Sisir K. Chattopadhyay

A quantitative in vitro focus assay for bovine papilloma virus.

Abstract A quantitative tissue culture assay for bovine papilloma virus (BPV) has been developed. Extracts of bovine papillomas which contained either BPV-1 or BPV-2 induced distinct foci in two mouse cell lines (NIH 3T3 and C127). Focus formation followed single-hit kinetics, suggesting that a single particle was required for focus induction. Preincubation of extracts with rabbit anti-BPV-1 serum neutralized their infectivity. The morphologically transformed cells contained BPV DNA sequences, grew in low serum and in soft agar, and were tumorigenic for nude mice.

Cellular origin and role of mink cell focus-forming viruses in murine thymic lymphomas

Embryo DNA of AKR mice contains several copies of intact mink cell focus-forming (MCF)-like env gene sequences with a proviral structure. In spontaneous thymic tumours, these MCF-like env sequences have regularly recombined with a specific region of the 3′ end of ecotropic env. This specific recombination is probably a critical event in the pathogenesis of spontaneous AKR tumours.

Origin of mink cytopathic focus-forming (MCF) viruses:Comparison with ecotropic and xenotropic murine leukemia virus genomes

Abstract Restriction endonuclease maps have been developed for the viral DNAs from nine xenotropic and eleven MCF murine leukemia viruses (MuLV) isolated from AKR and other mouse strains. In contrast to the highly related nature of ecotropic viral DNAs isolated from inbred mice and from M. m. molossinus , each xenotropic and MCF viral DNA was unique. Xenotropic MuLV DNAs could be divided into two classes, which correlated with previously reported serological and biochemical data; one xenotropic MuLV isolated from an AKR mouse showed features of both classes. Ecotropic viral DNA hybridized poorly or not at all to a 1.2 kbp segment of xenotropic viral DNA located in env 6.3–7.5 kbp from the left end of the viral DNAs. All MCF viral DNAs contained noneco-tropic sequences in a portion of the env gene region, but some MCF viruses were composed principally of nonecotropic sequences. The nonecotropic regions of the MCF viral DNAs were related to xenotropic MuLV DNA, but many MCF viral DNAs contained sequences not found either in xenotropic or ecotropic MuLV DNA. It was concluded that these MCF viruses probably arose via recombination between ecotropic MuLV and endogenous MuLV DNA sequences; sequences of recombination include a portion of env, but need not be limited to this region. The polytropic host range of MCF viruses may represent an endogenous viral function.

Pathology of Aging B6;129 Mice

Fifty male and 49 female B6;129 mice (wild-type, + / + ) were maintained until 2 years of age to study their age-related pathology. By 104—105 weeks, 14/50 (28%) of the males and 30/49 (61%) of the females were still alive. The most common contributing cause of morbidity or mortality was lymphoma. Lymphoma was observed in 21/50 (42%) of the males and 33/49 (67%) of the females with the most common sites being mesenteric lymph nodes, gut associated lymphoid tissue (Peyers patches), and spleen. The lymphoma most often appeared to arise in the mesenteric node. Immunohistochemistry revealed CD45R expression as well as infiltration by many CD3+ T cells. IgH gene rearrangements were found in typical mesenteric node lymphomas indicating B-cell origin. They bore similarities to the human T-cell rich, B-cell lymphomas. Other tumors included hepatocellular adenoma or carcinoma (male 12%, females 10%), lung alveolar Type II cell adenoma or carcinoma (male 32%, female 20%), thyroid follicular adenoma or carcinoma (male 2%, female 8%), ovarian tumors (17%), and endometrial tumors (6%). Nonneoplastic lesions included amyloidlike material in the nasal septum (male and female 100%), otitis media (male 84%, female 79%), epididymal epithelial karyomegaly (88%), melanosis (high incidences in various tissues including brain, parathyroid, and spleen), membranoproliferative glomerulonephritis (male 52%, female 71%), hyalinosis with extracellular crystals in several tissues (respiratory tract, gall bladder, stomach), islet cell hyperplasia (male 45%, female 29%) and esophageal dilation (male 10%, female 6%). The B6;129 mouse is a mouse with aging lesions similar to those in other mouse strains but with a characteristic common lymphoma.

Accelerated Appearance of Multiple B Cell Lymphoma Types in NFS/N Mice Congenic for Ecotropic Murine Leukemia Viruses

Spontaneous lymphomas occur at high frequency in NFS.V+ mice, strains congenic for ecotropic murine leukemia virus (MuLV) proviral genes and expressing virus at high titer. In the present study, a total of 703 NFS.V+ lymphomas were studied by histopathology, immunophenotypic analysis, immunoglobulin heavy chain or T cell receptor β chain rearrangements, and somatic ecotropic MuLV integrations; 90% of the lymphomas tested were of B cell lineage. Low-grade tumors included small lymphocytic, follicular, and splenic marginal zone lymphomas, while high-grade tumors comprised diffuse large-cell (centroblastic and immunoblastic types), splenic marginal zone, and lymphoblastic lymphomas. Comparison of mice of similar genetic background except for presence (NFS.V+) or absence (NFS.V−) of functional ecotropic MuLV genomes showed that NFS.V− clonal lymphomas developed at about one-half the rate of those occurring in NFS.V+ mice, and most were low-grade B cell lymphomas with extended latent periods. In NFS.V+ mice, clonal outgrowth, defined by Ig gene rearrangements, was associated with acquisition of somatic ecotropic proviral integrations, suggesting that, although generation of B cell clones can be virus independent, ecotropic virus may act to increase the rate of generation of clones and speed their evolution to lymphoma. The mechanism remains undefined, because only rare rearrangements were detected in several cellular loci previously associated with MuLV insertional mutagenesis.

Splenic Marginal Zone Lymphomas of Mice

Splenic marginal zone lymphomas (MZLs) have been found to occur at a high frequency in NFS.N mice congenic for high-expressing ecotropic murine leukemia virus (MuLV) genes from AKR and C58 mice. Based on morphological, immunological, and molecular studies of these mice, MZL is clearly recognizable as a distinct disease with a characteristic clinical behavior. MZL was staged according to the degree of accumulation and morphological change of cells within the splenic marginal zone, as follows: 1) a moderate increase in normal-looking MZ cells, judged to be prelymphomatous, and 2) MZL in three variants: i) distinct enlargement of MZ by normal-looking cells (MZL), ii) distinct enlargement of MZ by basophilic centroblast-like cells (MZL+), and iii) extensive splenic involvement by centroblast-like cells (MZL++). The rate of mitosis and apoptosis increases with lymphoma grade. In most cases, emergence of a dominant IgH clonal pattern in paired splenic biopsy and necropsy samples was correlated with progression. MZLs were transplantable and homed to the spleen. MZL may constitute a commonly occurring lymphoma type unrecognized, in part, because of the centroblastic morphology of high-grade MZL and possible overgrowth of lower-grade MZL by more aggressive follicular lymphomas.

Molecular phylogeny of Fv1

Abstract. Alleles at the Fv1 gene of inbred mice confer resistance to infection and spread of vertically or horizontally transmitted murine leukemia viruses (MuLV). The nucleotide sequence of Fv1 bears similarity to the gag of a human endogenous retrovirus, HERV-L, but is more closely related to the gag-coding sequence of a newly described class of HERV-L-related mouse endogenous retroviruses designated MuERV-L. Both observations suggest an origin of Fv1 from endogenous gag sequences. The molecular definition of Fv1 provided an opportunity to determine the phylogeny of the gene among wild mice and its relation to MuERV-L. PCR primers, chosen to include most of the coding region of Fv1 for both the n and b alleles, were used to amplify sequences from animals of the genus Mus, which were then sequenced. Closely related products were obtained from almost all animals examined that evolved after the separation from Rattus, in which the homologous gene was shown to be absent. A phylogenetic tree generated with Fv1 sequence data differs noticeably from that developed with sequence data from other genes. In addition, non-synonymous changes were found to be present twice as frequently as synonymous changes, a fact that departs from the standard behavior of a structural gene. These observations suggest that the Fv1 gene may have been subjected to possible horizontal transfers as well as to positive Darwinian selection.

Amplification and rearrangement of onc genes in mammalian species

Related eukaryotic species usually contain the same number of copies per cell of a given ‘unique sequence’ gene. In the few described exceptions, such as the preproinsulin1,2 and globin genes3–7, one or two additional copies per cell have been found in several related species, suggesting that germ-line amplification occurred millions of years ago in a common progenitor of these species. The transforming (onc) genes of retroviruses are a group of evolutionarily conserved genes for which at least 10 distinct members have been described (for a review see ref. 8). Sequences related to each onc have been identified in all vertebrate species tested, usually as one copy per haploid genome, although the rat has at least two different genes homologous to the onc of the rat-derived Harvey murine sarcoma virus (Ha-MuSV)9. In screening genomic DNA from several rodent species for sequences related to the onc of Ha-MuSV and to the closely related (but distinguishable) onc of Kirsten (Ki) MuSV10, we have now found evidence for relatively recent amplification of these genes. We report here that Mus pahari apparently contains at least 10 copies of Ha-MuSV-type onc, whereas most other Mus species contain only one or two copies. Similarly, Chinese hamsters (Cricetulus iseus) have about six copies of the Ki-MuSV-type onc compared with only one copy in Syrian hamsters (Mesocricetus auratus).

Combined histiologic and molecular features reveal previously unappreciated subsets of lymphoma in AKXD recombinant inbred mice

Hematopoietic neoplasms developing in AKXD recombinant inbred, NFS.V(+) and ICSBP knockout mice were assessed using morphologic, cytologic and molecular criteria that relate these disorders to human lymphoma and leukemia. Lymphoma types included precursor T-cell and B-cell lymphoblastic, small lymphocytic, splenic marginal zone, follicular, and diffuse large cell (DLCL). In addition to previously defined subtypes of DLCL composed of centroblasts or immunoblasts, two additional subtypes are defined here: lymphoblastic lymphoma like (LL) and lymphoma characterized by a histiocytic reaction (HS). DLCL(HS) were distinguished from true histiocytic lymphomas by the presence of clonal Ig gene rearrangements.

Lymphomas and High-Level Expression of Murine Leukemia Viruses in CFW Mice

ABSTRACT Historically, Swiss Webster mice of the CFW subline, both inbred and random-bred stocks, have been considered to have a low spontaneous occurrence of hematopoietic system tumors, and previous reports of infectious expression of murine leukemia viruses (MuLVs) have been rare and unremarkable. In marked contrast, in the present study of CFW mice from one source observed by two laboratories over a 2-year period, nearly 60% developed tumors, 85% of which were lymphomas, the majority of B-cell origin. All tumors tested expressed ecotropic MuLVs, and most expressed mink cell focus-inducing (MCF) MuLVs. Among normal mice of weanling to advanced age, over one-half were positive for ecotropic virus in tail or lymphoid tissues, and MCF virus was frequently present in lymphoid tissue, less often in tail. Patterns of ecotropic proviral integration indicated that natural infection occurred by both genetic and exogenous routes. Lymphomas were induced in NIH Swiss mice infected as neonates with tissue culture-propagated MuLVs isolated from normal and tumor tissue of CFW mice.