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      Mouse mammary tumour virus-like env nucleotide and p14 signal peptide are present in feline mammary carcinomas, but not in neoplastic or dysplastic canine mammary lesions

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          Abstract

          Mouse mammary tumour virus-like (MMTV-like) is suspected to be involved in human breast cancer and it has been hypothesized that companion animals might have a role in viral transmission. The aim of our study was to investigate the presence of MMTV-like nucleotide sequences and viral protein in a larger number of feline (FMCs) and canine mammary carcinomas (CMCs) by nested PCR and immunohistochemistry. Results showed that the presence of MMTV-like env sequence in FMCs was 7% (6/86), while all the CMCs and canine dysplastic lesions scored negative. All PCR-positive FMCs scored positive for the MMTV p14 signal peptide of the envelope precursor protein of the virus. In contrast, all PCR-negative FMCs and canine mammary lesions were also negative for immunohistochemistry analysis. Canine and feline normal mammary gland tissues scored negative for both PCR and MMTV-p14 protein. Multiple nucleotide alignment of MMTV-like env gene sequences isolated from cat showed 97% and 99% similarity with HMTV and MMTV, respectively, while the others two presented some polimorphisms. Particularly the sequences of one of these two tumors showed a polymorphism (c.7575 A> G), that causes a previously unreported amino acid substitution (Thr > Ala). In conclusion, the results of our study showed the presence of MMTV-like sequences and viral protein in some FMCs. Further studies are needed to understand whether this virus does play a role in the development of FMCs, if MMTV-like is an exogenous virus as these data suggest and, in such a case, how and from whom this virus was acquired.

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          Most cited references 27

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          Mouse DNA contamination in human tissue tested for XMRV

          Background We used a PCR-based approach to study the prevalence of genetic sequences related to a gammaretrovirus, xenotropic murine leukemia virus-related virus, XMRV, in human prostate cancer. This virus has been identified in the US in prostate cancer patients and in those with chronic fatigue syndrome. However, with the exception of two patients in Germany, XMRV has not been identified in prostate cancer tissue in Europe. Most putative associations of new or old human retroviruses with diseases have turned out to be due to contamination. We have looked for XMRV sequences in DNA extracted from formalin-fixed paraffin- embedded prostate tissues. To control for contamination, PCR assays to detect either mouse mitochondrial DNA (mtDNA) or intracisternal A particle (IAP) long terminal repeat DNA were run on all samples, owing to their very high copy number in mouse cells. Results In general agreement with the US prevalence, XMRV-like sequences were found in 4.8% of prostate cancers. However, these were also positive, as were 21.5% of XMRV-negative cases, for IAP sequences, and many, but not all were positive for mtDNA sequences. Conclusions These results show that contamination with mouse DNA is widespread and detectable by the highly sensitive IAP assay, but not always with less sensitive assays, such as murine mtDNA PCR. This study highlights the ubiquitous presence of mouse DNA in laboratory specimens and offers a means of rigorous validation for future studies of murine retroviruses in human disease.
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            Detection of mammary tumor virus env gene-like sequences in human breast cancer.

            Mouse mammary tumor virus (MMTV) has been related to human breast cancer (BC) in previous studies. Although suggestive sequence homology to MMTV has been described in BC DNA, the presence of human endogenous retroviruses (HERs) confounded these results. We have selected a 660-bp sequence of the MMTV env gene with very low homology to HER or to any other human or viral gene. We have searched for sequences homologous to it using the polymerase chain reaction. DNA was extracted from fresh or frozen tissues using primers and probes constructed to detect 660 bp; for paraffin-embedded tissues, we sought 250-bp sequences by similar methodology. The 660-bp sequence was detected in 121 (38.5%) of the 314 unselected BC samples, in cultured BC cells, in 2 (6.9%) of 29 breast fibroadenomas and in 2 (1.8%) of 107 breast specimens from reduction mammoplastias. The sequence was not found in normal tissues including breast, lymphocytes from BC patients, nor in other human cancers or cell lines. The 250-bp sequence was detected in 60 (39.7%) of the 151 BCs, and in 1 of 27 normal breast samples assayed from paraffin-embedded sections. Cloning and sequencing of the 660 bp and 250 bp demonstrated that they are 95-99% homologous to MMTV env gene, but not to the known HERs nor to other viral or human genes (< 18%). Southern blot analysis using labeled cloned sequences showed that the 660-bp sequences were present in low copy number as a 7-8-kb EcoRI fragment only in breast cancer samples and two breast cancer cell lines that were positive by PCR. These data indicate that 38-40% of human breast cancers contain gene sequences homologous to the MMTV env gene that are absent from other tumors and tissues. These MMTV env gene-like sequences may play a role in the etiology of a large proportion of human breast cancer.
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              SOME POSSIBLE EFFECTS OF NURSING ON THE MAMMARY GLAND TUMOR INCIDENCE IN MICE.

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                Author and article information

                Contributors
                Role: ConceptualizationRole: InvestigationRole: MethodologyRole: Writing – review & editing
                Role: Data curationRole: Investigation
                Role: Investigation
                Role: Investigation
                Role: Data curationRole: Investigation
                Role: Investigation
                Role: InvestigationRole: Writing – original draft
                Role: Investigation
                Role: ConceptualizationRole: SupervisionRole: Writing – review & editing
                Role: ConceptualizationRole: SupervisionRole: Writing – review & editing
                Role: ConceptualizationRole: Funding acquisitionRole: SupervisionRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                24 July 2018
                2018
                : 13
                : 7
                Affiliations
                [1 ] Fondazione Pisana per la Scienza, Pisa, Italia
                [2 ] Dipartimento di Scienze Veterinarie, Università di Pisa, Pisa, Italia
                [3 ] Dipartimento di Ricerca Traslazionale e delle Nuove Tecnologie in Medicina e Chirurgia, Università di Pisa, Pisa, Italia
                Baylor College of Medicine, UNITED STATES
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                ‡ These authors also contributed equally to this work.

                Article
                PONE-D-18-08397
                10.1371/journal.pone.0200839
                6057629
                30040851
                © 2018 Civita et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                Page count
                Figures: 4, Tables: 2, Pages: 12
                Product
                Funding
                Funded by: funder-id http://dx.doi.org/10.13039/501100007514, Università di Pisa;
                Award Recipient :
                This work was supported by Università di Pisa grant no. 599999_2017_Neoplasie_Mammarie to AP. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Medicine and Health Sciences
                Oncology
                Cancers and Neoplasms
                Carcinomas
                Research and Analysis Methods
                Database and Informatics Methods
                Bioinformatics
                Sequence Analysis
                Sequence Alignment
                Biology and Life Sciences
                Molecular Biology
                Molecular Biology Techniques
                Artificial Gene Amplification and Extension
                Polymerase Chain Reaction
                Research and Analysis Methods
                Molecular Biology Techniques
                Artificial Gene Amplification and Extension
                Polymerase Chain Reaction
                Research and Analysis Methods
                Computational Techniques
                Split-Decomposition Method
                Multiple Alignment Calculation
                Medicine and Health Sciences
                Oncology
                Cancers and Neoplasms
                Breast Tumors
                Breast Cancer
                Biology and Life Sciences
                Evolutionary Biology
                Evolutionary Systematics
                Phylogenetics
                Phylogenetic Analysis
                Biology and Life Sciences
                Taxonomy
                Evolutionary Systematics
                Phylogenetics
                Phylogenetic Analysis
                Computer and Information Sciences
                Data Management
                Taxonomy
                Evolutionary Systematics
                Phylogenetics
                Phylogenetic Analysis
                Research and analysis methods
                Database and informatics methods
                Bioinformatics
                Sequence analysis
                DNA sequence analysis
                Research and Analysis Methods
                Database and Informatics Methods
                Bioinformatics
                Sequence Analysis
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