Adult onset X-linked chronic granulomatous disease in a woman patient caused by a de novo mutation in paternal-origin CYBB gene and skewed inactivation of normal maternal X chromosome.

We report a 28-year-old woman patient suffering from refractory subcutaneous abscess. Stimuli-induced microbicidal reactive oxygen metabolites formation test of the patient's neutrophils revealed that only 9.6% of the neutrophils produced H2O2. DNA analysis of the CYBB that encodes gp91(phox) demonstrated that she was heterozygous for a nonsense mutation, 206Trp(TGG)/stop(TGA) and therefore, a diagnosis of adult onset X-linked chronic granulomatous disease was made. Our molecular biological study revealed that her disease was caused by a de novo mutation in the CYBB gene on the paternal-origin X-chromosome and a skewed inactivation of the normal maternal X-chromosome.

A missense mutation of the Toll-like receptor 3 gene in a patient with influenza-associated encephalopathy.

Toll-like receptors (TLRs) recognize pathogen-associated molecular patterns and mediate the activation of NF-kappaB and the production of proinflammatory cytokines, which is critical for the innate immune system. TLR3 recognizes both double-stranded RNA and the influenza A virus. Since influenza-associated encephalopathy is frequent in Japan and East Asia and its pathological mechanism remains unknown, we analyzed several genes including TLRs and the retinoic acid inducible gene I, which could be involved in the recognition of the RNA virus. In one of three patients with influenza-associated encephalopathy, we detected a novel missense mutation (F303S) in just the TLR3 gene. This was confirmed as a loss-of-function mutant in a dose-dependent manner by NF-kappaB and IFN-beta reporter assays using wild-type and mutant TLR3-transfected HEK293 cells. Our results imply that a mutation of the TLR3 gene could be one of the factors responsible for influenza-associated encephalopathy.

Clinical evaluation of interferon-gamma treatment to chronic granulomatous disease patients with splice site mutations.

IFN-gamma dependent increase of superoxide production by neutrophils was observed in three patients with Chronic Granulomatous disease from one family. The patients have the gp91-phox defect due to a splicing abnormality derived from a silent mutation adjacent to the third intron of CYBB gene. Apparent differences of splicing pattern of CYBB gene transcripts in patients' neutrophils were detected between 1 and 25 days after administration of IFN-gamma. Furthermore, the transcript containing all missing exons could be detected in all specimens after the treatment. The changes of splicing pattern in the transcripts and prolonged effect on superoxide generating ability of patients' neutrophils indicate that IFN-gamma induced an ability to correct abnormal splicing of CYBB gene transcripts in progenitor cells at least in part.

Newly established Askin tumor cell line and overexpression of focal adhesion kinase in Ewing sarcoma family of tumors cell lines.

Askin tumor is a malignant small round cell tumor that originates from the thoracopulmonary region and is a member of Ewing sarcoma family of tumors (ESFT). Only a few Askin tumor cell lines have been established. An Askin tumor cell line, designated MP-ASKIN-SA, was established from the left thoracic tumor of a 13-year-old Japanese boy. ESFT is known to have a high rate of distant metastases at diagnosis. The genes controlling the spread of ESFT cells, however, have not been elucidated. G-banding chromosome analysis revealed that the MP-ASKIN-SA cell line has complex chromosomal abnormalities including trisomy 8. The EWS/FLI1 chimeric transcript and c-myc overexpression were revealed by the reverse transcriptase-polymerase chain reaction and Northern blot analysis. Furthermore, we investigated the expression of the focal adhesion kinase (FAK) gene in the ESFT cell lines using Northern blot analysis. In addition to the MP-ASKIN-SA cell line, six Ewing sarcoma cell lines, one peripheral nerve sheath tumor cell line, and two Askin tumor cell lines were analyzed. All ESFT cell lines, including MP-ASKIN-SA, expressed five- to twenty-eight-fold-increased values of FAK, as compared with fibroblasts obtained from the bone marrow of a healthy volunteer. These results raise the possibility that the overexpression of c-myc and FAK are involved in the poor prognosis of ESFT.