趨化因子CCL20在促進骨巨細胞瘤增殖、遷移及溶骨破壞中的作用
發(fā)布時間：2018-11-11 22:00
【摘要】：骨巨細胞瘤(Giant-cell Tumor of Bone,GCTB)是常見的原發(fā)性骨腫瘤之一,又稱破骨細胞瘤。約占臨床全部骨腫瘤病例的5-6%,好發(fā)于長骨干骺端與胸椎、骶骨處。發(fā)病年齡多在20-40歲,女性更為多見。目前認為骨巨細胞瘤屬于交界性腫瘤,具有較強的局部浸潤生長能力和肺轉移可能。因其病灶以局部溶骨性破壞為主要特點,臨床上多以病理性骨折伴持續(xù)性疼痛為首發(fā)癥狀,同時可造成神經功能受損等不良后果,嚴重降低患者生活質量。該類型腫瘤對放療、化療敏感性均不強,目前的首選治療方案為手術切除腫瘤結合靶向藥物治療。然而骨巨細胞瘤具有高復發(fā)傾向,即使予以被認可的囊外切除術,其復發(fā)率仍高達41.7%。骨巨細胞瘤來源于骨髓中的間葉細胞,以廣泛存在的多核巨細胞(MNGC)為主要病理特點,其形態(tài)接近破骨細胞并表現出與破骨細胞類似的生物學行為和功能。研究認為骨巨細胞瘤中同時存在另兩種細胞類型,即成骨細胞來源的巨細胞腫瘤基質細胞(GCTSC)和單核細胞(MNHC)。其中GCTSC因具有無限增殖能力及刺激單核細胞形成多核巨細胞的能力,在骨巨細胞瘤溶骨性破壞過程中起到關鍵作用,被認為是骨巨細胞瘤中實際的腫瘤成分。因此對GCTSC的分化、增殖調控及其誘導多核巨細胞分化、遷移能力的研究可以為預防、診斷及治療骨巨細胞瘤提供新的方向。趨化因子家族包括四個亞族,分別為C、CC、CXC、CX3C。趨化因子CCL20又稱為巨噬細胞炎性蛋白-3α(MIP3α),屬CC亞族,其受體為CCR。其編碼DNA位于第二號染色體,CCL20 mRNA翻譯形成96個氨基酸的前體蛋白,后經加工形成70個氨基酸的成熟趨化因子CCL20。CCL20主要在肝臟、肺和淋巴組織中的單核細胞、T淋巴細胞、樹突狀細胞、內皮細胞中表達,可被多種細胞因子如腫瘤壞死因子α(TNFα)、白介素1(IL-1)、γ干擾素(IFN-γ)等誘導表達。參與炎癥細胞趨化、腫瘤細胞增殖、轉移等生理及病理過程。目前研究已證實趨化因子CCL20與乳腺癌、肝癌、結腸癌及胰腺癌等多種惡性腫瘤的侵襲和轉移密切相關。然而并無文獻報道其與骨巨細胞瘤的發(fā)生、發(fā)展是否存在關聯。在本實驗中,我們首先通過基因芯片分析比較了不同類型趨化因子在骨巨細胞瘤患者的腫瘤基質細胞及正常松質骨細胞中的表達差異,發(fā)現趨化因子CCL20表達明顯上調。再通過血清免疫學分析比較了趨化因子CCL20在骨巨細胞瘤患者血清的表達水平以及不同大小骨巨細胞瘤之間表達水平的差異。其次,通過細胞條件培養(yǎng)試驗驗證了CCL20自分泌促進GCTSC增殖的作用,通過細胞遷移試驗技術(Transwell)驗證了趨化因子CCL20對GCTSC遷移能力的影響,再以細胞條件培養(yǎng)、骨片試驗驗證了CCL20誘導破骨細胞形成的作用。最后,通過檢測破骨細胞相關Maker gene在CCL20刺激下的表達量變化探索了CCL20影響骨巨細胞瘤可能的信號傳導通路。我們首次證實了趨化因子CCL20在骨巨細胞瘤中表達明顯升高,并證實了趨化因子CCL20有促進GCTSC增殖、遷移及破骨細胞形成的作用,并發(fā)現其可能的作用機制。實驗結果如下:1.趨化因子CCL20在骨巨細胞瘤組織中表達明顯增高;2.趨化因子CCL20有促進GCTSC分化、增殖、遷移及破骨細胞形成的作用;3.趨化因子CCL20可能通過影響AKT磷酸化從而在骨巨細胞瘤中產生生物學作用。
[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.
【學位授予單位】：第二軍醫(yī)大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R738.1
本文編號：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.
【學位授予單位】：第二軍醫(yī)大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R738.1
【參考文獻】
相關期刊論文 前2條
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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