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Elevation of intact and proteolytic fragments of acute phase proteins constitutes the earliest systemic antiviral response in HIV-1 infection.

Kramer HB, Lavender KJ, Qin L, Stacey AR, Liu MK, di Gleria K, Simmons A, Gasper-Smith N, Haynes BF, McMichael AJ, Borrow P, Kessler BM - PLoS Pathog. (2010)

Bottom Line: Both A-SAA and VIRIP have anti-viral activity in vitro and quantitation of their plasma levels indicated that circulating concentrations are likely to be within the range of their inhibitory activity.Our results provide evidence for a first wave of host anti-viral defense occurring in the eclipse phase of AHI prior to systemic activation of other immune responses.Insights gained into the mechanism of action of acute-phase reactants and other innate molecules against HIV and how they are induced could be exploited for the future development of more efficient prophylactic vaccine strategies.

View Article: PubMed Central - PubMed

Affiliation: Henry Wellcome Building for Molecular Physiology, Nuffield Department of Clinical Medicine, University of Oxford, Oxford, Oxfordshire, United Kingdom.

ABSTRACT
The earliest immune responses activated in acute human immunodeficiency virus type 1 infection (AHI) exert a critical influence on subsequent virus spread or containment. During this time frame, components of the innate immune system such as macrophages and DCs, NK cells, beta-defensins, complement and other anti-microbial factors, which have all been implicated in modulating HIV infection, may play particularly important roles. A proteomics-based screen was performed on a cohort from whom samples were available at time points prior to the earliest positive HIV detection. The ability of selected factors found to be elevated in the plasma during AHI to inhibit HIV-1 replication was analyzed using in vitro PBMC and DC infection models. Analysis of unique plasma donor panels spanning the eclipse and viral expansion phases revealed very early alterations in plasma proteins in AHI. Induction of acute phase protein serum amyloid A (A-SAA) occurred as early as 5-7 days prior to the first detection of plasma viral RNA, considerably prior to any elevation in systemic cytokine levels. Furthermore, a proteolytic fragment of alpha-1-antitrypsin (AAT), termed virus inhibitory peptide (VIRIP), was observed in plasma coincident with viremia. Both A-SAA and VIRIP have anti-viral activity in vitro and quantitation of their plasma levels indicated that circulating concentrations are likely to be within the range of their inhibitory activity. Our results provide evidence for a first wave of host anti-viral defense occurring in the eclipse phase of AHI prior to systemic activation of other immune responses. Insights gained into the mechanism of action of acute-phase reactants and other innate molecules against HIV and how they are induced could be exploited for the future development of more efficient prophylactic vaccine strategies.

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Tandem mass spectrometry-based detection of the antiviral peptide VIRIP, derived from the C-terminus of AAT, in plasma during AHI.(A) MS/MS spectrum of precursor ion peptide 773.75 Da [M(ox)+3H]3+ identified as VIRIP 377–396 derived from AAT (Swissprot accession nr P01009) as detected in plasma panel 9012 (time point 5 d post-T0). Identified b- and y- fragment ions are shown. (B) LC-MS/MS based detection of the VIRIP precursor ion 773.75 Da [M(ox)+3H]3+ in plasma panels 63229 and 9012. Mass peak ion counts are shown in black, and viremic time points are indicated by the grey arrow. (C) Quantitation of synthetic VIRIP peptide by LC-MS/MS indicates that the amount of VIRIP detected in plasma is in the low micromolar range.
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ppat-1000893-g004: Tandem mass spectrometry-based detection of the antiviral peptide VIRIP, derived from the C-terminus of AAT, in plasma during AHI.(A) MS/MS spectrum of precursor ion peptide 773.75 Da [M(ox)+3H]3+ identified as VIRIP 377–396 derived from AAT (Swissprot accession nr P01009) as detected in plasma panel 9012 (time point 5 d post-T0). Identified b- and y- fragment ions are shown. (B) LC-MS/MS based detection of the VIRIP precursor ion 773.75 Da [M(ox)+3H]3+ in plasma panels 63229 and 9012. Mass peak ion counts are shown in black, and viremic time points are indicated by the grey arrow. (C) Quantitation of synthetic VIRIP peptide by LC-MS/MS indicates that the amount of VIRIP detected in plasma is in the low micromolar range.

Mentions: A more comprehensive analysis by liquid chromatography tandem mass spectrometry (LC-MS/MS) revealed a number of other plasma components including complement factors, apo-lipoproteins and alpha-1-antitrypsin (AAT) that are present in plasma during AHI (Table S1). The factors found to be elevated in plasma during AHI included a C-terminal peptide derived from AAT, residues 377–396, referred to as VIRIP (Table S1 and Fig. 4A), which was shown to inhibit HIV-1 entry into host cells by targeting the gp41 fusion peptide [10]. Seven plasma panels from HIV-infected individuals and five panels from uninfected controls were evaluated for the presence of VIRIP by tandem mass spectrometry in a semi-quantitative fashion. Ion counts detected for the precursor ion representing the expected molecular mass of VIRIP were correlated to viremia, and revealed an elevation of VIRIP coincident with and after the initial increase in viremia in two of the seven plasma donor panels from infected individuals, but none in the five controls (Fig. 4B). A semi-quantitative titration of VIRIP peptide by mass spectrometry indicated that the amount detected corresponds to an estimated value of 0.1–0.3µM of VIRIP in plasma at peak concentrations (Fig. 4C). Considering the sample loss during the isolation of VIRIP peptide from plasma, the effective VIRIP concentration will likely be in the range of low µg/ml, which is approximating the IC50 value at which VIRIP interferes with HIV-1 entry [10] (see also below).


Elevation of intact and proteolytic fragments of acute phase proteins constitutes the earliest systemic antiviral response in HIV-1 infection.

Kramer HB, Lavender KJ, Qin L, Stacey AR, Liu MK, di Gleria K, Simmons A, Gasper-Smith N, Haynes BF, McMichael AJ, Borrow P, Kessler BM - PLoS Pathog. (2010)

Tandem mass spectrometry-based detection of the antiviral peptide VIRIP, derived from the C-terminus of AAT, in plasma during AHI.(A) MS/MS spectrum of precursor ion peptide 773.75 Da [M(ox)+3H]3+ identified as VIRIP 377–396 derived from AAT (Swissprot accession nr P01009) as detected in plasma panel 9012 (time point 5 d post-T0). Identified b- and y- fragment ions are shown. (B) LC-MS/MS based detection of the VIRIP precursor ion 773.75 Da [M(ox)+3H]3+ in plasma panels 63229 and 9012. Mass peak ion counts are shown in black, and viremic time points are indicated by the grey arrow. (C) Quantitation of synthetic VIRIP peptide by LC-MS/MS indicates that the amount of VIRIP detected in plasma is in the low micromolar range.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2865525&req=5

ppat-1000893-g004: Tandem mass spectrometry-based detection of the antiviral peptide VIRIP, derived from the C-terminus of AAT, in plasma during AHI.(A) MS/MS spectrum of precursor ion peptide 773.75 Da [M(ox)+3H]3+ identified as VIRIP 377–396 derived from AAT (Swissprot accession nr P01009) as detected in plasma panel 9012 (time point 5 d post-T0). Identified b- and y- fragment ions are shown. (B) LC-MS/MS based detection of the VIRIP precursor ion 773.75 Da [M(ox)+3H]3+ in plasma panels 63229 and 9012. Mass peak ion counts are shown in black, and viremic time points are indicated by the grey arrow. (C) Quantitation of synthetic VIRIP peptide by LC-MS/MS indicates that the amount of VIRIP detected in plasma is in the low micromolar range.
Mentions: A more comprehensive analysis by liquid chromatography tandem mass spectrometry (LC-MS/MS) revealed a number of other plasma components including complement factors, apo-lipoproteins and alpha-1-antitrypsin (AAT) that are present in plasma during AHI (Table S1). The factors found to be elevated in plasma during AHI included a C-terminal peptide derived from AAT, residues 377–396, referred to as VIRIP (Table S1 and Fig. 4A), which was shown to inhibit HIV-1 entry into host cells by targeting the gp41 fusion peptide [10]. Seven plasma panels from HIV-infected individuals and five panels from uninfected controls were evaluated for the presence of VIRIP by tandem mass spectrometry in a semi-quantitative fashion. Ion counts detected for the precursor ion representing the expected molecular mass of VIRIP were correlated to viremia, and revealed an elevation of VIRIP coincident with and after the initial increase in viremia in two of the seven plasma donor panels from infected individuals, but none in the five controls (Fig. 4B). A semi-quantitative titration of VIRIP peptide by mass spectrometry indicated that the amount detected corresponds to an estimated value of 0.1–0.3µM of VIRIP in plasma at peak concentrations (Fig. 4C). Considering the sample loss during the isolation of VIRIP peptide from plasma, the effective VIRIP concentration will likely be in the range of low µg/ml, which is approximating the IC50 value at which VIRIP interferes with HIV-1 entry [10] (see also below).

Bottom Line: Both A-SAA and VIRIP have anti-viral activity in vitro and quantitation of their plasma levels indicated that circulating concentrations are likely to be within the range of their inhibitory activity.Our results provide evidence for a first wave of host anti-viral defense occurring in the eclipse phase of AHI prior to systemic activation of other immune responses.Insights gained into the mechanism of action of acute-phase reactants and other innate molecules against HIV and how they are induced could be exploited for the future development of more efficient prophylactic vaccine strategies.

View Article: PubMed Central - PubMed

Affiliation: Henry Wellcome Building for Molecular Physiology, Nuffield Department of Clinical Medicine, University of Oxford, Oxford, Oxfordshire, United Kingdom.

ABSTRACT
The earliest immune responses activated in acute human immunodeficiency virus type 1 infection (AHI) exert a critical influence on subsequent virus spread or containment. During this time frame, components of the innate immune system such as macrophages and DCs, NK cells, beta-defensins, complement and other anti-microbial factors, which have all been implicated in modulating HIV infection, may play particularly important roles. A proteomics-based screen was performed on a cohort from whom samples were available at time points prior to the earliest positive HIV detection. The ability of selected factors found to be elevated in the plasma during AHI to inhibit HIV-1 replication was analyzed using in vitro PBMC and DC infection models. Analysis of unique plasma donor panels spanning the eclipse and viral expansion phases revealed very early alterations in plasma proteins in AHI. Induction of acute phase protein serum amyloid A (A-SAA) occurred as early as 5-7 days prior to the first detection of plasma viral RNA, considerably prior to any elevation in systemic cytokine levels. Furthermore, a proteolytic fragment of alpha-1-antitrypsin (AAT), termed virus inhibitory peptide (VIRIP), was observed in plasma coincident with viremia. Both A-SAA and VIRIP have anti-viral activity in vitro and quantitation of their plasma levels indicated that circulating concentrations are likely to be within the range of their inhibitory activity. Our results provide evidence for a first wave of host anti-viral defense occurring in the eclipse phase of AHI prior to systemic activation of other immune responses. Insights gained into the mechanism of action of acute-phase reactants and other innate molecules against HIV and how they are induced could be exploited for the future development of more efficient prophylactic vaccine strategies.

Show MeSH
Related in: MedlinePlus