Investigating the role of CD56⁻ ‘atypical’ NK cells in Gambian children with asymptomatic malaria

Abstract

Malaria is a parasitic disease responsible for over 600,000 deaths a year, caused by Plasmodium parasites. In malaria-endemic regions, individuals endure repeated exposure to and are infected with the parasite throughout their lives. Despite this high exposure level, complete parasitic clearance from the bloodstream, known as sterile immunity, is rarely achieved. Instead, individuals from highly endemic regions develop ‘clinical immunity’, where they remain parasitemic but do not experience symptoms. This acquired tolerance is associated with afebrile infections and reflects a host immune system that controls parasitemia without triggering overt disease. While anti-malarial antibodies are recognized as a critical component of this protection, the contribution of cellular immune responses remains poorly understood.

Natural Killer (NK) cells are a diverse group of innate lymphoid cells with well-documented roles in early defense against infections. They exert cytotoxic effects on infected cells and can enhance immune clearance through antibody-dependent mechanisms. In malaria, NK cells have been shown to interact with malaria-specific antibodies, resulting in malaria killing and inhibition of parasite growth. Additionally, NK cells have been shown to undergo functional adaptation in malaria infections. Phenotypic and epigenetic changes characterize these ‘adaptive’ NK cells and are more prevalent in clinically immune and malaria-resistant immune profiles. Shifts in NK cell subpopulations, including expansions of specific populations, have been associated with reduced disease severity and symptomology in malaria infection. Among these, CD56- NK cells, considered ‘atypical’ NK cells due to a lack of canonical CD56 expression, have shown enhanced cytotoxic potential against malaria parasites and may contribute to anti-malarial immunity.

Although CD56- NK cells are less characterized, emerging evidence suggests they possess enhanced cytotoxic potential, particularly in antibody-dependent cellular cytotoxic (ADCC) pathways. As malaria-specific antibodies are strongly correlated with clinical immunity, these atypical NK cells may serve as an important effector cell population in antibody-mediated parasite clearance. We hypothesize that repeated malaria exposure drives the expansion and functional adaptation of CD56⁻ NK cells, giving them increased cytotoxic capabilities that contribute to parasite suppression and the prevention of symptomatic disease.

However, the role of atypical NK cells in clinical immunity to malaria is poorly defined, as little research has been done in this area. CD56⁻ NK cells have been classified by increased antibody-dependent cytotoxic ability, and anti-malaria antibodies have been closely linked to clinical immunity. I hypothesize that these atypical NK cells experience expansion and changes that lead to an adaptive-like phenotype in malaria infection, enabling more effective parasite killing.

This study aims to phenotype peripheral blood mononuclear cells (PBMCs) samples from afebrile, malaria-infected, and uninfected children from the Upper River Region of The Gambia using cytometric analysis. This approach will allow us to investigate the phenotype and potential functionality of CD56⁻ NK cells in the context of asymptomatic infection. Cytometry involves staining cells with antibodies conjugated to fluorophores, enabling the detection of surface and intracellular proteins. The cytometer will quantify the signals to determine the presence and intensity of marker expression.

Optimization of the cytometry panel and staining protocol is critical for accurate detection, particularly for rarer cell populations such as CD56⁻ NK cells. This involves carefully selecting fluorochromes, antibody titrations, alterations to staining protocols, and calibration of cytometer settings to ensure sensitivity, specificity, and reproducibility. Findings from this study will help clarify the contribution of atypical NK cells to clinical immunity.