- 學位論文
前十字韌帶斷裂後的早期介入被動及主動運動治療對於軟骨的保護作用：兔子模型
The protective effect of early passive and active therapeutic exercise on articular cartilage after anterior cruciate ligament transection in rabbits
摘要
背景：前十字韌帶斷裂是否需立即接受重建手術，對於未來產生退化性關節炎（Osteoarthritis, OA）目前仍有爭議。有兩個主要因子將直接影響軟骨的生成與退化，包含：（1）關節腔內生理變化、（2）關節表面承受力學後的變異。而在韌帶斷裂後，關節腔內產生的發炎反應及不穩定也許會促使軟骨退化。適當的運動治療在軟骨退化的早期介入，被證實對於軟骨的修復有所幫助，主要因為運動改變軟骨生長的微環境（microenvironments）。然而，儘管臨床上目前有許多前十字韌帶斷裂的治療方式，包含：保守的復健治療與手術治療，但對於關節軟骨的退化仍無法避免。此外，臨床診斷退化性關節炎常用為特殊檢查、X光或核磁共振影像，然而往往被評估到差異時，大部分已經是造成不可逆的軟骨傷害。因此，如何做好早期的預防是決定未來軟骨的重要關鍵。目前臨床上評估，共同評估方式為使用自我評估問卷、理學檢查或影像學檢查分析預後，卻鮮少研究探討關節內細胞與細胞外基質的變化。此外，前十字韌帶在不執行或等待執行重建術時，該執行何種運動治療，將對於軟骨具有保護性功能的可行性也尚未被探討。 目的：先建立兔子前十字韌帶斷裂模型，再執行不同的運動模式（被動式、主動式、合併式運動）探討對於軟骨細胞與細胞外基質的保護效益。 材料與方法：30隻公紐西蘭大白兔，手術將右膝前十字韌帶做人工切除（anterior cruciate ligament transection, ACLT），左膝則為控制組（僅將皮膚及關節囊切開，無切除韌帶）。動物隨機分配到四個治療組。少活動組（sedentary, SED）：未接受任何運動治療，僅於籠子內自由活動；連續被動運動組（continuous passive motion exercise, CPM）：於術後一週內介入連續性被動運動連續7天，每天15分鐘，治療後兔子回歸籠子自由活動；主動的跑步機運動組（active treadmill exercise, TRE）：兔子於術後第三、四週介入輕至中度強度跑步機運動治療，每次10分鐘，每週3天；被動與跑步機運動混合組（CPM+TRE）：先於第一週執行連續性被動運動，隨後兔子回歸籠子自由活動，再於第三週執行跑步機運動，連續執行兩週輕至中度強度跑步機運動治療。所有兔子於第五週犧牲，進行全膝軟骨分析，觀察組織學染色（細胞型態、細胞外基質排列與含量）、量化退化性關節炎分數、發炎性反應之生化分析。 結果：巨觀方面：磨損的嚴重情形為組TRE最嚴重，接著依序為SED、CPM+TRE、CPM組，此外，CPM組在關節炎巨觀量表中顯著性的優於其他三組。組織學方面：TRE組的軟骨退化程度最為嚴重，接著依序為SED、CPM+TRE組，而CPM組狀態最為良好。其中，在TRE組與SED組細胞型態（H&E staining）中顯示關節表面磨損、細胞排列不規則、軟骨細胞數量變少、軟骨細胞產生聚集效應（關節炎的前兆之一）、細胞外基質排列雜亂及糖胺聚醣（glycosaminoglycan, GAG）也有崩解的現象；相反地，若早期給予CPM則可保護軟骨關節表面，使其較為平滑、細胞型態及細胞外基質排列整齊和維持軟骨各層的細胞及組織含量。值得一提的是，若太早給主動的跑步機運動（亦即CPM+TRE組）發現關節表面已開始出現不規則、細胞型態有趨向關節炎的現象，細胞外基質排列開始不規則，其糖胺聚醣也開始流失。在發炎因子TNF-α及細胞凋亡因子Caspase-3的結果中，在早期執行CPM可以減少發炎反應，有較少的凋亡細胞存在，顯示達到保護軟骨的效果；但在其他組別則出現軟骨表淺層及中層有TNF-α與Caspase-3的反應，其中以TRE組表現最為明顯。 結論：當前十字韌帶斷裂後，早期介入CPM具有保護軟骨的作用，可保持關節軟骨表面的平整性、細胞的型態、減低軟骨發炎反應、細胞凋亡及維持細胞外基質的結構與含量；相對地，在尚未進行認帶重建手術前，無論是少活動或進行主動承重運動，有加速軟骨發炎、軟骨細胞凋亡及細胞外基質崩解的現象。 臨床意義：前十字韌帶斷裂且無執行（或等待執行）韌帶重建術的患者，建議積極在早期進行CPM保護軟骨的生長狀態。
並列摘要
Background: It remains controversial that whether or not exerting an anterior cruciate ligament (ACL) reconstruction surgery immediately after an ACL rupture that maybe lead to osteoarthritis (OA). Two main factors develop the cartilage repair or cartilage degradation: (1) changes of intra-articular physiological condition, (2) and changes of the mechanical properties on the articular surface. Once ligament damage, a cascade of corresponding in situ joint inflammation and instability in the knee joint may deteriorate cartilage degradation. Appropriate exercise therapies in an early intervention, changing the in situ microenvironments in the joint, have been suggested to promote healthy cartilage growth. Although numerous clinical treatments for ACL rupture were adopted by either conservative treatments or surgical treatments, but those managements for preventing the occurrence of OA is deemed to be limited. In clinical practice, the common assessments for outcomes typically comprise self-reported questionnaires, physical examinations or imaging analysis. However, a few studies investigate the changes of extracellular matrix and cell morphology of articular cartilage after ACL injury. Furthermore, whether or not (or waiting) to perform ACL reconstruction, the implementation of which exercise therapy is yet to be investigated for the feasibility protection of cartilage. Purpose: We first establish ACL rupture in the rabbit model, and subsequently perform diverse kinds of exercise treatments (i.e., continuous passive motion exercise, CPM; active treadmill exercise, TRE; or combination exercise) to investigate the effects of the protection of chondrocytes and extracellular matrix. Material & Methods: A total of 30 rabbits were used in this study. All animals received the ACL transection (ACLT) of right knee. Left knee was for the sham group (only open and close the joint without ACLT). The animals were randomly allocated to four groups：(1) SED group, rabbits just kept in the cage. (2) CPM group, rabbits performed the CPM exercise for continuously 7 days, 15 min/ day post ACLT. (3) TRE group, rabbits performed active treadmill exercise at the third week for two weeks after ACLT (10 min/day, 3 days/week, 2 weeks). (4) CPM+TRE group, rabbits performed the CPM at the first week and followed by active treadmill exercise at the third week for two weeks (the exercise protocol as aforementioned). All animals sacrificed at the fifth week. All knees were taken out for whole knee evaluations including gross appearance, histology, and OA quantitative scores as well as inflammatory reactions. Results: Regarding gross appearance, the TRE group had the most cartilage abrasion, followed by SED, CPM+TRE, and CPM. In addition, the CPM+TRE group was better than the SED group. The CPM group demonstrated the best cartilage smooth among others. Regarding histological aspects, the TRE group showed the most severe cartilage degeneration. Furthermore, the CPM+TRE group was better than the SED group. The CPM group showed the modest degeneration. On the basis of H&E staining, the TRE group showed much cartilage abrasion, disorganized, decreased of cartilage cells, appeared chondrocytes cluster, and decreased of GAG. In contrast, CPM in the early stage can protect the cartilage. The CPM group had the smoothest surface of cartilage, more oriented arrangement of cells, and had the most of GAG. In addition, loading in the early stage (the CPM+TRE group) will lead to osteoarthritis. We also found that the CPM group had the least TNF-α and caspase-3. However, the TRE group had the most TNF-α and caspase-3, particularly in superficial and middle layers of the cartilage. Conclusion：CPM in the early stage after ACL injury provides the protection of cartilage. In contrast, active treadmill exercise may lead to osteoarthritis.