探討溫感性水膠包覆高濃度血小板血漿及可塑型人工骨對於萎縮性不癒合骨折之療效

Study the Treatment of Atrophic Nonunion by Using Thermosensitive Hydrogel Encapsulating Platelet-Rich Plasma and Bone Substitute

何柏賢(Po-Hsien Ho)

指導教授 ： 陳志華

共同指導教授 ： 莊爾元(Er-Yuan Chuang)

臺北醫學大學/醫學工程學院/生醫材料暨組織工程研究所/碩士(2017年)

摘要

在臨床上，骨折後經過9個月無法癒合而在最後3個月完全無骨頭癒
合徵象稱為骨折不癒合，發生機率約佔骨折病患6%。然而，在這6%
的病患中會有46%的患者由於細菌感染、骨折缺損區域過大或是骨膜
受損造成萎縮型不癒合。一旦發生，常需多次手術治療。傳統之治療
方式包含多次手術及自體骨移植等。此種治療方式將會耗費治療時間
及成本，且自體骨移植之來源有限，會造成取骨處疼痛出血等各種併
發症。本研究目的是使用高濃度血小板血漿和骨替代物包覆在溫感性
水膠中來治療骨折萎縮型不癒合。在體外實驗中，我們發現高濃度血
小板血漿配合骨替代物，有助於MG-63細胞的生長。在體內實驗中，
我們利用大鼠骨折萎縮型不癒合的動物模式，並用溫感性水膠包覆高
濃度血小板血漿和骨替代物治療。溫感性水膠的逆溫性，可以在室溫
下形成膠狀，可有效填補補缺損的部位，碰到體溫後會固化並緩慢降
解並釋放高濃度血小板血漿內之生長因子，進而治療骨折萎縮型不癒
合。在影像分析和組織學分析也證實了骨頭的癒合。溫感性水膠包覆
高濃度血小板和骨替代物，確實可以促進骨折萎縮型不癒合的生長。
因此希望透過本次研究成果進而提供臨床上新的治療方法。

關鍵字

骨折不癒合 ； 萎縮型不癒合 ； 高濃度血小板血漿 ； 溫感性水膠 ； 骨替代物

並列摘要

Background:
Atrophic nonunion generally occurs in diaphysis of long bone fracture, which is a critical complication of fractures in clinical practice. The incidence of atrophic nonunion has been estimated to account for about 46% of nonunion cases, caused by bacterial infections, bone defects or periosteum damages. Atrophic nonunion represents a formidable challenge for orthopaedic surgeons. Their successful treatment is frequently time consuming, and requires numerous resources. Atrophic nonunion is associated with chronic pain, functional, psychosocial disability and causing an economic burden for patients. The purpose of this study therefore was to investigate the current advances in the enhancement of the fracture healing response and treatment of atrophic nonunion and provides a therapeutic opportunity.
Aims:
The aim of this study was to use the combination of platelet-rich plasma (PRP) and bone substitute as a technology platform to improve fracture healing by applying a thermosensitive hydrogel carrier system which could be attached within the atrophic nonunion part for further slower control-release.
Material and Methods:
10 ml of whole blood was collected from 10 healthy Sprague Dawley rats by cardiac puncture, in the presence of K2EDTA anticoagulant. Blood was centrifuged twice (1000×g for a period of 9 min at 4oC and 3000×g for a period of 20 min at 4oC) to get PRP. Biphasic calcium phosphate (BCP) bone substitute was purchased from Wiltrom medical company. The Industrial Technology Research Institute (ITRI) provided the thermosensitive hydrogel, also referred to as mPEG-PLGA/Box/mPEG-PLGA, which was synthesized by the ring-opening polymerization of monomers. The in vitro study, MG-63 cells was treated with nothing (control), PRP, BCP or PRP + BCP, and then analysis by MTT assay. In the in vivo study, the hydrogel was used to encapsulate nothing (control), PRP, BCP and PRP + BCP and then implanted to the animal model (N=4 per group) of atrophic nonunion. The bone defect of femur part was evaluated by micro-CT and stained with H & E staining 4 weeks after implanting the material to assess the extracellular matrix (ECM).
Results:
The in vitro study showed that combining PRP with bone substitute, could assist MG-63 proliferation as found by MTT assay. In the in vivo study, micro-CT data showed that PRP + BCP encapsulated in thermosensitive hydrogel could significantly promote bone regeneration of atrophic nonunion bone defect in the rat model (N=4 per group). The histological by H&E staining revealed that the combination of PRP + BCP encapsulated in thermosensitive hydrogel group led to enhanced proliferation of ECM.
Conclusions:
Combining PRP and BCP for encapsulation by thermosensitive hydrogel effectively improved the atrophic nonunion. This system can therefore be used as a tissue engineering strategy and provides a therapeutic opportunity for promoting atrophic nonunion regeneration that can have potentials applied in the for the future clinical use.

並列關鍵字

atrophic nonunion；bone regeneration；platelet-rich plasma；bone substitute；thermosensitive hydrogel

參考文獻

1. Creevey, W., et al., Fracture and dislocation classification compendium-2007. J Orthop Trauma, 2007. 21(10 Supplement): p. S1-S133.

2. Marsh, J., et al., Fracture and dislocation classification compendium-2007: Orthopaedic Trauma Association classification, database and outcomes committee. 2007, LWW.

3. Koval, K.J. and J.D. Zuckerman, Handbook of fractures. 2006.

4. Tzioupis, C. and P.V. Giannoudis, Prevalence of long-bone non-unions. Injury, 2007. 38: p. S3-S9.

5. Burchardt, H. and W. Enneking, Transplantation of bone. Surgical Clinics of North America, 1978. 58(2): p. 403-427.