In vitro carcinogenesis of hamster pancreatic duct cells: cellular and molecular alterations

Abstract

Neoplastic transformation of Syrian golden hamster (SGH) pancreatic duct cells was induced by in vitro treatment with the direct-acting carcinogens N-methylnitrosourea (MNU) and N-(2-hydroxypropyl)nitrosourea (HPNU), with subsequent selection by sustained culture in serum- and epidermal growth factor (EGF)-deprived medium. The present study examines the efficacy of serum and EGF deprivation as a selection pressure and the effect of the carcinogen dose, frequency and interval of exposure on tumorigenesis and K-ras mutation. Selection of carcinogen-initiated duct cells by serum and EGF deprivation is highly reproducible and effective, increasing the incidence of tumors from 26 to 93% for MNU or from 0 to 100% for HPNU. SGH pancreatic duct cells exposed to 0.5 mM MNU for 13 weeks (long-treatment schedule) produced K-ras mutations at codon 12 in six of six tumors. However, when cells were exposed to 0.125, 0.25 or 0.5 mM MNU daily for 5 days (short-treatment schedule), mutations of K-ras at codon 13 were identified in four of 16 tumors, the remaining 12 showing no mutations. Duct cells exposed to 0.5 mM HPNU by the short-treatment schedule produced K-ras mutations in codon 13 in six of six tumors, as contrasted to 12 tumors that developed from cells exposed to 0.125 or 0.25 mM HPNU, which all contained K-ras codon 12 mutations. The current experiments demonstrate that K-ras mutation in pancreatic carcinogenesis in vitro by MNU or HPNU can be modified by the nature and dose of the carcinogen as well as the frequency and duration of exposure.

Induction of adenocarcinoma from hamster pancreatic islet cells treated with N-nitrosobis(2-oxopropyl)amine in vitro

“…Moreover, according to our experience the growth of cells deriving from islets is significantly different from that of cultured ductal cells, in that ductal cells form glandular structures even in monolayer (27). Because it has been shown that BOP can transform hamster ductal cells in vitro (20,21), it was of interest to examine the effect of BOP on isolated pure islets in vitro in the system we have established (19). In this system, isolation and long-term maintenance of pure islets free of exocrine cell contamination was possible.…”

“…The number of viable cells, determined by the dye exclusion test, was counted at 24 h intervals using a hemocytometer. Population doubling time was calculated by plotting the mean cell number versus days in culture on a semi-logarithmic graph, as reported (21,22).…”

“…Although presently the origin of these cells from islet cell precursors or from transdifferentiation of islet cells is obscure, the model provided a unique opportunity to test the effect of BOP on cultured islets. Because it has been shown by us (20) and others (21) that BOP can transform isolated pancreatic ductal cells in vitro, we tested its effect on cultured hamster islets.…”

In Vitro Induction of Giant Cell Tumors from Cultured Hamster Islets Treated with N-Nitrosobis(2-Oxopropyl)amine

“…10 We and others have shown that in vitro treatment of hamster ductal cells with pancreatic carcinogens leads to malignant cell transformation and adenocarcinoma in vivo. 5,6 No giant cell carcinomas have ever been produced. On the contrary, treatment of isolated and purified hamster islets with BOP produced tumors of giant cell morphology.…”

“…Treatment of isolated hamster pancreatic ductal cells with BOP or methylnitrosourea (MNU) in vitro resulted in the induction of anaplastic cancer, but no giant cell tumors have been observed. 5,6 A similar treatment scheme of isolated purified hamster pancreatic islets with BOP, however, resulted in formation of anaplastic invasive cancers, which, in vitro and in vivo, presented patterns consistent with human pancreatic giant cell tumor. In this report we describe some biological patterns of this tumor compared to the human tumor.…”