Subphysiological concentrations of extracellular calcium sensitize normal human keratinocytes to UVB-induced apoptosis

Abstract Normal human keratinocytes are stimulated to proliferate in serum-free medium containing subphysiological concentrations of calcium (0.09 mM, low calcium). In this study, we examined the effect of increased levels of extracellular calcium (2.0 mM, normal calcium) on UVB-induced apoptosis. Apoptosis was assessed by changes in cellular morphology, annexind V-FITC flow cytometry, and the formation of internucleosomal DNA ladders. High doses of UVB induced keratinocytes grown in low calcium medium to undergo apoptosis. In contrast, keratinocytes grown for 72 h in normal calcium medium were completely resistant to UVB-induced apoptosis. No apoptosis was observed even at UVB doses as high as 1200 J/m². However, despite the lack of UVB-induced cell death, keratinocytes grown in normal calcium medium lost the ability to proliferate following high levels of UVB irradiation. High doses of UVB also increased the expression of the differentiation-specific proteins involucrin and cytokeratin 10 in a dose-dependent manner. In addition, growth in normal calcium medium lowered the UVB-induced stimulation of the p53 protein and altered the normal subcellular localization pattern of p53. UVB irradiation of human keratinocytes grown in normal calcium medium may be inducing further cell differentiation in the absence of overt cell death. Received: 16 April 1998 / Received after revision: 31 August 1998 / Accepted: 23 September 1998     

Calcipotriol induces apoptosis in psoriatic keratinocytes

In recent years, vitamin D3 analogues have become one of the most widely prescribed topical treatments for mild or moderate chronic plaque psoriasis. These molecules are effective and safe, but their exact mechanism of action is not completely understood. In vitro studies have shown that D3 analogues decrease proliferation and induce differentiation of keratinocytes, and have strong immunomodulating effects, but there are no conclusive data about apoptosis. The aim of this study was to evaluate differences in apoptotic response between lesional and perilesional keratinocytes of patients with psoriasis before and after treatment with calcipotriol, a synthetic vitamin D3 analogue. Keratinocytes were isolated from psoriatic plaques including lesional and perilesional skin, and cultured. Cells were treated with calcipotriol for 20 h and examined under confocal microscopy after staining with propidium iodide. The number of apoptotic cells after incubation with calcipotriol was significantly higher in lesional than in perilesional keratinocytes (P < 0.05) or non‐treated psoriatic keratinocytes (P < 0.05). In conclusion, calcipotriol seems to induce apoptosis in psoriatic keratinocytes.     

TWEAK/Fn14 activation induces keratinocyte proliferation under psoriatic inflammation

Tumor necrosis factor (TNF)‐like weak inducer of apoptosis (TWEAK) has been reported to induce keratinocyte apoptosis in vitro by engaging its sole receptor of fibroblast growth factor‐inducible 14 (Fn14). In this study, we explored the role of TWEAK/Fn14 pathway in the growth of psoriatic keratinocytes that is, however, characterized by suppressed apoptotic cell death. Skin tissues from the patients with psoriasis or healthy donors were determined for TWEAK and Fn14 expression, and primary keratinocytes were evaluated under the stimulation of psoriatic proinflammatory cytokines or plus TWEAK. The results showed that both TWEAK and Fn14 were highly expressed in psoriatic skins. Moreover, the stimulation of psoriatic cytokines enhanced Fn14 expression by keratinocytes in vitro, which expressed TNF receptor 2 predominantly and proliferated increasingly with the addition of TWEAK. Furthermore, TWEAK stimulation enhanced the synthesis of survivin, inhibitor of apoptosis protein 2 and cellular FLICE‐inhibitory protein in lesional keratinocytes. Therefore, TWEAK/Fn14 interaction prefers to enhance proliferation but not apoptosis of keratinocytes under psoriatic inflammation. The activation of nuclear factor‐κB signalling‐dependent anti‐apoptotic proteins and biased expression of TNF receptors may be responsible for such a novel principle in keratinocytes under psoriatic inflammation.     

Infliximab restores the balance between pro- and anti-apoptotic proteins in regressing psoriatic lesions

Background Psoriasis and psoriatic arthritis are treated very efficaciously with infliximab, a chimaeric human–murine antitumour necrosis factor (TNF)‐α antibody. As we reported earlier, infliximab, besides its anti‐inflammatory properties, induces a caspase‐independent programmed cell death of psoriatic keratinocytes. Objectives To elucidate this finding further, we investigated the epidermal expression of proteins involved in the mitochondria‐dependent (intrinsic) pathway of cell death. Methods Quantification of proteins with pro‐ (p53, AIF, Bax) and anti‐apoptotic functions (Bcl‐2, Bcl‐XL) and of NF‐κB was performed by means of immunohistochemistry and digital image analysis of the staining of nonlesional skin and lesional psoriatic skin from patients treated with infliximab at weeks 0, 2 and 6. Results Serial biopsies from psoriatic plaques of samples taken at days 0, 5, 14 and 21 of therapy demonstrated a significant downregulation of anti‐apoptotic proteins Bcl‐2, Bcl‐XL and NF‐κB during treatment and, in parallel, a significant upregulation of pro‐apoptotic proteins p53, Bax and AIF. These differences in expression correlated with decreases in epidermal thickness and clinical outcome (Psoriasis Area and Severity Index). At day 21, expression levels of apoptosis‐related proteins in lesional skin approximated those found in nonlesional skin. Conclusions Our data therefore suggest that TNF‐targeting agents may induce the regression of psoriasis at least in part by normalizing the expression of apoptosis‐related proteins in lesional keratinocytes.     

Apoptosis, P53 and Bcl-2 expression in response to topical calcipotriol therapy for psoriasis

BACKGROUND: The histopathologic changes characteristic of psoriasis might be related to suppressed apoptosis. The P53 and Bcl-2 proteins play a central role in the regulation of apoptosis. This study aimed to evaluate P53 and Bcl-2 expression and apoptotic cells in the psoriatic skin before and after topical calcipotriol therapy. METHODS: Skin biopsies were obtained from nonlesional and lesional skin of 10 patients with generalized plaque psoriasis before and after treatment with topical calcipotriol ointment. The P53 and Bcl-2 expression was evaluated using immunoperoxidase technique and apoptotic cells by the terminal deoxynucleotide transferase (TdT) mediated deoxyuridine triphosphate nick end labeling (TUNEL) method. RESULTS: After topical calcipotriol therapy, keratinocytes of psoriatic skin showed significant decrease of P53 (P = 0.002) and increase of Bcl-2 (P = 0.01) expression. On the other hand lymphocytes showed significant decrease of Bcl-2 (P = 0.01). There were no apoptotic cells before treatment but after calcipotriol therapy, apoptosis was more detectable in keratinocytes than in lymphocytes. CONCLUSIONS: The results of the study suggested that one of the actions of calcipotriol in psoriasis might be exerted through induction of apoptosis, especially of keratinocytes, through a P53-independent pathway. Meanwhile, suppression of Bcl-2 expression in lymphocytes may promote apoptosis of dermal lymphocytes leading to healing of psoriasis.     

Selenium protects primary human keratinocytes from apoptosis induced by exposure to ultraviolet radiation

The generation of reactive oxygen species has been implicated in ultraviolet radiation (UVR)-induced skin damage. In mice, increasing dietary selenium intake protects skin from UVR-induced DNA damage and photocarcinogenesis. We sought to determine whether selenium supplementation could protect keratinocytes from apoptosis resulting from exposure to broadband (TL20W/12) UVR. Unirradiated cultures contained 6.5 +/- 1% apoptotic cells; the maximum percentage of apoptotic cells (34 +/- 5%) was seen 16 h after UVR of 600 J/m(2). Under these conditions cell death from necrosis was 15 +/- 2.5% of the total cells. A 24-h preincubation with sodium selenite (10 nm(-1) microm) or selenomethionine (50 nm(-1) microm) protected cultured human keratinocytes from UVR-induced apoptosis. In primary keratinocytes the greatest reduction in apoptosis was found with 100 nm of either selenium compound (71% reduction in the numbers of total apoptotic cells; P < 0.01). Supplementation with 100-200 nm selenite or selenomethionine prevented UVR-induced apoptosis, but did not decrease the levels of UVR-induced p53, as measured by Western blotting. Collectively, this data suggests that selenium prevents UVR-induced cell death by inhibiting p53-independent cell death pathways.     

T‐lymphocyte‐induced,fas‐mediated apoptosis is associated with early keratinocyte differentiation

Please cite this paper as: T‐lymphocyte‐induced, fas‐mediated apoptosis is associated with early keratinocyte differentiation. Experimental Dermatology 2009. Abstract: The development of eczematous lesions is thought to be due in part to a breakdown in skin barrier function as a result of T lymphocytes (T cells) invading the skin causing epidermal keratinocyte apoptosis. In this study, we investigated the interaction of T cells and keratinocytes on apoptosis and terminal differentiation using an in vitro co‐culture system. Experiments were performed using the HaCaT keratinocyte cell line or normal human epidermal keratinocytes. Activated human peripheral blood‐derived T cells were found to induce Fas‐dependent keratinocyte apoptosis by up to sixfold. Increased Fas was associated with increased IFN‐γ. The T‐cell apoptotic signal was found to target preferentially keratinocytes in the very early stages of terminal differentiation, such as those with low levels of α6‐integrin expression, and result in subsequent increased caspase 3 activity. This observation was accompanied by a marked increase in keratinocyte ICAM‐1 expression and its ligand LFA‐1 on T cells. Our data suggest that T cells may initiate the onset of keratinocyte terminal differentiation making them more susceptible to Fas‐dependent cell death signals delivered by the T cells.     

Low-dose ultraviolet B radiation synergizes with TNF-alpha to induce apoptosis of keratinocytes

High-dose ultraviolet B (UVB) irradiation is known to induce apoptosis of keratinocytes, but low-dose UVB dose not. In this paper we present evidence that low-dose UVB can induce TNF-alpha-dependent apoptosis of keratinocytes. In our study, 5 mJ/cm(2) doses of UVB were not sufficient by themselves to induce apoptosis of cultured human keratinocytes, but 20 mJ/cm(2) doses of UVB were. The combination of 5 mJ/cm(2) doses of UVB and exogenous TNF-alpha (15 ng/ml) induced significant apoptosis of keratinocytes, although exogenous TNF-alpha without UVB did not. This phenomenon was accompanied by enhanced clustering of tumor necrosis factor receptor 1 (TNFR1). TNF-alpha's promotion of the induction of apoptosis by low-dose UVB was seen until 30 min after irradiation but not at 1 h. We confirmed this finding using a skin organ culture system. UVB (20 mJ/cm(2)), which did not induce transformation of epidermal keratinocytes into sunburn cells, induced apoptosis when TNF-alpha was added to the culture medium. These results suggest that one of the possible mechanisms of inducing keratinocyte apoptosis by low-dose UVB and TNF-alpha is that low-dose UVB augments ligand-binding-induced TNFR1 clustering, resulting in increased apoptotic cell death.     

Induction of apoptosis in human HaCaT keratinocytes

Although cell death by apoptosis has been recognized as an important control mechanism in the maintenance of tissue homeostasis and in the elimination of cells with damaged DNA, information on the induction and characteristics of apoptosis in keratinocytes is rather scarce. Apoptotic mechanisms may play an important role in normal and disturbed homeostasis of the skin. In the present study, we therefore investigated the effects of several potential inducers of apoptosis in the human keratinocyte cell line HaCaT. Apoptosis was assessed with respect to morphological changes by light and electron microscopic examinations and to DNA integrity by a specific ELISA. UVB irradiation induced the morphology and internucleosomal DNA fragmentation characteristic of apoptosis in a dose- and time-dependent manner. Interferon-γ caused DNA cleavage at the linker regions without producing morphological features consistent with apoptotic cell death. In contrast, treatment with dithranol and NP-40 resulted in necrotic alterations in the keratinocytes. Treatment with the calcium ionophore A23187 caused morphological changes which were similar to the characteristics of ‘nonapoptotic programmed cell death’. Dexamethasone, tumor necrosis factor-α, transforming growth factor-β, TPA, retinoic acid, the podophyllin derivative etoposide, the thromboxane A ₂ analogue U46619, cycloheximide, and the nitric oxide donors sodium nitroprusside and S-nitroso-glutathione, which are all known to induce apoptosis in other cell types, did not affect HaCaT keratinocytes. These results demonstrate that apoptosis can be induced in keratinocytes in vitro but the apoptosis differs from that in other cell types, such as haematopoietic cells, with regard to the type of inducer and/or the sensitivity of the target cells. Since keratinocytes are affected by numerous external and internal stimuli, they might posses several protective mechanisms to prevent apoptosis and to ensure the structural integrity of the outermost barrier of the body. Received: 28 November 1995     

银屑病患者角质形成细胞凋亡以及维A酸与榄香烯的影响

目的 探讨银屑病患者角质形成细胞凋亡以及维A酸与榄香烯对体外培养的角质形成细胞凋亡的影响.方法 用TUNEL法测定28例银屑病患者角质形成细胞凋亡;流式细胞仪和DNA片段化观察维A酸与榄香烯对体外培养的角质形成细胞凋亡的影响.结果 28例银屑病患者中19例有大量凋亡细胞分布于表皮各层,另9例较少,分布于棘细胞层和颗粒层.浓度为1、5、10、20μg/mL的维A酸和浓度为20、40、60、80μg/mL的榄香烯可促进人角质形成细胞株Colo16细胞凋亡.结论 银屑病患者角质形成细胞凋亡异常增多,一定浓度的维A酸与榄香烯可在实验室内促进人角质形成细胞株Colo-16细胞凋亡.     

Photoprotective effect of calcipotriol upon skin photoreaction to UVA and UVB

It has been shown that 1,25-dihydroxyvitamin D₃ has a photoprotective effect against UVB injury in mouse skin and cultured rat keratinocytes by induction of metallothionein (MT). Calcipotriol is a synthetic analogue of 1,25-dihydroxyvitamin D₃ with equipotent cell regulating properties, but with a lower risk of calcium-related side effects. The aim of the present study was to see whether calcipotriol has a photoprotective property both in vitro and in vivo. We examined the effect of calcipotriol on UV-induced damage of cultured human keratinocytes through a cell viability assay, and measurement of DNA synthesis by cultured keratinocytes, on UV-induced damage of mouse skin and on minimal erythema dose (MED). We found that calcipotriol was protective against UVB-induced reduction in DNA synthetic activity of cultured keratinocytes in relatively low doses (20 and 40 mJ/cm²) of UVB. With phototesting following application of calcipotriol, five subjects among 10 healthy volunteers and three among six psoriasis patients showed an increase in MED compared with the vehicle-treated site. These findings imply that calcipotriol may be photoprotective and that more extensive studies with various doses of UV irradiation and modes of calcipotriol delivery are required.     

Aberrant accumulation of p53 associates with Ki67 and mitotic count in benign skin lesions

Sixty-two skin samples from patients with a variety of benign disorders (20 cases of psoriasis, 14 cases of chronic dermatitis, 11 seborrhoeic keratoses, 11 cases of lichen planus), and seven normal skin samples, were stained immunohistochemically with a polyclonal antibody (CM-1) to p53, and a monoclonal antibody to Ki67, using the avidin-biotin complex method, p53-positive keratinocytes could be found in most of these lesions. The percentage of p53-positive cells was, however, far lower than usually seen in p53-positive malignant tumours. No p53 reactivity was observed in the normal skin samples. Variable Ki67 reactivity was observed in all skin samples. Overall, the number of Ki67-positive cells was higher in skin samples in which the proportion of p53-positive cells was high (>0.5% of total epidermal cell population) (P=0.004). This also applied separately to psoriatic and non-psoriatic lesions (P=0.028 and P=0.033, respectively). In cases with >10% of Ki67-positive cells, there were significantly more mitoses (P<0.001). This association applied to both psoriasis and the other lesions studied (P=0.024 and P <0.001, respectively). The results show that immunohistochemically detectable accumulation of p53 is a frequent finding in non-neoplastic skin lesions. As p53 positivity was associated with the proliferation marker Ki67, the accumulation of p53 is possibly a response to an increased proliferation rate of the keratinocytes in these skin diseases, or alternatively it may be associated with apoptosis.     

Programmed cell death of keratinocytes in infliximab-treated plaque-type psoriasis

BACKGROUND: Tumour necrosis factor (TNF)-alpha blockade using infliximab, a chimeric anti-TNF-alpha antibody, is an effective treatment for plaque-type psoriasis, inducing remission in about 80% of patients. OBJECTIVES: To examine infliximab-induced programmed cell death (PCD) of keratinocytes in psoriatic plaques on serial skin biopsy samples. METHODS: Five patients with moderate to severe plaque-type psoriasis received infliximab infusions intravenously (5 mg kg(-1)) at weeks 0, 2 and 6. Biopsies of nonlesional and lesional skin (days 0, 5, 14 and 21) were obtained. Conventional microscopy was used to examine the morphology of the psoriatic keratinocytes. In situ detection of apoptosis was performed by electron microscopy and by immunohistochemical staining with anti-p53 and anti-caspase-3 antibodies. Results Infusion of infliximab induced a clinical response in all five patients with psoriasis, with a mean Psoriasis Area and Severity Index improvement of 24.8% already at day 5. This was accompanied by significant histopathological changes in the skin biopsy samples after infliximab treatment. Light and electron microscopic evaluation revealed apoptosis-like morphological changes in lesional keratinocytes, i.e. nuclear condensation, chromatin fragmentation and cytoplasmic vesiculation, visible already after the first infusion. These damaged keratinocytes stained positively for p53, but not for active caspase-3. CONCLUSIONS: The effects of infliximab in psoriasis extend beyond merely anti-inflammatory actions, and may include caspase-independent PCD of lesional keratinocytes. The PCD of keratinocytes may be an important mechanism that could explain at least in part the rapid and sustained therapeutic effect of infliximab in psoriasis.     

The anti‐apoptotic protein G1P3 is overexpressed in psoriasis and regulated by the non‐coding RNA,PRINS

Please cite this paper as: The anti‐apoptotic protein G1P3 is overexpressed in psoriasis and regulated by the non‐coding RNA, PRINS. Experimental Dermatology 2010; 19: 269–278. Abstract: Psoriasis Susceptibility‐Related RNA Gene Induced by Stress (PRINS) is a non‐coding RNA overexpressed in lesional and non‐lesional psoriatic epidermis and induced by stress. Its function in healthy and psoriatic skin is still not known. Here, we report that PRINS regulates G1P3, a gene with anti‐apoptotic effects in keratinocytes. siRNA‐mediated inhibition of PRINS gene resulted in altered cell morphology and gene expression alterations, as demonstrated in a microarray experiment. One of the genes regulated by PRINS ncRNA was G1P3, an interferon‐inducible gene with anti‐apoptotic effects in cancer cells. Interestingly, we found that G1P3 was 400‐fold upregulated in hyperproliferative lesional and ninefold upregulated in non‐lesional psoriatic epidermis compared to healthy epidermis. In vitro, G1P3 protein levels were highest in proliferating keratinocytes and siRNA‐mediated downregulation of G1P3 resulted in increased cell apoptosis. These data indicate that G1P3 inhibits spontaneous keratinocyte apoptosis and hence its high expression in psoriatic skin may contribute to the development of psoriatic lesions. We hypothesize that the deregulation of the PRINS ncRNA may contribute to psoriasis and results in decreased sensitivity to spontaneous keratinocyte apoptosis via the regulation of G1P3.     

Apoptosis Is Induced by Anti-Fas Antibody Alone in Cultured Human Keratinocytes

Fas is a well-known cell surface receptor whose main function is the induction of apoptosis in many cell types including human keratinocytes. Several reports indicate that anti-Fas antibody can induce apoptosis in cultured keratinocytes after interferon gamma (IFNγ) pretreatment. Because IFNγ is synthesized by activated T cells, but not by keratinocytes, these results suggest that Fas may only be effective in apoptosis occurring in T-cell mediated inflammatory skin diseases. We hypothesized that Fas alone might mediate apoptosis in normal human keratinocytes without any other help and thus play a role in normal epidermal homeostasis. By using Cell Death Detection ELISA, we observed keratinocyte apoptosis 24 hours after anti-Fas antibody stimulation not only in IFNγ-pretreated conditions but also in non-pretreated conditions. Even though the percentage of cultured keratinocytes stained by anti-Fas antibody increased from 7.8 to 25.8% 24 hours after IFNγ stimulation, the apoptotic rate of the anti-Fas only group was the same as that of the anti-Fas plus IFNγ treated group. In both conditions, we have verified apoptotic phenomena in cultured keratinocytes in situ by TUNEL staining. Some apoptotic bodies were phagocytosed by neighboring keratinocytes. Fas-mediated apoptosis was not inhibited by the protein synthesis inhibitor cycloheximide and was enhanced by inhibitors of several protein kinases, including PKC and staurosporine. These results suggest that Fas-mediated apoptosis may play a role in both T cell-mediated skin diseases and normal epidermal homeostasis.     

Neutrophil elastase promotes proliferation of HaCaT cell line and transwell psoriasis organ culture model

Background Neutrophil elastase (NE) plays an important role in psoriasis. In this study we observed the effect of NE on the proliferation of HaCaT cells and transwell psoriasis organ culture model and investigated the mechanism. Methods HaCaT cells were treated with various concentrations NE (0, 0.1, 1, 10, 100 IU/l). In addition, the cells were co‐stimulated with 10 IU/l NE and 1 g/l sivelestat. Then, HaCaT cells proliferation and DNA synthesis were determined using methyl thiazolyl tetrazolium (MTT) and tritiated thymidine (3H‐TdR) assay respectively. Cell cycle distribution was measured using fluorescence activated cell sorting (FACS). Subsequently, we established cultured transwell psoriasis organ model in vitro. Then, the cultured transwell psoriasis organ model was treated with 10 IU/l NE. Immunohistochemistry was employed to detect the expression levels of Ki67 and p53 in the cultured transwell psoriasis organ model. Results MTT and 3H‐TdR incorporation assay suggested NE could remarkably promote the proliferation and DNA synthesis of HaCaT cell in a dose‐dependent manner. After NE treatment (10 IU/l) for 24 h, the cell fraction of HaCaT cell in G2 + S phase was increased significantly, whereas the cell fraction in G1 phase was reduced remarkably. Immunohistochemistry results revealed enhanced expression of both Ki67 and p53 genes in cultured transwell psoriasis organ model after NE treatment. Conclusions NE significantly promotes the proliferation of HaCaT cell. Meanwhile, it also up‐regulates the expression levels of Ki67 and p53 in psoriasis lesion tissue, which plays an important role in the pathogenesis of psoriasis lesion.     

Higher susceptibility to apoptosis following ultraviolet B irradiation of xeroderma pigmentosum fibroblasts is accompanied by upregulation of p53 and downregulation of Bcl-2

Apoptosis plays an important part as a defence mechanism in eliminating damaged cells. Among the complex factors which regulate apoptosis, the p53 tumour suppressor protein which is induced by DNA damage has been suggested to play a crucial part. Cells from xeroderma pigmentosum (XP) patients, which are defective in nucleotide excision repair, express higher levels of p53 and are highly susceptible to cell death after ultraviolet (UV) irradiation. To examine the relationships between DNA damage, p53 and apoptosis, normal and XP group A fibroblasts were exposed to UVB, and expressions of molecules involved in apoptosis were examined. Apoptosis of XP and normal cells was clearly detected at 48 h after irradiation with UVB at doses of 5 and 40 mJ/cm2, respectively. Cells were positive by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labelling (TUNEL) staining under these exposure conditions. At 6 h after irradiation, p53 protein expression was induced in normal and XP cells at minimal doses of 10 and 2.5 mJ/cm2, respectively. Bcl-2 protein, an inhibitor of apoptosis, was downregulated prior to cell death following UVB exposure at doses that induced apoptosis in both cell types. These results suggest that DNA damage due to UVB induces apoptosis by upregulating proapoptotic molecules such as p53, and by downregulating anti-apoptotic molecules such as Bcl-2.     

卡泊三醇对银屑病皮损中角质形成细胞凋亡的影响

从细胞凋亡角度探讨卡泊三醇(CPT)治疗银屑病(PS)的药理机制。采用末端标记(TUNEL)技术 ,分别检测了30例CPT治疗前后的PS患者和10名正常人皮肤标本的角质形成细胞凋亡。经CPT治疗后的PS皮损中的角质形成细胞凋亡数较治疗前明显增加(P<0.05)。CPT治疗PS的疗效可能与其增加角质形成细胞的凋亡有关。     

A low UVB dose, with the potential to trigger a protective p53-dependent gene program, increases the resilience of keratinocytes against future UVB insults

One protein central in the response of human keratinocytes to ultraviolet B damage is p53. By transactivating genes involved in either cell cycle arrest or DNA repair, p53 has a leading role in the recovery from this damage. Considering this role, we wished to investigate whether the triggering of a p53-dependent gene program by repetitive ultraviolet B (UVB) exposure can induce an adaptive response in human skin cells. In particular, we examined two p53-target genes, p21/WAF1 and p53R2, with a crucial role in p53-induced cell cycle arrest and p53-induced DNA repair respectively. Exposure to a mild UVB dose was able to induce an adaptive response in human keratinocytes, leading to increased survival of cells that maintain their capacity to repair DNA damage upon exposure to apoptotic doses of UVB. Our study indicates that this adaptation response is only achieved if the interval between subsequent UVB insults allows sufficient time for the p53-induced protective gene program to be induced. Our results also demonstrate that small but quickly recurring UVB exposures are as harmful as one intense, continual exposure to UVB irradiation. Future research will be oriented toward investigating alternative ways to induce an adaptive response without pre-exposing the cells to UV.     

Keratinocyte apoptosis in epidermal remodeling and clearance of psoriasis induced by UV radiation

Psoriasis is a common chronic skin disorder, but the mechanisms involved in the resolution and clearance of plaques remain poorly defined. We investigated the mechanism of action of UVB, which is highly effective in clearing psoriasis and inducing remission, and tested the hypothesis that apoptosis is a key mechanism. To distinguish bystander effects, equal erythemal doses of two UVB wavelengths were compared following in vivo irradiation of psoriatic plaques; one is clinically effective (311 nm) and one has no therapeutic effect on psoriasis (290 nm). Only 311 nm UVB induced significant apoptosis in lesional epidermis, and most apoptotic cells were keratinocytes. To determine clinical relevance, we created a computational model of psoriatic epidermis. Modeling predicted apoptosis would occur in both stem and transit-amplifying cells to account for plaque clearance; this was confirmed and quantified experimentally. The median rate of keratinocyte apoptosis from onset to cell death was 20 minutes. These data were fed back into the model and demonstrated that the observed level of keratinocyte apoptosis was sufficient to explain UVB-induced plaque resolution. Our human studies combined with a systems biology approach demonstrate that keratinocyte apoptosis is a key mechanism in psoriatic plaques clearance, providing the basis for future molecular investigation and therapeutic development.