Molecular Studies of Neutropenia in Barth Syndrome.

The Barth syndrome is a rare mitochondrial disease that causes dilated cardiomyopathy, skeletal myopathy, and severe neutropenia. It has been reported that at least two patients with Barth syndrome died from septicemia attributed to neutropenia. The disease is an X-linked autosomal recessive disorder caused by mutations in the G4.5 or TAZ (tafazzin) gene, which introduce truncations, substitution, or alternative splice sites. Hematological investigations revealed normal chemotaxis, but the presence cytoplasmic vacuoles in myeloid cells and maturation arrest of neutrophil development at the myelocyte stage. Thus, a defect in neutrophil formation appears to be the primary reason for neutropenia and the susceptibility to infections in the Barth syndrome patients. Recent studies were mostly focused on cardiac, genetic and metabolic abnormalities associated with the disorder. However, the neutropenia aspect of the Barth syndrome remains unclear. The TAZ gene mutations appear to truncate the tafazzin protein likely resulting in the loss of function. Although a drosophila model of the Barth syndrome was recently reported, the cellular or mouse models of this disorder are not available yet. Thus, the link between TAZ mutations and severe neutropenia remains unknown. We hypothesized that TAZ mutations, which lead to the loss of function of tafazzin protein, trigger impaired cell survival and reduced production of neutrophils and their neutrophil precursors, thus resulting in severe neutropenia in Barth syndrome. To test this hypothesis, we attempted to knock-down the expression of the tafazzin gene in human myeloid progenitor cells using TAZ-specific shRNA. Four shRNA sequences specific to exons 4 through 7 were used for transfection of human myeloid progenitor cells that were later examined by flow cytometry and Western blot analyses. At least 2 of the shRNA constructs resulted in a substantial down-regulation in the expression level of the tafazzin protein in transfected myeloid progenitor cells as determined by Western blot. Flow cytometry analyses revealed that the knock-down of TAZ gene expression was associated with approximately 40–50% increase in proportion of apoptotic annexin-positive cells compared with cells transfected with control scrambled shRNA. These data suggest that the loss of function of TAZ gene is cytotoxic to hematopoietic cells and that severe neutropenia is due to the accelerated apoptosis of myeloid progenitor cells in patients with Barth syndrome. Further studies needed to elucidate the specific signaling pathways and to identify potentially therapeutic agents capable of controlling accelerated apoptosis of myeloid cells in Barth syndrome.