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DeltaRR vaccination protects from KA-induced seizures and neuronal loss through ICP10PK-mediated modulation of the neuronal-microglial axis.

Laing JM, Aurelian L - Genet Vaccines Ther (2008)

Bottom Line: The CM from Delta PK-infected EOC2 and EOC20 cells did not contain IL-10, but it contained TNF-alpha and RANTES.IL-10 neutralization significantly (p < 0.01) decreased, but did not abrogate, the neuroprotective activity of the CM from Delta RR-infected microglial cultures indicating that ICP10PK modulates the neuronal-microglial axis, also through induction of various microglial neuroprotective factors.Protection was associated with a significant (p < 0.001) increase in the numbers of IL-10+ microglia (CD11b+) as compared to Delta PK-treated animals.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pharmacology and Experimental Therapeutics, University of Maryland, School of Medicine, Baltimore, MD 21201, USA. jlain001@umaryland.edu

ABSTRACT
Ischemic brain injury and epilepsy are common neurodegenerative diseases caused by excitotoxicity. Their pathogenesis includes microglial production of inflammatory cytokines. Our studies were designed to examine whether a growth compromised HSV-2 mutant (Delta RR) prevents excitotoxic injury through modulation of microglial responses by the anti-apoptotic HSV-2 protein ICP10PK. EOC2 and EOC20 microglial cells, which are differentially activated, were infected with Delta RR or the ICP10PK deleted virus (Delta PK) and examined for virus-induced neuroprotective activity. Both cell lines were non-permissive for virus growth, but expressed ICP10PK (Delta RR) or the PK deleted ICP10 protein p95 (Delta PK). Conditioned medium (CM) from Delta RR-, but not Delta PK-infected cells prevented N-methyl-D-aspartate (NMDA)-induced apoptosis of primary hippocampal cultures, as determined by TUNEL and caspase-3 activation (76.9 +/- 5.3% neuroprotection). Neuroprotection was associated with inhibition of TNF-alpha and RANTES and production of IL-10. The CM from Delta PK-infected EOC2 and EOC20 cells did not contain IL-10, but it contained TNF-alpha and RANTES. IL-10 neutralization significantly (p < 0.01) decreased, but did not abrogate, the neuroprotective activity of the CM from Delta RR-infected microglial cultures indicating that ICP10PK modulates the neuronal-microglial axis, also through induction of various microglial neuroprotective factors. Rats given Delta RR (but not Delta PK) by intranasal inoculation were protected from kainic acid (KA)-induced seizures and neuronal loss in the CA1 hippocampal fields. Protection was associated with a significant (p < 0.001) increase in the numbers of IL-10+ microglia (CD11b+) as compared to Delta PK-treated animals. Delta RR is a promising vaccination/therapy platform for neurodegeneration through its pro-survival functions in neurons as well as microglia modulation.

No MeSH data available.


Related in: MedlinePlus

ICP10PK induces IL-10 expression in ΔRR-infected microglia. EOC2 and EOC20 cells were mock-infected with PBS, or infected with ΔRR, ΔPK (moi = 5) or mock-infected with PBS and culture supernatants collected 1–72 hrs p.i. were assayed for IL-10 by ELISA, as described in Materials and Methods. Results are the mean of three independent experiments ± SD. (***p < 0.001 relative to ΔRR-infected).
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Figure 7: ICP10PK induces IL-10 expression in ΔRR-infected microglia. EOC2 and EOC20 cells were mock-infected with PBS, or infected with ΔRR, ΔPK (moi = 5) or mock-infected with PBS and culture supernatants collected 1–72 hrs p.i. were assayed for IL-10 by ELISA, as described in Materials and Methods. Results are the mean of three independent experiments ± SD. (***p < 0.001 relative to ΔRR-infected).

Mentions: To examine whether ΔRR infection induces the production of neuroprotective factors and verify the effect of the cell activation state on their production, EOC2 and EOC20 cells were infected with ΔRR or ΔPK (moi = 5) or mock-infected with PBS and the CM were assayed for IL-10 production by ELISA. We focused on IL-10, because: (i) it is a pleiotropic cytokine with neuroprotective activity [31], (ii) IL-10 inhibits the transcription and translation of TNF-α and RANTES in macrophages [33], and (iii) ICP10PK upregulates IL-10 production in T cells [34]. ΔRR induced IL-10 production in both EOC2 and EOC20 cells. The kinetics of IL-10 production appeared to be somewhat different for the two cell lines, but the maximal levels at 72 hrs p.i. were similar (Fig. 7). In EOC2 cells, IL-10 was first seen at 4 hrs p.i and production increased with time, reaching maximal levels at 48–72 hrs p.i. In EOC20 cells, IL-10 was also first seen at 4 hrs p.i., but production seemed to reflect a two-phase kinetics, reaching a plateau at 24–48 hrs p.i. and increasing again, with maximal levels apparently not yet reached at 72 hrs pi. IL-10 was not seen in CM from ΔPK infected EOC2 or EOC20 cells (Fig. 7), indicating that its production is induced by ICP10PK. This is consistent with previous reports that IL-10 is not produced in microglia infected with HSV-1 [35], which does not conserve a functional ICP10PK [17,36].


DeltaRR vaccination protects from KA-induced seizures and neuronal loss through ICP10PK-mediated modulation of the neuronal-microglial axis.

Laing JM, Aurelian L - Genet Vaccines Ther (2008)

ICP10PK induces IL-10 expression in ΔRR-infected microglia. EOC2 and EOC20 cells were mock-infected with PBS, or infected with ΔRR, ΔPK (moi = 5) or mock-infected with PBS and culture supernatants collected 1–72 hrs p.i. were assayed for IL-10 by ELISA, as described in Materials and Methods. Results are the mean of three independent experiments ± SD. (***p < 0.001 relative to ΔRR-infected).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2268933&req=5

Figure 7: ICP10PK induces IL-10 expression in ΔRR-infected microglia. EOC2 and EOC20 cells were mock-infected with PBS, or infected with ΔRR, ΔPK (moi = 5) or mock-infected with PBS and culture supernatants collected 1–72 hrs p.i. were assayed for IL-10 by ELISA, as described in Materials and Methods. Results are the mean of three independent experiments ± SD. (***p < 0.001 relative to ΔRR-infected).
Mentions: To examine whether ΔRR infection induces the production of neuroprotective factors and verify the effect of the cell activation state on their production, EOC2 and EOC20 cells were infected with ΔRR or ΔPK (moi = 5) or mock-infected with PBS and the CM were assayed for IL-10 production by ELISA. We focused on IL-10, because: (i) it is a pleiotropic cytokine with neuroprotective activity [31], (ii) IL-10 inhibits the transcription and translation of TNF-α and RANTES in macrophages [33], and (iii) ICP10PK upregulates IL-10 production in T cells [34]. ΔRR induced IL-10 production in both EOC2 and EOC20 cells. The kinetics of IL-10 production appeared to be somewhat different for the two cell lines, but the maximal levels at 72 hrs p.i. were similar (Fig. 7). In EOC2 cells, IL-10 was first seen at 4 hrs p.i and production increased with time, reaching maximal levels at 48–72 hrs p.i. In EOC20 cells, IL-10 was also first seen at 4 hrs p.i., but production seemed to reflect a two-phase kinetics, reaching a plateau at 24–48 hrs p.i. and increasing again, with maximal levels apparently not yet reached at 72 hrs pi. IL-10 was not seen in CM from ΔPK infected EOC2 or EOC20 cells (Fig. 7), indicating that its production is induced by ICP10PK. This is consistent with previous reports that IL-10 is not produced in microglia infected with HSV-1 [35], which does not conserve a functional ICP10PK [17,36].

Bottom Line: The CM from Delta PK-infected EOC2 and EOC20 cells did not contain IL-10, but it contained TNF-alpha and RANTES.IL-10 neutralization significantly (p < 0.01) decreased, but did not abrogate, the neuroprotective activity of the CM from Delta RR-infected microglial cultures indicating that ICP10PK modulates the neuronal-microglial axis, also through induction of various microglial neuroprotective factors.Protection was associated with a significant (p < 0.001) increase in the numbers of IL-10+ microglia (CD11b+) as compared to Delta PK-treated animals.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pharmacology and Experimental Therapeutics, University of Maryland, School of Medicine, Baltimore, MD 21201, USA. jlain001@umaryland.edu

ABSTRACT
Ischemic brain injury and epilepsy are common neurodegenerative diseases caused by excitotoxicity. Their pathogenesis includes microglial production of inflammatory cytokines. Our studies were designed to examine whether a growth compromised HSV-2 mutant (Delta RR) prevents excitotoxic injury through modulation of microglial responses by the anti-apoptotic HSV-2 protein ICP10PK. EOC2 and EOC20 microglial cells, which are differentially activated, were infected with Delta RR or the ICP10PK deleted virus (Delta PK) and examined for virus-induced neuroprotective activity. Both cell lines were non-permissive for virus growth, but expressed ICP10PK (Delta RR) or the PK deleted ICP10 protein p95 (Delta PK). Conditioned medium (CM) from Delta RR-, but not Delta PK-infected cells prevented N-methyl-D-aspartate (NMDA)-induced apoptosis of primary hippocampal cultures, as determined by TUNEL and caspase-3 activation (76.9 +/- 5.3% neuroprotection). Neuroprotection was associated with inhibition of TNF-alpha and RANTES and production of IL-10. The CM from Delta PK-infected EOC2 and EOC20 cells did not contain IL-10, but it contained TNF-alpha and RANTES. IL-10 neutralization significantly (p < 0.01) decreased, but did not abrogate, the neuroprotective activity of the CM from Delta RR-infected microglial cultures indicating that ICP10PK modulates the neuronal-microglial axis, also through induction of various microglial neuroprotective factors. Rats given Delta RR (but not Delta PK) by intranasal inoculation were protected from kainic acid (KA)-induced seizures and neuronal loss in the CA1 hippocampal fields. Protection was associated with a significant (p < 0.001) increase in the numbers of IL-10+ microglia (CD11b+) as compared to Delta PK-treated animals. Delta RR is a promising vaccination/therapy platform for neurodegeneration through its pro-survival functions in neurons as well as microglia modulation.

No MeSH data available.


Related in: MedlinePlus