auto immune blistering disease in skin

1. ADVANCES IN
IMMUNOLOGY IN AIBD
DR.HIMABINDU
ASI

2. INTRODUCTION
• The stratified squamous epithelium of the epidermis forms a continuous barrier
against the external environment
• InAIBDs ther e is the disruption of the intact skin barrier as a consequence of
blistering and erosions caused by production of autoantibodies against structural
proteins in the epidermis or at the epidermal-dermal junction.
• Immunologically, these conditions are driven by humoral and cellular autoimmune
responses directed against distinct target antigens and can be classed in three main
groups 1. pemphigoid and 2. pemphigus diseases as well as3. dermatitis herpetiformis
(DH)
• Over the past four decades, our knowledge of the pathophysiology of AIBDs has been
greatly advanced by demonstrating that passive transfer of antibodies against skin
antigens induce blisters in experimental animals models with clinical, histologic, and
immunopathogenic responses similar to those observed in human disease .
• Intraepidermal blistering found in pemphigus disorders are caused by autoantibodies
targeting cadherin proteins in desmosomes; subtypes pemphigus vulgaris antibodies
against (dsg)-3 and pemphigus foliaceus antibodies against desmoglein−1,
•

3. • In bullous pemphigoid (BP), autoantibodies target two hemidesmosome components
BP180 and BP230; and in
• epidermolysis bullosa acquisita (EBA) patients have autoantibodies target type VII
collagen anchoring fibrils.
• In DH patients, autoantibodies target tissue and epidermal transglutaminase (eTG)
protiens
• Diagnosis of AIBDs relies on direct immunofluorescence microscopy studies and
immunoserological assays
• Multiple mechanisms of skin barrier disruption and blister formation in AIBDs have
been described:
• in pemphigus disorders steric hindrance (the direct inhibition of protein-protein
binding by autoantibodies) and cell signaling events cause desmosomal instability,
• while complement and inflammatory cell activation mediated through Fc-signaling
cause keratinocyte death and blister formation in pemphigoid and epidermolysis
bullosa acquisita

4. • The pathogenesis of AIBDs can be divided into three phases:
• (i) the induction phase (loss of self-tolerance or the initiation of autoimmunity to
the target antigen),
• (ii) the maintenance phase (maintained production of autoantibodies) and
• (iii) the effector phase (autoantibody-mediated tissue damage).
• Specific mechanisms relating to these phases have been described for AIBDs,
including pemphigus disorders, BP, EBA, and DH.

5. INDUCTION OF AUTOIMMUNITY AGAINST SKIN
ANTIGENS
• There are multiple theories that explain how the loss of tolerance to self-antigens initially occurs
• majority of AIBDs are a product of several aberrant processes .
• Genetic factors play an important role, as specific skin blistering diseases have varying
prevalence in different populations and inherited human leukocyte antigen (HLA) types are
associated with autoreactivity to specific autoantigens
• Multiple HLA alleles have been identified which are associated with pemphigus vulgaris , BP ,
and EBA . class II HLA (e.g., HLA-DQβ1∗0301) BP Strong associations between MHC class II
haplotypes and anti-Dsg3 antibody production, such as HLADRB1*04:02 in Jewish populations ,
DQB1*05:03 in non-Jewish Caucasian populations , and DRB1*14 and DQB1*05:03 in Japanese
populations
• Genetic susceptibility is not limited to HLA types, as pemphigus vulgaris has been associated
with mutations in ST18 (a gene encoding a pro-apoptotic transcription factor) in certain
populations and experimental models of EBA have identified non-HLA murine gene loci that
confer susceptibility to disease development however further studies are required to extrapolate
these findings to clinical populations.

6. CELL DAMAGE
• Cell damage has been proposed as a common “triggering factor”— due to surgical
trauma UV radiation , neurological disorders and other pre-existing conditions , viral
infection , and radiotherapy have all been associated with disrupted skin barrier
function and development of AIBDs .
• Cell damage via necrosis or necroptosis releases a complex intracellular milieu into
the extracellular space which serves as a source of sensitizing autoantigens
additionally cell death results in the release of damage associated molecular patterns
which stimulate localized inflammation and wound healing processes .
• Normal healing responses following trauma cause infiltration of dendritic cells and
other antigen presenting cells which may also participate in autoimmune sensitization
of AIBDs.

7. EPITOPE SPREADING
• Epitope spreading is an inbuilt mechanism of the adaptive immune system that aids in
protecting against changing pathogens, however spreading from pathogenic to autologous
epitopes and molecular mimicry of similar epitopes may also contribute to the formation of
AIBDs .
• Fogo selvage, an endemic form of pemphigus foliaceus found in Brazilian populations, is
associated with a history of sand fly bites and characterized by autoantibodies against Dsg1.
These autoantibodies have shown cross reactivity to proteins present in sand fly saliva
• (PNP) pemphigus where tumor-associated antigens may become targeted in an effort to
destroy the tumor, however similar antigens may also be shared by keratinocytes
• PNP associated with lymphatic malignancies, including non-Hodgkin's lymphoma and chronic
lymphocytic leukemia., production and release of cytokines which can lead to over-stimulation
of humoral immunity and autoimmune reactions, including disruption of skin barrier and
development of AIBDs.
• Findings of autoimmune skin blistering in carcinoma patients has fuelled speculation that these
diseases may be triggered by an anti-tumor immune response
• Coeliac-disease associated skin blistering, known as DH, is caused by antibodies against gluten-
induced digestive enzyme tissue transglutaminase which undergo epitope spreading to cross-
react with epidermal transglutaminase (eTG) leading to the disruption of the skin barrier and
subsequent skin blistering .
• Epitope spreading may also contribute to the diversity of and disease progression of AIBds,

8. • AIBDs have been associated with the use of certain drugs which trigger pathogenesis through a
variety of mechanisms. One of the most well-described etiologies is BP in diabetic patients
taking dipeptidyl-peptidase 4 (DPP-4) inhibitors vildagliptin, teneligliptin, and linagliptin which
present with antibodies against the mid-portion of BP180.
• In addition, human leukocyte antigen-DQB1 was identified as the major haplotype in Japanese
DPP4i-BP
• It has been suggested that DPP-4 inhibition reduces plasmin production and alters BP180
cleavage, resulting in altered antigenicity of BP180 which is supported by the finding that
symptoms generally subside after drug discontinuation.
• The use of immune checkpoint inhibitors for cancer therapy has been linked to secondary
development of AIBD including mucous membrane pemphigoid and BP , likely a consequence
of nonspecific immune activation.
DPP4 is a member of the prolyl-oligopeptidase superfamily which cleaves incretins such as
glucosedependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1). The
suppression of DPP4 activity prolongs the GLP-1/GIP-dependent insulin secretion from
pancreatic beta cells that can be induced by increased serum glucose levels. Thus, DPP4i is
thought to have beneficial effects in reducing blood glucose levels without posing a
significant risk of hypoglycemia.

9. MAINTAINED AUTOANTIBODY PRODUCTION
• Autoantibodies are often present in healthy individuals but are generally non-pathogenic
IgM antibodies with low affinity present in low levels and do not alter skin barrier
homeostasis.
• For autoantibodies to gain pathogenicity, class-switching to IgG or IgA subtypes, somatic
mutation and increased production occur after exposure to self-antigen.
• Glycosylation and sialylation patterns on autoantibodies also contribute to pathogenicity
and antibodies stimulated via T-cell interaction within germinal centers exhibit reduced
sialyation patterns and are pro-inflammatory .
• In the context of AIBDs, the presence of self-antigen released by damaged keratinocytes
may stimulate production of autoantibodies, which in turn bind to healthy tissue to
stimulate skin barrier disruption and further tissue damage and promote more
autoantibody generation in a self-perpetuating cycle. Continued production of
pathogenic autoantibodies may be achieved by ongoing B cell activation and production
of short-lived plasma cells or the production of long-lived plasma cells which are
challenging to target .
• T-cell driven education within the germinal center appears to be a requirement for the
development of long-lived plasma cells, which gives rise to both lasting immunity against

10. • Murine models of EBA have also demonstrated the presence of plasma cells with
“intermediate” lifespans which contribute to autoantibody persistence .
• Alterations in cellular immune networks also contribute to maintained
autoantibody production and disease chronicity: Increased Th1 and Th17
cytokines and chemokines have been reported in patients with pemphigus
disorders , BP , and DH and
• changes in Treg populations are also associated with AIBDs with Tregs thought to
be protective against pathogenic autoantibody production .
• Higher frequencies of Th17 cells secreting IL-21 have been reported in
pemphigus lesions which form a tertiary lymph node like structure within the skin
and promote autoantibody production

11. • Desmogleins contain four cadherin repeats, EC1 to EC4, in their extracellular
domain. The N-terminal EC1 and EC2 domains are crucial for homophilic
adhesion and are the predominant regions targeted by auto-antibodies in
pemphigus , suggesting epitope-dependent determination of auto-antibody
pathogenicity. Indeed, pathogenic monoclonal anti-Dsg3 antibodies isolated from
PV patients and mouse models tend to target the N-terminus

12. • Desmogleins are expressed as intracellular precursor proteins that contain pro-
peptides on the external side of the N-terminus of the protein. After pro-peptide
cleavage, they are transported to the cell surface as the mature transmembrane
proteins that mediate cell adhesion.
• Interestingly, anti-Dsg1-reactive antibodies can be isolated from healthy donors.
However, these antibodies only react to the pro-peptide, but not the mature form, and
are thus non-pathogenic.
• Pathogenic auto-antibodies against the mature protein can be obtained only from PF
patients , suggesting an immunoregulatory mechanism that prevents the
development of pathogenic B cells in healthy individuals

13. International Immunology, Volume 31, Issue 7, July 2019, Pages 431–437, https://doi.org/10.1093/intimm/dxz030
The content of this slide may be subject to copyright: please see the slide notes for details.
FIG. 1. SCHEMATIC OVERVIEW. (A) PATHOGENESIS IN THE
PEMPHIGUS MURINE MODEL. * ACANTHOLYSIS IS A
PATHOLOGICAL CHANGE ...
Fig. 1. Schematic overview. (a) Pathogenesis in the
pemphigus murine model. * acantholysis is a pathological
change associated with pemphigus with intraepidermal
blistering. (b) Treg-mediated suppression of anti-Dsg3
antibody production in the pemphigus model. (c) Dsg3-
specific Th1 responses induce interface dermatitis in
mice. (d) Dsg3-specific Th17 responses induce
psoriasiform dermatitis in mice. (e) Ectopic expression of
epidermal autoantigens by squamous metaplasia allows
skin-specific T cells to attack the lungs in mice. This
mechanism is helpful to understand why bronchiolitis
obliterans is implicated in PNP, which can be
characterized as humoral and cellular autoimmunity to the
skin. (f) Central tolerance against Dsg3. Dsg3-specific
CD4⁺ T cells are deleted in the thymus in
the mice. (g) LCs expand Treg cell numbers and
suppress interface dermatitis in mice.

14. BP
the imbalance between autoreactive T helper (Th) and T regulatory (Treg) cells as well as a T cell-independent
activation of toll-like receptor (TLR) system may induce B cell stimulation, with consequent BP autoantibody
secretion. In parallel, the Th17 pathway activation appeared to maintain the inflammatory cascade started by
humoral hyperactivation, by triggering Th2 response, recruiting neutrophils and eosinophils and stimulating the
release of proinflammatory cytokines and proteolytic enzymes

15. • BP180 is the main BP
autoantigen and IgG
and/or IgE against this
antigen may be
detected in most
patients
• The immunodominant
non-collagenous
region 16A (NC16A) of
the BP180 ectodomain
is extensively
accepted to be the
main target for BP
autoantibodies.
• also domains of
BP180 outside of
NC16A, such as the C-
terminal domain of
BP180 and a 120-kDa
fragment of BP180
known as LAD-1,

16. • On direct immunofluorescence studiesIgGareusuallypredominant.
• Mihai et al. showed that IgG1 and IgG4 were the most frequent IgG subtypes
found in BP along the dermal-epidermal junction.
• By means of a cryosection assay, the same authors proved also that IgG1
promoted blister formation through complement fixation,
• whereas IgG4, which cannot fix the complement, activated leucocytes and
induced dermal-epidermal separation with a lower pathogenic potential as
compared to IgG1IgG directed against BP180 domains other than NC16A seemed
to be responsible of less inflammatory manifestations due to the fact that they
hardlyinduceanyBP180depletion
• Moreover, IgG reactivity with extracellular paralleled the severity of the disease
course

17. • higherAnti-BP180 IgE antibodies showed also pathogenic features; indeed, the
pathogenicity of anti-BP180 IgE antibodies has been demonstrated via an IgE
hybridoma to LABD97, a component of the shed ectodomain of BP180, which was
injected in severe combined immunodeficiency (SCID) mice with engrafted human
skin, with consequent development in the skin grafts of eosinophil infiltration and
histological subepidermal blisters resembling those of BP .
• Furthermore, the injection of total IgE isolated from two BP patients sera into human
skin grafted onto athymic nude mice induced erythematous plaques reminiscent of
those clinically seen in BP .
• Antibodies of the IgG class reacting against BP230,are usually found in a smaller
proportion of BP patients Although the actual pathogenic role of antiBP230
autoantibodies has been initially debated it has been more recently well-documented
and IgG reactivity with intracellular epitopes of BP230 has been shown to correlate to
BP severity . levels of anti-BP180 IgG were associated with increased 1-year mortality

18. LINKS BETWEEN BULLOUS PEMPHIGOID PATHOGENESIS
AND DISEASE-RELATED COMORBIDITIES
1.NEUROLOGIC AND NEURODEGENERATIVE DISORDERS
• Several epidemiological studies have confirmed that patients affected by
neurologic and neurodegenerative diseases, such as multiple sclerosis and
Alzheimer’s disease, have an increased risk of developing BP .
• Since BP180 is expressed at low levels in neuronal tissue ,
• neurodegeneration or neuroinflammation lead to loss of self-tolerance toward
BP180 and consequent BP development

19. 2.VITAMIN D HAS BEEN SHOWN TO HAVE AN IMMUNE
REGULATORY FUNCTION AND HYPOVITAMINOSIS D HAS
BEEN CORRELATED TO AN INCREASED RISK AIBD
• higher prevalence of vitamin D deficiency and lower 25-hydroxyvitamin D levels were
found in BP patients as compared to controls, inversely correlated to disease severity
in one study
• D3 has been revealed to elicit down-regulation of the BP230 gene through post-
transcriptional mechanisms independently by active protein synthesis
• non-toxic doses of calcitriol, reduced the release of the proinflammatory cytokines IL-
6 and IL-8 induced by purified human BP IgG from HaCaT cells
• A similarly beneficial effect of calcitriol has been recently reported also in mice with
experimental EBA

20. 3.THROMBOSIS
• a recent multicenter cohort study showing that BP patients have a 4-fold increased risk of
developing veno thromboembolism
• D-dimer and F1+2, two coagulation activation markers, have been observed to be increased in
blister fluid, lesional skin and plasma (90), with F1+2 correlating to the anti-BP180 levels
• extrinsic blood coagulation pathway activation may be regarded as a contributory mechanism
leading to local inflammation and blister formation in BP
• Fibrinolysis is inhibited in BP, chiefly due to increased plasminogen activator inhibitor type 1
(PAI-1) activity and plasma levels.
• recent observation that serum levels of soluble P-selectin, a marker of platelet activation, were
significantly higher in BP patients than controls led to hypothesize a role of platelets in disease
pathogenesis

21. AUTOANTIBODY-INDUCED TISSUE DAMAGE IN DIFFERENT CLINICAL
SETTINGS (A) Pemphigus disorders are caused by autoantibodies against
desmoglein (Dsg) proteins Dsg1 and Dsg3. Binding of anti-Dsg
destabilizes desmosomes to cause acantholysis of keratinocytes
within the epidermis and triggers keratinocyte signal transduction
events which promote inflammation, skin barrier disruption and
further skin blistering. (B) Blisters in bullous pemphigoid are caused
by anti-BP180 antibodies which bind hemidesmosomes on basal
keratinocytes and trigger complement activation and inflammatory
responses including ROS and protease release by neutrophils which
directly kill keratinocytes. Skin barrier disruption and skin blistering
is caused by keratinocyte death and sustained localized
inflammation. (C) Epidermolysis bullosa acquisita is caused by anti-
Collagen VII antibodies which bind fibrils that anchor
hemidesmosomes to the basement membrane. Deposition of IgG
induces complement activation via the classical pathway and
activation of neutrophils. Basal keratinocytes sustain damage via
action of neutrophil-derived ROS and proteases resulting in splitting
at the dermal-epidermal junction and skin barrier
disruption. (D) Dermatitis hepetiformis (DH) is caused by cross-
reactive antibodies that bind epidermal transglutaminase (eTG). eTG
is produced by keratinocytes and accumulates in the papillary dermis
where it forms immunogenic immune complexes with anti-eTG IgA
that trigger complement activation. Fibrin deposition and influx of
leukocytes (which damage keratinocytes via release of ROS and
proteases) cause the formation of neutrophilic abscesses which
develop into fluid-filled subepidermal blisters that disrupt the intact
skin barrier.
preclinical animal studies and some clinical evidence
elucidating the contributions of eosinophils , mast cells , Th17
and Treg cells to AIBD's, and these may represent novel
therapeutic targets.

22. • Unraveling the precise mechanisms of autoantibody-induced pathology has been
the focus of much research in recent years.
• Autoantibodies against Dsg proteins, as found in pemphigus diseases, were
initially thought to interfere with Dsg-Dsg interactions in desmosomes by steric
hindrance however evidence for
• direct effects of Dsg-anti-Dsg binding on intracellular signaling events was later
discovered, including the p38 mitogen-activated protein kinase pathway leading
to acantholysis.
• Anti-Dsg may also reduce the number of desmosomes by clustering Dsg on the
cell surface and interfering with normal turnover of desmosomal proteins,
thereby depleting desmosomes of Dsg and promoting acantholysis.
• In more recent years, other autoantibody species and non-Dsg interactions have
been identified as contributing to pemphigus pathology and have prompted the
hypothesis that multiple pathways may act synergistically to cause classical
pemphigus disease pathology

23. • BP, autoantibodies against BP180 and BP230 components of hemidesmosomes
produce blistering at the dermal-epidermal junction. Activation of the
complement cascade via classical and alternative pathways has been
demonstrated to contribute to skin barrier disruption and BP pathology and C3
deposition at the epidermal basement membrane is a common clinical finding .
• Complement activation induces inflammation and damages keratinocytes via
cytotoxic action of neutrophils leading to skin blistering and barrier disruption.
• Though complement-independent mechanisms of BP pathology have been since
described including direct activation of neutrophils via immune complex-FcγR
binding, complement activation is still a prevalent target for novel BP
therapeutics
• Autoreactive IgE and eosinophilia are common findings in BP patients and IgE
immune complexes binding and activating eosinophils thereby contributing to
blister formation has been shown in in vitro and in vivo studies , implicating
eosinophils as potential therapeutic targets in BP.

24. • Disease pathology in EBA is caused by predominantly IgG1 and IgG3 autoantibodies
that bind Collagen VII within anchoring fibrils at the dermal-epidermal junction.
• Like BP, complement activation is considered key to EBA pathogenesis however the
alternative pathway appears to be the dominant pathway behind experimental EBA
pathology
• Activation of complement induces inflammation, leukocyte extraversion, complement
activation and subsequent tissue damage and disruption of skin barrier via release of
ROS and proteases from neutrophils and other myeloid cells.
• Ex-vivo studies of patient serum incubated with healthy skin and donor neutrophils
exhibit loss of epidermal adherence, hence clearly indicating that antibodies mediate
clinical EBA blistering via neutrophil activation . The role of T cells in EBA pathology is
yet to be fully elucidated, however murine studies show that NKT and γδT cells likely
amplify tissue damage in EBA via interaction with immune complexes and neutrophils

25. • DH is characterized by accumulation of ant-eTG IgA antibodies within the
papillary dermis, however eTG is primarily expressed within superficial epidermal
layers .
• It is hypothesized that eTG may be released into the blood, where interaction with
IgA occurs in nearby dermal vessels; alternatively eTG may be deposited along
the basement membrane as a result of trauma, however further research is
required to confirm these hypotheses.
• Following IgA deposition, papillary abscesses characterized by a neutrophilic
infiltrate and fibrin accumulation form which develop into a split at the basement
membrane and subepidermal blistering.
• Patients with DH show reduced levels of anti-inflammatory IL-10 and reduced
Treg cell numbers in lesional skin compared to healthy skin which indicates the
role of Tregs in modulating local inflammatory responses in DH

28. important role for C5a-C5aR1-axis activation in the
development of BP