Non-heparan sulfate-binding interactions of endostatin/collagen XVIII in murine development.

Knobloch syndrome is characterized by a congenital generalized eye disease and cranial defect. Pathogenic mutations preferentially lead to a deletion or functional alteration of collagen XVIII's most C-terminal endostatin domain. Endostatin can be released from collagen XVIII and is a potent inhibitor of angiogenesis and tumor growth. We show differential expression of binding partners for endostatin, vascular endothelial growth factor (VEGF), and the collagen XV endostatin homologue in murine embryonal development using a set of alkaline phosphatase fusion proteins. Consistent with the human phenotype, vascular mesenchyme in the developing eye was identified as endostatin's primary target. While endostatin predominantly bound to blood vessels, the VEGF164 affinity probe labeled nonvascular tissues such as forebrain, hindbrain, the optic nerve, and the surface ectoderm of the future cornea. Strikingly increased staining specificity was observed with a non-heparin/heparan sulfate-binding endostatin probe. In contrast, elimination of the heparan sulfate binding site from VEGF led to complete loss of binding. The collagen XV endostatin homologue showed a highly restricted binding pattern. Oligomerization with endogenous endostatin was ruled out by use of collagen XVIII knockout mice. Our data provide strong evidence that collagen XVIII's C-terminal endostatin domain harbors a prominent tissue-binding site and that binding can occur in the absence of heparan sulfates in situ.     

Glial cell expression of hepatocyte growth factor in vitreoretinal proliferative disease

The hepatocyte growth factor (HGF) has been crucially implicated in the development of proliferative retinal diseases; however, it is unclear whether retinal glial cells express or respond to HGF. Therefore, we examined the expression of HGF and of the receptor for HGF, c-Met, by immunohistochemical costaining with glial fibrillary acidic protein (GFAP) in epiretinal membranes of patients with proliferative vitreoretinopathy (PVR) and proliferative diabetic retinopathy (PDR), respectively. Furthermore, it was determined whether cells of the human retinal glial cell line, MIO-M1, secrete HGF protein, and whether HGF stimulates proliferation and chemotaxis, and secretion of the vascular endothelial growth factor (VEGF). Neuroretinas of patients with PVR express elevated mRNA level for HGF in comparison to control retinas. In epiretinal membranes of patients with PVR or PDR, immunoreactivity for HGF and for c-Met, respectively, partially colocalized with immunoreactivity for GFAP. Fetal bovine serum and basic fibroblast growth factor, but not heparin-binding epidermal or platelet-derived growth factors, evoked HGF secretion by cultured retinal glial cells. HGF displayed only a marginal effect on cell proliferation while it stimulated chemotaxis. HGF promoted the secretion of VEGF, via activation of the phosphatidylinositol-3 kinase. It is concluded that glial cells in epiretinal membranes express both HGF protein and c-Met receptors. The results suggest an autocrine/paracrine role of HGF in glial cell responses during proliferative vitreoretinal disorders as well as in retinal neovascularization, by stimulating of VEGF release.     

c-Fos-dependent induction of the small ras-related GTPase Rab11a in skin carcinogenesis

Malignant transformation of mouse skin by tumor promoters and chemical carcinogens, such as the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA), is a multistage process leading to the formation of squamous cell carcinomas. It has been shown that mice lacking the AP-1 family member c-Fos exhibit an impaired transition from benign to malignant skin tumors. Here, we demonstrate enhanced expression of the small Ras-related GTPase Rab11a after short-term TPA treatment of mouse back skin. Expression of Rab11a in vivo and in vitro critically depended on c-Fos, because TPA application to the back skin of c-Fos-deficient mice and to mouse embryonic fibroblasts did not induce Rab11a mRNA or protein expression. Moreover, dexamethasone, which is a potent inhibitor of AP-1-mediated transactivation that exhibits anti-inflammatory and anti-tumor promoting activities, inhibited TPA-induced expression of Rab11a. Within the Rab11a gene promoter, we identified a functional AP-1 binding element that exhibited elevated c-Fos binding activity after TPA treatment of keratinocytes. Enhanced expression was not restricted to chemically induced mouse skin tumors but was also found in tumor specimens derived from patients with epithelial skin tumors. These data identify Rab11a as a novel, tumor-associated c-Fos/AP-1 target and may point to an as yet unrecognized function of Rab11a in the development of skin cancer.     

Peanut-induced anaphylaxis in children and adolescents: Data from the European Anaphylaxis Registry

Background

Peanut allergy has a rising prevalence in high-income countries, affecting 0.5%–1.4% of children. This study aimed to better understand peanut anaphylaxis in comparison to anaphylaxis to other food triggers in European children and adolescents.

Methods

Data was sourced from the European Anaphylaxis Registry via an online questionnaire, after in-depth review of food-induced anaphylaxis cases in a tertiary paediatric allergy centre.

Results

3514 cases of food anaphylaxis were reported between July 2007 - March 2018, 56% in patients younger than 18 years. Peanut anaphylaxis was recorded in 459 children and adolescents (85% of all peanut anaphylaxis cases). Previous reactions (42% vs. 38%; p = .001), asthma comorbidity (47% vs. 35%; p < .001), relevant cofactors (29% vs. 22%; p = .004) and biphasic reactions (10% vs. 4%; p = .001) were more commonly reported in peanut anaphylaxis. Most cases were labelled as severe anaphylaxis (Ring&Messmer grade III 65% vs. 56% and grade IV 1.1% vs. 0.9%; p = .001). Self-administration of intramuscular adrenaline was low (17% vs. 15%), professional adrenaline administration was higher in non-peanut food anaphylaxis (34% vs. 26%; p = .003). Hospitalization was higher for peanut anaphylaxis (67% vs. 54%; p = .004).

Conclusions

The European Anaphylaxis Registry data confirmed peanut as one of the major causes of severe, potentially life-threatening allergic reactions in European children, with some characteristic features e.g., presence of asthma comorbidity and increased rate of biphasic reactions. Usage of intramuscular adrenaline as first-line treatment is low and needs to be improved. The Registry, designed as the largest database on anaphylaxis, allows continuous assessment of this condition.

     

Determination of penetration profiles of topically applied substances by means of tape stripping and optical spectroscopy: UV filter substance in sunscreens

Penetration profiles of topically applied drugs and cosmetic products provide important information on their efficacy. The application of tape stripping in combination with UV/VIS spectroscopy is checked to determine the local position of topically applied substances inside the stratum corneum, the penetration profile. The amount of corneocytes removed with each tape strip is quantified via the particle-dependent absorption, the pseudoabsorption, in the visible spectral range. The concentration of a typical UV filter substance, 4-methylbenzylidene camphor, is determined by optical spectroscopy using the tape strips removed originally. In this case, a time-dependent increase in the absorbance must be taken into account. Laser scanning microscopic investigations confirm that the nonhomogeneous distribution of the filter substance, on the strips, can explain this spectroscopic behavior. When reaching a homogeneous distribution, the UV spectroscopic signal reflects the correct concentration. These spectroscopic values are compared with high performance liquid chromatography (HPLC) data. The values obtained with both methods for the concentrations of 4-methylbenzylidene camphor are in good agreement. The data obtained are used to illustrate the determination of a penetration profile of a UV filter substance. The results demonstrate that the described protocol is well suited to characterize, in a simple manner, topically applied substances that have a characteristic UV/VIS absorption band.     

Abbau von Mischsubstraten durch Hefen I. Substrate der Sulfitablauge

Two Candida strains were isolated from an industrial batch reactor for fermentation of waste sulfite liquor. Pure cultures of these yeasts assimilate single substrates contained in waste sulfite liquor with different specific activity. The respiration rate is influenced by some induction substrates, too. In a mixed substrate model corresponding to waste sulfite liquor the single substrates are metabolized by pure cultures in a specific sequential manner. These sequences change during degradation of mixed substrates by mixed cultures. The spectrum of metabolized substrates, however, is not reduced. This fact is important for energy balance or for waste water contamination.     

Glial bridges and Schwann cell migration during chronic demyelination in the C.N.S.

Summary The formation of fibrotic bridges from subpial astrocytes into the subarachnoid space of the spinal cord and the migration of Schwann cells to the central nervous system (C.N.S.) is appraised in chronically demyelinated C.N.S. lesions. Spinal cord tissue was studied from inbred, Strain 13 guinea pigs with chronic experimental allergic encephalomyelitis (EAE). It has been found that uncommitted Schwann cells are present around remyelinated fibres in nerve root entry zones, between meningeal cells at a distance from the roots and along blood vessels within the spinal cord parenchyma. It is speculated that these cells migrate via the above route to the C.N.S. In the present model, this invasion might be aided by glial fibrosis, a process which leads to surface irregularities in the spinal cord, an extensive extracellular space and possible breaches in the glia limitans through which Schwann cells might penetrate.     

Influence of selected wound dressings on PMN elastase in chronic wound fluid and their antioxidative potential in vitro

Exudates from non-healing wounds contain elevated levels of proteolytic enzymes, like elastase from polymorphonuclear granulocytes (PMN elastase), reactive oxygen species (ROS) and reactive nitrogen species (RNS). The overproduction of proteolytic enzymes leads to reduced concentrations of growth factors and proteinase inhibitors, resulting in an imbalance between degradation and remodelling processes. Thus, the reduction of protein-degrading enzymes and scavenging of ROS and RNS seem to be suitable ways to support the healing process of chronic stagnating wounds. The aim of this study was to test selected wound dressings from different biomaterials (collagen, oxidized regenerated cellulose (ORC) and ORC/collagen mixture), regarding their antioxidative potential in vitro and their influence on the concentration and activity of PMN elastase in chronic wound fluid. Antioxidant capacity of the investigated wound dressing was determined by a pholasin-based chemiluminescent assay. PMN elastase concentration was determined by means of ELISA. Enzyme activities could be measured by a fluorescence assay. As the presented data demonstrates, all tested materials showed antioxidant capacity. In addition, the investigated materials were able to reduce the concentration and activity of PMN elastase. Beside other aspects, such as biocompatibility, biodegradability, fluid absorption and clinical effects (e.g. angiogenesis and microcirculation), the understanding of these properties may help to support the further refinement of wound dressings for improved wound healing.     

Hematopoietic progenitor cells and cellular microenvironment: behavioral and molecular changes upon interaction

Cell-cell contact between stem cells and cellular determinants of the microenvironment plays an essential role in controlling cell division. Using human hematopoietic progenitor cells (CD34+/CD38-) and a stroma cell line (AFT024) as a model, we have studied the initial behavioral and molecular sequel of this interaction. Time-lapse microscopy showed that CD34+/CD38- cells actively migrated toward and sought contact with stroma cells and 30% of them adhered firmly to AFT024 stroma through the uropod. CD44 and CD34 are colocalized at the site of contact. Gene expression profiles of CD34+/CD38- cells upon cultivation with or without stroma for 16, 20, 48, or 72 hours were analyzed using our human genome cDNA microarray. Chk1, egr1, and cxcl2 were among the first genes upregulated within 16 hours. Genes with the highest upregulation throughout the time course included tubulin genes, ezrin, c1qr1, fos, pcna, mcm6, ung, and dnmt1, genes that play an essential role in reorganization of the cytoskeleton system, stabilization of DNA, and methylation patterns. Our results demonstrate directed migration of CD34+/CD38- cells toward AFT024 and adhesion through the uropod and that upon interaction with supportive stroma, reorganization of the cytoskeleton system, regulation of cell division, and maintenance of genetic stability represent the most essential steps.