Cyclic neutropenia and severe congenital neutropenia in patients with a shared ELANE mutation and paternal haplotype: evidence for phenotype determination by modifying genes.

BACKGROUND: Cyclic neutropenia (CN) and severe congenital neutropenia (SCN) are disorders of neutrophil production that differ markedly in disease severity. Mutations of the ELANE gene (the symbol recently replacing ELA2) are considered largely responsible for most cases of CN and SCN, but specific mutations are typically associated with one or the other. PROCEDURE: We performed ELANE genotyping on all individuals and paternal sperm in an SCN kindred with eight SCN progeny of a sperm donor and six different mothers. RESULTS: One patient with CN had the same S97L ELANE mutation as seven patients with the SCN phenotype. The mutant allele was detected in the donor's spermatozoa, representing 18% of the ELANE gene pool, but not in DNA from his lymphocytes, neutrophils, or buccal mucosa, indicating gonadal mosaicism. CONCLUSIONS: The coexistence of CN and SCN phenotypes in this kindred with a shared paternal haplotype strongly suggests both a role for modifying genes in determination of congenital neutropenia disease phenotypes, and the classification of CN and SCN within a spectrum of phenotypes expressing varying degrees of the same disease process.

Mosaic tetraploidy and transient GFI1 mutation in a patient with severe chronic neutropenia.

This report presents the case of a 15-year-old male with severe chronic neutropenia, leukopenia, and persistent tetraploid mosaicism in the bone marrow and peripheral blood. His father had mild neutropenia and bone marrow tetraploidy. Flow cytometric analysis of DNA content peripheral blood showed tetraploidy in 20% of granulocytes and 15% of monocytes. Sequence analysis of the ELA2 gene was normal, but the GFI1 gene exhibited transient appearance of single base changes the coding region and promoter. We speculate that an underlying genetic defect, inherited in an autosomal dominant pattern, leads to both disordered mitosis and neutropenia in this kindred.

Neutrophil elastase and granulocyte colony-stimulating factor receptor mutation analyses and leukemia evolution in severe congenital neutropenia patients belonging to the original Kostmann family in northern Sweden.

BACKGROUND AND OBJECTIVES: Severe congenital neutropenia (SCN) or Kostmann syndrome was originally reported to be an autosomal recessive disease of neutrophil production causing recurrent, life-threatening infections. Mutations in the neutrophil elastase gene (ELA-2) have previously been identified in patients with sporadic or autosomal dominant SCN. DESIGN AND METHODS: We studied 14 individuals (four patients with SCN and ten close relatives) belonging to the original Kostmann family in northern Sweden for mutations in the ELA-2 and the granulocyte colony-stimulating factor (G-CSF) receptor genes. RESULTS: One patient belonging to the original Kostmann family harbored a novel heterozygous ELA-2 mutation (g.2310T-->A;Leu92His) that was not inherited from her parents. The mutation was identified in DNA isolated from both whole blood and skin fibroblasts, suggesting a sporadic de novo mutation. As a young adult this patient sequentially acquired two mutations in the gene for the G-CSF receptor (G-CSFR) and therefore recently received a hematopoietic stem cell transplant, due to the risk of evolution to leukemia. Moreover, another patient developed acute leukemia and was treated with transplantation. No pathogenic ELA-2 or G-CSFR gene mutations were found in this patient or the other two patients, nor in any healthy relative. INTERPRETATION AND CONCLUSIONS: Our data are the first to document leukemia evolution and G-CSFR gene mutations in the original Kostmann kindred. In addition, our findings indicate that ELA-2 mutations are not the primary cause of SCN in the Swedish Kostmann family.

Periodontal status in two siblings with severe congenital neutropenia: diagnosis and mutational analysis of the cases.

BACKGROUND: Severe congenital neutropenia (SCN), also known as Kostmann syndrome, was originally reported as an autosomal recessive disease of neutrophil production. The disease is characterized by a maturation arrest of neutrophil precursors at the promyelocytic stage of differentiation and by extremely low levels of mature neutrophils in peripheral blood. METHODS: A 6-year-old male presented with a complaint of gingival swelling and bleeding, and swelling at the left side of his face. Upon clinical examination, severe inflammation of all gingival tissues was apparent, and a periapical abscess with mobility was noted on the left mandibular second molar. Medical and dental histories revealed numerous recurrent bacterial infections associated with oral and non-oral tissues. His medical history with recurrent infections led us to evaluate his 3-year-old sister to determine the status of her oral health. Inflammation of her oral tissues and recurrent bacterial infections were apparent. Their consanguineous parents were in good health. To assist in identifying possible systemic diseases underlying the inflammatory situation in the siblings, consultations were requested from the Pediatric Hematology Department at Selcuk University and Pediatric Oncology Department at Gulhane Military Medical Academy. RESULTS: Based on absolute neutrophil count (< or =200/mm(3)) and bone marrow aspiration findings consistent with early maturation arrest in myelopoiesis, the cases were diagnosed as SCN. No chromosomal abnormality was detected upon cytogenetic examination. Sequencing analysis also revealed no mutation in the neutrophil elastase or growth factor independent-1 (GFI-1) genes in these patients. Severe periodontal disease, attachment loss, and mobility for over 50% of the deciduous teeth were noted. Within 6 months, the male sibling lost all of his deciduous teeth due to periapical and periodontal infections. His sister presented with tooth mobility for all mandibular incisors. Monthly visits, including scaling, polishing, and 0.2% chlorhexidine digluconate irrigation were performed to support their oral hygiene and to avoid recurrent oral infections. We have been able to stabilize these patients' periodontal conditions during a 2-year follow-up period. CONCLUSION: This case report emphasizes the role of periodontists and pediatric dentists in the diagnosis of diseases linked with neutrophil and other systemic disorders and highlights the need to optimize the health of oral tissues with regular appointments.

Kostmann syndrome: severe congenital neutropenia associated with defective expression of Bcl-2, constitutive mitochondrial release of cytochrome c, and excessive apoptosis of myeloid progenitor cells.

Kostmann syndrome, or severe congenital neutropenia (SCN), is an autosomal recessive disorder of neutrophil production. To investigate the potential role of apoptosis in SCN, bone marrow aspirates and biopsies were obtained from 4 patients belonging to the kindred originally described by Kostmann and 1 patient with SCN of unknown inheritance. An elevated degree of apoptosis was observed in the bone marrow of these patients, and a selective decrease in B-cell lymphoma-2 (Bcl-2) expression was seen in myeloid progenitor cells. Furthermore, in vitro apoptosis of bone marrow-derived Kostmann progenitor cells was increased, and mitochondrial release of cytochrome c was detected in CD34(+) and CD33(+) progenitors from patients, but not in controls. Administration of granulocyte colony-stimulating factor (G-CSF) restored Bcl-2 expression and improved survival of myeloid progenitor cells. In addition, cytochrome c release was partially reversed upon incubation of progenitor cells with G-CSF. In sum, these studies establish a role for mitochondria-dependent apoptosis in the pathogenesis of Kostmann syndrome and yield a tentative explanation for the beneficial effect of growth factor administration in these patients.

Cellular and molecular abnormalities in severe congenital neutropenia predisposing to leukemia.

Severe congenital neutropenia (SCN) is a rare hematological disease characterized by a selective decrease in the level of circulating neutrophils in peripheral blood, maturation arrest at the promyelocyte stage of differentiation in the bone marrow, recurrent severe infections, and evolution to acute myelogenous leukemia (AML). Cellular and molecular studies of 12 SCN patients, including 5 patients that evolved to develop AML, revealed impaired proliferative characteristics and accelerated apoptosis of bone marrow progenitor cells in SCN compared with 11 healthy controls as demonstrated by flow cytometry analysis. Sequencing analysis revealed heterozygous deletion or substitution mutations in the neutrophil elastase (NE) gene in 9 of 12 patients but not in healthy controls. Expression of various NE mutants, but not normal NE, resulted in accelerated apoptosis of human promyelocytic HL-60 progenitor cells, similar to impaired survival observed in patients' cells. Bone marrow-derived primitive CD34(+) and CD33(+)/CD34(-) progenitor cells from SCN patients evolving to AML, all with mutations in the granulocyte colony-stimulating factor receptor (G-CSFR) gene, demonstrated normal cell survival, whereas more differentiated CD15(+)/CD33(-)/CD34(-) cells negative for mutant G-CSFR gene, continue to exhibit accelerated apoptosis. These data demonstrate that impaired survival of bone marrow myeloid progenitor cells, probably driven by expression of mutant NE, is the cellular mechanism responsible for neutropenia in SCN. Furthermore, our results suggest that acquired G-CSFR mutations may initiate signaling events that override the pro-apoptotic effect of mutant NE in primitive progenitor cells, resulting in an expansion of the abnormal AML clone.

Mutant elastase in pathogenesis of cyclic and severe congenital neutropenia.

Severe neutropenia is characterized by maturation arrest of myeloid cells at the promyelocyte stage of hematopoiesis. We reported that accelerated apoptosis of bone marrow myeloid progenitor cells was observed in both cyclic (CN) and severe congenital neutropenia (SCN). Short and long-term cultures of bone marrow CD34+ cells revealed reduced production of multipotent progenitors in SCN. In contrast, production of these cells was slightly elevated in CN compared with CD34+ cells from healthy volunteers. Production of myeloid-committed progenitor cells was significantly reduced in both CN and SCN. FACS analysis of CD34+ cells revealed G /G cell cycle arrest in SCN but not in CN.(0) (1) All CN patients and more than 90% of SCN patients have mutation in the neutrophil elastase (NE) gene. Molecular modeling of NE tertiary structure indicates that mutations observed in SCN are primarily located around the glycosylation sites, whereas CN mutations affect predominantly the active site. Transient expression of CN- or SCN-specific mutant NE cDNA results in impaired survival of human myeloid progenitor cells compared with control cells transfected with intact NE cDNA. We hypothesize that abnormal processing and subcellular localization of mutant NE might predetermine the etiology of cyclic or severe congenital neutropenia.