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The Effect of Ascitic Fluid of Patients After Partial Hepatectomy on DNA Synthesis of Adult Rat and Noncancerous Human Hepatocytes in Primary Culture
Yoshiaki Yamane*† and Tetsuo Mura‡
*Division of Chemistry, Institute of Steroid Research, †First Department of Surgery and ‡Department of Pathological Biochemistry, Faculty of Medicine, Tottori University, Yonago 683, Japan
We studied the effect of ascitic fluid of patients who underwent partial hepatectomy for hepatocellular carcinoma (HCC) on DNA synthesis of normal adult rat and noncancerous human hepatocytes in primary culture. Ascitic fluid stimulated DNA synthesis of hepatocytes in a concentration-dependent manner. The mitogenic activity of 10% (vol/vol) ascitic fluid plus 0.1 µmol/L insulin was five times in the controls, and its potency was the same as that of epidermal growth factor (EGF, 10 ng/mL) plus norepinephrine (10 µmol/L) and insulin. Human noncancerous hepatocytes also responded to ascitic fluid (2.3-fold the activity in the controls, P < 0.05). This factor was subsequently further purified about 580-fold from ascitic fluid by a combination of ultrafiltration, cationic exchange chromatography on an S-Sepharose column, and heparin-Sepharose CL-6B affinity chromatography. The partially purified factor was a heat- and acid-labile cationic protein with a molecular weight between 60,000 and 102,000 by gel filtration on a Sephadex G-150 column. Additionally, this growth-promoting fraction could be made to increase 2-fold by having it coexist in serum-free culture with low-molecular-weight fractions having a molecular weight between 10,000 and 45,000 by gel filtration on Sephadex G-75. These results suggest that mitogenic and comitogenic substances exist in the ascitic fluid of patients after partial hepatectomy for HCC, and may lead to a clarification of the activation mechanism of growth stimulant in HCC patients.
Key words: ascites; hepatocellular carcinoma; hepatocyte growth-stimulating factor; partial hepatectomy; rat and human hepatocytes