Department of Physiology, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan.
Bone. 2011 Feb;48(2):286-97. doi: 10.1016/j.bone.2010.09.020. Epub 2010 Sep 18.
Cyclooxygenase-2 (COX-2) is thought to be an inducible enzyme, but increasing reports indicate that COX-2 is constitutively expressed in several organs. The status of COX-2 expression in bone and its physiological role remains undefined. Non-selective non-steroidal anti-inflammatory drugs (NSAIDs) and selective COX-2 inhibitors, which commonly suppress COX-2 activity, were reported to suppress osteoblast proliferation via Akt/FOXO3a/p27(Kip1) signaling, suggesting that COX-2 may be the key factor of the suppressive effects of NSAIDs on proliferation. Although Akt activation correlates with PTEN deficiency and cell viability, the role of COX-2 on PTEN/Akt regulation remains unclear. In this study, we hypothesized that COX-2 may be constitutively expressed in osteoblasts and regulate PTEN/Akt-related proliferation. We examined the localization and co-expression of COX-2 and p-Akt in normal mouse femurs and in cultured mouse (mOBs) and human osteoblasts (hOBs). Our results showed that osteoblasts adjacent to the trabeculae, periosteum and endosteum in mouse femurs constitutively expressed COX-2, while COX-2 co-expressed with p-Akt in osteoblasts sitting adjacent to trabeculae in vivo, and in mOBs and hOBs in vitro. We further used COX-2 siRNA to test the role of COX-2 in Akt signaling in hOBs; COX-2 silencing significantly inhibited PTEN phosphorylation, enhanced PTEN activity, and suppressed p-Akt level and proliferation. However, replenishment of the COX-2 enzymatic product, PGE2, failed to reverse COX-2-dependent Akt phosphorylation. Furthermore, transfection with recombinant human COX-2 (rhCOX-2) significantly reversed COX-2 siRNA-suppressed PTEN phosphorylation, but this effect was reduced when the enzymatic activity of rhCOX-2 was blocked. This finding indicated that the effect of COX-2 on PTEN/Akt signaling is not related to PGE2 but still dependent on COX-2 enzymatic activity. Conversely, COX-1 silencing did not affect PTEN/Akt signaling. Our findings provide new insight into bone physiology; namely, that COX-2 is constitutively expressed in osteoblasts in the dynamic bone growth area, which facilitates osteoblast proliferation via PTEN/Akt/p27(Kip1) signaling.
环氧化酶-2（COX-2）被认为是一种诱导酶，但越来越多的报道表明 COX-2 在几个器官中持续表达。骨中 COX-2 表达的状态及其生理作用尚不清楚。非选择性非甾体抗炎药（NSAIDs）和选择性 COX-2 抑制剂，通常抑制 COX-2 活性，据报道通过 Akt/FOXO3a/p27(Kip1)信号抑制成骨细胞增殖，表明 COX-2 可能是 NSAIDs 对增殖抑制的关键因素。尽管 Akt 激活与 PTEN 缺乏和细胞活力相关，但 COX-2 对 PTEN/Akt 调节的作用仍不清楚。在这项研究中，我们假设 COX-2 可能在成骨细胞中持续表达并调节 PTEN/Akt 相关增殖。我们检查了正常小鼠股骨中 COX-2 和 p-Akt 的定位和共表达，以及培养的小鼠（mOBs）和成骨细胞（hOBs）。我们的结果表明，小鼠股骨中靠近小梁、骨膜和骨内膜的成骨细胞持续表达 COX-2，而体内小梁附近的成骨细胞中 COX-2 与 p-Akt 共表达，在体外的 mOBs 和 hOBs 中也是如此。我们进一步使用 COX-2 siRNA 测试 COX-2 在 hOBs 中 Akt 信号的作用；COX-2 沉默显著抑制了 PTEN 的磷酸化，增强了 PTEN 的活性，并抑制了 p-Akt 水平和增殖。然而，补充 COX-2 的酶产物 PGE2 未能逆转 COX-2 依赖性 Akt 磷酸化。此外，转染重组人 COX-2（rhCOX-2）显著逆转了 COX-2 siRNA 抑制的 PTEN 磷酸化，但当 rhCOX-2 的酶活性被阻断时，这种效应降低。这一发现表明 COX-2 对 PTEN/Akt 信号的影响与 PGE2 无关，但仍依赖于 COX-2 的酶活性。相反，COX-1 沉默不影响 PTEN/Akt 信号。我们的发现为骨骼生理学提供了新的见解；即在动态骨生长区域，成骨细胞中持续表达 COX-2，通过 PTEN/Akt/p27(Kip1)信号促进成骨细胞增殖。
