胫骨内接种建立乳腺癌骨转移大鼠模型的特点

目的研究胫骨内接种MRMT-1细胞制作的乳腺癌骨转移大鼠模型在行为学、影像学、核医学、病理学和分子生物学等方面的特点。方法使用雌性SD大鼠,随机分为假手术组和模型组,使用胫骨内注射法制成乳腺癌骨转移模型。造模后第19天时进行疼痛测定;第21天取材,测定肿瘤体积,通过影像技术评估骨质缺损程度,核医学测定骨矿物质含量(BMC)和骨密度(BMD),HE染色观察形态,抗酒石酸酸性磷酸酶(TRAP)染色并计数破骨细胞,免疫组化法测定增殖细胞核抗原(PCNA)、护骨素(OPG)和核因子kB受体活化因子配体(RANKL),荧光实时定量RT-PCR测定甲状旁腺激素相关蛋白(PTHrP)。结果模型组在造模后第19天已出现机械痛觉超敏、机械痛觉过敏和热痛觉过敏(P0.01)。第21天取材后胫骨影像评分升高(P0.01),BMD下降(P0.05);肉眼观察肿瘤生长明显(P0.01),镜下可见溶骨病变为主的混合性骨质破坏;破骨细胞数量和活性增加(P0.01),PTHrP、OPG水平与OPG/RANKL比值均下降(P0.05、P0.01),而RANKL无明显变化。结论乳腺癌骨转移大鼠模型具有疼痛和骨质破坏的表现,但未表现出PTHrP和RANKL升高,其损伤途径是通过抑制OPG破坏了OPG-RANKL-RANK系统的平衡,引起破骨细胞过度激活,造成骨吸收作用亢进。

Characteristics of rat model of bone metastasis from breast cancer

Objective To study the eharacteristies of rat model of bone metastasis from breast cancer, which was de- veloped through intra-tibial injection of MRMT-1 rat breast cancer cells. Method Female Sprague-Dawley rats were ran- domized into sham operation group and model group. Sham operation group was intra-tibially injected with normal saline, while model group was injected with the MRMT-1 cells to develop model. On day 19 after operation, allodynia and hyper- algesia were measured. All rats were killed on day 21, and then tibias were collected to measure the volume of tumors, as bone mineral density （BMD） and bone mineral capacity （BMC） were also determined. After HE staining, tartrate-resist-ant acid phosphatase （TRAP） staining and immunohistochemistry staining, the bone pathological changes, osteoclast count, as well as semi-quantitatively measure expressions of proliferating cell nuclear antigen （ PCNA）, osteoprotegerin （OPG） and receptor activator of nuclear factor-KB ligand （RANKL） were observed and investigated, respectively. By re- al-time quantitative RT-PCR, expression of parathyroid hormone related protein （PTHrP） was determined. Result Rats receiving intra-tibial injections of MRMT-1 cells displayed the development of mechanical allodynia and hyperalgesia （P 〈 0. 05 or P 〈0. 01 ）. Bone destruction was found by image examination （P 〈0. 05 or P 〈0. 01 ）. Mixed bone lesion was ob- served through HE staining sections. Osteoclast enhanced （ P 〈 0. 01 ）, PTHrP and OPG decreased （ P 〈 0. 05 or P 〈 0. 01 ）, but RANKL remained unchanged. Conclusion The model of bone metastasis from breast cancer shows character- istics of cancer pain and bone lesion. The injury mechanism is to break the balance of OPG-RANKL-RANK system by in- hibiting OPG, these changes would cause excessive activation of osteoclasts, which then induces hyperfunction of bone re- sorption.