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Toll-like receptor responses to Peste des petits ruminants virus in goats and water buffalo.

Dhanasekaran S, Biswas M, Vignesh AR, Ramya R, Raj GD, Tirumurugaan KG, Raja A, Kataria RS, Parida S, Elankumaran S, Subbiah E - PLoS ONE (2014)

Bottom Line: Upon stimulation of PBMC with synthetic TLR3 and TLR7 agonists or PPRV, the levels of proinflammatory cytokines were found to be significantly higher while immunosuppressive interleukin (IL) 10 levels were lower in PPRV resistant Kanni and Salem Black breeds and water buffalo at transcriptional level, correlating with reduced viralloads in infected PBMC.Pre-treatment of Vero cells with human IFNα resulted in reduction of PPRV replication, confirming the role of IFNα in limiting PPRV replication.Treatment with IRS66, a TLR7 antagonist, resulted in the reduction of IFNα levels, with increased PPRV replication confirming the role of TLR7.

View Article: PubMed Central - PubMed

Affiliation: Department of Animal Biotechnology, Madras Veterinary College, Tamil Nadu Veterinary and Animal Sciences University, Chennai, Tamil Nadu, India.

ABSTRACT
Ovine rinderpest or goat plague is an economically important and contagious viral disease of sheep and goats, caused by the Peste des petits ruminants virus (PPRV). Differences in susceptibility to goat plague among different breeds and water buffalo exist. The host innate immune system discriminates between pathogen associated molecular patterns and self antigens through surveillance receptors known as Toll like receptors (TLR). We investigated the role of TLR and cytokines in differential susceptibility of goat breeds and water buffalo to PPRV. We examined the replication of PPRV in peripheral blood mononuclear cells (PBMC) of Indian domestic goats and water buffalo and demonstrated that the levels of TLR3 and TLR7 and downstream signalling molecules correlation with susceptibility vs resistance. Naturally susceptible goat breeds, Barbari and Tellichery, had dampened innate immune responses to PPRV and increased viral loads with lower basal expression levels of TLR 3/7. Upon stimulation of PBMC with synthetic TLR3 and TLR7 agonists or PPRV, the levels of proinflammatory cytokines were found to be significantly higher while immunosuppressive interleukin (IL) 10 levels were lower in PPRV resistant Kanni and Salem Black breeds and water buffalo at transcriptional level, correlating with reduced viralloads in infected PBMC. Water buffalo produced higher levels of interferon (IFN) α in comparison with goats at transcriptional and translational levels. Pre-treatment of Vero cells with human IFNα resulted in reduction of PPRV replication, confirming the role of IFNα in limiting PPRV replication. Treatment with IRS66, a TLR7 antagonist, resulted in the reduction of IFNα levels, with increased PPRV replication confirming the role of TLR7. Single nucleotide polymorphism analysis of TLR7 of these goat breeds did not show any marked nucleotide differences that might account for susceptibility vs resistance to PPRV. Analyzing other host genetic factors might provide further insights on susceptibility to PPRV and genetic polymorphisms in the host.

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Induction of cytokine genes of goat PBMCs with TLR3 and TLR7 agonists and PPRV.A) Fold changes in mRNA expression levels of IL1β, IL6, IL8, IL10, IL12p40, TNFα, IFNγ and IFNα in goat PBMC stimulated with a) poly I:C, b) imiquimod or c) infected with PPRV d) Goat and buffalo PBMCs infected with PPRV. Fold change was determined by the 2−ΔΔCt formula. An upregulation in TNFα expression levels, consistent with a downregulation in IL10 levels, is observed in Kanni and Salem Black breeds of all treatment groups. In addition, upregulation of IFNα and IFNγ was observed in the PBMC of Kanni and Salem Black breeds after PPRV infection. PPRV stimulation resulted in an upregulation of IL1β and IFNα in buffalo PBMC and IL10, IL12p40 and IFNγ in goat PBMCs. Bars with the same superscript do not differ significantly. Significance is indicated when p<0.05. Values represent mean ± SD of 5 individual animals per goat breed.
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pone-0111609-g004: Induction of cytokine genes of goat PBMCs with TLR3 and TLR7 agonists and PPRV.A) Fold changes in mRNA expression levels of IL1β, IL6, IL8, IL10, IL12p40, TNFα, IFNγ and IFNα in goat PBMC stimulated with a) poly I:C, b) imiquimod or c) infected with PPRV d) Goat and buffalo PBMCs infected with PPRV. Fold change was determined by the 2−ΔΔCt formula. An upregulation in TNFα expression levels, consistent with a downregulation in IL10 levels, is observed in Kanni and Salem Black breeds of all treatment groups. In addition, upregulation of IFNα and IFNγ was observed in the PBMC of Kanni and Salem Black breeds after PPRV infection. PPRV stimulation resulted in an upregulation of IL1β and IFNα in buffalo PBMC and IL10, IL12p40 and IFNγ in goat PBMCs. Bars with the same superscript do not differ significantly. Significance is indicated when p<0.05. Values represent mean ± SD of 5 individual animals per goat breed.

Mentions: To understand the mechanistic basis for this differential permissiveness for virus replication in different goat breeds and goat vs water buffalo, we analyzed the downstream effector molecules of TLR3 and TLR7 engagement. PBMC stimulated with poly I:C, or imiquimod and further infected with PPRV were analyzed for the expression of IL1β, IL6, IL8, IL10, IL12p40, TNFα, IFNγ and IFNα mRNA (Figure 4A–D). Though upregulation of both pro- and anti-inflammatory cytokines were observed in all goat breeds, levels of TNFα was seen to be consistently higher in Kanni and Salem black breeds in all treatment groups (poly I:C, imiquimod and PPRV). This effect was prominent in the case of PPRV infected PBMC, where levels of TNFα and more importantly, IFNα and IFNγ, were significantly higher in Kanni and Salem black breeds as compared to Barbari (Figure 4C). Significant differences were not observed between these breeds and Tellicherry in the levels of pro-inflammatory cytokine mRNA. Consistent with this, levels of the immunosuppressive cytokine IL10 were significantly lower in these breeds than in Barbari (Figure 3c). Lower mRNA expression levels of IL10 in Kanni and Salem black breeds were also observed on stimulating TLR3 and TLR7 with poly I:C and imiquimod (Figure 4A, B).


Toll-like receptor responses to Peste des petits ruminants virus in goats and water buffalo.

Dhanasekaran S, Biswas M, Vignesh AR, Ramya R, Raj GD, Tirumurugaan KG, Raja A, Kataria RS, Parida S, Elankumaran S, Subbiah E - PLoS ONE (2014)

Induction of cytokine genes of goat PBMCs with TLR3 and TLR7 agonists and PPRV.A) Fold changes in mRNA expression levels of IL1β, IL6, IL8, IL10, IL12p40, TNFα, IFNγ and IFNα in goat PBMC stimulated with a) poly I:C, b) imiquimod or c) infected with PPRV d) Goat and buffalo PBMCs infected with PPRV. Fold change was determined by the 2−ΔΔCt formula. An upregulation in TNFα expression levels, consistent with a downregulation in IL10 levels, is observed in Kanni and Salem Black breeds of all treatment groups. In addition, upregulation of IFNα and IFNγ was observed in the PBMC of Kanni and Salem Black breeds after PPRV infection. PPRV stimulation resulted in an upregulation of IL1β and IFNα in buffalo PBMC and IL10, IL12p40 and IFNγ in goat PBMCs. Bars with the same superscript do not differ significantly. Significance is indicated when p<0.05. Values represent mean ± SD of 5 individual animals per goat breed.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4219731&req=5

pone-0111609-g004: Induction of cytokine genes of goat PBMCs with TLR3 and TLR7 agonists and PPRV.A) Fold changes in mRNA expression levels of IL1β, IL6, IL8, IL10, IL12p40, TNFα, IFNγ and IFNα in goat PBMC stimulated with a) poly I:C, b) imiquimod or c) infected with PPRV d) Goat and buffalo PBMCs infected with PPRV. Fold change was determined by the 2−ΔΔCt formula. An upregulation in TNFα expression levels, consistent with a downregulation in IL10 levels, is observed in Kanni and Salem Black breeds of all treatment groups. In addition, upregulation of IFNα and IFNγ was observed in the PBMC of Kanni and Salem Black breeds after PPRV infection. PPRV stimulation resulted in an upregulation of IL1β and IFNα in buffalo PBMC and IL10, IL12p40 and IFNγ in goat PBMCs. Bars with the same superscript do not differ significantly. Significance is indicated when p<0.05. Values represent mean ± SD of 5 individual animals per goat breed.
Mentions: To understand the mechanistic basis for this differential permissiveness for virus replication in different goat breeds and goat vs water buffalo, we analyzed the downstream effector molecules of TLR3 and TLR7 engagement. PBMC stimulated with poly I:C, or imiquimod and further infected with PPRV were analyzed for the expression of IL1β, IL6, IL8, IL10, IL12p40, TNFα, IFNγ and IFNα mRNA (Figure 4A–D). Though upregulation of both pro- and anti-inflammatory cytokines were observed in all goat breeds, levels of TNFα was seen to be consistently higher in Kanni and Salem black breeds in all treatment groups (poly I:C, imiquimod and PPRV). This effect was prominent in the case of PPRV infected PBMC, where levels of TNFα and more importantly, IFNα and IFNγ, were significantly higher in Kanni and Salem black breeds as compared to Barbari (Figure 4C). Significant differences were not observed between these breeds and Tellicherry in the levels of pro-inflammatory cytokine mRNA. Consistent with this, levels of the immunosuppressive cytokine IL10 were significantly lower in these breeds than in Barbari (Figure 3c). Lower mRNA expression levels of IL10 in Kanni and Salem black breeds were also observed on stimulating TLR3 and TLR7 with poly I:C and imiquimod (Figure 4A, B).

Bottom Line: Upon stimulation of PBMC with synthetic TLR3 and TLR7 agonists or PPRV, the levels of proinflammatory cytokines were found to be significantly higher while immunosuppressive interleukin (IL) 10 levels were lower in PPRV resistant Kanni and Salem Black breeds and water buffalo at transcriptional level, correlating with reduced viralloads in infected PBMC.Pre-treatment of Vero cells with human IFNα resulted in reduction of PPRV replication, confirming the role of IFNα in limiting PPRV replication.Treatment with IRS66, a TLR7 antagonist, resulted in the reduction of IFNα levels, with increased PPRV replication confirming the role of TLR7.

View Article: PubMed Central - PubMed

Affiliation: Department of Animal Biotechnology, Madras Veterinary College, Tamil Nadu Veterinary and Animal Sciences University, Chennai, Tamil Nadu, India.

ABSTRACT
Ovine rinderpest or goat plague is an economically important and contagious viral disease of sheep and goats, caused by the Peste des petits ruminants virus (PPRV). Differences in susceptibility to goat plague among different breeds and water buffalo exist. The host innate immune system discriminates between pathogen associated molecular patterns and self antigens through surveillance receptors known as Toll like receptors (TLR). We investigated the role of TLR and cytokines in differential susceptibility of goat breeds and water buffalo to PPRV. We examined the replication of PPRV in peripheral blood mononuclear cells (PBMC) of Indian domestic goats and water buffalo and demonstrated that the levels of TLR3 and TLR7 and downstream signalling molecules correlation with susceptibility vs resistance. Naturally susceptible goat breeds, Barbari and Tellichery, had dampened innate immune responses to PPRV and increased viral loads with lower basal expression levels of TLR 3/7. Upon stimulation of PBMC with synthetic TLR3 and TLR7 agonists or PPRV, the levels of proinflammatory cytokines were found to be significantly higher while immunosuppressive interleukin (IL) 10 levels were lower in PPRV resistant Kanni and Salem Black breeds and water buffalo at transcriptional level, correlating with reduced viralloads in infected PBMC. Water buffalo produced higher levels of interferon (IFN) α in comparison with goats at transcriptional and translational levels. Pre-treatment of Vero cells with human IFNα resulted in reduction of PPRV replication, confirming the role of IFNα in limiting PPRV replication. Treatment with IRS66, a TLR7 antagonist, resulted in the reduction of IFNα levels, with increased PPRV replication confirming the role of TLR7. Single nucleotide polymorphism analysis of TLR7 of these goat breeds did not show any marked nucleotide differences that might account for susceptibility vs resistance to PPRV. Analyzing other host genetic factors might provide further insights on susceptibility to PPRV and genetic polymorphisms in the host.

Show MeSH
Related in: MedlinePlus