Alterations of the Transforming Growth Factor‐(β Signaling Pathway in Hepatocellular Carcinomas Induced Endogenously and Exogenously in Rats

To elucidate involvement of the transforming growth factor‐β (TGF‐β) signaling pathway in endogenous and exogenous liver carcinogenesis, we investigated mutations of TGF‐β receptor type II (TGF‐βRII), Smad2 and Smad4 genes, and expression of TGF‐βRII in hepatocellular carcinomas (HCCs) induced by a choline‐deficient L‐amino acid‐defined (CDAA) diet and by N‐nitrosodiethyl‐amine (DEN). Male Fischer 344 rats received a CDAA diet continuously and HCCs were sampled after 75 weeks. Administration of DEN was followed by partial hepatectomy (PH), with colchicine to induce cell cycle disturbance and a selection pressure regimen, HCCs being obtained after 42 weeks. Total RNAs were extracted from individual HCCs and mutations in TGF‐PRH, Smad2 and Smad4 were investigated by reverse transcription (RT)‐polymerase chain reaction (PCR)‐restric‐tion‐single‐strand conformation polymorphism (SSCP) analysis followed by sequencing analysis. Mutations of Smad2 were detected in 2 out of 12 HCCs (16.7%) induced by the CDAA diet, a GGT‐to‐GGC transition (Gly to Gly) at codon 30 and a TCT‐to‐GCT (Ser to Ala) transversion at codon 118, without any TGF‐βRII or Smad4 alterations. No mutations of TGF‐βRII, Smad2 and Smad4 were encountered in eleven HCCs induced by the exogenous carcinogen. Semi‐quantitative RT‐PCR revealed reduced expression of TGF‐βRII in 2 HCCs (16.7%) without Smad2 mutations out of 12 HCCs induced by the CDAA diet and none of 11 induced by DEN. These results suggest that the TGF‐p signaling pathway may be disturbed in endogenous liver carcinogenesis in rats.