Research Fields | Bőrklinika

Short description of our research

PSORIASIS

The role of D type cyclins in keratinocyte cell cycle regulation and in psoriasis (Dr. Nóra Belső, Attila Bebes, Dr. Márta Széll, Dr. Zsuzsanna Bata-Csörgő)

The cell cycle regulatory D type cyclins have several isoforms. We know it from our previous investigations that high level expression of α5 integrin preceeds the entry of keratinocytes into the cell cycle. Therefore we hypothetized that α5 integrin contributes to keratinocyte cell cycle regulation. The aim of our work was to study the role of different D type cyclins in the G0-G1/S and G1/S transition of keratinocytes, in psoriasis and to reveal whether D type cyclins regulate the expression of α5 integrin.
We have demonstrated that D type cyclins have differential roles in keratinocyte cell cycle regulation and are upregulated in psoriasis, a multifactorial skin diseases characterized by hyperproliferating keratinocytes. Moreover, we showed that blocking the function of α5 integrin with a neutralizing antibody downregulated the expression of D1 cyclin in HaCaT keratinocytes.
In order to further examine D type cyclins in keratinocyte cell cycle regulation we silenced D1, D2 and D3 cyclins individually and together in HaCaT keratinocytes. The lack of one D type cyclin function had no effect on cell cycle regulation and cell morphology, but double or triple D type cyclin silencing resulted in G2/M arrest of the cells and the formation multinucleated HaCaT cell. To elucidate the gene expression changes due to D type cyclin double or triple silencing, a real-time RT-PC-based array method was applied. The results of this experimets showed a significant decrease in Ki67 mRNA expression, indicating a regulatory connection between D type cyclins and Ki67 in these cells. Moreover, silencing of Ki67 resulted in the appearance of multinucleated aberrant cells and a relevant decrease in the mRNA expressions of all D type cyclins. These data indicate that beside their well known function during the G0-G1/S transition, D type cyclins also regulate mitosis. This regulatory function of D type cyclins during mitosis may take place via influencing Ki67 expression. Although widely used as a proliferation marker, not much is known about the function of Ki67. The formation of multinucleated cells in Ki67 silenced HaCaT keratinocytes indicates that Ki67 also has a cell cycle regulatory function in mitosis. Our data suggest the existence of a regulatory cycle between D type cyclins and Ki67 which is key in mitosis regulation in keratinocytes.

The role of KGF/KGFR system in keratinocyte cell cycle regulation and in psoriasis. (Dr. Nóra Belső, Bernadett Kormos, Vas Krisztina, Dr. Márta Széll, Dr. Zsuzsanna Bata-Csörgő)

Beside immune dysregulation, data suggest that abnormal skin homeostasis maybe crucial in psoriasis development; in fact it may drive the immune response. In the past few years we focused on investigating the uninvolved psoriatic skin in order to uncover differences between the psoriatic and the healthy skin.
We applied tape stripping on non-lesional skin of psoriatic patients and on normal skin of healthy subjects and looked at changes in the epidermis and the dermis. We demonstrated that both keratinocyte growth factor (KGF) and its receptor, KGFR is expressed differentially in healthy and psoriatic uninvolved skin, even before injury occurs. KGF was expressed in the dermis and all layers of the epidermis in both normal and psoriatic uninvolved skin; staining intensity was much stronger in psoriatic tissue compared to normal. Six hours after tape stripping KGF mRNA increased 10-fold in normal dermis and only 5-fold in psoriatic. Twenty-four and 48 hours after tape stripping KGF expression slightly increased in healthy skin and no change was noted in the psoriatic uninvolved tissue. In healthy skin KGFR was expressed only in the basal, immediate suprabasal keratinocytes, but in the psoriatic uninvolved skin KGFR expression was detected in all layers of keratinocytes. Injury enhanced the intensity of expression, but did not change the localization of KGFR expression either in normal or in psoriatic epidermis. These data provide additional evidence to altered skin homeostasis in psoriasis.

Investigations on the role of professional immune cells in the pathogenesis of psoriasis (Dr. Ferenc Kovács-Sólyom, Képíró László, Dr. Rolland Gyulai)

According to our previous results, regulatory T cells are functionally defective in psoriasis, however, the reasons are unknown. In this study we investigated the possible role of interleukin-1 receptors in psoriasis pathogenesis, with emphasis on regulatory and effector T cells.
Effector (CD4+CD25-, Teff) and regulatory (CD4+CD25+CD127-, Treg) T cells were isolated from normal and psoriatic blood. Expression of type 1 (signal-transmitting) and type 2 (decoy) interleukin-1 receptors, and IL-1R antagonist (IL-1RN) mRNAs were determined by real-time RT-PCR. Cell surface, intracellular and soluble protein expression was investigated by flow cytometry and ELISA. Upon CD3/CD28 activation, IL-1R1 mRNA expression was increased in normal Teff and Treg cells. Interestingly, while CD3/CD28 activation did not influence IL-1R1 expression in psoriatic Treg cells, activation induced stronger IL-1R1 mRNA expression in psoriatic Teff cells compared to normals.
Expression of decoy receptor (IL-1R2 and soluble IL-1R2) mRNA was elevated in activated normal Teff and Treg cells. Psoriatic Teff and Treg cells, however, more rapidly and more intensively increased their IL-1R2 and sIL-1R2 expression. Expression of IL-1 receptor antagonist (IL-1RN) was moderately increased upon activation in both normal effector and regulatory T cells. However, expression was markedly downregulated in psoriatic Treg cells, and highly activated in psoriatic effector cells. IL-1R2 protein expression was constant in normal cells, but significantly increased in activated psoriatic effector cells compared to regulatory T cells. Our data suggest that differential expression of genes in the interleukin-1 pathway in normal and psoriatic T lymphocytes may underline some functional differences in these cells, and may have a role in psoriasis pathogenesis.
In another set of experiments we study the role of the TL1A molecule in the pathogenesis of psoriasis. The TNF superfamily member TL1A has an immunmodulatory effect on activated T-cells as a co-stimulator. It is known that TL1A plays an important role in the pathogenesis of autoimmune inflammatory disorders, such as Crohn’s disease and rheumatoid arthritis. Three isoforms of TL1A have has been described, however their specific functions are still unknown. We set out to determine the role of TL1A isoforms and DR3 receptor in the pathogenesis of psoriasis. We are going to detect of TL1A and DR3 in healthy, psoriatic lesional and non-lesional skin using immunohistochemical methods. mRNA expression of TL1A isoforms will be detected by real-time RT-PCR using specific primers. We aim to investigate the role of keratinocytes in the modulation of T-cells through the TL1A-DR3 pathway in psoriasis. This work is performed in co-operation with Dr. Vilmos Tubak, Institute of Biochemistry, Biological Research Center of the Hungarian Academy of Sciences.

Investigations on PRINS, a non-coding RNA contributing to psoriasis susceptibility (Dr. Márta Széll, Sarolta Bacsa, Dr. Zsuzsanna Bata-Csörgő)

We have recently identified a non-coding RNA, PRINS that plays a role in psoriasis susceptibility and in cellular stress response. In our ongoing investigations we are working on the functional characterization of this non-coding RNA molecule.
We have found that PRINS regulates G1P3, a gene with antiapoptotic 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 antiapoptotic effects in cancer cells. Interestingly, we found that G1P3 was 400-fold upregulated in hyperproliferative lesional and 9-fold 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.
To reveal the cellular function(s) of PRINS we searched for direct interacting partner(s) of this stress-induced ncRNA molecule, using a special ribonucleoprotein (RNP) purification kit which had been developed for RNA-target-search purposes. As a template, we used the 39mer sequence element of the PRINS in vitro RNA transcript that had effectively knocked down the expression of PRINS in our previous silencing experiments. In HaCaT cell lysates we identified nucleophosmin protein as a potential candidate for physical interaction with PRINS RNA. Nucleophosmin is an ubiquitously expressed nucleolar phosphoprotein which shuttles continuously between the nucleus and the cytoplasm. In immunohistochemical experiments we could detect that the expression of nucleophosmin is significantly elevated in the dividing cells of the basal layer of psoriatic involved skin samples compared to healthy, and psoriatic uninvolved samples. Our experimental data suggests that the PRINS ncRNA may provide a structural skeleton of an RNP complex playing a role in stress-induced cellular processes and that the abnormal functioning of this complex may contribute to the pathogenesis of psoriasis.
In the near future we plan to optimize the in situ detection of PRINS non-coding RNA and study its expression in various malignant, benign and inflammatory skin diseases. This project will be performed in co-operation with Eniko Sonkoly and Andor Pivarcsi in the Karolinska Institute, Stockholm, Sweden.

Identification and characterization of regulatory networks contributing to psoriasis susceptibility (Dr. Kornélia Szabó, Dr. Zsuzsanna Bata-Csörgő, Dr. Attila Dobozy, Dr. Márta Széll)

The non-lesional skin of psoriatic patients possess some inherent characteristics that are already present in the otherwise healthy-looking skin, and make them prone to develop the typical psoriatic symptoms in response to various stimuli.
Our major aim was to identify and characterize genes and proteins that are differentially expressed in uninvolved psoriatic and normal epidermis, and to discover regulatory networks that are responsible for these differences. To this end, a cDNA microarray experiment was performed where we compared the gene expression profile of 4 healthy and 4 psoriatic uninvolved epidermis samples upon T cell lymphokine induction in organotypic cultures. The cDNA microarray experiment identified 57 annotated genes with known functions, and another 11 expressed transcripts with unknown functions that were differentially regulated in psoriatic uninvolved and healthy epidermis following T cell lymphokine induction. 11 of the annotated genes have already been implicated in the pathogenesis of psoriasis. Pathway analysis using various software packages and public databases suggests that many of the identified genes play an important role in cellular processes; regulation of cell morphology, development and cell death, and also in the metabolism of small molecules and lipids. Our results can help to uncover basic mechanisms of this complex disease and lead to novel therapeutic approaches to prevent disease development.

Investigations on anti-α6 integrin antibodies in different subsets of patients with psoriasis vulgaris (Dr. Mária Kiss, Dr. Zsuzsanna Bata-Csörgő)

Integrins are cell surface receptors that are involved in cell-matrix adhesion and signaling. α6 integrin, a laminin receptor, contains 1050 amino acids present as a heavy (110 kDa) and a light chain (30 kDa) linked by a disulfide bond. The a6b4 integrin heterodimer, together with other members of the epidermal integrin family, is expressed by epidermal keratinocytes. This expression is restricted to the basal and suprabasal proliferative cell layers, both in the epidermis and in cultures of keratinocytes. α6β4 integrin is localized at the basal pole of the basal cells and coordinates a structural linkage between intracellular filaments and the extracellular matrix of the basement membrane. The a6b4 integrin plays a crucial role in the assembly of hemidesmosomes, and antibodies directed against α6β4 integrin induce a dermal-epidermal dissociation in vitro. It was earlier demonstrated that a6 integrin is a BP180 binding partner in the hemidesmosomal multimolecular complex and that this connection is essential for the functioning of hemidesmosomes. More recent evidence suggested a pathophysiological role for autoantibodies against α6 integrin in the subepidermal blister formation of oral pemphigoid, a subset of mucous membrane pemphigoid.
We hypothesize that abnormal laminin integrity described in psoriatic uninvolved skin may result in insufficient binding of α6β4 to its ligand and autoantibodies could be produced. The autoantobody production in psoriasis may then further enhance the structural abnormality of the dermo-epidermal junction, resulting in microwounds and a wound healing phenotype. With the use of PeptideStructure and PlotStructure software, four different antigenic epitopes for α6 integrin were predicted and their fusion recombinant constructs were prepared in an E. coli expression system. We are screening psoriatic patient’s sera for a6 integrin autoantibodies with the fusion recombinant proteins in an ELISA system.

The role of nerve growth factor (NGF) and cutaneous neuropeptides in the maintenance of skin homeostasis and in the pathogenesis of inflammatory skin disorders (Dr. Mária Kiss, Bernadett Kormos, Dr. Sándor Husz)

Nerve growth factor (NGF) is a very important and potent growth factor in the skin. Normal human keratinocytes synthesize and release high amounts of biologically active NGF. Additionally, human keratinocytes express both the high-affinity (TrkA) and the low-affinity (p75NTR) NGF receptors. NGF plays an important role in the maintenance of skin homeostasis and cell proliferation and its expression can be enhanced in pathological conditions, such as itching and psoriasis vulgaris. NGF production of keratinocytes is modulated by cutaneous neuropeptides. It is well documented that, in neuronal cells, the NGF pathway (neurotrophin pathway) plays a central role in the maintenance of the balance between neuronal cell proliferation, differentiation and apoptosis. NGF and its precursor proNGF are very important in this process. ProNGF elicits opposite biological effects to those of mature NGF; while mature NGF promotes cell survival and cell proliferation, proNGF can cause cell death. The aim of our investigations is to characterize the components of NGF/proNGF pathway in normal human keratinocytes. Additionally, we investigate the effects of cutaneous neuropeptides (substance P, calcitonin gene-related peptide, vasoactiv intestinal polipeptide, and galanin) on the function of NGF/proNGF pathway in human skin.


MELANOCYTE BIOLOGY AND MALIGNANT MELANOMA

Investigations on the proliferation and pigmentation of normal human adult epidermal melanocytes cultured in a novel chemical mitogen-free Mel-mix medium (Bernadett Kormos, Dr. Márta Széll, Dr. Gábor Szabad, Dr. Zsuzsanna Bata-Csörgő)

The standard way of in vitro culturing for normal human adult epidermal melanocytes includes the use of chemical mitogens, such as the tumor promoter TPA and the cAMP enhancer cholera toxin. This culture environment is not suited for in vitro experiments and in vivo therapy in many instances. Previously we have established a new culture system (Mel-mix) that avoids chemical mitogens and in which melanocytes are capable of a long-term and rapid proliferation.
Melanocytes cultured in Mel-mix medium lose their pigment content and decrease c-Kit and tyrosinase related protein-1 (TRP-1) differentiation marker expressions. The dendritic phenotype of melanocytes becomes bipolar in Mel-mixcultures. These data indicate that melanocytes dedifferentiate in this chemical-free culture. Comparison of the proliferation and senescence of melanocytes cultured in Mel-mix and in chemical mitogen containing media shows that the proliferation rate is higher in Mel-mix cultured cells, while the rate of senescence is similar in both cultures. A neural precursor marker, nestin is expressed by melanocytes cultured in Mel-mix medium, which provides further evidence for dedifferentiation. Currently we examine the effect of UVB on differentiated and dedifferentiated melanocytes and the possibilities using dedifferentiated melanocytes in therapy.

Identification and characterization of genetic and environmental predisposing factors for melanoma (Dr. Márta Széll, Dr. Zsanett Csoma, Dr. Klára Balogh, Hilda Polyánka, Dr. Judit Oláh)

Epidemiologic data suggest an increasing malignant melanoma incidence worldwide therefore identifying melanoma risk factors is of high interest of the society. Our primary aim is to identify predisposing factors for melanoma susceptibility.
In our previous studies we identified a novel CDKN2A intronic mutation (IVS1+37 G/C) in a Hungarian melanoma-prone family. To decide whether this intronic mutation has any effect on mRNA splicing and contributes to melanoma susceptibility, aberrant splice variant(s) of CDKN2A mRNA in the tissue samples of the effected family members will be detected and the exact role of the mutation in the regulation of splicing will be studied with the use of a de novo constructed CDKN2A minigene. This study is performed in an international collaboration with the laboratory of Professor Franco Pagani, ICGEB Institute, Trieste, Italy.
A study for gene-environmental interaction on the prevalence of melanoma predisposing pigmented skin lesions is also undertaken. This project is part of a twin study that aims to survey the effect of neonatal blue light treatment on the numbers of melanocytic nevi. Mutations of the melanoma-predisposing CDKN2A gene as well as polymorphisms of MCR1 and HAL genes will be studied. The latter two genes play regulatory roles in human pigmentation and UV-induced immunosuppression, respectively. The results of this survey will provide us valuable data on gene-environment interactions toward susceptibility to cutaneous malignant melanoma, the skin cancer with the worst prognostic factors.



ACNE VULGARIS

Studying the role of genetic and molecular factors in the pathogenesis of acne vulgaris (Dr. Kornélia Szabó, Dr. Gábor Tax, Andrea Tanácsné Bajkán)

Our major goal is to identify and characterize genetic and molecular factors that play a role in the pathogenesis of the most common dermatologic disease, acne vulgaris.
Around puberty adolescent individuals suffer from dramatic hormonal changes that lead to increased sebum secretion. As a result the otherwise commensal Propionibacterium acnes (P. acnes) bacterium exhibits extensive growth properties, and populate the skin follicles. The bacterium and its secreted enzymes can set off innate immune events in the surrounding skin cells, leading to enhanced gene expression of several downstream target genes: among them cytokines, chemokines and genes encoding proteins with antimicrobial properties. We are investigating the signal transduction mechanisms that are responsible for these expression changes in an in vitro keratinocyte culture system, and try to identify and analyze various genes that are playing important roles in these processes.
It is also known that there are great variations in the severity of individual acne symptoms and genetic predisposing and/or protective factors are playing an important role in these variations. At present we are working on the identification and detailed description of such factors. Currently we are studying known polymorphisms of genes that are proved to play important roles in acne pathogenesis (TNFA, and IL1A pro-inflammatory cytokine gene polymorphisms), and analyze them in retrospective case-control genetic studies. We are trying to answer the question of how these factors influence the regulation of genes, how do they affect the structure and/or function of the resulting proteins, and how all these changes lead to pathogenic processes and thus the initiation of acne vulgaris.
Experimental evidence suggests that altered gene expression of the TNFA gene caused by the P. acnes bacterium is a central event in the molecular pathogenesis of acne lesion formation. We are also investigating the exact course and kinetics of TNFα mRNA expression in in vitro keratinocyte culture system using molecular methods.
Our results can help the understanding of the molecular pathogenesis of a very common skin disease, acne vulgaris.


GENODERMATOSES

Identification and functional characterization of mutations in genodermatoses (Dr. Nikoletta Nagy, Dr. Márta Széll)

The technical development of the genetic investigative methods has significantly accelerated the discovery of the genetic background of several monogenic skin diseases. A recent new initiative at our Department is the investigation of the genetic background of monogenic skin diseases. There are already ongoing genetic investigations on the following diseases: neurofibromatosis, Brooke-Spiegler syndrome, Carney-complex, Darier disease and epidermoliysis bullosa, and our aim is to further widen the scale of the investigated diseases.
We have recently identified a novel CYLD mutation in a Hungarian pedigree affected by Brooke-Spiegler syndrome and spanning 3 generations. To reveal the pathogenic role of the newly identified mutation, functional studies were performed on CD4+ T lymphocytes of two patients since CYLD is highest expressed in this cell type. CD4+ cells of BSS patients were stimulated by TNFα and the activation of the NF-κB pathway was monitored by the expression of NEMO transcription factor. Our data demonstrated a significant reduction in the protein level of NEMO in the samples of the BSS patients compared to healthy controls. Our findings provide the possibility to carry out prenatal genetic screening and they might also contribute to the development of new gene therapy methods.


VENOUS LEG ULCER

Identification and characterization of genetic and molecular factors for leg ulcer development (Dr. Nikoletta Nagy, Dr. Gábor Szabad, Dr. Győző Szolnoky, Dr. Márta Széll)

Chronic non-healing venous leg ulcer is considered a multifactorial disorder. Several genetic, environmental and life style factors are suspected to play a role in the pathomechanism of the disease but the exact pathogenesis of venous leg ulcer is still not known. We are investigating the role of several genetic and molecular factors (fibroblast growth factor receptor 2, FGFR2; tumor necrosis factor A, TNFA; syndecane-4, SDC4; neurophilin-1, NRP1) in the pathomechanism of the disease.
Our results demonstrated that the expression of the FGFR2-IIIb isoform is related with the proliferation states of the keratinocytes and not with their differentiation state. The fact that FGFR2-IIIb exhibits a lower expression level in venous leg ulcer patients suggests a receptorial dysfunction which, through impairment of the proliferation phase of re-epithelization, leads to pathologic wound healing in these patients. A possible underlying mechanism for the abnormal expression of FGFR2-IIIb seen in venous leg ulcer patients may be the presence of the 2451 A/G 3’UTR SNP, which could alter the stability of the mRNA and result in decreased amounts of FGFR2 proteins and a receptor dysfunction. In this case, both the re-epithelization through FGFR2-IIIb and the angiogenesis through FGFR2-IIIc may be impaired.
A potential genetic factor contributing to venous leg ulcer development could be the A allele of the -308 TNFA SNP, though our data suggest that its association with leg ulceration is secondary and the primary association probably exists with obesity.
We alsodemonstrated decreased expression of SDC4 in the uninvolved dermis of venous leg ulcer patients with three independent methods (real time RT-PCR, Western blot and immunohistochemistry) and hypothetized that this misfunction influences dermal wound repair proccesses and thus predisposes to venous leg ulcer development.
In case of NRP1 we could not demonstrate any association with venous leg ulcer development neither with polymorphism studies nor with gene expression investigations.

Keratinocyte and fibroblast cultures for autologous transplantation (Dr. Anna Kenderessy-Szabó, Dr. Gábor Szabad, Dr. Zsuzsanna Bata-Csörgő)

Transplantation with autologous epidermal cells is an up-to-date therapeutic modality for the treatment of skin diseases characterized with epidermal lesions. For that we worked out and routinely use a culture system with proliferation inducing autologous factors that is free of any animal derived growth factors.


ALLERGIC RHINITIS

Genomic studies in allergic rhinitis (Dr. Márta Széll, Dr. Edina Garaczi, Hilda Polyánka)

The aim of this study was to determine whether forkhead box P3 (FOXP3) polymorphisms contribute to allergic rhinitis (AR) in a Central-European population, the Hungarians, similarly as it was found in Han Chinese. A case-control study was performed and the genotype distribution of the rs3761548 FOXP3 polymorphism was analyzed separately in females and in males. The results demonstrated that females homozygous for the rare FOXP3 rs3761548 allele (A/A) are protected against AR; otherwise, females who are either wild types (C/C) or heterozygote carriers (C/A) of the rare allele are more susceptible to AR (OR [95%CI] = 2.089 [1,095; 3.988]). We were able to confirm the findings of Zhang et al. in a geographically and ethnically distinct population, the Hungarians, and revealed that the rs3761548 SNP is a marker of a haplotype in these two populations. This is not done in Sub-Saharan Africans, suggesting that this haplotype was fixed after early modern humans left Africa.
We have shown previously that the mixed UVA-UVB-visible light intranasal phototherapy (rhinophototherapy) is effective in the treatment of allergic rhinitis. We aimed to investigate whether genetic factors contribute to the individual differences seen in the efficacy of rhinophototherapy. It is well known that TNFα plays a key role in the light induced immunosuppression, therefore we performed our genetic studies on the TNFA gene. Our working hypothesis was futher supported by the results of a Japanese group that demonstrated the contribution of certain TNFA polymorphisms and microsatellites in the UV-induced immunosuppression of skin. Fifty-seven allergic rhinitis patients were enrolled into the study. Patients were subdivided into two groups according to their response to rhinophototherapy (good responders vs. weak responders). We found that the -863 polymorphism of TNFA gene showed a significant association (p=0,0068) with the individual response to rhinophototherapy: those who carry the rare allele all exhibited good response to the treatment. Next we aimed to investigate the molecular background of our finding. We asked whether the rare A allele was able to contribute to the good response in rhinophototherapy. We constructed chimaeric constructs with the wild type (C/C) and rare (A/A) – 863 alleles of TNFA and luciferase riporter gene and transfected these constructs into keratinocytes. The keratinocytes were irradiated with UV-B and luciferase activity was measured. According to our results the rare allele resulted in a higher level of reporter gene activity compared to the wild type allele. Results of our in vitro experiments suggest that the rare -863A allele of TNFA confers a higher level of TNFA gene expression and contributes to an elevated level of responsiveness in rhinophototherapy.


PHOTOBIOLOGY STUDIES

The role of huCOP1 in keratinocyte UV response (Dr. Ágnes Kinyó, Attila Bebes, Dr. Márta Széll, Dr. Mária Kiss, Dr. Zsuzsanna Bata-Csörgő, Dr. Lajos Kemény)

The aim of our photobiology studies is to identify and characterize molecular factors playing role in the cellular response of keratinocytes to UVB irradiation.
UVB irradiation has been shown to trigger a broad range of changes in gene expression in human skin; however, factors governing these events are still not well understood. We have shown that human constitutive photomorphogenic protein-1 (huCOP1), an E3 ligase, contributes to the orchestration of UVB response of keratinocytes. We have demonstrated that (i) huCOP1 protein is expressed both in the nucleus and in the cytoplasm of cultured keratinocytes, (ii) UVB reduces the levels of the huCOP1 mRNA and protein, and (iii) induces changes in the subcellular localization of huCOP1. Finally, we also showed that gene-specific silencing of huCOP1 induces the accumulation of the tumor suppressor p53 protein, which is further increased after UVB irradiation. Currently we are studying the expression of two other p53 regulating proteins, Pirh2 and MDM2 in keratinocytes and their role in keratinocytes UVB response. This work is performed in co-operation with Dr. Ferenc Nagy, Institute of Plant Biology, Biological Research Center of the Hungarian Academy of Sciences.

The role of ABCG2 transporter in keratinocyte light response (Attila Bebes, Dr. Márta Széll, Dr. Zsuzsanna Bata-Csörgő, Dr. Lajos Kemény)

We also perform photobiology studies on the ABCG2 xenobiotic transporter. A systematic study of xenobiotic transporters in human keratinocytes drew our attention to the ABCG2 molecule participating in the photoprotection of keratinocytes. Xenobiotic transporters of the ABC protein superfamily play an important part in maintaining the biochemical barrier of various tissues, but the roles of these transporters in the skin are not known. We found the ABCC4 and ABCG2 transporters highly expressed in proliferating keratinocytes. Abrogation of ABCC4 and ABCG2 functions, however, did not affect the proliferation of HaCaT keratinocytes. Whereas both transporters were overexpressed in psoriatic lesional epidermis compared to normal skin, ABCC4 was localized to hyperproliferating basal keratinocytes, while ABCG2 to the differentiating suprabasal cells. The pattern of ABCG2 protein expression induced in ultraviolet light-irradiated healthy human epidermis resembled psoriatic lesions. Specific inhibition of ABCG2 with the non-toxic fumitremorgin C analog Ko-134 in ultraviolet light-irradiated keratinocytes significantly decreased cell viability in vitro, indicating that ABCG2 contributes to cellular stress response. Moreover, ABCG2 inhibition in HaCaT keratinocytes subjected to photodynamic treatment resulted in a substantial decrease in cell viability, suggesting that targeting ABCG2 can enhance the efficacy of photodynamic therapy used in various dermatological disorders. This project is performed in co-operation with SOLVO Biotechnology ZRt.