886 HTERT PROMOTER GENE POLYMORPHISM AND THE RISK OF HEPATOCELLULAR CARCINOMA (HCC) IN PATIENTS WITH CHRONIC HEPATITIS B

Background and Aims: Many investigations have revealed that a low recurrence rate of hepatocellular carcinoma (HCC) is associated with high serum albumin levels in patients; therefore, high levels of serum albumin comprise a major indicator of a favorable prognosis. The mechanism inhibiting the proliferation of HCC has not yet been elucidated, so we investigated the effect of serum albumin on HCC cell proliferation. Methods: Hep3B was cultured in MEM with no serum or containing 5 g/dl bovine albumin. As the control samples, Prionex (Polysciences, Inc.) was added to generate the same osmotic pressure as albumin. After 24-hour incubation, the expressions of afetoprotein (AFP), p53, p21, and p57 were evaluated with real-time PCR using total RNA extracted from the liver. Protein expressions and the phosphorylation of Rb (retinoblastoma) were determined by Western blot analysis using total protein extracted from the liver. For flow cytometric analysis of the cell cycle, FACS analysis was performed. The percentages of cell cycle distribution were evaluated by PI staining, and all samples were analyzed employing FACScalibur (BD) with appropriate software (ModFit LT, BD). Results: The mRNA levels of AFP relative to Alb(−): Alb(−), Alb(+), and Prionex, were 1.0.7±0.2 (p < 0.001 for Alb(−)), and 1±0.3, respectively. The mRNA levels of p21 were 1, 1.58±0.4 (p = 0.007 for Alb(−) and p=0.004 for Prionex), and 0.8±0.2, respectively. The mRNA levels of p57 were 1, 4.4±1.4 (p = 0.002 for Alb(−) and Prionex), and 1.0±0.1, respectively. The protein expression levels of Rb were similar within all culture media. The phosphorylation of P807/811 and P780 of Rb protein was reduced in Alb(+). More cells in the G0/G1 phase and less cells in S and G2/M phases were obtained in Alb(+) than in Alb(−) (G0/G1: 45.2, 49.5, 44.7%; G2/M: 44.3, 41.5, 43.0%; S: 10.5, 7.4, 17.2%, Alb(−), Alb(+), Prionex, respectively). The same results were obtained in HepG2. Conclusions: The presence of albumin in serum reduces the phosphorylation of Rb proteins and enhances the expression of p21 and p57, following an increase in the G0/G1 cell population, and suppresses cell proliferation. These results suggest that albumin itself suppresses the proliferation of hepatocellular carcinoma.