甲胎蛋白mRNA在肝癌组织中的表达及其意义

目的 :探讨甲胎蛋白 (AFP) m RNA在肝癌组织中的表达及其临床意义。方法 :用巢式逆转录聚合酶链反应 (nestedRT- PCR)检测 19例肝细胞癌患者癌组织 AFP m RNA。结果 :16例 (88.9% )患者癌组织 AFP m RNA呈阳性表达 ,其外周血 AFP m RNA亦为阳性 ;而阴性表达的 3例其外周血 AFP m RNA和血清 AFP亦为阴性。癌组织 AFP m RNA阳性率在血清 HBs Ag阳性组 (93.8% )、合并肝硬化组 (10 0 % )和血清 AFP≥ 2 5 μg/ L 组 (10 0 % )高于 HBs Ag阴性组 (33.3% )、无合并肝硬化组 (5 7.1% )和 AFP<2 5 μg/ L 组 (5 7.1% ) (P <0 .0 5 )。相关分析显示外周血 AFP m RNA阳性率和血清 AFP水平均与癌组织 AFP m RNA阳性表达呈正相关 (r =0 .6 413,P <0 .0 5 ;r=0 .6 0 2 4,P <0 .0 5 )。癌组织 AFP m RNA阳性表达和血清 HBs Ag阳性表达及合并有肝硬化呈正相关 (r =0 .6 5 0 3,P <0 .0 5 ;r =0 .5 90 8,P <0 .0 5 ) ,而与有无门静脉癌栓或肝外转移及癌灶大小无相关性 (P >0 .0 5 )。结论 :大多数肝细胞癌的 AFP基因处于活化状态。外周血 AFP m RNA阳性表达和血清 AFP水平均与癌组织 AFP m

HBsAg阳性甲胎蛋白mRNA在肝癌组织中临床意义

目的:探讨甲胎蛋白mRNA在HB sA g阳性肝癌组织中表达的临床意义。方法:用巢式逆转录聚合酶链反应检测38例HB-sA g阳性肝细胞癌患者癌组织甲胎蛋白mRNA。结果:32例(84.2%)患者癌组织甲胎蛋白mRNA呈阳性表达,其外周血甲胎蛋白mR-NA亦为阳性;而阴性表达的6例(15.8%)其外周血甲胎蛋白mRHA和血清甲胎蛋白亦为阴性。HB sA g阳性癌组织甲胎蛋白mRNA阳性率在合并肝硬化组为100%,合并血清甲胎蛋白>25 ug/L组为100%、无合并肝硬化组为57.1%,<25 ug/L组为57.1%,P<0.05。相关分析显示外周血甲胎蛋白mRNA阳性率和血清甲胎蛋白水平均于癌组织甲胎蛋白mRNA阳性表达呈正相关。癌组织甲胎蛋白RNA阳性表达与合并肝硬化呈正相关,而与有无门静脉癌栓或肝外转移及癌灶大小无相关性。结论:大多数HB sA g阳性肝细胞癌的甲胎蛋白基因处于活化状态,其外周血甲胎蛋白mRNA阳性表达和血清甲胎蛋白水平均与癌组织甲胎蛋白mRNA阳性表达有关。

肝细胞癌及癌旁组织中白蛋白mRNA和甲胎蛋白mRNA的表达

目的 :检测肝细胞癌及癌旁组织白蛋白 m RNA和甲胎蛋白 m RNA表达 ,为外周血肝细胞癌转移指标提供依据。方法 :用 RT- PCR技术检测 18例肝细胞癌和 13例癌旁肝组织中白蛋白 m RNA和甲胎蛋白 m RNA。结果 :1白蛋白 m RNA在 18例癌及 13例癌旁组织中均有高表达 ;2白蛋白 m RNA检出率明显高于第一轮 PCR甲胎蛋白 m RNA(P<0 .0 0 1) ;3白蛋白 m RNA和甲胎蛋白 m RNA可同时在癌组织中表达。结论 :选择检测白蛋白 m RNA敏感性高 ,可以避免漏检不表达或表达很低的甲胎蛋白 m RNA的肝细胞癌转移。

肝癌患者外周血甲胎蛋白mRNA表达水平与肝外转移及预后的关系

目的 探讨肝癌患者外周血甲胎蛋白mRNA表达水平与肝外转移及预后的关系。方法 采用realtime PCR(RT PCR)技术 ,定量检测 5 6例肝细胞癌患者外周血甲胎蛋白mRNA的表达水平。结果 5 6例肝癌患者外周血甲胎蛋白mRNA的平均表达水平为 (346 4 4 .6± 94 90 5 .4 )copies/ μgRNA ,伴有肝外转移的 13例 10 0 %为阳性 ,平均水平为 (1385 6 9.2± 15 94 99.3)copies/ μgRNA。 结论 伴有肝外转移的肝癌患者外周血甲胎蛋白mRNA呈高水平表达 ,甲胎蛋白mRNA可作为诊断肝癌远处转移的重要指标之一。

外周血甲胎蛋白mRNA的检测及其临床意义

目的 :寻找肝细胞癌微小转移的标志物。方法 :用巢式反转录聚合酶链反应 (Nested- RT- PCR)技术检测 6 5例肝细胞癌、2 1例非肝癌的恶性肿瘤、2 2例慢性乙型肝炎或肝硬化患者及 2 1例健康志愿者外周静脉血中的甲胎蛋白 (AFP) m RNA。结果 :AFP- m RNA在健康志愿者、非肝癌恶性肿瘤患者外周血中均为阴性 ;在肝细胞癌组外周血中 AFP- m RNA的检出率 (4 4/6 5 ,6 7.7% )明显高于慢性乙型肝炎或肝硬化组 (2 /2 2 ,9.1% ,P<0 .0 1)。AFP- m RNA检出率与临床分期、门静脉癌栓、肝外转移显著相关。在 12例血清 AFP<2 5 μg/L 的肝癌患者血中 ,7例 (5 8.3% )可检出 AFP- m RNA。结论 :AFP- m RNA可作为血循环中有肝癌细胞或肝细胞的标志物 ,在肝癌患者 ,阳性预示有血源性转移的可能 ;并且 AFP- m RNA对血清

原发性肝癌患者外周血中AFPmRNA检测的临床意义

通过定量检测AFPmRNA在原发性肝癌(PHC)患者外周血中的表达,探讨AFPmRNA作为PHC患者早期诊断和微转移指标的可能性。建立荧光定量聚合酶链式反应(FQ-PCR),检测58例外周血AFPmRNA的表达量。其中,30例PHC患者中19例AFP≥20ng/m l,23例外周血中AFPmRNA有表达;肝癌患者外周血中AFPmRNA的基因拷贝数与肝癌患者有肝内外转移有明显相关,而肝炎及健康对照组外周血中AFPmRNA均未表达,肝硬化组有一例表达。应用荧光定量PCR检测PHC外周血中AFPmRNA表达情况,可以作为早期反映PHC患者外周血中是否存在肝癌细胞和发生微转移的早期灵敏指标。

肝细胞癌病人外周血AFP-mRNA含量与肝外转移间的关系研究

目的定量检测肝细胞癌(HCC)病人外周血AFP-mRNA,并探讨其拷贝数与有无肝外转移之间的关系。方法以FQ-PCR方法检测肝癌有肝外转移组(10例)和无肝外转移组(15例)外周血单个核细胞AFP-mRNA。结果有肝外转移组拷贝数(4.7×105±36.2)copies/μg RNA无肝外转移组(3.9×102±25.4)copies/μg RNA,二者之间有明显差异。结论AFP-mRNA拷贝数作为判断有无肝外转移有一定的价值。

肝癌患者手术前后血液中AFP-mRNA的变化及意义

目的 探讨手术操作是否会导致肝癌细胞的血行播散。方法 选择 14例无远处转移并行肝癌切除术的肝癌患者 ,提取患者术前和术后外周血标本 ,于术中肿瘤切除后提取门静脉和肝静脉血标本。采用巢式RT PCR方法检测血中AFP mRNA的表达 ,观察手术前后血液中AFP mRNA表达阳性率及表达水平的变化。结果 术前外周血、手术切除肿瘤后门静脉血和术后外周血中AFP mRNA表达阳性率分别为 42 .9% ,42 .9%和 35 .7% ,术中切除肿瘤后肝静脉血中的表达阳性率为 5 7.1% ,虽高于前三者 ,但无统计学差异 ,但AFP mRNA表达水平明显高于术前及术后外周血和术中门静脉血 (P<0 .0 5 )。结论 手术中有包括肝癌细胞在内的肝脏来源的细胞经肝静脉进入血循环。

原发性肝细胞癌患者外周血AFP mRNA的表达及其临床意义

采用逆转录聚合酶链式反应(RT-PCR)检测78例原发性肝细胞癌(HCC)患者及109例其它疾病组患者外周血AFP mRNA的表达,同时用微粒子酶免疫法检测血清AFP水平。HCC外周血AFP mRNA的阳性率为61.5%,其它疾病组阳性率为1.8%,对照组均为阴性。HCC患者外周血中,AFP mRNA的阳性率与肝内静脉癌栓形成、肝内外转移以及与肿瘤分期、大小有关。作为肿瘤微转移检测的灵敏指标,外周血AFP mRNA能更好反映肿瘤的早期转移。

静脉血中微量肝癌细胞的检测方法

用聚合酶链反应法检测肝细胞癌病人静脉血中微量癌细胞，设计了两对特异引物，ＰＣＲ反应后出现与已知序列相符的１０１ｂｐ条带，并用Ｐｓｔｌ限制性内切酶消化确认其特异性。套式ＰＣＲ灵敏度达０．０１ｆｇ重组ＤＮＡ，在５ｍｌ静脉血中有１～１０个癌细胞即可识别，且重复性好。该方法可广泛地用于肝癌的早期诊断及转移机制的基础研究。

Expression and location of α-fetoprotein during rat colon development

AIM： To investigate the expression of α-fetoprotein （AFP）, a cancer-associated fetal glycoprotein, and its involvement during rat colon development. METHODS： Colons from Sprague-Dawley rat fetuses, young and adult （8 wk old） animals were used in this study. Expression levels of AFP in colons of different development stage were detected by reversetranscriptase PCR （RT-PCR） and Western blotting. To identify the cell location of AFP in the developing rat colons, double-immunofluorescent staining was performed using antibodies to specific cell markers and AFP, respectively. RESULTS： The highest levels of AFP mRNA were detected in colons of rats at embryonic day 18.5 （e18.5）. Compared to e18.5 d, the AFP expression was significantly decreased during rat development （85% for e20.5, P 〈 0.05, 58% for postnatal day 0.5 （P〈0.5）, P 〈 0.05, 37% for P7, P 〈 0.05, 24% for P14, P 〈 0.05, and 11% for P21, P 〈 0.05） and undetected in adult rats. Only the 72-kDa isoform of AFP was detected by Western blotting, the expression pattern was similar to AFP mRNA and conformed to the results of mRNA expression. The AFP positive staining was identical to different distribution patterns in fetuses, young and adult animals and positive staining for both AFP and vimentin was overlapped in mesenchymal cells at each stage tested. CONCLUSION： This study has for the first time demonstrated that AFP is localized in the mesenchyme of rat colon from the embryo to the weaning stage by immunofluorescence and presents 72-kDa isoform in the developing rat colons by Western blotting. The dynamic expression of AFP in the various developmental stages of the colon indicates that AFP might be involved in many aspects of colon development.

α-fetoprotein,vascular endothelial growth factor receptor-1 and early recurrence of hepatoma

AIM:To investigate whether α-fetoprotein (AFP) and vascular endothelial growth factor receptor (VEGFR)-1 correlate with early recurrence of hepatoma/hepatocel-lular carcinoma (HCC).METHODS:From 2000 to 2005,114 consecutive pa-tients with HCC underwent primary curative hepatecto-my.The mean age was 60.7 (8.7) years and 94 patients were male.The median follow-up period was 71.2 mo (range:43-100 mo).Immediately prior to commencing laparotomy,5 mL bone marrow was aspirated from thesternum and collected in citrate-coated test tubes.The initial 2 mL of bone marrow aspirate was discarded in each case.AFP mRNA and VEGFR-1 mRNA in the bone marrow and peripheral blood (BM-and PH-AFP mRNA and BM-and PH-VEGFR-1 mRNA,respectively) were measured by real-time quantitative reverse transcription polymerase chain reaction.As normal controls,VEGFR-1 mRNA in the bone marrow and peripheral blood was also measured in 11 living liver donors.These data were evaluated for any correlation with early recurrence,comparing clinical and pathological outcomes.RESULTS:The cut-off value of the BM-AFP mRNA and PH-AFP mRNA level in patients with HCC was set at 1.92 × 10-7 and zero,respectively,based on data from the controls.A total of 34 (29.8%) and six (5.4%) patients were positive for BM-AFP mRNA and PH-AFP mRNA,respectively.The BM-VEGFR-1 mRNA levels in all HCC patients were higher than those in the normal con-trols,and this was the case also for PH-VEGFR-1mRNA.The 25-percentile values for the BM-and PH-VEGFR-1 mRNA in HCC patients were used as the cut-off values for assigning the patients into two groups based on these transcript levels.The High group for BM-VEG-FR-1 mRNA contained 81 (71.1%) HCC cases and the Low group was assigned 33 (28.9%) patients.These numbers for PH-VEGFR-1mRNA were 78 (75.0%) and 26 (25.0%),respectively.HCC recurred in 80 patients;in the remnant liver in 48 cases,in the remnant liver and remote tissue in 20,and in the remote tissue alone in 12.BM-AFP mRNA-positive cases showed a signifi-cantly higher rate of early recurrence (within 1 year of surgical treatment) compared with BM-AFP mRNA-negative patients (P=0.0091).Patients were classified into four groups according to the level/status of their BM-VEGFR-1 and BM-AFP mRNA as follows:group A (n=23),BM-VEGFR-1/BM-AFP mRNA=low/negative;group B (n=57) high/negative;group C (n=10) low/positive;group D (n=24),high/positive.This classifi-cation was found to correlate with a recurrence of thisdisease within 1 year (P=0.0228).The disease-free survival curve of group A was significantly better than that of groups B,C or D (P=0.0437,P=0.0325,P=0.0225).No other classification (i.e.,PH-VEGF-R1/BM-AFP,BM-VEGF-R1/PH-AFP,and PH-VEGF-R1/PH-AFP mRNA) showed such a correlation.CONCLUSION:The evaluation of BM-AFP and BM-VEG-FR-1 mRNA in patients with HCC may be a valuable pre-dictor of disease recurrence following curative resection.

肝细胞癌在血中微小转移的检测

目的 肝细胞癌患者术后复发常是术前不能检出微转移灶或术中瘤细胞释放入血之故。建立早期检出肝细胞癌 (HCC)血中微小转移的方法并评价其临床意义。方法 采用巢式RT PCR检测肝细胞特异白蛋白 (ALB)mRNA和甲胎蛋白 (AFP)mRNA在HCC患者外周血有核细胞成分及组织标本中的表达情况。并以肝癌细胞系HepG2 、SMMC772 1,乳腺癌细胞MCF 7作为阳性、阴性对照。结果 在肝组织 ,无论是癌、癌旁还是正常组织 ,ALBmRNA的表达均为阳性 (8/8) ;而AFPmRNA在癌组织7/8例表达 ,癌旁 5 /8例表达。外周血表达的情况为 :同一组标本ALBmRNA的阳性率为 75 .6 % (34 /4 5 ,P <0 .0 1) ,AFPmRNA的阳性率为 5 7.8% (2 6 /4 5 ,P <0 .0 1)。在有肝外转移和无肝外转移分组中 ,ALBmRNA和AFPmRNA的表达分别为 10 0 .0 % ,5 4.2 %和 90 .5 % ,2 9.2 %。结论 在HCC患者外周血有核细胞成分中检测ALBmRNA和AFPmRNA的表达 ,是预测转移、复发的简捷手段 ,若ALB与AFP二者联合使用 。