Seroprevalence of IgG antibodies against Bordetella pertussis in healthy individuals aged 4-24 years in Turkey.

The distribution of IgG antibodies to Bordetella pertussis was investigated in serum samples from 550 subjects, aged 4-24 years, to determine the optimal age for booster immunisation. Levels of antibody to B. pertussis antigens were determined using an ELISA that measures a mixture of pertussis toxin, filamentous haemagglutinin and lipopolysaccharide. Geometric mean titres of anti-pertussis antibodies in subjects aged 4-6 years were significantly lower than those in other age groups, which reflects waning immunity following vaccination. High positive titres in older children and adolescents suggested acquired B. pertussis infection, and booster doses at the ages of 7 and 15 years are therefore suggested.

Evaluation of human antibody responses to diphtheria toxin subunits A and B in various age groups.

This study aimed to evaluate human antibody responses to diphtheria toxin subunits in various age groups. Antibodies against the intact diphtheria toxin and the diphtheria toxin subunits A and B were evaluated in 1319 individuals using a double-antigen ELISA. Although high levels of protection (83.6%, 95% CI 79.2-87.4) were found in children and adolescents, the middle-aged adult population was less protected (28.8%, 95% CI 24.3-33.6). An increase in age was associated with a decrease in the frequency of protected individuals in the 0-39-year age group (p <0.001). Anti-subunit B levels correlated well (p <0.01) with levels of antibodies against the intact toxin. In children aged < or =16 years, the intervals at which the peaks in geometric mean titres of anti-subunit B antibodies were observed were found to correlate with the ages at which booster doses are administered. Overall, males appeared to be more protected than females (OR 1.67, 95% CI 1.34-2.08, p <0.001). A small group of individuals had antibody levels of > or =0.1 IU/mL against the intact toxin, but did not have protective antibody against subunit B. Determination of anti-subunit B antibody levels should help in evaluating the effectiveness of diphtheria boosters and other aspects of diphtheria immunity.

Serum levels of sL-selectin and tumour necrosis factor-alpha in children with type 1 diabetes mellitus.

Type 1 diabetes mellitus (DM) is a result of inflammation and destruction of alpha-cells in the pancreatic islet cells. The aim of this study is to evaluate the associations of diabetes with soluble L-selectin (sL-selectin) and tumour necrosis factor-alpha (TNF-alpha) in children with type 1 DM; and also to evaluate the associations of these parameters with the disease period, glycaemic control state and puberty stage. Serum sL-selectin and TNF-alpha levels were measured in 44 children with type 1 DM and 44 healthy children. Neither the patients nor the control group showed significant difference between the levels of sL-selectin and TNF-alpha (sequence mean 12.17+/-1.62 ng/ml vs. 12.62+/-1.56 ng/ml and 7.27+/-3.1 pg/ml vs. 7.88+/-2.7 pg/ml). There was no statistically significant difference between children with duration of diabetes longer than 5 years and children with duration of diabetes shorter than 1 year. There was also no statistically significant difference between poor glycaemic control and good-acceptable glycaemic control patients. The present results indicate that sL-selectin and TNF-alpha serum levels are not increased and cannot be used as prognostic predictors in type 1 DM; and also sL-selectin and TNF-alpha do not change with the disease period, glycaemic control state and puberty stage.

Development of a rapid, single-step procedure using protein G affinity chromatography to deplete fetal calf serum of its IgG and to isolate murine IgG1 monoclonal antibodies from supernatants of hybridoma cells.

Fetal calf serum (FCS) was depleted of its immunoglobulin G (IgG) in a rapid procedure using protein G affinity chromatography. 20 ml of FCS was depleted of its IgG in less than 80 min by applying 5 ml of FCS to a 1 ml HiTrap protein G Sepharose column followed by appropriate elution. Various concentrations of IgG-depleted FCS (G-FCS) were used in RPMI-1640 medium to grow the mouse hybridoma cell lines CAy-G (anti-HBs IgG1 mAb producing hybridoma cell) and CAy-M (anti-HBs IgM mAb producing hybridoma cell), which secreted hepatitis B virus surface antigen (HBsAg)-reactive IgG1 and IgM monoclonal antibodies (mAbs), respectively. Antibody production and cell growth were used as indices to compare the efficacy of RPMI/G-FCS with that of RPMI/FCS and serum/protein-free Hybri Max (Sigma, MO, USA) hybridoma medium. MAb production and cell growth of CAy-G and CAy-M hybridoma cell lines in RPMI/G-FCS were similar to culture in RPMI/FCS and significantly better than culture in Hybri Max. We found that G-FCS was superior to whole FCS as a culture supplement for the purification of IgG1 mAbs. IgG1 mAbs were isolated in a single-step procedure using protein G affinity chromatography, from the supernatant of CAy-G hybridoma cells cultured in RPMI/10% G-FCS (RPMI-1640 medium supplemented with 10% G-FCS). SDS-PAGE analysis revealed that the purity of IgG isolated from the supernatant of CAy-G cells cultured in RPMI/10% G-FCS was more than 99%.

Alpha interferon treatment in myasthenia gravis: effects on natural killer cell activity.

The efficacy of recombinant interferon-alpha (rIFN alpha), on natural killer (NK) cell cytotoxic activity, CD3+, CD4+, CD8+, CD56+, HLA-DR+ lymphocyte counts, anti-acetylcholine receptor antibody (AChR Ab) levels, single fibre electromyography findings (SFEMG) and clinical course were evaluated in patients with myasthenia gravis (MG). During the IFN alpha treatment (3 mu, subcutaneous, 3 times a week), NK cell cytotoxicity and CD4+/8+ ratio increased, NK cell count remarkably decreased, and no significant clinical or SFEMG changes were observed. This preliminary open study in MG patients has demonstrated enhanced NK activity per unit NK cell after IFN alpha therapy. Although lymphocyte phenotypes and NK function approached normal levels during therapy, a higher dose of IFN alpha may be required for a significant clinical response. It has been also concluded that 6 months of IFN alpha therapy seems to be safe in MG, though in patients with malignancy, IFN alpha may cause increased autoimmunity, AChR positivity and MG.

Tumor necrosis factor (TNF) induction from monocyte/macrophages by Candida species.

Candida albicans was studied for its capacity to induce TNF production from mouse peritoneal macrophages (PM phi). TNF activities in the culture supernatants of Candida-stimulated PM phi and human peripheral blood monocytes were assessed by L 929 bioassay and ELISA respectively. C. albicans induced TNF production from PM phi and human peripheral blood monocytes in a dose-dependent manner. Although the capacity was lesser than live form, heat-killed C. albicans was also found to be capable of stimulating PM phi to induce TNF. The filtered supernatant of 24 h cultured live C. albicans had no effects on TNF production from PM phi. Saccharomyces cerevisiae-extracted mannan, a yeast cell wall antigen, induced TNF production from PM phi in a dose-dependent manner. Thus, the effect of C. albicans on TNF production from PM phi was seemed to be directly related to the presence of the yeast cell wall itself. Compatible with these data, when various candida species (C. albicans, C. tropicalis, C. pseudotropicalis. C. lusitaniae, C. krusei, C. parapsilosis, C. guilliermondii, C. stellatoidea, C. glabrata) and S. cerevisiae were compared to each other, at a concentration of 2 x 10(6) yeast cells/ml from each species, it was observed that TNF inducing capacities varied. Among the species used in this study, C. guilliermondii and C. glabrata, of which the yeast cell size were the smallest ones, were found to be less potent than that of others to induce TNF from PM phi.

Comparison of the effects of Salmonella minnesota Re595 lipopolysaccharide, lipid A and monophosphoryl lipid A on nitric oxide, TNF-alpha, and IL-6 induction from RAW 264.7 macrophages.

Lipopolysaccharide (LPS) exhibits a wide variety of bioactivities. Although it was generally proposed that the lipid A component represented the active center responsible for most of the bioactivities of LPS, a variety of lipid A partial structures and analogues were reported to have different properties. Lipopolysaccharide of the Re595 mutant of Salmonella minnesota is lack of O and part of the core polysaccharide (2 keto-3-deoxyoctanate (KDO) left on lipid A). Re595 lipid A (LA) and Re595 monophosphoryl lipid A (MPLA) differ in structure from Re595 LPS by lacking KDO and KDO plus phosphoryl group respectively. Whether these lipid A-common Re595 LPS preparations differed in activities, we investigated their effects on nitric oxide (NO), TNF-alpha, IL-6, and IL-12 induction from murine macrophage cell line RAW 264.7. RAW 264.7 cells (2 x 10(5) cells ml(-1)) were stimulated with these LPS preparations at 1 microg ml(-1) for 48 h. Re595 LPS, Re595 LA and Re595 MPLA significantly induced NO, TNF-alpha and IL-6 production; NO, TNF-alpha and IL-6 inducing capacities were in the order of LPS = LA > MPLA, LPS = LA = MPLA, and LPS = LA > MPLA respectively. However, these preparations did not induce IL-12 production from RAW cells even when stimulated in combination with IFN-gamma (20 U ml(-1)). IFN-gamma itself also induced NO, TNF-alpha and IL-6 production from RAW 264.7 cells. When RAW 264.7 cells were stimulated with IFN-gamma plus any of these preparations, effects were additive and synergistic for NO and IL-6 responses respectively. But TNF-alpha responses of RAW cells against these preparations were almost equal when cultured alone or in combination with IFN-gamma. Pre-treatment of RAW cells either with LPS, LA or MPLA at low concentration (0.1 microg ml(-1)) for 60 min before pulsing with IFN-gamma (20 IU ml(-1)) plus LPS (1 microg ml(-1)) for an additional 48 h, significantly (P < 0.01) decreased NO response. Although to a lesser extent, TNF-alpha and IL-6 responses were also decreased. Complete inhibition of NO inducing effect of these LPS preparations was achieved with polymyxin B at 40 microg ml(-1). But the concentration of polymyxin B to get a significant (P < 0.05) inhibitory effect on LPS was four times higher than that for LA or MPLA. Unexpectedly, polymyxin B also inhibited INF-gamma-induced NO production from RAW cells in a dose-dependent fashion. These findings suggested that effect of LPS was dependent, at least in part, on both the LPS polysaccharide chain length and the hydrophilic portion of LPS. In addition, not only LPS but also LA and MPLA exert either enhancing or suppressive effects, depending on their concentrations and the timing of their addition with respect to co-stimulators.

Effect of allergen specific immunotherapy (IT) on natural killer cell activity (NK), IgE, IFN-gamma levels and clinical response in patients with allergic rhinitis and asthma.

Allergen specific immunotherapy (IT) has been widely used for many years as a specific treatment of allergic diseases. A variety of changes in immunological parameters have been described but it still remains uncertain as to which of them is responsible for the improvement of symptoms. The aim of our study was to evaluate the effect of IT on natural killer (NK) cell activity, IL-4, IFN-gamma, IgE levels and skin test reactivity in addition to clinical efficacy. Thirty-one patients with allergic rhinitis and asthma were selected according to positive history, skin prick tests to Dermatophagoides pteronyssinus (Der p) or grass pollens, presence of specific IgE antibodies in sera and clinical findings, and were submitted to one year of placebo-controlled IT. Total IgE, specific IgE, IL-4 and IFN-gamma levels were measured by using ELISA method. Standard chromium 51 release assay was used to measure NK cell cytotoxic activity against the human leukemic cell line, K562 target cells. Mean symptom and medication scores, skin test reactivity and histamine sensitivity were significantly decreased in the patients given IT at the end of the first year when compared with the placebo group. However, there was neither a significant reduction in total and specific IgE levels nor a significant increase in IFN-gamma levels at the first year of IT. IL-4 levels were only measured at the beginning of the study because of the very low levels. A decrease in NK cell activity was found in patients treated with grass pollen extracts after 12 months when compared with Der p and placebo group. No signs of major local or systemic side effects due to IT were seen in patients during the study. Although significant clinical efficacy of specific IT with standardized extracts has been demonstrated in allergic rhinitis and asthmatic patients at the end of the first year of IT, no significant changes in immunological parameters were observed. However we conclude that a decrease in NK cell cytotoxic activity during IT has to be taken into account in the follow-up of patients.

Determination of tetanus antibodies by a double-antigen enzyme-linked immunosorbent assay in individuals of various age groups.

In this study, tetanus immunity was determined in 549 randomly chosen individuals of various age groups in Ankara, Turkey. Antibody levels in sera of the individuals were measured using a double-antigen enzyme-linked immunosorbent assay. Overall, 66.5% (95%CI, 62.4-70.4) of the population studied was found to have basic protection (>or=0.01 IU/ml) against tetanus. Protective levels of tetanus antibodies declined progressively with age. The rate of protection in children and adolescents (aged<20 years) exceeded 90%, while only 16.3% (95%CI, 8.9-26.2) of those over 60 years of age were protected. Females over 60 years of age were less immune than males of the same age group (p=0.034). Although the rates of protection in children and adolescents are regarded as satisfactory, the rates among adults are low. Preventive measures against tetanus should therefore focus on scheduled booster immunization for adults as well as children.

The assessment of functional status in Turkish children with cerebral palsy (a preliminary study).

INTRODUCTION: Cerebral palsy (CP) is one of the most common causes of disability in childhood leading to functional limitations. Assessment of the functional limitations is important to determine the severity of the disability in CP and to evaluate the benefit of the rehabilitation programme. However, the results of the measurements show variations according to different sociocultural characteristics. The Functional Independence Measure of Children (WeeFIM) had not been studied in Turkish children previously. The aims of this study were to evaluate the functional disability of Turkish children with CP by using WeeFIM and to compare the results with those of healthy counterparts. METHODS: A total of 86 children aged 24 months to 120 months were included in the study. Forty-five children with CP and 41 healthy children representing the controls were evaluated with WeeFIM. Both children with CP and healthy controls were categorized into four groups according to their chronological age. The variations in the WeeFIM subsets scores (self-care, sphincter control, transfers and locomotion, communication and social cognition) and total WeeFIM scores in children with CP and healthy controls were analysed. RESULTS: The children with CP had lower WeeFIM scores than healthy controls. The sphincter control subset scores of children with CP increased as they grew up. There was no statistically significant difference in all WeeFIM subset scores and the total WeeFIM scores among the four age groups of children with CP. CONCLUSION: The WeeFIM appears to be a useful instrument for measuring the disability of Turkish children with CP. However, studies with wider series are needed to generalize our results.

The effect of omeprazole on human natural killer cell activity.

1. Omeprazole, an antiulcer drug, inhibits the gastric acid pump via blocking the parietal cell (H+ + K+)-ATPase. Omeprazole was also reported to have an inhibitory action on polymorphonuclear neutrophil activities. In the present study the potential effect of omeprazole on human natural killer cell (NK) activity was investigated. 2. Omeprazole decreased NK cytotoxic activity in a dose-dependent manner. 3. Degraded omeprazole showed a similar action. 4. In vitro NK inhibitory action of omeprazole and its acid-degraded form was observed at the concentrations equal and higher than 18 microM (micromolar). 5. NK inhibitory action of omeprazole was recovered to 75% by washing away of the agent. 6. Omeprazole decreased the conjugate formation of effector and target cells by 50% at the concentration of 288 microM

Endomysium antibodies in the diagnosis of celiac disease in short-statured children with no gastrointestinal symptoms.

BACKGROUND: Endomysium antibodies (EmAb) are strongly associated with untreated celiac disease and are suggested to be diagnostic. The aim of the present study was to assess the value of anti-EmAb for celiac disease screening in children with short stature. METHODS: In 84 children with height less than the third percentile for age, preliminary work-ups were made to find a cause for their short stature and then their serum was assayed for anti-EmAb by indirect immunofluorescence tests using monkey esophagus. RESULTS: Seven children were strongly positive for EmAb and all had positive histologic findings for celiac disease. CONCLUSIONS: Our results show that there is an association with occult celiac disease and idiopathic short stature and that the serum anti-EmAb test is useful in identifying such cases.

Effect of a Staphylococcus epidermidis-extracted slime factor on human natural killer cell activity.

A slime factor produced by Staphylococcus epidermidis was a complex glycoconjugate extracted by the phenol extraction method. The potential stimulatory or inhibitory capacity of the phenol-extracted slime (PES) was tested on human natural killer cell cytotoxic activity. Various concentrations of the PES preparation were incubated with the effector cells 30 min before and during the assay period. The PES factor inhibited natural killer cell cytotoxic activity at a concentration of 250 micrograms/ml and at higher concentrations (p < 0.05). The inhibition of natural killer cell cytotoxic activity may probably be related to the complex composition of the slime substance.