Binding of avian IgY to type VII collagen does not activate complement and leucocytes and fails to induce subepidermal blistering in mice.

BACKGROUND: Epidermolysis bullosa acquisita (EBA) is a severe autoimmune skin disease characterized by tissue-bound and circulating autoantibodies to type VII collagen, the major component of anchoring fibrils. When passively transferred into mice, rabbit IgG against type VII collagen induces Fc-dependent activation of complement, the recruitment of leucocytes into the skin, and subepidermal blistering. In addition to these inflammatory mechanisms, clinical and experimental evidence suggests that antibodies against type VII collagen might induce blisters by disrupting the ligand function of type VII collagen by an Fc-independent mechanism. OBJECTIVES: To study noninflammatory mechanisms of blister formation in experimental EBA. METHODS: We generated chicken IgY antibodies directed to recombinant type VII collagen and examined their pathogenic activity using ex vivo and animal models. RESULTS: Mice injected with these chicken IgY antibodies showed binding of the antibodies to the dermal-epidermal junction of skin sections. However, IgY antibodies did not fix complement C3 in enzyme-linked immunosorbent assay and immunofluorescence complement-binding assays. In addition, IgY antibodies did not induce dermal-epidermal separation ex vivo. Following their passive transfer into Balb/c mice, chicken IgY antibodies against type VII collagen bound at the dermal-epidermal junction, but, in contrast to rabbit IgG, did not fix complement C3, recruit granulocytes, or induce skin blisters. CONCLUSIONS: These findings demonstrate that binding of avian IgY to type VII collagen is not pathogenic in vivo and strongly suggest that in experimental EBA, antibodies to type VII collagen induce blisters not by direct disruption of the ligand function of type VII collagen, but rather by an Fc-dependent engagement of humoral and cellular inflammatory factors.

Neonatal platelets from cord blood and peripheral blood.

The haemostatic system in neonates is different from that of adults, possibly contributing to an increased incidence of bleeding disorders, such as intracranial hemorrhage. In this study, we analyzed platelets from cord blood and peripheral blood, collected at three time points after delivery from 20 term and 37 preterm neonates as well as blood from 20 healthy adults. Platelet membrane glycoproteins (GP) were quantified and P-selectin expression and PAC-1 binding ability before and after stimulation with TRAP were analyzed by whole blood flow cytometry. We found no significant differences in neonatal platelets from cord blood and peripheral blood within the first 24h of life. Platelets from infants less than 30 weeks of gestation expressed lower levels of GP (33271+/-9381 vs. 44085+/-17287 for GPIIIa, P<0.05) and were less reactive than platelets from term newborns (4.3+/-3.3 vs. 20.1+/-11.8% PAC-1 positive platelets after stimulation with TRAP, P<0.05). A significantly lower level of GPIIb/IIIa expression on platelets from peripheral blood was seen in term newborns as well as preterm infants, compared to adults. There was only a partial enhancement in the degranulation ability (alpha-granules) (13.4+/-12.3 vs. 50.3+/-16.1% P-selectin positive platelets, P<0.05) and no significant increase for PAC-1 binding (13.6+/-10.9 vs. 15.3+/-5.9% PAC-1 positive platelets, P=0.8) during the first 12 days of life. In conclusion, we could demonstrate that neonatal platelet reactivity increases with gestational age.