Estradiol formation by human osteoblasts via multiple pathways: Relation with osteoblast function

Abstract

The importance of estrogens in bone metabolism is illustrated by the accelerated bone loss and increase in osteoporotic fractures associated with postmenopausal estrogen deficiency. In this study, the expression and activity of the enzymes involved in estrogen metabolism in human osteoblastic cells were investigated in relation to differentiation of these cells. PCR reactions using mRNA from an in vitro differentiating human cell line (SV-HFO) were performed to assess mRNA expression of the enzymes aromatase, different subtypes of 17beta-hydroxysteroid dehydrogenase (17beta-HSD), and steroid sulfatase. Aromatase, sulfatase, and 17beta-HSD type 2 and 4 were found to be expressed throughout differentiation. Expression of 17beta-HSD type 3, however, was relatively weak, except for early time points in differentiation. Type 1 17beta-HSD expression was not detected. Aromatase activity decreased during differentiation, as was demonstrated by the conversion of androstenedione (A) and testosterone (T) into estrone (E(1)) and estradiol (E(2)), respectively. The 17beta-HSD isozymes catalysing a reductive reaction convert androstenedione and estrone into testosterone and estradiol, respectively. Their activity declined with differentiation. Analysis of 17beta-HSD activity indicated both oxidative (E(2) to E(1); T to A) and reductive (E(1) to E(2); A to T) metabolism at all stages of osteoblast differentiation. Both activities declined as cells moved toward a differentiating mineralizing phenotype. However, the oxidative reaction was increasingly in favor of the reductive reaction at all times during differentiation. Sulfatase activity, as demonstrated by the conversion of estrone-sulfate into estrone, was constant during differentiation. In conclusion, we have demonstrated that all enzymes necessary for estrogen metabolism are expressed and biologically active in differentiating human osteoblasts. The activity of aromatase and 17beta-HSD was found to be dependent on the stage of cell differentiation. In addition, human osteoblasts effectively convert estradiol into estrone. The efficacy of osteoblasts to synthesize estradiol may determine the ultimate change in rate of bone turnover after menopause, as well as the development of osteoporosis. Moreover, the enzymes involved in the metabolism of estradiol may form a target for intervention.

Aromatase activity of human mesenchymal stem cells is stimulated by early differentiation, vitamin D and leptin

“…In addition to gonads, this enzyme is found in different organs, including adipose tissue, brain, skin, endothelium and bone. Skeletal cells also express a number of other enzymes implicated in sex steroid metabolism (Schweikert et al 1980, Janssen et al 1999, Compston 2002, Ishida et al 2002, Issa et al 2002, supporting the concept that active androgens and oestrogens can be synthesized within the bone marrow cells from circulating C19 precursors. Thus, besides contributing to the circulating oestrogen pool, the oestrogen synthesized within bone tissue compartments may be locally active in a paracrine or intracrine way (Labrie et al 1997, Simpson 2000, Simpson & Davis 2001.…”

“…Aromatase has been reported to be expressed in hMSCs (Heim et al 2004), in osteoblast or osteoblast-like cells from foetal and adult tissues (Purohit et al 1992, Tanaka et al 1993, Schweikert et al 1995, Sasano et al 1997, Janssen et al 1999, in articular cartilage chondrocytes, in adipocytes adjacent to bone trabeculae, in osteocytes (Sasano et al 1997) and in macrophage/osteoclast-like cells (Shozu et al 1997). The expression of CYP 19 has been shown to be regulated by differential promoter usage, depending on the tissue context.…”

“…In cultures of bonederived osteoblast or osteoblast-like cells, the regulation of aromatase expression has been studied mainly at the transcriptional level, showing that dexamethasone, vitamin D, testosterone and phytoestrogen genistein, among others, may function as regulatory factors of CYP19 expression (Tanaka et al 1996, Jakob et al 1997, Shozu & Simpson 1998, Shozu et al 2000, Enjuanes et al 2003, Heim et al 2004. Further, transcription of CYP19 has been reported to be induced by physiological or pathological conditions, such as bone differentiation and fractures (Lea et al 1997, Janssen et al 1999, Heim et al 2004, pointing to the importance that local oestrogen generation may have for adequate triggering and ensuing of the differentiation pathway.…”

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

“…The SV-HFO cell line was established from human fetal calvaria, and retains the features of human osteoblasts well (13). Aromatase activity and gene transcript were detected in SV-HFO (14). Aromatase activity and CYP19 gene transcript were up-regulated in a Dex-dependent manner.…”

Regulation of CYP19 gene expression in primary human osteoblasts: effects of vitamin D and other treatments

“…Aromatase activity has been detected in several human adult and fetal tissues, and pathologic samples (9,10). With regard to bone, aromatase activity and/or aromatase mRNA have been detected in tissue samples (11 -13), in human cultured osteoblasts (14,15), in human osteosarcoma cell lines HOS, U2OS and MG63 (4 -6, 14, 16), in differentiating human osteoblastic cells such as SV-HFO (17), and in SV40-immortalized human osteoblasts (5,6). A fairly recent study (15) has shown that promoter usage and regulation of aromatase gene expression in osteoblast-like cells may be specific and different from those of other tissues such as placenta, ovary and adipose tissue.…”