Saso K, Moehren G, Higashi K, Hoek J B
Department of Pathology, Anatomy and Cell Biology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
Gastroenterology. 1997 Jun;112(6):2073-88. doi: 10.1053/gast.1997.v112.pm9178701.
BACKGROUND & AIMS: Long-term ethanol intake suppresses liver regeneration in vivo and ethanol interferes with epidermal growth factor (EGF)-induced DNA synthesis in vitro. Therefore, the effects of long-term ethanol treatment on EGF-activated signaling reactions in rat hepatocytes were investigated.
Hepatocytes from long-term ethanol-fed rats and pair-fed controls were stimulated with EGF (0.5-20 nmol/L) for 15-120 seconds. Tyrosine phosphorylation of EGF receptor (EGFR), Shc, and phospholipase-C gamma1 (PLC gamma), and growth factor receptor binding protein 2 (Grb2) coprecipitation with EGFR and Shc were analyzed by Western blotting.
EGFR autophosphorylation was suppressed at all EGF concentrations in ethanol-fed cells compared with pair-fed cells, without significant differences in total EGFR protein or EGFR tyrosine kinase activity detected in cell lysates, suggesting that intracellular factors suppressed EGFR function. EGF-induced PLC gamma tyrosine phosphorylation and inositol 1,4,5-trisphosphate (InsP3) formation were suppressed, but cytosolic [Ca2+]c elevation was little affected, indicating enhanced InsP3-mediated intracellular Ca2+ release in ethanol-fed cells. Grb2 binding to EGFR was suppressed, but EGF-induced Shc tyrosine phosphorylation and Grb2 association with Shc were not significantly decreased.
Long-term ethanol feeding suppressed EGF-induced receptor autophosphorylation in rat hepatocytes with differential inhibition of downstream signaling processes mediated by PLC gamma, Shc, and Grb2. Altered patterns of downstream signals emanating from EGFR may contribute to deficient liver regeneration in chronic alcoholism.
长期摄入乙醇会抑制体内肝脏再生，且乙醇在体外会干扰表皮生长因子（EGF）诱导的DNA合成。因此，本研究探讨了长期乙醇处理对大鼠肝细胞中EGF激活的信号反应的影响。
用EGF（0.5 - 20 nmol/L）刺激长期乙醇喂养大鼠和配对喂养对照大鼠的肝细胞15 - 120秒。通过蛋白质免疫印迹法分析EGF受体（EGFR）、Shc和磷脂酶Cγ1（PLCγ）的酪氨酸磷酸化，以及生长因子受体结合蛋白2（Grb2）与EGFR和Shc的共沉淀情况。
与配对喂养的细胞相比，在所有EGF浓度下，乙醇喂养细胞中的EGFR自磷酸化均受到抑制，而在细胞裂解物中检测到的总EGFR蛋白或EGFR酪氨酸激酶活性无显著差异，这表明细胞内因子抑制了EGFR功能。EGF诱导的PLCγ酪氨酸磷酸化和肌醇1,4,5 - 三磷酸（InsP3）形成受到抑制，但胞质[Ca2+]c升高受影响较小，这表明乙醇喂养细胞中InsP3介导的细胞内Ca2+释放增强。Grb2与EGFR的结合受到抑制，但EGF诱导的Shc酪氨酸磷酸化以及Grb2与Shc的结合未显著降低。
长期乙醇喂养抑制了大鼠肝细胞中EGF诱导的受体自磷酸化，并对由PLCγ、Shc和Grb2介导的下游信号传导过程产生不同程度的抑制。EGFR发出的下游信号模式改变可能导致慢性酒精中毒时肝脏再生不足。
