Publications by authors named "Amir M Hossini"

26 Publications

  • Page 1 of 1

Metabolic Disorders/Obesity Is a Primary Risk Factor in Hidradenitis Suppurativa: An Immunohistochemical Real-World Approach.

Dermatology 2021 Jul 22:1-9. Epub 2021 Jul 22.

Departments of Dermatology, Venereology, Allergology and Immunology, Dessau Medical Center, Brandenburg Medical School Theodor Fontane and Faculty of Health Sciences Brandenburg, Dessau, Germany.

Background: Hidradenitis suppurativa (HS) is an inflammatory, potentially scarring disease of the hair follicle, affecting the apocrine gland-bearing skin areas. The major comorbid disorders associated with the occurrence or the aggravation of the disease are obesity and smoking. Numerous efforts to dissociate these factors led to controversial results.

Objectives: To assess the importance of metabolic disorders/obesity, smoking/environmental toxins, and inflammation in HS by utilizing the differential expression of major relevant protein markers in lesional skin of obese/smoking versus non-obese/non-smoking HS patients.

Methods: Lesional skin specimens deriving from two groups of HS patients (BMI >30 and smokers, n = 12 vs. BMI <30 and non-smokers, n = 10) were stained with antibodies raised against irisin, PPARγ, and IGF-1R, which correlate with metabolic disorders/obesity, EGFR and AhR, associated with smoking, and IL-17, IL-17R, and S100A8, as markers of inflammation.

Results: Metabolic disorders/obesity-related markers exhibited marked differential expression between the two groups, while smoking-associated markers a limited one. IL-17R expression was stronger in obese/smokers, and S100A8 staining exhibited intense strong immunoreactivity in both groups without significant difference.

Conclusions: The notion that obesity plays a role in HS development appears to be supported by the prominent regulation of the associated lesional biomarkers. Tobacco smoking might contribute less to HS than previously suspected.
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http://dx.doi.org/10.1159/000517017DOI Listing
July 2021

3D-SeboSkin Model for Human ex vivo Studies of Hidradenitis Suppurativa/Acne Inversa.

Dermatology 2021 Jun 2:1-8. Epub 2021 Jun 2.

Departments of Dermatology, Venereology, Allergology, and Immunology, Dessau Medical Center, Brandenburg Medical School Theodor Fontane and Faculty of Health Sciences Brandenburg, Dessau, Germany.

Background: Hidradenitis suppurativa/acne inversa (HS) is a chronic, recurrent inflammatory skin disease. Its pivotal pathogenetic event is believed to be the occlusion of the hair follicle generating a perifollicular lympho-histiocytic inflammation. However, knowledge of the exact HS pathogenesis requires further research.

Objective: To develop a human HS model applicable in preclinical research which could help to understand the pathophysiology of HS and to determine the action of therapeutic candidates.

Methods: The 3D-SeboSkin technology was applied to maintain explants of involved and uninvolved skin of HS patients ex vivo for 3 days. Detection of differential expression of previously detected HS biomarkers was performed by immunohistochemistry in a group of female patients (n = 9, mean age 37.2 ± 8.4 years).

Results: The application of the 3D-SeboSkin model preserved the structural integrity of lesional and perilesional HS skin ex vivo, as previously described for healthy skin. Moreover, the HS 3D-SeboSkin setting maintained the differential expression and pattern of several HS biomarkers (S100A9, KRT16, SERPINB3) in epidermal and dermal tissue and the appendages.

Conclusion: We have validated HS 3D-SeboSkin as a reproducible, human model, which is appropriate for preclinical lesional and perilesional HS skin studies ex vivo.
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http://dx.doi.org/10.1159/000515955DOI Listing
June 2021

Topical application of autophagy-activating peptide improved skin barrier function and reduced acne symptoms in acne-prone skin.

J Cosmet Dermatol 2021 Mar 7;20(3):1009-1016. Epub 2020 Aug 7.

Research Division, Incospharm Corp. Daejeon, South Korea.

Background: Recent studies about the important roles of autophagy signaling in sebaceous lipogenesis and epidermal differentiation suggest potential benefits of autophagy activation in acne.

Aims: To investigate the effects of an autophagy activator on acne-prone skin.

Methods: Autophagy signaling in human immortalized SZ95 sebocytes, normal human epidermal keratinocytes, and 3D reconstituted skin was examined. Effects of an autophagy-activating peptide on sebaceous lipogenesis were measured by fluorescence microscopic analysis. The clinical efficacy in acne-prone skin was evaluated through an eight-week, double-blind, randomized, vehicle-controlled study. Changes in skin surface lipid compositions were further analyzed.

Results: In cultured sebocytes and keratinocytes, the investigated autophagy-activating peptide increased LC3-II expression, indicating a stimulation of autophagy signaling. Testosterone and linoleic acid treatment induced lipogenesis in cultured sebocytes and is further inhibited by the autophagy activator peptide treatment. Increased expression of differentiation marker proteins in cultured keratinocytes was also observed by autophagy-activating peptide. In clinical study, reduction of closed comedones and the amount of skin surface lipids as well as of trans-epidermal water loss (TEWL) were observed in acne-prone skin after autophagy-activating peptide application. In addition, reduction of squalene and increase in cholesterol were observed after an 8-week application.

Conclusions: Topical application of an autophagy activator downregulated sebaceous lipogenesis and improved the skin barrier function. Considering the important roles of sebum and skin barrier function in acne pathogenesis, autophagy activation might represent a new therapeutic option in early forms of acne.
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http://dx.doi.org/10.1111/jocd.13636DOI Listing
March 2021

Autophagy regulates lipid production and contributes to the sebosuppressive effect of retinoic acid in human SZ95 sebocytes.

J Dermatol Sci 2020 May 18;98(2):128-136. Epub 2020 Apr 18.

Department of Dermatology, Yonsei University College of Medicine, Seoul, Republic of Korea. Electronic address:

Background: Autophagy is a catabolic process for eliminating damaged organelles or proteins to maintain cellular homeostasis. Recently, lipids have been demonstrated to be targets for autophagosomal degradation. Therefore, autophagy might be involved in sebaceous gland homeostasis, however, relevant data are lacking.

Objectives: We investigated the role of autophagy in sebaceous lipogenesis and its regulatory mechanisms in human SZ95 sebocytes. We also examined the possible role of autophagy in 13-cis-retinoic acid (13-cis-RA)-mediated sebosuppression.

Methods: Autophagy markers expression was examined by immunohistochemistry in normal and acne lesional skin. SZ95 sebocytes were treated with autophagy inhibitors under starvation or treated with a combination of testosterone and linoleic acid (testosterone/LA), with or without autophagy inducer rapamycin or 13-cis-RA. Lipids were assessed by BODIPY and quantitative Nile Red staining. Autophagy-related gene 7 small interference RNA was used to confirm the role of autophagy on the sebosuppressive effect of rapamycin or 13-cis-RA.

Results: Autophagy markers were strongly expressed in the maturing sebaceous gland cells in healthy skin, whereas downregulated in the acne-involved sebaceous glands. Testosterone/LA or insulin-like growth factor-1 inhibited starvation-induced sebocyte autophagy. Pharmacological inhibition of autophagy led to increased sebaceous lipid accumulation. Contrary, rapamycin inhibited the testosterone/LA-induced lipogenesis and expression of fatty acid synthesis genes via activating the autophagy pathway. 13-cis-RA increased autophagy in SZ95 sebocytes, partly via FoxO1 activation, and inhibition of autophagy abolished the sebosuppressive effect of 13-cis-RA.

Conclusions: Autophagy plays an important role in the modulation of lipogenesis in human sebocytes and is involved in the sebostatic effect of 13-cis-RA.
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http://dx.doi.org/10.1016/j.jdermsci.2020.04.001DOI Listing
May 2020

Aryl Hydrocarbon Receptor Modulates the Expression of TNF-α and IL-8 in Human Sebocytes via the MyD88-p65NF-κB/p38MAPK Signaling Pathways.

J Innate Immun 2019 27;11(1):41-51. Epub 2018 Jul 27.

Department of Dermatology, Renji Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai,

Activation of Toll-like receptor (TLR)-2 and subsequent inflammatory response contribute to lesion development in acne vulgaris. A cross-talk between aryl hydrocarbon receptor (AhR), a cytosolic receptor protein that responds to environmental and physiological stress, and TLRs has recently been reported. In this study, we explored the possible role of AhR in the effects induced on cultured human SZ95 sebocytes by peptidoglycan (PGN), a classic TLR2 agonist. PGN-induced secretion of inflammatory factors TNF-α and IL-8 in human SZ95 sebocytes was suppressed after knockdown of AhR and pretreatment with the AhR antagonist CH223191. In addition, the AhR agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) enhanced TNF-α and IL-8 secretion in PGN-pretreated sebocytes. Furthermore, PGN-induced expression of myeloid differentiation factor 88 (MyD88), phospho-p38MAPK (p-p38MAPK), and p-p65NF-κB was strengthened by TCDD and repressed by CH223191. AhR inhibition by transfecting shRNA blocked the ability of PGN to stimulate phosphorylation of p38MAPK and p65NF-κB in SZ95 sebocytes. Overall, these data demonstrate that AhR is able to modulate PGN-induced expression of TNF-α and IL-8 in human SZ95 sebocytes involving the MyD88-p65NF-κB/p38MAPK signaling pathway, which probably indicates a new mechanism in TLR2-mediated acne.
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http://dx.doi.org/10.1159/000491029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6738280PMC
February 2020

Effects of Extracellular Calcium and 1,25 dihydroxyvitamin D3 on Sebaceous Gland Cells In vitro and In vivo.

Acta Derm Venereol 2017 Mar;97(3):313-320

Department of Dermatology, Venereology, Allergology and Immunology, Dessau Medical Center, DE-06847 Dessau, Germany.

Calcium and 1,25 dihydroxyvitamin D3 (1,25(OH)2D3) are promoters of epithelial cell functions; however their effects on sebaceous glands are unknown. In this study, morphology, ultrastructure, cell numbers, lipid synthesis and apoptosis of SZ95 sebocytes were assessed in vitro under different concentrations of extracellular calcium with or without 1,25(OH)2D3. Moreover, serum calcium and 1,25(OH)2D3 levels were assessed in acne and non-acne patients (controls). Under conditions of low extracellular calcium, lipogenesis and cell detachment were observed. Increasing extracellular calcium enhanced sebocyte numbers, induced epithelial morphology and reduced lipogenesis. Moreover, a reduction in extracellular calcium reduced E-cadherin and enhanced caspase 3/7 activity (apoptosis), whereas calcium chelation by EGTA (ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'-tetraacetic acid) resulted in enhanced lipogenesis. 1,25(OH)2D3 decreased sebaceous lipogenesis, but also induced signs of autophagy. In the clinical study, patients and controls exhibited normal serum calcium levels. Younger acne patients presented lower 1,25(OH)2D3 levels than did older ones. In conclusion, extracellular calcium and 1,25(OH)2D3 regulate sebocyte morphology, increase cell numbers, decrease sebaceous lipogenesis and induce cell autophagy in vitro. The increased ionized calcium and the reduced 1,25(OH)2D3 levels detected in the serum of younger patients with acne may contribute respectively to increased sebaceous gland volume and enhanced lipogenesis.
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http://dx.doi.org/10.2340/00015555-2525DOI Listing
March 2017

PI3K/AKT Signaling Pathway Is Essential for Survival of Induced Pluripotent Stem Cells.

PLoS One 2016 3;11(5):e0154770. Epub 2016 May 3.

Departments of Dermatology, Venereology, Allergology and Immunology, Dessau Medical Center, Dessau, Germany.

Apoptosis is a highly conserved biochemical mechanism which is tightly controlled in cells. It contributes to maintenance of tissue homeostasis and normally eliminates highly proliferative cells with malignant properties. Induced pluripotent stem cells (iPSCs) have recently been described with significant functional and morphological similarities to embryonic stem cells. Human iPSCs are of great hope for regenerative medicine due to their broad potential to differentiate into specialized cell types in culture. They may be useful for exploring disease mechanisms and may provide the basis for future cell-based replacement therapies. However, there is only poor insight into iPSCs cell signaling as the regulation of apoptosis. In this study, we focused our attention on the apoptotic response of Alzheimer fibroblast-derived iPSCs and two other Alzheimer free iPSCs to five biologically relevant kinase inhibitors as well as to the death ligand TRAIL. To our knowledge, we are the first to report that the relatively high basal apoptotic rate of iPSCs is strongly suppressed by the pancaspase inhibitor QVD-Oph, thus underlining the dependency on proapoptotic caspase cascades. Furthermore, wortmannin, an inhibitor of phosphoinositid-3 kinase / Akt signaling (PI3K-AKT), dramatically and rapidly induced apoptosis in iPSCs. In contrast, parental fibroblasts as well as iPSC-derived neuronal cells were not responsive. The resulting condensation and fragmentation of DNA and decrease of the membrane potential are typical features of apoptosis. Comparable effects were observed with an AKT inhibitor (MK-2206). Wortmannin resulted in disappearance of phosphorylated AKT and activation of the main effector caspase-3 in iPSCs. These results clearly demonstrate for the first time that PI3K-AKT represents a highly essential survival signaling pathway in iPSCs. The findings provide improved understanding on the underlying mechanisms of apoptosis regulation in iPSCs.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0154770PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4854383PMC
July 2017

Ex vivo human skin and SZ95 sebocytes exhibit a homoeostatic interaction in a novel coculture contact model.

Exp Dermatol 2015 Jul 4;24(7):497-502. Epub 2015 May 4.

Departments of Dermatology, Venereology, Allergology and Immunology, Dessau Medical Center, Dessau, Germany.

The sebaceous gland displays key functions of the human skin, such as hormone synthesis in situ, antimicrobial activity and participation to inflammatory responses. Consequently, there is an emerging need of advanced in vitro models to study complex interactions between the sebaceous gland and the other skin compartments. Despite the evolution of both full-skin organ culture and reconstructed three-dimensional skin models, no satisfactory solutions have been provided for the integration of sebaceous glands and/or sebaceous gland cells in those models, probably due to their problematic maintenance both in vitro and ex vivo. We have developed a coculture model of explant skin in direct contact with immortalized SZ95 sebocytes, which resulted in overall improved structural integrity of the epidermis, higher percentage of proliferating basal epidermal cells and reduced apoptosis of differentiating keratinocytes after 6 days, as detected by Ki67 and TUNEL staining, respectively. Furthermore SZ95 sebocytes exhibited morphological and biochemical signs of normal differentiation and lipid accumulation, while interleukin-6 expression in the supernatant of the cocultures was decreased in comparison with the control. The data provide evidence of a beneficial interaction between sebocytes and skin explants and provide the rationale for their integration in future three-dimensional skin models.
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http://dx.doi.org/10.1111/exd.12712DOI Listing
July 2015

Induced pluripotent stem cell-derived neuronal cells from a sporadic Alzheimer's disease donor as a model for investigating AD-associated gene regulatory networks.

BMC Genomics 2015 Feb 14;16:84. Epub 2015 Feb 14.

Institute for Stem Cell Research and Regenerative Medicine, Heinrich Heine University Duesseldorf, Moorenstr. 5, 40225, Duesseldorf, Germany.

Background: Alzheimer's disease (AD) is a complex, irreversible neurodegenerative disorder. At present there are neither reliable markers to diagnose AD at an early stage nor therapy. To investigate underlying disease mechanisms, induced pluripotent stem cells (iPSCs) allow the generation of patient-derived neuronal cells in a dish.

Results: In this study, employing iPS technology, we derived and characterized iPSCs from dermal fibroblasts of an 82-year-old female patient affected by sporadic AD. The AD-iPSCs were differentiated into neuronal cells, in order to generate disease-specific protein association networks modeling the molecular pathology on the transcriptome level of AD, to analyse the reflection of the disease phenotype in gene expression in AD-iPS neuronal cells, in particular in the ubiquitin-proteasome system (UPS), and to address expression of typical AD proteins. We detected the expression of p-tau and GSK3B, a physiological kinase of tau, in neuronal cells derived from AD-iPSCs. Treatment of neuronal cells differentiated from AD-iPSCs with an inhibitor of γ-secretase resulted in the down-regulation of p-tau. Transcriptome analysis of AD-iPS derived neuronal cells revealed significant changes in the expression of genes associated with AD and with the constitutive as well as the inducible subunits of the proteasome complex. The neuronal cells expressed numerous genes associated with sub-regions within the brain thus suggesting the usefulness of our in-vitro model. Moreover, an AD-related protein interaction network composed of APP and GSK3B among others could be generated using neuronal cells differentiated from two AD-iPS cell lines.

Conclusions: Our study demonstrates how an iPSC-based model system could represent (i) a tool to study the underlying molecular basis of sporadic AD, (ii) a platform for drug screening and toxicology studies which might unveil novel therapeutic avenues for this debilitating neuronal disorder.
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http://dx.doi.org/10.1186/s12864-015-1262-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4344782PMC
February 2015

Identification of biomarkers of human skin ageing in both genders. Wnt signalling - a label of skin ageing?

PLoS One 2012 30;7(11):e50393. Epub 2012 Nov 30.

Departments of Dermatology, Venereology, Allergology and Immunology, Dessau Medical Center, Dessau, Germany.

The goal of our work has been to investigate the mechanisms of gender-independent human skin ageing and examine the hypothesis of skin being an adequate model of global ageing. For this purpose, whole genome gene profiling was employed in sun-protected skin obtained from European Caucasian young and elderly females (mean age 26.7±4 years [n1 = 7] and 70.75±3.3 years [n2 = 4], respectively) and males (mean age 25.8±5.2 years [n3 = 6] and 76±3.8 years [n4 = 7], respectively) using the Illumina array platform. Confirmation of gene regulation was performed by real-time RT-PCR and immunohistochemistry. 523 genes were significantly regulated in female skin and 401 genes in male skin for the chosen criteria. Of these, 183 genes exhibited increased and 340 decreased expression in females whereas 210 genes showed increased and 191 decreased expression in males with age. In total, 39 genes were common in the target lists of significant regulated genes in males and females. 35 of these genes showed increased (16) or decreased (19) expression independent of gender. Only 4 overlapping genes (OR52N2, F6FR1OP2, TUBAL3 and STK40) showed differential regulation with age. Interestingly, Wnt signalling pathway showed to be significantly downregulated in aged skin with decreased gene and protein expression for males and females, accordingly. In addition, several genes involved in central nervous system (CNS) ageing (f.i. APP, TAU) showed to be expressed in human skin and were significanlty regulated with age. In conclusion, our study provides biomarkers of endogenous human skin ageing in both genders and highlight the role of Wnt signalling in this process. Furthermore, our data give evidence that skin could be used as a good alternative to understand ageing of different tissues such as CNS.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0050393PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3511529PMC
May 2013

Mutual regulation of Bcl-2 proteins independent of the BH3 domain as shown by the BH3-lacking protein Bcl-x(AK).

PLoS One 2012 10;7(4):e34549. Epub 2012 Apr 10.

Department of Dermatology and Allergy, Skin Cancer Center, University Medical Center Charité, Berlin, Germany.

The BH3 domain of Bcl-2 proteins was regarded as indispensable for apoptosis induction and for mutual regulation of family members. We recently described Bcl-x(AK), a proapoptotic splice product of the bcl-x gene, which lacks BH3 but encloses BH2, BH4 and a transmembrane domain. It remained however unclear, how Bcl-x(AK) may trigger apoptosis.For efficient overexpression, Bcl-x(AK) was subcloned in an adenoviral vector under Tet-OFF control. The construct resulted in significant apoptosis induction in melanoma and nonmelanoma cell lines with up to 50% apoptotic cells as well as decreased cell proliferation and survival. Disruption of mitochondrial membrane potential, and cytochrome c release clearly indicated activation of the mitochondrial apoptosis pathways. Both Bax and Bak were activated as shown by clustering and conformation analysis. Mitochondrial translocation of Bcl-x(AK) appeared as an essential and initial step. Bcl-x(AK) was critically dependent on either Bax or Bak, and apoptosis was abrogated in Bax/Bak double knockout conditions as well by overexpression of Bcl-2 or Bcl-x(L). A direct interaction with Bcl-2, Bax, Bad, Noxa or Puma was however not seen by immunoprecipitation. Thus besides BH3-mediated interactions, there exists an additional way for mutual regulation of Bcl-2 proteins, which is independent of the BH3. This pathway appears to play a supplementary role also for other proapoptotic family members, and its unraveling may help to overcome therapy resistance in cancer.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0034549PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3323539PMC
October 2012

Mitochondrial-associated cell death mechanisms are reset to an embryonic-like state in aged donor-derived iPS cells harboring chromosomal aberrations.

PLoS One 2011 14;6(11):e27352. Epub 2011 Nov 14.

Molecular Embryology and Aging Group, Department of Vertebrate Genomics, Max Planck Institute for Molecular Genetics, Berlin, Germany.

Somatic cells reprogrammed into induced pluripotent stem cells (iPSCs) acquire features of human embryonic stem cells (hESCs) and thus represent a promising source for cellular therapy of debilitating diseases, such as age-related disorders. However, reprogrammed cell lines have been found to harbor various genomic alterations. In addition, we recently discovered that the mitochondrial DNA of human fibroblasts also undergoes random mutational events upon reprogramming. Aged somatic cells might possess high susceptibility to nuclear and mitochondrial genome instability. Hence, concerns over the oncogenic potential of reprogrammed cells due to the lack of genomic integrity may hinder the applicability of iPSC-based therapies for age-associated conditions. Here, we investigated whether aged reprogrammed cells harboring chromosomal abnormalities show resistance to apoptotic cell death or mitochondrial-associated oxidative stress, both hallmarks of cancer transformation. Four iPSC lines were generated from dermal fibroblasts derived from an 84-year-old woman, representing the oldest human donor so far reprogrammed to pluripotency. Despite the presence of karyotype aberrations, all aged-iPSCs were able to differentiate into neurons, re-establish telomerase activity, and reconfigure mitochondrial ultra-structure and functionality to a hESC-like state. Importantly, aged-iPSCs exhibited high sensitivity to drug-induced apoptosis and low levels of oxidative stress and DNA damage, in a similar fashion as iPSCs derived from young donors and hESCs. Thus, the occurrence of chromosomal abnormalities within aged reprogrammed cells might not be sufficient to over-ride the cellular surveillance machinery and induce malignant transformation through the alteration of mitochondrial-associated cell death. Taken together, we unveiled that cellular reprogramming is capable of reversing aging-related features in somatic cells from a very old subject, despite the presence of genomic alterations. Nevertheless, we believe it will be essential to develop reprogramming protocols capable of safeguarding the integrity of the genome of aged somatic cells, before employing iPSC-based therapy for age-associated disorders.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0027352PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3215709PMC
March 2012

Skin and brain age together: The role of hormones in the ageing process.

Exp Gerontol 2010 Oct 16;45(10):801-13. Epub 2010 Aug 16.

Department of Dermatology, Dessau Medical Center, Germany.

The importance of the endocrine environment in the initiation of the ageing process has been elucidated in several in vivo and in vitro studies. Changes in endocrine pathways accompany healthy ageing, these include the growth hormone/insulin-like growth factor-I axis (somatopause) and that of sexual hormones, namely estradiol (menopause), testosterone (andropause), and dehydroepiandrosterone and its sulphate (adrenopause). The clinical significance of these changes is variable and results in morphological and functional alterations of all organ systems including the skin and the central nervous system. Moreover, the pathogenesis of age-associated diseases such as epithelial skin cancer and neurodegenerative diseases has been partly attributed to the lack of hormones. Several studies have been conducted in an attempt to reverse the ageing process and clinical signs by substitution of the serum hormone levels in older individuals, however the benefits of hormone replacement therapy, if any, are still controversial. On the other hand, recent data suggest that skin is a window to the human organism and represents an adequate model for ageing research, also implying the use of skin samples for evaluating the ageing status of the central nervous system.
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http://dx.doi.org/10.1016/j.exger.2010.08.005DOI Listing
October 2010

Efficient and selective tumor cell lysis and induction of apoptosis in melanoma cells by a conditional replication-competent CD95L adenovirus.

Exp Dermatol 2010 Aug;19(8):e56-66

Department of Dermatology and Allergy, HTCC - Skin Cancer Center Charité, Charité-Universitätsmedizin Berlin.

The high mortality of melanoma demands the development of new strategies, and gene therapy may be considered provided improvements in efficacy and selectivity. Overexpression of the death ligand CD95L/FasL has been shown in previous studies as highly effective for apoptosis induction in melanoma cells. For efficient and selective targeting of melanoma, a conditional replication-competent adenoviral vector was constructed (Ad5-FFE-02), which drives CD95L expression by a tetracycline-inducible promoter. For restricting its replication to melanoma cells, the adenoviral E1A gene is controlled by a tyrosinase-derived promoter. Furthermore, adenoviral E1B was deleted and a mutated E1A was used to preferentially support replication in tumor cells. Proving its high selectivity and efficiency, strong expression of E1A and doxycycline-dependent induction of CD95L were characteristic for tyrosinase-positive melanoma cells after Ad5-FFE-02 transduction, whereas absent in non-melanoma cell lines. Importantly, Ad5-FFE-02-mediated cell lysis was restricted to melanoma cells, and induction of apoptosis was found only in tyrosinase and CD95 expressing cells. Finally, the combination of adenoviral replication and CD95L-mediated apoptosis resulted in an enhanced repression of melanoma cell growth. This new adenoviral vector may provide a basis for an efficient targeting of melanoma.
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http://dx.doi.org/10.1111/j.1600-0625.2009.00977.xDOI Listing
August 2010

Expression and function of bcl-2 proteins in melanoma.

Curr Genomics 2008 Sep;9(6):409-19

Charité - Universitätsmedizin Berlin, Department of Dermatology and Allergy, Skin Cancer Center Charité, Berlin, Germany.

Bcl-2 proteins are critical regulators of mitochondrial membrane permeability and the proapoptotic mitochondrial pathway. The family encloses pro- and antiapoptotic factors encoded by over 15 genes, which frequently give rise to alternative splice products. Antiapoptotic, proapoptotic multidomain, and proapoptotic BH3-only proteins are characterized by the presence of at least one of four Bcl-2 homology domains (BH 1-4). Their expression and activities are controlled by survival pathways as MAP kinases and protein kinase B/Akt, which are in touch with a number of transcription factors. In melanoma, the mitochondrial apoptosis pathways and Bcl-2 proteins appear of particular importance for apoptosis resistance, which has been addressed in clinical trials applying antisense-Bcl-2. Overexpression or induction of proapoptotic Bcl-2 proteins as well as the use of small molecule mimetics for the proapoptotic BH3 domain are further promising strategies.
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http://dx.doi.org/10.2174/138920208785699571DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2691663PMC
September 2008

Apoptosis induction by Bcl-2 proteins independent of the BH3 domain.

Biochem Pharmacol 2008 Dec 22;76(11):1612-9. Epub 2008 Aug 22.

Charité-Universitätsmedizin Berlin, Department of Dermatology and Allergy, Skin Cancer Center Charité, Charité Campus Mitte, Charitéplatz 1, Berlin, Germany.

Bcl-2 proteins, characterized by up to four Bcl-2 homology domains (BH1-BH4), are critical regulators of the mitochondrial proapoptotic pathway. Three major subgroups have been described, namely antiapoptotic proteins, proapoptotic multidomain and BH3-only proteins. These are basic for present models explaining the regulation of the mitochondrial outer membrane permeability. However, several Bcl-2 proteins have been described that do not fit into these models, due to their atypical domain structure or due to their ability to induce apoptosis independently of BH3. These proteins are indicators for new mechanisms in apoptosis control by Bcl-2 proteins, which may supply additional targets for novel therapeutic approaches.
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http://dx.doi.org/10.1016/j.bcp.2008.08.013DOI Listing
December 2008

Apoptosis pathways and oncolytic adenoviral vectors: promising targets and tools to overcome therapy resistance of malignant melanoma.

Exp Dermatol 2008 Jan;17(1):1-11

Department of Dermatology and Allergy, Skin Cancer Center, Charité- Universitätsmedizin Berlin, Berlin, Germany.

In the last decades melanoma incidence has been increasing worldwide, while mortality remained on a high level. Until now, there is no suitable therapy for metastasized melanoma, which could lead to a significant increase in overall survival. Apoptosis deficiency is supposed to be a critical factor for therapy resistance, and previous work has characterized the basic mechanisms of apoptosis regulation in melanoma. Genes and strategies suitable for efficient induction of apoptosis in melanoma cells were identified, which are based on proapoptotic Bcl-2 proteins (Bcl-x(S), Bcl-x(AK), Bik/Nbk and Bax) as well as on tumor necrosis factor (TNF)-related death ligands (CD95L/Fas ligand and TNF-related apoptosis-inducing ligand, TRAIL). Proapoptotic genes may be employed in improved gene therapeutic strategies, based on conditional oncolytic adenoviral vectors.
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http://dx.doi.org/10.1111/j.1600-0625.2007.00655.xDOI Listing
January 2008

Overcoming apoptosis deficiency of melanoma-hope for new therapeutic approaches.

Drug Resist Updat 2007 Dec 3;10(6):218-34. Epub 2007 Dec 3.

Charité-Universitätsmedizin Berlin, Department of Dermatology and Allergy, Skin Cancer Center Charité, Charité Campus Mitte, Charitéplatz 1, 10117 Berlin, Germany.

The increased incidence of malignant melanoma in the last decades, its high mortality and pronounced therapy resistance pose an enormous challenge. Important therapeutic targets for melanoma are the induction of apoptosis and suppression of survival pathways. Preclinical studies have demonstrated the efficacy of pro-apoptotic Bcl-2 proteins and of death receptor ligands to trigger apoptosis in melanoma cells. In the clinical setting, BH3 domain mimics and death receptor agonists are therefore considered as promising, specific novel treatments to add to the conventional pro-apoptotic strategies such as chemo- or radiotherapy. However, constitutively activated survival pathways, in particular the mitogen-activated protein kinases, protein kinase B/Akt and nuclear factor (NF)-kappaB, all may work in concert to prevent effective therapy. Thus, selective biologicals developed with the aim to inhibit pro-survival signaling are currently tested in melanoma. For highly therapy-resistant tumors such as melanoma, development of novel drug combinations will be essential, and combinations of survival inhibitors and pro-apoptotic mediators appear most promising. The challenge of the near future will be to make a rational choice of the multiple possible combinations and protocols. This review gives a critical overview of proteins involved in melanoma chemoresistance, which are targets for current drug development leading to the best choice for future trials.
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http://dx.doi.org/10.1016/j.drup.2007.09.001DOI Listing
December 2007

Inhibition of the alternative complement activation pathway in traumatic brain injury by a monoclonal anti-factor B antibody: a randomized placebo-controlled study in mice.

J Neuroinflammation 2007 May 2;4:13. Epub 2007 May 2.

Department of Trauma and Reconstructive Surgery, Charité University Medical School, Campus Benjamin Franklin, Berlin, Germany.

Background: The posttraumatic response to traumatic brain injury (TBI) is characterized, in part, by activation of the innate immune response, including the complement system. We have recently shown that mice devoid of a functional alternative pathway of complement activation (factor B-/- mice) are protected from complement-mediated neuroinflammation and neuropathology after TBI. In the present study, we extrapolated this knowledge from studies in genetically engineered mice to a pharmacological approach using a monoclonal anti-factor B antibody. This neutralizing antibody represents a specific and potent inhibitor of the alternative complement pathway in mice.

Methods: A focal trauma was applied to the left hemisphere of C57BL/6 mice (n = 89) using a standardized electric weight-drop model. Animals were randomly assigned to two treatment groups: (1) Systemic injection of 1 mg monoclonal anti-factor B antibody (mAb 1379) in 400 mul phosphate-buffered saline (PBS) at 1 hour and 24 hours after trauma; (2) Systemic injection of vehicle only (400 mul PBS), as placebo control, at identical time-points after trauma. Sham-operated and untreated mice served as additional negative controls. Evaluation of neurological scores and analysis of brain tissue specimens and serum samples was performed at defined time-points for up to 1 week. Complement activation in serum was assessed by zymosan assay and by murine C5a ELISA. Brain samples were analyzed by immunohistochemistry, terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) histochemistry, and real-time RT-PCR.

Results: The mAb 1379 leads to a significant inhibition of alternative pathway complement activity and to significantly attenuated C5a levels in serum, as compared to head-injured placebo-treated control mice. TBI induced histomorphological signs of neuroinflammation and neuronal apoptosis in the injured brain hemisphere of placebo-treated control mice for up to 7 days. In contrast, the systemic administration of an inhibitory anti-factor B antibody led to a substantial attenuation of cerebral tissue damage and neuronal cell death. In addition, the posttraumatic administration of the mAb 1379 induced a neuroprotective pattern of intracerebral gene expression.

Conclusion: Inhibition of the alternative complement pathway by posttraumatic administration of a neutralizing anti-factor B antibody appears to represent a new promising avenue for pharmacological attenuation of the complement-mediated neuroinflammatory response after head injury.
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http://dx.doi.org/10.1186/1742-2094-4-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1867808PMC
May 2007

Pharmacological complement inhibition at the C3 convertase level promotes neuronal survival, neuroprotective intracerebral gene expression, and neurological outcome after traumatic brain injury.

Exp Neurol 2006 Jun 20;199(2):454-64. Epub 2006 Mar 20.

Department of Trauma and Reconstructive Surgery, Charité-University Medical School, Campus Benjamin Franklin, Hindenburgdamm 30, 12200 Berlin, Germany.

The complement system represents an important mediator of neuroinflammation in traumatic brain injury. We have previously shown that transgenic mice with central nervous system-targeted overexpression of Crry, a potent murine complement inhibitor at the level of C3 convertases, are protected from complement-mediated neuropathological sequelae in brain-injured mice. This knowledge was expanded in the present study to a pharmacological approach by the use of a recombinant Crry molecule (termed Crry-Ig) which was recently made available in a chimeric form fused to the non-complement fixing mouse IgG1 Fc region. In a standardized model of closed head injury in mice, the systemic injection of 1 mg Crry-Ig at 1 h and 24 h after trauma resulted in a significant neurological improvement for up to 7 days, as compared to vehicle-injected control mice (P < 0.05, repeated measures ANOVA). Furthermore, the extensive neuronal destruction seen in the hippocampal CA3/CA4 sublayers in head-injured mice with vehicle injection only was shown to be preserved - to a similar extent as in "sham"-operated mice - by the posttraumatic injection of Crry-Ig. Real-time RT-PCR analysis revealed that the post-treatment with Crry-Ig resulted in a significant up-regulation of candidate neuroprotective genes in the injured hemisphere (Bcl-2, C1-Inh, CD55, CD59), as compared to the vehicle control group (P < 0.01, unpaired Student's t test). Increased intracerebral Bcl-2 expression by Crry-Ig treatment was furthermore confirmed at the protein level by Western blot analysis. These data suggest that pharmacological complement inhibition represents a promising approach for attenuation of neuroinflammation and secondary neurodegeneration after head injury.
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http://dx.doi.org/10.1016/j.expneurol.2006.01.033DOI Listing
June 2006

Selective induction of apoptosis in melanoma cells by tyrosinase promoter-controlled CD95 ligand overexpression.

J Invest Dermatol 2005 Jan;124(1):221-8

Department of Dermatology, Charite-Universitätsmedizin Berlin, Campus Benjamin Franklin, Berlin, Germany.

Induction of apoptosis has been demonstrated previously by overexpression of CD95 ligand (CD95L) in cultured human melanoma cells. For in vivo approaches based on CD95L, however, targeted expression is a prerequisite and tyrosinase promoters have been considered for selection. Luciferase reporter gene assays performed for a representative panel of melanoma cell lines characterized by strong (SK-Mel-19), moderate (SK-Mel-13, MeWo), weak (A-375), and missing expression (M-5) of endogenous tyrosinase revealed high tyrosinase promoter activities in SK-Mel-19, SK-Mel-13, and MeWo, but only weak activities in A-375 and M-5 as well as in non-melanoma cell lines. After transfection of a CMV promoter CD95L expression construct, melanoma cells were found highly sensitive, as compared with non-melanoma cells. By applying a tyrosinase promoter CD95L construct, apoptosis was selectively induced in SK-Mel-19, SK-Mel-13, MeWo as well as in A-375, which was characterized by high CD95 surface expression and high sensitivity to agonistic CD95 activation. M5 and non-melanoma cell lines remained uninfluenced. Also, resistance to agonistic CD95 activation seen in MeWo characterized by weak CD95 surface expression was overcome by overexpression of CD95L. Our investigations provide evidence that tyrosinase promoter CD95L constructs may be of value for selective induction of apoptosis in therapeutic strategies for melanoma.
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http://dx.doi.org/10.1111/j.0022-202X.2004.23572.xDOI Listing
January 2005

CD95/Fas signaling in human melanoma cells: conditional expression of CD95L/FasL overcomes the intrinsic apoptosis resistance of malignant melanoma and inhibits growth and progression of human melanoma xenotransplants.

Oncogene 2003 Dec;22(57):9131-41

Department of Dermatology, Charité-Universitätsmedizin Berlin, Campus Benjamin Franklin, 14195 Berlin, Germany.

The significance of CD95/Fas ligand expression by melanoma cells has remained a controversial matter in recent years. On the other hand, CD95 activation may represent a powerful tool for eliminating tumor cells. Here, we demonstrate expression of CD95 in 15/17 human melanoma cell lines analysed, but complete lack of CD95 ligand (CD95L). Overexpression of CD95 in a tetracycline-inducible expression system enhanced melanoma cell sensitivity to CD95 ligation but was unable to trigger apoptosis by itself. In clear contrast, all melanoma cells tested responded with increased apoptosis to conditional expression of CD95L (2-10-fold), both after transient and after stable transfection. Activation of caspase-8, Bid cleavage, cytochrome c release and caspase-3 activation followed after CD95L induction indicating a functional CD95-signaling cascade. CD95L was also able to enhance the proapoptotic effect of chemotherapeutics applied in parallel. Nude mouse experiments revealed that tumorigenicity was lost when melanoma xenografts were triggered to express CD95L. In addition, further progression of pre-existing melanomas was inhibited and even regression was seen after induction of CD95L expression. Due to these data, transfection of CD95L proofs as a highly efficient tool against melanoma cells in vitro and in vivo, and targeted expression of CD95L may thus represent a suitable strategy for melanoma therapy.
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http://dx.doi.org/10.1038/sj.onc.1207228DOI Listing
December 2003

Conditional expression of exogenous Bcl-X(S) triggers apoptosis in human melanoma cells in vitro and delays growth of melanoma xenografts.

FEBS Lett 2003 Oct;553(3):250-6

Charité - Universitätsmedizin Berlin, Campus Benjamin Franklin, Department of Dermatology, Fabeckstrasse 60-62, 14195 Berlin, Germany.

The Bcl-2-related proteins Bcl-X(L) and Bcl-X(S) represent alternative splice products and exert opposite activities in the control of apoptosis, but their significance for melanoma is not yet clear. Applying the tetracycline-inducible expression system Tet-On, we found overexpression of Bcl-X(S) by itself sufficient to induce apoptosis in vitro in stably transfected human melanoma cell lines. Combination with proapoptotic agents such as etoposide, pamidronate, and ceramide resulted in additive proapoptotic effects, whereas Bcl-X(L) protected from apoptosis caused via CD95/Fas stimulation. In nude mice growth of melanoma xenotransplants derived from stably transfected cells was significantly reduced after induction of Bcl-X(S) by doxycycline. Our results indicate that Bcl-X proteins are of major importance for control of apoptosis in malignant melanoma.
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http://dx.doi.org/10.1016/s0014-5793(03)01017-2DOI Listing
October 2003

The role of nuclear factor-kappa B and melanogenesis in tumor necrosis factor-alpha-induced apoptosis of normal human melanocytes.

Skin Pharmacol Appl Skin Physiol 2002 Sep-Oct;15(5):321-9

Department of Dermatology, University Medical Center Benjamin Franklin, Free University of Berlin, Berlin, Germany.

The proapoptotic potential of tumor necrosis factor-alpha (TNF-alpha) has been demonstrated for various cell types, whereas nuclear factor-kappaB (NF-kappaB) is known to support the transcription of prosurvival genes. In the present study, investigation of normal human melanocytes revealed induction of apoptosis after TNF-alpha treatment (100 U/ml) in only 3 out of 11 cultures analyzed, whereas 8 cultures remained largely resistant. In sensitive cultures, NF-kappaB binding activity was found increased after TNF-alpha treatment; apoptosis-resistant cells were characterized by relatively high basic NF-kappaB binding activities and did not show NF-kappaB activation after TNF-alpha treatment. Inhibition of NF-kappaB by a specific inhibitor, Bay-11, either induced apoptosis itself or resistant melanocyte cultures became sensitive to TNF-alpha treatment. No correlation was found between apoptosis sensitivity and the expression of TNF receptor-1 or the expression of Bax, Bcl-2 and Bcl-X(L). A strong correlation, however, was found regarding the pigmentation degree, as high pigmentation correlated with apoptosis resistance and sensitive melanocyte cultures were weakly pigmented. These data may indicate that in cultured melanocytes, high levels of melanogenesis lead to an increase in oxidative stress which itself causes NF-kappaB activation. NF-kappaB mediates the transcription of antiapoptotic factors which may block TNF-alpha-induced apoptosis at early steps of the signal cascade.
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http://dx.doi.org/10.1159/000064536DOI Listing
November 2002

Bcl-2 overexpression prevents apoptosis induced by ceramidase inhibitors in malignant melanoma and HaCaT keratinocytes.

FEBS Lett 2002 Apr;516(1-3):47-52

Department of Dermatology, University Medical Center Benjamin Franklin, The Free University of Berlin, 14195, Berlin, Germany.

We examined the biological effects of the ceramide analogues (1S,2R)-2-N-myristoylamino-1-phenyl-1-propanol (D-e-MAPP) and (1R,2R)-2-N-myristoylamino-1-(4-nitrophenyl)-1,3-propandiol (D-NMAPPD) on human HaCaT keratinocytes and human melanoma cells. We could demonstrate that D-e-MAPP and D-NMAPPD are able to suppress acid ceramidase activity. The elevation of the endogenous level of ceramide is followed by induction of apoptosis and suppression of proliferation in HaCaT keratinocytes. Moreover, we recently identified a group of human melanoma cell populations which are heterogeneously susceptible to C2-ceramide-mediated apoptosis. Studies with these melanoma cells revealed correlation between ceramide-mediated apoptosis and D-NMAPPD-induced apoptosis, confirming the effect of this inhibitor on ceramide signaling in human melanoma cells. These findings suggest ceramidase inhibitors as a potential new therapeutical class of antiproliferative and cytostatic drugs.
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http://dx.doi.org/10.1016/s0014-5793(02)02472-9DOI Listing
April 2002
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