Detection of four human milk groups with respect to Lewis blood group dependent oligosaccharides

Neutral oligosaccharides in human milk samples from approximately 50 women were analysed applying a recently developed high-pH anion-exchange chromatographic method. Three different oligosaccharide patterns could be detected in accordance with milk groups that had been already described. These oligosaccharide groups correspond to the Lewis blood types Le(a−b+), Le(a+b−) and Le(a−b−). In addition to these oligosaccharide patterns, a new carbohydrate pattern was detected in a milk sample from a Le(a−b−) individual. Here, only nonfucosylated oligosaccharides and compounds bearing a1,3 linked fucosyl residues were found, whereas structures with a1,2 and a1,4 fucosyl linkages were missing. This finding led to the hypothesis that there are four different oligosaccharide milk groups that fit well to the genetic basis of the Lewis blood group system. This revised version was published online in November 2006 with corrections to the Cover Date.     

6-OHDA-induced ectopia of external granule cells in the subarachnoid space covering the cerebellum

The present report describes the genesis, development and topographical distribution of ectopic cells of the external granular layer in the subarachnoid space covering the rat cerebellum. Following one intracisternal injection to newborn rats of 100 micrograms 6-hydroxydopamine (6-OHDA), the meningeal cells degenerate and are removed by phagocytosis within 24 h post injection (p.i.), leaving the cerebellar cortex without a pia-arachnoid cover. Defects appear in the basal lamina investing the cerebellar cortex 3 to 5 days p.i., and both external granule cells and 'sprouts' from Bergmann-glia endfeet grow into the subarachnoid space. The latter form large, flat glial lamellae and cover extensive areas of the denuded cerebellar surface, although they do not form a glial scar over the exposed neuropil of the cerebellar cortex. The numbers of ectopic external granule cells increase within the subarachnoid space both by proliferation and a continuous efflux of cells from the cerebellar cortex. They migrate, aggregate, and ultimately develop into granule, stellate and basket cells, the morphology of which is indistinguishable from their counterparts in situ; they make specific afferent and efferent connections, both among themselves and with the underlying cerebellar cortex and brainstem. The distribution of ectopic external granule cells and their derivatives is restricted to the anterior vermal fissures and the vermal-hemispheric junctions. The present results indicate that external granule cells and their derivatives are capable of both differentiating normally and surviving in the subarachnoid space if they become associated with glial cells and establish synaptic connections.     

Molecular organization of the outer membrane of Salmonella typhimurium different release of lipopolysaccharide from wild type and lipopolysaccharide mutant cells by EDTA treatment

Cells of Salmonella typhimurium wild type and of several well defined lipopolysaccharide mutants were treated with EDTA. The percentage release of lipopolysaccharide and phospholipid was determined. The results obtained show that the release of lipopolysaccharide by EDTA declines along with the gradually diminishing chain length of the lipopolysaccharide, althought the total amount of lipopolysaccharide was found to increase at the same time in the respective mutants. Implications of these findings for the organization of the outer membrane are discussed.     

The bulk of ATP is associated to proteins in the living cell: a release kinetics study

The analysis of the nonionic detergent induced release kinetics of intracellular components is a promising approach to obtain information on the possible molecular interactions inside the living cell. The molecular interactions of ATP were studied by monitoring the nonionic detergent-induced release kinetics of ATP from cultured human cells. The observed rapid mobilization rate in the presence of Triton X 100 and the retarded release in the presence of Brij 58 were in good agreement with the detergent-induced release kinetics of proteins and K+ reported earlier. These data are consistent with the idea that the bulk of ATP is associated to the protein macromolecules of the living cell.     

Targeted disruption of the porcine immunoglobulin kappa light chain locus

Inactivation of the endogenous pig immunoglobulin (Ig) loci, and replacement with their human counterparts, would produce animals that could alleviate both the supply and specificity issues of therapeutic human polyclonal antibodies (PAbs). Platform genetics are being developed in pigs that have all endogenous Ig loci inactivated and replaced by human counterparts, in order to address this unmet clinical need. This report describes the deletion of the porcine kappa (??) light chain constant (C??) region in pig primary fetal fibroblasts (PPFFs) using gene targeting technology, and the generation of live animals from these cells via somatic cell nuclear transfer (SCNT) cloning. There are only two other targeted loci previously published in swine, and this is the first report of a targeted disruption of an Ig light chain locus in a livestock species. Pigs with one targeted C?? allele (heterozygous knockout or ±) were bred together to generate C?? homozygous knockout (?/?) animals. Peripheral blood mononuclear cells (PBMCs) and mesenteric lymph nodes (MLNs) from C?? ?/? pigs were devoid of ??-containing Igs. Furthermore, there was an increase in lambda (??) light chain expression when compared to that of wild-type littermates (C?? +/+). Targeted inactivation of the Ig heavy chain locus has also been achieved and work is underway to inactivate the pig lambda light chain locus.     

Localisation of the Vacuolar Proton Pump (V-H+-ATPase) and Carbonic Anhydrase II in the Human Eccrine Sweat Gland

The localisation of the vacuolar proton pump (V-H+ -ATPase) and the enzyme carbonic anhydrase II (CAII) was investigated in the human eccrine sweat gland employing standard immunohistochemical techniques after antigen retrieval using microwave heat treatment and high pressure. The high-pressure antigen retrieval unmasked the presence of V-H+ -ATPase in the clear cells of the secretory coil, with a distribution similar to that previously observed for CAII. However, the dark cells were unreactive to both antibodies. In addition, heat and high-pressure antigen retrieval demonstrated the presence of CAII in the apical zone of luminal cells of the reabsorptive duct, a location not previously reported. The localisation of V-H+ -ATPase and CAII in the secretory coil clear cells suggests that the formation of HCO3- and H+ by carbonic anhydrase II and the transport of H+ by V-H+ -ATPase may play an role in sweat fluid secretion. Their presence at the apex of the duct cells indicates involvement in ductal ion reabsorption.     

Meta-analysis identified the TNFA -308G > A promoter polymorphism as a risk factor for disease severity in patients with rheumatoid arthritis

Introduction

The goal of this study is to investigate whether the -308G > A promoter polymorphism in the tumor necrosis factor alpha (TNFA) gene is associated with disease severity and radiologic joint damage in a large cohort of patients with rheumatoid arthritis (RA).

Methods

A long-term observational early RA inception cohort (n = 208) with detailed information about disease activity and radiologic damage after 3, 6 and 9 years of disease was genotyped for the TNFA -308G > A promoter polymorphism (rs1800629). A longitudinal regression analysis was performed to assess the effect of genotype on RA disease severity and joint damage. Subsequently, a meta-analysis, including all publically available data, was performed to further test the association between joint erosions and the TNFA polymorphism. To learn more about the mechanism behind the effect of the polymorphism, RNA isolated from peripheral blood from RA patients (n = 66) was used for TNFA gene expression analysis by quantitative PCR.

Results

Longitudinal regression analysis with correction for gender and disease activity showed a significant difference in total joint damage between GG and GA+AA genotype groups (P = 0.002), which was stable over time. The meta-analysis, which included 2,053 patients, confirmed an association of the genetic variant with the development of erosions (odds ratio 0.78, 95% CI 0.62, 0.98). No significant differences in TNFA gene expression were observed for the different genotypes, confirming earlier findings in healthy individuals.

Conclusions

Our data confirm that the TNFA -308G > A promoter polymorphism is associated with joint damage in patients with RA. This is not mediated by differences in TNFA gene expression between genotypes.