趋化因子CCL20在促进骨巨细胞瘤增殖、迁移及溶骨破坏中的作用
发布时间：2018-11-11 22:00
【摘要】：骨巨细胞瘤(Giant-cell Tumor of Bone,GCTB)是常见的原发性骨肿瘤之一,又称破骨细胞瘤。约占临床全部骨肿瘤病例的5-6%,好发于长骨干骺端与胸椎、骶骨处。发病年龄多在20-40岁,女性更为多见。目前认为骨巨细胞瘤属于交界性肿瘤,具有较强的局部浸润生长能力和肺转移可能。因其病灶以局部溶骨性破坏为主要特点,临床上多以病理性骨折伴持续性疼痛为首发症状,同时可造成神经功能受损等不良后果,严重降低患者生活质量。该类型肿瘤对放疗、化疗敏感性均不强,目前的首选治疗方案为手术切除肿瘤结合靶向药物治疗。然而骨巨细胞瘤具有高复发倾向,即使予以被认可的囊外切除术,其复发率仍高达41.7%。骨巨细胞瘤来源于骨髓中的间叶细胞,以广泛存在的多核巨细胞(MNGC)为主要病理特点,其形态接近破骨细胞并表现出与破骨细胞类似的生物学行为和功能。研究认为骨巨细胞瘤中同时存在另两种细胞类型,即成骨细胞来源的巨细胞肿瘤基质细胞(GCTSC)和单核细胞(MNHC)。其中GCTSC因具有无限增殖能力及刺激单核细胞形成多核巨细胞的能力,在骨巨细胞瘤溶骨性破坏过程中起到关键作用,被认为是骨巨细胞瘤中实际的肿瘤成分。因此对GCTSC的分化、增殖调控及其诱导多核巨细胞分化、迁移能力的研究可以为预防、诊断及治疗骨巨细胞瘤提供新的方向。趋化因子家族包括四个亚族,分别为C、CC、CXC、CX3C。趋化因子CCL20又称为巨噬细胞炎性蛋白-3α(MIP3α),属CC亚族,其受体为CCR。其编码DNA位于第二号染色体,CCL20 mRNA翻译形成96个氨基酸的前体蛋白,后经加工形成70个氨基酸的成熟趋化因子CCL20。CCL20主要在肝脏、肺和淋巴组织中的单核细胞、T淋巴细胞、树突状细胞、内皮细胞中表达,可被多种细胞因子如肿瘤坏死因子α(TNFα)、白介素1(IL-1)、γ干扰素(IFN-γ)等诱导表达。参与炎症细胞趋化、肿瘤细胞增殖、转移等生理及病理过程。目前研究已证实趋化因子CCL20与乳腺癌、肝癌、结肠癌及胰腺癌等多种恶性肿瘤的侵袭和转移密切相关。然而并无文献报道其与骨巨细胞瘤的发生、发展是否存在关联。在本实验中,我们首先通过基因芯片分析比较了不同类型趋化因子在骨巨细胞瘤患者的肿瘤基质细胞及正常松质骨细胞中的表达差异,发现趋化因子CCL20表达明显上调。再通过血清免疫学分析比较了趋化因子CCL20在骨巨细胞瘤患者血清的表达水平以及不同大小骨巨细胞瘤之间表达水平的差异。其次,通过细胞条件培养试验验证了CCL20自分泌促进GCTSC增殖的作用,通过细胞迁移试验技术(Transwell)验证了趋化因子CCL20对GCTSC迁移能力的影响,再以细胞条件培养、骨片试验验证了CCL20诱导破骨细胞形成的作用。最后,通过检测破骨细胞相关Maker gene在CCL20刺激下的表达量变化探索了CCL20影响骨巨细胞瘤可能的信号传导通路。我们首次证实了趋化因子CCL20在骨巨细胞瘤中表达明显升高,并证实了趋化因子CCL20有促进GCTSC增殖、迁移及破骨细胞形成的作用,并发现其可能的作用机制。实验结果如下:1.趋化因子CCL20在骨巨细胞瘤组织中表达明显增高;2.趋化因子CCL20有促进GCTSC分化、增殖、迁移及破骨细胞形成的作用;3.趋化因子CCL20可能通过影响AKT磷酸化从而在骨巨细胞瘤中产生生物学作用。
[Abstract]:Giant cell tumor of bone (GCTB) is one of the most common primary bone tumors, or osteoclast. accounting for 5-6% of the total clinical bone tumor cases, and is good for the long-shaft bone-bone end and the thoracic vertebra and the bone-bone. The age of the disease is more than 20-40 years, and the female is more. At present, the giant cell tumor of the bone belongs to the borderline tumor, and has stronger local infiltration and growth ability and the possibility of lung metastasis. Because of the local osteolytic destruction of its focus, it is mainly characterized by pathological fracture with persistent pain as the first symptom, and can cause the adverse effects of nerve function damage and the like, and the quality of life of the patient is seriously reduced. The type of tumor is less sensitive to radiotherapy and chemotherapy, and the current preferred treatment scheme is to treat the tumor in combination with the targeted medicine. However, giant cell tumor of bone has a tendency of high recurrence, and the recurrence rate of giant cell tumor is 45.7%. Giant cell tumor of bone is derived from the mesenchymal cells in the bone marrow, and is the main pathological feature of the widely present multinucleated giant cell (MNGC), which is close to osteoclast and exhibits similar biological behavior and function as osteoclast. It is considered that there are two other types of cells in the bone giant cell tumor, namely, the giant cell tumor stromal cells (GCTSC) and the mononuclear cells (MNHC) of the osteoblast origin. GCTSC plays a key role in the process of osteolytic destruction of the giant cell tumor of the bone, which is considered to be the actual tumor component in the bone giant cell tumor. Therefore, the study of the differentiation, proliferation and regulation of GCTSC and the induction of multi-nuclear giant cell differentiation can provide a new direction for the prevention, diagnosis and treatment of giant cell tumor of bone. The chemokine family includes four subfamilies, C, CC, CXC, and CX3C, respectively. Chemokine CCL20 is also called macrophage inflammatory protein-3-(MIP3-1), which belongs to CC subfamily and its receptor is CCR. The encoded DNA is located in the second chromosome, and the CCL20 mRNA is translated to form a precursor protein of 96 amino acids, and the mature chemokines CCL20, CCL20, which form 70 amino acids, are mainly expressed in the monocytes, T-lymphocytes, dendritic cells, and the endothelial cells in the liver, the lung and the lymphoid tissue, can be expressed by various cytokines such as tumor necrosis factor IX (TNF antigen), interleukin 1 (IL-1), interferon (IFN-1), and the like. It is involved in the physiological and pathological processes of the chemotaxis of the inflammatory cells, the proliferation and metastasis of the tumor cells, and the like. The present study has shown that the chemokine CCL20 is closely related to the invasion and metastasis of various malignant tumors, such as breast cancer, liver cancer, colon cancer and pancreatic cancer. However, there is no literature to report whether it is associated with the occurrence and development of giant cell tumor of bone. In this experiment, we first analyzed the expression of the different types of chemokines in the tumor matrix cells and normal cancellous bone cells in the bone giant cell tumor patients by gene chip analysis, and the expression of the chemokine CCL20 was found to be up-regulated. The expression level of the serum of the chemokine CCL20 in the bone giant cell tumor and the difference of the expression level between the giant cell tumors of different sizes were compared by the serum immunological analysis. Secondly, the effect of the CCL20 autocrine on the proliferation of GCTSC was verified by the cell condition culture test. The effect of the chemokine CCL20 on the GCTSC migration ability was verified by the cell migration test (Transwell), and the effect of the CCL20 on the formation of osteoclasts was verified by the cell culture and the bone fragment test. Finally, through the detection of the expression of osteoclast-related Maker gene under the stimulation of the CCL20, the possible signal transduction pathway of the bone giant cell tumor of the bone is explored by the CCL20. We first confirmed that the expression of the chemokine CCL20 in the giant cell tumor of the bone was significantly increased, and it was confirmed that the chemokine CCL20 had the effect of promoting the proliferation, migration and osteoclast formation of the GCTSC, and found its possible mechanism of action. The experimental results are as follows: 1. The expression of the chemokine CCL20 in the tissue of the giant cell tumor of the bone was significantly increased; Chemokine CCL20 has the effect of promoting the differentiation, proliferation, migration and osteoclast formation of GCTSC; The chemokine CCL20 may play a biological role in the bone giant cell tumor by affecting the phosphorylation of AKT.
【学位授予单位】：第二军医大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R738.1
本文编号：2326262
[Abstract]:Giant cell tumor of bone (GCTB) is one of the most common primary bone tumors, or osteoclast. accounting for 5-6% of the total clinical bone tumor cases, and is good for the long-shaft bone-bone end and the thoracic vertebra and the bone-bone. The age of the disease is more than 20-40 years, and the female is more. At present, the giant cell tumor of the bone belongs to the borderline tumor, and has stronger local infiltration and growth ability and the possibility of lung metastasis. Because of the local osteolytic destruction of its focus, it is mainly characterized by pathological fracture with persistent pain as the first symptom, and can cause the adverse effects of nerve function damage and the like, and the quality of life of the patient is seriously reduced. The type of tumor is less sensitive to radiotherapy and chemotherapy, and the current preferred treatment scheme is to treat the tumor in combination with the targeted medicine. However, giant cell tumor of bone has a tendency of high recurrence, and the recurrence rate of giant cell tumor is 45.7%. Giant cell tumor of bone is derived from the mesenchymal cells in the bone marrow, and is the main pathological feature of the widely present multinucleated giant cell (MNGC), which is close to osteoclast and exhibits similar biological behavior and function as osteoclast. It is considered that there are two other types of cells in the bone giant cell tumor, namely, the giant cell tumor stromal cells (GCTSC) and the mononuclear cells (MNHC) of the osteoblast origin. GCTSC plays a key role in the process of osteolytic destruction of the giant cell tumor of the bone, which is considered to be the actual tumor component in the bone giant cell tumor. Therefore, the study of the differentiation, proliferation and regulation of GCTSC and the induction of multi-nuclear giant cell differentiation can provide a new direction for the prevention, diagnosis and treatment of giant cell tumor of bone. The chemokine family includes four subfamilies, C, CC, CXC, and CX3C, respectively. Chemokine CCL20 is also called macrophage inflammatory protein-3-(MIP3-1), which belongs to CC subfamily and its receptor is CCR. The encoded DNA is located in the second chromosome, and the CCL20 mRNA is translated to form a precursor protein of 96 amino acids, and the mature chemokines CCL20, CCL20, which form 70 amino acids, are mainly expressed in the monocytes, T-lymphocytes, dendritic cells, and the endothelial cells in the liver, the lung and the lymphoid tissue, can be expressed by various cytokines such as tumor necrosis factor IX (TNF antigen), interleukin 1 (IL-1), interferon (IFN-1), and the like. It is involved in the physiological and pathological processes of the chemotaxis of the inflammatory cells, the proliferation and metastasis of the tumor cells, and the like. The present study has shown that the chemokine CCL20 is closely related to the invasion and metastasis of various malignant tumors, such as breast cancer, liver cancer, colon cancer and pancreatic cancer. However, there is no literature to report whether it is associated with the occurrence and development of giant cell tumor of bone. In this experiment, we first analyzed the expression of the different types of chemokines in the tumor matrix cells and normal cancellous bone cells in the bone giant cell tumor patients by gene chip analysis, and the expression of the chemokine CCL20 was found to be up-regulated. The expression level of the serum of the chemokine CCL20 in the bone giant cell tumor and the difference of the expression level between the giant cell tumors of different sizes were compared by the serum immunological analysis. Secondly, the effect of the CCL20 autocrine on the proliferation of GCTSC was verified by the cell condition culture test. The effect of the chemokine CCL20 on the GCTSC migration ability was verified by the cell migration test (Transwell), and the effect of the CCL20 on the formation of osteoclasts was verified by the cell culture and the bone fragment test. Finally, through the detection of the expression of osteoclast-related Maker gene under the stimulation of the CCL20, the possible signal transduction pathway of the bone giant cell tumor of the bone is explored by the CCL20. We first confirmed that the expression of the chemokine CCL20 in the giant cell tumor of the bone was significantly increased, and it was confirmed that the chemokine CCL20 had the effect of promoting the proliferation, migration and osteoclast formation of the GCTSC, and found its possible mechanism of action. The experimental results are as follows: 1. The expression of the chemokine CCL20 in the tissue of the giant cell tumor of the bone was significantly increased; Chemokine CCL20 has the effect of promoting the differentiation, proliferation, migration and osteoclast formation of GCTSC; The chemokine CCL20 may play a biological role in the bone giant cell tumor by affecting the phosphorylation of AKT.
【学位授予单位】：第二军医大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R738.1
【参考文献】
相关期刊论文 前2条
1 ;Chemokines and hepatocellular carcinoma[J];World Journal of Gastroenterology;2010年15期
2 Claudia Rubie;Vilma Oliveira Frick;Mathias Wagner;Christina Weber;Bianca Kruse;Katja Kempf;Jochen Knig;Bettina Rau;Martin Schilling;;Chemokine expression in hepatocellular carcinoma versus colorectal liver metastases[J];World Journal of Gastroenterology;2006年41期
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