Neutrophil dysfunction in a family with a SAPHO syndrome-like phenotype

Abstract

SAPHO syndrome (synovitis, acne, pustulosis, hyperostosis, osteitis) is an inflammatory disorder of the bone, skin, and joints. We describe a family with multiple affected members who segregate a SAPHO syndrome-like phenotype, and we report the results of neutrophil studies and candidate gene analysis. We obtained written informed consent and a family history and reviewed medical records. We collected DNA and sequenced candidate genes, and we performed functional studies on neutrophils isolated from the proband and her mother. The pedigree segregated chronic osteomyelitis and cutaneous inflammation in a pattern that suggested an autosomal-dominant disorder. No coding sequence mutations were detected in PSTPIP1, PSTPIP2, LPIN2, SH3BP2, or NCF4. Analysis of neutrophil function in the proband, including nitroblue tetrazolium tests, myeloperoxidase assays, neutrophil chemotaxis, and neutrophil chemotaxis assays, revealed no identifiable abnormalities. However, an abnormality in the luminol, but not the isoluminol, respiratory burst assays following stimulation with phorbol myristate acetate (PMA) was detected in neutrophils isolated from the affected proband. Internal oxidant production was also reduced in the proband and her mother when neutrophils were treated with fMLP with or without platelet-activating factor, PMA alone, or tumor necrosis factor alpha alone. This family segregates a disorder characterized by chronic inflammation of the skin and bone. Functional differences in neutrophils exist between affected individuals and controls. The biologic significance of this defect remains unknown. Identification of the gene defect will help identify an immunologic pathway that, when dysregulated, causes inflammation of the skin and bone.

Figure 1.

Pedigree. The arrow points to the proband. Solid symbols denote individuals with both chronic multifocal osteomyelitis and skin inflammation. Hatched symbols denote individuals with skin inflammation. Gray symbols denote individuals with other inflammatory disorders (see last paragraph of Case Reports).

Figure 2.

Respiratory burst in patient and control neutrophils in response to increasing doses of phorbol myristate acetate (PMA). The luminol and isoluminol assays can be used to differentiate extracellular oxidant production and intracellular oxidant production by primary neutrophils. Primary neutrophils were purified using Polymorphprep and treated with increasing doses of PMA (3.1 ng/ml, 6 ng/ml, and 31 ng/ml). A and B, Area under the burst curve for internal and external oxidant production by the luminol assay (A) or isoluminol assay (B) in patient and control neutrophils at varying doses of PMA. Values are the mean ± SEM of 4 experiments. * = P < 0.05 versus control neutrophils, by analysis of variance. C, Representative burst curve for both luminol and isoluminol after stimulation with 31 ng/ml of PMA in patient and control neutrophils. Control neutrophils isolated from healthy donors exhibit robust oxidant production both internally (luminol) and externally (isoluminol), while patient neutrophils exhibit a defect in only internal oxidant production. Shown in C are the results of 1 experiment, representative of >4 experiments performed.

Figure 3.

Flow cytometry analysis of oxidant production in neutrophils. Shown is the relative oxidative burst in neutrophils isolated from the patient and from her mother and in control neutrophils treated with fMLP, platelet-activating factor (PAF) plus fMLP, phorbol myristate acetate (PMA), or tumor necrosis factor α (TNFα). Data are the results from 3 experiments with the patient’s neutrophils and 1 experiment with neutrophils isolated from the patient’s mother. Values are the mean ± SEM of 3 experiments for the patient. Neutrophils from both the patient and her mother showed reduced oxidative burst compared with control neutrophils (P < 0.05 by analysis of variance). un = unstimulated neutrophils.