TY - JOUR
T1 - Cyclooxygenase-2 gene disruption promotes proliferation of murine calvarial osteoblasts in vitro
AU - Xu, Zheng
AU - Choudhary, Shilpa
AU - Okada, Yosuke
AU - Voznesensky, Olga
AU - Alander, Cynthia
AU - Raisz, Lawrence
AU - Pilbeam, Carol
N1 - Funding Information:
Grant funding: RO1DK48361 and the Donaghue Foundation.
Funding Information:
This work was funded by NIH RO1DK48361 and an award from the Donaghue Foundation of Connecticut to CP.
PY - 2007/7
Y1 - 2007/7
N2 - Cyclooxygenase-2 (COX-2) is highly expressed in osteoblasts, and COX-2 produced prostaglandins (PGs) can increase osteoblastic differentiation in vitro. The goal of this study was to examine effects of COX-2 expression on calvarial osteoblastic proliferation and apoptosis. Primary osteoblasts (POBs) were cultured from calvariae of COX-2 wild-type (WT) and knockout (KO) mice. POB proliferation was evaluated by 3H-thymidine incorporation and analysis of cell replication and cell cycle distribution by flow cytometry. POB apoptosis was evaluated by annexin and PI staining on flow cytometry. As expected, PGE2 production and alkaline phosphatase (ALP) activity were increased in WT cultures compared to KO cultures. In contrast, cell numbers were decreased in WT compared to KO cells by day 4 of culture. Proliferation, measured on days 3-7 of culture, was 2-fold greater in KO than in WT POBs and associated with decreased Go/G1 and increased S cell cycle distribution. There was no significant effect of COX-2 genotype on apoptosis under basal culture conditions on day 5 of culture. Cell growth was decreased in KO POBs by the addition of PGE2 or a protein kinase A agonist and increased in WT POBs by the addition of NS398, a selective COX-2 inhibitor. In contrast, differentiation and cell growth in marrow stromal cell (MSC) cultures, evaluated by ALP and crystal violet staining respectively, were increased in MSCs from WT mice compared to MSCs from KO mice, and exogenous PGE2 increased cell growth in KO MSC cultures. We conclude that PGs secondary to COX-2 expression decrease osteoblastic proliferation in cultured calvarial cells but increase growth of osteoblastic precursors in MSC cultures.
AB - Cyclooxygenase-2 (COX-2) is highly expressed in osteoblasts, and COX-2 produced prostaglandins (PGs) can increase osteoblastic differentiation in vitro. The goal of this study was to examine effects of COX-2 expression on calvarial osteoblastic proliferation and apoptosis. Primary osteoblasts (POBs) were cultured from calvariae of COX-2 wild-type (WT) and knockout (KO) mice. POB proliferation was evaluated by 3H-thymidine incorporation and analysis of cell replication and cell cycle distribution by flow cytometry. POB apoptosis was evaluated by annexin and PI staining on flow cytometry. As expected, PGE2 production and alkaline phosphatase (ALP) activity were increased in WT cultures compared to KO cultures. In contrast, cell numbers were decreased in WT compared to KO cells by day 4 of culture. Proliferation, measured on days 3-7 of culture, was 2-fold greater in KO than in WT POBs and associated with decreased Go/G1 and increased S cell cycle distribution. There was no significant effect of COX-2 genotype on apoptosis under basal culture conditions on day 5 of culture. Cell growth was decreased in KO POBs by the addition of PGE2 or a protein kinase A agonist and increased in WT POBs by the addition of NS398, a selective COX-2 inhibitor. In contrast, differentiation and cell growth in marrow stromal cell (MSC) cultures, evaluated by ALP and crystal violet staining respectively, were increased in MSCs from WT mice compared to MSCs from KO mice, and exogenous PGE2 increased cell growth in KO MSC cultures. We conclude that PGs secondary to COX-2 expression decrease osteoblastic proliferation in cultured calvarial cells but increase growth of osteoblastic precursors in MSC cultures.
KW - Apoptosis
KW - Marrow stromal cells
KW - Mitogenesis
KW - Nonsteroidal anti-inflammatory drugs
KW - Prostaglandins
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U2 - 10.1016/j.bone.2007.03.009
DO - 10.1016/j.bone.2007.03.009
M3 - Article
C2 - 17467356
AN - SCOPUS:34250192194
VL - 41
SP - 68
EP - 76
JO - Bone
JF - Bone
SN - 8756-3282
IS - 1
ER -