Cross-reactivity of autoantibodies from patients with epidermolysis bullosa acquisita with murine collagen VII

The pathomechanism of antibody-mediated tissue damage in autoimmune diseases can be best studied in experimental models by passively transferring specific autoantibodies into animals. The reproduction of the disease in animals depends on several factors, including the cross-reactivity of patient autoantibodies with the animal tissue. Here, we show that autoantibodies from patients with epidermolysis bullosa acquisita (EBA), a subepidermal autoimmune blistering disease, recognize multiple epitopes on murine collagen VII. Indirect immunofluorescence microscopy revealed that EBA patients’ IgG cross-reacts with mouse skin. Overlapping, recombinant fragments of murine collagen VII were used to characterize the reactivity of EBA sera and to map the epitopes on the murine antigen by ELISA and immunoblotting. The patients’ autoantibody binding to murine collagen VII triggered pathogenic events as demonstrated by a complement fixing and an ex vivo granulocyte-dependent dermal–epidermal separation assay. These findings should greatly facilitate the development of improved disease models and novel therapeutic strategies.     

Chemotactic and random movement of cord-blood granulocytes

Summary Chemotactic responsiveness and random movement of cord-blood granulocytes were studied with a modified Boyden's method. Cord-blood granulocytes were less active chemotactically than granulocytes from healthy children and adults, whereas the random filter movement of the cells from all three sources was about the same. In cord sera, concentrations of cell directed chemotaxis inhibitors were equal to those in sera from other age groups. Compared with the situation in healthy children and adults, the generation of chemotactic factors in cord-blood sera was impaired. This impairment was not related to an increased activity of chemotactic factor inactivators. Measurement of the cyclic nucleotide levels in granulocytes from cord-blood and from children belonging to various age groups revealed that the cord granulocytes have significantly lower concentrations of cAMP and cGMP, which could have been responsible for the decreased chemotactic responsiveness.     

Mutation of FIG4 causes neurodegeneration in the pale tremor mouse and patients with CMT4J

Membrane-bound phosphoinositides are signalling molecules that have a key role in vesicle trafficking in eukaryotic cells. Proteins that bind specific phosphoinositides mediate interactions between membrane-bounded compartments whose identity is partially encoded by cytoplasmic phospholipid tags. Little is known about the localization and regulation of mammalian phosphatidylinositol-3,5-bisphosphate (PtdIns(3,5)P2), a phospholipid present in small quantities that regulates membrane trafficking in the endosome-lysosome axis in yeast. Here we describe a multi-organ disorder with neuronal degeneration in the central nervous system, peripheral neuronopathy and diluted pigmentation in the 'pale tremor' mouse. Positional cloning identified insertion of ETn2beta (early transposon 2beta) into intron 18 of Fig4 (A530089I17Rik), the homologue of a yeast SAC (suppressor of actin) domain PtdIns(3,5)P2 5-phosphatase located in the vacuolar membrane. The abnormal concentration of PtdIns(3,5)P2 in cultured fibroblasts from pale tremor mice demonstrates the conserved biochemical function of mammalian Fig4. The cytoplasm of fibroblasts from pale tremor mice is filled with large vacuoles that are immunoreactive for LAMP-2 (lysosomal-associated membrane protein 2), consistent with dysfunction of the late endosome-lysosome axis. Neonatal neurodegeneration in sensory and autonomic ganglia is followed by loss of neurons from layers four and five of the cortex, deep cerebellar nuclei and other localized brain regions. The sciatic nerve exhibits reduced numbers of large-diameter myelinated axons, slowed nerve conduction velocity and reduced amplitude of compound muscle action potentials. We identified pathogenic mutations of human FIG4 (KIAA0274) on chromosome 6q21 in four unrelated patients with hereditary motor and sensory neuropathy. This novel form of autosomal recessive Charcot-Marie-Tooth disorder is designated CMT4J.     

Chemotactic and random movement of cord-blood granulocytes

Chemotactic responsiveness and random movement of cord-blood granulocytes were studied with a modified Boyden's method. Cord-blood granulocytes were less active chemotactically than granulocytes from healthy children and adults, whereas the random filter movement of the cells from all three sources was about the same. In cord sera, concentrations of cell directed chemotaxis inhibitors were equal to those in sera from other age groups. Compared with the situation in healthy children and adults, the generation of chemotactic factors in cord-blood sera was impaired. This impairment was not related to an increased activity of chemotactic factor inactivators. Measurement of the cyclic nucleotide levels in granulocytes from cord-blood and from children belonging to various age groups revealed that the cord granulocytes have significantly lower concentrations of cAMP and cGMP, which could have been responsible for the decreased chemotactic responsiveness.