發(fā)布時間：2018-10-25 06:30

【摘要】：背景和目的外周T細胞淋巴瘤(PTCL)是一組異質性強、侵襲性高的非霍奇金淋巴瘤,缺乏規(guī)范統(tǒng)一的治療方案,預后較其他類型淋巴瘤差。尋找新的分子標記物對于深入探索其成瘤機制、研發(fā)新藥都有重要意義。近幾年國外基于高通量基因組層面的研究首次揭示TH2細胞分化指標轉錄因子GATA3可能在PTCL發(fā)生發(fā)展中起到作用。本研究擬進行GATA3的預后研究,并在體外條件下探索GATA3對腫瘤增殖、浸潤等惡性行為的影響,為淋巴瘤發(fā)病機理研究提供新的實驗證據(jù)。方法回顧性收集PTCL患者的福爾馬林固定石蠟包埋的病理標本及臨床資料,所有病例均為2007年至2013年間由北京協(xié)和醫(yī)院病理科診斷的初治患者。對GATA3、TBX21、CD68進行免疫組化染色,分析其表達與患者臨床特征及預后的關系。在體外條件下通過慢病毒轉染法建立GATA3穩(wěn)定敲低的Hut78淋巴瘤細胞系,應用流式細胞檢測技術研究敲低細胞系的凋亡變化,應用實時定量PCR技術檢測敲低細胞系的細胞因子表達,通過條件培養(yǎng)方法研究敲低細胞系對M2巨噬細胞分化的影響。結果本研究共納入109例初治PTCL患者,其中PTCL非非特指為60例。在PTCL組織的腫瘤細胞中54例GATA3表達陽性(49.5%),47例TBX21表達陽性(43.1%),57例CD68表達陽性(52.3%)。GATA3和CD68與PTCL的不良預后相關。K-M曲線法發(fā)現(xiàn),GATA3陽性組的中位總生存期(OS)為120天,GATA3陰性組的中位OS為511天(Log-rank, P=0.000); CD68陽性組的中位OS為180天,CD68陰性組的中位OS為450天(Log-rank, P=0.037)。多因素分析提示ECOG評分≥2分、GATA3陽性為PTCL預后不良的獨立危險因子。相關性分析提示GATA3高表達與出現(xiàn)B癥狀(Pearson P=0.006)、CD68高表達相關(Pearson P0.01)。體外研究篩選出Hut78為GATA3高表達的T細胞淋巴瘤細胞系。應用慢病毒轉染shGATA3序列成功后,與對照組相比,Hut78-shGATA3細胞的GATA3 mRNA和蛋白表達量均明顯下調,早期凋亡階段細胞百分比明顯增高。在細胞因子方面,Hut78-shGATA3轉錄IL-4、IL-5、IL-13及VEGF的mRNA水平均明顯下降。應用淋巴瘤細胞上清誘導U937向巨噬細胞分化,研究發(fā)現(xiàn)與對照組相比,Hut78-shGATA3上清誘導巨噬細胞向M2分化的比例顯著下降。結論在PTCL中,GATA3陽性患者較GATA3陰性患者OS明顯縮短,且與ECOG評分≥2分一并為預后不良的獨立影響因素,提示GATA3可以作為PTCL患者評估預后的因素。GATA3表達量與CD68表達量呈正相關,提示(GATA3可能促進了巨噬細胞在PTCL腫瘤組織中浸潤。從絕對例數(shù)來看,GATA3高表達與嗜血綜合征的發(fā)生相關。體外實驗證實,T淋巴瘤細胞系Hut78的GATA3表達量顯著增高。GATA3敲低后,T細胞淋巴瘤細胞的早期凋亡比例增加,TH2相關細胞因子及VEGF表達水平下降,且誘導巨噬細胞向M2型分化的能力降低。GATA3可能通過這些途徑對PTCL患者預后產生影響。
[Abstract]:Background and objective Peripheral T-cell lymphoma (PTCL) is a group of non-Hodgkin 's lymphoma with strong heterogeneity and high invasiveness. Searching for new molecular markers is of great significance for further exploring its tumorigenesis mechanism and developing new drugs. In recent years, foreign studies based on high-throughput genome level for the first time revealed that TH2 cell differentiation index transcription factor GATA3 may play a role in the development of PTCL. This study is intended to study the prognosis of GATA3 and to explore the effects of GATA3 on malignant behaviors such as tumor proliferation and invasion in vitro, providing new experimental evidence for the study of the pathogenesis of lymphoma. Methods the pathological specimens and clinical data of formalin fixed paraffin embedded PTCL patients were collected retrospectively. All the patients were newly diagnosed by the Department of Pathology of Peking Union Hospital from 2007 to 2013. Immunohistochemical staining was used to analyze the relationship between the expression of GATA3,TBX21,CD68 and clinical features and prognosis. GATA3 stable knockout Hut78 lymphoma cell lines were established by lentivirus transfection in vitro. Apoptosis of knockout cell lines was studied by flow cytometry and cytokine expression was detected by real-time quantitative PCR. The effect of knock-down cell line on M 2 macrophage differentiation was studied by conditioned culture. Results A total of 109 patients with PTCL were enrolled in this study, including 60 patients with PTCL. 54 cases (49.5%) were positive for GATA3, 47 (43.1%) for TBX21 and 57 (52.3%) for CD68 in PTCL. GATA3 and CD68 were correlated with the poor prognosis of PTCL. K-M curve showed that the median total survival time of GATA3 positive group was 120 days, and that of GATA3 negative group was 120 days. The median OS of CD68 positive group was 180 days and the median OS of CD68 negative group was 450d (Log-rank, P0. 037). Multivariate analysis showed that ECOG score 鈮，

本文編號：2292902