To analyze gene expression of a-fetoprotein (AFP) and AFP gene expression is shown during massive liver albumin in hepatocellular carcinoma (HCC), messenger necrosis 7 and in hepatocarcinogenesis. On the other RNAs (mRNAs) of these proteins in six human hepatoma hand, albumin gene expression begins with the initiacell lines and in 30 cases of HCC were quantitatively tion of liver organogenesis and increases gradually analyzed by competitive reverse transcription (RT) folwith development. [8][9][10][11] Albumin transcription is known lowed by polymerase chain reaction (PCR). The tranto be modulated by various factors such as hormones, scriptional levels of both AFP and albumin genes in the acute phase reaction, extracellular matrix, heat, HepG2 and Huh 7 cell lines were 10 10 copies/mg RNA, in and serum colloid osmolarity. 12 Previous studies recontrast to approximately 10 5 copies/mg RNA in HLE and vealed the modulation of AFP gene transcription in HLF cell lines. AFP and albumin mRNA levels in three several hepatoma cell lines of rodents. 13,14 However, normal livers were 10 5 and 10 10 transcripts/mg RNA, respectively. In 30 cases with HCC AFP mRNA level in transcriptional levels of AFP and albumin genes in huneoplasm was 10 to 10 5 -fold enhanced as compared with man hepatocellular carcinoma (HCC) and the biological that of nonneoplastic portion, and correlated with sesignificance of the varied AFP transcription have not rum AFP level and tumor size (P õ .01). In contrast, been clarified yet.
albumin mRNA level was not reduced in the neoplasms
In the present study, we have developed competitive presenting enhanced AFP mRNA levels, indicating that reverse transcription followed by polymerase chain re-AFP and albumin gene expression in situ is not necessaraction (RT-PCR) assays to evaluate messenger RNA ily mutually exclusive. Prospective analysis revealed (mRNA) levels of both AFP and albumin in tissue specithat an increased serum AFP was shown at the time of men obtained by biopsy. In addition, both AFP and recurrence among patients with enhanced AFP mRNA albumin mRNA levels in human HCC were quantitated levels in neoplasm only, indicating that AFP mRNA levels in neoplasm could be a clinically predictable tool. in comparison with those of nonneoplastic portion to (HEPATOLOGY 1996;23:1384-1392.)
elucidate AFP and albumin gene expression in human liver carcinogenesis. In humans, a-fetoprotein (AFP) and albumin genes
PATIENTS AND METHODS
are present in tandem, about 14.5 kbp apart, on chromosome 4q11-q13 and consist of 15 exons. 1-5 AFP is Cell Lines. Six human hepatoma cell lines were used.
an oncofetal protein, and its gene expression is shown HepG2 15 was obtained from Riken Cell Bank (Tokyo, Japan). mainly by hepatocytes and endodermal cells of the vis-Huh 7, 16 Huh 6, 17 PLC/PRF/5, 18 HLE, and HLF 19 were kind ceral yolk sac in the early stages of fetal liver developgifts of the Japanese Cancer Resource Bank (Tokyo, Japan). ment, but repressed soon after birth. 6 Reactivation of The cells were plated at a density of 5 1 10 5 cells/cm 2 in plastic flasks (25 cm 2 ; Corning Costar, Cambridge, MA) in RPMI 1640 medium (GIBCO BRL, Gaithersburg, MD) supplemented with 1% fetal calf serum (GIBCO BRL), and incu-Abbreviations: AFP, a-fetoprotein; HCC, hepatocellular carcinoma; RTbated at 37ЊC in a humidified atmosphere of 95% air: 5% CO 2 PCR, reverse-transcription polymerase chain reaction; mRNA, messenger for 48 hours. Medium was then changed and the cells were RNA.