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Involvement of Cytokines, DNA Damage, and Reactive Oxygen Intermediates in Ultraviolet Radiation-Induced Modulation of Intercellular Adhesion Molecule-1 Expression
Involvement of Cytokines, DNA Damage, and Reactive Oxygen Intermediates in Ultraviolet Radiation-Induced Modulation of Intercellular Adhesion Molecule-l Expression Jean Krutmann and Markus Grewe Clinical and Experimental Photo dennatology, Department of Dermatology, Heinrich-Heme-University, Dusseldorf, Gcnnany
By virtue of its capacity to serve as a counter-receptor
tern, UVR exposure of human keratinocytes leads to
for lyntphocyte function-associated antigen-1. inter cellular adhesion molecule-1 (ICAM-1) plays a pivotal
the release of IL-1a, which in turn up-regulates the expression of IL-1 receptor type 1 molecules on the
role in generation and maintenance of immunologic!
keratinocyte surface, thereby increasing the sensitiv
inflammatory skin diseases by mediating leukocyte!
ity of these cells toward IL-1a. As a consequence,
keratinocyte adhesion. Ultraviolet radiation (UVR)
irradiated keratinocytes are capable of responding to
may exert both antiinflammatory effects (e.g .• UV
endogenously produced IL-1a by increasing ICAM-1
phototherapy)
and
proinflammatory
effects
(e.g.,
expression. Modulation of keratinocyte ICAM-1 ex
triggering of photosensitive skin diseases) on human
pression after UVR exposure may be observed after
skin. Recent evidence indicates that UVR-induced
both short-wave UVR (UVB; 280-320 nm) and long
changes of keratinocyte ICAM-1 expression consti
wave UVR (UVA1; 340-400 nm). The photobiologic
tute the molecular basis for these ambivalent prop
mechanisms underlying UVB versus UVAt radiation
erties of UVR , as UVR is able to exert two separate
induced ICAM-1 modulation have been found to
and even opposite effects on ICAM-1 expression. As
differ.
an antiinflammatory effect, UVR may inhibit cyto
radiation-induced modulation of ICAM-1 expression
kine-induced up-regulation of keratinocyte ICAM-1
appears to he mediated via the induction of DNA
Although not completely
delineated,
UVB
expression, whereas induction of ICAM-1 expression
damage, whereas UVAt radiation effects involve the
by UVR represents a proinflammatory activity. This
generation of reactive oxygen intermediates. ] Invest
latter effect is mediated by an autocrine mechanism
Dermatol 105:67S-70S, 1995
involving interleukin (IL)-1a. In this autocrine sys--
U
--- - -------
ltraviolet radiation (UVR) exposure has a major
(ICAM-1) by UVR are of particular interest. Identical doses of
impact on the generation as well as the course of
UVBR or UVA 1 R may exert either anti- or proinflammatory
in human
effects at the level of ICAM-l expression. Thus, UVR-induced
skin [1]. By using short-wave UVR (UVB; 280-
modulation of ICAM-l expression represents a photoimmunologic
inflammatory/immunologic reactions
model system to study the ambivalent nature of UVR-induced
320 nm) and/or long-wave UVR (UVA1; 340-
400 nm) as a therapeutic tool, inflammatory skin diseases including
immunomodulation.
psoriasis and atopic dermatitis may be efficiently treated [2,3]. On the other hand, UVB radiation (UVBR) and/or UVA1 radiation
the physical interaction of keratinocytes with leukocytes, and
(UVA1R)
Keratinocyte ICAM-l expression is an important prerequisite for
are well-known triggers for a variety of photosensitive
down- or up-regulation of JCAM-1 expression produced by UVR
skin diseases, such as polymorphous light eruption and lupus
profoundly affects the course of inflammatory/immune responses in
erythematosus [4,5]. In recent years, substantial progress has been
the skin [6-8]. Accordingly, UVR may inhibit cytokine-induced
made in understanding the molecular basis of these ambivalent effects associated with UVR exposure. In this regard, studies on the
antiinflammatory effect is thought to account for the elfectiveness of
phot oimmunologic and photobiologic mechanisms operative in the regulation of the expression of intercellular adhesion molecule-l
other hand, induction of keratinocyte ICAM-l expression by UVR
keratinocyte ICAM-l expression both ill !litro and ill !lillO, * and this UVR phototherapy in inflammatory skin diseases [9,10]. On the has heen proposed to represent an important trigger mechanism that may have relevance in the pathogenesis of photosensitive lupus
Reprint requests to: Dr. Jean Krutmann, Clinical and Experimental Photodermatology, Department of Dermatology, Heinrich-Heine-Univer sity, Moorenstrasse 5, 0-40225 Dusseldorf, G erman y .
Abbreviations: IL-llU/IJ, necrosis factor receptor.
interlcukin-l receptor type
0022-202X/95/S09.50
•
I1I1; TNFR,
SSDHJ022-202X(95)OOl S4-D
-*
M,
Kauer
M,
Copyright
67S
it)
---- ------
K, Grether-Beck S, KrutmannJ: u ltrav iol et B rad iation (UVBR):
Gyufko
A novel antiinflammatory property of inhibition of kcratinocyte (KC) ICAM-1 (ab str) . J bwest Den/wt,,/ 103:428, 1994.
tumor
•
Stege H, Grewe
expression in vivo in human skin
1995 by The Society for Investigative Dermatology , Inc.
68S
KRUTMANN AND GREWE
THE JOURNAL OF INVESTIGATIVE DERMATOLO(;\
erythematosus and polymorphous light eruption [5,7,10,11]. In this review, we summarize the current knowledge about the mechanisms by which UVER and UVA1R regulate ICAM-l expression. From these studies, a more generalized concept emerges concerning the molecular basis of UVBR- versus UVAIR-induced immunomodulation, in which UVBR exerts its effects Ilia the induction of DNA damage, whereas UVAIR induced immunomodulation involves the generation of reactive oxygen intermediates.
UVR INHIBITS THE CYTOKlNE-INDUCED EXPRESSION OF ICAM-l Keratinocyte ICAM-1 expression is markedly enhanced upon stim ulation of cells with the proinflammatory cytokines interferon (IFN)-y and tumor necrosis factor (TNF)-a, and this up-regulation may be efficiently blocked both ill vitro and in vivo* if keratinocytes are exposed to sublethal doses of UVBR before cytokine stimula tion [9,10]. Further characterization of this antiinflammatory effect revealed that UVER inhibited ICAM-l expression at a pretransla tional level by suppressing IFN-y-induced up-regulation of ICAM-l mRNA steady-state levels* [12]. In addition, UVBR induced inhibition of keratinocyte ICAM-l expression was found to be transient in nature [12]. This conclusion was derived from ill vitro as well as ill I';VO* studies demonstrating that cytokine-induced ICAM-l mRNA and protein expression were inhihited by UVER if irradiated cells were stimulated with cytokines immediately after UVBR exposure. In contrast, cytokine stimulation effectively up regulated ICAM-1 expression in irradiated cells if cytokines were added after a certain period following UVB irradiation. Specifically, restoration of cytokine responsiveness in irradiated cells was ob served for IFN-y as early as 12 h after irradiation, whereas restoration of TNF-a responsiveness required a longer restoration period of 24- 48 h [12]. Delayed restoration ofTNF-a responsive ness in irradiated keratinocytes was most likely due to UVBR induced modulation of keratinocyte expression of the 55-kD tumor necrosis factor receptor (TNFR). This receptor molecule, which mediates TNF-a-induced ICAM-l expression in human keratino cytes, was down-regulated in UVB-irradiated keratinocytes shortly (0 -12 h) after UVER exposure by an autocrine mechanism involv ing the ligand TNF-a itself, whereas 24 h after irradiation, 55-kD TNFR expression was restored [ 13 14]. The UVE doses required to modulate 55-kD TNFR and ICAM-l expression in keratinocytes were identical, and the time courses of both UVBR-induced regulatory events are consistent with a cause-effect relation. It is therefore probable that down-regulation of 55-kD TNFR expres sion contributes to the failure of UVB-irradiated, TNF-a-stimu lated keratinocytes to up-regulate ICAM-t expression, whereas 55-kD TNFR reexpression at 24 h at least partially explains restoration of TNF-a responsiveness in irradiated cells. In contrast to TNF-a, IFN-y is not produced by UVE-irradiated keratinocytes, and thus, IFN-y-mediated down-regulation of IFN-y receptors in irradiated keratinocytes does not occur. These differences in the kinetics of cytokine receptor expression may explain the earlier restoration of IFN-y responsiveness as compared with TNF-a responsiveness. From a phototherapeutic point of view, it is remarkable that after restoration of cytokine responsiveness in irradiated keratinocytes, UVBR-induced inhibition of cytokine-mediated ICAM-l expres sion could be reinduced both ill vitro and ill vivo* by subsequent UVBR exposures. Taken together, these ill vitro and ill 1'ivo observations indicate that UVER-induced inhibition of cytokine mediated up-regulation of keratinocyte ICAM-l expression ac counts at least in part for the effectiveness of UV phototherapy in inflammatory skin diseases. ,
INTERLEUKIN (IL)-la MEDIATES UVR-INDUCED KERATINOCYTE ICAM-l EXPRESSION BY AN AUTOCRINE MECHANISM INVOLVING IL-l RECEPTOR TYPE I In addition to suppressing cytokine-mediated ICAM-I expression, UVBR was found to up-regulate constitutive ICAM-I expression in human keratinocytes ill vitro as well as ill vi1'ot [10,12 ]. UVR induced up-regulation of keratinocyte ICAM-l expression required a 24-48-h incubation period, indicating an indirect, possibly autocrine mechanism. In keeping with this hypothesis was the observation that UVER-induced ICAM-l expression could be mimicked in unirradiated, transformed human keratinocytes (KB cells) if these cells were cultured in the presence of conditioned medium that had been harvested from UVE-irradiated cells [12]. It is interesting that addition of neutralizing antibodies directed against TNF-a to conditioned medium and/or to UVB-irradiated keratinocytes (KB cells as well as normal human keratinocytes) failed to hlock this ICAM-I-inducing activity, although in the same experiments these antibodies were highly efficient in inhibiting several other autocrine effects, which were mediated by UVER induced, endogenously produced TNF-a [14,15J. The failure of TNF-a to mediate UVBR-induced ICAM-l expression could not be explained by UVBR-induced modulation of keratinocyte 55-kD TNFR expression, because at this later time point (24- 48 h after irradiation), 55-kD TNFR expression was not only restored, but was even increased above baseline levels in UVB-irradiated cell> [14]. From these studies, it was concluded that UVER-induced ICAM-l expression was mediated by a soluble factor, which wa, produced in increased amounts by keratinocytes after UVER exposure and which was distinct from TNF-a. Recent studies using long-term cultured, normal human kerati nocytes have demonstrated this mediator to be identical to the cytokine IL-la,:J: which mediates UVER-induced ICAM-1 expres sion 1'ia an autocrine mechanism involving two consecutive step' (Fig 1). In a first step (0-24 h after irradiation), UVER-induced. endogenously produced IL-la was found to increase keratinocyte expression of the IL-l receptor type 1 (IL-lRI). § The IL-J RI is the receptor molecule responsible for signaling IL-l-mediated effects in human keratinocytes, and IL-la-induced up-regulation of IL-IRI expression in keratinocytes leads to a markedly increased capacity of human keratinocytes to respond to IL-1a. This increase in IL-1 responsiveness was found to be an indispensable prerequisite for keratinocytes to be able to up-regulate ICAM-l expression after IL-la stimulation. Accordingly, IL-la failed to induce ICAM-l expression in resting, unirradiated keratinocytes, but was able to do so if keratinocytes were either UVE irradiated:J: or left unirradiated but primed with IL-la to increase IL-IRI expression.§ There is increasing evidence that under certain conditions, human keratino cytes may also express IL-1 receptor type II (IL-1RII).§ In contrast to IL-1RI, which functions as the signaling receptor for IL-1, IL-1RII serves as a decoy receptor by limiting rather than promot ing IL-l-mediated responses [16]. Under in vitro conditions, we have not been able to detect IL-lRII expression in unirradiated or irradiated normal human keratinocytes.§ This was in contrast to transformed keratinocytes, in which IL-IRII expression was rapidly induced by UVBR.§ Moreover, UVBR-induced IL-1RII up-reg ulation did not prevent, but clearly limited, UVBR-induced ICAM-l up-regulation. The nonsignaling IL-IRII appears to be expressed in human skin in more differentiated keratinocytes, the t Norris DA, Bennion SD, Middleton MH: Further definition of the effects of ultraviolet light on ICAM-l (CD54) expression by human keratinocytes (abstr).) iIlllest Denllato/ 94:560, 1990. t KrunnalmJ, Trefzer U, Kapp A, Sch6pfE. Luger TA: Interleukin-la, but not tumor necrosis factor a, mediates ultraviolet B rarnation-induced ICAM-l expression in human keratinocytes (abstr).) ]'lJIest Dern1ato/ 98:578, 1992. § Grewe M. Gyutko K, Budnik A. Olaizola-Horn S, Krutmann J: Expression. regulation, and function of interleukin 1 receptors type I and II in nonnal and transfonned human keratinocytes (manuscript submitted for publication).
VOL.
105, NO.
I, SUPPLEMENT,
JULY
UV EFFECTS ON
1995
ICAM-!
EXPRESSION
69S
IL-la
UVB
UVA1 IL-la
PLASMA Ii�d peroxidation f--- Vi!. E ----� --------,---.--������==��-M EMBRANEL----' o,
i
IL-lRI
Reactive Oxygen Intermediates
SOD� GSH
.-->
es�
•
CYTOPLASM eICAM-l
'0,
Transcriptionfactor-activation
mRNA
� NUCLEUS
ICAM-)
second step
first step
2.
Figure
1. Mechanism of UVBR-induced up-regulation of keratino ICAM-l expression. Upon irradiation, keratinocytes secrete IL-la, which hinds in an autocrine manner to IL-lRI, which in tum leads to increased synthesis and expressiun uf IL-llU molecules (first step). Up-regulation of JL-1RI expression increases the sensitivity ofkeratillocytes toward IL-Ia, and these cells are now able to respond to endogenously produced IL-l a by Figure cyte
up-regulating ICAM-l mRNA and surface expression (second step).
The proposed photobiologic mechanism of
induced up-regulation ofkeratinocyte
ICAM-1
UVA1 R
expression. UVAIR
leads to the generation of free oxygen intermediates, which either directly and lor
pia
the gcneration of lipid peroxides induce
rCAM-1 mRNA
and
surface expression. SOD, superoxide dismutase; GSH, glutathione.
do so in transformed cells. ** In contrast, UVBR regulated ICAM-l expression in both normal and transformed keratinocytes, indicat
cells that receive the largest UV dose [17]. It is therefore tempting to speculate that within the upper layers of the epidermis, IL-l RII may help to prevent overshooting responses to UVBR-induced IL-1 ex.
In keeping with this hypothesis is the observation by
Bennion et al that keratinocyte ICAM-1 expression in UV-irradi ated skin or in lesional skin of patients with subacute cutaneous lupus erythematosus does not follow a UV dose-dependent gradi ent, but rather is diffuse throughout the epidermis.'\! Further studies are required to determine the role of IL-1 ex for UVR-induced ICAM-l expression ill vivo in human skin. In this regard, it will be of particular interest to assess whether the same two-step, IL-IO' mediated autocrine loop identified ill pitro will also mediate UVR induced keratinocyte ICAM-l expression in UVR-triggered skin diseases such as polymorphous light eruption and lupus erythematosus. UVER AND UVA1R BOTH AFFECT ICAM-1 EXPRESSION ' ALTHOUGH BY DIFFERENT PHOTOBIOLOGIC MECHANISMS expression,
i.e.,
inhibition of cytokine-induced ICAM-1 up-regulation early after UVR exposure and induction of constitutive ICAM-1 expression at later time points, could be observed not only upon exposure of cells (UVA1;
but also upon irradiation with long-wave UVAR
340- 400
nm). ** Although the immunologic mechanisms
involved in UVBR-induced and UVAIR-induced modulation of ICAM-l expression were found to be essentially identical, it now appears that the photobiologic processes underlying these immu noregulatory events differ markedly. In this regard, a key observa tion was made when UVA 1 R-induced modulation of ICAM-l expression was compared in normal l'erSIIS transformed human keratinocytes. Specifically, UVA1 R was perfectly capable of mod ulating ICAM-1 expression in normal keratinocytes, but failed to
'II
Bennion SO, Middleton
MH,
David-Bajar
KM,
expression in three diseases indicate different triggers of disease.
J IIIl'cst
(in press).
H, Budnik A, Luscher P, Grewe M, Tyrrell RM, Krutmann J: Ultraviolet Al radiation induced immunomodulation is mediated via the generation of singlet oxygcn: UV AI radiation effects on keratinocyte ICAM-! expression (manuscript submitted **
Olaizola-Horn S, Grether-Beck S, Christuph
for publication).
induced modulation of ICAM-I expression. The availability of UVAIR-resistant and UVA1R-sensitive ke ratinocyte populations allowed us to study further the photobio logic
mechanisms
underlying
UVA1 R-induced
modulation
of
rCAM-1 expression. Studies on the nature of the resistance of transformed human keratinocytes towards UVAIR-induced immu Ilomodulation revealed that the capacity of UVAIR to exert regulatory elfects on the expression of ICAM-l in human kerati nocytes strictly depended on the thiol state of the irradiated cell. Accordingly, UVAIR-resistant, transformed human keratinocytes had a threefold higher content of endogenous glutathione as compared with nonnal keratinocytes. Even more important, reduc tion of endogenous glutathione levels rendered UVAIR-resistant keratinocytes UVA1 R sensitive, and these cells behaved essentially induced ICAM-l expression. ** This finding prompted us to analyze the involvement of reactive oxygen intermediates in UVAl R-induced immunomodulation by exposing keratillocytes to UVA1R in the presence of substances that either inhibited or enhanced the formation of reactive oxygen species.
From these studies, it appeared that UVAIR-induced
modulation of keratinocyte rCAM-l
expression was primarily
mediated hy the generation of singlet oxygen. * * In addition, circumstantial evidence exists that UVAIR-induced lipid peroxi dation
may
be
involved
in UVAIR-induced modulation of
rCAM-l expression, as treatment of keratinocytes with vitamin E was found to inhibit both UVAIR-induced lipid peroxidation and UVAIR-induced ICAM-1 induction in human keratinocytes.** Based on these experiments, we propose that UV AIR-induced modulation of human keratinocyte ICAM-l expression is mediated
via the generation of reactive oxygen intermediates, in particular Brice S, Norris DA:
Varied patterns of epidermal intercellular adhesiun molecule-l (ICAM - l )
Deflllatof
UVBR- and UVA1R-induced immunomodulation, whereas trans fomled cells were sensitive only to UVBR-, and not to UVAIR
the same as normal human keratinocytes with regard to UVAIR
Biphasic modulation of keratinocyte ICAM-l
to UVER,
ing that normal human keratinocytes were sensitive to both
singlet oxygen, which either directly andlor via the generation of lipid peroxides may influence the transcriptional control of the ICAM-l gene
(Fig 2).
Whether identical mechanisms also account
for other UVAIR-induced immunomodulatory effects (e.g., cytokine production) in human keratinocytes remains to be determined [18]. The fact that UVER, in contrast to UVAIR, was capable of regulating ICAM-l expression in both normal and transformed human keratinocytes indicated that UVBR-induced immunomodu-
70S
KRUTMANN AND GREWE
TH E JOURN AL OF INVESTIGATIVE DERMATOLOGY
present within the cytoplasm may additionally contribute to UVBR-induced immunomodulatory eifects [23,241.
UVB
Work disCl/sscd ill this review was sl/pported ill part by grants Kr 871/3-1, 3-2,
alld
3-3 from the Delltsche Forscllllllgsgellleillsclwft; EV5V-CT-91-0034 from the COllllllissioll of the EI/ropean COlllmlf"ities program; alld by B,itish-Germatl
ARC
Project ",,,,,ber 504. We thank Dr. SlIsalllle Grether-Beck, Diisseldorf; for carifl/I/J' reading the manuscript. -------
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ICAW-l mRNA
-.l
.......
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KrutmannJ, Czech W, Diepgen T, Niedner R, Kapp A, Schopf E: High-dose
Inc., Oxford, 1995 1:191-194,1993 UVAI therapy of patients with atopic deTInatitis.] Am Acad Dennatol26:225230, 1992
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Holzle E, Plewig G, Hofmann C. Roser-Maas E: Polymorphous light emption.
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Norris DA: Photoimmunology of lupus erythematosus. In: Krutmann j, Elmcb
Experimental reproduction of skin lesions.] Am Acad Den11atoI7:111-125, 1982 CA (eds.).
PJlOwirmmIHoJogy. Blackwell Science Publications Inc., Oxford.
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Kupper TS: Mechanisms of cutaneous inflammation. Interaction between epi dennal cytukines, adhesion molecules� and leukocytes. Arch Dennatoi t 25: 1406-1412,1989
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Norris DA: Cytokine modulation of adhesion molecules in the regulation of immunologic cytotoxicity to epidermal targets.] T1lvest Demwto/ 95: 111S-120S. 1990
lCA - 1 -�-�mRNA
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KrutmannJ: Regulatory interactions between epidermal cell adhesion molecules and cytokines. In: Luger TA, Schwarz T (cds.). Epidermal C)'tokiltes and Growth Factors. Marcel Dekker, New York, 1994, pp 415-432
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KrutmannJ, Kock A, Schauer E, Parlow F. Kapp A, Forster E, Schopf E, Luger TA: Tumor necrosis factor {3 and ultraviolet radiation are potent regulators of human keratinocyte lCAM-l expression.] bll'est DmllatoI95:127-131, 1990
](1.
Norris OA, Lyons ME, Middleton MH, YohnJ], Kashihara-Sawami M: Ultra violet radiation can either suppress or induce expression of intercellular adhesion molecule-I (ICAM-I) on the surface of cultured human kcratino cytes.] IIII'est DermatoI95:132-138, 1990
DNA damage in UVBR induced inhibition ofICAM-l exp ression. Constitutive ICAM-l mRNA and surface expression in human keratinocytes is low, but may be significantly induced upon stimulation of unirradiated cells with IFN-y (bottom). Induction of DNA damage in keratinocytes by UVBR effectively prevents IFN-y-mediated up-regulation of ICAM-l mRNA and surface expression (top). Figure
3. A
possible model for the role of
II.
Norris PG, BarkerJNWN, Allen MH, Leiferman KM, MacDonald OM, Haskard DO. HawkJLM: Adhesion molecule expression in polymorphic light eruption. ] Invest Dennatol 99:504-508, 1994
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Krutmann J, Czech W, Parlow F, Trefzer U. Kapp A, Schopf E, Luger TA: Ultraviolet radiation effects on human kcratinocyte ICAM-1 expression: UV induced inhibition of cytokinc-induced ICAM-l mRNA expression is transient, differentially rcstored for IFN -y versus TNFa. and followed by ICAM-l induction via a TNFa-like pathway.] 1Il1'est Dennatol 98:923-928, 1992
13.
latory eifects did not depend on glutathione levels in keratinocytes, and thus involved mechanisms alternative to those identified for UVAiR-induced ICAM-l regulation. In studies comparing nornlal cells and xeroderma pigmentosum cells, which are deficient in DNA excision repair, we have observed that generation of DNA photoproducts was required for the ability of UVBR to inhibit cytokine-mediated ICAM-l up-regulation (Fig 3) [19]. Specifi cally, dose-response experiments revealed that in xeroderma pig mentosum cells, 2-3-fold lower UVBR doses were required to inhibit IFN--y--induced ICAM-l up-regulation to an extent equiv alent to that observed in normal cells [19]. In addition, in xero derma pigmentosum complementation group D cells, no restora tion of IFN-y responsiveness was observed for up to 48 h after exposure, but responsiveness was restored in xeroderma pigmen tosum complementation group C cells after 24 h. This was in marked contrast to normal cells, in which inhibition of IFN--y- induced ICAM-l expression was restored 12 h after UVBR exposure [19J. Ongoing studies are directed at assessing the fol lowing: 1) the involvement of a specific photoproduct in UVBR induced irnmunomodulation, and 2) whether the generation of photoproducts per se or rather the formation of DNA excision products or the activation of repair processes is required for UVBR-induced modulation of ICAM-l expression. These studies corroborate and extend previous work in animal models that suggested the relevance of DNA as a chromophore for UVBR induced immunomodulation [20-22]. At present, however, it is not possible to exclude that target molecules outside of the nucleus and
Trefzer U. Brockhaus M, Loetscher H. Parlow F, Kapp A. Schopf E. Krutmann
J:
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ICAM-l
expression.] Invest Dermatol 97:911-916,1991 14.
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Bohnert E, Jung EG: Evidence that DNA damage is a mediate in
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Yarosh DB: The role of DNA danlage and lTV-induced cytokines in skin cancer.
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Devary Y. Rosette C, DiDonato JA, Karin M: Nf-kB activation by uLtraviolet light is not dependent on a nuclear signal. Sriellce 26 1:1442-1445, 1993
By virtue of its capacity to serve as a counter-receptor
tern, UVR exposure of human keratinocytes leads to
for lyntphocyte function-associated antigen-1. inter cellular adhesion molecule-1 (ICAM-1) plays a pivotal
the release of IL-1a, which in turn up-regulates the expression of IL-1 receptor type 1 molecules on the
role in generation and maintenance of immunologic!
keratinocyte surface, thereby increasing the sensitiv
inflammatory skin diseases by mediating leukocyte!
ity of these cells toward IL-1a. As a consequence,
keratinocyte adhesion. Ultraviolet radiation (UVR)
irradiated keratinocytes are capable of responding to
may exert both antiinflammatory effects (e.g .• UV
endogenously produced IL-1a by increasing ICAM-1
phototherapy)
and
proinflammatory
effects
(e.g.,
expression. Modulation of keratinocyte ICAM-1 ex
triggering of photosensitive skin diseases) on human
pression after UVR exposure may be observed after
skin. Recent evidence indicates that UVR-induced
both short-wave UVR (UVB; 280-320 nm) and long
changes of keratinocyte ICAM-1 expression consti
wave UVR (UVA1; 340-400 nm). The photobiologic
tute the molecular basis for these ambivalent prop
mechanisms underlying UVB versus UVAt radiation
erties of UVR , as UVR is able to exert two separate
induced ICAM-1 modulation have been found to
and even opposite effects on ICAM-1 expression. As
differ.
an antiinflammatory effect, UVR may inhibit cyto
radiation-induced modulation of ICAM-1 expression
kine-induced up-regulation of keratinocyte ICAM-1
appears to he mediated via the induction of DNA
Although not completely
delineated,
UVB
expression, whereas induction of ICAM-1 expression
damage, whereas UVAt radiation effects involve the
by UVR represents a proinflammatory activity. This
generation of reactive oxygen intermediates. ] Invest
latter effect is mediated by an autocrine mechanism
Dermatol 105:67S-70S, 1995
involving interleukin (IL)-1a. In this autocrine sys--
U
--- - -------
ltraviolet radiation (UVR) exposure has a major
(ICAM-1) by UVR are of particular interest. Identical doses of
impact on the generation as well as the course of
UVBR or UVA 1 R may exert either anti- or proinflammatory
in human
effects at the level of ICAM-l expression. Thus, UVR-induced
skin [1]. By using short-wave UVR (UVB; 280-
modulation of ICAM-l expression represents a photoimmunologic
inflammatory/immunologic reactions
model system to study the ambivalent nature of UVR-induced
320 nm) and/or long-wave UVR (UVA1; 340-
400 nm) as a therapeutic tool, inflammatory skin diseases including
immunomodulation.
psoriasis and atopic dermatitis may be efficiently treated [2,3]. On the other hand, UVB radiation (UVBR) and/or UVA1 radiation
the physical interaction of keratinocytes with leukocytes, and
(UVA1R)
Keratinocyte ICAM-l expression is an important prerequisite for
are well-known triggers for a variety of photosensitive
down- or up-regulation of JCAM-1 expression produced by UVR
skin diseases, such as polymorphous light eruption and lupus
profoundly affects the course of inflammatory/immune responses in
erythematosus [4,5]. In recent years, substantial progress has been
the skin [6-8]. Accordingly, UVR may inhibit cytokine-induced
made in understanding the molecular basis of these ambivalent effects associated with UVR exposure. In this regard, studies on the
antiinflammatory effect is thought to account for the elfectiveness of
phot oimmunologic and photobiologic mechanisms operative in the regulation of the expression of intercellular adhesion molecule-l
other hand, induction of keratinocyte ICAM-l expression by UVR
keratinocyte ICAM-l expression both ill !litro and ill !lillO, * and this UVR phototherapy in inflammatory skin diseases [9,10]. On the has heen proposed to represent an important trigger mechanism that may have relevance in the pathogenesis of photosensitive lupus
Reprint requests to: Dr. Jean Krutmann, Clinical and Experimental Photodermatology, Department of Dermatology, Heinrich-Heine-Univer sity, Moorenstrasse 5, 0-40225 Dusseldorf, G erman y .
Abbreviations: IL-llU/IJ, necrosis factor receptor.
interlcukin-l receptor type
0022-202X/95/S09.50
•
I1I1; TNFR,
SSDHJ022-202X(95)OOl S4-D
-*
M,
Kauer
M,
Copyright
67S
it)
---- ------
K, Grether-Beck S, KrutmannJ: u ltrav iol et B rad iation (UVBR):
Gyufko
A novel antiinflammatory property of inhibition of kcratinocyte (KC) ICAM-1 (ab str) . J bwest Den/wt,,/ 103:428, 1994.
tumor
•
Stege H, Grewe
expression in vivo in human skin
1995 by The Society for Investigative Dermatology , Inc.
68S
KRUTMANN AND GREWE
THE JOURNAL OF INVESTIGATIVE DERMATOLO(;\
erythematosus and polymorphous light eruption [5,7,10,11]. In this review, we summarize the current knowledge about the mechanisms by which UVER and UVA1R regulate ICAM-l expression. From these studies, a more generalized concept emerges concerning the molecular basis of UVBR- versus UVAIR-induced immunomodulation, in which UVBR exerts its effects Ilia the induction of DNA damage, whereas UVAIR induced immunomodulation involves the generation of reactive oxygen intermediates.
UVR INHIBITS THE CYTOKlNE-INDUCED EXPRESSION OF ICAM-l Keratinocyte ICAM-1 expression is markedly enhanced upon stim ulation of cells with the proinflammatory cytokines interferon (IFN)-y and tumor necrosis factor (TNF)-a, and this up-regulation may be efficiently blocked both ill vitro and in vivo* if keratinocytes are exposed to sublethal doses of UVBR before cytokine stimula tion [9,10]. Further characterization of this antiinflammatory effect revealed that UVER inhibited ICAM-l expression at a pretransla tional level by suppressing IFN-y-induced up-regulation of ICAM-l mRNA steady-state levels* [12]. In addition, UVBR induced inhibition of keratinocyte ICAM-l expression was found to be transient in nature [12]. This conclusion was derived from ill vitro as well as ill I';VO* studies demonstrating that cytokine-induced ICAM-l mRNA and protein expression were inhihited by UVER if irradiated cells were stimulated with cytokines immediately after UVBR exposure. In contrast, cytokine stimulation effectively up regulated ICAM-1 expression in irradiated cells if cytokines were added after a certain period following UVB irradiation. Specifically, restoration of cytokine responsiveness in irradiated cells was ob served for IFN-y as early as 12 h after irradiation, whereas restoration of TNF-a responsiveness required a longer restoration period of 24- 48 h [12]. Delayed restoration ofTNF-a responsive ness in irradiated keratinocytes was most likely due to UVBR induced modulation of keratinocyte expression of the 55-kD tumor necrosis factor receptor (TNFR). This receptor molecule, which mediates TNF-a-induced ICAM-l expression in human keratino cytes, was down-regulated in UVB-irradiated keratinocytes shortly (0 -12 h) after UVER exposure by an autocrine mechanism involv ing the ligand TNF-a itself, whereas 24 h after irradiation, 55-kD TNFR expression was restored [ 13 14]. The UVE doses required to modulate 55-kD TNFR and ICAM-l expression in keratinocytes were identical, and the time courses of both UVBR-induced regulatory events are consistent with a cause-effect relation. It is therefore probable that down-regulation of 55-kD TNFR expres sion contributes to the failure of UVB-irradiated, TNF-a-stimu lated keratinocytes to up-regulate ICAM-t expression, whereas 55-kD TNFR reexpression at 24 h at least partially explains restoration of TNF-a responsiveness in irradiated cells. In contrast to TNF-a, IFN-y is not produced by UVE-irradiated keratinocytes, and thus, IFN-y-mediated down-regulation of IFN-y receptors in irradiated keratinocytes does not occur. These differences in the kinetics of cytokine receptor expression may explain the earlier restoration of IFN-y responsiveness as compared with TNF-a responsiveness. From a phototherapeutic point of view, it is remarkable that after restoration of cytokine responsiveness in irradiated keratinocytes, UVBR-induced inhibition of cytokine-mediated ICAM-l expres sion could be reinduced both ill vitro and ill vivo* by subsequent UVBR exposures. Taken together, these ill vitro and ill 1'ivo observations indicate that UVER-induced inhibition of cytokine mediated up-regulation of keratinocyte ICAM-l expression ac counts at least in part for the effectiveness of UV phototherapy in inflammatory skin diseases. ,
INTERLEUKIN (IL)-la MEDIATES UVR-INDUCED KERATINOCYTE ICAM-l EXPRESSION BY AN AUTOCRINE MECHANISM INVOLVING IL-l RECEPTOR TYPE I In addition to suppressing cytokine-mediated ICAM-I expression, UVBR was found to up-regulate constitutive ICAM-I expression in human keratinocytes ill vitro as well as ill vi1'ot [10,12 ]. UVR induced up-regulation of keratinocyte ICAM-l expression required a 24-48-h incubation period, indicating an indirect, possibly autocrine mechanism. In keeping with this hypothesis was the observation that UVER-induced ICAM-l expression could be mimicked in unirradiated, transformed human keratinocytes (KB cells) if these cells were cultured in the presence of conditioned medium that had been harvested from UVE-irradiated cells [12]. It is interesting that addition of neutralizing antibodies directed against TNF-a to conditioned medium and/or to UVB-irradiated keratinocytes (KB cells as well as normal human keratinocytes) failed to hlock this ICAM-I-inducing activity, although in the same experiments these antibodies were highly efficient in inhibiting several other autocrine effects, which were mediated by UVER induced, endogenously produced TNF-a [14,15J. The failure of TNF-a to mediate UVBR-induced ICAM-l expression could not be explained by UVBR-induced modulation of keratinocyte 55-kD TNFR expression, because at this later time point (24- 48 h after irradiation), 55-kD TNFR expression was not only restored, but was even increased above baseline levels in UVB-irradiated cell> [14]. From these studies, it was concluded that UVER-induced ICAM-l expression was mediated by a soluble factor, which wa, produced in increased amounts by keratinocytes after UVER exposure and which was distinct from TNF-a. Recent studies using long-term cultured, normal human kerati nocytes have demonstrated this mediator to be identical to the cytokine IL-la,:J: which mediates UVER-induced ICAM-1 expres sion 1'ia an autocrine mechanism involving two consecutive step' (Fig 1). In a first step (0-24 h after irradiation), UVER-induced. endogenously produced IL-la was found to increase keratinocyte expression of the IL-l receptor type 1 (IL-lRI). § The IL-J RI is the receptor molecule responsible for signaling IL-l-mediated effects in human keratinocytes, and IL-la-induced up-regulation of IL-IRI expression in keratinocytes leads to a markedly increased capacity of human keratinocytes to respond to IL-1a. This increase in IL-1 responsiveness was found to be an indispensable prerequisite for keratinocytes to be able to up-regulate ICAM-l expression after IL-la stimulation. Accordingly, IL-la failed to induce ICAM-l expression in resting, unirradiated keratinocytes, but was able to do so if keratinocytes were either UVE irradiated:J: or left unirradiated but primed with IL-la to increase IL-IRI expression.§ There is increasing evidence that under certain conditions, human keratino cytes may also express IL-1 receptor type II (IL-1RII).§ In contrast to IL-1RI, which functions as the signaling receptor for IL-1, IL-1RII serves as a decoy receptor by limiting rather than promot ing IL-l-mediated responses [16]. Under in vitro conditions, we have not been able to detect IL-lRII expression in unirradiated or irradiated normal human keratinocytes.§ This was in contrast to transformed keratinocytes, in which IL-IRII expression was rapidly induced by UVBR.§ Moreover, UVBR-induced IL-1RII up-reg ulation did not prevent, but clearly limited, UVBR-induced ICAM-l up-regulation. The nonsignaling IL-IRII appears to be expressed in human skin in more differentiated keratinocytes, the t Norris DA, Bennion SD, Middleton MH: Further definition of the effects of ultraviolet light on ICAM-l (CD54) expression by human keratinocytes (abstr).) iIlllest Denllato/ 94:560, 1990. t KrunnalmJ, Trefzer U, Kapp A, Sch6pfE. Luger TA: Interleukin-la, but not tumor necrosis factor a, mediates ultraviolet B rarnation-induced ICAM-l expression in human keratinocytes (abstr).) ]'lJIest Dern1ato/ 98:578, 1992. § Grewe M. Gyutko K, Budnik A. Olaizola-Horn S, Krutmann J: Expression. regulation, and function of interleukin 1 receptors type I and II in nonnal and transfonned human keratinocytes (manuscript submitted for publication).
VOL.
105, NO.
I, SUPPLEMENT,
JULY
UV EFFECTS ON
1995
ICAM-!
EXPRESSION
69S
IL-la
UVB
UVA1 IL-la
PLASMA Ii�d peroxidation f--- Vi!. E ----� --------,---.--������==��-M EMBRANEL----' o,
i
IL-lRI
Reactive Oxygen Intermediates
SOD� GSH
.-->
es�
•
CYTOPLASM eICAM-l
'0,
Transcriptionfactor-activation
mRNA
� NUCLEUS
ICAM-)
second step
first step
2.
Figure
1. Mechanism of UVBR-induced up-regulation of keratino ICAM-l expression. Upon irradiation, keratinocytes secrete IL-la, which hinds in an autocrine manner to IL-lRI, which in tum leads to increased synthesis and expressiun uf IL-llU molecules (first step). Up-regulation of JL-1RI expression increases the sensitivity ofkeratillocytes toward IL-Ia, and these cells are now able to respond to endogenously produced IL-l a by Figure cyte
up-regulating ICAM-l mRNA and surface expression (second step).
The proposed photobiologic mechanism of
induced up-regulation ofkeratinocyte
ICAM-1
UVA1 R
expression. UVAIR
leads to the generation of free oxygen intermediates, which either directly and lor
pia
the gcneration of lipid peroxides induce
rCAM-1 mRNA
and
surface expression. SOD, superoxide dismutase; GSH, glutathione.
do so in transformed cells. ** In contrast, UVBR regulated ICAM-l expression in both normal and transformed keratinocytes, indicat
cells that receive the largest UV dose [17]. It is therefore tempting to speculate that within the upper layers of the epidermis, IL-l RII may help to prevent overshooting responses to UVBR-induced IL-1 ex.
In keeping with this hypothesis is the observation by
Bennion et al that keratinocyte ICAM-1 expression in UV-irradi ated skin or in lesional skin of patients with subacute cutaneous lupus erythematosus does not follow a UV dose-dependent gradi ent, but rather is diffuse throughout the epidermis.'\! Further studies are required to determine the role of IL-1 ex for UVR-induced ICAM-l expression ill vivo in human skin. In this regard, it will be of particular interest to assess whether the same two-step, IL-IO' mediated autocrine loop identified ill pitro will also mediate UVR induced keratinocyte ICAM-l expression in UVR-triggered skin diseases such as polymorphous light eruption and lupus erythematosus. UVER AND UVA1R BOTH AFFECT ICAM-1 EXPRESSION ' ALTHOUGH BY DIFFERENT PHOTOBIOLOGIC MECHANISMS expression,
i.e.,
inhibition of cytokine-induced ICAM-1 up-regulation early after UVR exposure and induction of constitutive ICAM-1 expression at later time points, could be observed not only upon exposure of cells (UVA1;
but also upon irradiation with long-wave UVAR
340- 400
nm). ** Although the immunologic mechanisms
involved in UVBR-induced and UVAIR-induced modulation of ICAM-l expression were found to be essentially identical, it now appears that the photobiologic processes underlying these immu noregulatory events differ markedly. In this regard, a key observa tion was made when UVA 1 R-induced modulation of ICAM-l expression was compared in normal l'erSIIS transformed human keratinocytes. Specifically, UVA1 R was perfectly capable of mod ulating ICAM-1 expression in normal keratinocytes, but failed to
'II
Bennion SO, Middleton
MH,
David-Bajar
KM,
expression in three diseases indicate different triggers of disease.
J IIIl'cst
(in press).
H, Budnik A, Luscher P, Grewe M, Tyrrell RM, Krutmann J: Ultraviolet Al radiation induced immunomodulation is mediated via the generation of singlet oxygcn: UV AI radiation effects on keratinocyte ICAM-! expression (manuscript submitted **
Olaizola-Horn S, Grether-Beck S, Christuph
for publication).
induced modulation of ICAM-I expression. The availability of UVAIR-resistant and UVA1R-sensitive ke ratinocyte populations allowed us to study further the photobio logic
mechanisms
underlying
UVA1 R-induced
modulation
of
rCAM-1 expression. Studies on the nature of the resistance of transformed human keratinocytes towards UVAIR-induced immu Ilomodulation revealed that the capacity of UVAIR to exert regulatory elfects on the expression of ICAM-l in human kerati nocytes strictly depended on the thiol state of the irradiated cell. Accordingly, UVAIR-resistant, transformed human keratinocytes had a threefold higher content of endogenous glutathione as compared with nonnal keratinocytes. Even more important, reduc tion of endogenous glutathione levels rendered UVAIR-resistant keratinocytes UVA1 R sensitive, and these cells behaved essentially induced ICAM-l expression. ** This finding prompted us to analyze the involvement of reactive oxygen intermediates in UVAl R-induced immunomodulation by exposing keratillocytes to UVA1R in the presence of substances that either inhibited or enhanced the formation of reactive oxygen species.
From these studies, it appeared that UVAIR-induced
modulation of keratinocyte rCAM-l
expression was primarily
mediated hy the generation of singlet oxygen. * * In addition, circumstantial evidence exists that UVAIR-induced lipid peroxi dation
may
be
involved
in UVAIR-induced modulation of
rCAM-l expression, as treatment of keratinocytes with vitamin E was found to inhibit both UVAIR-induced lipid peroxidation and UVAIR-induced ICAM-1 induction in human keratinocytes.** Based on these experiments, we propose that UV AIR-induced modulation of human keratinocyte ICAM-l expression is mediated
via the generation of reactive oxygen intermediates, in particular Brice S, Norris DA:
Varied patterns of epidermal intercellular adhesiun molecule-l (ICAM - l )
Deflllatof
UVBR- and UVA1R-induced immunomodulation, whereas trans fomled cells were sensitive only to UVBR-, and not to UVAIR
the same as normal human keratinocytes with regard to UVAIR
Biphasic modulation of keratinocyte ICAM-l
to UVER,
ing that normal human keratinocytes were sensitive to both
singlet oxygen, which either directly andlor via the generation of lipid peroxides may influence the transcriptional control of the ICAM-l gene
(Fig 2).
Whether identical mechanisms also account
for other UVAIR-induced immunomodulatory effects (e.g., cytokine production) in human keratinocytes remains to be determined [18]. The fact that UVER, in contrast to UVAIR, was capable of regulating ICAM-l expression in both normal and transformed human keratinocytes indicated that UVBR-induced immunomodu-
70S
KRUTMANN AND GREWE
TH E JOURN AL OF INVESTIGATIVE DERMATOLOGY
present within the cytoplasm may additionally contribute to UVBR-induced immunomodulatory eifects [23,241.
UVB
Work disCl/sscd ill this review was sl/pported ill part by grants Kr 871/3-1, 3-2,
alld
3-3 from the Delltsche Forscllllllgsgellleillsclwft; EV5V-CT-91-0034 from the COllllllissioll of the EI/ropean COlllmlf"ities program; alld by B,itish-Germatl
ARC
Project ",,,,,ber 504. We thank Dr. SlIsalllle Grether-Beck, Diisseldorf; for carifl/I/J' reading the manuscript. -------
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DNA damage in UVBR induced inhibition ofICAM-l exp ression. Constitutive ICAM-l mRNA and surface expression in human keratinocytes is low, but may be significantly induced upon stimulation of unirradiated cells with IFN-y (bottom). Induction of DNA damage in keratinocytes by UVBR effectively prevents IFN-y-mediated up-regulation of ICAM-l mRNA and surface expression (top). Figure
3. A
possible model for the role of
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Krutmann J, Czech W, Parlow F, Trefzer U. Kapp A, Schopf E, Luger TA: Ultraviolet radiation effects on human kcratinocyte ICAM-1 expression: UV induced inhibition of cytokinc-induced ICAM-l mRNA expression is transient, differentially rcstored for IFN -y versus TNFa. and followed by ICAM-l induction via a TNFa-like pathway.] 1Il1'est Dennatol 98:923-928, 1992
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latory eifects did not depend on glutathione levels in keratinocytes, and thus involved mechanisms alternative to those identified for UVAiR-induced ICAM-l regulation. In studies comparing nornlal cells and xeroderma pigmentosum cells, which are deficient in DNA excision repair, we have observed that generation of DNA photoproducts was required for the ability of UVBR to inhibit cytokine-mediated ICAM-l up-regulation (Fig 3) [19]. Specifi cally, dose-response experiments revealed that in xeroderma pig mentosum cells, 2-3-fold lower UVBR doses were required to inhibit IFN--y--induced ICAM-l up-regulation to an extent equiv alent to that observed in normal cells [19]. In addition, in xero derma pigmentosum complementation group D cells, no restora tion of IFN-y responsiveness was observed for up to 48 h after exposure, but responsiveness was restored in xeroderma pigmen tosum complementation group C cells after 24 h. This was in marked contrast to normal cells, in which inhibition of IFN--y- induced ICAM-l expression was restored 12 h after UVBR exposure [19J. Ongoing studies are directed at assessing the fol lowing: 1) the involvement of a specific photoproduct in UVBR induced irnmunomodulation, and 2) whether the generation of photoproducts per se or rather the formation of DNA excision products or the activation of repair processes is required for UVBR-induced modulation of ICAM-l expression. These studies corroborate and extend previous work in animal models that suggested the relevance of DNA as a chromophore for UVBR induced immunomodulation [20-22]. At present, however, it is not possible to exclude that target molecules outside of the nucleus and
Trefzer U. Brockhaus M, Loetscher H. Parlow F, Kapp A. Schopf E. Krutmann
J:
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ICAM-l
expression.] Invest Dermatol 97:911-916,1991 14.
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Bohnert E, Jung EG: Evidence that DNA damage is a mediate in
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Yarosh DB: The role of DNA danlage and lTV-induced cytokines in skin cancer.
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Devary Y. Rosette C, DiDonato JA, Karin M: Nf-kB activation by uLtraviolet light is not dependent on a nuclear signal. Sriellce 26 1:1442-1445, 1993