副甲狀腺素類似物對退化性關節炎的影響

退化性關節炎(osteoarthritis, OA)是一種好發於負重關節的退化性疾病。當關節炎發生時，關節軟骨因磨損而喪失細胞外間質(extracellular matrix)，例如第II型膠原蛋白與葡萄糖胺聚醣(glycosaminoglycan, GAG)等。由於造成退化性關節炎的病因與機制不明，使得退化性關節炎的治療成效有限。目前在細胞學的研究上發現，在退化性關節炎發生時，成熟的軟骨細胞會走向終止分化(terminal differentiation)，同時表現出第X型膠原蛋白並逐漸走向凋亡，此一現象與胚胎期軟骨細胞進行軟骨內骨化的情況相類似。研究指出副甲狀腺素相關胜肽(parathyroid hormone related peptide, PTHrP)具有抑制軟骨細胞肥大，抑制終止分化，並維持軟骨細胞軟骨化的能力。副甲狀腺素(parathyroid hormone, PTH)與PTHrP兩者均可透過PTH接受器1型(PTHR1)活化蛋白激酶A (protein kinase A, PKA)及蛋白激酶C (protein kinase C, PKC)的訊息傳遞路徑。根據本實驗室先前的研究結果顯示，PTH類似物(1-34)能抑制人類關節軟骨細胞走向終止分化；並在大鼠退化性關節炎模式中，證明PTH(1-34)顯著抑制退化性關節炎的發生。然而，其詳細的調控機制則尚未釐清。近期研究指出，軟骨細胞內cAMP被活化時，能抑制軟骨細胞外間質被分解，顯示cAMP對抑制細胞外間質的流失是一重要的訊息路徑。故本研究之目的是利用僅能活化cAMP/PKA路徑的PTH類似物(1-31)，來釐清退化性關節炎進程的抑制是否透過cAMP/PKA的訊息路徑。實驗是以papain誘發退化性關節炎的大鼠後，分別給予PTH(1-31)與PTH(1-34)之治療作比較，以釐清cAMP/ PKA訊息路徑在OA發生與治療上的重要性。實驗結果發現PTH(1-31)能抑制葡萄糖胺聚醣及第II型膠原蛋白之流失，並阻止第X型膠原蛋白表現及抑制細胞凋亡，顯示PTH (1-31)具有抑制退化性關節炎的進行，此外，在本研究中發現，PTH (1-31)單獨處理於正常關節軟骨後並不影響葡萄糖胺聚醣及第II型膠原蛋白之表現，亦無觀察到有細胞凋亡之現象，顯示PTH (1-31)對於正常關節軟骨細胞功能應無影響，且PTH (1-31)對於關節軟骨之效應類似於PTH (1-34)的實驗結果，此外更進一步發現PTH (1-31)抑制第X型膠原蛋白表現及細胞凋亡的效果更顯著，故推論cAMP/PKA訊息路徑可能是抑制軟骨細胞走向終止分化進而抑制退化性關節炎之主要路徑；但PTH (1-34)對增加COL II表現之結果較PTH (1-31)顯著，推論PKC訊息傳遞路徑在軟骨分化亦有貢獻。

關鍵字

退化性關節炎；副甲狀腺素

並列摘要

Osteoarthritis (OA) is a degenerative joint disease and associated with changes in glycosaminoglycan (GAG) and collagen levels, but the mechanism remains to be defined. During OA progression, articular chondrocytes undergo terminal differentiation as that occurs during endochondral ossification. Previous studies showed that parathyroid hormone-related peptide (PTHrP) can suppress terminal differentiation of chondrocytes during endochondral ossification. Our previous finding emphasized that PTH (1-34) can suppress the progress of terminal differentiation in cultured human articular chondrocytes (HACs) and papain-induced OA in rats. PTH and PTHrP share the same receptor PTHR1, and subsequently stimulates both adenylate cyclase (PKA pathway) and phospholipase-C (PKC pathway). Previous reports showed that PTH (1-31) had high affinity to bind to PTHR1 and stimulated adenylate cyclase but not phospholipase-C. Therefore, in this study we hypothesized that PTH (1-31) suppress terminal differentiation of articular chondrocytes through cAMP/PKA signaling, and thus suppress OA progression. OA was induced by intra-articular injections of papain solution and cystein in the right knees of rats and left knee of each rat was as the contralateral control. The rats were divided into five groups: OA, PTH (1-34), PTH (1-31), OA+PTH (1-34), and OA+PTH (1-31). Our data showed that the PTH (1-31) treatment suppressed the papain-induced OA changes. The Safranin-O-Fast-Green staining showed that GAG level reduced in OA group, but not in the OA+ PTH (1-34) and the OA+PTH (1-31) groups. The immunohistochemistry analysis indicated that the level of COL II showed a marked increase in the OA+ PTH (1-34) and the OA+ PTH (1-31) groups; the COL X expression was less in the OA+ PTH (1-34) and the OA+ PTH (1-31) groups compared with the OA group. The level of GAG, COL II and COL X of the contralateral control cartilage were not significantly different among the five groups. In the present study, we found that PTH (1-31) and PTH (1-34) showed the similar effect on suppressing GAG loss in the papain-induced OA rat model. And, we further found that PTH (1-31) significantly suppressed the COL X expression and chondrocytes apoptosis than PTH (1-34). But, PTH (1-34) was more effective than PTH (1-31) on maintaining the COL II content in OA rats. On the other hand, PTH (1-31) did not affect the intact articular cartilage. According to the results, we suggest that the suppressive effect of PTH (1-31) on COL X expression and apoptosis in articular chondrocytes might due to cAMP/ PKA signaling pathway, but not PKC signaling pathway which may contribute to maintaining chondrocyte differentiation.