| 1. |
- Fossum, Caroline, et al.
(författare)
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PCV2 on the spot-A new method for the detection of single porcine circovirus type 2 secreting cells
- 2014
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Ingår i: Journal of Virological Methods. - : Elsevier BV. - 0166-0934 .- 1879-0984. ; 196, s. 185-192
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Tidskriftsartikel (refereegranskat)abstract
- A porcine circovirus type 2 SPOT (PCV2-SPOT) assay was established to enumerate virus-secreting lymphocytes obtained from naturally infected pigs. The assay is based on the same principle as general ELISPOT assays but instead of detecting cytokine or immunoglobulin secretion, PCV2 particles are immobilized and detected as filter spots. The method was used to evaluate the influence of various cell activators on the PCV2 secretion in vitro and was also applied to study the PCV2 secretion by lymphocytes obtained from pigs in healthy herds and in a herd afflicted by postweaning multisystemic wasting disease (PMWS). Peripheral blood mononuclear cells (PBMCs) obtained from a pig with severe PMWS produced PCV2-SPOT5 spontaneously whereas PBMCs obtained from pigs infected subclinically only generated PCV2-SPOT5 upon in vitro stimulation. The PCV2 secretion potential was related to the PCV2 DNA content in the PBMCs as determined by two PCV2 real-time PCR assays, developed to differentiate between Swedish PCV2 genogroups 1 (PCV2a) and 3 (PCV2b). Besides the current application these qPCRs could simplify future epidemiological studies and allow genogroup detection/quantitation in dual infection experiments and similar studies. The developed PCV2-SPOT assay offers a semi-quantitative approach to evaluate the potential of PCV2-infected porcine cells to release PCV2 viral particles as well as a system to evaluate the ability of different cell types or compounds to affect PCV2 replication and secretion. (C) 2013 Elsevier B.V. All rights reserved.
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| 2. |
- Hu, Kefei, et al.
(författare)
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Nanoparticulate Quillaja saponin induces apoptosis in human leukemia cell lines with a high therapeutic index
- 2010
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Ingår i: International Journal of Nanomedicine. - 1176-9114 .- 1178-2013. ; 5:1, s. 51-62
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Tidskriftsartikel (refereegranskat)abstract
- Saponin fractions of Quillaja saponaria Molina (QS) have cytotoxic activity against cancer cells in vitro, but are too toxic to be useful in the clinic. The toxic effect was abolished by converting QS fractions into stable nanoparticles through the binding of QS to cholesterol. Two fractions of QS were selected for particle formation, one with an acyl-chain (ASAP) was used to form killing and growth-inhibiting (KGI) particles, and the other without the acyl-chain (DSAP) was used to formulate blocking and balancing effect (BBE) particles. KGI showed significant growth inhibiting and cancer cell-killing activities in nine of 10 cell lines while BBE showed that on one cell line. The monoblastoid lymphoma cell line U937 was selected for analyzing the mode of action. Low concentrations of KGI (0.5 and 2 microg/mL) induced irreversible exit from the cell cycle, differentiation measured by cytokine production, and eventually programmed cell death (apoptosis). Compared to normal human monocytes, the U937 cells were 30-fold more sensitive to KGI. The nontoxic BBE blocked the cell killing effect of KGI in a concentration-dependent manner. In conclusion, the formulation of QS into nanoparticles has the potential of becoming a new class of anticancer agents.
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| 3. |
- Timmusk, Sirje, et al.
(författare)
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Regulator of G protein signalling 16 is target for a porcine circorvirus type 2 protein
- 2009
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Ingår i: Journal of General Virology. - : Microbiology Society. - 0022-1317 .- 1465-2099. ; 90, s. 2425-2436
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Tidskriftsartikel (refereegranskat)abstract
- Interaction studies have suggested that the non-structural protein encoded by open reading frame 3 (ORF3) of porcine circovirus type 2 (PCV2) binds specifically to a regulator of G protein signalling (RGS) related to human RGS16 (huRGS16). The full-length clone of RGS16 was generated from porcine cells and sequence analysis revealed a close relationship to huRGS16 and murine RGS16. In vitro pull-down experiments verified an interaction between porcine RGS16 (poRGS16) and ORF3 from PCV2. Using GST-linked ORF3 proteins from three different genogroups of PCV2 and from porcine circovirus type 1 (PCV1) in the pull-down experiments indicated that there were differences in their ability to bind poRGS16. Quantitative RT-PCR demonstrated that the expression of poRGS16 mRNA could be induced by a number of cell activators including mitogens (LPS and PHA), interferon inducers (ODN 2216 and poly I : C) and the neurotransmitter norepinephrine. Immunofluorescence labelling confirmed the induced expression of poRGS16 at the protein level and suggested that the PCV2 ORF3 protein co-localized with poRGS16 in LPS-activated porcine PBMC. Furthermore, poRGS16 appeared to participate in the translocation of the ORF3 protein into the cell nucleus, suggesting that the observed interaction may play an important role in the infection biology of porcine circovirus.
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