【摘要】：目的探討多發(fā)性骨髓瘤(MM)患者的循環(huán)骨髓瘤細胞(CMCs)數(shù)量和髓內瘤細胞(MMCs)的相關性以及CMCs與MM臨床相關指標的關系,評價CMCs反映骨髓瘤細胞負荷及病情進展的可行性。建立一套免疫磁珠(IMB)富集、篩選CMCs的方法,用于監(jiān)測MM的MRD和復發(fā)。方法(1)CMCs和MMCs的相關性分析:應用流式細胞術(FCM)檢測82例MM患者與30例非漿細胞相關疾病的貧血患者的CMCs和MMCs的比例,探討兩者相關性,分別比較兩者在各療效組中的差異,分析CMCs與MM臨床相關指標的關系。(2)用骨髓瘤細胞系U266建立細胞濃度梯度模型,FCM分別檢測每個模擬濃度點時抗APC磁珠分選前后的U266細胞比值,即直接FCM組和IMB-FCM組,根據(jù)抗APC磁珠靶向抗原CD38-APC/CD138-APC的不同,兩組又分為CD38-直接FCM組和CD138-直接FCM組以及CD38-IMB-FCM組和CD138-IMB-FCM組。分別比較IMB-FCM組和相應的直接FCM組U266細胞的比值,確定IMB-FCM檢測瘤細胞的敏感度。(3)收集32例MM患者的骨髓和外周血標本,分為骨髓直接FCM組、外周血CD38-直接FCM組、外周血CD38-IMB-FCM組、外周血CD138-直接FCM組、外周血CD138-IMB-FCM組,分別比較外周血IMB-FCM組與相應的外周血/骨髓直接FCM組檢測瘤細胞的比值及CMCs的陽性率。結果(1)初發(fā)及復發(fā)組和PR組的CMCs陽性檢出率分別為71.4%、64.5%,兩組中CMCs與MMCs比例均呈正相關(P0.05);MMCs和CMCs比例在初發(fā)及復發(fā)、PR、CR組中依次降低(P0.0083)。MMCs和CMCs在CR組及對照組中均未檢測出。(2)與CMCs陰性組相比,CMCs陽性組的β2-MG升高、中度以上貧血、sCrea升高、腎功能異常及骨質破壞的頻率明顯升高,呈現(xiàn)出較高的DS、ISS分期(P0.05)。經(jīng)多因素Logistic回歸分析,β2-MG升高、中度以上貧血是CMCs存在的獨立風險因子(P0.05)。(3)在每個模擬濃度點CD38/CD138-IMB-FCM組檢測U266細胞的比例均分別顯著高于相應的直接FCM組(P0.05),直接FCM、IMB-FCM檢測U266細胞的敏感度分別為0.01%、0.001%。(4)瘤細胞比值在骨髓直接FCM組分別顯著高于外周血CD38/CD138-IMB-FCM組,在外周血CD38/CD138-IMB-FCM組分別顯著高于相應的外周血-直接FCM組(P0.0167);CD38/CD138-IMB-FCM組檢測CMCs陽性率(90.6%,87.5%)分別顯著高于相應的直接FCM組(65.6%)(P0.05)。結論(1)CMCs可反映MM患者的腫瘤負荷及病情進展;相比MMCs,檢測CMCs某種程度可以替代反復抽取骨髓檢測,提高患者依從性。β2-MG升高和中度以上貧血是CMCs存在的獨立風險因子。(2)IMB-FCM能提高CMCs的陽性檢出率,敏感度比FCM高10倍。
[Abstract]:Objective to investigate the correlation between the number of circulating myeloma cells (CMCs) and intramedullary tumor cell (MMCs) in patients with multiple myeloma (MM), and the relationship between CMCs and MM, and to evaluate the feasibility of CMCs in reflecting the cell load and progression of myeloma. A set of immunomagnetic bead (IMB) enrichment and CMCs screening method was established to monitor the MRD and recurrence of MM. Methods (1) correlation analysis of CMCs and MMCs: the proportion of CMCs and MMCs in 82 patients with MM and 30 anemia patients with non-plasma cell related diseases were detected by flow cytometry (FCM). The relationship between CMCs and clinical parameters of MM was analyzed. (2) the cell concentration gradient model was established by using myeloma cell line U266. FCM was used to detect the ratio of U266 cells before and after the separation of anti-APC beads at each simulated concentration point, that is, direct FCM group and IMB-FCM group. According to the difference of CD38-APC/CD138-APC, the two groups were divided into CD38- direct FCM group, CD138- direct FCM group, CD38-IMB-FCM group and CD138-IMB-FCM group. The ratio of U266 cells in IMB-FCM group and corresponding direct FCM group was compared to determine the sensitivity of IMB-FCM in detecting tumor cells. (3) the bone marrow and peripheral blood samples of 32 patients with MM were collected and divided into three groups: bone marrow direct FCM group, peripheral blood CD38- direct FCM group, peripheral blood CD38-IMB-FCM group. The ratio of tumor cells and the positive rate of CMCs in peripheral blood IMB-FCM group and corresponding peripheral blood / bone marrow direct FCM group were compared between peripheral blood CD138- direct FCM group and peripheral blood CD138-IMB-FCM group. Results (1) the positive rate of CMCs in the primary and recurrent group and PR group was 71.4% and 64.5%, respectively. There was a positive correlation between CMCs and MMCs in the two groups (P0.05). The proportion of); MMCs and CMCs decreased in turn in the PR,CR group (P0.0083). MMCs and CMCs were not detected in CR group and control group). (2) 尾 2-MG in CMCs positive group was higher than that in CMCs negative group. Moderate anemia, sCrea increased, renal dysfunction and bone destruction frequency increased significantly, showing a higher DS,ISS stage (P0.05). By multivariate Logistic regression analysis, 尾 2-MG increased. Moderate anemia is an independent risk factor for CMCs (P0.05). (3). The percentage of U266 cells detected in CD38/CD138-IMB-FCM group was significantly higher than that in direct FCM group at each simulated concentration point (P0.05). The sensitivity of direct FCM,IMB-FCM detection for U266 cells was 0.01and 0.001 respectively (4) tumor cells were detected. The ratio in bone marrow direct FCM group was significantly higher than that in peripheral blood CD38/CD138-IMB-FCM group. The positive rate of CMCs in CD38/CD138-IMB-FCM group was significantly higher than that in peripheral blood direct FCM group (P0.0167), and the positive rate of CMCs in CD38/CD138-IMB-FCM group (90.6%) was significantly higher than that in direct FCM group (65.6%) (P0.05). Conclusion (1) CMCs can reflect the tumor load and disease progression of MM patients, compared with MMCs, detection of CMCs, it can replace repeated bone marrow detection. Increased 尾 2-MG and moderate anemia were independent risk factors for CMCs. (2) IMB-FCM could increase the positive detection rate of CMCs, and the sensitivity was 10 times higher than that of FCM.
【學位授予單位】：鄭州大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R733.3;R730.43

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