肝癌外周血AFP mRNA的测定及意义

Detection of AFP mRNA in Peripheral Blood of Patients with Hepatocellular Carcinoma by Nested Reverse Transcription Polymerase Chain Reaction

目的建立一种灵敏的巢式逆转录聚合酶链反应法（RT－PCR），用于检测外周血中的AFPmRNA。方法将患者分成四组：原发性肝细胞癌组38例，慢性肝炎组9例，肝炎后肝硬化组12例，正常对照组10例。通过采肝素抗凝外周血、分离单个核细胞、抽提RNA、逆转录合成cDNA、设计AFP基因引物来进行巢式PCR扩增。结果所有正常人外周血中AFPmRNA全部阴性，原发性肝细胞癌组的外周血中AFPmRNA阳性率（23／38，60．5％）明显高于慢性肝炎组（2／12，16．7％，P＜0．01）和肝硬化组（1／9，11.1％，P＜0．01）。有远处转移灶的肝癌病例其外周血AFPmRNA全部阳性（8／8，100％）。结论RT－PCR技术检测外周血AFPmRNA可作为反映肝癌细胞血行播散的指标。

Objective A sensitive nested reverse transcription polymerase chainreaction(RT-PCR) was established for detection of AFP mRNA. Methods Heparinizedblood samples were obtained from 38 patients with hepatocellular carcinoma (HCC), 9patients with chronic hepatitis, 12 patients with liver cirrhosis and 10 normal subjects.Mononuclear cells were separated from blood. Total RNA was extracted by the acid -guanidium - phenol - chloroform method. The first strand of cDNA was synthesized from analiquot of the RNA using Moloney murine leukemia virus reverse transcriptase and randomhexamers. The two pairs of primers chosen from the sequences of AFP were used for thetwo - step PCR. Rcsults AFP mRNA in perlpheral blood was not detected in all normalsubjects, but it was detected in 23 of 38 (6O.5 % ) cases with HCC. The positive rate in patientswith HCC was higher than the cases with cirrhosis (2/l2, 16.7%, P<0.O1) and the cases withchronic hepatitis (1/9, 11.1 %, P<0.Ol). AFP mRNA was found in the peripheral blood of all 8HCC patients with distant metastasis. Conclusion The detection of AFP mRNA inperipheral blood may be helpful for clinical detection of metastasis of HCC.