University of Groningen
Apoptotic cell clearance in Systemic Lupus Erythematosus (SLE)
Reefman, Esther
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CHAPTER 2
CHAPTER 2
ABSTRACT
Cell death by apoptosis is a physiological process that enables the elimination of
cells without causing an inflammatory response. In self renewing tissue like the
epidermal layers of the skin, cell numbers are tightly regulated by a delicate balance
between proliferation, differentiation and cell death. Besides cell death by terminal
differentiation in normal skin, cell death can also be induced by exposure to sunlight.
This paper will review the different forms of cell death in the skin and discuss the role
of apoptosis in diseases like skin cancer, psoriasis and systemic lupus erythematosus
(SLE).
16
INTRODUCTION
Apoptosis is a form of programmed cell death which plays a crucial role in
development and homeostasis of the human body. Especially in continuously renewing
tissues like the skin, cell growth and cell death has to be tightly regulated to ensure
correct tissue function and prevent inappropriate excessive growth.
The skin is composed of three distinct layers. The deepest layer is the subcutaneous fat
which is covered by the dermis. The dermis contains blood vessels, nerves, hair
follicles and sweat glands. The top layer is called the epidermis, a continuously
renewing multilayered epithelium with a high turnover rate. The epidermis is
composed of three cell types: Langerhans cells (LC), melanocytes and keratinocytes.
Keratinocytes are the cells which form the, by far, largest part of the epidermis. Basal
and suprabasal keratinocytes together constitute the rapidly proliferating compartment
known as the stratum basale. Terminal keratinocyte differentiation is initiated upon
detachment from the underlying basement membrane into the stratum spinosum (1).
Keratinocytes move upwards through the stratum spinosum into the stratum
granulosum where cells loose their nucleus. The end product is a mechanically rigid,
crosslinked, keratinous envelope in the stratum corneum and is shed as an enucleate
corneocyte. Langerhans cells are the residential dendritic cells of the epidermis.
Melanocytes produce pigment largely composed of melanin and are located in the
basal layer of the epidermis.
Regulation of cell death is very important in the homeostasis of the epidermis.
The turnover rate of keratinocytes is very high and basal and suprabasal keratinocytes
are constantly dividing and finally dying after terminal differentiation in the stratum
granulosum. Furthermore, skin is continuously exposed to external factors as
chemicals and sunlight which can damage keratinocytes. Cell death ensures that
critically damaged keratinocytes are removed to prevent dysfunction of these cells.
ROLE OF APOPTOSIS IN TERMINAL DIFFERENTIATION
During the process of terminal differentiation keratinocytes eventually die.
There is some controversy whether terminal differentiation of keratinocytes is a variant
of apoptosis. Some histological features of terminally differentiating keratinocytes
resemble cellular changes associated with apoptosis, for example, nuclear pyknosis
and fragmentation into nucleosome-sized pieces (2). Both processes also share
activation of endonucleases (3), caspase-3 activity (4) and degradation of DNA.
However several reports also point at the differences between terminal differentiation
and apoptosis. DNA fragmentation is hardly seen in terminally differentiated
keratinocytes (3). We could not detect the active form of caspase-3 in the skin of
healthy controls (5). Furthermore, Gandarillas et al showed in vitro that keratinocytes
undergo terminal differentiation without going into apoptosis (6). Finally, in diseases
characterized by defects in growth, differentiation or adhesion no increase of apoptotic
nuclei were detected in affected epidermis compared to normal skin. In combination
these data strongly suggest that in keratinocytes terminal differentiation and apoptosis
are distinct cellular events.
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Apoptosis in the skin
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SUNLIGHT INDUCED APOPTOSIS IN SKIN
Apoptosis in the skin can be induced by chemicals and ionizing radiation,
however most relevant is exposure of the skin to sunlight. Ultraviolet radiation (UVR)
is a strong inducer of apoptosis in the skin. Apoptotic epidermal keratinocytes, known
as sunburn cells (SBC), were originally described by their distinctive appearance, i.e.
pyknotic nuclei and a shrunken, eosinophilic cytoplasm (7). These features make SBC
easily recognizable in hematoxylin eosin (H&E) stained sections. SBC can be detected
as early as 8 hours after UVR exposure with maximal numbers at 24-48 hours and
disappearance by 60-72 hours (8;9). In vitro, apoptosis will finally result in
permeabilization of the plasma membrane, known as secondary necrosis, which may
lead to inflammation. However, it is believed that this will not occur under
physiological circumstances in vivo due to rapid clearance of apoptotic cells (10). In
the skin, loss of desmosomes on apoptotic keratinocytes (11) will result in more rapid
migration towards the stratum corneum. Subsequently, shedding will clear a large part
of SBC from the skin. In addition, infiltrating macrophages (12), Langerhans cells (13)
and keratinocytes (14) internalize SBC before they reach the stratum corneum.
UV light (200-400 nm) is sub-divided according to wavelength in UVA (315-400
nm), UVB (280-315nm) and UVC (200-280 nm) light. UVC is absorbed by the
stratospheric ozone layer, therefore sunlight usually contains only UVA and UVB.
UVA penetrates through epidermal and dermal layers of the skin, and is weakly
absorbed by bio-molecules. In contrast, UVB does not penetrate much further than the
epidermal layer and is strongly absorbed by DNA and protein. UVB is therefore the
most effective inducer of SBC in the skin (15;16). UVB-induced apoptosis has been
recognized as a complex mechanism in which a variety of signaling pathways are
involved; 1) apoptosis induced by direct DNA-damage, 2) death receptor mediated
apoptosis, and 3) apoptosis via formation of reactive oxygen species (ROS). These
pathways will be discussed below (Fig 1).
1. Apoptosis induced by direct DNA-damage
UVB-induced DNA damage has long been proposed to be the only mediator of
UVB-induced cell death. Absorbance of UVB energy by DNA results in two main
types of lesions known as cyclobutane pyrimidine dimers (CPD) and <6-4>
photoproducts. CPDs comprise 70-80% of all UV photoproducts. The majority of
DNA lesions are removed by nucleotide excision repair (NER), which excises short
single stranded DNA nucleotide stretches (24-32 bases) around the damaged site and
subsequently replaces the excised DNA lesion (17). Increased numbers of SBC are
found in patients suffering from xeroderma pigmentosum, a disease in which genetic
defects disable the nucleotide excision repair process (18). The crucial role of DNA
damage in UVB-induced apoptosis has been confirmed by in vivo studies in humans
showing enhancement of DNA repair by the topical application of repair enzymes
reduces the formation of SBC (19).
Cells with sustained DNA damage show a dramatic rise in their level of tumor
suppressor gene p53 within minutes (20). This upregulation was found to be
proportional to the amount of CPD introduced into genomic DNA by UVB. P53 acts
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Apoptosis in the skin
Figure 1: Schematic representation of the three signaling pathways involved in UVB-induced
apoptosis. 1) Direct DNA-damage upregulates expression and function of p53, which activates
Bax/Bak resulting in cytochrome c release from the mitochondria, subsequently leading to apoptotic
protease activating factor (Apaf-1), caspase-9 and caspase-3 activation and execution of apoptosis.
P53 can also act as a transcription factor which is able to induce the transcription and expression
of death receptors. 2) Death receptor clustering leads to recruitment and activation of FADD which
activates caspase-8 which in turn activates caspase-3. 3) Direct formation of reactive oxygen
species (ROS) results in lipid peroxidation of the mitochondrial membrane and subsequent release
of cytochrome c eventually leading to caspase-3 activation. Bcl-2 and Bcl-XL can inhibit cytochrome
c release while inhibitors of apoptosis (IAPs) can inhibit activation of caspases.
as a transcription factor, inducing transcription of a number of genes whose products
trigger cell-cycle arrest during G1 phase. This allows the cell to repair DNA prior to
DNA synthesis, reducing the DNA mutation rate (8). In mice lacking functional p53
lower amounts of SBC are induced (8). When DNA damage is not repaired sufficiently
apoptosis is initiated by induction of pro-apoptotic factors like Bax or Bak resulting in
release of cytochrome c from mitochondria. An apoptosome is formed by binding of
cytochrome c to apoptotic protease activating factor 1 (Apaf-1) and subsequently
procaspase 9. This critical event leads to the initiation of the apoptotic cascade by
activation of caspase-9 and caspase-3 through cleavage (21). Activation of these
executer caspases in turn leads to cleavage of critical cellular substrates and DNA
fragmentation. This mitochondrial pathway can be regulated by the anti-apoptotic Bcl2 family of proteins (Bcl-2, Bcl-XL) which prevent release of cytochrome c from
19
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mitochondria (22) and inhibitors of apoptosis (IAPs) which prevent caspase cleavage
(23). UV exposure leads to down-regulation of Bcl-2 in human skin, promoting
apoptosis (24). Over-expression of Bcl-XL and Bcl-XS in the epidermis of transgenic
mice contributes to cell survival after UV irradiation (25). UV induced DNA damage
has also been shown to enhance export and transcriptional activation of death receptors
and their ligands through mainly p53-dependent pathways (26;27).
2. Death receptor mediated UV-induced apoptosis
Death receptors are members of the tumor necrosis factor (TNF) receptor
superfamily, including TNF receptor-1, the TNFα related apoptosis inducing ligand
(TRAIL) receptors and Fas (CD95). Although all of these receptors have been linked
to UV-induced apoptosis (27-29), Fas mediated cell death seems to play the most
prominent role (30).
Normal skin keratinocytes express Fas (31). Expression is upregulated in chronically
sun-exposed keratinocytes (32) but also after a single-dose of UVB radiation (33),
demonstrating its involvement in sun-induced damage. CD95 mediates apoptosis via
binding of its ligand, FasL (CD95L), which may be present in soluble form or
expressed on the membrane of cells. FasL is also constitutively expressed by
keratinocytes, and expression is upregulated after UV exposure (29;34). With respect
to UVB-induced apoptosis, Aragene et al demonstrated a direct and ligand
independent activation of Fas, by clustering of Fas at the cell membrane (35).
Clustering of Fas, ligand dependent or independent, leads to activation of the
intracellular death domains which, in turn, can bind Fas-associated protein (FADD)
triggering a cascade of proteases, starting with caspase-8. FADD recently has also
been shown to be upregulated by UV irradiation (36). Caspase-8 will cleave the
executer caspase-3 which drives the apoptotic events as described before.
3. Cell stress induced by UVB-irradiation
Reducing DNA damage and blocking death receptor signaling does not block
apoptosis completely, indicating that at least a third independent signaling pathway is
involved (30). UVB is known to be a potent inducer of reactive oxygen species (ROS)
in the cytoplasm. ROS have been shown to initiate cellular damage and apoptosis (3740). ROS can have multiple effects on a cell, as it can damage DNA by inducing strand
breaks, alter bases like, 8-hydroxyguanine, modify transcriptional binding sites on the
DNA (41) and cause lipid peroxidation. The cytotoxic potential of ROS lies in the
latter effect. Lipid peroxidation changes the structure of plasma membranes (42) and
damages the inner mitochondrial membrane leading to cytochrome c release,
subsequently resulting in apoptosis (40). Mitochondria are not only a target for ROS
but may also function as a source for ROS. In the late phase of UV-induced apoptosis,
mitochondrial dysfunction leads to enrichment of ROS in the cytoplasm (43). Radical
scavengers are able to partially inhibit UVB-induced apoptosis, although direct DNA
damage induction and CD95 signaling are not affected by these scavengers (30).
Therefore, ROS contributes independently to UVB-mediated cell death.
20
ROLE OF UV-INDUCED APOPTOSIS IN PATHOGENESIS AND TREATMENT OF DISEASE
UV induced apoptosis in the skin is known to be involved in the pathogenesis of
multiple diseases. Defects in apoptosis may lead to hyperproliferation or even
development of malignant cells. However, apoptotic cells themselves may also be
involved in generating pathology. Next, we will discuss the three major diseases
associated with UV induced apoptosis.
Cancer induction
Skin cancer is a very common form of cancer among white Caucasians. Chronic
repeated exposures to sunlight are epidemiologically shown to be the main cause of
skin cancers (44). Skin malignancies are broadly divided in melanoma and nonmelanoma skin cancers. Non-melanoma skin cancers (NMSC) occur in sun-exposed
sites and include basal cell and squamous cell carcinomas, and are derived from
keratinocytes. Melanomas are derived from melanocytes and arise from naevi in both
sun exposed as well as non-exposed skin.
UVB in sunlight acts as a skin carcinogenic inducer as well as a promoter,
through the induction of DNA damage. Unrepaired photoproducts can lead to the
incorporation of base mismatches. Mutations occur most commonly when a
photoproduct contains a cytosine. Among CPDs, thymine-cytosine (T-C) and cytosinecytosine (C-C) are shown to be most mutagenic, while thymine-thymine dimers (T-T)
are poorly mutagenic. The <6-4> photoproducts are very efficiently repaired within 3
hours after formation and are therefore less mutagenic (45). As mentioned earlier, UV
radiation can generate reactive oxygen species (ROS) which can cause oxidative
damage to the DNA bases, forming 8-hydroxy-guanine which will result in G to T
transversions (46). Deletions and chromosomal aberrations can also be induced by
UVB irradiation in mammalian cells (47;48). A gene commonly mutated after chronic
UV exposure is the tumor suppressor p53 (49;50). As discussed above, this protein
plays a central role in induction of apoptosis via direct DNA damage and ROS
formation. The mutations can result in loss of p53 function leading to defective repair
of UV induced DNA damage (51) and a decreased apoptotic response (52). Mutations
that result in decreased apoptosis are associated with clonal expansion of these cells
leading to tumor formation. Additionally, UV exposure causes transient immune
suppression which may also facilitate mutant cells to grow without elimination
(53;54).
Psoriasis
Psoriasis is a common skin disorder characterized by hyperplasia and
incomplete differentiation of epidermal keratinocytes (55). Aberrant expression of
many regulatory molecules involved in proliferation might play a role in the
pathogenesis of psoriasis. Bcl-XL has been shown to be over-expressed in all layers of
the psoriatic epidermis (56). Furthermore, in vitro studies demonstrated that human
keratinocytes derived from psoriatic plaques are resistant to induced apoptosis
compared to keratinocytes derived from normal skin (57). However, several groups
reported that most keratinocytes in psoriatic epidermis had double strand breaks in
their DNA, which might reflect an apoptotic process (56;58). Kwashima et al recently
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Figure 2: Schematic representation of the role of UVB-induced apoptosis in pathogenesis and
treatment of disease. A) Cancer: UVB induces damage to DNA which can lead to the formation of
mutations in the tumor suppressor gene, p53. Loss of p53 function can result in a decrease of
apoptotic signals leading to the formation of a neoplastic keratinocyte. B) Psoriasis: In the
pathogenesis of psoriasis keratinocytes are hyperproliferative and/or defective in terminal
differentiation. Increased Bcl-XL expression is involved in inhibition of apoptosis in psoriasitic skin.
Furthermore, infiltrating T cells producing pro-inflammatory cytokines (IL-2, TNFα, IFNγ) are also
involved in pathogenesis of psoriasis. Phototherapy, used for psoriatic lesions, induces apoptosis in
keratinocytes and T cells directly, and by upregulation of FasL on keratinocytes. IL-10 produced
after irradiation also induces immunosuppression. Combined these effects will resolve psoriatic
lesions. C) SLE: Apoptotic keratinocytes induced by UVB express nuclear and cytosolic antigens
(with or without posttranslational alteration induced by apoptosis) on their cell membrane. Autoantibodies present in SLE recognize these antigens. Inflammatory cells like residential Langerhans
cells (LC), or macrophages (Mф) recruited in the initial phase after irradiation, respond in a proinflammatory manner due to interaction of Fcγ-receptors and autoantibodies. This might lead to
further recruitment of more inflammatory cells from the blood to the site where apoptotic cells
reside and thus inflammatory lesions develop.
22
showed that the increase in numbers of DNA double strand breaks in psoriatic lesions
was not the result of apoptotic processes but caused by active DNA replication (59).
Together these reports show that defects in apoptosis might play an important role in
the pathogenesis of psoriasis. Psoriasis is also accompanied by infiltration of activated
T lymphocytes in the dermis and epidermis, which in turn produce multiple
inflammatory cytokines, like IFNγ, IL-2, and TNFα. Whether the defects in apoptosis
and differentiation are secondary to this influx is still unclear. UVB phototherapy is
one of the most common treatments for extensive psoriasis and can induce long-lived
clinical remissions. The main therapeutic effect of UVB irradiation seems to result
from of depleting T cells in lesions from epidermis and dermis. Apoptosis seems to be
the primary mechanism involved in this depletion (60). Although UVB can induce
apoptosis in T lymphocytes and keratinocytes, at the doses used for treatment mainly T
lymphocytes are targeted (61). Apoptosis of T cells can be induced directly by UVB or
indirectly via upregulation of FasL in keratinocytes (29). Besides induction of
apoptosis in immune competent cells, UVB can also modulate immune responses,
especially by release of mediators from keratinocytes, IL-10 in particular (62). IL-10
is normally decreased in psoriatic skin and upon systemic administration of IL-10 antipsoriatic effects are seen (63). Beneficial effects of UVB phototherapy can therefore
be the result of both elimination of T-cells, keratinocytes by apoptosis and immune
modulation by IL-10.
Systemic Lupus Erythematosus
Systemic Lupus Erythematosus (SLE) is a systemic autoimmune disease
characterized by the presence of autoantibodies to nuclear and cytoplasmic antigens in
conjunction with a wide range of clinical manifestations. Photosensitivity is one of the
characteristics of SLE affecting 10-50% of patients (64). Most, but not all, cutaneous
lupus lesions occur in light-exposed areas and can be triggered by sunlight exposure.
Sunlight exposure even can induce systemic disease activity. The processes inducing
cutaneous and systemic inflammatory lesions have not been elucidated, but in recent
years apoptotic cells have been suggested to be one of the major factors involved.
Accumulation of apoptotic cells due to an increased rate of apoptosis, decreased
elimination of apoptotic cells or a combination of both has been hypothesized to be an
important factor in the development of inflammatory lesions as seen in the skin
(65;66). During the apoptotic process intracellular constituents are excessively
presented to the immune system due to cell-surface expression of intracellular
constituents and/or posttranslational alteration of cellular proteins during apoptosis
(67-70). In SLE, autoantibodies are present which are specific for these antigens and
binding to the apoptotic cell may change its clearance. In the skin of SLE patients
deposition of IgG is often found at the dermal-epidermal junction, the so called lupusband, or in the epidermal layer (71). Increased numbers of apoptotic cells have been
detected in LE-skin lesions (72). Keratinocytes from SLE patients have been shown to
be more vulnerable to UV light in vitro (73). Additionally, Baima et al demonstrated
an increase in Fas expression in keratinocytes in LE lesions, while Bcl-2 expression
was decreased (72). We have recently investigated the induction of apoptosis in vivo in
SLE skin after UVB irradiation. Although SLE patients were more sensitive for UVB
23
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Apoptosis in the skin
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similar numbers of apoptotic cells were induced in SLE patients and controls.
However, the infiltrate seen after exposure was more extensive in 25% of the patients,
and even persisted after ten days when hardly any infiltrate was seen in the other
patients and controls. After three days, in some patients, inflammatory lesions could be
detected in the vicinity of apoptotic cells. We are currently characterizing the nature of
this infiltrate to elucidate the mechanisms involved. However, these data indicate a
more pro-inflammatory reaction after irradiation with UVB which is associated with
apoptotic cells. We speculate this might be caused by the opsonization of apoptotic
keratinocytes by autoantibodies and subsequently altered clearance by macrophages
and/or other phagocytes.
CONCLUDING REMARKS
Due to daily exposure to sunlight, apoptosis in the skin is an every day event. In
this way we are protected from cancer cell formation. However, when apoptotic
mechanisms fail due to defects or mutations in key apoptotic proteins, pathological
processes take over, like in skin cancer and psoriasis. Apoptotic cells themselves can
also be the source for pathology to develop. This might be the case in SLE, where
binding of auto-antibodies to apoptotic cells may lead to a altered recognition and
clearance of these cells, leading to inflammation. Understanding these processes can
result in improvement of current therapies.
24
Apoptosis in the skin
Reference List
2. Maruoka,Y., Harada,H., Mitsuyasu,T., Seta,Y., Kurokawa,H., Kajiyama,M., and
Toyoshima,K. 1997. Keratinocytes become terminally differentiated in a process involving
programmed cell death. Biochem.Biophys.Res.Commun. 238:886-890.
3. McCall,C.A. and Cohen,J.J. 1991. Programmed cell death in terminally differentiating
keratinocytes: role of endogenous endonuclease. J.Invest Dermatol. 97:111-114.
4. Allombert-Blaise,C., Tamiji,S., Mortier,L., Fauvel,H., Tual,M., Delaporte,E., Piette,F.,
DeLassale,E.M., Formstecher,P., Marchetti,P. et al. 2003. Terminal differentiation of human
epidermal keratinocytes involves mitochondria- and caspase-dependent cell death pathway.
Cell Death.Differ. 10:850-852.
5. Reefman.E, H.Kuiper, M.Jonkman, C.G.M.Kallenberg, and M.Bijl. 2004. Irradiation of
Systemic lupus erythematosus skin: sensitivity, apoptosis induction and inflammation. 4th
international congress on autoimmunity (Abstr.)
6. Gandarillas,A., Goldsmith,L.A., Gschmeissner,S., Leigh,I.M., and Watt,F.M. 1999. Evidence
that apoptosis and terminal differentiation of epidermal keratinocytes are distinct processes.
Exp.Dermatol. 8:71-79.
7. Daniels,F., Jr., Brophy,D., and Lobitz,W.C., Jr. 1961. Histochemical responses of human skin
following ultraviolet irradiation. J.Invest Dermatol. 37:351-357.
8. Murphy,G., Young,A.R., Wulf,H.C., Kulms,D., and Schwarz,T. 2001. The molecular
determinants of sunburn cell formation. Exp.Dermatol. 10:155-160.
9. Woodcock,A. and Magnus,I.A. 1976. The sunburn cell in mouse skin: preliminary quantitative
studies on its production. Br.J.Dermatol. 95:459-468.
10. Proskuryakov,S.Y., Konoplyannikov,A.G., and Gabai,V.L. 2003. Necrosis: a specific form of
programmed cell death? Exp.Cell Res. 283:1-16.
11. Weiske,J., Schoneberg,T., Schroder,W., Hatzfeld,M., Tauber,R., and Huber,O. 2001. The fate
of desmosomal proteins in apoptotic cells. J.Biol.Chem. 276:41175-41181.
12. Cooper,K.D., Duraiswamy,N., Hammerberg,C., Allen,E., Kimbrough-Green,C., Dillon,W.,
and Thomas,D. 1993. Neutrophils, differentiated macrophages, and monocyte/macrophage
antigen presenting cells infiltrate murine epidermis after UV injury. J.Invest Dermatol.
101:155-163.
13. Reis e Sousa, Stahl,P.D., and Austyn,J.M. 1993. Phagocytosis of antigens by Langerhans cells
in vitro. J.Exp.Med. 178:509-519.
14. Sharlow,E.R., Paine,C.S., Babiarz,L., Eisinger,M., Shapiro,S., and Seiberg,M. 2000. The
protease-activated receptor-2 upregulates keratinocyte phagocytosis. J.Cell Sci. 113 ( Pt
17):3093-3101.
15. Kumakiri,M., Hashimoto,K., and Willis,I. 1977. Biologic changes due to long-wave
ultraviolet irradiation on human skin: ultrastructural study. J.Invest Dermatol. 69:392-400.
25
CHAPTER 2
1. Green,H. 1977. Terminal differentiation of cultured human epidermal cells. Cell 11:405-416.
CHAPTER 2
16. Young,A.R. 1987. The sunburn cell. Photodermatol. 4:127-134.
17. de Laat,W.L., Jaspers,N.G., and Hoeijmakers,J.H. 1999. Molecular mechanism of nucleotide
excision repair. Genes Dev. 13:768-785.
CHAPTER 2
18. Petit-Frere,C., Capulas,E., Lowe,J.E., Koulu,L., Marttila,R.J., Jaspers,N.G., Clingen,P.H.,
Green,M.H., and Arlett,C.F. 2000. Ultraviolet-B-induced apoptosis and cytokine release in
xeroderma pigmentosum keratinocytes. J.Invest Dermatol. 115:687-693.
19. Stege,H., Roza,L., Vink,A.A., Grewe,M., Ruzicka,T., Grether-Beck,S., and Krutmann,J. 2000.
Enzyme plus light therapy to repair DNA damage in ultraviolet-B-irradiated human skin.
Proc.Natl.Acad.Sci.U.S.A 97:1790-1795.
20. Kastan,M.B., Onyekwere,O., Sidransky,D., Vogelstein,B., and Craig,R.W. 1991. Participation
of p53 protein in the cellular response to DNA damage. Cancer Res. 51:6304-6311.
21. Sitailo,L.A., Tibudan,S.S., and Denning,M.F. 2002. Activation of caspase-9 is required for
UV-induced apoptosis of human keratinocytes. J.Biol.Chem. 277:19346-19352.
22. Hockenbery,D., Nunez,G., Milliman,C., Schreiber,R.D., and Korsmeyer,S.J. 1990. Bcl-2 is an
inner mitochondrial membrane protein that blocks programmed cell death. Nature 348:334336.
23. Deveraux,Q.L., Takahashi,R., Salvesen,G.S., and Reed,J.C. 1997. X-linked IAP is a direct
inhibitor of cell-death proteases. Nature 388:300-304.
24. Washio,F., Ueda,M., Ito,A., and Ichihashi,M. 1999. Higher susceptibility to apoptosis
following ultraviolet B irradiation of xeroderma pigmentosum fibroblasts is accompanied by
upregulation of p53 and downregulation of bcl-2. Br.J.Dermatol. 140:1031-1037.
25. Pena,J.C., Fuchs,E., and Thompson,C.B. 1997. Bcl-x expression influences keratinocyte cell
survival but not terminal differentiation. Cell Growth Differ. 8:619-629.
26. Bennett,M., Macdonald,K., Chan,S.W., Luzio,J.P., Simari,R., and Weissberg,P. 1998. Cell
surface trafficking of Fas: a rapid mechanism of p53-mediated apoptosis. Science 282:290293.
27. Kibitel,J., Hejmadi,V., Alas,L., O'Connor,A., Sutherland,B.M., and Yarosh,D. 1998. UVDNA damage in mouse and human cells induces the expression of tumor necrosis factor alpha.
Photochem.Photobiol. 67:541-546.
28. Qin,J.Z., Bacon,P., Panella,J., Sitailo,L.A., Denning,M.F., and Nickoloff,B.J. 2004. Low-dose
UV-radiation sensitizes keratinocytes to TRAIL-induced apoptosis. J.Cell Physiol 200:155166.
29. Leverkus,M., Yaar,M., and Gilchrest,B.A. 1997. Fas/Fas ligand interaction contributes to UVinduced apoptosis in human keratinocytes. Exp.Cell Res. 232:255-262.
30. Kulms,D., Zeise,E., Poppelmann,B., and Schwarz,T. 2002. DNA damage, death receptor
activation and reactive oxygen species contribute to ultraviolet radiation-induced apoptosis in
an essential and independent way. Oncogene 21:5844-5851.
26
Apoptosis in the skin
32. Filipowicz,E., Adegboyega,P., Sanchez,R.L., and Gatalica,Z. 2002. Expression of CD95 (Fas)
in sun-exposed human skin and cutaneous carcinomas. Cancer 94:814-819.
33. Bang,B., Rygaard,J., Baadsgaard,O., and Skov,L. 2002. Increased expression of Fas on human
epidermal cells after in vivo exposure to single-dose ultraviolet (UV) B or long-wave UVA
radiation. Br.J.Dermatol. 147:1199-1206.
34. Ouhtit,A., Gorny,A., Muller,H.K., Hill,L.L., Owen-Schaub,L., and Ananthaswamy,H.N. 2000.
Loss of Fas-ligand expression in mouse keratinocytes during UV carcinogenesis. Am.J.Pathol.
157:1975-1981.
35. Aragane,Y., Kulms,D., Metze,D., Wilkes,G., Poppelmann,B., Luger,T.A., and Schwarz,T.
1998. Ultraviolet light induces apoptosis via direct activation of CD95 (Fas/APO-1)
independently of its ligand CD95L. J.Cell Biol. 140:171-182.
36. Kim,P.K., Weller,R., Hua,Y., and Billiar,T.R. 2003. Ultraviolet irradiation increases FADD
protein in apoptotic human keratinocytes. Biochem.Biophys.Res.Commun. 302:290-295.
37. Masaki,H., Atsumi,T., and Sakurai,H. 1995. Detection of hydrogen peroxide and hydroxyl
radicals in murine skin fibroblasts under UVB irradiation. Biochem.Biophys.Res.Commun.
206:474-479.
38. Peus,D., Vasa,R.A., Beyerle,A., Meves,A., Krautmacher,C., and Pittelkow,M.R. 1999. UVB
activates ERK1/2 and p38 signaling pathways via reactive oxygen species in cultured
keratinocytes. J.Invest Dermatol. 112:751-756.
39. Tyrrell,R.M. 1995. Ultraviolet radiation and free radical damage to skin. Biochem.Soc.Symp.
61:47-53.
40. Farber,J.L. 1994. Mechanisms of cell injury by activated oxygen species. Environ.Health
Perspect. 102 Suppl 10:17-24.
41. Floyd,R.A. 1990. Role of oxygen free radicals in carcinogenesis and brain ischemia. FASEB J.
4:2587-2597.
42. de Kok,T.M., ten Vaarwerk,F., Zwingman,I., van Maanen,J.M., and Kleinjans,J.C. 1994.
Peroxidation of linoleic, arachidonic and oleic acid in relation to the induction of oxidative
DNA damage and cytogenetic effects. Carcinogenesis 15:1399-1404.
43. Cadenas,E. and Davies,K.J. 2000. Mitochondrial free radical generation, oxidative stress, and
aging. Free Radic.Biol.Med. 29:222-230.
44. Woodhead,A.D., Setlow,R.B., and Tanaka,M. 1999. Environmental factors in nonmelanoma
and melanoma skin cancer. J.Epidemiol. 9:S102-S114.
45. Brash,D.E. and Haseltine,W.A. 1982. UV-induced mutation hotspots occur at DNA damage
hotspots. Nature 298:189-192.
46. Shibutani,S., Takeshita,M., and Grollman,A.P. 1991. Insertion of specific bases during DNA
synthesis past the oxidation-damaged base 8-oxodG. Nature 349:431-434.
27
CHAPTER 2
31. Oishi,M., Maeda,K., and Sugiyama,S. 1994. Distribution of apoptosis-mediating Fas antigen
in human skin and effects of anti-Fas monoclonal antibody on human epidermal keratinocyte
and squamous cell carcinoma cell lines. Arch.Dermatol.Res. 286:396-407.
CHAPTER 2
47. Emri,G., Wenczl,E., Van Erp,P., Jans,J., Roza,L., Horkay,I., and Schothorst,A.A. 2000. Low
doses of UVB or UVA induce chromosomal aberrations in cultured human skin cells. J.Invest
Dermatol. 115:435-440.
CHAPTER 2
48. Horiguchi,M., Masumura,K.I., Ikehata,H., Ono,T., Kanke,Y., and Nohmi,T. 2001. Molecular
nature of ultraviolet B light-induced deletions in the murine epidermis. Cancer Res. 61:39133918.
49. Brash,D.E., Rudolph,J.A., Simon,J.A., Lin,A., McKenna,G.J., Baden,H.P., Halperin,A.J., and
Ponten,J. 1991. A role for sunlight in skin cancer: UV-induced p53 mutations in squamous
cell carcinoma. Proc.Natl.Acad.Sci.U.S.A 88:10124-10128.
50. Ziegler,A., Leffell,D.J., Kunala,S., Sharma,H.W., Gailani,M., Simon,J.A., Halperin,A.J.,
Baden,H.P., Shapiro,P.E., Bale,A.E. et al. 1993. Mutation hotspots due to sunlight in the p53
gene of nonmelanoma skin cancers. Proc.Natl.Acad.Sci.U.S.A 90:4216-4220.
51. Smith,M.L., Chen,I.T., Zhan,Q., O'Connor,P.M., and Fornace,A.J., Jr. 1995. Involvement of
the p53 tumor suppressor in repair of u.v.-type DNA damage. Oncogene 10:1053-1059.
52. Hill,L.L., Ouhtit,A., Loughlin,S.M., Kripke,M.L., Ananthaswamy,H.N., and OwenSchaub,L.B. 1999. Fas ligand: a sensor for DNA damage critical in skin cancer etiology.
Science 285:898-900.
53. Guzman,E., Langowski,J.L., and Owen-Schaub,L. 2003. Mad dogs, Englishmen and
apoptosis: the role of cell death in UV-induced skin cancer. Apoptosis. 8:315-325.
54. Fisher,M.S. and Kripke,M.L. 1977. Systemic alteration induced in mice by ultraviolet light
irradiation and its relationship to ultraviolet carcinogenesis. Proc.Natl.Acad.Sci.U.S.A
74:1688-1692.
55. McKay,I.A. and Leigh,I.M. 1995. Altered keratinocyte growth and differentiation in psoriasis.
Clin.Dermatol. 13:105-114.
56. Wrone-Smith,T., Johnson,T., Nelson,B., Boise,L.H., Thompson,C.B., Nunez,G., and
Nickoloff,B.J. 1995. Discordant expression of Bcl-x and Bcl-2 by keratinocytes in vitro and
psoriatic keratinocytes in vivo. Am.J.Pathol. 146:1079-1088.
57. Wrone-Smith,T., Mitra,R.S., Thompson,C.B., Jasty,R., Castle,V.P., and Nickoloff,B.J. 1997.
Keratinocytes derived from psoriatic plaques are resistant to apoptosis compared with normal
skin. Am.J.Pathol. 151:1321-1329.
58. Laporte,M., Galand,P., Fokan,D., de Graef,C., and Heenen,M. 2000. Apoptosis in established
and healing psoriasis. Dermatology 200:314-316.
59. Kawashima,K., Doi,H., Ito,Y., Shibata,M.A., Yoshinaka,R., and Otsuki,Y. 2004. Evaluation
of cell death and proliferation in psoriatic epidermis. J.Dermatol.Sci. 35:207-214.
60. Ozawa,M., Ferenczi,K., Kikuchi,T., Cardinale,I., Austin,L.M., Coven,T.R., Burack,L.H., and
Krueger,J.G. 1999. 312-nanometer ultraviolet B light (narrow-band UVB) induces apoptosis
of T cells within psoriatic lesions. J.Exp.Med. 189:711-718.
28
Apoptosis in the skin
Krueger,J.G., Wolfe,J.T., Nabeya,R.T., Vallat,V.P., Gilleaudeau,P., Heftler,N.S., Austin,L.M.,
and Gottlieb,A.B. 1995. Successful ultraviolet B treatment of psoriasis is accompanied by a
reversal of keratinocyte pathology and by selective depletion of intraepidermal T cells.
J.Exp.Med. 182:2057-2068.
62. Nishigori,C., Yarosh,D.B., Ullrich,S.E., Vink,A.A., Bucana,C.D., Roza,L., and Kripke,M.L.
1996. Evidence that DNA damage triggers interleukin 10 cytokine production in UVirradiated murine keratinocytes. Proc.Natl.Acad.Sci.U.S.A 93:10354-10359.
63. Asadullah,K., Sterry,W., Stephanek,K., Jasulaitis,D., Leupold,M., Audring,H., Volk,H.D., and
Docke,W.D. 1998. IL-10 is a key cytokine in psoriasis. Proof of principle by IL-10 therapy: a
new therapeutic approach. J.Clin.Invest 101:783-794.
64. Provost,T.T. and Flynn,J.A. 2001. Cutaneous Medicine, Chapter 5:Lupus Erythematosus.
Elsevier Science, London, UK. 41-81.
65. Bijl,M., Limburg,P.C., and Kallenberg,C.G. 2001. New insights into the pathogenesis of
systemic lupus erythematosus (SLE): the role of apoptosis. Neth.J.Med. 59:66-75.
66. Reefman,E., Dijstelbloem,H.M., Limburg,P.C., Kallenberg,C.G., and Bijl,M. 2003. Fcgamma
receptors in the initiation and progression of systemic lupus erythematosus. Immunol.Cell
Biol. 81:382-389.
67. Casciola-Rosen,L.A., Anhalt,G., and Rosen,A. 1994. Autoantigens targeted in systemic lupus
erythematosus are clustered in two populations of surface structures on apoptotic
keratinocytes. J.Exp.Med. 179:1317-1330.
68. Saegusa,J., Kawano,S., Koshiba,M., Hayashi,N., Kosaka,H., Funasaka,Y., and Kumagai,S.
2002. Oxidative stress mediates cell surface expression of SS-A/Ro antigen on keratinocytes.
Free Radic.Biol.Med. 32:1006-1016.
69. Utz,P.J., Hottelet,M., Schur,P.H., and Anderson,P. 1997. Proteins phosphorylated during
stress-induced apoptosis are common targets for autoantibody production in patients with
systemic lupus erythematosus. J.Exp.Med. 185:843-854.
70. Utz,P.J. and Anderson,P. 1998. Posttranslational protein modifications, apoptosis, and the
bypass of tolerance to autoantigens. Arthritis Rheum. 41:1152-1160.
71. Halberg,P., Ullman,S., and Jorgensen,F. 1982. The lupus band test as a measure of disease
activity in systemic lupus erythematosus. Arch.Dermatol. 118:572-576.
72. Baima,B. and Sticherling,M. 2001. Apoptosis in different cutaneous manifestations of lupus
erythematosus. Br.J.Dermatol. 144:958-966.
73. Furukawa,F., Itoh,T., Wakita,H., Yagi,H., Tokura,Y., Norris,D.A., and Takigawa,M. 1999.
Keratinocytes from patients with lupus erythematosus show enhanced cytotoxicity to
ultraviolet radiation and to antibody-mediated cytotoxicity. Clin.Exp.Immunol. 118:164-170.
29
CHAPTER 2
61.