.4 sequence that was transfected into SV-HFO was activated by FSK synergistically with Dex. Conclusions: Synergistic up-regulation of aromatase activity, CYP19 gene transcript, and promoter I.4 activity were Dex-dependent and not up-regulated by FSK alone. The results of this work may form the basis of bone-specific estrogen-replacement therapy that increases the estrogen concentration in bone tissue only.

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Forskolin up-regulates aromatase (CYP19) activity and gene transcripts in the human adrenocortical carcinoma cell line H295R

Journal of Endocrinology ◽

10.1677/joe.0.1800125 ◽

2004 ◽

Vol 180 (1) ◽

pp. 125-133 ◽

Cited By ~ 31

Author(s):

M Watanabe ◽

S Nakajin

Keyword(s):

Cell Line ◽

Adrenocortical Carcinoma ◽

Carcinoma Cell ◽

Carcinoma Cell Line ◽

Adrenal Tumors ◽

Aromatase Activity ◽

Gene Transcript ◽

Aromatase Expression ◽

Cyp19 Gene ◽

Gene Transcripts

A number of conditions related to sex-reversal in boys and men and precocious puberty in girls are caused by estrogen-secreting adrenal tumors. In these tumors, cytochrome P450 aromatase (aromatase) that is encoded in the CYP19 gene is expressed at high levels. To investigate the molecular mechanism of aromatase expression in these adrenal tumors, we characterized the activity, gene transcript and genomic promoter region of aromatase in the human adrenocortical carcinoma cell line H295R. Aromatase activity and the transcript of the CYP19 gene were highly up-regulated by forskolin, but not by dexamethasone. The results from exon I-specific reverse transcriptase (RT)-PCR and the transfection of reporter constructs suggested that promoter I.3 and promoter II were activated in H295R. Deletion and mutation analysis suggested that cAMP response element-like sequence (CLS) and steroidogenic factor-1 (SF-1) motif, were critical for the activation of promoter II. The results of this work should provide the basis for the molecular analysis of aromatase expression in adrenocortical cells.

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Effect of β-agonist on the dexamethasone-induced expression of aromatase by the human monocyte cells

Endocrine Connections ◽

10.1530/ec-16-0099 ◽

2017 ◽

Vol 6 (2) ◽

pp. 82-88 ◽

Cited By ~ 2

Author(s):

Masatada Watanabe ◽

Shuji Ohno ◽

Hiroshi Wachi

Keyword(s):

Atopic Dermatitis ◽

Human Skin ◽

Human Monocyte ◽

Skin Thickness ◽

Aromatase Activity ◽

Gene Transcript ◽

Aromatase Expression ◽

Estrogen Synthesis ◽

Peripheral Monocyte

Emerging evidence suggests that sex steroids are important for human skin health. In particular, estrogen improves skin thickness, elasticity and moisture of older women. The major source of circulating estrogen is the ovary; however, local estrogen synthesis and secretion have important roles in, for example, bone metabolism and breast cancer development. We hypothesized that infiltrated peripheral monocytes are one of the sources of estrogen in skin tissues. We also hypothesized that, during atopic dermatitis under stress, a decline in the hypothalamus–pituitary–adrenal axis (HPA) and facilitation of the (hypothalamus)–sympathetic–adrenomedullary system (SAM) attenuates estrogen secretion from monocytes. Based on this hypothesis, we tested aromatase expression in the human peripheral monocyte-derived cell line THP-1 in response to the synthetic glucocorticoid dexamethasone (Dex), the synthetic β-agonist isoproterenol (Iso) and the β-antagonist propranolol (Pro). Dex mimics glucocorticoid secreted during excitation of the HPA, and Iso mimics catecholamine secreted during excitation of the SAM. We found that aromatase activity and the CYP19A1 gene transcript were both upregulated in THP-1 cells in the presence of Dex. Addition of Iso induced their downregulation and further addition of Pro rescued aromatase expression. These results may suggest that attenuation of estrogen secretion from peripheral monocytes could be a part of the pathology of stress-caused deterioration of atopic dermatitis. Further examination using an in vitro human skin model including THP-1 cells might be a valuable tool for investigating the therapeutic efficacy and mechanism of estrogen treatment for skin health.

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Effect of epidermal growth factor and prostaglandin on the expression of aromatase (CYP19) in human adrenocortical carcinoma cell line NCI-H295R cells

Journal of Endocrinology ◽

10.1677/joe.1.06214 ◽

2006 ◽

Vol 188 (1) ◽

pp. 59-68 ◽

Cited By ~ 13

Author(s):

Masatada Watanabe ◽

Mariko Noda ◽

Shizuo Nakajin

Keyword(s):

Growth Factor ◽

Epidermal Growth Factor ◽

Cell Line ◽

Adrenocortical Carcinoma ◽

Carcinoma Cell ◽

Aromatase Activity ◽

Gene Transcript ◽

Aromatase Expression ◽

H295r Cells ◽

Epidermal Growth

We investigated the effects of epidermal growth factor (EGF) and prostaglandins (PG) on the expression of aromatase (CYP19) in human adrenocortical carcinoma cell line NCI-H295R cells. EGF significantly increased aromatase activity and CYP19 gene transcript in NCI-H295R cells. Exon PII was selected from among several tissue-specific exon I regions. Promoter II that abuts on exon PII was activated by EGF. PGE2 also significantly increased aromatase activity, CYP19 gene transcript, and promoter II activity. The results of experiments using protein kinase (PK) inhibitors suggest that the cAMP–PKA signaling pathway is involved in the up-regulation of aromatase expression by EGF. PGE2 activated promoter II activity in 4 h, while12 h was required for its activation by EGF. In addition, PGE2 was secreted from NCI-H295R cells in response to EGF. Selective agonists for prostaglandin receptors EP1 and EP2 significantly increased aromatase activity, which was decreased by the corresponding antagonists. Finally, antagonists for EP1 and EP2 inhibited the up-regulation of aromatase expression following EGF. These results suggest that PGE2 secondarily acts as an autocrine signal in the up-regulation of aromatase expression by EGF in NCI-H295R cells.

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