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S E C T I O N

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DISORDERS OF EPIDERMAL AND DERMAL-EPIDERMAL COHESION AND VESICULAR AND BULLOUS DISORDERS

C H A P T E R 5 1

Epidermal and Epidermal-Dermal Cohesion

Leena Bruckner-TudermanJohn R. Stanley

The cell-cell and cell–basement mem-brane adhesion in the epidermis providesthe skin with its resistance against envi-ronmental influences; epidermal integrityis required for protection of the entireorganism against mechanical, physical, ormicrobial noxae. The major cellular struc-tures involved are the desmosomes atcell-cell junctions in the epidermis andthe hemidesmosome–basement mem-brane adhesion complexes and relatedstructures at the epidermal-dermaljunction. Ultrastructurally, the hemides-mosome closely resembles one-half ofthe desmosome; however, at the molecu-lar level, these two structures are distinct.Both represent specifically organized as-semblies of intracellular and transmem-brane molecules, which on one hand an-chor cytoskeletal filaments to the cellplasma membrane and on the other handthe plasma membrane to extracellularstructures, i.e., another cell or the ex-tracellular matrix. Our knowledge ofthe desmosomal, hemidesmosomal, andbasement membrane molecules has ex-panded drastically in the last 10 to 15years due to the great power of both mo-lecular genetics and proteomics. Aftersome of the proteins were initially identi-fied as autoantigens in pemphigus andpemphigoid, a multitude of moleculeshave now been characterized at both pro-tein and gene levels, and their expression,

regulation, and functions have been dis-cerned. The antigenic epitopes in differ-ent autoimmune blistering skin diseaseshave been carefully mapped and, to date,mutations in at least 18 different geneshave been shown to underlie heritabledisorders of epidermal or epidermal-dermal adhesion in humans and mice.Morphologic, molecular and functionalaspects of these adhesion structures aredelineated in this chapter.

EPIDERMAL COHESION

Ultrastructure of Desmosomes

Desmosomes (also called

maculae ad-haerentes

) are the major cell adhesionjunction of epithelial tissues; however,they are also found in myocardium anda few other tissues.

1–3

Not only do theyserve as a sort of spot weld of the kerat-inocytes but they also anchor interme-diate filaments (keratin in the case ofepidermis and mucous membranes) tothe cell surface. In serving these func-tions, they form a three-dimensionalscaffolding of the intermediate filamentsthat are stretched from the nuclear en-velope to the cell surface.

4

This scaffold-ing is critical to stabilize epidermalstructure in the face of external trauma,and its collapse results in cytolysis ofkeratinocytes (see Chaps. 47 and 60).

Ultrastructural cross-section of thedesmosome shows a symmetric struc-ture consisting of two apposing denseplaques inside the membranes that arethemselves separated by approximately30 nm (Fig. 51-1A).

5

This intercellularcenter has been called the

desmoglea

(from Greek, “desmosomal glue”), be-cause it is thought to provide the adhe-sion that keeps cells together at thisjunction.

6

In high resolution, there is athin electron dense midline in the centerof the desmoglea. The dense plaque in-side the cell actually is formed of an

outer very dense plaque bordering themembrane, and an inner less denseplaque that intertwines with keratin in-termediate filaments.

Biochemical Characterization of Desmosomes

The molecular components of the vari-ous desmosomal substructures havebeen biochemically characterized andcloned (see Fig. 51-1B).

4

These proteinsfall into three major gene families:plakins (e.g., desmoplakin), armadilloproteins (e.g., plakoglobin and plakophi-lins), and desmosomal cadherins (des-mogleins and desmocollins). Plakins andarmadillo proteins are in the plaque, anddesmosomal cadherins are transmem-brane proteins whose extracellularamino-termini comprise the desmoglea.The major inner plaque component is

desmoplakin

, which exists in two RNAsplice variants, desmoplakin I and II.

7,8

Desmoplakin is part of the plakin genefamily,

9

which includes bullous pemphi-goid antigen 1 and plectin, plaque pro-teins of the hemidesmosome, as well asenvoplakin and periplakin, whose func-tions in the desmosome are not as yetwell defined. Similar to other desmo-somal components, desmoplakin is amodular protein, with different modulesfulfilling different functions. The centralpart of one desmoplakin molecule coilsaround the central part of another to forma rod-like center. The carboxy-terminusbinds to keratin filaments,

10

and theamino terminus binds to plakoglobin.

11

Desmoplakin, therefore, provides themajor link between the keratin filamentsand the desmosomal plaque.

Plakoglobin

is the second major proteinin desmosomal plaques.

12

A member ofthe armadillo gene family that also in-cludes plakophilins,

13

it is a modular pro-tein with repeating homologous units.

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The first indication of plakoglobin bind-ing to a desmosomal protein (desmo-glein) came from studies with pemphigus(see Chap. 52) sera in which plakoglobinwas co-precipitated with desmoglein byboth pemphigus vulgaris and foliaceussera.

14

Desmoglein 3

and 1 are the pemphi-gus vulgaris and foliaceus antigens, re-spectively, that the sera bound, and plako-

globin was co-precipitated because of itsbinding to these desmogleins. Various do-mains of plakoglobin modulate its bind-ing to desmogleins and desmocollins.

15–17

Other domains bind to desmoplakin,thus linking desmogleins and desmocol-lins to desmoplakin.

11

Plakophilins

, addi-tional plakin proteins in the desmosome,bind various desmosomal components,and may aid in clustering and stabilizingthem.

18,19

Finally, desmogleins and desmocol-lins provide the link between the extra-cellular desmoglea and the intracellularplaque.

20–22

These glycoproteins are inthe cadherin supergene family, and,similar to other members of this family,are thought to mediate calcium-depen-dent adhesion. In humans, there arefour genes encoding desmogleins 1, 2,3, and 4. The desmoglein 3 gene wasactually discovered and cloned becauseit was the antigen in pemphigus vul-garis (see Chap. 52).

23

These isoformshave different tissue distributions withdesmogleins 1 and 3 being predominantin epidermis and mucous membranes.Within these tissues, the desmogleinsalso have unique distributions, whichhas major implications for understand-ing the pathophysiology of pemphigus(see Chap. 52). Similarly, there are fourgenes encoding desmocollins I, II, III,and IV, which in addition have alterna-tive RNA splice variants, and, similar todesmogleins, each have unique tissue

specific distributions.

21,24–26

These des-mosomal cadherins play a major role inproviding cell adhesion in the epidermisand mucous membranes. Similar to theother desmosomal components, des-mosomal glycoproteins are modularwith different parts of the moleculesproviding different functions. Their cy-toplasmic tails bind plakoglobin andother armadillo proteins and their ex-tracellular domains provide adhesionwith desmosomal cadherins expressedon apposing cells.

Function and Pathology of Desmosomes

As described in Biochemical Character-ization of Desmosomes, the major func-tion of desmosomes is thought to be celladhesion. Animal models, autoimmuneskin diseases, and the blisters caused byexfoliative toxin from staphylococciconfirm this function (Table 51-1; seeChaps. 52 and 178).

27

For example, amouse genetically engineered with atargeted deletion of desmoglein 3 devel-ops mucous membrane and skin blistersfrom loss of cell adhesion in the deepepidermis, similar to pemphigus vul-garis patients.

28

Patients with pemphi-gus vulgaris and foliaceus develop lossof cell adhesion and resultant blisters inthe epidermis due to anti-desmoglein 3and 1 antibodies, respectively (Fig. 51-2).

EPIDERMAL AND EPIDERMAL-

DERMAL COHESION

AT A GLANCE

■

Desmosomes, hemidesmosomes, and the epidermal basement membrane are providers of epidermal and epidermal-dermal cohesion.

■

They represent distinct molecular adhe-sion complexes.

■

The major components of desmosomes belong to three major gene families: the plakins, armadillo proteins, and desmo-somal cadherins.

■

The hemidesmosomal components com-prise plakin homologues, integrins, and collagenous transmembrane proteins.

■

All basement membranes contain collagen IV, laminins, nidogens, and perlecan.

■

Functional specificity of basement mem-branes is provided by additional tissue-specific glycoproteins.

■

In addition to their structural roles, des-mosomes, hemidesmosomes, and the epidermal basement membrane are bio-logically active in cellular signaling.

■

Mutations in the genes encoding the above proteins cause hereditary skin dis-eases; ranging from hypotrichosis and keratoderma to epidermolysis bullosa and Kindler syndrome.

■

Protein components of desmosomes, hemidesmosomes, and epidermal base-ment membrane are targeted in autoim-mune skin blistering diseases of the pemphigus or pemphigoid group and in epidermolysis bullosa acquisita.

�

FIGURE 51-1

Electron microscopic image (

A

) and simplified schematic diagram (

B

) of a desmosome(not drawn to scale). dg = desmoglea; dm = dense midline; dp = desmoplakin; dsc = desmocollins; dsg= desmoglein; idp = inner dense plaque; kf = keratin filaments; odp = outer dense plaque; pg = plako-globin; pkp = plakophilins; pm = plasma membrane. (Electron micrograph used with permission fromKathleen Green and with permission of Elsevier, adopted from Yin T, Green KJ: Regulation of desmosomeassembly and adhesion.

Semin Cell Develop

Biol

15

:665, 2004.)

A B

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Exfoliative toxins from staphylococcispecifically cleave desmoglein 1 to causeblisters from loss of cell adhesion in thesuperficial epidermis in bullous impe-tigo and staphylococcal scalded skinsyndrome (see Chap. 178).

29

More recent data suggests that thefunction of desmosomes is much morecomplicated than simple cell adhesion.For example, desmosomal molecules areimportant in development and differen-tiation.

30,31

A targeted genetic deletionof desmoglein 2 causes early embryoniclethality in mice, and prevents prolifera-tion of embryonic stem cells.

32

Inappro-priate expression of desmoglein 3 in theupper epidermis results in hyperprolif-eration, abnormal differentiation, andbarrier defects.

33,34

Mice with a targeteddeletion of desmocollin I also have ab-normal differentiation and a defectiveepidermal barrier.

35

In addition, the ar-madillo components of the desmosomemay mediate or modulate signal trans-

duction.

31

The original member of thearmadillo family,

Armadillo

, was discov-ered in Drosophila as a member of theWnt signaling pathway. Another mem-ber of the family,

β

-catenin, is both acomponent of the adherens junction,another cell adhesion junction, andplays a critical role in Wnt signaling inepithelial cells.

β

-Catenin, in this laterfunction, translocates to the nucleus andcomplexes with other molecules toform a transcription factor. Plakoglobin,a member of this family, may also servea similar dual role (adhesion and signal-ing), but exactly how remains to be de-termined. Similarly, plakophilins maylocalize to both desmosomes and trans-locate to the nucleus, suggesting theyalso may be involved in signaling andtranscription.

36–38

These complexities of the function ofdesmosomal components may accountfor the various disease manifestations ofpatients with mutations in desmosomegenes (see Table 51-1).

39,40

Desmoglein 1and desmoplakin mutations are associ-ated with striate palmar plantar kerato-derma.

41,42

Desmoglein 4 mutations arefound in autosomal recessive hypotricho-sis.

43

Autosomal recessive desmoplakinmutations can cause not only kerato-derma but woolly hair and left ventric-ular cardiomyopathy.

44

Plakoglobinmutations result in Naxos disease,keratoderma, woolly hair, and arrhyth-mogenic right ventricular cardiomyopa-thy.

45

Plakophilin 1 mutations result in adisease that includes skin fragility and ec-todermal dysplasia.

46

Interestingly, someof these genetic diseases of different des-mosomal components have overlapping

clinical features, suggesting that theymay share certain final common patho-physiologic pathways.

The exact pathophysiologic mecha-nisms relating the mutations in thesegenetic diseases to the phenotypes havenot been well worked out. However,these diseases clearly indicate the im-portance of desmosomes and their com-ponents beyond just cell adhesion.

EPIDERMAL-DERMAL COHESION

Structural and Functional Characteristics of Basement Membranes

Basement membranes underlie epithelialand endothelial cells and separate themfrom each other or from the adjacentstroma. Another form of basement mem-brane surrounds smooth muscle or nervecells. The physiologic functions of base-ment membranes are diverse: In the vari-ous organ systems they provide supportfor differentiated cells, maintain tissue ar-chitecture during remodeling and repair,and, in some cases, acquire specializedfunctions, including the ability to serve asselective permeability barriers (e.g., theglomerular basement membrane or theblood-brain barrier) or acquire strong ad-hesive properties, like the basement mem-brane at the dermal-epidermal junction, orthat surrounding smooth muscle cells,which provide the tissues resistanceagainst shearing forces. All of these charac-teristics of basement membranes are alsoused during development and differentia-tion of multicellular organisms (Box 51-1).

TABLE 51-1

Desmosomal Targets in Skin Disease

D

ESMOSOME

C

OMPONENT

A

UTOIMMUNE

T

ARGET

G

ENETIC

T

ARGET

Desmosomal cadherins Desmoglein 1

a

PF SPPK (AD)Desmoglein 3 PV, PNPDesmoglein 4 PF,

b

PV

b

AR hypotrichosisDesmocollins SPD

47

Desmosomal plaque proteins Desmoplakin PNP SPPK (AD); keratoderma, woolly hair, left ventricular cardio-myopathy (AR)

Other plakins

c

PNPPlakoglobin Naxos disease: keratoderma, woolly hair, right ventricular

cardiomyopathy (AR)Plakophilins Plakophilin 1: skin fragility and ectodermal dysplasia (AR)

Stratum corneum desmosome protein

Corneodesmosin Hypotrichosis simplex of the scalp (AD)

47

a

Also targeted by exfoliative toxin in bullous impetigo and staphylococcal scalded-skin syndrome.

b

Anti-desmoglein 1 in these sera cross-react with desmoglein 4.

c

Including envoplakin, periplakin, and bullous pemphigoid antigen 1.AD = autosomal dominant; AR = autosomal recessive; PF = pemphigus foliaceus (see Chap. 52); PNP = paraneoplastic pemphigus (see Chap. 53); PV = pemphigus vulgaris (see Chap. 52); SPD = subcorneal pustular dermatosis (see Chap. 34); SPPK = striate palmoplantar keratoderma (see Chap. 48).

�

FIGURE 51-2

Indirect immunofluorescencestaining of human epidermis with pemphigusserum that contains autoantibodies to desmo-glein 3.

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Ultrastructurally, basement mem-branes most often appear as trilaminarstructures, consisting of a central elec-tron-dense region, known as the

laminadensa

, adjacent on either side to an ap-parently less-dense area, known as the

lamina lucida

or

lamina rara

. The laminalucida directly abuts the plasma mem-branes of the adherent cells. The rela-tive size of each of these regions variesamong basement membranes of differ-ent tissues, among the basement mem-branes of the same tissue at differentages, and as a consequence of diseases.For example, the trilaminar glomerularbasement membrane in humans variesfrom 240 nm to 340 nm in width,whereas the bilaminar basement mem-brane of the dermal-epidermal junctionmeasures 50 nm to 90 nm. This ultra-structure demonstrates that basementmembranes serve as substrates for theattachment of cells and fix their polarity.Their continuity throughout the variousorgan systems stabilizes the establishedtissue orientations and provides a tem-plate for orderly repair after traumaticinjury. Major disruptions in the base-ment membrane result in the formationof scar tissue and the loss of function inthat area.

Different basement membranes con-tain both common and unique compo-nents. All share a basic network struc-ture to which specific macromoleculeshave been appended. These moleculesare responsible for the specialized struc-tures and functions of different base-ment membranes. The basic constitu-ents of these structures are collagen IV,laminins, nidogens, and proteoglycansof the perlecan type, which all are highlyconserved, although the isoforms, thenumber of the polypeptide subunits, andtheir individual structures vary amongspecies.

47,48

The nearly ubiquitous distri-bution of heparan sulfate proteoglycansin all basement membranes suggeststhat these serve as selective permeabilitybarriers in multiple locations, includingthe kidney and the blood-brain barrier.Ultrafiltration may be especially impor-

tant during development and morpho-genesis of all tissues.

Basement membranes also providephysical separation between epitheliaand their underlying extracellular matri-ces. This barrier is especially importantin the containment of tumors. With theexception of certain cells of the immunesystem, non-malignant cells seldomcross a basement membrane. Benign tu-mor cells, when placed on isolated am-niotic basement membranes, are unableto cross this barrier in vitro. In contrast,malignant cells bind the basementmembrane, regionally disrupt its struc-ture at the site of the attachment, andmigrate through the rupture. Lamininsand integrins mediate the tumor-cellbinding, and the basement membranedissolution is catalyzed by metallopro-teases produced by the tumor cell. Theabsence of distinguishable basementmembranes in tumor biopsies has beenan indicator of malignancy, and thereappears to be a high correlation be-tween metastasis and basement mem-brane degradation. These observationsunderline the importance of the basallamina as an obstacle to cell migration.

By binding biologically active signal-ing molecules, basement membranesregulate a multitude of biologic events.The constituent proteoglycans can bindgrowth factors that can be released fromthe complexes by tissue proteinases.Thus, the basement membranes are po-tent regulators of cell adhesion and mi-gration, cytoskeleton and cell form, celldivision, differentiation and polariza-tion, and apoptosis.

49

ULTRASTRUCTURE OF THE DERMAL-EPIDERMAL JUNCTION

The dermal-epidermal junction is an ex-ample of a highly complex form of base-ment membrane,

50,51

which underlies thebasal cells and extends into the upper lay-ers of the dermis (Fig. 51-3A). This base-ment membrane is continuous along theepidermis and skin appendages, includingsweat glands, hair follicles, and sebaceous

glands. The dermal-epidermal junctioncan be divided into three distinct zones.The first zone contains the keratin fila-ment-hemidesmosome complex of thebasal cells and extends through the laminalucida to the lamina densa. The plasmamembranes of the basal cells in this regioncontain numerous electron-dense platesknown as hemidesmosomes. The intra-cellular architecture and organization ofthe basal cells are maintained by keratinintermediate filaments, 7 to 10 nm in di-ameter that course through the basal cellsand insert into the desmosomes andhemidesmosomes. External to the plasmamembrane is a 25- to 50-nm–wide laminalucida that contains anchoring filaments, 2to 8 nm in diameter, originating in theplasma membrane and inserting into thelamina densa. The anchoring filamentscan be seen throughout the lamina lucidabut they are concentrated in the regions ofthe hemidesmosomes. Ultrastructurally,the anchoring filaments appear to securethe epithelial cells to the lamina densa.

The existence of the lamina lucida invivo has been questioned. When the ul-trastructure of the basement membraneis evaluated after high-pressure preser-vation techniques, the lamina densa ap-

�

FIGURE 51-3 A.

Ultrastructure of the humandermal-epidermal junction as visualized by trans-mission electron microscopy after standard fixationand embedding protocols. af = anchoring fila-ment; AF = anchoring fibril; AP = anchoringplaque; BM = basement membrane; Hd = hemi-desmosome; Ld = lamina densa; Ll = lamina lu-cida. (Bar = 200 nm.)

B.

Ultrastructure of the hu-man dermal-epidermal junction by transmissionelectron microscopy following protocols usinghigh-pressure fixation and embedding techniques.Note the dense character of both the basementmembrane and the subjacent papillary dermis. An-choring filaments, anchoring fibrils, and anchoringplaques are not distinguishable. (Both photos usedwith permission from Douglas R. Keene, MD, Shri-ners Hospital, Portland, Oregon.)

A

B

Box 51-1

Major Functions of Basement Membranes

■

Scaffold for tissue organization and template for tissue repair.

■

Selective permeability barriers. The renal basement membranes serve for the ultrafiltration of plasma, and other basement membranes also demonstrate selective filtration.

■

Physical barriers between different types of cells or between cells and their underlying extracellu-lar matrix.

■

Firmly link an epithelium to its underlying matrix or to another cell layer and provide polarity.

■

Regulate cellular functions.

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451

pears intimately associated with theepithelial cell surface.

50,52

When the der-mal-epidermal junction is similarly pre-pared, no distinct lamina lucida is seen(see Fig. 51-3B). This suggests that thelamina lucida may result from shrinkageof the cell surface away from the laminadensa due to dehydration. The appear-ance of anchoring filaments spanningthe lamina lucida may then result fromthe firm attachment of constituents of thelamina densa at the hemidesmosomethat is subsequently pulled from thelamina densa by shrinkage. Other com-ponents that are also tightly fixed to thekeratinocyte plasma membrane, eitherat the hemidesmosomes or at other sitesalong the membrane between thehemidesmosomes, may similarly be-come displaced into the shrinkagespace. Regardless of its actual occur-rence in vivo, the evaluation of thelamina lucida by standard electron mi-croscopy techniques has allowed identi-fication of specific structures that wouldotherwise have been difficult to detect.In addition, the morphologic term

lam-ina lucida

remains practical in the scien-tific communication and continues tobe used.

The second zone, the lamina densa,appears as an electron-dense amor-phous structure 20 nm to 50 nm inwidth. The dermal-epidermal laminadensa is similar in appearance to analo-gous structures in other organs. At highmagnification, it has a granular-fibrousappearance.

50

The major molecularcomponents of the lamina densa are col-lagen IV, nidogen-entactin, perlecan,and laminins, which all can polymerizeto networks of variable thickness.

53

The subbasal lamina contains mi-crofibrillar structures. Two of these arereadily distinguishable. The first struc-ture, known as

anchoring fibrils

, appearsas condensed fibrous aggregates 20 nmto 75 nm in diameter.

54

At high resolu-tion, they appear to have a nonperiodiccross-striated banding pattern (see Fig.51-3A). The length of the anchoringfibril is difficult to measure because ofits random orientation in relation to theplane of the section. In toad skin, thesestructures have lengths of approxi-mately 800 nm. The anchoring fibrils inhuman skin appear to be somewhatshorter. The ends of the anchoringfibrils appear less tightly packed, givinga somewhat frayed appearance. Theproximal end inserts into the basal lam-ina, and the distal end is integrated intothe fibrous network of the dermis.

55

Many of the anchoring fibrils originat-

ing at the lamina densa loop back intothe lamina densa in a horseshoe-likemanner; others insert their oppositeends into amorphous-appearing struc-tures, termed

anchoring plaques

.

55

Thesestructures are believed to be indepen-dent “islands” of electron-dense mate-rial, although some controversy exists inthe literature.

56

Anchoring fibrils are pri-marily aggregates of collagen VII.

57

Tubular fibrillin-containing micro-fibrils, 10 nm to 12 nm in diameter, arealso localized in the sublamina densa re-gion. These are elastic-related fibers, be-cause elastic components of the dermisare formed from microfibrillar andamorphous components.

58

The micro-fibrillar component in the presence ofabundant amorphous component isknown as the

elastic fiber

. In the papil-lary dermis, the microfibrils insert intothe basal lamina perpendicular to thebasement membrane and extend intothe dermis, where they gradually mergewith the elastic fibers to form a plexusparallel to the dermal-epidermal junc-tion. These two elastic components ap-pear to be continuous with the elastic fi-bers present deep within the reticulardermis.

58

The gradient of an increasingamorphous component from the basallamina into the reticular dermis mayrepresent a system of increasingly ma-ture elastic fibers.

In summary, the ultrastructure of thedermal-epidermal junction strongly sug-gests that the lamina densa functions asa structural scaffold for the attachmentof the epidermal cells at one surface, se-cured by anchoring filaments extendingfrom the lamina densa to the hemides-mosomes. The latter also serve as inser-tion points for intracellular keratin fila-ments that form scaffolding for thebasal cells. On the opposite surface, theextracellular matrix suprastructures ofthe dermis are firmly attached to thelamina densa. The interaction of differ-ent dermal fibers with the basal laminaappears to be mediated by the anchor-ing fibrils. The elastic system of the der-mis inserts directly into the basal laminavia the microfibrils. Thus, the dermal-epidermal junction provides a continu-ous series of attachments between thereticular dermis and the cytoskeleton ofthe basal cells. These observationssuggest four major functions for the epi-dermal basement membrane: (1) a struc-tural foundation for the secure attach-ment and polarity of the epidermalbasal cells; (2) a barrier function, sepa-rating the components of the epidermisand the dermis; (3) firm attachment of

the dermis to the epidermis through acontinuous system of structural ele-ments; and (4) modification of cellularfunctions, such as organization of thecytoskeleton, differentiation, or rescuefrom apoptotic signaling via outside-insignalling mechanisms.

BIOCHEMICAL CHARACTERIZATION OF THE BASEMENT MEMBRANE

Basement membranes contain collage-nous and non-collagenous glycoproteinsand proteoglycans. The unusual aminoacids hydroxyproline and hydroxylysineare found almost exclusively in col-lagens;

59

the hydroxyproline contentsuggests that collagens account for 40percent to 65 percent of the total base-ment membrane protein. All basementmembranes contain collagen IV as themajor collagen, laminin, nidogen, and theheparan sulfate proteoglycan perlecan, ir-respective of tissue source (Box 51-2). Inaddition, many other tissue specific gly-coproteins have been found in basementmembranes, including different laminins,fibulins, and fibronectin.

47–49

In contrast to the ubiquitous compo-nents within all basement membranes,some molecules demonstrate a restricteddistribution. Certain collagen IV iso-forms, collagens VII and XVII, or severallaminins, are not ubiquitous. For exam-ple, the

α

3 chain of collagen IV is local-ized in the basement membrane of thekidney and lung, but not in those of theskin and blood vessels. In contrast, col-lagens VII and XVII are associated withthe squamous epithelia of skin but arenot found in glomerular and alveolarbasement membranes. Differences inmacromolecular composition are re-sponsible for morphologic and func-tional variance of basement membranes.

Ubiquitous Components of Basement Membranes

COLLAGEN IV

Collagen IV is a heterotri-mer of three

α

chains.

48,59 Each of thesecontains three distinct domains (Fig.51-4A): the N-terminal cysteine-rich (7-S)

Box 51-2

Ubiquitous Components of Basement Membranes

■ Collagen IV■ Laminins■ Nidogens■ Perlecan

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452

domain, a central triple-helical domain,and a C-terminal globular domain (NC-l). The trimer composition is deter-mined by the NC-l domains, and the αchains are linked to each other by cova-lent interactions through these do-mains.60 The triple helix of collagen IVis long and contains several sites atwhich glycine is not present in everythird position.59 These discontinuitiesresult in increased flexibility in the col-lagen IV helix, but also render it suscep-tible to a variety of proteases.

The characteristic fibril structure of theinterstitial collagens results from aggre-gation of molecules in a staggered array,producing a cross-banding pattern inelectron microscope after appropriatestaining. In contrast, the ultrastructure ofthe lamina densa indicates no periodiccross-striations, suggesting that the ag-gregates of collagen IV are different. Thesuprastructure of collagen IV has beenpartially elucidated by rotary shadowingelectron microscopy that indicated thatthe major interactions among collagen IVmolecules occur at their amino- and car-boxyl-terminal domains, and by lateralassociation of their triple helices (see

Fig. 51-4A).47,48,59 Covalent interactionsamong 7-S regions of different moleculesare the basis for the specialized networkcharacteristic of basement membranes(see Fig. 51-4B). The individual 7-S re-gions overlap in both the parallel and an-tiparallel directions, producing a charac-teristic four-legged “spider” form. TheNC-l domains at the end of each leg ofthe spider interact with the NC-l domainof the adjacent aggregates (see Fig. 51-4B).Association is stabilized by covalentbonds. These end-to-end interactions re-sult in an extended two-dimensional net-work that is the basis of basement mem-brane organization.

The high flexibility of the basementmembrane network structure makes thepossibility of interactions with other col-lagens or noncollagenous molecules veryattractive. An open meshwork of col-lagen IV with, for example, laminins orperlecan, can be easily visualized (Fig.51-5).49 The implied porosity of thisstructure would then be limited by thesize of the pores in the collagen networkand by structural elements associatedwith it. This model of the basementmembrane structure allows considerable

mechanical stability while retainingphysiologic flexibility. These propertiesof strength and elasticity would be ex-pected for a dynamic surface, such asthat seen in the epidermal-dermal junc-tion and surrounding blood vessels.

Collagen IV molecules in differentbasement membranes contain geneti-cally distinct but structurally homologous

� FIGURE 51-5 A. Representation of the net-works formed by the ubiquitous components of thebasement membranes. Monomeric collagen IV(Col-IV) self-assembles into dimers and tetramersthat further aggregate into a complex lattice. Lami-nins self-polymerize into networks. Perlecan canoligomerize in vitro, and the glycosaminoglycanside chains interact with the Col-IV framework. Ni-dogen-entactin is thought to bind components ofall three networks and also fibulins. Nidogen there-fore plays a central role as a stabilizer of the lam-ina densa framework. Individual molecules are notdrawn to scale. (Drawing used with permissionfrom Peter Yurchenco, MD, Robert Wood JohnsonMedical School, Piscataway, N.J, U.S.A.) B. Rotaryshadowing image of a quick-freeze, deep-etchreplica of Col-IV polymers. The replica shows anextensive, branching and anastomosing networkwith occasional globular structures (arrowhead ),which can be visualized as a model for the struc-ture of the lamina densa. (Photo provided by Toshi-hiko Hayashi, PhD, University of Tokyo, Japan. Seealso Nakazato K et al: Gelation of lens capsule typeIV collagen solution at a neutral pH. J Biochem120:889, 1996.)

A

B

� FIGURE 51-4 Images of basement membrane molecules visualized by rotary shadowing. A. Col-lagen IV monomer and a dimer resulting from aggregation of C-terminal NC-1 domains. B. Collagen IVtetramer (“spider”) demonstrating the 7-S domain with the four protruding molecules and their large ter-minal NC-1 domains. C. Laminin 1 (laminin 111) molecules. D. Nidogen molecules. E. Procollagen VII.The NC-1 and NC-2 regions are indicated. F. Laminin 5 (laminin 332) molecules. (All micrographs wereprovided by Douglas R. Keene, MD, Shriners Hospital, Portland, Oregon.)

A

B

C D

E

F

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α chains. The α1 and α2 chains are ubiq-uitous, but the α3, α4, α5, and α6 chainsshow restricted distribution among tis-sues.61 The chain organization and dis-criminatory interactions between theNC-1 domains govern network assem-bly in the basement membranes.60 Twonetworks, namely α1-α2–containing andα3-α4-α5–containing networks, exist inthe glomerular basement membrane.Smooth muscle basement membraneshave an α1-α2-α5-α6–containing net-work in addition to the classic α1-α2network. In the skin, the α1-α2–contain-ing collagen IV network dominateswithin the dermal-epidermal junction,but α1-α2-α5-α6–containing network isalso likely to be present.62 Taken to-gether, the six chains of collagen IV aredistributed in three major networks, α1-α2, α3-α4-α5, and α1-α2-α5-α6, whosechain composition is determined by theNC-l domains.

The α3(IV) chain is the antigen recog-nized by the circulating autoantibodiesin the Goodpasture syndrome,61 and au-toantibodies directed against the NC-ldomain of the α5(IV) collagen chainwere reported in a case with sub-epider-mal bullous eruptions and renal insuffi-ciency.63 Structural aberrations in thegenes encoding the α3, α4, α5, and α6chains cause different forms of Alportsyndrome, a genetic disease character-ized by nephritis and deafness.61 Recentanalyses of mouse mutants revealed un-expected phenotypes of mutations inthe mouse Col4α1 and Col4α2 genes en-coding the α1 and α2 chains (i.e., abnor-malities of the kidney, the brain, andthe eye). Peripheral glomerulopathy inthe kidney, porencephaly, a rare neuro-logic disease characterized by degener-ative cavities in the brain, intracerebralhemorrhages, and a spectrum of eyeabnormalities affecting the iris, cornea,retinal arterioles, and optic nerve wereobserved.64,65 Subsequently, a Col4α1mutation was identified in a humanfamily with small-vessel disease, whichoften underlies ischemic strokes andintracerebral hemorrhages.66 Interest-ingly, despite the ubiquitous presenceof the α1 and α2 chains of collagen IVin basement membranes, aberrationsdid not occur in all basement mem-branes, suggesting varying roles forcollagen IV in different basementmembranes.

Laminins. Laminins are very large glyco-proteins (600 to 950 kd) within the lam-ina lucida/lamina densa of all basementmembranes.49 Three types of subunit

chains have been designated α (200 kdto 400 kd), β (220 kd), and γ (155 kd to200 kd) chains, and each laminin is a tri-meric aggregate of one α, β, and γ chain.The trimers have semirigid and ex-tended structures, which appear as anasymmetric cross in rotary shadowingelectron microscopy (see Fig. 51-4C).The long arm of the cross is approxi-mately 125 nm in length; the short armsare variable. Each laminin molecule isdivided into globular and rod-like do-mains that have been individually impli-cated in various functions, such as ag-gregation with itself and with othercomponents of the lamina densa (Fig.51-6), cell attachment and spreading,neurite outgrowth, or cellular differenti-ation.49,67 The C-terminal LG domain ofthe α chain, at the foot of the long armof the laminin cross, harbors the bindingsite for integrins.49,67

To date, 15 laminin isoforms have beenidentified. These represent different tri-meric combinations of five distinct αchains, three β chains and three γ chainsknown so far. Historically, laminins werenamed as laminin-1 to laminin-15, in theorder of their discovery, but this classifi-cation had grown quite impractical, withthe need to memorize the numbers. Anew, simplified nomenclature is based onthe chain composition and the number ofeach α, β, and γ chain (Table 51-2).68 Forexample, the classic “prototype” laminin-

1, with α chain composition α1β1γ1, isnow called laminin 111. The major lami-nin of the epidermal basement mem-brane, the previous laminin-5, with αchain composition α3β3γ2, is now calledlaminin 332.

The α2 chain containing laminins arepresent primarily within the basementmembranes of the muscle fibers, nerves,neuromuscular junction, and glomeru-lus. Mutations in the α2 chain causeprogressive muscular dystrophy and asignificant decrease in the amount ofbasement membrane accumulated sur-rounding muscle cells.67 The absence ofthe basement membrane leads to pro-gressive degeneration of the muscle dueto cell death. Therefore, the predictionis that laminins, and basement mem-branes in general, are required to pre-vent apoptosis by the cell types theysurround.67 The α3 chain is involvedin epithelial adhesion (see Epithelial-Specific Basement Membrane Compo-nents), and the α4 and α5 chains arefound in a variety of tissues includingendothelia, epithelia, neuromuscularjunction, and glomerulus.69 Laminin 511is present in the basement membrane ofthe epidermis and the hair follicles,where it is believed to be involved indevelopmental signaling.70 The distribu-tion of the β2 chain is largely restrictedto the neuromuscular junction, but it isalso found in nonmuscle tissues such as

� FIGURE 51-6 A schematic representation of laminin molecules. Each laminin is a heterotrimer ofan α, a β, and a γ chain. On the left, the classic prototype laminin 111 consisting of α1β1γ1 chainsis shown. The N-terminal short arm of each chain is free, the long C-termini fold to a coiled-coil andform the long arm. The distal C-terminus of the α chain contains five globular LG domains, whichharbor the integrin-binding site. Laminin 332, the previous laminin 5, exists in two forms, 3A32 and3B32. These represent splice variants of the α chain, the short variant is 3A and the “full length”chain 3B. The N-termini of the β3 and γ2 chains are proteolytically processed to yield mature lami-nin 332.

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the kidney glomerulus and the capillarybasal lamina.67 The β3 chain is involvedin epithelial adhesion (see Epithelial-Specific Basement Membrane Compo-nents). Three γ chain variants—γ1, γ2,and γ3—are known. The γ2 chain isfound only in laminin 332 in the skin(see Epithelial-Specific Basement Mem-brane Components). The γ3 chain, acomponent of laminins 423 and 523,binds nidogens 1 and 2 and is localizedin basement membrane zones of adultand embryonic brain, kidney, skin, mus-cle, and testis. It is present in muchlower concentrations than the γ1 chain,a fact that may indicate highly special-ized functions.71 The functions of alllaminins are not yet fully understood,but by interacting with integrins andother cell surface components lamininscontrol cellular activities such as adhe-sion, migration, proliferation, and polar-ity in a wide variety of organs.67,68

Nidogens. Two nidogens, nidogen 1 and2, also known as entactin 1 and 2, are dis-tinct gene products. Both are relativelysmall molecules (see Fig. 51-4D), whichbind laminins at a specific site within theγ1 and the γ3 chain,71 but also collagen IV,perlecan, and fibulins. It has been pro-posed that nidogens act as connecting el-ements between the collagen IV and

laminin networks and integrate otherbasement membrane components intothis specialized extracellular matrix.47,48

Targeted ablation of nidogens in miceshowed that the loss of either isoformhas no effect on basement membrane for-mation and organ development, but lackof both results in severe defects of thelung and the heart72 that are not compati-ble with life. Interestingly, despite theubiquitous presence of nidogens in base-ment membranes, aberrations did not oc-cur in all basement membranes, suggest-ing distinct roles for nidogens in differentbasement membranes.72

Heparan Sulfate Proteoglycans. Anotherclass of ubiquitous integral basementmembrane constituents are the pro-teoglycans.73 Three proteoglycans arecharacteristically present in vascular andepithelial basement membranes: perle-can, agrin, and bamacan. They consistof a core protein of various lengths, andthe first two carry primarily heparansulfate side chains, whereas the lattercarries primarily chondroitin sulfate.The name perlecan is derived from its ro-tary shadowing appearance reminiscentof a string of pearls. Perlecan representsa complex multidomain proteoglycanwith enormous dimensions and a num-ber of post-translational modifications.74

Knockout mice lacking perlecan exhib-ited abnormalities in many tissues, in-cluding basement membranes, andembryonic lethality. The basementmembranes deteriorated in regions un-der increased mechanical stress, such asmyocardium or skin, resulting in lethalcardiac abnormalities and skin blister-ing.73,74 Agrin is a major heparan sulfateproteoglycan of neuromuscular junc-tions and renal tubular basement mem-branes. Bamacan is likely to be thechondroitin sulphate proteoglycan pre-viously identified in many basementmembranes. The proteoglycans are ca-pable of interactions with several otherbasement membrane components andare believed to contribute to the overallarchitecture of the basement membraneas well as provide tissue-specific func-tions. The high sulfate content of theproteoglycans makes them highly nega-tively charged and hydrophilic, and thecharge density is responsible for provid-ing the selective permeability of the glo-merular basement membrane.

Syndecans are transmembrane hepa-ran sulfate proteoglycans present onmost cell types, including basal kerati-nocytes of the epidermis. The extracel-lular domains have affinity for lamininsand, presumably through these interac-tions, they regulate a variety of biologicprocesses, ranging from growth factorsignaling, cell adhesion, and cytoskeletalorganization, to infection of cells withmicroorganisms.75

Fibulins. Fibulins are a family of sixhighly conserved, calcium-binding extra-cellular matrix proteins. They are locatedin vessel walls, basement membranes,and microfibrillar structures and theyhave overlapping binding sites for a vari-ety of ligands, both basement membraneproteins and components of the intersti-tial connective tissues.76 Therefore, fibu-lins are believed to function as inter-molecular bridges that stabilize thesupramolecular organization of extracel-lular membrane structures, such as elasticfibers and basement membranes. Geneticdefects of the genes encoding fibulin 4and 5 cause different forms of cutis laxa.77

EPITHELIAL-SPECIFIC BASEMENT MEMBRANE COMPONENTS

The dermal-epidermal junction of skin isan excellent example of specific diver-gence in basement membrane structure.The structural components of hemides-mosomes, anchoring filaments, and an-choring fibrils in the basement mem-

TABLE 51-2New Laminin Nomenclaturea

OLD NAMECHAIN COMPOSITION NEW NAME TISSUE DISTRIBUTION

Laminin-1 α1β1γ1 Laminin 111 Developing epithelia

Laminin-2 α2β1γ1 Laminin 211 Muscle, nerves

Laminin-3 α1β2γ1 Laminin 121 Myotendinous junction

Laminin-4 α2β2γ1 Laminin 221 Neuromuscular junction, glomerulus

Laminin-5 α3β3γ2 Laminin 332A Stratified epithelia

Laminin-6 α3β1γ1 Laminin 311A Stratified epithelia

Laminin-7 α3β2γ1 Laminin 321A Amnion, maybe other stratified epithelia

Laminin-8 α4β1γ1 Laminin 411 Endothelia, nerves, smooth muscle, adipose tissue

Laminin-9 α4β2γ1 Laminin 421 Endothelia, neuromuscular junction, smooth muscle, glomerulus, adipose tissue

Laminin-10 α5β1γ1 Laminin 511 Mature epithelia and endothelia, smooth muscle

Laminin-11 α5β2γ1 Laminin 521 Mature epithelia and endothelia, smooth muscle, neuromuscular junction, glomerulus

Laminin-12 α2β1γ3 Laminin 213 Muscle

Laminin-14 α4β2γ3 Laminin 423 Retina, central nervous system, kidney, testis

Laminin-15 α5β2γ3 Laminin 523 Retina, central nervous system, muscle, kidney

aSee references 68 and 69.

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brane zone are quite well characterized(for review, see refs. 47–51, 78, 79). Acartoon depicting the relative locations ofthe proteins found at the dermal-epider-mal junction is shown in Figure 51-7.These proteins are listed in Table 51-3and discussed in the following sections.

Hemidesmosomes

Ultrastructurally, the hemidesmosomeclosely resembles one-half of the des-

mosome at cell-cell junctions in the epi-dermis. However, the components ofthese two structures are distinct. Char-acterization of the hemidesmosomalproteins was initially aided by the useof autoantibodies in serum of patientswith bullous pemphigoid (Fig. 51-8),which recognized antigens with a massfrom 120 kd to 230 kd.27,80,81 Cloning ofthe cDNAs for the molecules helpedidentify three distinct polypeptidechains: 230 kd, 180 kd, and 120 kd. The

230-kd protein, BPAG1 (bullous pem-phigoid antigen 1, or BP230), is a coiled-coil dimeric protein with homology toplakins, which bind intermediate fila-ments.80,82 BPAG1 is the major compo-nent of the hemidesmosomal innerdense plaque. In a transgenic mousemodel, deletions in BPAG1 caused epi-dermolysis bullosa simplex78 but no hu-man mutations have been found sofar.83 The 180-kd protein, previouslycalled BPAG2 or BP180, and the major

� FIGURE 51-7 Model of the hypothetical relationships of molecules within the dermal-epidermal junction basement membrane. The illustration depictslaminin 332 as the bridge between the transmembrane hemidesmosomal integrin α6β4 and the collagen VII NC-1 domain. The tight binding of laminin 332 toα6β4 and to collagen VII provides the primary resistance to frictional forces. The transmembrane collagen XVII also participates in this stabilization, becauseits extracellular domain also binds laminin 332. Within the epithelial cell, the transmembrane elements bind the proteins of the hemidesmosomal denseplaque, bullous pemphigoid antigen (BPAG)1 and plectin, which then associate with the keratins. Collagen XVII binds BPAG1, integrin α6β4, and plectin, andintegrin α6β4 binds plectin. The laminin 5-6 complex is shown within the basement membrane between hemidesmosomes, bound by integrin α3β1, and as-sociated with the intracellular proteins kindlin, talin, and vinculin. This complex presumably maintains basement membrane stability. In vitro, integrin α3β1,kindlin, talin, and vinculin, and another transmembrane collagen, type XIII, are localized to the focal contacts, which may function as the link between thebasement membrane and the epithelial cortical actin network. In the lamina densa, collagen IV and perlecan networks are stabilized by nidogen. Anchoringfibrils are secured to the lamina densa by the NC-1 domain of collagen VII. The fibrils project into the dermis and either terminate in anchoring plaques or loopback to the lamina densa. The anchoring fibril network entraps dermal fibrils, thus securing the adhesion of the lamina densa to the papillary dermis. None ofthe molecules is drawn to scale.

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antigen in bullous pemphigoid, is atransmembrane collagen now known ascollagen XVII, where the collagenous do-main is extracellular.84–87 In fact, col-lagen XVII is a prototype of the novelprotein family of collagenous trans-membrane proteins, type II transmem-brane molecules with one or more col-lagenous stretches in the extracellulardomain.88 Mutations in collagen XVIIcause junctional epidermolysis bullosa(see Chap. 60), indicating that it stabi-lizes interactions of basal keratinocyteswith the basement membrane. Its intra-cellular ligands are plectin, BPAg-1 andβ4 integrin, and the extracellular ligandsα6 integrin and laminin 332.78,79 The120-kd protein is the ectodomain of col-lagen XVII,85,87,88 which is shed from

the cell surface by proteinases of theADAM (A disintegrin-like and metallo-proteinase-containing) family89 throughcleavage within the juxtamembranousNC-16 domain (Fig. 51-9). Plectin, an-other dimeric plakin homologue, is also acomponent of the hemidesmosome.78

However, its tissue distribution is notlimited to hemidesmosome-containingbasement membranes. Mutations ofplectin result in epidermolysis bullosasimplex and progressive muscular dys-trophy and, in some cases, junctional epi-dermolysis bullosa with pyloric atresia(see Chap. 60), indicating a role for plec-tin in the stability of cell-basement mem-brane adhesion in a variety of tissues.

One key component of the hemides-mosome is the integrin α6β4.

78 It has a

high affinity for laminin 332, and there-fore is essential to integration of thehemidesmosome with the underlyingbasement membrane and stroma.78,79

Mutations in either the α6 or β4 chainsresult in dermal-epidermal instability. Inmice, null mutations cause severe junc-tional epidermolysis bullosa.78 In hu-mans, different mutations in the α6 or β4chains are associated with variably se-vere skin blistering associated with py-loric atresia.79,83

A member of the widely expressed cellsurface transmembrane proteins of thetetraspanin family, CD151, is also a com-ponent of the hemidesmosome. It formscomplexes with α3β1 and α6β4 integrins atthe basolateral surface of basal keratino-cytes and stabilizes their functions by pro-

TABLE 51-3Hemidesmosomal and Basement Membrane Zone (BMZ) Targets in Skin Disease

HEMIDESMOSOME/BMZ COMPONENT AUTOIMMUNE TARGET GENETIC TARGET

Cytoskeletal proteins Keratin 5 and 14 EBS

Hemidesmosomal plaque proteins Bullous pemphigoid antigen 1/BP230 BPPlectin BP, CP EBS-MD

Other intracellular adhesion complex proteins Kindlin-1 KS

Hemidesmosomal transmembrane components Collagen XVII/BP180 BP, CP, LAD, PG JEB-non-Herlitzα6β4 integrin BP, CP JEB-PACD151 Pretibial EB, nephritis, deaf-

ness, β-thalassemia minor

Anchoring filament proteins Laminin 332/laminin-5 CP JEB-HerlitzJEB–non-Herlitz

Ectodomain of collagen XVII LAD, BP JEB-non-Herlitz

Anchoring fibril proteins Collagen VII EBA DEB

BP = bullous pemphigoid (see Chap. 54); CP = cicatricial pemphigoid (see Chap. 55); DEB = dystrophic EB (see Chap. 60 for all); EB = epidermolysis bullosa; EBA = EB acquisita (see Chap. 58); EBS = EB simplex; EBS-MD = EBS with muscular dystrophy; JEB= junctional EB; JEB-PA = JEB with pyloric atresia; KS = Kindler syn-drome; LAD = linear immunoglobulin A dermatosis (see Chap. 56); PG = pemphigoid gestationis (see Chap. 57).

� FIGURE 51-8 A. Skin blisters in bullous pemphigoid result from antibody-mediated loss of function of the adhesion molecule collagen XVII. B. Indirect im-munofluorescence staining of human skin with a pemphigoid serum that contains autoantibodies to collagen XVII.

A B

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viding a framework for the spatial organi-zation of the different components.90

Dissociation of the CD151-integrin com-plexes permits remodeling of epithelialcell interactions with the basement mem-brane and cell migration.91,92 CD151-nullmice have apparently normal tissue mor-phology, including hemidesmosomes, butexhibit some functional aberrations, e.g.,poor keratinocyte migration in explantcultures.93 A rare human genetic condi-tion delivered indirect information on thefunctions of CD151. In addition to the ex-pression of CD151 in several tissues likethe kidney and the skin, its gene also en-codes the MER2 blood group antigen onerythrocytes. Homozygous CD151-nullmutations were identified in three MER2-negative patients, who also presentedwith hereditary nephritis, sensorineuraldeafness, pretibial epidermolysis bullosa,and β-thalassemia minor.94 These symp-toms suggest that CD151 is important forthe assembly of the basement membranein the kidney, skin, and inner ear andplays a role in erythropoiesis.

Other Epidermal Adhesion Complexes

In addition to the hemidesmosomalcomponents, other adhesion moleculesare known to be present at the basolat-eral aspect of basal keratinocytes, for ex-ample integrin α3β1, the receptor for the

laminin 332–311 complex in the base-ment membrane between the hemides-mosomes,95 and another transmembranecollagen, type XIII.96 Since these proteinsare localized to focal contacts in vitro, to-gether with vinculin and talin, they arepredicted to function as the link betweenthe basement membrane and the epithe-lial cortical actin network. Recently, anovel intracellular component of thiscomplex, kindlin-1, was identified by thegenetic studies. The kindlin-1 gene,KIND1, is mutated in the Kindler syn-drome,97–99 a disorder with skin blister-ing in infancy, progressive poikiloderma,skin atrophy, pigment anomalies, and,occasionally, skin cancer. In the epider-mis, kindlin-1 is expressed at the basolat-eral surface of basal keratinocytes, and incultivated epithelial cells it co-localizeswith vinculin, a marker of integrin-medi-ated cell-matrix adhesions, and actin mi-crofilaments.97,100 These observationssuggest that kindlin-1 is necessary for thestability of the dermal-epidermal junc-tion and that, in addition to hemidesmo-somes, tethering the actin cytoskeletonto cell-matrix adhesions offers alternativemeans to anchor basal epithelial cells tothe basement membrane.79

Anchoring Filaments

The anchoring filaments contain laminin332 and the ectodomain of collagen

XVII,67,68,79 two ligands that interactwith each other through noncovalentbonds.101 The ectodomain of collagenXVII, which protrudes from the plasmamembrane into the lamina lucida, has aloop structure consistent with its role asan anchoring filament protein.102 Themolecular characteristics of this collagenare delineated in Hemidesmosomes.84–88

Laminin 332 is a disulfide-bonded com-plex of α3, β3, and γ2 chains. The twosplice variants of the α3 chain, α3A andα3B, associate with the α3 and γ2chains to form laminin 3A32 and 3B32(see Fig. 51-6), previously known aslaminin-5.68 Rotary shadowing imagingindicates that laminin 332 has a rod-likestructure terminating in the globular re-gions (see Fig. 51-4F), a shape consistentwith its role as an anchoring filamentprotein. The individual chains are con-siderably truncated relative to otherlaminin chains, and this truncation is re-flected in the loss of the structuresequivalent to the short arms of otherlaminins. Additionally, the α3 and γ2chains are proteolytically processed af-ter secretion from the keratinocyte, fur-ther trimming the short arms.103,104 TheC-terminus of the α chain, longer thanthat of the β and γ chains, comprises fiveglobular LG modules, LG1 through LG5,which interact with cell surface recep-tors. The α3β1 and α6β4 integrins haveaffinity for the LG1-3 domains,105

whereas the LG4-5 tandem has affinityfor syndecans and β-dystroglycan onthe keratinocyte surface.106 The LG4-5modules are proteolytically cleaved inmost laminins, a process which maymodulate interactions with cell surfacereceptors.67,68 Laminin 332 forms co-valent complexes with laminin 311(α3β1γ1), binds to the NC-1 domain ofcollagen VII, the anchoring fibrilprotein79 and to the distal ectodomainof collagen XVII.101 Genetic evidencedemonstrates that laminin 332 is essen-tial in keratinocyte adhesion, as nullmutations in any of its componentα3, β3, or γ2 chains result in severe Her-litz junctional epidermolysis bullosa(see Chap. 60).83 Targeted disruption ofthe mouse lama3 gene prevented thesynthesis of both laminin 332 and 311molecules and resulted in abnormalhemidesmosomes, lack of survival ofmutant epithelial cells, severe junctionalblistering, abnormalities of ameloblastdifferentiation in developing teeth, andperinatal lethality.107 Therefore, the se-verity of the laminin 332 null phenotypereflects the loss of its ability to bridgethe hemidesmosomes and the anchor-

� FIGURE 51-9 Schematic representation of collagen XVII and its ectodomain shedding. Collagen XVII isa hemidesmosomal transmembrane protein with an intracellular N-terminus. The extracellular C-terminus(ectodomain) contains several collagenous subdomains (brown) and intervening non-collagenous se-quences (beige). The ectodomain can be shed from the cell surface by proteinases of the ADAMs (A disin-tegrin-like and metalloproteinase-containing) family, themselves transmembrane proteins. Thus, the 180-kd full-length molecule yields a shorter soluble ectodomain of 120 kd. The 180-kd full-length molecule isthe classic bullous pemphigoid antigen-2, and the ectodomain is the 120-kd linear immunoglobulin A (IgA)dermatosis antigen. Further degradation of the C-terminus of the ectodomain results in the 97-kd linear IgAdermatosis antigen.

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ing fibrils, resulting in a separationwithin the lamina lucida.

Epithelial Lamina Densa

The basement membrane beneath andbetween the hemidesmosomes containsthe α1-α2–containing collagen IV net-work, probably some α1-α2-α5-α6–containing collagen IV network, as wellas nidogen, perlecan, and laminin α3and α5-containing molecules.47–49,61

The N-terminus of the α3 chain lacksthe so-called LN modules, which allowself-polymerization of other lamininisoforms. However, the α3 chain canassociate with the β1 and γ1 chains oflaminin 311, previously laminin-6,which has the unique property of form-ing disulfide-bonded dimers with lami-nin 332.67,68 It is probable that the majorα3-containing laminin in the laminadensa between hemidesmosomes is thelaminin 332-311 complex.67,68,79 As thelaminin α3 chain is a ligand for integrinα3β1 present between hemidesmo-somes, binding of laminin 332-311 com-plex to the intracellular actin cytoskele-ton is likely to be mediated by integrinα3β1. This is consistent with studies inmice in which targeted ablation of theintegrin α3 chain causes loss of the base-ment membrane between hemidesmo-somes but not beneath them.108 The N-termini of laminin 332 bind to collagenVII, a component of the anchoringfibrils located in the sub-lamina densa,so that anchoring filaments and fibrilsare directly connected.109 The laminin332-311 complex and laminin 511(α5β1γ1), previously laminin-10, containa γ1 chain and can therefore bind ni-dogen 1 and 2 and the collagen IV net-work.70,71 Further, nidogen 1 and fibulin1 and 2 were shown to be ligands for re-combinant N-terminal γ2 chain of lami-nin 332.110 These interactions are im-portant for the integration of laminin332 into the extracellular matrix beforethe maturation of the γ2 chain,103 as asubstantial portion of N-terminus of theγ2 chain is cleaved in human adult skin.79

Yet another link, which strengthens der-mal-epidermal cohesion, is provided bymolecular interactions between perlecanwithin the lamina densa and fibrillin 1 inthe microfibrils.111

Anchoring Fibrils

Collagen VII is the major component ofthe anchoring fibrils.55,57,112 The col-lagen VII molecule is distinguished fromother collagens in that it has a very long

triple-helical domain, 450 nm in length.Globular domains exist at both ends ofthe triple helix, and the N-terminal do-main NC-1 is very large and trident-like(see Fig. 51-4E). The smaller C-propep-tide, NC-2, is believed to facilitate theformation of the antiparallel, cen-trosymmetric dimers,113 before it is re-moved by the metalloproteinase bonemorphogenetic protein 1114 to yield amature collagen VII. The dimers are co-valently cross-linked through disulfidebonds at the carboxy terminus, and theyaggregate laterally to form the anchor-ing fibrils.112 The fibrils are further stabi-lized by tissue transglutaminase, whichcatalyzes the formation of covalent γ-glutamyl-ε-lysine cross-links.115

The NC-1 domain of collagen VIIbinds to laminin-5 and collagen IV withinthe lamina densa (see Fig. 51-3A).116

The triple helical domains of an antipar-allel collagen VII dimer make the lengthof the anchoring fibril. It extends per-pendicularly from the lamina densa andeither loops back into the lamina densaor inserts into the anchoring plaques.55–57

The anchoring plaques are electron-dense structures that contain collagen IVand laminin 332, and perhaps otherbasement membrane components, butwhich are believed to be independent ofthe lamina densa itself.55 They are dis-tributed randomly in the papillary der-mis below the lamina densa and areinterrelated by additional anchoringfibrils. The anchoring fibril network

forms a scaffold that entraps largenumbers of dermal fibrils, securing thelamina densa to the subjacent der-mis.55–57,112 In the acquired form ofepidermolysis bullosa, epidermolysisbullosa acquisita (see Chap. 58), and inbullous systemic lupus erythematosus(see Chap. 156), autoantibodies targetmainly the NC-1 domain of collagenVII.117,118

Mutations in COL7A1, the gene en-coding collagen VII, result in dystrophicepidermolysis bullosa (see Chap. 60).As of 2006, more than 300 COL7A1mutations have been found in both re-cessive and dominant forms of dystro-phic epidermolysis bullosa (Fig. 51-10),and the spectrum of biologic and clini-cal phenotypes is much broader thananticipated.79,83,119 In two differentmouse models, complete or partial defi-ciency of collagen VII recapitulated theclinical and morphologic characteristicsof recessive dystrophic epidermolysisbullosa in humans.120,121 These micewill be useful for testing of moleculartherapy strategies for dystrophic epider-molysis bullosa. A number of such ther-apies, ranging from protein122 therapyto transposon-mediated gene trans-fer,123 have been considered. Gene aug-mentation in an ex vivo approach cur-rently seems to be most promising as afirst step, in particular for recessive dys-trophic epidermolysis bullosa.124–127

This strategy comprises genetic correc-tion of patient keratinocytes in vitro,

� FIGURE 51-10 Skin fragility and blistering in dystrophic epidermolysis bullosa. Functional deficiencyof collagen VII as a result of mutations in the COL7A1 gene leads to trauma-induced separation of theepidermis and the dermis. The blister roof is below the lamina densa, leading to scar formation on heal-ing of the blisters.

CHAPTER 52■

PEMPHIGUS

459

and their cultivation into epithelialsheets for grafting onto the patient’sskin. Results with a canine dystrophicepidermolysis bullosa model seempromising: transduction of epidermalcells with a retroviral vector containingthe entire collagen VII cDNA enhancedexpression and deposition of collagenVII at the dermal-epidermal junction.126

In another approach, nonviral genetransfer using cotransfection of C31integrase and human collagen VII re-sulted in the expression of the proteinin collagen VII–deficient DEB keratino-cytes.124 As fibroblasts also contributeto the synthesis of collagen VII, theirsuitability for gene therapy approachesis under investigation.128,129

Cellular Origin of the Dermal-Epidermal Basement Membrane

The basement membrane constituentsare products of both epithelial and mes-enchymal cells. In vitro modeling ofbasement membrane formation clearlyshows that, under at least some condi-tions, the dermal-epidermal junctionbasement membrane is contributed toby both tissue compartments, and ithas been proposed that differentiatedfibroblasts exist adjacent to epithelialtissues in vivo, which produce base-ment membrane components and assist

in basement membrane assembly.130,131

Of the known basement membranecomponents, only laminins 332 (lami-nin-5) and 311A (laminin-6) are exclu-sively produced by the epidermis. Epi-thelial cells also manufacture most ofcollagen VII, whereas mainly mesen-chymal cells synthesize collagen IV, ni-dogen, perlecan, and the laminin α2chain.131 Because the mesenchymalproducts are translocated to the baso-lateral epithelial surface where theycondense, that surface must provide thelocalization cues. Integrins α6β4 andα3β1 and collagen XVII have been im-plicated in this process, suggesting thatthe laminins coordinate basementmembrane polymerization.67,68,101 Asnidogen is required for stabilization ofthe basement membrane, and becauseit is a mesenchymal product, it is likelythat the dermis is essential to develop-ment of basement membranes.133 Stud-ies using different skin equivalent cul-ture models have demonstrated that atight interplay between fibroblasts andkeratinocytes, in terms of both theirmatrix production and secretion of sol-uble signals, regulates the formation ofthe dermal-epidermal basement mem-brane.132,134 An interesting regulatorystep may be added by dermal enzymes(e.g., bone morphologic protein 1),which process epithelial cell products,such as laminin 332 and procollagen

VII, to mature basement membranemolecules.103,104,114

KEY REFERENCES

The full reference list for all chapters is available at www.digm7.com.

4. Yin T, Green KJ: Regulation of desmo-some assembly and adhesion. SeminCell Develop Biol 15:665, 2004

27. Payne AS et al: Desmosomes and dis-ease: pemphigus and bullous impetigo.Curr Opin Cell Biol 16:536, 2004

39. McGrath JA: Keratinocyte adhesion andthe missing link: from Dowling-Mearato Hay-Wells. Clin Exp Dermatol 26:296,2001

48. Timpl R: Macromolecular organizationof basement membranes. Curr Opin CellBiol 8:618, 1996

61. Hudson BG et al: Alport’s syndrome,Goodpasture’s syndrome, and type IVcollagen. N Engl J Med 348:2543, 2003

68. Aumailley M, et al: A simplified lamininnomenclature. Matrix Biol 24:326, 2005

73. Iozzo RV: Basement membrane pro-teoglycans: from cellar to ceiling. NatRev Mol Cell Biol 6:646, 2005

78. Koster J, Borradori L, Sonnenberg A:Biology of the hemidesmosomes, inHandbook of Experimental Pharmacology,vol 165, edited by J Behrens, WJ Nelson.Berlin, Springer, 2004, p 243

80. Yancey KB: The pathophysiology ofautoimmune blistering diseases. J ClinInvest 115:825, 2005

83. Has C, Bruckner-Tuderman L: Molecu-lar and diagnostic aspects of geneticskin fragility. J Dermatol Science 44:129,2006

C H A P T E R 5 2

PemphigusJohn R. Stanley

The term pemphigus refers to a group ofautoimmune blistering diseases of skinand mucous membranes that are char-acterized histologically by intraepider-mal blisters due to acantholysis (i.e.,separation of epidermal cells from eachother) and immunopathologically by invivo bound and circulating immuno-globulin G (IgG) directed against the cellsurface of keratinocytes.1 The nosologyof this group of diseases is outlined inTable 52-1. Essentially, pemphigus canbe divided into four major types: vul-garis, foliaceus, paraneoplastic (seeChap. 53), and IgA pemphigus (see

Chap. 34). In pemphigus vulgaris (PV),the blister occurs in the deeper part ofthe epidermis, just above the basallayer, and in pemphigus foliaceus (PF),also called superficial pemphigus, the blis-ter is in the granular layer.2

The history of the discovery of pem-phigus, and its various forms, is coveredin Lever’s classic monograph Pemphigusand Pemphigoid.2 Both PV and PF displaya spectrum of disease. Various pointsalong these spectra have been givenunique names, but because the presen-tation of these diseases is fluid, patients’disease usually crosses these artificialdesignations over time. Thus, patientswith PV may present with more local-ized disease, one form of which is calledpemphigus vegetans of Hallopeau. Thismay become slightly more extensiveand may merge into pemphigus vege-tans of Neumann. Finally, with more se-vere disease, full-blown PV may appear.Similarly, patients with PF may present

with more localized disease, repre-sented by pemphigus erythematosus.However, these patients often go on tomore widespread PF.

TABLE 52-1Classification of Pemphigus

TYPE FORM

Pemphigus vulgaris

Pemphigus vegetans: LocalizedDrug-induced

Pemphigus foliaceus

Pemphigus erythemato-sus: LocalizedFogo selvagem: EndemicDrug-induced

Paraneoplastic pemphigus IgA pemphigus Sub-corneal pustular

dermatosisIntraepidermal neutro-philic IgA dermatosis

IgA = immunoglobulin A.