ABSTRACT
BACKGROUND: The molecular mechanism of class-switch recombination (CSR) in human subjects has not been fully elucidated. The CSR-induced mutations occurring in the switch region of the IgM gene (Smu-SHMs) in in vitro CSR-activated and in vivo switched B cells have been analyzed in mice but not in human subjects. OBJECTIVE: We sought to better characterize the molecular mechanism of CSR in human subjects. METHODS: Smu-SHMs were analyzed in vitro and in vivo by using healthy control subjects and patients with molecularly defined CSR defects. RESULTS: We found that Smu-SHMs can be induced in vitro by means of CSR activation in human subjects. We also found large amounts of Smu-SHMs in in vivo class-switched memory B cells, smaller (although significant) amounts in unswitched memory B cells, and very low amounts in naive B cells. In class-switched memory B cells a high frequency of Smu-SHMs was found throughout the Smu. In unswitched memory B cells, the Smu-SHM frequency was significantly decreased in the 5' part of the Smu. The difference between switched and unswitched B cells suggests that the extension of somatic hypermutation (SHM) to the 5' upstream region of the Smu might be associated with the effective induction of CSR. The analysis of the pattern of mutations within and outside the WRCY/RGYW (W, A/T; R, A/G; and Y, C/T) motifs, as well as the Smu-SHMs, in CD27(+) B cells from CD40 ligand (CD40L)-, activation-induced cytidine deaminase (AID)-, and uracil-DNA glycosylase (UNG)-deficient patients revealed the dependence of Smu-SHM on CD40L, AID, UNG, and the mismatch repair system in human subjects. CONCLUSION: CD40L-, AID-, UNG-, and mismatch repair system-dependent Smu-SHMs and extension to the 5' region of Smu are necessary to accomplish effective CSR in human subjects.
Subject(s)
CD40 Ligand/immunology , Cytidine Deaminase/immunology , Immunoglobulin Class Switching , Immunoglobulin M/genetics , Somatic Hypermutation, Immunoglobulin , Uracil-DNA Glycosidase/immunology , 5' Flanking Region , Adult , Amino Acid Motifs , B-Lymphocytes/immunology , CD40 Ligand/deficiency , CD40 Ligand/genetics , Child , Cytidine Deaminase/deficiency , Cytidine Deaminase/genetics , Gene Expression Regulation , Humans , Immunoglobulin M/immunology , Immunologic Memory , Infant , Male , Molecular Sequence Data , Uracil-DNA Glycosidase/deficiency , Uracil-DNA Glycosidase/geneticsABSTRACT
OBJECTIVE: To assess the feasibility of T-cell receptor excision circles (TRECs) quantification for neonatal mass screening of severe combined immunodeficiency (SCID). STUDY DESIGN: Real-time PCR based quantification of TRECs for 471 healthy control patients and 18 patients with SCID with various genetic abnormalities (IL2RG, JAK3, ADA, LIG4, RAG1) were performed, including patients with maternal T-cell engraftment (n = 4) and leaky T cells (n = 3). RESULTS: TRECs were detectable in all normal neonatal Guthrie cards (n = 326) at the levels of 10(4) to 10(5) copies/microg DNA. In contrast, TRECs were extremely low in all neonatal Guthrie cards (n = 15) and peripheral blood (n = 14) from patients with SCID, including those with maternal T-cell engraftment or leaky T cells with hypomorphic RAG1 mutations or LIG4 deficiency. There were no false-positive or negative results in this study. CONCLUSION: TRECs quantification can be used as a neonatal mass screening for patients with SCID.
Subject(s)
DNA Repair/genetics , Neonatal Screening/methods , Receptors, Antigen, T-Cell/genetics , Severe Combined Immunodeficiency/diagnosis , Severe Combined Immunodeficiency/genetics , Adolescent , Adult , Child , Child, Preschool , Feasibility Studies , Female , Humans , Infant , Infant, Newborn , Male , Middle Aged , Predictive Value of Tests , Reverse Transcriptase Polymerase Chain Reaction , Ribonuclease P/blood , Severe Combined Immunodeficiency/blood , Young AdultABSTRACT
The authors describe a patient with osteosarcoma in whom a brain abscess developed after autologous peripheral stem cell transplantation. Serologic markers of fungal infection were negative, but fungal DNA was detected in the cerebrospinal fluid (CSF) by panfungal polymerase chain reaction (PCR) assay using primers derived from fungal 18S ribosomal RNA (rRNA) genes. The sequence of PCR products on the panfungal assay was identical to the 18S rRNA genes of Aspergillus species. The combination of sequence analysis and panfungal PCR assay could be useful in the diagnosis of cerebral aspergillosis.
Subject(s)
DNA, Fungal/cerebrospinal fluid , Neuroaspergillosis/diagnosis , Polymerase Chain Reaction/methods , Adolescent , Base Sequence , Female , Humans , Molecular Diagnostic Techniques , Molecular Sequence Data , Osteosarcoma/complications , Osteosarcoma/therapy , Peripheral Blood Stem Cell Transplantation/adverse effects , Sequence Analysis, DNA , Transplantation, AutologousABSTRACT
We report the MRI findings of a 2-year-old boy with recurrent herpes simplex encephalitis (HSE). At the age of 14 months, the patient developed a high fever that lasted over 1 week and he did not receive appropriate treatment. At 6 months after the fever, MRI showed marked atrophic changes in both deep temporal lobes with hyperintensity in the hippocampi and parahippocampal gyri. Thirteen months after the first episode of the fever, the patient was diagnosed with recurrent HSE by polymerase chain reaction assay of the CSF; MRI at this time revealed diffuse cortical swelling. Hyperintensity on T2-weighted images was noted in the occipito-parietal cortex bilaterally, the left thalamus, the subcortical white matter and the splenium of the corpus callosum. Recurrence of HSE may be more common in infants than previously thought. It is important to consider the possibility of recurrent HSE and to understand that MRI findings in HSV1 encephalitis in infants and young children appear to differ from those observed in neonates, older children and adults.
