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An AGM model for changes in complement during pregnancy: neutralization of influenza virus by serum is diminished in late third trimester.

Mayer AE, Parks GD - PLoS ONE (2014)

Bottom Line: The relative contributions of these components to the anti-viral response are difficult to dissect because most humans have pre-existing influenza-specific Abs.Strikingly, we found that circulating levels of C3, C3a, and C4 are diminished late in gestation relative to nonpregnant animals, and while neutralization capacity and serum C3a return to normal shortly after parturition, C3 and C4 levels do not.This AGM model system will enable further studies of the role of physiologic and hormonal changes in downregulating C'-mediated anti-viral immunity during pregnancy, and it will permit the identification of therapeutic targets to improve outcomes of influenza virus infection in pregnant women.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Wake Forest School of Medicine, Winston-Salem, NC 27101, United States of America.

ABSTRACT
Pregnant women in the third trimester are at increased risk of severe influenza disease relative to the general population, though mechanisms behind this are not completely understood. The immune response to influenza infection employs both complement (C') and antibody (Ab). The relative contributions of these components to the anti-viral response are difficult to dissect because most humans have pre-existing influenza-specific Abs. We developed the African green monkey (AGM) as a tractable nonhuman primate model to study changes in systemic innate immunity to influenza during pregnancy. Because the AGMs were influenza-naïve, we were able to examine the role of C' in influenza virus neutralization using serum from non-pregnant animals before and after influenza infection. We determined that serum from naïve AGMs neutralized influenza via C', while post-infection neutralization did not require C', suggesting an Ab-mediated mechanism. The latter mimicked neutralization using human serum. Further, we found that ex vivo neutralization of influenza with both naïve and influenza-immune AGM serum occurred by virus particle aggregation and lysis, with immune serum lysing virus at a much higher rate than naïve serum. We hypothesized that the anti-influenza C' response would diminish late in AGM pregnancy, corresponding with the time when pregnant women suffer increased influenza severity. We found that influenza neutralization capacity is significantly diminished in serum collected late in the third trimester. Strikingly, we found that circulating levels of C3, C3a, and C4 are diminished late in gestation relative to nonpregnant animals, and while neutralization capacity and serum C3a return to normal shortly after parturition, C3 and C4 levels do not. This AGM model system will enable further studies of the role of physiologic and hormonal changes in downregulating C'-mediated anti-viral immunity during pregnancy, and it will permit the identification of therapeutic targets to improve outcomes of influenza virus infection in pregnant women.

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Effect of HI on the neutralization capacity of pre- and post-immune sera from AGM experimentally infected with PR8.Two AGMs were inoculated with PR8 as described in the Materials and Methods. Pre-immune sera (d 0) and sera collected at d 14 post-infection were tested at the indicated fold dilutions for in vitro neutralization of PR8-GFP as described in Fig. 2. Results are expressed as the mean plus SD from triplicate experiments to determine the percentage of GFP-positive cells compared to in vitro control infections (which lacked serum treatment). *** p<0.0001 as compared to matching serum treatment for the same animal at day 0 (shown on day 14 values) based on t-tests on log transformed neutralization percentages.
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pone-0112749-g004: Effect of HI on the neutralization capacity of pre- and post-immune sera from AGM experimentally infected with PR8.Two AGMs were inoculated with PR8 as described in the Materials and Methods. Pre-immune sera (d 0) and sera collected at d 14 post-infection were tested at the indicated fold dilutions for in vitro neutralization of PR8-GFP as described in Fig. 2. Results are expressed as the mean plus SD from triplicate experiments to determine the percentage of GFP-positive cells compared to in vitro control infections (which lacked serum treatment). *** p<0.0001 as compared to matching serum treatment for the same animal at day 0 (shown on day 14 values) based on t-tests on log transformed neutralization percentages.

Mentions: The capacity of sera to neutralize PR8 was tested using the PR8-GFP flow cytometry assay described above. As shown in Fig. 4, pre-immune sera showed dose-dependent neutralization of PR8-GFP and no neutralization was seen with HI sera. Interestingly, post-immune sera (normal and HI) were better at neutralizing PR8 than any Hu serum sample tested, as evidenced by complete neutralization of PR8-GFP out to a 1∶640 dilution (Fig. 4). The finding that PR8 was neutralized by HI post- but not pre-immune sera demonstrated that C′ is less important for ex vivo influenza neutralization after AGMs have been exposed to influenza virus.


An AGM model for changes in complement during pregnancy: neutralization of influenza virus by serum is diminished in late third trimester.

Mayer AE, Parks GD - PLoS ONE (2014)

Effect of HI on the neutralization capacity of pre- and post-immune sera from AGM experimentally infected with PR8.Two AGMs were inoculated with PR8 as described in the Materials and Methods. Pre-immune sera (d 0) and sera collected at d 14 post-infection were tested at the indicated fold dilutions for in vitro neutralization of PR8-GFP as described in Fig. 2. Results are expressed as the mean plus SD from triplicate experiments to determine the percentage of GFP-positive cells compared to in vitro control infections (which lacked serum treatment). *** p<0.0001 as compared to matching serum treatment for the same animal at day 0 (shown on day 14 values) based on t-tests on log transformed neutralization percentages.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4237339&req=5

pone-0112749-g004: Effect of HI on the neutralization capacity of pre- and post-immune sera from AGM experimentally infected with PR8.Two AGMs were inoculated with PR8 as described in the Materials and Methods. Pre-immune sera (d 0) and sera collected at d 14 post-infection were tested at the indicated fold dilutions for in vitro neutralization of PR8-GFP as described in Fig. 2. Results are expressed as the mean plus SD from triplicate experiments to determine the percentage of GFP-positive cells compared to in vitro control infections (which lacked serum treatment). *** p<0.0001 as compared to matching serum treatment for the same animal at day 0 (shown on day 14 values) based on t-tests on log transformed neutralization percentages.
Mentions: The capacity of sera to neutralize PR8 was tested using the PR8-GFP flow cytometry assay described above. As shown in Fig. 4, pre-immune sera showed dose-dependent neutralization of PR8-GFP and no neutralization was seen with HI sera. Interestingly, post-immune sera (normal and HI) were better at neutralizing PR8 than any Hu serum sample tested, as evidenced by complete neutralization of PR8-GFP out to a 1∶640 dilution (Fig. 4). The finding that PR8 was neutralized by HI post- but not pre-immune sera demonstrated that C′ is less important for ex vivo influenza neutralization after AGMs have been exposed to influenza virus.

Bottom Line: The relative contributions of these components to the anti-viral response are difficult to dissect because most humans have pre-existing influenza-specific Abs.Strikingly, we found that circulating levels of C3, C3a, and C4 are diminished late in gestation relative to nonpregnant animals, and while neutralization capacity and serum C3a return to normal shortly after parturition, C3 and C4 levels do not.This AGM model system will enable further studies of the role of physiologic and hormonal changes in downregulating C'-mediated anti-viral immunity during pregnancy, and it will permit the identification of therapeutic targets to improve outcomes of influenza virus infection in pregnant women.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Wake Forest School of Medicine, Winston-Salem, NC 27101, United States of America.

ABSTRACT
Pregnant women in the third trimester are at increased risk of severe influenza disease relative to the general population, though mechanisms behind this are not completely understood. The immune response to influenza infection employs both complement (C') and antibody (Ab). The relative contributions of these components to the anti-viral response are difficult to dissect because most humans have pre-existing influenza-specific Abs. We developed the African green monkey (AGM) as a tractable nonhuman primate model to study changes in systemic innate immunity to influenza during pregnancy. Because the AGMs were influenza-naïve, we were able to examine the role of C' in influenza virus neutralization using serum from non-pregnant animals before and after influenza infection. We determined that serum from naïve AGMs neutralized influenza via C', while post-infection neutralization did not require C', suggesting an Ab-mediated mechanism. The latter mimicked neutralization using human serum. Further, we found that ex vivo neutralization of influenza with both naïve and influenza-immune AGM serum occurred by virus particle aggregation and lysis, with immune serum lysing virus at a much higher rate than naïve serum. We hypothesized that the anti-influenza C' response would diminish late in AGM pregnancy, corresponding with the time when pregnant women suffer increased influenza severity. We found that influenza neutralization capacity is significantly diminished in serum collected late in the third trimester. Strikingly, we found that circulating levels of C3, C3a, and C4 are diminished late in gestation relative to nonpregnant animals, and while neutralization capacity and serum C3a return to normal shortly after parturition, C3 and C4 levels do not. This AGM model system will enable further studies of the role of physiologic and hormonal changes in downregulating C'-mediated anti-viral immunity during pregnancy, and it will permit the identification of therapeutic targets to improve outcomes of influenza virus infection in pregnant women.

Show MeSH
Related in: MedlinePlus