Single-Cell Meta-Analysis of Neutrophil Activation in Kawasaki Disease and Multisystem Inflammatory Syndrome in Children Reveals Potential Shared Immunological Drivers

Abstract

BACKGROUND: Kawasaki disease (KD) and multisystem inflammatory syndrome in children (MIS-C) share similar clinical manifestations, including cardiovascular complications, suggesting similar underlying immunopathogenic processes. Aberrant neutrophil activation may play a crucial role in the shared pathologies of KD and MIS-C; however, the associated pathogenic mechanisms and molecular drivers remain unknown. METHODS: We performed a single-cell meta-analysis of neutrophil activation with 103 pediatric single-cell transcriptomic peripheral blood mononuclear cell data across 9 cohorts, including healthy controls, KD, MIS-C, compared with dengue virus infection, juvenile idiopathic arthritis, and pediatric celiac disease. We used a series of computational analyses to investigate the shared neutrophil transcriptional programs of KD and MIS-C that are linked to systemic damage and cardiac pathologies, and suggested Food and Drug Administration–approved drugs to consider as KD and MIS-C treatment. RESULTS: We meta-analyzed 521 950 high-quality cells. We found that blood signatures associated with risks of cardiovascular events are enriched in neutrophils of KD and MIS-C. We revealed the expansion of CD177 + neutrophils harboring hyperactivated effector functions in both KD and MIS-C, but not in healthy controls or in other viral-, inflammatory-, or immune-related pediatric diseases. KD and MIS-C CD177 + neutrophils had highly similar transcriptomes, marked by conserved signatures and pathways related to molecular damage. We found the induction of a shared neutrophil expression program, potentially regulated by SPI1 (Spi-1 proto-oncogene), which confers enhanced effector functions, especially neutrophil degranulation. CD177 and shared neutrophil expression program expressions were associated with acute stages and attenuated during KD intravenous immunoglobulin treatment and MIS-C recovery. Network analysis identified hub genes that correlated with the high activation of CD177 + neutrophils. Disease-gene association analysis revealed that the CD177 + neutrophil shared neutrophil expression program was associated with the development of coronary and myocardial disorders. Last, we identified and validated TSPO (translocator protein) and S100A12 (S100 calcium-binding protein A12) as main molecular targets, for which the Food and Drug Administration–approved drugs methotrexate, zaleplon, metronidazole, lorazepam, clonazepam, temazepam, and zolpidem, among others, are primary candidates for drug repurposing. CONCLUSIONS: Our findings indicate that CD177 + neutrophils may exert systemic pathological damage contributing to the shared morbidities in KD and MIS-C. We uncovered potential regulatory drivers of CD177 + neutrophil hyperactivation and pathogenicity that may be targeted as a single therapeutic strategy for either KD or MIS-C.

Exploration of common genomic signatures of Systemic juvenile rheumatoid arthritis and Kawasaki disease

“…Acute medium-vessel vasculitis, known as KD, primarily affects infants older than six months and children under ve years old. It is the leading cause of acquired heart disease in children and predisposes them to the formation of coronary artery aneurysms [12] . Clinical signs of KD and SJIA are comparable, indicating similar underlying immunopathogenic mechanisms.…”

“…However, an overabundance of neutrophils may cause several illnesses, including systemic lupus erythematosus, asthma, rheumatoid arthritis, ischemia-reperfusion injury, and vascular injury, among other tissue damage and disorders [31,32] . Neutrophils signi cantly increase during the acute phase of KD and damage endothelial cells by secreting oxygen free radicals and elastases in children with acute KD, and was also associated with the development of coronary and myocardial disorders [12] . In a similar vein, neutrophil self-activation levels rise markedly in SJIA and then fall throughout the remission phase in accordance with the illness [33] .…”

Hypovitaminosis D and Leukocytosis to Predict Cardiovascular Abnormalities in Children with Kawasaki Disease: Insights from a Single-Center Retrospective Observational Cohort Study

“…Elevated levels of semaphorin 4D have been observed in KD patients, particularly those with CVAs, and have been correlated with increased levels of metalloproteinases and different pro-inflammatory cytokines [ 27 ]. Inter alia, patients with KD, especially those who do not respond to conventional therapies and tend to develop CVAs, show overexpression of CD177 on neutrophil cell membranes: CD177 is a marker of both neutrophil activation and degranulation and appears to be specifically associated with KD, as it significantly decreases after treatment with IVIGs [ 28 , 29 ].…”

DataSheet_1.pdf

“…TLR2 is required to activate LCWE-induced inflammatory response in a MyD88-dependent manner, activating the NF-kB signaling pathway and the production of several inflammatory cytokines, including IL-1b (34). Neutrophils activated and seem to play an essential role in KD immunopathogenesis (61,62). When neutrophils are activated, they secret enzymes and proteases, such as neutrophil elastase, to neutralize and eliminate pathogenic threats.…”

“…However, overactivation and uncontrolled recruitment can lead to tissue damage, destroying the intima and media layers of the CA wall and contributing to lesion formation. In KD, most circulating neutrophils are activated and are a major source of IL-1b (61,63). Neutrophils infiltrate the media layer of CA during the acute phase of KD (26), and KD patients with giant CAA have increased levels of calprotectin (S100A8-9), which is primarily secreted by neutrophils, years after disease onset (64).…”