Suzuki K, Rapuano B E, Bockman R S
Hospital for Special Surgery, New York, New York, USA.
J Bone Miner Res. 1997 Apr;12(4):561-7. doi: 10.1359/jbmr.1997.12.4.561.
Gene expression of the matrix-degrading enzyme collagenase-1 in rabbit synoviocytes and human fibroblasts is down-regulated by prostaglandin E1 (PGE1) through a cyclic adenosine monophosphate (cAMP)-dependent pathway. In the current study, we examined the role of protein kinase A (PKA) in the PGE1-mediated effect on collagenase-1 gene expression. Collagenase-1 gene expression was rapidly induced several-fold above control both by a phorbol ester, 12-o-tetradecanoyl phorbol 13 acetate, and interleukin-1 beta (IL-1 beta) in HIG-82 synoviocytes. Treatment with PGE1 and forskolin increased PKA activity in the HIG-82 cells within 15 minutes of adding the stimulating agents. Two inhibitors of PKA, the isoquinoline-sulfonamide derivative, H-89 and a cAMP analog, RpcAMP, blocked the ability of PGE1 to down-regulate collagenase-1 gene expression. However, if PGE1 was added from 6 h to 30 minutes before the PKA inhibitor H-89, collagenase-1 gene expression was inhibited. Constitutive PKA activity was increased in HIG-82 synoviocytes stably transfected with an expression vector pCMV.C alpha that caused the HIG-82 cells to overexpress an active catalytic subunit of PKA. Cells stably transfected with an inactive, mutated C-alpha-variant showed no change in PKA activity. Collagenase-1 mRNA levels in TPA-stimulated cells were reduced to baseline levels in the pCMV.C alpha but not in the mutated C-alpha-transfected cells. These data show the importance of PKA in regulating collagenase-1 gene expression in a synoviocyte cell line.
基质降解酶胶原酶 -1 在兔滑膜细胞和人成纤维细胞中的基因表达可通过环磷酸腺苷（cAMP）依赖性途径被前列腺素 E1（PGE1）下调。在本研究中，我们检测了蛋白激酶 A（PKA）在 PGE1 介导的对胶原酶 -1 基因表达的影响中的作用。在 HIG - 82 滑膜细胞中，佛波酯、12 - O - 十四烷酰佛波醇 13 - 乙酸酯以及白细胞介素 -1β（IL -1β）均可使胶原酶 -1 基因表达迅速诱导至对照水平以上数倍。在添加刺激剂后的 15 分钟内，用 PGE1 和福斯高林处理可增加 HIG - 82 细胞中的 PKA 活性。PKA 的两种抑制剂，异喹啉 - 磺酰胺衍生物 H - 89 和一种 cAMP 类似物 RpcAMP，可阻断 PGE1 下调胶原酶 -1 基因表达的能力。然而，如果在 PKA 抑制剂 H - 89 之前 6 小时至 30 分钟添加 PGE1，则胶原酶 -1 基因表达会受到抑制。在用表达载体 pCMV.Cα稳定转染的 HIG - 82 滑膜细胞中，组成型 PKA 活性增加，该载体使 HIG - 82 细胞过表达 PKA 的活性催化亚基。用无活性的突变 C -α变体稳定转染的细胞中 PKA 活性无变化。在 TPA 刺激的细胞中，pCMV.Cα转染细胞中的胶原酶 -1 mRNA 水平降至基线水平，而在突变 C -α转染细胞中则未降低。这些数据表明 PKA 在调节滑膜细胞系中胶原酶 -1 基因表达方面具有重要作用。
