单核-巨噬细胞表达的胸苷磷酸化酶增强去氧氟尿苷抗结肠癌细胞活性  

Thymidine phosphorylase expressed in monocyte-macrophages enhanced anticancer effect of 5'-deoxy-5-fluorouridine on colorectal carcinoma cells

作　　者：张继民[1] 刘明姬[2] 溝井賢幸 椎葉健一 佐佐木巌 松野正纪[3] 

机构地区：[1]广州医学院第二附属医院胃肠外科,510260 [2]中山大学黄埔医院麻醉科 [3]日本東北大学医学部第一外科

出　　处：《中华医学杂志》2004年第9期718-724,共7页National Medical Journal of China

基　　金：日本文部省博士科研基金资助项目 ;教育部留学人员科研启动基金资助项目 (教外司留 [2 0 0 2 ] 2 47号 )

摘　　要：目的　探讨结肠癌组织中胸苷磷酸化酶 (dThdPase)的表达 ,检定单核 巨噬细胞表达的dThdPase对抗癌药物 5′ 脱氧氟尿苷 (5’ DFUR)的转化调控作用。方法　对 4 0例结肠癌标本进行免疫组织化学染色 ,分析其细胞的dThdPas表达 ,并测定癌组织和周围正常组织的dThdPase活性。应用ELISA法分别检测 4株结肠癌细胞LS174T、CloneA、Colo32 0、MIP10 1和 2株类巨噬细胞THP 1、U937的dThdPase蛋白含量。MTT法测定 4株结肠癌细胞对 5 Fu和 5′ DFUR的半数有效浓度 (IC50 )。把定量5′ DFUR加入培养基中同类巨噬细胞或单核细胞一起培养 2 4h后 ,取其上清液二倍稀释加入大肠癌细胞中测定IC50 有无改变。并在加入定量 5′ DFUR的类巨噬细胞培养液中检测 5 Fu的生成。结果　结肠癌组织中dThdPase活性明显高于周围正常肠组织 ,表达dThdPase阳性细胞主要是肿瘤相关巨噬细胞 (TAM)。 4株结肠癌细胞中仅LS174T检出 0 5单位 /mg的dThdPase蛋白 ;而THP 1和U937则分别为 18 2单位 /mg和 19 3单位 /mg。全部大肠癌细胞对 5′ DFUR的IC50 均明显高于 5 Fu(P <0 0 0 0 1)。但同THP 1、U937或单核细胞一起培养 2 4h后 ,其IC50 明显下降 ,仅相当于原来的 5 4 %～4 1 8% (P <0 0 0 1)。从含定量 5′ DFUR的THP 1、U937的培养?Objective To detect the thymidine phosphorylase (dThdPase) expression in colorectal carcinoma tissue,and clarify whether dThdPase expressed in macrophage-like cell lines,and monocytes from human peripheral blood can modulate the anticancer effect of 5'-deoxy-5-fluorouridine (5'-DFUR) on colorectal carcinoma cells.Methods Forty specimens resected from 40 patients with colorectal carcinoma were immunohistochemically stained by the monoclonal antibodies 654-1 (anti-dThdPase) and PG-M1 (anti-macrophage marker CD68). Then morphometrical analysis and positive cell counting were performed. In 27 of 40 specimens,dThdPase activity analysis was assayed by HPLC. The dThdPase level were also measured by ELISA in 4 colorectal cancer cell lines,LS174T,Clone A,Colo320,MIP101,and 2 macrophage- like cell lines,THP-1,U937. After estimated the drug sensitivities of each colorectal carcinoma cell both to 5′-DFUR and 5-Fu by MTT assay, THP-1,U937,or monocytes isolated from human blood were incubated in the medium containing 5-Fu or 5′-DFUR for 24 hours,respectively. Then the supernatant was collected and 2-fold serially diluted with the medium,in which the macrophage-like cells,or monocytes were also cultivated for 24h without anticancer agents. Using the serially diluted medium,MTT assay were also carried out on 4 colorectal carcinoma cell lines. Each experiment was repeated six times and the means of IC50 value±standard errors (mean±S.E.M.) were calculated. Finally,THP-1 or U937 cells were cultured in medium containing 400 μmol/L of 5′-DFUR for 24 hours,then the supernatants were collected and the amount of generated 5-Fu was measured by HPLC.Results dThdPase activities was significantly increased (139.7 μg·5-Fu·h -1 ·ml -1 ±61.5 μg·5-Fu·h -1 ·ml -1 ) in colorectal carcinoma tissues compared with adjacent normal tissues (42.2 μg·5-Fu·h -1 ·ml -1 ±21.4 μg·5-Fu·h -1 ·ml -1 ), P <0.001 . In immunohistochemical analysis,it was confirmed that most cells expressed dThdPase-positi

关 键 词：单核-巨噬细胞表达 胸苷磷酸化酶 去氧氟尿苷 结肠癌 细胞活性 巨噬细胞 单核细胞 

分 类 号：R735.3[医药卫生—肿瘤]