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Interferon-alpha/beta receptor-mediated selective induction of a gene cluster by CpG oligodeoxynucleotide 2006.

Kato A, Homma T, Batchelor J, Hashimoto N, Imai S, Wakiguchi H, Saito H, Matsumoto K - BMC Immunol. (2003)

Bottom Line: Oligodeoxynucleotides containing unmethylated CpG motifs (CpG ODN) are known to exert a strong adjuvant effect on Th1 immune responses.Blocking with mAb against IFN-alpha/beta receptor strongly inhibited the induction of these IFN-inducible genes by CpG ODN.This study provides new information regarding the possible immunomodulatory effects of CpG ODN in vivo via an IFN-alpha/beta receptor-mediated paracrine pathway.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Allergy & Immunology, National Research Institute for Child Health & Development, Tokyo, Japan. atkato@nch.go.jp

ABSTRACT

Background: Oligodeoxynucleotides containing unmethylated CpG motifs (CpG ODN) are known to exert a strong adjuvant effect on Th1 immune responses. Although several genes have been reported, no comprehensive study of the gene expression profiles in human cells after stimulation with CpG ODN has been reported.

Results: This study was designed to identify a CpG-inducible gene cluster that potentially predicts for the molecular mechanisms of clinical efficacy of CpG ODN, by determining mRNA expression in human PBMC after stimulation with CpG ODN. PBMCs were obtained from the peripheral blood of healthy volunteers and cultured in the presence or absence of CpG ODN 2006 for up to 24 hours. The mRNA expression profile was evaluated using a high-density oligonucleotide probe array, GeneChip. Using hierarchical clustering-analysis, out of a total of 10,000 genes we identified a cluster containing 77 genes as having been up-regulated by CpG ODN. This cluster was further divided into two sub-clusters by means of time-kinetics. (1) Inflammatory cytokines such as IL-6 and GM-CSF were up-regulated predominantly 3 to 6 hours after stimulation with CpG ODN, presumably through activation of a transcription factor, NF-kappaB. (2) Interferon (IFN)-inducible anti-viral proteins, including IFIT1, OAS1 and Mx1, and Th1 chemoattractant IP-10, were up-regulated predominantly 6 to 24 hours after stimulation. Blocking with mAb against IFN-alpha/beta receptor strongly inhibited the induction of these IFN-inducible genes by CpG ODN.

Conclusion: This study provides new information regarding the possible immunomodulatory effects of CpG ODN in vivo via an IFN-alpha/beta receptor-mediated paracrine pathway.

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Related in: MedlinePlus

Effects of blocking mAbs against IFN receptors on up-regulation of mRNA expression by CpG ODN. PBMC were stimulated with IFN-α2 (200 IU/ml), IFN-γ (100 IU/ml) or CpG ODN 2006 (2 μg/ml) in the presence or absence of blocking mAbs against IFN-α/β receptor or IFN-γ receptor for 6 hours. The mRNA levels of IFIT1 (A), IP-10 (B) and OAS1 (C) were determined by SYBR® green-based real-time quantitative RT-PCR. The results are shown as the mean ± SEM of three independent experiments and expressed as -fold increase above the mRNA level in the control cells. In this experiment, up-regulation of IFIT1 and IP-10 by CpG ODN 2006 was not blocked by the addition of the isotype-matched control mAbs (data not shown). * p < 0.05 vs CpG ODN 2006 stimulated. NS: not significant.
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Figure 4: Effects of blocking mAbs against IFN receptors on up-regulation of mRNA expression by CpG ODN. PBMC were stimulated with IFN-α2 (200 IU/ml), IFN-γ (100 IU/ml) or CpG ODN 2006 (2 μg/ml) in the presence or absence of blocking mAbs against IFN-α/β receptor or IFN-γ receptor for 6 hours. The mRNA levels of IFIT1 (A), IP-10 (B) and OAS1 (C) were determined by SYBR® green-based real-time quantitative RT-PCR. The results are shown as the mean ± SEM of three independent experiments and expressed as -fold increase above the mRNA level in the control cells. In this experiment, up-regulation of IFIT1 and IP-10 by CpG ODN 2006 was not blocked by the addition of the isotype-matched control mAbs (data not shown). * p < 0.05 vs CpG ODN 2006 stimulated. NS: not significant.

Mentions: To elucidate whether IFN-inducible genes such as IFIT1, OAS1 and IP-10 were up-regulated directly by CpG ODN or as a result of autocrine/paracrine pathways of IFNs, blocking experiments were performed using anti-IFN receptor mAbs. Up-regulation of mRNA for IFIT1 by IFN-α2 and CpG ODN was significantly inhibited by the addition of anti-IFN-α/β receptor mAb (Fig. 4). With respect to IP-10, mRNA expression was up-regulated by IFN-α2, IFN-γ and CpG ODN. Induction of IP-10 by IFN-α2 or IFN-γ was significantly inhibited by the addition of mAbs specific for each IFN receptor (p < 0.05). The induction by CpG ODN was significantly inhibited by anti-IFN-α/β receptor mAb, whereas it was partially inhibited, though not statistically significant, by anti-IFN-γ receptor mAb (Fig. 4).


Interferon-alpha/beta receptor-mediated selective induction of a gene cluster by CpG oligodeoxynucleotide 2006.

Kato A, Homma T, Batchelor J, Hashimoto N, Imai S, Wakiguchi H, Saito H, Matsumoto K - BMC Immunol. (2003)

Effects of blocking mAbs against IFN receptors on up-regulation of mRNA expression by CpG ODN. PBMC were stimulated with IFN-α2 (200 IU/ml), IFN-γ (100 IU/ml) or CpG ODN 2006 (2 μg/ml) in the presence or absence of blocking mAbs against IFN-α/β receptor or IFN-γ receptor for 6 hours. The mRNA levels of IFIT1 (A), IP-10 (B) and OAS1 (C) were determined by SYBR® green-based real-time quantitative RT-PCR. The results are shown as the mean ± SEM of three independent experiments and expressed as -fold increase above the mRNA level in the control cells. In this experiment, up-regulation of IFIT1 and IP-10 by CpG ODN 2006 was not blocked by the addition of the isotype-matched control mAbs (data not shown). * p < 0.05 vs CpG ODN 2006 stimulated. NS: not significant.
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Related In: Results  -  Collection

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Figure 4: Effects of blocking mAbs against IFN receptors on up-regulation of mRNA expression by CpG ODN. PBMC were stimulated with IFN-α2 (200 IU/ml), IFN-γ (100 IU/ml) or CpG ODN 2006 (2 μg/ml) in the presence or absence of blocking mAbs against IFN-α/β receptor or IFN-γ receptor for 6 hours. The mRNA levels of IFIT1 (A), IP-10 (B) and OAS1 (C) were determined by SYBR® green-based real-time quantitative RT-PCR. The results are shown as the mean ± SEM of three independent experiments and expressed as -fold increase above the mRNA level in the control cells. In this experiment, up-regulation of IFIT1 and IP-10 by CpG ODN 2006 was not blocked by the addition of the isotype-matched control mAbs (data not shown). * p < 0.05 vs CpG ODN 2006 stimulated. NS: not significant.
Mentions: To elucidate whether IFN-inducible genes such as IFIT1, OAS1 and IP-10 were up-regulated directly by CpG ODN or as a result of autocrine/paracrine pathways of IFNs, blocking experiments were performed using anti-IFN receptor mAbs. Up-regulation of mRNA for IFIT1 by IFN-α2 and CpG ODN was significantly inhibited by the addition of anti-IFN-α/β receptor mAb (Fig. 4). With respect to IP-10, mRNA expression was up-regulated by IFN-α2, IFN-γ and CpG ODN. Induction of IP-10 by IFN-α2 or IFN-γ was significantly inhibited by the addition of mAbs specific for each IFN receptor (p < 0.05). The induction by CpG ODN was significantly inhibited by anti-IFN-α/β receptor mAb, whereas it was partially inhibited, though not statistically significant, by anti-IFN-γ receptor mAb (Fig. 4).

Bottom Line: Oligodeoxynucleotides containing unmethylated CpG motifs (CpG ODN) are known to exert a strong adjuvant effect on Th1 immune responses.Blocking with mAb against IFN-alpha/beta receptor strongly inhibited the induction of these IFN-inducible genes by CpG ODN.This study provides new information regarding the possible immunomodulatory effects of CpG ODN in vivo via an IFN-alpha/beta receptor-mediated paracrine pathway.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Allergy & Immunology, National Research Institute for Child Health & Development, Tokyo, Japan. atkato@nch.go.jp

ABSTRACT

Background: Oligodeoxynucleotides containing unmethylated CpG motifs (CpG ODN) are known to exert a strong adjuvant effect on Th1 immune responses. Although several genes have been reported, no comprehensive study of the gene expression profiles in human cells after stimulation with CpG ODN has been reported.

Results: This study was designed to identify a CpG-inducible gene cluster that potentially predicts for the molecular mechanisms of clinical efficacy of CpG ODN, by determining mRNA expression in human PBMC after stimulation with CpG ODN. PBMCs were obtained from the peripheral blood of healthy volunteers and cultured in the presence or absence of CpG ODN 2006 for up to 24 hours. The mRNA expression profile was evaluated using a high-density oligonucleotide probe array, GeneChip. Using hierarchical clustering-analysis, out of a total of 10,000 genes we identified a cluster containing 77 genes as having been up-regulated by CpG ODN. This cluster was further divided into two sub-clusters by means of time-kinetics. (1) Inflammatory cytokines such as IL-6 and GM-CSF were up-regulated predominantly 3 to 6 hours after stimulation with CpG ODN, presumably through activation of a transcription factor, NF-kappaB. (2) Interferon (IFN)-inducible anti-viral proteins, including IFIT1, OAS1 and Mx1, and Th1 chemoattractant IP-10, were up-regulated predominantly 6 to 24 hours after stimulation. Blocking with mAb against IFN-alpha/beta receptor strongly inhibited the induction of these IFN-inducible genes by CpG ODN.

Conclusion: This study provides new information regarding the possible immunomodulatory effects of CpG ODN in vivo via an IFN-alpha/beta receptor-mediated paracrine pathway.

Show MeSH
Related in: MedlinePlus