Plasticity of natural killer cells in pregnant patients infected with SARS-CoV-2 and their neonates during childbirth

Abstract

The COVID-19 pandemic has occurred due to infection caused by the SARS-CoV-2 coronavirus, which impacts gestation and pregnancy. In SARS-CoV-2 infection, only very rare cases of vertical transmission have been reported, suggesting that fetal immune imprinting due to a maternal infection is probably a result of changes in maternal immunity. Natural killer (NK) cells are the leading maternal immune cells that act as a natural defense system to fight infections. They also play a pivotal role in the establishment and maintenance of pregnancy. While peripheral NK cells display specific features in patients infected with SARS-CoV-2 in the general population, information remains elusive in pregnant mothers and neonates. In the present study, we analyzed the characteristics of NK cells isolated from both neonatal umbilical cord blood and maternal peripheral blood close to the time of delivery. Phenotype and functions were compared in 18 healthy pregnant women and 34 COVID-19 patients during pregnancy within an ongoing infection (PCR⁺; N = 15) or after recovery (IgG⁺PCR^-; N = 19). The frequency of NK cells from infected women and their neonates was correlated with the production of inflammatory cytokines in the serum. The expression of NKG2A and NKp30, as well as degranulation of NK cells in pregnant women with ongoing infection, were both negatively correlated to estradiol level. Furthermore, NK cells from the neonates born to infected women were significantly decreased and also correlated to estradiol level. This study highlights the relationship between NK cells, inflammation, and estradiol in patients with ongoing infection, providing new insights into the impact of maternal SARS-CoV-2 infection on the neonate.

Keywords: COVID-19; SARS-CoV-2; natural killer cells; neonates; pregnancy.

Figure 1

Study outline and hormone production. (A). Study outline of the recruitment of neonates and mothers. (B) Production of progesterone and estradiol in the sera of mothers. Data are shown for healthy controls (N= 18; C: crosses), recovered patients (N= 21; R: open squares), and patients with ongoing infection (N=16; O: circles). Open circles represent the asymptomatic patients and closed circles the symptomatic patients. Black lines represent the median. *p < 0.05; **p < 0.001.

Figure 2

Characteristic of NK cells in mothers and their neonates. (A) Absolute values of CD3^-CD56⁺ NK cells in mothers (maternal blood) and their neonates (cord blood). Data are shown for healthy controls (N= 18; C: crosses), recovered patients (N= 19; R: open squares), and patients with ongoing infection (N=15; O: circles). (B) Significant correlation of the absolute values of CD3^-CD56⁺ NK between mothers (maternal blood) and their neonates (cord blood). for the groups of recovered (R: open squares) and ongoing infection (O: circles). (C) Frequency of CD3^-CD56⁺ NK cells in mothers (maternal blood) and their neonates (cord blood). (D) Significant correlation between the frequency of CD3^-CD56⁺ NK cells in neonates (cord blood) born to mothers with ongoing (O: circles) infection and the production of estradiol in mothers. (E) Frequency of CD56^bright subpopulation of NK cells gated on CD3^-CD56⁺ NK cells. (F) Frequency of HLA-DR⁺-activated NK cells gated on CD3^-CD56⁺ NK cells in mothers (maternal blood) and their neonates (cord blood). Frequency’ data are shown for healthy controls (N= 10; C: crosses), recovered patients (N= 10; R: open squares), and patients with ongoing infection (N=13; O: circles). Open circles represent the asymptomatic patients and closed circles the symptomatic patients. Black lines represent the median. *p < 0.05; **p < 0.001; ****p < 0.00001; NS, non significant.

Figure 3

Pattern of receptor expression on CD3^-CD56⁺ NK cells. (A) Frequency of NKG2A, NKG2C, and NKp30 on NK cells gated on CD3^-CD56⁺ NK cells from mother (maternal blood) and their neonates (cord blood). (B) Correlation between the frequency of NK cell receptors and the production of estradiol in mothers (maternal blood) with ongoing infection (O: circles). (C) Frequency of NKp46 and DNAM-1 on NK cells gated on CD3^-CD56⁺ NK cells from mother (maternal blood) and their neonates (cord blood). Data are shown for healthy controls (N= 10; C: crosses), recovered patients (N= 10; R: squares), and patients with ongoing infection (N=13; O: circles). Open circles represent the asymptomatic patients and closed circles the symptomatic patients. Black lines represent the median. *p < 0.05; **p < 0.001.

Figure 4

Degranulation and production of cytokines by CD3^-CD56⁺ NK cells. (A) Degranulation of NK cells from mothers (maternal blood) and their neonates (cord blood), measured by cell surface expression of CD107a, in the presence of standard K562 target cells (ratio 1:1). (B) Correlation between CD107a and the production of estradiol in mothers (maternal blood) with ongoing infection (O: circles). (C) Intracellular production of IFN-γ and TNF-α in untreated cells (UT), after IL-12+IL-18 overnight stimulation (+IL-12/IL-18) or in the presence of standard K562 target cells (ratio 1:1). Data are shown NK cells of mothers (maternal blood) and their neonates (cord blood) from healthy controls (N= 10; C: crosses), recovered patients (N= 10; R: squares), and patients with ongoing infection (N=13; O: circles). Open circles represent the asymptomatic patients and closed circles the symptomatic patients. Black lines represent the median.

Figure 5

Production of cytokines in the sera of mothers and their neonates. (A) Production of IL-8 in the sera of mothers (maternal blood) and their neonates (cord blood). (B) Correlation between IL-8 production and the frequency of NK cells in mothers (maternal blood) with ongoing infection (O: circles). (C) Production of IL-22 in the sera of mothers (maternal blood) and their neonates (cord blood). (D) Correlation between IL-22 production and the frequency of NK cells in neonates born to mothers with ongoing infection (cord blood (O)). (E) Correlation between production of IL-22 production in neonates (cord blood) born to mothers with ongoing infection (cord blood (O)) and the production of estradiol in mothers with ongoing infection. Data are shown for healthy controls (N= 18; C: crosses), recovered patients (N= 19; R: squares), and patients with ongoing infection (N=15; O: circles). Open circles represent the asymptomatic patients and closed circles the symptomatic patients. Black lines represent the median.