1 alpha,25-dihydroxyvitamin D3 regulates the expression of carbonic anhydrase II in nonerythroid avian bone marrow cells.

Abstract

la,25-Dihydroxyvitamin D3 [1,25(OH)2D3], the active metabolite of the steroid hormone vitamin D, is a potent regulator of macrophage and osteoclast differentiation. The mature osteoclast, unlike the circulating monocyte or the tissue macrophage, expresses high levels of carbonic anhydrase H (CAH). This enzyme generates protons and bicarbonate from water and carbon dioxide and is involved in bone resorption and acid-base regulation. To test whether 1,25(OH)2D3 could induce the differentiation of myelomonocytic precursors toward osteoclasts rather than macrophages, we analyzed its effects on the expression of CAll in bone marrow cultures containing precursors common to both cell types. The expression of CAU was markedly increased by 1,25(OH)2D3 in a doseand time-dependent manner. In bone marrow, this increase occurred at the mRNA and protein levels and was detectable as early as 24 hr after stimulation. 1,25(OH)2D3 was also found to induce CAH expression in a transformed myelomonocytic avian cell line. These results suggest that 1,25(OH)2D3 regulates the level at which myelomonocytic precursors express CAU, an enzyme that is involved in the function of the mature osteoclast. la,25-Dihydroxyvitamin D3 [1,25(OH)2D3] is the active metabolite of the steroid hormone vitamin D (1). This metabolite regulates the differentiation of osteoclasts and macrophages, two cell types that are thought to be derived from a common myelomonocytic precursor (2-6). 1,25(OH)2D3 induces an increase in bone resorption in vivo (1, 7) and in organ cultures (8, 9) but has no effect on the resorptive activity of isolated osteoclasts (10, 11). The increase in bone resorption is believed to be mediated primarily by means of an effect upon the proliferation and/or differentiation of osteoclast precursors within the hematopoietic bone marrow (12-15), leading to an increase in the number of mature osteoclasts (7,9 (13,14,19,20) and increases the proportion ofthe cells that can resorb bone (14).Taken together, these results suggest that this steroid hormone may specifically induce the differentiation of myelomonocytic precursors toward the macrophage and/or osteoclast lineage.One of the genes that is expressed at high levels in the osteoclast (21-23), but not in peripheral monocytes or mature macrophages (24), is the gene encoding carbonic anhydrase II (CAII). CAII is an enzyme that reversibly generates bicarbonate and protons from carbon dioxide and water (25). This enzyme plays an essential role in acid-secreting cells such as the kidney tubule intercalated cell (26), the gastric mucosa oxyntic cell (27), and the osteoclast (21). In humans, defective CAII is associated with cerebral calcification, tubular acidosis, and osteopetrosis (28), providing genetic evidence for CAII involvement in bone resorption. An analogue of the human disease has been produced by chemical mutagenesis in the mouse, where CAII deficiency is associated with renal tubular acidosis and vascular calcification but no major bone defects (29).We have investigated wheth...

Osteoclasts express high levels of pp60c-src in association with intracellular membranes.

“…Although our results do not address the issue of pp60~-'~'s functional role in bone resorption, they indicate that the protein is specifically expressed at high levels in the osteoclast, as shown by immunocytochemistry, immunoblotting, and src kinase activity. Induction of differentiation of bone marrow mononuclear cells toward an osteoclast-like phenotype by treatment with 1,25(OH):D3 (4,5,33) led to higher levels of pp60 ~ .... than in untreated cells, further suggesting that osteoclasts are characterized by elevated c-src expression. These results clearly demonstrate a direct link between the osteoclast and c-src expression, and strongly suggest, but by no means prove, that the osteoclast may be the site of the specific defect in c-src-mice that leads to impaired osteoclast function and osteopetrosis.…”

v-erbA overexpression is required to extinguish c-erbA function in erythroid cell differentiation and regulation of the erbA target gene CAII.

“…Preliminary evidence indeed suggests that the CAII gene is regulated by retinoic acid in normal bone marrow progenitors (C. Schr6der and H. Beug,unpubl.). Other studies have demonstrated that CAII is also regulated by T3, vitamin D3, and androgen (Harkonen and V/i/in~inen 1988;Hillstrom-Shapiro et al 1989;Billecocq et al 1990;Pain et al 1990). However, no cis-acting hormoneresponse element has been identified thus far in the regulatory region of the CAII gene.…”

A New Experimental System for the Extended Application of Cyclic Hydrostatic Pressure to Cell Culture

“…The calculated HCO 3 − output from the ABL5 was used as a measure of proper humidification. Assuming that BMPCs cells do not produce their own bicarbonate through carbonic anhydrase [23], increases or decreases in HCO 3 − can indicate a change in the concentration of media components due to water evaporation or condensation, respectively.…”