Defects of B-cell terminal differentiation in patients with type-1 Kabuki syndrome

Kabuki syndrome (KS) is a complex multi-system developmental disorder associated with mutation of genes encoding histone-modifying proteins. In addition to craniofacial, intellectual, and cardiac defects, KS is also characterized by humoral immune deficiency and autoimmune disease, yet no detailed molecular characterization of the KS-associated immune phenotype has previously been reported

Hypohidrotic Ectodermal Dysplasia and Immunodeficiency with Coincident NEMO and EDA Mutations

Ectodermal dysplasias (ED) are uncommon genetic disorders resulting in abnormalities in ectodermally derived structures. Many ED-associated genes have been described, of which ectodysplasin-A (EDA) is one of the more common. The NF-κB essential modulator (NEMO encoded by the IKBKG gene) is unique in that mutations result in severe humoral and cellular immunologic defects in addition to ED. We describe three unrelated kindreds with defects in both EDA and IKBKG resulting from X-chromosome crossover. This demonstrates the importance of thorough immunologic consideration of patients with ED even when an EDA etiology is confirmed, and raises the possibility of a specific phenotype arising from coincident mutations in EDA and IKBKG

Aberrant maturation of mutant perforin underlies the clinical diversity of hemophagocytic lymphohistiocytosis

Missense mutations in perforin, a critical effector of lymphocyte cytotoxicity, lead to a spectrum of diseases, from familial hemophagocytic lymphohistiocytosis to an increased risk of tumorigenesis. Understanding of the impact of mutations has been limited by an inability to express human perforin in vitro. We have shown, for the first time to our knowledge, that recombinant human perforin is expressed, processed appropriately, and functional in rat basophilic leukemia (RBL) cells following retroviral transduction. Subsequently, we have addressed how perforin missense mutations lead to absent perforin detection and impaired cytotoxicity by analyzing 21 missense mutations by flow cytometry, immunohistochemistry, and immunoblot. We identified perforin missense mutations with partial maturation (class 1), no apparent proteolytic maturation (class 2), and no recognizable forms of perforin (class 3). Class 1 mutations exhibit lytic function when expressed in RBL cells and are associated with residual protein detection and variable cytotoxic function in affected individuals, suggesting that carriers of class 1 alleles may exhibit more subtle immune defects. By contrast, class 3 mutations cause severely diminished perforin detection and cytotoxicity, while class 2 mutations have an intermediate phenotype. Thus, the pathologic mechanism of perforin missense mutation likely involves a protein dosage effect of the mature protein

Elevated Granzyme B in cytotoxic lymphocytes is a signature of immune activation in hemophagocytic lymphohistiocytosis

Patients with hemophagocytic lymphohistiocytosis (HLH) exhibit immune hyper-activation as a consequence of genetic defects in secretory granule proteins of cytotoxic T lymphocytes (CTL) and natural killer (NK) cells. Murine models of HLH demonstrate significant activation of CTL as central to the disease pathogenesis, but evaluation of CTL and NK activation in children with HLH or inflammatory conditions is not well described. CD8 T cells only express granzyme B (GrB) following stimulation and differentiation into CTL; therefore, we reasoned that GrB expression may serve as a signature of CTL activation. It is unknown whether human NK cells are similarly activated in vivo, as marked by increased granule proteins. Perforin and GrB levels are measured in both CTL and NK cells by flow cytometry to diagnose perforin deficiency. We retrospectively compared GrB expression in blood samples from 130 children with clinically suspected and/or genetically defined HLH to age-matched controls. As predicted, CD8 expressing GrB cells were increased in HLH, regardless of genetic etiology. Remarkably, the GrB protein content also increased in NK cells in patients with HLH and decreased following immunosuppressive therapy. This suggests that in vivo activation of NK cells occurs in hyper-inflammatory conditions. We conclude that increased detection of GrB in CTL and NK are an immune signature for lymphocyte activation in HLH, irrespective of genetic subtype and may also be a useful measure of immune activation in other related conditions