Late diagnosis of chronic granulomatous disease.

Modern era advancements in medical care, with improved treatment of infections, can result in delayed diagnosis of congenital immunodeficiencies. In this study we present a retrospective cohort of 16 patients diagnosed with Chronic Granulomatous Disease (CGD) at adulthood. Some of the patients had a milder clinical phenotype, but others had a classic phenotype with severe infectious and inflammatory complications reflecting a profoundly impaired neutrophil function. It is therefore of great importance to investigate the individual journey of each patient through different misdiagnoses and the threads which led to the correct diagnosis. Currently the recommended definitive treatment for CGD is hematopoietic stem cell transplantation (HSCT). Although survival of our patients to adulthood might argue against the need for early HSCT during infancy, we claim that the opposite is correct, as most of them grew to be severely ill and diagnosed at a stage when HSCT is debatable with potentially an unfavorable outcome. This cohort stresses the need to increase awareness of this severe congenital immunodeficiency among clinicians of different specialties who might be treating undiagnosed adult patients with CGD.

Chronic granulomatous disease of childhood: an unusual cause of recurrent uncommon infections in a 61-year-old man.

Chronic granulomatous disease (CGD) is a rare congenital immunodeficiency that affects 1 : 250,000 of the population, which is characterized by recurrent bacterial and fungal infections and by granuloma formation. We investigated a 61-year-old man presented with a 20-year history of a relapsing skin rash appearing as mildly pruritic and erythematous plaques affecting various body regions. Cutaneous biopsies were taken and sent for histology and tissue culture. Leucocyte function was assessed by determining the generation of reactive oxygen species. Bactericidal activity was assessed in the presence of autologous and homologous sera. Western blotting was performed for protein analysis of the reduced nicotinamide adenine dinucleotide phosphate oxidase system, and mutation screening was carried out using PCR amplification and sequence analysis. Examination of biopsies obtained from lesional skin indicated a suppurative granulomatous process. Tissue cultures grew Aspergillus nidulans and Aspergillus fumigatus (confirmed by PCR). A. nidulans has often been associated with CGD, and the leucocyte function tests supported this diagnosis. Direct DNA sequencing led to the identification of a hemizygous missense novel mutation in CYBB (c.907C>T), which predicts a p.His303Tyr amino-acid substitution in gp91-phox, thus confirming the diagnosis of CGD. In conclusion, we report a case of a rare inherited immunodeficiency, CGD, in a 61-year-old man, and describe the novel hemizygous missense mutation underlying the condition. Mild forms of usually fatal immunodeficiencies should be considered when assessing the occurrence of unusual infectious diseases in apparently healthy people.

Human mast cells release Interleukin-8 and induce neutrophil chemotaxis on contact with activated T cells.

BACKGROUND: Mast cells have recently been shown to control neutrophil recruitment during T-cell mediated cutaneous DTH reaction in vivo through TNF-alpha and MIP-2, the functional murine analogue of human IL-8. Although the nature of signals transmitted from T cells which activate mast cells has not yet been defined, we hypothesized that a direct cross-talk (i.e. heterotypic adhesion) between these two cell populations exists, as has previously been reported. AIMS: The present study was aimed at gaining insight into the functional role of mast cell-T cell contact in expression and release of IL-8, and its effect on neutrophil chemotaxis. METHODS: The IL-8 gene expression was identified by Affymetrix GeneChip arrays, validated by RT-PCR and the protein measured by ELISA. Chemotaxis was evaluated by using a modified Boyden chamber assay. RESULTS: Mast cells were found to express and release significantly higher concentrations of IL-8 on incubation with membranes obtained from activated, as compared to resting T cells. Supernatants obtained from these activated mast cells induced significant neutrophil chemotaxis that was inhibited by neutralizing mAb to IL-8. CONCLUSIONS: Thus, activated T cells, on heterotypic adhesion to mast cells, deliver the necessary signals for the latter to release cytokines and chemokines necessary for cell migration to sites of antigen challenge, thereby facilitating T-cell mediated inflammatory processes.

Early immunological defects in comatose patients after acute brain injury.

OBJECT: The aim of this prospective study was to evaluate the phagocytic, humoral, and cellular arms of the immune system in comatose patients shortly after severe brain injury and to compare the findings with those reported earlier in patients in a persistent vegetative state. The study was conducted in intensive care units and immunology laboratories of university-affiliated hospitals in central Israel. METHODS: The study group consisted of 14 men aged 16 to 65 years who were comatose as a result of acute brain injury due to mechanical trauma. All were studied within 72 hours of injury. Brain damage was severe in all cases (Glasgow Coma Scale score < 8). Healthy age- and sex-matched volunteers served as simultaneous controls. Infections arose in nine (75%) of the 12 patients in whom data were available; the cumulative mortality rate was 38% (five of 13 patients in whom outcome data were available). Every patient exhibited one or more defects in at least one arm of the immune system. Significant deficiencies were noted in neutrophil superoxide release, immunoglobulin (Ig)G, IgG1, IgM, C1q, C2, properdin, alternate C pathway, T cells, T helper cells, T suppressor cells, and natural killer cells. In an earlier series of patients examined by the authors months after the primary insult, these impairments were absent in most of the patients in the vegetative state. CONCLUSIONS: Significant deficiencies of the immune system, particularly the cellular arm, are precipitated by severe brain injury within 72 hours of the event. These impairments probably play a role in the high rate of complicating infections and multiple organ failure. Together with earlier findings, the results of this study indicate that if brain-injured patients survive these hazards, their immune system will eventually recover.

In vitro effect of advanced glycation end-products on human polymorphonuclear superoxide production.

BACKGROUND: Advanced glycation end-products (AGEs) are elevated in the sera of diabetic patients. The latter are prone to severe bacterial infections. Advanced glycation end-products have been shown to modulate immune competent cell activities. In this study we examined the in vitro effect of advanced glycation end-products on superoxide anion generation by human polymorphonuclear leukocytes. MATERIALS AND METHODS: Advanced glycation end-products were prepared by incubation of bovine serum albumin (BSA) with glucose for 90 days. Superoxide production was measured as the superoxide dismutase-inhibitable reduction of ferricytochrome c. The effect of advanced glycation end-products on superoxide production was evaluated in both baseline (nonstimulated) and stimulated (by either formyl-methionyl-leucyl-phenylalanine, or phorbol-myristate-acetate) polymorphonuclear leukocytes. RESULTS: The baseline superoxide production of polymorphonuclear leukocytes was significantly increased by advanced glycation end-products in a dose-dependent manner. In contrast, in stimulated polymorphonuclear leukocytes advanced glycation end-products significantly inhibited superoxide production, again in a dose-dependent manner. This inhibitory effect of advanced glycation end-products was observed after dialyzing AGE-BSA, thereby eliminating the possible influence of reactive carbohydrates. No modification of superoxide production was seen with BSA and only a mild inhibitory effect of glucose at high concentrations. CONCLUSIONS: Advanced glycation end-products depress superoxide production by stimulated polymorphonuclear leukocytes. As superoxide plays an essential role in bactericidal activity, this polymorphonuclear leukocyte dysfunction may be a contributory factor to the increased prevalence and severity of bacterial infection seen in diabetic patients.

Effect of granulocyte and granulocyte macrophage colony stimulating factors (G-CSF and GM-CSF) on neonatal neutrophil functions.

Although there are many studies on the effect of granulocyte and granulocyte-macrophage colony stimulating factors (G/GM-CSF) on adult neutrophil functions, there is little information regarding their influence on neonatal cells. We studied the in vitro effect of G/GM-CSF on neutrophil chemotaxis, polarization, and superoxide anion generation in 47 neonates compared with 35 adults. We found that G-CSF and GM-CSF significantly enhanced the chemotaxis of newborn infants' neutrophils, normalizing their chemotactic defect [from 35 +/- 7 cells/field (mean +/- SE) to 49 +/- 5 cells/field with G-CSF, p < 0.05 and to 55 +/- 4 cells/field with GM-CSF, p < 0.001]. It is notable that the maximal neutrophil response to the cytokines was observed particularly in the newborn infants with severe impairment in their chemotactic activity. Statistical analysis of the data showed a significant inverse correlation, which supported this observation (r = -0.6, p < 0.02 for G-CSF; r = -0.76, p < 0.001 for GM-CSF). The reduced polarization of neonatal compared with adult cells [71 +/- 5% versus 86 +/- 2% (mean +/- SE), p < 0.05], was corrected by CSF-priming (to 87 +/- 4% with G-CSF and to 92 +/- 2% with GM-CSF, p < 0.05). In addition, the neutrophil superoxide generation was significantly improved in both groups following the CSF-priming. GM-CSF and G-CSF gave comparable results in all functions studied except that GM-CSF improved superoxide release to a greater extent. This study shows a significant improvement of the neonatal neutrophil functions following in vitro CSF-priming and contributes to a better understanding of the neonatal neutrophil behavior when treated with G/GM-CSF.

Neutrophil function response to aerobic and anaerobic exercise in female judoka and untrained subjects.

OBJECTIVES: Recent studies have indicated reduced immunity in trained athletes. AIM: To assess the effects of aerobic and anaerobic exercise on the phagocytic process in 18-26 year old trained female judoka (n = 8) and untrained controls (n = 7). METHODS: Each subject participated randomly in two different testing sessions (aerobic, 20 minutes of treadmill running at 70-80% of maximal heart rate; anaerobic, Wingate anaerobic test). Venous blood samples were drawn before, immediately after, and 24 hours after each session. RESULTS: There were no significant differences in basal values of net chemotaxis (chemotaxis--random migration), bactericidal activity, and superoxide anion release between the judoka and the untrained women. There was a significant decrease in net chemotaxis 24 hours after the aerobic exercise in both the judoka (from 64 (19) to 39 (13) cells/field, p < 0.02) and the untrained controls (from 60 (7) to 47 (12) cells/field, p < 0.05). Bactericidal activity and superoxide anion release did not change significantly after aerobic exercise in either group. There were no significant changes in net chemotaxis, bactericidal activity, and superoxide anion release after anaerobic exercise in either the judoka or untrained women. CONCLUSIONS: The decrease in net chemotaxis after aerobic, but not after anaerobic, exercise, suggests that net chemotaxis is affected by the combination of exercise intensity and duration, and not by the exercise intensity itself. Similar effects of both exercise sessions in the judoka and the untrained women suggest that training had no effect on neutrophil function response to aerobic and anaerobic exercises.

Aspects of leukocyte function and the complement system following aerobic exercise in young female gymnasts.

Recent studies have reported reduced immunity in trained athletes. Scant information exists on changes in the immune function among trained children. The purpose of this study was to assess the effect of aerobic exercise on the phagocytic process of neutrophils and the complement system in young athletes. Subjects included prepubertal elite female gymnasts (n = 7) and untrained girls (n = 6) aged 10-12 years. Venous blood was withdrawn before, immediately post and 24 h following a 20-min run at a heart rate of 170-180 beats.min-1. Neutrophil random migration, chemotactic activity, bactericidal function and PMA/FMLP-stimulated superoxide anion release as well as various complement components were assessed. Net chemotaxis was found reduced (P < 0.05) 24 h following exercise (58 +/- 11 vs. 36 +/- 11 cells/field in gymnasts and 47 +/- 7 vs. 42 +/- 8 cells/field in untrained girls pre- and 24 h post-exercise, respectively). The basal values, as well as post-exercise values of bactericidal activity were lower (P < 0.05) in gymnasts as compared with the control group (0.8 +/- 0.3, 0.8 +/- 0.2 and 0.8 +/- 0.1 log decrease of colonies in gymnasts at pre-, immediately post-, and 24 h post-exercise, respectively and 1.1 +/- 0.1, 1.1 +/- 0.1 and 1.0 +/- 0.2 log decrease of colonies in controls, respectively). No significant effect on the bactericidal activity was observed in either group following exercise. The addition of homologous sera did not correct the bactericidal activity. PMA-stimulated superoxide anion release decreased (P < 0.05) among gymnasts immediately following exercise (5.7 +/- 0.4 vs. 4.4 +/- 1.0 mmol O2/10(6) PMN.min) and remained low 24 h later. The same trend was observed in FMLP-stimulated neutrophils but the data were not significant. Significantly decreased levels (P < 0.05) of the early complement components (C1Q, C1R) were also found following exercise (1.34 +/- 0.64 vs. 1.27 +/- 0.28 and 1.09 +/- 0.07 vs. 1.02 +/- 0.06 pre- and post-exercise in gymnasts and untrained, respectively). Furthermore, consistently lower C2 and C3 were observed in gymnasts compared with controls. Neutrophil dysfunction as well as impairment of the complement system seem to occur following exercise.

Neonatal neutrophil inflammatory responses: parallel studies of light scattering, cell polarization, chemotaxis, superoxide release, and bactericidal activity.

Neutrophil dysfunction among newborn infants, especially those born prematurely, is well recognized, but the mechanism responsible for this phenomenon is yet to be clarified. In this study, we evaluated the stimulus response coupling in neutrophils from 90 healthy newborns and 96 healthy adults in an effort to establish whether defective neonatal neutrophil function is a result of impaired signal perception or immature responsiveness. Measurement of rapid- and slow-light scattering responses (LSR) to 1 microM FMLP stimulation revealed that neonatal neutrophils have about one-half the corresponding responsiveness of adult cells (rapid-LSR: 6.1 +/- 3.1 arbitrary light intensity units vs. 12.0 +/- 2.8, P < .001; and slow-LSR: 5.0 +/- 2.5 vs. 9.1 +/- 2.0; P < .001). The same markedly reduced activity was observed in newborn neutrophil chemotaxis and bactericidal activity in comparison with adult cells. Nevertheless, low FMLP concentrations (less than 1 nM) induced no difference in cell polarization between newborn and adult neutrophils, yet at higher FMLP concentrations, the newborn revealed significantly reduced cell polarization. Our data suggest that newborn infants bear a fully functional FMLP signal perception but lack the full capacity of inflammatory responsiveness.

Leukocyte function in chronic myeloproliferative disorders.

The myeloproliferative disorders (MPD) are clonal diseases that originate from a transformed stem cell and involve all myeloid lineage. The affected cells have both proliferative and functional impairment. Therefore, we evaluated and compared neutrophil function in 31 patients with polycythemia vera (PV), idiopathic myelofibrosis (MF), chronic myeloid leukemia (CML), and essential thrombocytosis (ET). Neutrophil chemotaxis, random migration, bactericidal activity and superoxide anion release in these patients were simultaneously compared to those of 31 healthy controls. In this study, chemotactic activity was significantly impaired in patients with PV and CML as compared to controls (M+/-SE: 42 +/- 6 vs. 69+/- 5 cells/field; p<0.005 and 47+/-7 vs. 68+/- 5; p<0.05, respectively). The assessment of the bactericidal activity of neutrophils showed no impairment in most of the patients. In the CML group, the serum had a very strong "lytic" effect on bacteria, possibly due to the high levels of serum lysozyme (22 +/- 2 microgram/ml). The superoxide anion release was found to be normal in most of the patients. Nevertheless, in 25% of PV patients the superoxide production was impaired (less than 60% of the simultaneous controls). In ET most patients had normal neutrophil function. Regarding the effect of treatment, neutrophil chemotactic activity was found to be significantly reduced in the hydrea-treated patients, as compared to the non- treated patients (p<0.001) or healthy controls (<0.0001). We conclude that disturbances in neutrophil function are present in patients with various MPDs, except ET. This probably reflects abnormal maturation of ancessors of the damaged stem cells. Nevertheless, we should keep in mind that therapy itself could affect neutrophil functions. This matter should be studied more extensively. Although infections are not common in MPD disorders, they occasionally occur. It is possible that impairment in the phagocytic function contribute to the development of infections in patients with myeloproliferative disorders.

Cellular and humoral immune response to exercise among gymnasts and untrained girls.

Recent studies reported reduced immunity in athletes following exercise. Physical activity affects both cellular and humoral immune functions. Scant information exists on exercise-induced changes in the immune system among children. The purpose of the present study was to investigate the effect of aerobic exercise on several aspects of cellular and humoral functions among 10-12 year-old highly trained female gymnasts (n = 7) and untrained girls (n = 6). All girls were pre-pubertal. Venous blood samples were drawn before, immediately after and 24 h following 20 min of treadmill running (heart rate 170-180 beats.min-1). White blood cells' number rose significantly following exercise and remained elevated for 24 h. The increase in leukocyte number was due to an increase in granulocytes as well as an increase in lymphocytes and monocytes. While neutrophil count returned to basal values after 24 h, lymphocytes and monocytes number remained elevated 24 h following exercise. Exercise resulted in a significant elevation of T cell lymphocytes, T helpers, T suppressors and natural killer cells. All values returned to normal after 24 h. There were no changes in B cell lymphocytes following exercise. Exercise had no effect on serum IgA, IgM, IgE, IgG and sub-types of IgG (IgG1, IgG2, IgG3 and IgG4). No differences were observed between gymnasts and untrained girls. In summary, the exercise-induced changes in cellular and humoral immune functions among the girls were generally similar to those described in adults. Whether the transitory effects of exercise on the immune system are related to increased susceptibility to illness is still questionable.

The effect of feeding dried tomato vines to beef cattle.

Dried Tomato vines (DTV) are used as a feedstuff in some beef cattle in Israel, despite the literature citation that tomato vines contain potentially harmful steroid alkaloids. A small-scale feeding trial over 42 d examined possible deleterious effects of feeding DTV, compared with wheat straw, in beef cows. No differences in hematological values, serum parameters for body weight were seen between the 2 groups. Steroid alkaloid content of the DTV was not examined, but toxic levels of nitrates were found. The main practical hazard in feeding DTV would appear to be from their potentially high nitrate content.

Some aspects of the humoral and neutrophil functions in post-comatose nawareness patients.

Post-comatose unawareness (PCU) is one of the possible outcomes of severe brain injury. Patients with severe brain injury have an increased susceptibility to severe nosocomial infections for multifactorial reasons, including immune suppression at different levels. We studied different immunological aspects in 11 PCU patients. Impaired humoral immunity was found in 27% of them. Two patients had decreased haemolytic activity of the classical complement pathway, associated with decreased levels of the components C1q, C1r and C4. Another patient had very low levels of IgG2 and IgG4. The neutrophil killing activity was impaired in these three patients, but was completely restored with the addition of a heterologous serum, suggesting a humoral defect. Neutrophils showed normal chemotaxis and random migration, and the superoxide anion release by neutrophils was also found to be normal. Understanding the immunological events in PCU patients contributes to a better and more intensive therapeutic approach, which might accelerate the rehabilitation of these patients.

Improved chemotactic ability of neonatal polymorphonuclear cells induced by mild membrane rigidification.

Membrane lipid fluidity of peripheral blood polymorphonuclear cells (PMNs) of 24 newborn infants, 2-4 days after birth, was determined by steady-state fluorescence polarization with 1,6-diphenyl 1,3,5-hexatriene (DPH) as a probe and compared with that of PMNs from 23 adults. Measurements with intact cells, which correspond to all cellular lipid domains, did not display any statistically significant difference between PMNs of the two groups. However, application of bixinoyl glucosamine, a membrane-impermeable fluorescence quencher, revealed that the PMN plasma membrane of the newborn is about 23% more fluid than that of the adult. Total cholesterol-to-phospholipid ratio of newborn PMNs was found to be lower by about 10% than that of the adult, which could account for the difference in their plasma membrane fluidity. The possible implication of this finding for the deficit in chemotactic ability of leukocytes from newborns was tested with neonatal PMNs that have incorporated cholesteryl hemisuccinate (CHS), an efficient plasma membrane rigidifier. In all neonatal PMNs tested a mild incorporation of CHS (0.5-1 min incubation in 50 micrograms/ml dispersion) caused a significant improvement in their net chemotaxis, from an average value of 28 +/- 7 to 43 +/- 11. Longer incubations with CHS caused a gradual decrease in chemotactic ability that approached the basal level after about 5 min incubation. The net chemotaxis in adult PMNs was significantly higher than that of neonatal PMNs (72 +/- 13) and was gradually inhibited by incorporation of CHS without any initial augmentation. Based on these results it was estimated that about 27% of the chemotactic deficit of neonatal PMNs is mediated by their immature fluid membrane.