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IgG4 autoantibodies induce dermal-epidermal separation.

Mihai S, Chiriac MT, Herrero-González JE, Goodall M, Jefferis R, Savage CO, Zillikens D, Sitaru C - J. Cell. Mol. Med. (2007 Sep-Oct)

Bottom Line: In these diseases, IgG1 and IgG3 isotypes, but not the IgG4 subclass, are thought to trigger inflammatory pathways resulting in tissue damage.Purified IgG4 did not fix complement, but, interestingly, like IgG1, activated leucocytes and induced dermal-epidermal separation.The potential of IgG4 autoantibodies to induce Fc-dependent dermal-epidermal separation was significantly lower compared to IgG1.

View Article: PubMed Central - PubMed

Affiliation: Department of Dermatology, University of Lübeck, Lübeck, Germany.

ABSTRACT
Bullous pemphigoid (BP) is a sub-epidermal autoimmune blistering disease associated with autoantibodies to the dermal-epidermal junction (DEJ). Patients' autoantibodies induce dermal-epidermal separation when co-incubated with cryosections of human skin and leucocytes from healthy volunteers. IgG autoantibodies trigger complement and/or leucocyte activation resulting in specific pathology in several autoimmune conditions. In these diseases, IgG1 and IgG3 isotypes, but not the IgG4 subclass, are thought to trigger inflammatory pathways resulting in tissue damage. The capacity of IgG4 autoantibodies to mediate tissue damage has not yet been demonstrated. In this study, we isolated IgG1 and IgG4 autoantibodies from bullous pemhigoid patients'serum and analysed their blister-inducing potential in our cryosection assay. As expected, complement-fixing IgG1 autoantibodies induced sub-epidermal splits in this experimental model. Purified IgG4 did not fix complement, but, interestingly, like IgG1, activated leucocytes and induced dermal-epidermal separation. The potential of IgG4 autoantibodies to induce Fc-dependent dermal-epidermal separation was significantly lower compared to IgG1. Our results demonstrate that IgG4 autoantibodies are able to activate leucocytes and point to a hitherto less recognized function of IgG4. Moreover, for the first time, we clearly demonstrate that BP IgG4 autoantibodies have the capacity to induce leucocyte-dependent tissue damage.

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IgG4 autoantibodies recruit and activate leucocytes. Sections of human skin were incubated with IgG1 and IgG4 autoantibodies from a BP patient, as well as with IgG from a healthy control. Subsequent addition of leucocytes from healthy donors leads to leucocyte attachment at the dermal–epidermal junction in sections treated with patient's IgG1 (A) and IgG4 autoantibodies (B), but not in sections incubated with control IgG (C). Activation of leucocytes, as revealed by the reduction of nitro blue tetrazolium (NBT) to formazan (dark precipitates), is induced by purified IgG1 (D) and IgG4 autoantibodies (E), but not by control IgG (F) (all magnifications, ×400). (G) Cryosections of human skin were treated with IgG1 (n = 3) and IgG4 (n = 5) antibodies (four sections/antibody preparation). Subsequently, leucocytes from healthy donors were incubated for 90 min with the cryosections. Deposition of formazan is represented as means ± SEM of the percent of the total length of the dermal–epidermal junction (DEJ) for each section.
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fig03: IgG4 autoantibodies recruit and activate leucocytes. Sections of human skin were incubated with IgG1 and IgG4 autoantibodies from a BP patient, as well as with IgG from a healthy control. Subsequent addition of leucocytes from healthy donors leads to leucocyte attachment at the dermal–epidermal junction in sections treated with patient's IgG1 (A) and IgG4 autoantibodies (B), but not in sections incubated with control IgG (C). Activation of leucocytes, as revealed by the reduction of nitro blue tetrazolium (NBT) to formazan (dark precipitates), is induced by purified IgG1 (D) and IgG4 autoantibodies (E), but not by control IgG (F) (all magnifications, ×400). (G) Cryosections of human skin were treated with IgG1 (n = 3) and IgG4 (n = 5) antibodies (four sections/antibody preparation). Subsequently, leucocytes from healthy donors were incubated for 90 min with the cryosections. Deposition of formazan is represented as means ± SEM of the percent of the total length of the dermal–epidermal junction (DEJ) for each section.

Mentions: The capacity of IgG1 and IgG4 autoantibodies to activate leucocytes at the basement membrane was examined using frozen sections of human skin incubated with antibody preparations from three and five BP patients, respectively, and, subsequently, with leucocytes from healthy volunteers. Surprisingly, in addition to IgG1 autoantibodies (Fig. 3A), IgG4 (Fig. 3B) also induced an attachment of leucocytes to the DEJ. To determine whether leucocytes that attach to the DEJ become activated, cryosections that had been previously treated with IgG4 and IgG1 autoantibodies were incubated with leucocytes in the presence of nitro blue tetrazolium. After a 90 min incubation, dark-blue precipitates of reduced nitro-blue tetrazolium (formazan) were found at the DEJ of sections treated with IgG1 autoantibodies (Fig. 3D and G). Interestingly, in sections incubated with purified IgG4 autoantibodies, deposition of formazan along the basement membrane was also observed in an antibody dose-dependent manner (Fig. 3E and G). In sections treated with control IgG, no attachment of cells and no precipitates of formazan at the DEJ were found (Fig. 3C and F). Using the Fisher's exact test, IgG4 autoantibodies induced significantly less formazan deposition when compared with IgG1 autoantibodies (P= 0.023).


IgG4 autoantibodies induce dermal-epidermal separation.

Mihai S, Chiriac MT, Herrero-González JE, Goodall M, Jefferis R, Savage CO, Zillikens D, Sitaru C - J. Cell. Mol. Med. (2007 Sep-Oct)

IgG4 autoantibodies recruit and activate leucocytes. Sections of human skin were incubated with IgG1 and IgG4 autoantibodies from a BP patient, as well as with IgG from a healthy control. Subsequent addition of leucocytes from healthy donors leads to leucocyte attachment at the dermal–epidermal junction in sections treated with patient's IgG1 (A) and IgG4 autoantibodies (B), but not in sections incubated with control IgG (C). Activation of leucocytes, as revealed by the reduction of nitro blue tetrazolium (NBT) to formazan (dark precipitates), is induced by purified IgG1 (D) and IgG4 autoantibodies (E), but not by control IgG (F) (all magnifications, ×400). (G) Cryosections of human skin were treated with IgG1 (n = 3) and IgG4 (n = 5) antibodies (four sections/antibody preparation). Subsequently, leucocytes from healthy donors were incubated for 90 min with the cryosections. Deposition of formazan is represented as means ± SEM of the percent of the total length of the dermal–epidermal junction (DEJ) for each section.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4401274&req=5

fig03: IgG4 autoantibodies recruit and activate leucocytes. Sections of human skin were incubated with IgG1 and IgG4 autoantibodies from a BP patient, as well as with IgG from a healthy control. Subsequent addition of leucocytes from healthy donors leads to leucocyte attachment at the dermal–epidermal junction in sections treated with patient's IgG1 (A) and IgG4 autoantibodies (B), but not in sections incubated with control IgG (C). Activation of leucocytes, as revealed by the reduction of nitro blue tetrazolium (NBT) to formazan (dark precipitates), is induced by purified IgG1 (D) and IgG4 autoantibodies (E), but not by control IgG (F) (all magnifications, ×400). (G) Cryosections of human skin were treated with IgG1 (n = 3) and IgG4 (n = 5) antibodies (four sections/antibody preparation). Subsequently, leucocytes from healthy donors were incubated for 90 min with the cryosections. Deposition of formazan is represented as means ± SEM of the percent of the total length of the dermal–epidermal junction (DEJ) for each section.
Mentions: The capacity of IgG1 and IgG4 autoantibodies to activate leucocytes at the basement membrane was examined using frozen sections of human skin incubated with antibody preparations from three and five BP patients, respectively, and, subsequently, with leucocytes from healthy volunteers. Surprisingly, in addition to IgG1 autoantibodies (Fig. 3A), IgG4 (Fig. 3B) also induced an attachment of leucocytes to the DEJ. To determine whether leucocytes that attach to the DEJ become activated, cryosections that had been previously treated with IgG4 and IgG1 autoantibodies were incubated with leucocytes in the presence of nitro blue tetrazolium. After a 90 min incubation, dark-blue precipitates of reduced nitro-blue tetrazolium (formazan) were found at the DEJ of sections treated with IgG1 autoantibodies (Fig. 3D and G). Interestingly, in sections incubated with purified IgG4 autoantibodies, deposition of formazan along the basement membrane was also observed in an antibody dose-dependent manner (Fig. 3E and G). In sections treated with control IgG, no attachment of cells and no precipitates of formazan at the DEJ were found (Fig. 3C and F). Using the Fisher's exact test, IgG4 autoantibodies induced significantly less formazan deposition when compared with IgG1 autoantibodies (P= 0.023).

Bottom Line: In these diseases, IgG1 and IgG3 isotypes, but not the IgG4 subclass, are thought to trigger inflammatory pathways resulting in tissue damage.Purified IgG4 did not fix complement, but, interestingly, like IgG1, activated leucocytes and induced dermal-epidermal separation.The potential of IgG4 autoantibodies to induce Fc-dependent dermal-epidermal separation was significantly lower compared to IgG1.

View Article: PubMed Central - PubMed

Affiliation: Department of Dermatology, University of Lübeck, Lübeck, Germany.

ABSTRACT
Bullous pemphigoid (BP) is a sub-epidermal autoimmune blistering disease associated with autoantibodies to the dermal-epidermal junction (DEJ). Patients' autoantibodies induce dermal-epidermal separation when co-incubated with cryosections of human skin and leucocytes from healthy volunteers. IgG autoantibodies trigger complement and/or leucocyte activation resulting in specific pathology in several autoimmune conditions. In these diseases, IgG1 and IgG3 isotypes, but not the IgG4 subclass, are thought to trigger inflammatory pathways resulting in tissue damage. The capacity of IgG4 autoantibodies to mediate tissue damage has not yet been demonstrated. In this study, we isolated IgG1 and IgG4 autoantibodies from bullous pemhigoid patients'serum and analysed their blister-inducing potential in our cryosection assay. As expected, complement-fixing IgG1 autoantibodies induced sub-epidermal splits in this experimental model. Purified IgG4 did not fix complement, but, interestingly, like IgG1, activated leucocytes and induced dermal-epidermal separation. The potential of IgG4 autoantibodies to induce Fc-dependent dermal-epidermal separation was significantly lower compared to IgG1. Our results demonstrate that IgG4 autoantibodies are able to activate leucocytes and point to a hitherto less recognized function of IgG4. Moreover, for the first time, we clearly demonstrate that BP IgG4 autoantibodies have the capacity to induce leucocyte-dependent tissue damage.

Show MeSH
Related in: MedlinePlus