The important role of laminin 332 and of the α6β4 integrin in epidermal–dermal adhesion was early confirmed by the finding that mutations affecting either of these molecules cause JEB, a genetic disorder that affects skin and mucous membranes [17]. It is characterised by a mesenchymal–epithelial separation within the lamina lucida and by hemidesmosomal abnormalities. JEB is generally divided into three subtypes: Herlitz (lethal), non-Herlitz (nonlethal) and JEB with pyloric atresia [4, 29, 35, 114]. All of them are inherited in an autosomal recessive manner [93] except for a single case recently reported [2]. Herlitz JEB is caused by a complete absence of laminin 332 [6, 65, 81], while non-Herlitz is caused by missense mutations leading to a reduction in functional laminin 332 or complete absence of collagen XVII [6, 65, 81, 115]. JEB with pyloric atresia is caused by a genetic mutation of α6 or β4 subunits that are the main receptors for laminin 332 located beneath HDs [91, 114]. The majority of mutations identified to date in both H-JEB and nH-JEB reside in the three genes that encode the αβγ chains of laminin 332, LAMA3α, LAMB3 and LAMC2, respectively [74, 111]. Normally, these three genes are expressed during all developmental stages of the skin including all steps of adhesion, proliferation and differentiation [31]. Nonsense or frameshift mutations of any of the three laminin 332 genes causing a premature termination codon (PTC) usually lead to a decrease in transcript levels due to nonsense-mediated mRNA decay [40]. If such mRNAs are translated, truncated polypeptides are produced that are incapable of participating in the formation of a functional heterotrimer. Other mutations that may occur in the laminin 332 genes include missense mutations, exon-skipping mutations and in-frame insertions or deletions. In these cases, translation of the resulting mRNA results in structurally imperfect protein production, which still allows secretion of more or less functional laminin 332 [96]. Most mutations related to the JEB are found in the LAMB3 gene and mainly in the exons encoding the N-terminal part of the β3 chain. The LAMB3 portion coding for the C-terminal α-helical domain, which is involved in subunit assembly, contains much less mutations [73, 97].

There are particularly two recurrent “hotspot” LAMB3 mutations, R635X and R42X, leading to PTCs which account for half of all LAMB3 mutations and result from a C-to-T transition [49, 73, 82]. In the LAMC2 gene, most of the mutations are found within the LE domain and the L4 module and rarely in the C-terminal α-helical domain [96]. Some mutations are also found within the LG1–LG5 globular domains of the LAMA3 gene. Particularly, the LG3 mutation K1299X, which affects the charged amino acid lysine at position 1299, is related to a severe phenotype seen in the homozygous human patients [45, 73, 78]. As the α3 chain is found in laminins 311 and 321, mutations in LAMA3 gene also affect these two laminins’ functions [96]. Based on the clinical severity, JEB patients with laminin 332 mutations have been classified into the Herlitz and non-Herlitz types, and there are some general “rules” regarding phenotype–genotype correlations [30]. In general terms, patients harbouring PTC causing mutations on both alleles (homozygous or compound heterozygous) suffer from the severe H-JEB [1, 81]. As a result of these mutations, the synthesis of one laminin 332 chain is abolished and no functional trimeric laminin 332 is produced [66, 80, 83]. The PTC mutations in H-JEB result in the absence of laminin 332 within the dermal–epidermal junction as confirmed by staining with laminin 332–specific monoclonal antibodies [39, 68, 113]. Examination of skin biopsies from H-JEB patients by electron microscopy reveals small rudimentary or absent HDs, as well as cleavages through the lamina lucida [103]. The fact that laminin 332 is involved in the regulation of motility and proliferation of keratinocytes predicts that its absence can affect the normal wound-healing process causing chronic erosions and formation of granulation tissue. All resulting laminin 332 defects cause a generalised blistering and early death of the patient, usually during the first year of life. The epidermis of an affected infant can be extensively peeled away only on simple handling, while in many cases there is death by overwhelming infection, caloric deprivation or from organ failure deriving from complications of the disease [73]. nH-JEB patients often carry a PTC on one allele and a missense or in-frame splice site mutation on the alternate allele. In these cases there is often a milder phenotype due to the production of partial or abnormal laminin 332. This is in accordance with immunofluorescence mapping studies showing diminished expression of laminin 332 within the dermal–epidermal junction of these patients [116]. The nH-JEB phenotype is characterised by lifelong blistering in a distribution that predominates in sites exposed to friction, trauma or heat, atrophic scars, hypopigmentation or hyperpigmentation at sites of healed blisters, incomplete alopecia, dystrophic nails and dental abnormalities [24, 30, 116]. The course and prognosis of nH-JEB depend on how severely affected the target protein is. Thus, some patients have a mild phenotype and normal lifespan (nH-JEB localised), and others have a severe disease and an increased risk of squamous carcinoma development and death (nH-JEB generalised) [46, 47].

Mucous membrane pemphigoid (MMP) is a heterogeneous group of rare autoimmune blistering diseases and is characterised by the presence of autoantibodies to various components of the dermal–epidermal anchoring complex causing erosive lesions followed by scarring of the skin and mucous membranes [50]. On the basis of the 2002 international consensus [19], MMP includes blistering diseases with preferential mucous membrane involvement. Patients with MMP may exhibit linear deposits of IgG and/or IgA autoantibodies and/or complement fragments in epithelial basement membranes. Subtypes of MMP correspond to the nature of the antigen targeted such as BP230 and BP180, laminin 332 and laminin 311, type VII collagen or the α6 and β4 integrin subunits [see Caux and Prost, Part III, Chapter 14; 50]. Early studies revealed that laminin 332 is a target antigen in about 25 % of MMP patients and most patients have IgG antibodies against the α-subunit of the heterotrimer [48, 55] that recognise both laminin 332 and laminin 311 [20]. The pathogenicity of anti-laminin 332 antibodies has been documented in vitro and in vivo. Passive transfer of anti-laminin 332 IgG to neonatal or adult mice induced subepidermal blisters of the skin and mucous membranes that mimicked clinical, histological and immunopathologic features seen in MMP patients [58]. Mice injection of Fab fragments directed against laminin 332 produced the same results as well as injection in mice lacking complement, mast cells or T cells, suggesting that the antibodies induced epidermal detachment in a noninflammatory and direct manner [53, 58]. An experimental human skin graft model was also used and revealed that patients’ anti-laminin 332 autoantibodies induced subepidermal blisters [54]. Assaying for anti-laminin 332 reactivity is of particular importance as 25 % of patients with laminin 332-specific antibodies are suspected to develop a malignancy [28, 59]. While direct immunofluorescence microscopy of patients’ epidermis and mucosal epithelium is still the gold standard for the diagnosis of pemphigoid diseases, diagnosis can be made serologically today [57, 95]. Patients with anti-laminin 332 antibodies display circulating IgG autoantibodies that bind to the dermal side of 1 M salt-split skin by indirect immunofluorescence [59]. Western blot analysis has revealed that anti-laminin 332 autoantibodies most often react with the α3 chain [48, 55] and less frequently with the β3 and/or the γ2 chain [33, 34, 55, 64, 102]. However, the exact mechanism is not known yet; it is most likely that antibodies against the α3 chain are disrupting the interaction with the keratinocyte integrins. Moreover, it is possible that autoantibodies against the β3 and γ2 chains may disrupt the interaction between laminin 332 and type VII collagen, as laminin-β3 is thought to mediate this binding [21, 87]. Neither specific epitopes nor laminin domains targeted by anti-laminin 332 autoantibodies have been precisely identified so far, and the use of recombinantly produced laminin domains will probably be helpful in such autoantibodies’ characterisation. Immunoprecipitation with radiolabelled human keratinocytes has been reported to be the most sensitive technique for the detection of serum autoantibodies against laminin 332, and immunoblotting using human keratinocytes’ ECM or purified laminin 332 was used as a convenient method to identify the laminin subunit involved [20, 41, 43, 56]. Enzyme-linked immunosorbent (ELISA)–based assays will probably soon replace these time- and reagent-consuming methods. Some studies conducted so far have reported patients’ serum samples analysis in ELISA assays using either purified laminin 332 or keratinocyte ECM as a substrate [11, 12, 56]. The most recent study [12] reported the analysis of serum samples from 154 patients with MMP and 89 control individuals. The authors found that 20 % of the MMP patients tested had serum anti-laminin 332 autoantibodies. Interestingly, these appeared to be present in a subset of patients with a severe form of the disease.