第二型成骨蛋白及雙氯苯雙胍己烷應用於氧化鋯人工牙根之骨整合能力與抗菌效果評估

Abstract

鈦金屬是目前市面上牙科植體材料的主流，其擁有良好的生物相容性、抗腐蝕強度、機械物理性質和優異的骨整合作用(Osseointegration)。但鈦金屬同時也存在著金屬材料本身所無法改善的缺陷，例如金屬色澤在前牙區較不美觀、鈦金屬長時間存在人體所造成鈦金屬離子在人體中的累積效應、金屬離子造成病人的過敏反應等等問題。因此近幾年來，市面上已經出現以陶瓷製作的植體，其目的在於取代鈦金屬植體無法改善的缺陷。而植體的成功與否，又以骨整合作用佔關鍵地位，特別是陶瓷植體目前皆為一件式設計(One piece)，較易提早承受咬合壓力，因此藉由改良的表面處理技術促進骨整合在早期癒合(Early healing phase)便發生對陶瓷植體是否成功將影響甚鉅。進一步探索植體失敗的因素中，植體周圍炎(Peri-implantitis)亦是常見的原因之一，若能減少植體周圍炎的發生，應也有助於骨整合作用的發生。 本實驗的目的是評估本地生產的之氧化鋯植體及鈦金屬植體在動物實驗模式下，藉由台大化工系陳賢燁教授團隊研發出來的新式表面鍍膜的方法進行植體表面改質，觀察對骨整合作用及抗菌效果的影響。並利用未脫鈣磨片的標本處理技術，搭配骨螢光標定，觀察氧化鋯植體與鈦金屬植體骨癒合模式之差異。 本研究組別共分為七組，第一組為市售氧化鋯植體(Zr-I3608)；第二組為自行研發單純進行鍍膜處理之氧化鋯植體；第三組為自行研發以浸泡方式進行鍍膜處理之氧化鋯植體，有螺紋處接枝第二型成骨蛋白(Bone morphogenetic protein II)，螺紋以上接枝雙氯苯雙胍己烷(Chlorhexidine)；第四組為自行研發以鍍膜方式進行表面改質之氧化鋯植體，有螺紋處接枝第二型成骨蛋白，螺紋以上接枝雙氯苯雙胍己烷；第五組為自行研發以鍍膜方式進行表面改質之氧化鋯植體，有螺紋處接枝史他汀類藥物(Simvastatin)，螺紋以上接枝雙氯苯雙胍己烷；第六組為市售鈦金屬植體(A1012-3508)；第七組為自行研發以鍍膜方式進行表面改質之鈦金屬植體，有螺紋處接枝第二型成骨蛋白，螺紋以上接枝雙氯苯雙胍己烷。觀測的時間點依序為植體植入後第12週、第8週、第4週和第1週。本實驗係採用了5隻米格魯犬(Beagle dog)，隨機植入下顎骨雙側無牙區，植牙手術進行時，以不同顏色之骨染劑進行骨螢光標定，以觀察不同時間點之骨新生。實驗期間以植體動搖度分析儀Periotest進行非侵入性之植體穩定度測試。於動物犧牲後取得標本，並進行植體穩定度 Periotest(PTV)、放射線影像、斷層掃描、組織切片及骨頭螢光標定觀察等方式分析植體周圍鍍膜處理對骨新生程度及抗菌效果的影響。 實驗結果的部份，牙菌斑堆積指數(Mombelli index)只有4週組之第四組以鍍膜方式進行表面改質之氧化鋯植體與第七組以鍍膜方式進行表面改質之鈦金屬植體有顯著差異(p=0.027)，其餘組別之各時間點均無顯著差異。植體動搖度(PTV)只有4週組之第一組市售氧化鋯植體(Zr-I3608)與第四組以鍍膜方式進行表面改質之氧化鋯植體有顯著差異(p=0.028)，其餘組別之各時間點均無顯著差異。放射線影像之齒槽骨吸收率(BRR)分析各組各時間點並無顯著差異。電腦斷層掃描分析之骨量、骨-植體接觸面積與礦物質化骨密度亦無顯著差異，然可觀察到鈦金屬植體之表現皆較氧化鋯植體為優。未脫鈣磨片之骨-植體接觸比(BIC)只有第一組之1週組與4週組(p=0.038)、第七組之8週組與12週組(p=0.027)、第三組與第七組之所有時間點(p=0.026)有顯著差異，其餘組別之時間點均無顯著差異。骨螢光標定影像、未脫鈣磨片與脫鈣染色玻片的觀察顯示兩種植體具有相似之骨新生過程，然鍍膜接枝生醫材料之組別較對照組而言並無較顯著之骨新生效果。 本次實驗結果可見氧化鋯植體與鈦金屬植體具有類似之生物相容性與骨癒合效果，然其表現較不穩定。而鍍膜接枝生醫材料於影像學上的觀察發現確有其效果，但於實驗數據上，或因樣本數少，並未達到統計學上的顯著差異。期待此氧化鋯植牙及該鍍膜接枝技術能進一步改進以有更好之臨床應用與發展。

Dental-implant-related prosthesis and dentures are already very common in the modern treatment projects. And titanium is the mainstream of implant materials. It has good biocompatibility, corrosion-resistant strength, mechanical and physical properties and excellent osseointegration. But there are also some defects that cannot be improved because of it’s a metal material like metal color exposure in esthetic area, titanium-ion accumulation in human body, titanium-related hypersensitivity and Galvanic effect. In recent years, there are dental implants made of ceramic. And osseointegration plays an important role in success of dental implant. Especially, all the ceramic dental implant is one piece designed and more likely to withstand an early occlusion force. Thus early healing process has great influence. Research goal of this study is to evaluate effects of BMP-2 and chlorhexidine in osseointegration and antibacterial of zirconia implant. Surface treatment of the titanium and zirconia implant was performed with parylene coating. We observed the un-decalcified group and decalcified group with microscope to evaluate the effects on osseointegration and antibacterial. According to the research protocol, we have seven groups: zirconia implant blank control group (group 1, n = 8), zirconia implant parylene coating only group (group 2, n = 8), zirconia implant bathing with BMP-2 and chlorhexidine group (group 3, n = 3), zirconia implant parylene coating with BMP-2 and chlorhexidine group (group 4, n = 11), zirconia implant parylene coating with statin and chlorhexidine group (group 5, n = 11), titanium implant blank control group (group 6, n = 8), titanium implant parylene coating with BMP-2 and chlorhexidine group (group 7, n = 11). Five 1-year-old Beagle dogs were enrolled in this research protocol. We removed 4 premolars and the first molar of the dogs, waited 6 to 8 weeks for extraction wound healing. Then implant surgery was performed on edentulous ridge both left side and ride with randomized assignment. There are four time points surgery, namely 12 weeks, 8 weeks, 4 weeks and 1 week. During the surgery, implant stability was measured with Periotest, dental plaque was recorded with Mombelli index and subcutaneous bone dye was injected. After sacrifice, radiographic analysis, computed-tomography analysis, biopsy interpretation, fluorescence labeling was performed and observed with microscope. Result of the study showed significant difference between group 4 and group 7 at 4-week time point in Mombelli index (p = 0.027); group 1 and group 4 at 4-week time point in Periotest value (p = 0.028); 1-week time point and 4-week time point in gropu 1 (p = 0.038), 8-week time point and 12-week time point in group 7 (p = 0.027), group 3 and group 7 (p = 0.026) in bone-implant contact. No significant difference was found in bone resorption rate, bone volumn, intersection surface and bone mineralized density. In the fluorescein-labeled bone section, un-decalcified sample with Stevenel’s blue and Alizarin red and decalcified sample with H E stain, we found there was similar bone healing pattern between titanium implant and zirconia implant. Though there was no significant difference, titanium implant group showed better osseointegration and worse anti-plaque attachment effect than zirconia implant group. And there was no significant improvement in parylene coating group with BMP-2, statin and chlorhexidine in osseointegration and anti-bacterial effect though image study showed developmental potential. Based on the finding of the results, we concluded zirconia implant had good biocompatibility and osseointegration. Parylene coating with or without BMP-2, statin and chlorhexidine showed improvement in image study though there was no significant difference. Further investigation on zirconia implant and multi-functional parylene coating may be needed for advanced application and development in the future.