摘要目的 探讨微小RNA-21(miR-21)在弥漫性大B细胞淋巴瘤(DLBCL)细胞系和患者血清中的表达及其在DLBCL早期诊断、基因分型及预后判断中的意义.方法 用实时荧光定量聚合酶链反应( FQ-PCR)从细胞水平检测9种DLBCL细胞系(OCI-Ly1、OCI-Ly3、OCI-Ly4、OCI-Ly7、OCI-Ly8、OCI-Ly10、OCI-Ly18、OCI-Ly19和HBL)中miR-21的表达,及临床确诊的62例DLBCL患者(DLBCL组)、50名健康体检者(健康对照组)血清miR-21表达水平；同时,随访62例DLBCL患者无复发生存期(RFS),应用Kaplan-Meier生存曲线分析62例DLBCL患者血清miR-21表达与预后的关系.结果 健康对照组B淋巴细胞miR-21的相对表达量为1.04±0.02,9种DLBCL细胞系miR-21 相对表达量依次为2.30±0.35、237.97±56.19、5.27±0.83、3.40±0.30、11.22±2.70、133.55±16.78、6.63±0.24、4.91±0.37、81.59±6.64,9种DLBCL细胞系的miR-21表达水平均高于健康对照组(t值分别为7.3、13.7、2 1.0、6.2、8.8、13.6、6.5、39.5、18.1,P均＜0.01)；且ABC亚型细胞系(OCI-Ly3、OCI-Ly10、HBL)显著高于GCB亚型细胞系(OCI-Ly1、OCI-Ly4、OCI-Ly7、OCI-Ly8、OCI-Ly18、OCI-Ly19；t=11.18,P＜0.01).DLBCL组血清中miR-21的表达量为21.38( 10.26 ～45.21),健康对照组为1.87(1.05 ～3.97),DLBCL组血清miR-21的表达显著高于健康对照组(U=168,P=0.000).DLBCL患者中GCB亚型miR-21的表达为18.30(7.32～33.46),ABC亚型miR-21的表达为28.68( 14.92～98.44),ABC亚型miR-21的表达高于GCB亚型(U=336,P=0.043).此外,GCB型Ⅰ+Ⅱ期患者血清miR-21表达为24.75( 16.08 ～50.38),而Ⅲ+Ⅳ期为11.96(4.10～21.05)；ABC型Ⅰ+Ⅱ期患者血清miR-21表达为47.49( 25.65～295.41),而Ⅲ+Ⅳ期为16.66(5.35～44.30),Ⅰ+Ⅱ期明显高于Ⅲ+Ⅳ期(GCB型U=62,P=0.013;ABC型U=53,P=0.014).随访DLBCL患者的RFS,miR-21高表达患者预后明显好于miR-21低表达患者(U=259,P=0.035).结论 DLBCL患者血清miR-21高表达,且与DLBCL临床分级分期、基因型和预后判断密切相关,miR-21有可能成为DLBCL早期诊断、基因分型及预后判断的新分子标志.

abstractsObjective To study the expression of microRNA-21 ( miR-21 )in serum of patient with diffuse large B cell lymphoma (DLBCL) and DLBCL cell lines and validate the significance of miR-21 in early diagnosis,genotyping and prognosis estimates of DLBCL.Methods miR-21 expression were detected by fluorescent quantity polymerase chain reaction (FQ-PCR)in 9 lymphoma cell lines (OCI-Ly1,OCI-Ly3,OCI-Ly4,OCI-Ly7,OCI-Ly8,OCI-Ly10,OCI-Ly18,OCI-Ly19 and HBL),the serum from DLBCL patients (n =62) and health controls (n =50 ).Kaplan-Meier survival analysis was carried out during the relapsefree survival period of DLBCL patients to explore the relationship between the prognosis and microRNA expression level.Results Real time FQ-PCR result indicated that miR-21 expression was higher in DLBCL cell lines than that in normal B cells (BC).miR-21 expression in normal B cell and 9 DLBCL cell lines separately were 1.04 ± 0.02,2.30 ± 0.35,237.97 ± 56.19,5.27 ± 0.83,3.40 ± 0.30,11.22 ± 2.70,133.55 ± 16.78,6.63 ±0.24,4.91 ±0.37 and 81.59 ±6.64.Compared with BC,the expression of miR-21 were higher in all 9 DLBCL cell lines ( t =7.3,13.7,21.0,6.2,8.8,13.6,6.5,39.5,18.1 ;P ＜ 0.01 ).miR-21 expression segregates with specific molecular subgroups of DLBCL The expression was higher in the ABC type cell lines (OCI-Ly3,OCI-Ly10,HBL) than GCB type cell lines (OCI-Ly1,OCI-Ly4,OCI-Ly7,OCI-Ly8,OCI-Ly18,OCI-Ly19;t =11.18,P ＜ 0.01 ).Consistent with the cell line models,miR-21 expression levels were higher in serum from DLBCL patients [21.38 (10.26-45.21 )] than from controls [1.87 ( 1.05-3.97 ),U =168,P =0.000],and the levels were higher in DLBCL cases with an ABC-type [28.68 ( 14.92-98.44 )] than those in GCB-type [18.30 ( 7.32-33.46 ),U =336,P =0.043].MiR-21 expression levels were different in sera from different clinical stage DLBCL patients.The miR-21 level in serum of patients with subgroup ABC and subgroup GCB in stage Ⅰ and Ⅱ were 47.49( 25.65-295.41 ) and 24.74( 16.08-50.38) respectively and in stage Ⅲ and Ⅳ were 16.66 ( 5.35-44.30 ) and 11.96 ( 4.10-21.05) respectively.The levels were higher in DLBCL cases with Ⅰ -Ⅱ stage than those with Ⅲ-Ⅳ stage (U =62,P =0.013 in GCB type; U =53,P =0.014 in ABC type).Moreover,compare with relapse-free survival in DLBCL patients,high miR-21 expression was associated with well prognosis ( U =259,P =0.035).Conclusions MiR-21 is high expression in DLBCL cell lines and DLBCL patients serum.miR-21 level in sera from DLBCL patients is associated with clinical stage,molecular subgroup and prognosis estimates.MiR-21 may serve as a new biomarker to early detection,genotyping and prognosis estimates of DLBCL.