Munehiro Yokoyama

Attenuated ALK5 receptor expression in human pancreatic cancer: Correlation with resistance to growth inhibition

Transforming growth factor‐β (TGF‐β) receptors constitute a family of transmembrane proteins that bind TGF‐β ligands. In this study we assessed the growth responsiveness to TGF‐β1 in pancreatic cancer cell lines and characterized the levels of expression of TGF‐β receptors in these cell lines and in human pancreatic cancer tissues. COLO 357 cells were most sensitive to the growth inhibitory actions of TGF‐β1, PANC‐1 cells exhibited moderate sensitivity, Hs766T cells exhibited slight sensitivity and MIA PaCa‐2 and T3M4 cells were resistant to TGF‐β1. Only COLO 357 cells expressed high levels of ALK5, the major type I TGF‐β receptor (TβRI). Hs766T and PANC‐1 cells expressed high levels of SKRI, another TβRI subtype. Only MIA PaCa‐2 cells did not exhibit the type II TGF‐β receptor (Tβ‐RII) transcript, whereas type III TGF‐β receptor (Tβ‐RIII) mRNA levels were elevated in this cell line and in HS766T cells. All the cell lines expressed TGF‐β1, but TGF‐β2 and TGF‐β3 mRNA levels were variable. ALK5 and SKRI mRNA levels were 6.8‐ and 9‐fold greater in the pancreatic tumors in comparison with the corresponding levels in the normal pancreas. However, in the cancer cells, ALK5 immunoreactivity was faint, whereas TβRII immunoreactivity was focal and intense. Conversely, in ductal cells adjacent to cancer cells ALK5 immunoreactivity was strong, whereas TβRII immunoreactivity was weak. Since ALK5 heterodimerization with TβRII is crucial for TGF‐β‐mediated signaling, our findings suggest that low levels of ALK5 in pancreatic cancer cells within a tumor may protect against growth inhibition.

Co‐expression of heparin‐binding EGF‐like growth factor and related peptides in human gastric carcinoma

Heparin‐binding epidermal growth factor (EGF)‐like growth factor (HB‐EGF) is a member of the EGF family of polypeptide growth factors, which includes EGF, transforming growth factor α (TGF‐α), amphiregulin (AR) and betacellulin (BTC). To assess the potential role of HB‐EGF in human gastric carcinomas, the expression of HB‐EGF and EGF receptor (EGF‐R) was examined in normal and cancerous gastric tissues and cultured gastric cancer cell lines. By Northern blot analysis, there was a 4.7‐fold increase in HB‐EGF mRNA levels in human gastric cancers compared with normal gastric tissues. There was a concomitant 3.9‐fold increase in EGF‐R mRNA levels in these cancers. Immunostaining revealed co‐localization in 72% of the cancer cells of HB‐EGF and EGF‐R. AR and BTC moieties were not evident by Northern blot analysis. However, using PCR, both AR and BTC mRNA species were demonstrated in normal and cancerous gastric tissues. By Northern blot analysis, HB‐EGF, TGF‐α, AR, BTC and EGF‐R mRNA moieties were co‐expressed in KATO III and NCI‐N87 gastric cancer cell lines. Furthermore, HB‐EGF, EGF and TGF‐α enhanced the growth of both cell lines in a dose‐dependent manner. Our findings suggest that HB‐EGF is relatively abundant in human gastric cancers and that co‐expression of the EGF ligand family may lead to excessive activation of EGF‐R in this disorder.

Alteration of Fibroblast Growth Factor and Receptor Expression After Acute Pancreatitis in Humans

After an episode of acute pancreatitis (AP) in humans, the pancreas exhibits varying degrees of fibrosis, acinar cell regeneration, and the formation of tubular complexes. The mechanisms underlying these changes are unknown. By using Northern blot analysis and immunohistochemistry, we assessed the expression and distribution of fibroblast growth factors (FGFs) and their receptor in human pancreatic tissues obtained from patients with AP. By comparison with the normal pancreas, acidic FGF (aFGF) and FGF-receptor 1 (FGFR-1, flg), mRNA levels were significantly decreased in human pancreatic tissues, which were obtained approximately 8 days after onset of AP. In the normal pancreas, acidic FGF, basic FGF, and FGFR-1 immunoreactivity was present at low to moderate levels in a heterogeneous pattern in the cytoplasm of acinar and ductal cells. Two patterns of immunoreactivity were observed in the AP group. In regions that have undergone necrosis, there was complete loss of aFGF, bFGF, and FGFR-1 immunostaining. In contrast, in the regenerating areas, aFGF and bFGF were readily evident in exocrine-type cells, in association with a marked increase in FGFR-1 immunoreactivity. These findings indicate that FGF/receptor expression is altered after AP, and raise the possibility that FGFs may be involved in the process of pancreatic exocrine regeneration during recovery from AP.

Single-strand conformation polymorphism analysis of the epidermal growth factor receptor at codon 497.

A variant human epidermal growth factor (EGF) receptor (HER) with a transition (G to A) at codon 497, resulting in a substitution of a lysine for an arginine, was recently demonstrated in several human pancreatic cancer cell lines. In the present study, we compared the frequency of expression of wild-type (HER497R) and variant (HER497K) EGF receptors in normal and malignant pancreatic tissue samples using single-strand conformation polymorphism analysis. Both receptors were expressed in normal (42%) and malignant (46%) tissues. Three samples (two normal and one cancer) expressed only HER497K. The cancer sample that was homozygous for HER497K exhibited strong EGF receptor immunore-activity. The high frequency of HER497K expression suggests that it is due to a relatively common polymorphism in the HER coding region. Its presence in some of the cancer samples suggests that, like the wild-type receptor, HER497K also has a role in pancreatic cancer.