Childhood allergy is preceded by an absence of gut lactobacilli species and higher levels of atopy-related plasma chemokines.

Alterations in the composition and reduced diversity of the infant microbiome are associated with allergic disease in children. Further, an altered microbiota is linked to immune dysregulation, including skewing of different T helper (Th) subsets, which is also seen in atopic individuals. The aim of this study was, therefore, to investigate the associations between gut lactobacilli and Th-related plasma factors in allergy development during childhood. A total of 194 children with known allergy status at 1 year of age were followed to 10 years of age. We used real-time polymerase chain reaction (PCR) to investigate the presence of three lactobacilli species (Lactobacillus casei, L. paracasei, L. rhamnosus) in infant fecal samples (collected between 1 week and 2 months of age) from a subgroup of children. Plasma chemokines and cytokines were quantified at 6 months and at 1, 2, 5 and 10 years of age with Luminex or enzyme-linked immunosorbent assay (ELISA). Fractional exhaled nitrogen oxide (FeNO) was measured and spirometry performed at 10 years of age. The data were analysed by non-parametric testing and a logistic regression model adjusted for parental allergy. An absence of these lactobacilli and higher levels of the chemokines BCA-1/CXCL13, CCL17/TARC, MIP-3α/CCL20 and MDC/CCL22 in plasma at 6 months of age preceded allergy development. The presence of lactobacilli associated with lower levels of atopy-related chemokines during infancy, together with higher levels of interferon (IFN)-γ and lower FeNO during later childhood. The results indicate that the presence of certain lactobacilli species in the infant gut may influence allergy-related parameters in the peripheral immune system, and thereby contribute to allergy protection.

Pregnancy-associated inflammatory markers are elevated in pregnant women with systemic lupus erythematosus.

During normal pregnancy a dampening in T cell-mediated immunity is compensated by an increased pro-inflammatory activity. Likewise, the autoimmune disease systemic lupus erythematosus (SLE) is associated with inflammatory activity and pregnancy complications occur frequently in women with SLE. The aim of this study was to elucidate how SLE influences the chemokine and cytokine balance during and after pregnancy. Blood samples were taken from pregnant women with or without SLE at second and third trimester and 8-12 weeks after pregnancy. Cytokines (interleukin (IL)-1ß, IL-2, IL-4, IL-6, IL-10, IL-12p70, IL-17A, TNF, IFN-γ and IFN-α), chemokines (CXCL8/IL-8, CXCL9/MIG, CXCL10/IP-10, CCL2/MCP-1, CCL5/RANTES and CCL17/TARC), soluble IL-6 receptor (sIL-6R) and soluble glycoprotein 130 (gp130) were measured in serum using cytometric bead array (CBA) or enzyme-linked immunosorbent assay (ELISA). Women with SLE had increased serum concentrations of CXCL8/IL-8, CXCL9/MIG, CXCL10/IP-10 and IL-10 compared to controls both during and after pregnancy. Further, when dividing the patients based on disease activity, the women with active disease had the highest levels. Importantly, women with SLE seemed to respond to pregnancy in a similar way as controls, since the changes of cytokines and chemokines over the course of pregnancy were similar but with overall higher levels in the patient group. In conclusion, changes in pro- and anti-inflammatory serum components during pregnancy in women with SLE, occurring on top of already more pro-inflammatory levels, might increase their risk for pregnancy complications and flares. How their children are affected by this heightened inflammatory milieu during pregnancy needs further investigation.

Cytokine Profiles and Antibody Responses to Plasmodium Falciparum Malaria Infection in Individuals Living in Ibadan; Southwest Nigeria

Background: The ability of the host immune system to efficiently clear Plasmodium falciparum parasites during a malaria infection depends on the type of immune response mounted by the host. Study design: In a cross-sectional study; we investigated the cellular-and antibody responses in individuals with P. falciparum infection; in an attempt to identify immunological signs indicative of the development of natural immunity against malaria in Ibadan; Nigeria. Levels of IL-10; IL-12(p70); IFN-a; and IgM; IgG and IgG1-4 subclasses in the serum of 36 symptomatic children with microscopically confirmed malaria parasitaemia and 54 asymptomatic controls were analysed by ELISA. Results: IFN-a and IL-10 were significantly higher in the symptomatic children (p=0.009; p=0.025 respectively) than in the asymptomatic controls but no differences were seen for IL-12(p70). Estimated higher ratios of IFN-a/IL-10 and IFN-a/IL-12 were also observed in the symptomatic children while the asymptomatic controls had higher IL-12/IL-10 ratio. The mean concentration levels of anti-P. falciparum IgG1; IgG2; IgG3 antibodies were statistically significantly higher in the individuals 5 years of age than 5 years while anti-P. falciparum IgG3 antibodies were notably low in 5 years category. Children 5 years had higher IgM antibodies than IgG and the expression of IgG subclasses increased with age. Conclusion: Taken together; malaria infection is on a delicate balance of pro- and anti-inflammatory cytokines. The higher levels of IFN-a seen in the symptomatic children (6months) may be instrumental in immune-protection against malaria by limiting parasite replication. The observed variations in immunoglobulin subclass levels were age- dependent and exposure-related

Development and clinical evaluation of a second-generation voice prosthesis (Provox 2), designed for anterograde and retrograde insertion.

Prosthetic voice restoration has considerably improved the results of vocal rehabilitation after total laryngectomy, and is presently the method of choice for many health-care providers treating laryngectomized patients. The Provox voice prosthesis, developed in the Netherlands Cancer Institute, is an indwelling device that has been applied in recent years with regular success. Its retrograde replacement method, using a disposable guide wire, assures reliable, atraumatic positioning of the prosthesis in the tracheoesophageal fistula. However, the method sometimes may be uncomfortable for the patient; therefore an adapted prosthesis and new replacement equipment were developed, which enable bidirectional insertion, i.e. not only in the traditional retrograde manner through the pharynx, but especially in an anterograde manner through the stoma. This second-generation voice prosthesis (Provox 2) was studied in a prospective clinical trial in 44 patients (33 experienced patients, seven first-time replacements and four primary insertions). The study demonstrated that the anterograde insertion with the Provox 2 system was applicable in all patients, making the voice prosthesis even easier to handle than with the traditional retrograde method. A stenosis of the pharyngoesophageal segment no longer interfered with the replacement. In addition, the patients judged the new method as being favourable, reporting significantly less discomfort during the replacement procedure (paired Student's t-test: p < 0.0001). Furthermore, the adapted voice prosthesis could be removed from the tracheoesophageal fistula without excessive force (mean 7.9 N, range 6.0-14.0 N), more easily than the original Provox (mean 20.9 N, range 5.5-25.0 N). It can be concluded that this second-generation indwelling voice prosthesis (Provox 2) seems to be a further improvement in the application of this voice rehabilitation system, not only simplifying the replacement procedure, but also diminishing the discomfort for the patient.

Hydroxylation of salicylate by microsomal fractions and cytochrome P-450. Lack of production of 2,3-dihydroxybenzoate unless hydroxyl radical formation is permitted.

Attack by hydroxyl radicals (.OH) upon salicylate (2-hydroxybenzoate) leads to formation of both 2,3-dihydroxybenzoate (2,3-DHB) and 2,5-dihydroxybenzoate (gentisate, 2,5-DHB). It has been suggested that formation of 2,3-DHB from salicylate is a means of monitoring .OH formation. Production of 2,3-DHB and 2,5-DHB by liver microsomal fractions and isoforms of cytochrome P-450 was investigated. Liver microsomes prepared from variously treated rats and rabbits catalysed the formation of 2,5-DHB but not 2,3-DHB. Formation of 2,5-DHB was inhibited by CO, metyrapone and SKF-525A, but not by the .OH scavengers mannitol and formate or by the iron chelator desferrioxamine. Purified P-450s IIE1, IIB4 or IA2 from rabbit liver microsomes, reconstituted together with NADPH-cytochrome P-450 reductase, led to formation of equal amounts of 2,3-DHB and 2,5-DHB in reactions that were almost completely inhibited by mannitol or formate. Addition of Fe3+/EDTA either to microsomes or to membranes containing reconstituted P-450 caused formation of approximately equal amounts of 2,3-DHB and 2,5-DHB, consistent with an .OH-dependent attack on salicylate. The data indicate that the microsomal P-450 system catalyses hydroxylation of salicylate to 2,5-DHB, but not formation of 2,3-DHB. Hence measurement of 2,3-DHB might provide a means of monitoring .OH formation. Care must be taken in studies of substrate hydroxylation by microsomes or reconstituted P-450 systems to avoid artefacts resulting from .OH generation.

Role of ethanol-inducible cytochrome P450 (P450IIE1) in catalysing the free radical activation of aliphatic alcohols.

Incubation of rat liver microsomes with 1-propanol and 1-butanol in the presence of NADPH and of the spin trapping agent 4-pyridyl-1-oxide-t-butyl nitrone (4-POBN) allowed the detection of free radical intermediates tentatively identified as 1-hydroxypropyl and 1-hydroxybutyl radical, respectively. Microsomes isolated from rats treated chronically with ethanol (EtOH) or with the combination of starvation and acetone treatment (SA), exhibited a two-fold increase in the ESR signal intensity as compared to untreated controls, whereas no increase was observed in phenobarbital-induced (PB) microsomes. Consistently, in reconstituted membrane vesicles, ethanol-inducible cytochrome P450IIE1 was twice as active as phenobarbital-inducible P450IIB1 in producing 1-butanol free radicals. In the microsomal preparations from EtOH and SA pretreated rats the addition of antibodies against cytochrome P450IIE1, but not of preimmune IgGs, lowered the ESR signal of 1-butanol radicals by more than 50%. The same antibodies decreased the free radical production by untreated microsomes by 35-40%, but were ineffective on microsomes from PB-treated animals. This indicated that cytochrome P450IIE1 is the major enzyme responsible for the free radical activation of alcohols in control and ethanol-fed rats. The generation of 1-hydroxybutyl radicals by EtOH microsomes was inhibited by 40, 48 and 68%, respectively, by the addition of isoniazid, tryptamine and octylamine, compounds known to specifically affect the NADPH oxidase activity of this isoenzyme. This effect was not due to the scavenging of the alcohol radical since none of these compounds affected the ESR signals originated from 1-butanol in a xanthine-xanthine oxidase system. When added to reconstituted membrane vesicles isoniazid, tryptamine and octylamine also decreased 1-butanol radical formation by P450IIE1 by 54, 38 and 66%, respectively. Such an inhibition corresponded to the effect exerted by the same compounds on O2- release from P450IIE1 containing vesicles. These results indicate that the capacity of cytochrome P450IIE1 to reduce oxygen is related to its ability to generate alcohol free radicals and suggest that ferric cytochrome P450-oxygen complex might act as oxidizing species toward alcohols.

Cytochrome P-450-dependent formation of reactive oxygen radicals: isozyme-specific inhibition of P-450-mediated reduction of oxygen and carbon tetrachloride.

1. Ethanol-inducible P450 IIE1 exhibits a high rate of oxygen consumption and oxidase activity. The enzyme is selectively distributed in the liver centrilobular area, the acinar region specifically destroyed after treatment with P450 IIE1 substrates/inducers such as ethanol, carbon tetrachloride, chloroform, N-nitrosodimethylamine and paracetamol. 2. Twenty substrates and ligands for cytochrome P450 IIB4 and P450 IIE1 were evaluated for their ability to inhibit microsomal and reconstituted NADPH-dependent oxidase activity, and the P450 IIE1-catalysed reduction of carbon tetrachloride to chloroform. Type I ligands and substrates did not inhibit the processes whereas nitrogen-containing compounds such as octylamine, cimetidine, imidazole and tryptamine inhibited NADPH oxidation and H2O2 formation in microsomes from starved and acetone-treated rats by around 50%. 3. Tryptamine, octylamine, isoniazid and p-chloroamphetamine inhibited reconstituted P450 IIE1-dependent oxidase activity with half maximal effects at 14-170 microM. 4. Isoniazid, cimetidine and tryptamine inhibited the P450 IIE1-dependent reduction of carbon tetrachloride, whereas acetone was without effect. 5. The oxygen dependency of microsomal oxidase activity exhibited high-affinity and low-affinity phases, with partial saturation at 20 microM of O2. 6. It is concluded that microsomal oxidase activity takes place at physiological concentrations of O2 and that isozyme-specific type II ligands compete with oxygen or carbon tetrachloride for reduction by P-450 haem.