Natural expression of feline type-C virus genomes. Prevalence of detectable FeLV and RD-114 GS antigen,type-C particles and infectious virus in postnatal and fetal cats

The group-specific (gs) antigen of RD-114, an endogenous type-C virus of domestic cats, was detected by complement fixation (CF) in two of 100 fetal cats and was not detected in any tumors or spleens of postnatal cats. RD-114 virus was isolated in vitro in RD human sarcoma cells from two of 31 normal cat fetuses but not from any postnatal cat tumors or spleens, with one exception. Feline leukemia virus (FeLV) gs antigen was detected by CF and type-C particles were seen by electron microscopy (EM) in high prevalence in young cats with lymphoma (65%), anemia (75%) and infectious peritonitis (25%). FeLV gs antigen, type-C particles and infectious FeLV were occasionally (10-20%) found in cat fetuses. The general absence of FeLV gs antigen in the endometrium of pregnant cats and the low prevalence of detectable FeLV gs antigen in cat fetuses suggest that the epigenetic transplacental spread of infectious FeLV is not a common occurrence in domestic cats. However, the isolation of FeLV from two pairs of fetal littermates and from the endometria and spleens of both mother cats indicates that such an epigenetic transmission of FeLV can occasionally occur. There was an excellent correlation between detection of RD-114 and FeLV gs antigen by CF tests in fetal and postnatal tissues and the subsequent isolation in vitro of the corresponding virus. In lymphoma- and carcinoma-bearing cats over 10 years of age, FeLV gs antigen, type-C particles and infectious virus were seldom found. RD-114 and FeLV gs antigens or infectious virus were never detected in the same tissues, the same cat or the same litter. These findings are discussed in relation to the natural history of these two different feline type-C virus genomes.     

Shortened telomere length and increased telomerase activity in hamster pancreatic duct adenocarcinomas and cell lines

Recently, shortened telomere length and increased telomerase activity have been demonstrated in various human cancers. In the study reported here, we ascertained whether gene changes are characteristic of pancreatic cancers. Hamster duct carcinomas and cell lines were investigated by Southern blot analysis for telomere restriction fragment (TRF) length and by the telomeric repeat amplification protocol (TRAP) assay for telomerase activity. Comparison with normal pancreas and spleen revealed shortened TRF length and markedly increased telomerase activity in primary pancreatic duct carcinomas induced by the rapid-production model as well as in a transplantable carcinoma and the cell lines. The enzyme level was 86.0–215.7 times the low levels found in control pancreas and spleen tissues. Late-passage Syrian hamster embryo cells, known to be immortalized and tumorigenic, had shorter TRFs than the original cells in primary culture did. These results indicate that hamster pancreatic duct carcinoma cells are immortalized, with the potential for proliferation ad infinitum, and provide a model for basic therapeutic research into the substances targeting telomerase. Mol. Carcinog. 18:153–159, 1997. © 1997 Wiley-Liss, Inc.     

Purification and analysis of glycoproteins bearing blood group-A determinants from hamster pancreatic ductal adenocarcinomas.

Although the hamster pancreas does not express A, B or H blood group antigens, all hamster pancreatic ductal adenocarcinomas induced by treatment with N-nitrosobis(2-oxopropyl)amine express blood group-A antigen. Thus, the acquisition of blood group-A antigen expression in this system is a cancer-associated alteration. We have purified three major blood group-A antigen bearing glycoproteins (gp120, gp135 and gp150) from hamster pancreatic cancer cell membrane preparations using affinity chromatography on DBA (Dolichos biflorus) agglutinin-agarose. When assayed by immunoblotting, gp120 and gp135 showed strong blood group-A reactivity, which was removed by treating membrane samples with peptide-N-glycosidase F. Blood group-A reactivity was unchanged by treatment of the membrane fractions with endoglycosidases F and H. In addition, these two glycoproteins bearing blood group-A antigen also bound L-PHA (Phaseolus vulgaris leucoagglutinin). These results demonstrate that gp120 and gp135 express blood group-A antigen on Asn-linked multi-antennary complex type glycan structures. The gp150 showed weak blood group-A expression. This is the first demonstration of the neoexpression of cancer-associated blood group-A determinants which reside on Asn-linked glycan structures.     

Typical type-C virus in human leukemia.

The virus HL-23, recently isolated from a patient with myelogenous leukemia, was compared by electron microscopy with virus-like particles observed in human tissues and with several mammalian type-C virus particles. It closely resembled the typical type-C virus particles of murine, feline, and nonhuman primate species, and differed from all particles previously reported in human tissues, including the atypical human placental particle.     

Interaction between Epstein-Barr virus and type-C virus in human cells.

The interaction between EBV and type-C viruses was studied in our FVNC experimental system, in which EBV and type-C viral genomes are contained in each cell. The data indicate that the human lymphoid FVNC cells are sensitive to both EBV and type-C virus exposure, showing high frequencies of induction of both repressed viral genomes. The two different viral genomes may be associated with different chromosomes in individual cells.     

Oncogenicity of gibbon type-C myelogenous leukemia virus

Young gibbons that were experimentally inoculated with cell-free gibbon ape leukemia virus (GaLV) and developed peristent viremia subsequently developed chronic granulocytic leukemia (CGL) with associated multifocal bone lesions and metastases. An 8-month-old gibbon inoculated with 10⁵ tissue culture infectious virus (TCIV) developed acute myeloproliferative disease with associated bone lesions after a latency of 5 months, while a 9-month-old gibbon inoculated with 10³ TCIV developed CGL after an II-month latency. The clinical symptoms associated with the onset of leukemia were an increased number of leukocytes which were predominantly mature granulocytes, development of anemia, and multifocal bone lesions. Terminally, the animals had elevated immature granulocytes in the blood, cellular bone marrow with a predominant number of immature granulocytes, and hepatosplenomegaly. The gibbon with CGL had metastatic growth in the spleen and lung. Two 14-month-old gibbons that were inoculated with 10³ TCIV and developed persistent neutralizing antibody to the virus infection remained free of hematopoietic disease, as did uninoculated animals. The fact that only animals with persistent viremia developed leukemia supports the oncogenicity of GaLV in gibbons.     

Cyclooxygenase-2 expression in human pancreatic adenocarcinomas

Cyclooxygenase-2 (COX-2) expression is up-regulated in several types of human cancers and has also been directly linked to carcinogenesis. To investigate the role of COX-2 in pancreatic cancer, we evaluated COX-2 protein expression in primary human pancreatic adenocarcinomas (n = 23) and matched normal adjacent tissue (n = 11) by immunoblot analysis. COX-2 expression was found to be significantly elevated in the pancreatic tumor specimens compared with normal pancreatic tissue. To examine whether the elevated levels of COX-2 protein observed in pancreatic tumors correlated with the presence of oncogenic K-ras, we determined the K-ras mutation status in a subset of the tumors and corresponding normal tissues. The presence of oncogenic K-ras did not correlate with the level of COX-2 protein expressed in the pancreatic adenocarcinomas analyzed. These observations were also confirmed in a panel of human pancreatic tumor cell lines. Furthermore, in the pancreatic tumor cell line expressing the highest level of COX-2 (BxPC-3), COX-2 expression was demonstrated to be independent of Erk1/2 activation. The lack of correlation between COX-2 and oncogenic K-ras expression suggests that Ras activation may not be sufficient to induce COX-2 expression in pancreatic tumor cells and that the aberrant activation of signaling pathways other than Ras may be required for up-regulating COX-2 expression. We also report that the COX inhibitors sulindac, indomethacin and NS-398 inhibit cell growth in both COX-2-positive (BxPC-3) and COX-2-negative (PaCa-2) pancreatic tumor cell lines. However, suppression of cell growth by indomethacin and NS-398 was significantly greater in the BxPC-3 cell line compared with the PaCa-2 cell line (P = 0.004 and P < 0.001, respectively). In addition, the three COX inhibitors reduce prostaglandin E(2) levels in the BxPC-3 cell line. Taken together, our data suggest that COX-2 may play an important role in pancreatic tumorigenesis and therefore be a promising chemotherapeutic target for the treatment of pancreatic cancer.     

Isolation of a B-tropic type-C virus from reticulum cell neoplasms induced in BALB/c mice by SJL/J type-C virus.

D1-murine leukemia virus (MuLV), an N-tropic type-C virus isolated from a spontaneous reticulum cell neoplasm, type B (RCN-B) of an SJL/J mouse was propagated in NIH Swiss mouse embryo cell cultures. When injected into BALB/c mice 1 day after neonatal thymectomy, 30% of the inoculated mice developed RCN-B in 5 months, whereas none of the uninoculated controls did. From the spleen and lymph node extracts of all RCN-B-bearing mice tested, B-tropic type-C viruses (designated E1-MuLV) were isolated in high titers (10-5 minus 10-6 XC plaque-forming units/ml). Only low titers (10-1 minus 10-2 XC plaque-forming units/ml) of N- or B-tropic viruses were isolated from those thymectomized mice, inoculated but nontumorous, whereas only N-tropic viruses were detected in the uninoculated thymectomized mice. No virus was isolated from the nonthymectomized, untreated controls. Antigenically, the viral envelope antigen (VEA) of E1-MuLV was distinct from those of DU-MuLV, xVEA, or Gross-VEA, but some cross reaction with AKR-MuLV-VEA was observed. The relationship of D1-MuLV to E1-MuVL with respect to oncogenesis and viral genome activation was discussed.     

Fibrin deposition in autochthonous Syrian hamster pancreatic adenocarcinomas induced by the chemical carcinogen N-nitroso-bis(2-oxopropyl)amine

Immunofluorescence studies have demonstrated the presence of fib (a group of fibrinogen- and fibrin-related proteins that react with antibodies raised against fibrinogen) in the stroma of several transplantable animal and autochthonous human tumors. Acceptance of these reports was tempered by the possibility of artifactual clotting and fibrinolysis associated with tumor removal or tumor transplantation and by the relatively poor histology inevitable when immunofluorescence is performed on frozen tissue sections. An immunoperoxidase study therefore was undertaken of the ductal pancreatic carcinomas induced in female LGV Syrian hamsters by N-nitroso-bis(2-oxopropyl)amine [(BOP) CAS: 60599-38-4]. Artifactual clotting and fibrinolysis associated with tumor removal were avoided by systemic anticoagulation and antifibrinolysis. Fibronectin and residual fib were prominent components of tumor stroma. Prominent fib deposits also were found in a new location: the basement membrane zones of atypical pancreatic ducts and invasive carcinomas. In contrast, fib deposits were never found in the basement membranes of blood vessels, nerves, or pancreatic acini of BOP-treated or normal animals, or in the ductal basement membranes in the normal pancreas. Ducts with marked atypicality and invasive pancreatic carcinomas frequently exhibited discontinuous basement membrane staining for fib, which often paralleled loss of staining for the integral basement membrane proteins--type IV collagen and laminin. Loss of acquired fib basement membrane staining with malignant disease progression may serve as a new marker for local tumor invasion.