Extracellular calcium (Ca2+o)-sensing receptor in a mouse monocyte-macrophage cell line (J774): potential mediator of the actions of Ca2+o on the function of J774 cells.
The calcium-sensing receptor (CaR) is a G protein-coupled receptor that plays key roles in extracellular calcium ion (Ca2+o) homeostasis in parathyroid gland and kidney. Macrophage-like mononuclear cells appear at sites of osteoclastic bone resorption during bone remodeling and may play a role in the "reversal" phase following osteoclastic resorption and preceding bone formation. Bone resorption produces substantial local increases in Ca2+o that could provide a signal for bone marrow mononuclear cells in the vicinity, leading us to investigate whether such mononuclear cells express the CaR. In this study, we used the mouse J774 cell line, which exhibits a pure monocyte-macrophage phenotype. Both immunocytochemistry and Western blot analysis, using polyclonal antisera specific for the CaR, detected CaR protein in J774 cells. The use of reverse transcriptase-polymerase chain reaction with CaR-specific primers, including a set of intron-spanning primers, followed by nucleotide sequencing of the amplified products, also identified CaR transcripts in J774 cells. Exposure of J774 cells to high Ca2+o (2.8 mM or more) or the polycationic CaR agonist, neomycin (100 microM), stimulated both chemotaxis and DNA synthesis in J774 cells. Therefore, taken together, our data strongly suggest that the monocyte-macrophage cell line, J774, possesses both CaR protein and mRNA very similar, if not identical, to those in parathyroid and kidney.
Yamaguchi
T
T
Department of Medicine, Brigham and Women's Hospital, and Harvard Medical School, Boston, Massachusetts 02115, USA.
Kifor
O
O
Chattopadhyay
N
N
Bai
M
M
Brown
E M
EM
D000595:Amino Acid Sequence; D000818:Animals; D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002633:Chemotaxis; D004247:DNA / Q000096:biosynthesis; D007150:Immunohistochemistry; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D009000:Monocytes / Q000378:metabolism*; D010641:Phenotype; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction
D002135:Calcium-Binding Proteins; D012333:RNA, Messenger; D004247:DNA