scholarly journals Prion infection, transmission, and cytopathology modeled in a low-biohazard human cell line

2020 ◽  
Vol 3 (8) ◽  
pp. e202000814
Author(s):  
Merve Avar ◽  
Daniel Heinzer ◽  
Nicolas Steinke ◽  
Berre Doğançay ◽  
Rita Moos ◽  
...  
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Human Cell ◽  
Prion Diseases ◽  
Human Cell Line ◽  
Proteinase K ◽  
Murine Cell ◽  
Prion Infection ◽  
Infected Cells ◽  
Bona Fide

Transmission of prion infectivity to susceptible murine cell lines has simplified prion titration assays and has greatly reduced the need for animal experimentation. However, murine cell models suffer from technical and biological constraints. Human cell lines might be more useful, but they are much more biohazardous and are often poorly infectible. Here, we describe the human clonal cell line hovS, which lacks the human PRNP gene and expresses instead the ovine PRNP VRQ allele. HovS cells were highly susceptible to the PG127 strain of sheep-derived murine prions, reaching up to 90% infected cells in any given culture and were maintained in a continuous infected state for at least 14 passages. Infected hovS cells produced proteinase K–resistant prion protein (PrPSc), pelletable PrP aggregates, and bona fide infectious prions capable of infecting further generations of naïve hovS cells and mice expressing the VRQ allelic variant of ovine PrPC. Infection in hovS led to prominent cytopathic vacuolation akin to the spongiform changes observed in individuals suffering from prion diseases. In addition to expanding the toolbox for prion research to human experimental genetics, the hovS cell line provides a human-derived system that does not require human prions. Hence, the manipulation of scrapie-infected hovS cells may present fewer biosafety hazards than that of genuine human prions.

scholarly journals Prion infection, transmission and cytopathology modelled in a low-biohazard human cell line

2020 ◽  
Author(s):  
Merve Avar ◽  
Daniel Heinzer ◽  
Nicolas Steinke ◽  
Berre Doğançay ◽  
Rita Moos ◽  
...  
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Human Cell ◽  
Prion Diseases ◽  
Human Cell Line ◽  
Proteinase K ◽  
Murine Cell ◽  
Prion Infection ◽  
Infected Cells ◽  
Bona Fide

AbstractTransmission of prion infectivity to susceptible murine cell lines has simplified prion titration assays and has greatly reduced the need for animal experimentation. However, murine cell models suffer from technical and biological constraints. Human cell lines might be more useful, but they are much more biohazardous and are often poorly infectible. Here we describe the human clonal cell line hovS, which lacks the human PRNP gene and expresses instead the ovine PRNP VRQ allele. HovS cells were highly susceptible to the PG127 strain of sheep-derived murine prions, reaching up to 90% infected cells in any given culture, and were maintained in a continuous infected state for at least 14 passages. Infected hovS cells produced proteinase K-resistant prion protein (PrPSc), pelletable PrP aggregates and bona fide infectious prions capable of infecting further generations of naïve hovS cells and mice expressing the VRQ allelic variant of ovine PrPC. Infection in hovS led to prominent cytopathic vacuolation akin to the spongiform changes observed in individuals suffering from prion diseases. In addition to expanding the toolbox for prion research to human experimental genetics, the hovS cell line provides a human-derived system that does not require human prions. Hence, the manipulation of scrapie-infected hovS cells may present fewer biosafety hazards than that of genuine human prions.

scholarly journals Extended Analysis of the In Vitro Tropism of Porcine Endogenous Retrovirus

2000 ◽  
Vol 74 (1) ◽  
pp. 49-56 ◽  
Author(s):  
Carolyn A. Wilson ◽  
Susan Wong ◽  
Matthew VanBrocklin ◽  
Mark J. Federspiel
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Human Cell ◽  
Human Cell Line ◽  
Endogenous Retrovirus ◽  
Human Cells ◽  
Productive Infection ◽  
Porcine Endogenous Retrovirus ◽  
Productive Replication

ABSTRACT We previously reported that mitogenic activation of porcine peripheral blood mononuclear cells resulted in production of porcine endogenous retrovirus(es) (PERV[s]) capable of productively infecting human cells (C. Wilson et al., J. Virol. 72:3082–3087, 1998). We now extend that analysis to show that additional passage of isolated virus, named here PERV-NIH, through a human cell line yielded a viral population with a higher titer of infectious virus on human cells than the initial isolate. We show that in a single additional passage on a human cell line, the increase in infectivity for human cells is accounted for by selection against variants carrying pig-tropic envelope sequences (PERV-C) as well as by enrichment for replication-competent genomes. Sequence analysis of the envelope cDNA present in virions demonstrated that the envelope sequence of PERV-NIH is related to but distinct from previously reported PERV envelopes. The in vitro host range of PERV was studied in human primary cells and cell lines, as well as in cell lines from nonhuman primate and other species. This analysis reveals three patterns of susceptibility to infection among these host cells: (i) cells are resistant to infection in our assay; (ii) cells are infected by virus, as viral RNA is detected in the supernatant by reverse transcription-PCR, but the cells are not permissive to productive replication and spread; and (iii) cells are permissive to low-level productive replication. Certain cell lines were permissive for efficient productive infection and spread. These results may prove useful in designing appropriate animal models to assess the in vivo infectivity properties of PERV.

scholarly journals Epigenetic Modifier Mutations in the LL-100 Panel

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 5271-5271
Author(s):  
Hilmar Quentmeier ◽  
Claudia Pommerenke ◽  
Hans G. Drexler
Keyword(s):  
Amino Acid ◽  
Drug Development ◽  
Cell Line ◽  
Cell Lines ◽  
Human Cell ◽  
Conflicts Of Interest ◽  
Human Cancer ◽  
Human Cell Line ◽  
Dominant Negative ◽  
Epigenetic Modifier

Abstract The NCI-60 human cell line panel, developed for use in drug development comprises sixty human cancer cell lines derived from nine different tissues. Only six cell lines of the NCI-60 were derived from blood cancers. Therefore, most forms and subtypes of leukemia and lymphoma are not represented in the NCI-60 panel. To respond to this apparent gap, we suggest the novel LL-100 panel, 100 leukemia and lymphoma cell lines representing the major leukemia/lymphoma entities, for basic research and drug development. Whole exome sequencing and RNA sequencing were performed to identify mutations in 100 cell lines. Here we list the 100 cell lines, ordered by subtype and show mutations in epigenetic modifier genes. We found cell lines with mutations in ASXL1, EZH2, IDH1, TET2 and in DNMT3A. Hitherto, cell line OCI-AML3 was the only human cell line described with a DNMT3A mutation. Twenty-two percent of patients with acute myeloid leukemia contain DNMT3A mutations and the median overall survival with DNMT3A mutations is shorter than without. Most DNMT3A mutations are heterozygous and alter amino acid R882, R882H being the most common DNMT3A mutation in AML. Exogenously mutant murine R878H (equivalent to human R882H) inhibits DNMT3A activity in a dominant negative manner. We describe here that the AML cell line SET-2 carries a heterozygous G to A transition at chr2_25234373 (hg38) which leads to the DNMT3A R882H amino acid substitution. Chip-based methylation analysis revealed that the described DNMT3A targets IRF8, KLF2, HOXA11 and HOXB2 are hypomethylated in cell lines OCI-AML3 (DNMT3A R882C) and in SET-2 (DNMT3A R882H). These data suggest that SET-2 is a novel model cell line for functional analysis of the DNMT3A R882 mutation and a first gain in knowledge through data mining the LL-100 panel. Disclosures No relevant conflicts of interest to declare.

scholarly journals Application of PMCA to screen for prion infection in a human cell line used to produce biological therapeutics

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Adam Lyon ◽  
Charles E. Mays ◽  
Frank Borriello ◽  
Glenn C. Telling ◽  
Claudio Soto ◽  
...  
Keyword(s):  
Cell Line ◽  
Human Cell ◽  
Human Cell Line ◽  
Prion Infection ◽  
Biological Therapeutics

scholarly journals CHEMICAL AND PHARMACOLOGICAL EVALUATION OF MANGLICOLOUS LICHEN GRAPHIS AJAREKARII PATW. & C. R. KULK.

2019 ◽  
Vol 57 (3) ◽  
pp. 300
Author(s):  
Polimati Haritha ◽  
Sunil Kumar Patnaik ◽  
Vinay Bharadwaj Tatipamula
Keyword(s):  
Free Radicals ◽  
Cell Line ◽  
Cell Lines ◽  
Human Cell ◽  
Ethanolic Extract ◽  
Human Cell Line ◽  
Chemical Examination ◽  
Cytotoxicity Studies ◽  
Normal Human Cell ◽  
Normal Human

The chemical examination of ethanolic extract of manglicolous lichen Graphis ajarekarii (Ga-Et) resulted in isolation of three known metabolites – chiodectonic acid (1), graphenone (2) and graphisquinone (3). All the isolates (1-3) and Ga-Et were screened against DPPH and superoxide free radicals, six different cancer cell lines and one normal human cell line (NHME). This work is the first report of antioxidant and cytotoxicity studies on the isolated metabolites (1-3).

scholarly journals Characterization of a new case of XMLV (Bxv1) contamination in the human cell line Hep2 (clone 2B)

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Vincent Loiseau ◽  
Richard Cordaux ◽  
Isabelle Giraud ◽  
Agnès Beby-Defaux ◽  
Nicolas Lévêque ◽  
...  
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Human Cell ◽  
Leukemia Virus ◽  
Human Cell Line ◽  
Chromosome 8 ◽  
Catalog Number ◽  
E6 And E7 ◽  
Human Papillomavirus 18

Abstract The use of misidentified cell lines contaminated by other cell lines and/or microorganisms has generated much confusion in the scientific literature. Detailed characterization of such contaminations is therefore crucial to avoid misinterpretation and ensure robustness and reproducibility of research. Here we use DNA-seq data produced in our lab to first confirm that the Hep2 (clone 2B) cell line (Sigma-Aldrich catalog number: 85011412-1VL) is indistinguishable from the HeLa cell line by mapping integrations of the human papillomavirus 18 (HPV18) at their expected loci on chromosome 8. We then show that the cell line is also contaminated by a xenotropic murine leukemia virus (XMLV) that is nearly identical to the mouse Bxv1 provirus and we characterize one Bxv1 provirus, located in the second intron of the pseudouridylate synthase 1 (PUS1) gene. Using an RNA-seq dataset, we confirm the high expression of the E6 and E7 HPV18 oncogenes, show that the entire Bxv1 genome is moderately expressed, and retrieve a Bxv1 splicing event favouring expression of the env gene. Hep2 (clone 2B) is the fourth human cell line so far known to be contaminated by the Bxv1 XMLV. This contamination has to be taken into account when using the cell line in future experiments.

MEIC Evaluation of Acute Systemic Toxicity

1998 ◽  
Vol 26 (1_suppl) ◽  
pp. 131-183 ◽  
Author(s):  
Cecilia Clemedson ◽  
Marianne Andersson ◽  
Yasunobu Aoki ◽  
Frank A. Barile ◽  
Anna Maria Bassi ◽  
...  
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Human Cell ◽  
Protein Denaturation ◽  
Human Cell Line ◽  
Background Information ◽  
In Vitro Toxicity ◽  
Toxicity Data ◽  
Relative Significance

Results from tests on the Multicentre Evaluation of In Vitro Cytotoxicity (MEIC) reference chemicals 31–50 in 67 different in vitro toxicity assays are presented in this paper as a prerequisite to in vitro/in vivo comparisons for all MEIC in vitro toxicity data in forthcoming papers, i.e. the final MEIC evaluation of the relevance of the tests. With the aim of increasing knowledge about the relative significance of some in vitro methodological factors, the strategies and methods of the preceding parts in the MEIC series (Parts II and III) were again employed to enable comparative cytotoxicity analysis of the new in vitro results presented in this paper. A principal components analysis (PCA) of the results from tests of the 20 chemicals in 67 assays demonstrated a dominating first component describing as much as 74% of the variance in the toxicity data, indicating a similar ranking of the cytotoxicities of the chemicals in most of the tests. The influence on the general variability of the results of a few, key methodological factors was also evaluated by using linear regression comparisons of the results of all pairs of methods available in the study, i.e. methods which were similar in all respects except for the factor being analysed. Results from this “random probe” analysis were: a) the cytotoxicities of 11 of the 20 chemicals increased considerably with exposure time (> 10 times over 4–168 hours); b) in general, human cell line toxicity was well predicted by cytotoxicity in animal cells; c) prediction of human cell line toxicity by most ecotoxicological tests was only fairly good; d) 14 comparisons of similar assays with different cell lines showed similar toxicities (mean R2 = 0.83); e) nine comparisons of similar assays employing different primary cultures and cell lines shared similar toxicities (mean R2 = 0.71); and f) 16 comparisons of similar assays with different growth/viability endpoints showed similar toxicities (mean R2 = 0.71). Results b, d, e and f must contribute to the PCA-documented high general similarity of the in vitro toxicity data. Results a and c, together with factors which were not analysed, such as different protocols and inter-laboratory variability of tests, could explain the 26% dissimilarity. To provide background information to the planned final MEIC evaluation of the relevance of the 61 methods in which all 50 chemicals have been tested, an additional PCA was made of the 50 chemical-61 assay in vitro database (from Parts II and III and the present paper). This supplementary PCA demonstrated an 80% similarity of results. Compared with the previous analysis of the tests of the first 30 MEIC reference chemicals (MEIC Part III), the present analysis of the tests of the last 20 MEIC chemicals indicates a somewhat higher variation in the results. Correspondingly, some deviating endpoint measurements and cell line responses were demonstrated by the pairwise comparisons in the present study. As a result, the analysis revealed a high correlation (R2 = 0.73) between the average human cell line toxicity and the results from a new protein denaturation test. These preliminary results suggest that intracellular protein denaturation may be a frequently occurring mechanism in basal cytotoxicity.

Sign in / Sign up

Export Citation Format

Share Document