Background/Purpose: Congenital heart block (CHB) may develop in the fetus of women with Ro/SSA autoantibodies. The mothers are commonly diagnosed with Sjögren’s syndrome or SLE. During pregnancy, the antibodies are transported across placenta and bind to the fetal heart where inflammation develops, leading to the fibrosis and calcification that cause the permanent disruption of impulse propagation. The mechanism by which the antibodies initiate inflammation is however not understood. In the mother, Ro/SSA antibodies may induce type I IFN production, and we recently described an upregulation of type I IFN-regulated genes in PBMC as well as an increase in circulating IFN-alpha also in the Ro/SSA exposed newborns. IFN-alpha is known to influence leukocytes, expand and activate NK cells, and in this study, we, therefore, analyzed the immune cell populations in cord blood of anti-Ro/SSA exposed neonates and evaluated the influence of IFN-alpha on fetal cardiac cells.

Methods: Maternal and cord blood was sampled at birth from healthy controls (HC) (n=9) and Ro/SSA positive pregnancies (n=13). PMBC were prepared and used for microarray analysis and for flow cytometry to define CD19⁺ B cell (CD27^–IgD⁺ naïve, CD27⁺IgD^– memory, CD27⁺IgD⁺ marginal zone), CD3⁺ T cell (CD8⁺, CD4⁺) subpopulations and CD16⁺CD56⁺ NK cells. Cardiomyocyte cultures were established from human fetal tissues (n=5) and stimulated by medium with or without IFN-alpha for 6 hours before mRNA preparation and microarray analysis.

Results: In the Ro/SSA positive mothers, an increase in naïve B cells, but decrease in memory and marginal zone cells was observed, confirming previous reports for non-pregnant Sjögren’s syndrome and SLE. Surprisingly, Ro/SSA exposed neonates presented an expanded population of NK cells (p=0.02), the presence of which was influenced by immunomodulatory treatment of the mother (neonates of non-treated mothers p=0.002, neonates of treated mothers p=ns compared to HC). No other differences in the T or B cell subsets analyzed were observed. Microarray analysis of PBMC revealed a type II IFN signature with high IFNγ, the prototype cytokine produced by activated NK cells, in Ro/SSA exposed neonates compared to HC. Finally, stimulation of fetal cardiomyocytes with IFN-alpha induced upregulation of MICA and MICB (major histocompatibility complex (MHC) class I chain related sequence A and B), which are ligands for activating NK cell receptors.

Conclusion: Our data demonstrate an expansion of NK cells and their activity markers in Ro/SSA exposed neonates, as well as an upregulation of activating NK cell receptors in fetal cardiac cells after IFN-alpha exposure, indicating that NK cell related effector mechanisms such as antibody-dependent cell cytotoxicity (ADCC) may be a central mechanism by which the inflammation is initiated in CHB. The expansion of NK cells in neonates at risk for CHB is a novel observation, and implicates fetal innate immune mechanisms in the pathogenesis.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/rossa-autoantibody-exposed-neonates-have-an-expansion-of-nk-cells-and-a-discernible-type-ii-ifn-signature-with-high-ifn%ce%b3-in-peripheral-blood/