Loss of B Cells in Patients with Heterozygous Mutations in IKAROS.

BACKGROUND: Common variable immunodeficiency (CVID) is characterized by late-onset hypogammaglobulinemia in the absence of predisposing factors. The genetic cause is unknown in the majority of cases, and less than 10% of patients have a family history of the disease. Most patients have normal numbers of B cells but lack plasma cells. METHODS: We used whole-exome sequencing and array-based comparative genomic hybridization to evaluate a subset of patients with CVID and low B-cell numbers. Mutant proteins were analyzed for DNA binding with the use of an electrophoretic mobility-shift assay (EMSA) and confocal microscopy. Flow cytometry was used to analyze peripheral-blood lymphocytes and bone marrow aspirates. RESULTS: Six different heterozygous mutations in IKZF1, the gene encoding the transcription factor IKAROS, were identified in 29 persons from six families. In two families, the mutation was a de novo event in the proband. All the mutations, four amino acid substitutions, an intragenic deletion, and a 4.7-Mb multigene deletion involved the DNA-binding domain of IKAROS. The proteins bearing missense mutations failed to bind target DNA sequences on EMSA and confocal microscopy; however, they did not inhibit the binding of wild-type IKAROS. Studies in family members showed progressive loss of B cells and serum immunoglobulins. Bone marrow aspirates in two patients had markedly decreased early B-cell precursors, but plasma cells were present. Acute lymphoblastic leukemia developed in 2 of the 29 patients. CONCLUSIONS: Heterozygous mutations in the transcription factor IKAROS caused an autosomal dominant form of CVID that is associated with a striking decrease in B-cell numbers. (Funded by the National Institutes of Health and others.).

Asthma: best treatment options.

LEARNING OBJECTIVES: To understand the principles of asthma management in patients of different ages, and to outline major avoidance measures, classes of medications available, and indications for immunotherapy (IT). DATA SOURCES: MEDLINE and major textbooks of allergy and immunology. CONCLUSIONS: There has been an understanding of asthma as a chronic inflammatory disease for many decades but recent information about mediators and cytokines has led to new therapies and better understanding of the effects of IT and other preventive measures. The best treatment for any individual patient depends upon many factors and is a decision to be made between the patient and the physician. It seems that the best response to IT occurs in those who are highly sensitive to unavoidable allergens, and who experienced a late-phase asthmatic response to the allergen initially. New data suggest that IT should be considered early, as it may prevent progression of asthma to more severe, less reversible disease. The best therapy results in the ablation of airway early- and late-phase reactivity.

Immunoglobulin deficiency in children with a sudden overwhelming infection.

BACKGROUND: A small percentage of previously healthy children develop a sudden overwhelming infection (SOI) that rapidly progresses and results in shock and, occasionally, death. Some of these children may have an undetected Ig deficiency. OBJECTIVE: The aim of this study was to evaluate the incidence of Ig deficiency in children with a SOI. METHODS: A case series study was conducted in a university hospital and included 18 children who either died in the emergency room or required admission to the pediatric intensive care unit secondary to a SOI. Two age-matched control groups included children hospitalized to regular floor beds with an infectious process (infected control group) or a noninfectious process (noninfected control group). Serum left from the initial blood draw, before fluid resuscitation, was collected and stored at -70 degrees C. Total IgG, IgG subclasses, IgM, and IgA were assayed by rate nephelometry in a blinded fashion. RESULTS: In the study group, one of six children under 1 year of age had low Ig levels in comparison with two of nine control patients. In those children over the age of 1 year, 8 of 12 patients (67%) had low Ig levels as compared with 2 of 19 controls (11%) by Fisher's exact test, P = .002. Of those patients with Ig deficiencies, three of eight had isolated IgG deficiency, two of eight had combined IgG and IgA deficiency, three of eight had combined IgG and IgM deficiency. CONCLUSIONS: Children over the age of 1 year who present with a SOI have a significantly higher incidence of Ig deficiencies compared with age-matched controls.

Bacteremia and skin/bone infections in two patients with X-linked agammaglobulinemia caused by an unusual organism related to Flexispira/Helicobacter species.

Two patients with Bruton's X-linked agammaglobulinemia are described with bacteremia and skin/bone infection due to an organism which by 16S rRNA gene sequence analysis was most closely related to "Flexispira" rappini (and thus designated a Flexispira-like organism, FLO) and more distantly related to the Helicobacter species. The organism required microaerobic conditions and, supplemental H(2) gas for growth and was reliably stained with acridine orange. In common with Helicobacter cinaedi infections, the focus of the FLO infection was in one case in the blood vessels or lymphatics of an extremity and in the other case in the skin and adjacent bone of an extremity. In both cases, prolonged IV antibiotic therapy was necessary to clear the infection. The susceptibility of XLA patients to FLO infection appears to be related to the fact that XLA is associated with severe B cell (humoral) immunodeficiency and thus these patients have difficulty with intravascular or intralymphatic infection. These findings elucidate the nature of FLO infections in humans and point the way to their detection and treatment.

Recurrent bacteremia caused by a "Flexispira"-like organism in a patient with X-linked (Bruton's) agammaglobulinemia.

Helicobacter spp., except for Helicobacter cinaedi, have only rarely been reported in cases of septicemia. A patient with X-linked (Bruton's) agammaglobulinemia was found to have persistent sepsis with a Helicobacter-like organism despite multiple courses of antibiotics. His periods of sepsis were associated with leg swelling thought to be consistent with cellulitis. The organism was fastidious and required a microaerophilic environment containing H(2) for growth. Optimal growth was observed at 35 to 37 degrees C on sheep blood, CDC anaerobe, and Bordet-Gengou agars. Serial subcultures every 4 to 5 days were required to maintain viability. The organism was strongly urease positive and showed highest relatedness to Helicobacter-like organisms with the vernacular name "Flexispira rappini" by 16S rRNA gene sequence analysis. Genomic DNA hybridization studies, however, found 24 to 37% relatedness to "F. rappini" and even less to other Helicobacter spp. Although the organism phenotypically resembles "Flexispira" and Helicobacter, it is thought to represent a new taxon. The patient's infection was eventually cleared with a prolonged (5-month) course of intravenous imipenem and gentamicin.

Quantitative DNA ploidy analysis of breast carcinoma: a study of the effects of joint photographer experts group (JPEG) compression on DNA ploidy images.

Telepathology usage in the past has typically been a qualitative procedure rather than a quantitative measurement. DNA ploidy using image analysis has been favorably compared to DNA ploidy analysis by flow cytometry in numerous publications. A step from DNA ploidy analysis using conventional image analysis to DNA ploidy analysis using stored images allows DNA ploidy analysis by image cytometry to become a powerful tool in telepathology. Remote DNA ploidy analysis using stored images has an impact on the field of pathology, as not every hospital or laboratory can afford to perform this type of specialized testing. However, images have large data files and require lengthy transmission times over communication systems to other computers. Joint Photographer Experts Group (JPEG) compression is a computer algorithm that allows the file size of an image to be reduced in order to decrease transmission times to another computer. A study was initiated to investigate the effects of JPEG compression on images of Feulgen stained breast tumor touch preps and the resulting DNA ploidy histograms.

The design and use of a computer-based digital image acquisition, management, and communications system for conferencing in pathology.

The Roche ImageManager is a PC computer-based system, running under Windows 3.1. This system allows the storage, viewing, printing, and transmission of, and conferencing on large sets of high resolution, color images from various sources. Images may be acquired at resolutions up to 3072 x 2320 pixels under software control, and transmitted for conferencing or stored in the database using JPEG compression. Each image is stored with associated information, text notes, annotations, and a "thumbnail" image. The stored information may be searched for matching images, for example representative images of a particular disease for training, or images to be reviewed for a particular patient. Images may be transmitted with all associated information between sites which have telephone service, worldwide. Support is being added for LAN use with a multi-user database, ISDN and Switch56 communications, and remote microscope control. Interactive conferencing allows rapid transmission of low resolution images for specimen scanning, viewing of multiple high resolution still images, and real-time interactive pointers for detailed discussions of image features. The ImageManager is being used and evaluated for conferencing and training purposes. A study was performed by Roche Biomedical Laboratories to compare the ability of pathologists to make a diagnosis on complex cases using transmitted images vs. routine microscopic examination. The effectiveness of the system in providing images of a sufficient quality to allow diagnosis in a private practice setting will be presented.

Quantitative DNA analysis: a comparison of conventional DNA ploidy analysis and teleploidy.

JPEG compression can be used on images for DNA ploidy analysis if careful consideration is given to the level of compression used for file storage. The amount of JPEG compression possible may vary depending on the type of tissue analyzed, however, a compromise may be reached for all types of tissue. JPEG compression should not go over a level of 70 for DNA analysis as this would result in possibly erroneous IOD calculation and erroneous DNA ploidy analysis. Also, the resulting file quality is so poor that even visual analysis is not possible. With careful training of personnel in cell selection, remote DNA ploidy analysis would be an effective tool for standardization and quality control in the pathology laboratory. By using remote DNA ploidy analysis, it is possible for hospitals to consolidate their workload, and make DNA ploidy analysis by image cytometry a cost effective test in the laboratory. Proficiency testing would also become possible as all laboratories performing DNA ploidy analysis would receive the same fields of view for testing. DNA ploidy analysis by image cytometry using stored images could be a versatile tool for the telepathology community.

Detection of Epstein-Barr virus and cytomegalovirus in patients with chronic fatigue.

Patients with chronic fatigue as a major complaint frequently present with recurrent sore throat, and on physical examination they have hyperemia and lymphoid hyperplasia of the pharyngeal area. Pharyngeal scrapings were obtained from 41 such patients and analyzed for Epstein-Barr virus or cytomegalovirus DNA by colorimetric in situ hybridization. Results were compared with healthy control subjects matched for age and sex. Epstein-Barr virus-DNA was detected more frequently in male patients, 5/9 (55.6%), than controls, 0/6 (0%), but there was no difference in frequency in female patients, 4/32 (12.5%), than control subjects, 1/29 (3.4%). Cytomegalovirus-DNA was detected infrequently in patients and controls, 13% versus 22% respectively. The presence of EBV-DNA did not correlate with antibody titers nor with the complaint of sore throat. Four of the five males who had positive EBV-DNA in the pharyngeal smears have now recovered.

Cerebral lateralization and histamine skin test asymmetries in humans.

Although asymmetries exist in both cerebral and somatic systems, their relationship is not well understood. One hypothesis suggests that some processes determining cerebral hemispheric dominance also affect immune systems. We employed histamine, a final common mediator for the immediate hypersensitivity immune response, to examine the relationship of skin response to cerebral lateralization (ie, left versus right brain dominance). Histamine skin wheal responses on the left and right forearms were measured using computerized planimetry in 176 consecutive patients undergoing allergy skin testing. Cerebral lateralization was determined by personal and family handedness via questionnaires. Subjects were classified as right handed (RH) with only RH relatives, RH with non-RH relatives, or non-RH (ie, left-handed or ambidextrous). Left/right arm asymmetries in wheal area were demonstrated and were related to cerebral dominance, but could not be explained by simple arm use. Right-handed subjects with only RH relatives had greater wheal size on the right arm, RH subjects with non-RH relatives had no significant left/right difference in wheal size, and non-RH subjects had greater wheal size on the left arm. Further, gender differences in these asymmetries appeared to be present for subjects with mixed cerebral dominance. The results suggest an association between process affecting cerebral and somatic asymmetries, and offer new insight into brain/body interactions.