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Sustained Immune Complex-Mediated Reduction in CD16 Expression after Vaccination Regulates NK Cell Function

View Article: PubMed Central - PubMed

ABSTRACT

Cross-linking of FcγRIII (CD16) by immune complexes induces antibody-dependent cellular cytotoxicity (ADCC) by natural killer (NK) cells, contributing to control of intracellular pathogens; this pathway can also be targeted for immunotherapy of cancerous or otherwise diseased cells. However, downregulation of CD16 expression on activated NK cells may limit or regulate this response. Here, we report sustained downregulation of CD16 expression on NK cells in vivo after intramuscular (but not intranasal) influenza vaccination. CD16 downregulation persisted for at least 12 weeks after vaccination and was associated with robust enhancement of influenza-specific plasma antibodies after intramuscular (but not intranasal) vaccination. This effect could be emulated in vitro by co-culture of NK cells with influenza antigen and immune serum and, consistent with the sustained effects after vaccination, only very limited recovery of CD16 expression was observed during long-term in vitro culture of immune complex-treated cells. CD16 downregulation was most marked among normally CD16high CD57+ NK cells, irrespective of NKG2C expression, and was strongly positively associated with degranulation (surface CD107a expression). CD16 downregulation was partially reversed by inhibition of ADAM17 matrix metalloprotease, leading to a sustained increase in both CD107a and CD25 (IL-2Rα) expression. Both the degranulation and CD25 responses of CD57+ NK cells were uniquely dependent on trivalent influenza vaccine-specific IgG. These data support a role for CD16 in early activation of NK cells after vaccination and for CD16 downregulation as a means to modulate NK cell responses and maintain immune homeostasis of both antibody and T cell-dependent pathways.

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Immune complex-induced shedding and sustained downregulation of NK cell CD16 expression. (A) Downregulation of CD16 requires IgG-replete human plasma and TIV antigen. PBMC were cultured for 5 h in FCS, IgG-depleted human plasma or IgG-replete plasma in the presence or absence of TIV. (B) ImageStream™ analysis of NK cells after 5 h culture with TIV plus human plasma (containing anti-influenza IgG) or FCS. Examples are shown of CD56dim CD57+ NK cells stained for surface (external) and intracellular (internal) CD16. (C) Time course for recovery of CD16 expression in vitro. Expression of CD16 was tracked by flow cytometry up to 18 days after culture of PBMC from five separate donors with TIV for 18 h in the presence of 1% human plasma (containing anti-influenza IgG). CD16 expression is shown for CD56bright, CD56dimCD57−, and CD56dim CD57+ NK cell subsets. Trend analysis was performed using a one-way repeated measures ANOVA (*p < 0.05).
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Figure 2: Immune complex-induced shedding and sustained downregulation of NK cell CD16 expression. (A) Downregulation of CD16 requires IgG-replete human plasma and TIV antigen. PBMC were cultured for 5 h in FCS, IgG-depleted human plasma or IgG-replete plasma in the presence or absence of TIV. (B) ImageStream™ analysis of NK cells after 5 h culture with TIV plus human plasma (containing anti-influenza IgG) or FCS. Examples are shown of CD56dim CD57+ NK cells stained for surface (external) and intracellular (internal) CD16. (C) Time course for recovery of CD16 expression in vitro. Expression of CD16 was tracked by flow cytometry up to 18 days after culture of PBMC from five separate donors with TIV for 18 h in the presence of 1% human plasma (containing anti-influenza IgG). CD16 expression is shown for CD56bright, CD56dimCD57−, and CD56dim CD57+ NK cell subsets. Trend analysis was performed using a one-way repeated measures ANOVA (*p < 0.05).

Mentions: IgG-dependent downregulation of CD16 may be due to internalization of CD16–IgG–Ag complexes or to cleavage of CD16 at the cell surface and shedding into the extracellular milieu. Downregulation of CD16 did not occur in cells cultured in FCS or in either IgG-depleted or IgG-replete human plasma in the absence of TIV antigen. CD16 downregulation was seen only when cells were cultured in IgG-replete plasma with TIV antigen but not when TIV antigen was added to cultures containing FCS or IgG-depleted plasma (Figure 2A). To determine whether CD16 was internalized after IgG–Ag cross-linking, NK cells were incubated with TIV plus IgG-replete (immune) human plasma or FCS for 5 h and analyzed by Imagestream™ for extracellular and intracellular CD16 (Figure 2B). Surface staining for CD16 was clearly visible on CD56dim CD57+ NK cells cultured with TIV + FCS but not on cells cultured with TIV and immune human plasma. In neither case was CD16 detected intracellularly, indicating that loss of CD16 from the cell surface was not accompanied by internalization of intact CD16.


Sustained Immune Complex-Mediated Reduction in CD16 Expression after Vaccination Regulates NK Cell Function
Immune complex-induced shedding and sustained downregulation of NK cell CD16 expression. (A) Downregulation of CD16 requires IgG-replete human plasma and TIV antigen. PBMC were cultured for 5 h in FCS, IgG-depleted human plasma or IgG-replete plasma in the presence or absence of TIV. (B) ImageStream™ analysis of NK cells after 5 h culture with TIV plus human plasma (containing anti-influenza IgG) or FCS. Examples are shown of CD56dim CD57+ NK cells stained for surface (external) and intracellular (internal) CD16. (C) Time course for recovery of CD16 expression in vitro. Expression of CD16 was tracked by flow cytometry up to 18 days after culture of PBMC from five separate donors with TIV for 18 h in the presence of 1% human plasma (containing anti-influenza IgG). CD16 expression is shown for CD56bright, CD56dimCD57−, and CD56dim CD57+ NK cell subsets. Trend analysis was performed using a one-way repeated measures ANOVA (*p < 0.05).
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Figure 2: Immune complex-induced shedding and sustained downregulation of NK cell CD16 expression. (A) Downregulation of CD16 requires IgG-replete human plasma and TIV antigen. PBMC were cultured for 5 h in FCS, IgG-depleted human plasma or IgG-replete plasma in the presence or absence of TIV. (B) ImageStream™ analysis of NK cells after 5 h culture with TIV plus human plasma (containing anti-influenza IgG) or FCS. Examples are shown of CD56dim CD57+ NK cells stained for surface (external) and intracellular (internal) CD16. (C) Time course for recovery of CD16 expression in vitro. Expression of CD16 was tracked by flow cytometry up to 18 days after culture of PBMC from five separate donors with TIV for 18 h in the presence of 1% human plasma (containing anti-influenza IgG). CD16 expression is shown for CD56bright, CD56dimCD57−, and CD56dim CD57+ NK cell subsets. Trend analysis was performed using a one-way repeated measures ANOVA (*p < 0.05).
Mentions: IgG-dependent downregulation of CD16 may be due to internalization of CD16–IgG–Ag complexes or to cleavage of CD16 at the cell surface and shedding into the extracellular milieu. Downregulation of CD16 did not occur in cells cultured in FCS or in either IgG-depleted or IgG-replete human plasma in the absence of TIV antigen. CD16 downregulation was seen only when cells were cultured in IgG-replete plasma with TIV antigen but not when TIV antigen was added to cultures containing FCS or IgG-depleted plasma (Figure 2A). To determine whether CD16 was internalized after IgG–Ag cross-linking, NK cells were incubated with TIV plus IgG-replete (immune) human plasma or FCS for 5 h and analyzed by Imagestream™ for extracellular and intracellular CD16 (Figure 2B). Surface staining for CD16 was clearly visible on CD56dim CD57+ NK cells cultured with TIV + FCS but not on cells cultured with TIV and immune human plasma. In neither case was CD16 detected intracellularly, indicating that loss of CD16 from the cell surface was not accompanied by internalization of intact CD16.

View Article: PubMed Central - PubMed

ABSTRACT

Cross-linking of Fc&gamma;RIII (CD16) by immune complexes induces antibody-dependent cellular cytotoxicity (ADCC) by natural killer (NK) cells, contributing to control of intracellular pathogens; this pathway can also be targeted for immunotherapy of cancerous or otherwise diseased cells. However, downregulation of CD16 expression on activated NK cells may limit or regulate this response. Here, we report sustained downregulation of CD16 expression on NK cells in vivo after intramuscular (but not intranasal) influenza vaccination. CD16 downregulation persisted for at least 12&thinsp;weeks after vaccination and was associated with robust enhancement of influenza-specific plasma antibodies after intramuscular (but not intranasal) vaccination. This effect could be emulated in vitro by co-culture of NK cells with influenza antigen and immune serum and, consistent with the sustained effects after vaccination, only very limited recovery of CD16 expression was observed during long-term in vitro culture of immune complex-treated cells. CD16 downregulation was most marked among normally CD16high CD57+ NK cells, irrespective of NKG2C expression, and was strongly positively associated with degranulation (surface CD107a expression). CD16 downregulation was partially reversed by inhibition of ADAM17 matrix metalloprotease, leading to a sustained increase in both CD107a and CD25 (IL-2R&alpha;) expression. Both the degranulation and CD25 responses of CD57+ NK cells were uniquely dependent on trivalent influenza vaccine-specific IgG. These data support a role for CD16 in early activation of NK cells after vaccination and for CD16 downregulation as a means to modulate NK cell responses and maintain immune homeostasis of both antibody and T cell-dependent pathways.

No MeSH data available.


Related in: MedlinePlus