Significance and Pathogenesis of Basal Keratinocyte Herniations in Psoriasis

Madalene C. Y. Heng, M.B ., F.R.A.C.P., Suni G. Kloss , B.A. , Craig S. Kuehn, B.S. , and David G. Chase, Ph.D. Department of Medicine, Division of Dermatology, U CLA San Fernando Valley Internal Medicine Program, Veterans Administration Medica l Center, Sepul veda, California, U .S.A. .

U sing transmISSIOn electron microscopy, we studied, quantitatively, b asal keratinocy te h erniations (BKH) in re­lation to the other basem ent membrane zone changes in psoriatic lesions of varying clinical activity, and in p so­riasiform skin diseases. BKH appears to correlate with dis­ease activity. They d o not occur p assively as a result of the formation of ga ps in the basal lamina . BKH in active p so-

Cytoplasmic processes fro m basal keratinocytes pro­truding into the dermis through gaps in the basal lamina, have been 'observed by several investigators [1 ,2] in involved psoriatic skin. We have subse­

. quently referred to these as basa l keratinocyte her­niations or BKH [3,4]. However, their role in psoriasis remains unclear. Loss of integrity of the kerat inocyte basement membrane has been described in psoriasis [1 ,5-7]. and BKH have been thought to occur passively as a consequence of the formation of ga ps in the basal lamina [2]. In this study, we have attempted to show, by a series of studies, that the frequency of BKH co rrelated with clinical activity; that the presence ofBKH could serve as a marker for active psorias is; that BKH in active disease were associated with electron-lucent areas , suggestive of proteolytic autodiges­tion; and that their apparent association with Langerhans cells, neutrophils, dermal macrophages, and endothelial cells suggests that these cells, w hich are know n producers of proteolytic en­zymes , may playa role in BKH formation.

PATIENTS AND METHODS

Patient Population All the patients and controls attended the Outpatient Dermatology Clinics at the Veterans Administration Medical Center, at Sepulveda, California. They were divided into the following groups. .

Patiel'lls: Biopsies were obtained fro m 36 psoriatic patients. C lin­ical detai ls have been summarized in Table I. Biopsies were taken from acute, untreated psoriatic plaques (4 patients), pustular le­sions (3 patients), eruptive and/or generalized psoriasis (3 pa­tien ts), treated patients with plaques without areas of resolution (5 patients), treated patien ts with plaques containing areas of res­olution (3 patients), resolving generalized psoriasis (2 patients),

Manuscript received July 31, 1985 ; accepted for publication February 18, 1986.

Supported by a Veterans Adminis tration Merit Proposal Grant. Reprint requests to: Madalene C. Y. Heng, M .B., Division of Der­

matology (1 11 F) , Veterans Administration M edical Center, 16111 Plum­mer Street, Sepulveda, Califo rnia 91343.

Abbreviations: BKH: basal keratinocyte hern i a~ion(s) BMZ: basement membrane zone

ria sis are associated with electron-lucent areas suggestive of proteolytic enzyme release. Their apparent association with Langerhans cells, neutrophils , macrophages, and endothelial cells may point to these cells as the source of proteolytic enzymes in psoriasis. BKH may prove to be a useful marker for clinical psoriasis. J Invest D ermatol 87:362-366, 1986

completely resolved lesions, marked only by hyperpigm entation (6 biopsies), and from uninvolved skin (10 biopsies) at least 10 cm from the nea rest psoriatic lesion .

Controls: The control population provided biopsies of normal skin from 4 nonpsoriati c patients (age- and sex-matched) , and from the involved skin of 10 patients with the following pso­riasiform skin diseases: seborrheic dermatitis (2 patients), pityri­asis rubra pilaris (1 patient) , and 1 patient each with the following: contact eczema (rubber chemicals), generalized eczema from the­ophylline, pustular eczema palms and soles, primary irritant der­matitis from Campho-phenique, contact dermatitis (nickel). li­chen simplex chronicus , and li chen planus.

Light and Electron Microscopy Four-millimeter punch biop­sies were taken and divided 10ngitudinal1y with a sharp scalpel. One hiM of each biopsy was fixed in 10% neutral buffered for­malin and processed for light microscopy. The paraffin sections were stained with hematoxylin and eosin. The other half w as fixed in 2.5% glutaraldehyde, buffered to pH 7.3 with 0. 1 M

sodium phosphate, postfixed in osmium tetroxide, treated en bloc with tanmc acid, dehydrated in alcohol and propylene oxide, and embedded in a mixture of Epon 812 and Araldite 502. Silver sections were cut on a Sorval MT 2B ultramicrotome with a diamond knife (Dupont) and examined with a Philips EM 201 transmission electron microscope .

Analysis of Data The frequency of BKH was determined as follows. A series of slightly overlapping electron micrographs were taken of a 150-250 ,uin length of basement membrane for each patient and control, and printed at a constant magnification of 17,000 x. The basement membrane lengths were measured on a graphics tablet connected to an Apple II plus microcomputer. The number of herniations and other basement membrane zone (BMZ) changes were counted as the length of basal lamina was measured with the graphics tablet.

RESULTS

Analysis of the data obtained from psoriatic lesions of varying clinical activity shows that BKH (Fig la) were most numerous in untreated eruptive, p!lstuJar, generalized psoriasis (9.1 ± 1.4 BKH per 200,um basal lamina length) , less numerous (3.8 ± 0.7 BKH per 200 ,um basal lamina length) in treated psoriatic lesions

0022-202X/86/S03.S0 Copyright © 1986 by The Society for Investigative Dermatology, Inc.

362

VOL. 87, NO.3 SEPTEMBER 1986 BASAL KERATINOCYTE HERNIATIONS IN PSORIASIS 363

Table I. Frequency of Basal Keratinocyte H erniation (BKH) and Basement Membrane Zone Abnormalities in Psoriasis and Controls

Lamina Lucida Basal Lamina C hanges Dilatations BKH

N o. of Patients (no .l200 f..Lm basal lamina length)

(no .l200 f..Lm basal (no.l200 f..Lm basal C linical Status (biopsies) Gaps Thinning Reduplication lamina length) lamina length)

Psoriasis: Treatedlresolving 10 13.5 :±: 1. 6 34.9 :±: 4.9 30.0 :±: 2.6 21.4 :±: 2.1 3.8 :±: 0.7 Untreated 10 40.6 :±: 1.9 76.5 ± 5.3 11.1 :±: 1.3 41.0 :±: 1.7 9. 1 :±: 1.4 Completely resolved 6 5.4 ± 0.6 19.8 ± 0.9 59.3 :±: 0.9 5.4 :±: 0.8 0 Uninvolved 10 2.2 :±: 0.2 9.7 ± 0.9 10.8 :±: 0.9 4.9 :±: 0.4 0

Controls (nonpsoriatics): Pityriasis rubra pilaris 1 52.2 102. 6 Eczema (all types) 6 13.6 26.2 Lichen planus 1 12.0 29.0 Normal skin 4 0.2

without areas of resolution, very infrequent in treated psoriatic plaques with areas of resolution (1.2 ± 0.4 BKH per 200 ,um epidermal length), and absent in areas that showed complete clin­ical resolution . They were absent from uninvolved psoriatic skin, from normal non psoriatic skin, and from a number of common dermatoses. These resuits have been summarized in Table I. For

Figure 1. a, Electron micrograph showing a basal keratinocyte herniation (BKH) con­sisting of a cytoplasmic process from the basal keratinocyte (BK), protruding through a gap (between arrowheads) in the basal lam­ina, into the dermis (D) . Note the e1ectron­lucent area (A) adjacent to the BKH, sugges­tive of proteolytic autodigestion. LL = di l­atation of the lamina lucida; LD = lamina densa. Bar = 1 f..L. b, Electron micrograph of a healing psoriatic plaque, showing re­duplication of the lamina densa (LD) . Note also a gap (between arrowheads) in the basal lamina, and multiple dilatations of the lam­ina lucida (LL). E = epidermis, D = der­m is. Bar = 1 f..L. c, Electron micrograph of a treated psoriatic plaque (steroids), show­ing a BKH of the central variety, protruding into the dermis (D) through a gap (between arrowheads) in the lamina densa (LD). Note localized dilatations (LL) of the lamina lu­cida. BK = basal keratinocyte. Bar = 1 f..L.

1.0

3.6 51.0 1 33.8 16.0 0 39.6 18.7 0

1.1 0.8 0

the purpose of statistical analysis, the psoriatic group was divided into 4 subgroups: involved untreated, treatedlresolving, com­pletely resolved, and uninvolved. An analysis of variance shows that the treated group is significantly different (p < 0.01) from that of the treated, resolved, and uninvolved groups with regard to BKH frequency .

364 HENG ET AL

An examination of the distribution of BKH in relation to the other BMZ changes, such as gaps (Fig 1b) , thinned areas (Fig 1a) in the lamina densa, and dilatations of the lam ina lucida (Fig 1b) revealed that altho ugh these BMZ abnormalities were, in general, most numerous in the involved psoriatic lesions , there was no correlation between the number of gaps in the lamina densa and the frequency ofBKH found (Table I). These BMZ changes were also found in all the control psoriasiform lesions studied . In pit­yriasis rosea, BKH were noted , but in a fre quency far lower than that found in psoriasis. Many ga ps were noted, particularly in seborrheic derm atitis, w ithout the presence of a single BKH. In psoriasis and many of the control diseases, Langerhans cells, lym­phocytes, and neutrophils were seen in the process of crossing the basement membrane, although such observations were more frequently found in psoriasi s. Multil ayered basal lamina (redu­plication, Fig 1 b) were most prominent in the resolving and com­pletely resolved psoriatic lesions (Table I) , and were frequentl y seen in the contro l biopsies (Table I) .

The BKH observed in biopsies from the patients were further examined and divided in to 2 subgroups according to whether or not they were associated with electron-lucent areas in their im­mediate dermal aspect (Figs 2,3). T he data have been summarized in Table II. It was found that virtuall y all the BKH fro m act ive, untreated psoriasis were associated w ith electron-lucent areas in their dermal aspect. Surprisingly, even BKH from healing pso­riatic lesions were associated with similar electron-lucent areas in all but 3 structures (Ta ble II ). The 3 BKH with minimal or no surrounding electron lucency (Fig 1e) were found in 1 patient who had received prolonged topical steroid therapy.

It was also noted that BKH were associated w ith the follo wing cell types: (1) Langerhans cells in the process of cross ing the basal membrane; (2) lymphocytes in the process of crossing the base­ment membrane; (3) derm al macro phages; (4) neutrophils (Fig 2); and (5) endothelial cells (Fig 3). M any of these cells had cy­topl asmic extens ions toward the BKH. Occasionally, BKH were noted to be in contact with localized areas of electron lucency, which were lined by cytoplasm from the neutrophil (Fig 2) or endothelial cell (Fig 3).

DISCUSSION

Our data in Table I show that BKH freq uency correlates with clinical activity in psoriatic lesions. Although there was some correlation of the o ther basement mem brane changes with clinical activity, the gaps in the basal lamina and lamina lucida dilatations

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were also found in uninvolved and healed psoriatic skin as well as in a number of control diseases, and are, therefore, not as useful as BKH as markers for the presence of the clinical psoriatic lesion. The presence of BKH also in pityriasis rubra pilaris, a disease characterized by hyperproli fera tion as is psoriasis, and the pres­ence of BKH-like structures in a num ber of tumors [8-11) would suggest that BKH may playa role in hyperproliferation . By pro­viding a means whereby epidermal and stro mal elements can react more intimately , over a larger surface area, it is possible that BKH m ay result in enhancem ent of the epidermal-stromal in­teraction that results in chronicity of the clinical lesion. The BKH may provide perpetuation of epidermal-stromal interaction that results in chronicity of the clinical lesion.

The widespread occurrence of gaps in the lamina densa and other BMZ abnormalities in psoriasis and in the psoriasiform diseases studied suggests that these changes may be the nonspecific consequence of the inflammatory process. Proteolytic enzym es released from macrophages, neutrophils, an d endothelial cells [14-1 6) are capable of causing basement membrane dissolution and may be responsible for ga ps in the basal lamina. In addition , chymotrypsin-like enzymes released from the epidermis and dam­aged mast cells in the dermis may result in dilatory changes ob­served in the lamina lucida [17). The predominance of BKH in psoriatic lesions sugges ts the value of these structures as a marker for active psoriasis, and investigation into their formation m ay, therefore, lead to a deeper understanding of psoriasis .

The work of Sugrue and Hay [1 8) suggests that proteolytic enzymes may have been responsible for the formation of BKH­like structures. U sing corneal epithelia in tissue culture, these investigators found that when a trypsin-collagenase mixture was added to their culture medium, the basal surface of their epithelia developed persistent "blebs" that protruded throu gh gaps in the basal lamina. In addition, they observed that when they added laminin , fib ronectin, and type I collagen , i.e., the ingredients that have been digested by their enzyme mixture, to their culture medium, the blebs retracted spontaneously within 2-6 h , re­gardless of the continued presence of gaps in the basal lamina . In addition, there is evidence that the release of proteolytic enzymes in psoriasis may be excessive [1 9,20). Our observations of the intimate association of BKH with localized areas of electron lu­cency, apparently produced by neutrophils (Fig 2) and endothelial cells (Fig 3), are in support of these concepts. We suggest that the localized areas of electron lucency may represent areas of autodigestion by proteolytic enzymes released from these cells. Further support of the involvement of proteolytic enzymes in

Figure 2. Electron photomicrograph showing a BKH closely associa ted with a localized electron-lucent area (P), lined by cytoplasm fro m an underlying neutrophil (N). The gap in the basal lamina is indi­cated by the space between arrowheads. E = epidermis. Bar = 1 Jl. .

VOL. 87. NO.3 SEPTEMBER 1986

Figure 3. Electron photomicrograph showing close contact and association be­tween a BKH, and localized electron-lu­cent areas (P) lined by cytoplasm from a dermal endothelial cell (E). A = basal ke­ratinocyte. The gap in the basal lamina is indicated by the space betweell arrowheads. Bar = 1 J.L .

disease activity in psoriasis is provided by our recent observations [21] that psoriatic individuals with comcomitant ai-antitrypsin deficiency had a more severe manifestation of the disease . This proteolytic inhibitor, together with az-macroglobulin, provides more than 90% of the antiproteolytic activity in serum [22] . We have, moreover, observed that in these psoriatic patients with associated ai-antitrypsin deficiency, the morphology of some of the BKH were noted to be altered to be broader at the base or to consist of multiple polypoid protrusions into the dermis [23]. lending further support to the concept that the release of excessive proteolytic enzymes m ay playa role in BKH formation in pso­naSlS .

BASAL KERATINOC YTE HERNIATION S IN PSORIASIS 365

It is of interest that individual endotheli al cell s migrating toward an angiogenic stimulus ha ve been noted not to possess a basement membrane [24], and we ha ve reported their presence in psoriasis induced by tape-stripping [25]. Endothelial cells are also ca pable of degrading type [ collagen [26]. which is thought to playa central ro le in maintaining the integrity of the basement mem­brane [1 8,27]. predominantly by reducin g the degrad ation of basal lamina proteoglycan [27]. It is interesting to note in tape-stripped uninvolved psoriatic skin that although Langerhans cell s and lym­phocytes were noted to cross the basement membrane as early as 2 nl.in after tape-stripping [25]. BKH were observed only after 1-2 weeks . On the other hand , endotheli al cell proliferation and

Table II. Association of Basal Keratinocyte Herniations (BKH) and Gaps in Basal Lamina (BL) with Electron-Lucent Areas in Dermis

Psoriasis Involved:

Untreated Treated.

Uninvolved Resolved Seborrheic Dermatitis Eczema (all types) Lichen planus Pi tyriasis rubra pi laris

'ElL = electron-lucent. ' Highly significant (p < 0.001).

No .l200 J.LlTI

Association with ElL Areas'

4 o o o 2 2 o o

Gaps in BL

BL Length

Not Associated with ElL

Areas

46 76 17 14 34 51 6

27

No.l200 J.Lm

Association with ElL Areas

58" 9 o o o o o I

BKH

BL Length

Not Associated with ElL

Areas

o 3 o o o o o o

366 HENG ET AL

intraepidermal neutrophils were noted just prior to the formation of BKH [25], sugges tin g, perhaps, that these cellular types may playa deflI1itive role in BKH formation. More studies exploring these concepts are, therefore, indicated .

We ackllowledge til e editorial skills of Eletl ll or M. Rllssell .

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