大鼠复合型放射性下颌骨骨坏死动物模型建立及观察

doi:10.3877/cma.j.issn.1674-1366.2016.01.005

目的

依据颌骨放射性骨坏死（ORNJ）新的临床诊断标准，探讨建立可模拟ORNJ不同临床分期的复合型动物模型的方法。

方法

健康SD雄性大鼠36只，随机分为6组：a₁、b₁、c₁为实验组，a₀、b₀、c₀组为对照组；根据生物学等效公式，实验组采用电子直线加速器对大鼠左侧下颌区进行照射，7 Gy/次，1次/d，照射5 d；照射后6周，处死a₁、a₀组大鼠，并拔除b₁、b₀、c₁、c₀组大鼠左侧下颌磨牙；照射12、18周后，分别处死b₁、b₀组大鼠和c₁、c₀组大鼠并取下颌骨标本。对所有标本进行组织学观察和micro-CT扫描与骨组织形态学分析。使用SPSS 18.0软件包对数据进行统计学分析。

结果

实验组照射区可见a₁组门齿生长障碍、b₁组死骨暴露、c₁组咬合错乱，化脓性感染与病理性骨折等症状；对照组无症状。组织学观察可见，实验组照射区骨髓腔与牙周膜内，炎性细胞浸润、纤维组织增生，b₁、c₁组还可见游离死骨；实验组门齿牙根与皮质骨渐进性破坏，c₁组部分大鼠门齿牙根结构消失，周围皮质骨不连续；实验组空白骨陷窝增多，骨陷窝内骨细胞减少。micro-CT结果显示：相对骨体积（BV/TV）：a₁（0.401 ± 0.053）、a₀（0.615 ± 0.037）、b₁（0.309 ± 0.041）、b₀（0.717 ± 0.040）、c₁（0.200 ± 0.026）、c₀（0.724 ± 0.041），实验组相对骨体积（BV/TV）均低于对照组；实验组间BV/TV值相比，a₁> b₁>c₁，P_a1、b1=0.012、P_b1、c1=0.034、P_c1、a1=0.004，差异有统计学意义（P<0.05）。

结论

建立了一种能够模拟出轻度无症状（a₁）、中度死骨暴露（b₁）与重度联合症状（c₁）三种不同ORNJ分型的复合型动物模型，为针对性研究ORNJ治疗方法提供了实验平台。

关键词: 骨坏死，放射性, 颌, 模型，动物, 纤维化, 骨组织形态学分析

Objective

To establish an osteoradionecrosis of jaws (ORNJ) rat model which could be used to simulate ORNJ at different clinical stages, according to the new clinical diagnostic standards.

Methods

Thirty six male SD rats were divided into 6 groups randomly. For group a₁, b₁ and c₁, the left mandible of each rat weas irradiated at doses of 7.0 Gy for 5 fractions, which were equivalent to total doses of 70 Gy. Group a₀, b₀ and c₀ were sham irradiated, serving as control groups. Six weeks after irradiation, rats in group a₀ and a₁ were sacrificed, meanwhile, left mandibular molars of rats in group b₁, b₀, c₁ and c₀ were extracted. 12 weeks and 18 weeks after irradiation, the rats in group b₀, b₁ and in group c₀, c₁ were sacrificed respectively. All the rats′ mandibles were taken. All samples were examined by clinical manifestation, micro-CT and histology methods. The data was analyzed with SPSS 18.0 software package.

Results

Alopecia and retardation of central incisor growth, at the irradiated site were seen in group a₁. Exposed necrotic bone was found in all irradiated mandibles of group b₁ and c₁. Reduced condyle volume, dental sepsis and obviously loose, necrotic bone of the irradiated mandible were found in group c₁. Fibrosis and inflammtions were found in medulla in all irradiated groups. In group b₁, small pieces of sequestrum were seen around the tooth root. And large sequestrum, pathological fracture, overwhelming fibrosis and disrupted incisor root were seen in group c₁. Significant increment of empty lacunae and decrement of osteocytes were observed in all irradiated groups, accompanying with the decreased BV/TV: a₁ (0.401 ± 0.053) , a₀ (0.615 ± 0.037) , b₁ (0.309 ± 0.041) , b₀ (0.717 ± 0.040) , c₁ (0.200 ± 0.026) , c₀ (0.724 ± 0.041) , P_a1、b1= 0.012, P_b1、c1= 0.034, P_c1、a1= 0.004, a₁>b₁>c₁, results were accepted as statistically significant at a value of P<0.05.

Conclusions

A combined rat model that can stimulate three different stages of ORNJ was successfully established. This model could be served as a new platform for future studies on pathogenesis and treatment of ORNJ.

Key words: Osteoradionecrosis, Jaw, Models, Animal, Fibrosis, Bone histomorphometric analysis

图1 SD大鼠放疗处理方法

图2 放疗后SD大鼠照射区的大体表现

表1 各组SD大鼠micro-CT相对骨体积分析结果

组别	鼠数（只）	BV/TV(%)	P值
a₁组	6	0.401 ± 0.053	P _a1、a0=0.045^a	P _a0、b0 = 0.115^b	P _a1、b1 = 0.012^c
a₀组	6	0.615 ± 0.037	P _a1、a0=0.045^a	P _a0、b0 = 0.115^b	P _a1、b1 = 0.012^c
b₁组	6	0.309 ± 0.041	P _b1、b0=0.002^a	P _b0、c0 = 0.914^b	P _b1、c1 = 0.034^c
b₀组	6	0.717 ± 0.040	P _b1、b0=0.002^a	P _b0、c0 = 0.914^b	P _b1、c1 = 0.034^c
c₁组	6	0.200 ± 0.026	P _c1、c0<0.001^a	P _c0、a0=0.096^b	P _c1、a1=0.004^c
c₀组	6	0.724 ± 0.041	P _c1、c0<0.001^a	P _c0、a0=0.096^b	P _c1、a1=0.004^c

表1 各组SD大鼠micro-CT相对骨体积分析结果

组别	鼠数（只）	BV/TV(%)	P值
a₁组	6	0.401 ± 0.053	P _a1、a0=0.045^a	P _a0、b0 = 0.115^b	P _a1、b1 = 0.012^c
a₀组	6	0.615 ± 0.037	P _a1、a0=0.045^a	P _a0、b0 = 0.115^b	P _a1、b1 = 0.012^c
b₁组	6	0.309 ± 0.041	P _b1、b0=0.002^a	P _b0、c0 = 0.914^b	P _b1、c1 = 0.034^c
b₀组	6	0.717 ± 0.040	P _b1、b0=0.002^a	P _b0、c0 = 0.914^b	P _b1、c1 = 0.034^c
c₁组	6	0.200 ± 0.026	P _c1、c0<0.001^a	P _c0、a0=0.096^b	P _c1、a1=0.004^c
c₀组	6	0.724 ± 0.041	P _c1、c0<0.001^a	P _c0、a0=0.096^b	P _c1、a1=0.004^c

图3 各组SD大鼠左侧下颌骨micro-CT图像

图4 SD大鼠放疗后照射区皮质骨骨细胞凋亡与髓腔纤维化的组织学观察

图5 SD大鼠放疗后照射区渐进性骨破坏的组织学观察（苏木精-伊红）

表2 各组SD大鼠下颌骨标本骨陷窝内骨细胞与空白骨陷窝计数

组别	鼠数（只）	骨陷窝内骨细胞（个）	空白骨陷窝（个）	P值
组别	鼠数（只）	骨陷窝内骨细胞（个）	空白骨陷窝（个）	骨陷窝内骨细胞			空白骨陷窝
a₁组	6	28.67 ± 3.15	3.57 ± 2.76	P _a1、a0 = 0.002^a	P _a0、b0=0.232^b	P _a1、b1 = 0.044^c	P _a1、a0=0.007^a	P _a0、b0 = 0.849^b	P _a1、b1 = 0.340^c
a₀组	6	62.00 ± 4.66	0.68 ± 0.95	P _a1、a0 = 0.002^a	P _a0、b0=0.232^b	P _a1、b1 = 0.044^c	P _a1、a0=0.007^a	P _a0、b0 = 0.849^b	P _a1、b1 = 0.340^c
b₁组	6	41.00 ± 6.06	5.16 ± 4.30	P _b1、b0 = 0.014^a	P _b0、c0=0.092^b	P _b1、c1 = 0.762^c	P _b1、b0=0.013^a	P _b0、c0 = 0.459^b	P _b1、c1 = 0.866^c
b₀组	6	68.83 ± 3.39	0.58 ± 1.05	P _b1、b0 = 0.014^a	P _b0、c0=0.092^b	P _b1、c1 = 0.762^c	P _b1、b0=0.013^a	P _b0、c0 = 0.459^b	P _b1、c1 = 0.866^c
c₁组	6	41.00 ± 4.51	4.37 ± 3.70	P _c1、c0 = 0.030^a	P _c0、a0 = 0.711^b	P _c1、a1=0.026^c	P _c1、c0=0.014^a	P _c0、a0=0.315^b	P _c1、a1 = 0.519^c
c₀组	6	59.17 ± 4.75	0.87 ± 0.61	P _c1、c0 = 0.030^a	P _c0、a0 = 0.711^b	P _c1、a1=0.026^c	P _c1、c0=0.014^a	P _c0、a0=0.315^b	P _c1、a1 = 0.519^c

表2 各组SD大鼠下颌骨标本骨陷窝内骨细胞与空白骨陷窝计数

组别	鼠数（只）	骨陷窝内骨细胞（个）	空白骨陷窝（个）	P值
组别	鼠数（只）	骨陷窝内骨细胞（个）	空白骨陷窝（个）	骨陷窝内骨细胞			空白骨陷窝
a₁组	6	28.67 ± 3.15	3.57 ± 2.76	P _a1、a0 = 0.002^a	P _a0、b0=0.232^b	P _a1、b1 = 0.044^c	P _a1、a0=0.007^a	P _a0、b0 = 0.849^b	P _a1、b1 = 0.340^c
a₀组	6	62.00 ± 4.66	0.68 ± 0.95	P _a1、a0 = 0.002^a	P _a0、b0=0.232^b	P _a1、b1 = 0.044^c	P _a1、a0=0.007^a	P _a0、b0 = 0.849^b	P _a1、b1 = 0.340^c
b₁组	6	41.00 ± 6.06	5.16 ± 4.30	P _b1、b0 = 0.014^a	P _b0、c0=0.092^b	P _b1、c1 = 0.762^c	P _b1、b0=0.013^a	P _b0、c0 = 0.459^b	P _b1、c1 = 0.866^c
b₀组	6	68.83 ± 3.39	0.58 ± 1.05	P _b1、b0 = 0.014^a	P _b0、c0=0.092^b	P _b1、c1 = 0.762^c	P _b1、b0=0.013^a	P _b0、c0 = 0.459^b	P _b1、c1 = 0.866^c
c₁组	6	41.00 ± 4.51	4.37 ± 3.70	P _c1、c0 = 0.030^a	P _c0、a0 = 0.711^b	P _c1、a1=0.026^c	P _c1、c0=0.014^a	P _c0、a0=0.315^b	P _c1、a1 = 0.519^c
c₀组	6	59.17 ± 4.75	0.87 ± 0.61	P _c1、c0 = 0.030^a	P _c0、a0 = 0.711^b	P _c1、a1=0.026^c	P _c1、c0=0.014^a	P _c0、a0=0.315^b	P _c1、a1 = 0.519^c

[1]	Ko C, Citrin D. Radiotherapy for the management of locally advanced squamous cell carcinoma of the head and neck[J]. Oral Dis，2009，15(2):121-132.
[2]	Nabil S, Samman N. Incidence and prevention of osteoradionecrosis after dental extraction in irradiated patients：a systematic review[J]. Int J Oral Maxillofac Surg，2011，40(3):229-243.
[3]	Harris M. The conservative management of osteoradionecrosis of the mandible with ultrasound therapy[J]. Br J Oral Maxillofac Surg，1992，30(5):313-318.
[4]	Epstein JB, Wong FL, Stevenson-Moore P. Osteoradionecrosis：clinical experience and a proposal for classification[J]. J Oral Maxillofac Surg，1987，45(2):104-110.
[5]	Marx RE. Osteoradionecrosis：a new concept of its pathophysiology[J]. J Oral Maxillofac Surg，1983，41(5):283-288.
[6]	Delanian S, Martin M, Housset M，et al. Iatrogenic fibrosis in cancerol-ogy（1）：descriptive and physiopathological aspects[J]. Bull Cancer，1993，80(3):192-201.
[7]	Delanian S, Lefaix JL. The radiation-induced fibroatrophic process：therapeutic perspective via the antioxidant pathway[J]. Radiother Oncol，2004，73(2):119-131.
[8]	Chiao TB, Lee AJ. Role of pentoxifylline and vitamin E in attenuationof radiation-induced fibrosis[J]. Ann Pharmacother，2005，39(3):516-522.
[9]	Delanian S, Depondt J, Lefaix JL，et al. Major healing of refractory mandible osteoradionecrosis after treatment combining pentoxifylline and tocopherol：a phaseⅡ trial[J]. Head Neck，2005，27(2):114-123.
[10]	Delanian S, Chatel C, Porcher R，et al. Complete restoration of refractory mandibular osteoradionecrosis by prolonged treatment with a pentoxifylline-tocopherol-clodronate combination（PENTOCLO）：a phaseⅡ trial[J]. Int J Radiat Oncol Biol Phys，2011，80(3):832-839.
[11]	McLeod NM, Pratt CA, Mellor TK，et al. Pentoxifylline and tocopherol in the management of patients with osteoradionecrosis，the Portsmouth experience[J]. Br J Oral Maxillofac Surg，2012，50(1):41-44.
[12]	Lyons A, Osher J, Warner E，et al. Osteoradionecrosis—a review of current concepts in defining the extent of the disease and a new classification proposal[J]. Br J Oral Maxillofac Surg，2014，52(5):392-395.
[13]	Sønstevold T, Johannessen AC, Stuhr L. A rat model of radiation injury in the mandibular area[J]. Radiat Oncol，2015(10):129.
[14]	Tchanque-Fossuo CN, Monson LA, Farberg AS，et al. Dose-response effect of human equivalent radiation in the murine mandible：partⅠ. A histomorphometric assessment[J]. Plast Reconstr Surg，2011，128(1):114-121.
[15]	Cohen M, Nishimura I, Tamplen M，et al. Animal model of radiogenic bone damage to study mandibular osteoradionecrosis[J]. Am J Otolaryngol，2011，32(4):291-300.
[16]	Fenner M, Park J, Schulz N，et al. Validation of histologic changes induced by external irradiation in mandibular bone. An experimental animal model[J]. J Craniomaxillofac Surg，2010，38(1):47-53.
[17]	Regaud C. Sur la necrose des os attenté par un processus cancereux et traites par les radiaions[J]. Compt Rend Soc Biol，1922(87):427.
[18]	Meyer OT, Dannenberg AM Jr. Radiation，infection，and macrophage function. Ⅱ. Effect of whole body radiation on the number of pulmonary alveolar macrophages and their levels of hydrolytic enzymes[J]. J Reticuloendothel Soc，1970，7(1):79-90.
[19]	Spiegelberg L, Djasim UM, van Neck HW，et al. Hyperbaric oxygen therapy in the management of radiation-induced injury in the head and neck region：a review of the literature[J]. J Oral Maxillofac Surg，2010，68(8):1732-1739.
[20]	Marx RE. A new concept in the treatment of osteoradionecrosis[J]. J Oral Maxillofac Surg，1983，41(6):351-357.
[21]	Bennett MH, Feldmeier J, Hampson N，et al. Hyperbaric oxygen therapy for late radiation tissue injury[J]. Cochrane Database Syst Rev，2012(5):CD005005.
[22]	Madrid C, Abarca M, Bouferrache K. Osteoradionecrosis：an update[J]. Oral Oncol，2010，46(6):471-474.
[23]	Chrcanovic BR, Reher P, Sousa AA，et al. Osteoradionecrosis of the jaws—a current overview—part 1：physiopathology and risk and predisposing factors[J]. J Oral Maxillofac Surg，2010，14(1):3-16.
[24]	Niehoff P, Springer IN, Açil Y，et al. HDR brachytherapy irradiation of the jaw - as a new experimental model of radiogenic bone damage[J]. J Craniomaxillofac Surg，2008，36(4):203-209.
[25]	Tamplen M, Trapp K, Nishimura I，et al. Standardized analysis of mandibular osteoradionecrosis in a rat model[J]. Otolaryngol Head Neck Surg，2011，145(3):404-410.
[26]	Armin BB, Hokugo A, Nishimura I，et al. Brachytherapy-mediated bone damage in a tat model investigating maxillary osteoradionecrosis[J]. Arch Otolaryngol Head Neck Surg，2012，138(2):167-171.
[27]	Kurihashi T, Iwata H, Nasu M，et al. Experimental study on wound healing of alveolar bone sockets in the rat maxilla after x-ray irradiation[J]. Odontology，2002，90(1):35-42.
[28]	Gallagher KK, Deshpande S, Tchanque-Fossuo CN，et al. Role of parathyroid hormone therapy in reversing radiationinduced nonunion and normalization of radiomorphometrics in a murine mandibular model of distraction osteogenesis[J]. Head Neck，2013，35(12):1732-1737.
[29]	Lee W, Lee BD, Lee KK，et al. A magnetic resonance imaging study on changes in rat mandibular bone marrow and pulp tissue after high-dose irradiation[J]. Imaging Sci Dent，2014，44(1):43-52.
[30]	Ardran GM. Bone destruction not demonstrable by radiography[J]. Br J Radiol，1951，24(278):107-109.

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Establishment and observation of a combined rat model of osteoradionecrosis

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Establishment and observation of a combined rat model of osteoradionecrosis