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1.
Int J Mol Sci ; 24(20)2023 Oct 19.
Article in English | MEDLINE | ID: mdl-37895015

ABSTRACT

Enhanced glycolysis (Warburg effect) driven by the BRAF oncogene, dysregulated GAPDH expression, and activation of the PI3K/AKT/mTOR signaling pathway may significantly contribute to the resistance-targeted therapy of BRAF-mutated melanomas. Therefore, we aimed to study for the first time the anti-tumor activity of the GAPDH inhibitor GP-2250 in BRAF-mutated melanoma cell lines and benign melanocytes. We employed three melanoma cell lines and one primary melanocyte cell line (Ma-Mel-61a, Ma-Mel-86a, SH-4 and ATCC-PCS-200-013, respectively), which were exposed to different GP-2250 doses. GP-2250's effects on cell proliferation and viability were evaluated by means of the BrdU and MTT assays, respectively. The RealTime-Glo Annexin V Apoptosis and Necrosis Assay was performed for the evaluation of apoptosis and necrosis induction. RT-PCR and western blotting were implemented for the determination of AKT and STAT3 gene and protein expression analyses, respectively. The melanoma cell lines showed a dose-dependent response to GP-2250 during BrDU and MTT testing. The RealTime-Glo Annexin V assay revealed the heterogenous impact of GP-2250 on apoptosis as well as necrosis. With respect to the melanoma cell lines Ma-Mel-86a and SH-4, the responses and dosages were comparable to those used for the MTT viability assay. Using the same dose range of GP-2250 administered to melanoma cells, however, we observed neither the noteworthy apoptosis nor necrosis of GP-2250-treated benign melanocytes. The gene expression profiles in the melanoma cell lines for AKT and STAT3 were heterogenous, whereby AKT as well as STAT3 gene expression were most effectively downregulated using the highest GP-2250 doses. Immunoblotting revealed that there was a time-dependent decrease in protein expression at the highest GP-2250 dose used, whereas a time- as well as dose-dependent AKT decrease was predominantly observed in Ma-Mel-61a. The STAT3 protein expression of Ma-Mel-86a and SH-4 was reduced in a time-dependent pattern at lower and moderate doses. STAT3 expression in Ma-Me-61a was barely altered by GP-2250. In conclusion, GP-2250 has anti-neoplastic effects in BRAF-mutated melanoma cell lines regarding tumor cell viability, proliferation, and apoptosis/necrosis. GP-2250 is able to downregulate the gene and protein expression of aberrant tumorigenic pathways in melanoma cell lines. Since GP-2250 is a GAPDH inhibitor, the substance may be a promising combination therapy for tumors presenting the Warburg effect, such as melanoma.


Subject(s)
Antineoplastic Agents , Melanoma , Humans , Annexin A5 , Antineoplastic Agents/pharmacology , Apoptosis/genetics , Bromodeoxyuridine/metabolism , Cell Line, Tumor , Cell Proliferation , Melanocytes/metabolism , Melanoma/drug therapy , Melanoma/genetics , Melanoma/pathology , Necrosis/drug therapy , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins B-raf/metabolism , Proto-Oncogene Proteins c-akt/metabolism
4.
Dermatology ; 233(2-3): 129-135, 2017.
Article in English | MEDLINE | ID: mdl-28750404

ABSTRACT

BACKGROUND: The pathogenesis of hidradenitis suppurativa (HS), with its complex inflammatory network, is still elusive. Imbalances in DNA methylation can lead to genome destabilization and have been assumed to play a role in inflammatory diseases. Global DNA methylation and hydroxymethylation have not been studied in HS yet. OBJECTIVE: We conducted this study to investigate the global DNA methylation and hydroxymethylation status in lesional and perilesional HS skin compared to healthy controls. METHODS: Immunohistochemical analysis was performed for 5-methylcytosine (5-mC) and 5-hydroxymethylcytosine (5-hmC) in 30 lesional and 30 corresponding healthy-appearing perilesional HS tissue samples. We included 30 healthy subjects as an interindividual control group. RESULTS: 5-hmC levels were significantly lower in healthy-appearing perilesional (p < 0.0001) and lesional HS skin (p < 0.0001) when compared to healthy controls. There was no significant difference between lesional HS skin and perilesional HS skin regarding 5-hmC levels (p = 0.6654). In contrast to 5-hmC, 5-mC staining showed no significant changes between the 3 groups. Univariate analysis revealed no significant association between patients' characteristics, disease severity, and the levels of 5-mC and 5-hmC. CONCLUSION: Our findings indicate that imbalances in DNA hydroxymethylation may play a role in the pathogenesis of HS rather than DNA methylation. Further studies are warranted to investigate the significance of DNA hydroxymethylation and the regulating enzymes in HS in order to advance our knowledge of the inflammatory network in this disease.


Subject(s)
5-Methylcytosine/analogs & derivatives , DNA Methylation , Epigenesis, Genetic , Hidradenitis Suppurativa/genetics , 5-Methylcytosine/metabolism , Case-Control Studies , Female , Humans , Immunohistochemistry , Male , Skin
5.
Postepy Dermatol Alergol ; 34(4): 299-305, 2017 Aug.
Article in English | MEDLINE | ID: mdl-28951703

ABSTRACT

INTRODUCTION: Apart from neutrophils, other immune cells may play a significant pathogenetic role in cutaneous leukocytoclastic vasculitis (CLV). AIM: To investigate lymphocytes and related immunological factors in patients with CLV requiring systemic glucocorticosteroid treatment. MATERIAL AND METHODS: Fourteen patients with severe idiopathic CLV were treated with systemic prednisolone in a tapered dose regimen. Ten healthy individuals served as controls. At baseline and post-treatment, we studied inducer/helper and suppressor/cytotoxic T lymphocytes, B lymphocytes, natural killer cells, CD4+CD25++CD127- cells, CD4+CD25+CD39+ cells and FOXP3, transforming growth factor ß1 (TGF-ß1) and interleukin-10 (IL-10) mRNA levels in the blood using flow cytometry and real time polymerase chain reaction (RT-PCR), respectively. On immunohistochemistry, we studied CD4, CD8, granzyme B, TGF-ß1, and IL-10. RESULTS: Flow cytometry did not show significant differences. The RT-PCR revealed that TGF-ß1 mRNA expression was significantly higher after therapy when compared to baseline and controls. On immunohistology, baseline CLV lesions showed significantly more CD4+ lymphocytes than post-treated CLV and controls. CD8+ expression was significantly higher after therapy when compared to baseline and controls. Baseline granzyme B was significantly increased when compared to treated CLV and controls. The IL-10 expression of treated CLV was significantly increased when compared to baseline CLV and; baseline CLV IL-10 expression was significantly increased as compared to controls. CONCLUSIONS: Circulating T regulatory cells do not play a significant role in the pathogenesis of CLV. T helper cells and granzyme B seem to be involved in the inflammatory cutaneous process of CLV. A resolution of CLV observed after glucocorticosteroid treatment may be mediated via up-regulation of TGF-ß1 and IL-10 in different compartments.

6.
Pharmacogenet Genomics ; 26(5): 248-53, 2016 May.
Article in English | MEDLINE | ID: mdl-26967570

ABSTRACT

Fumaric acid esters (FAE) are beneficial in the treatment of psoriasis. However, about a third of psoriasis patients do not respond to FAE. We aimed to determine whether glutathione-S-transferase (GST) M1 and GSTP1 polymorphisms are associated with treatment outcome in psoriasis patients treated with FAE. We studied 84 psoriasis patients who were treated with FAE for 3 months. FAE nonresponders were defined as having psoriasis area and severity improvement index less than 50% after 3-month therapy. GSTM1 genotyping for gene deletion and GSTP1 exon 5 105 Ile→Val polymorphisms were assessed using a high-resolution melting analysis. A dropout rate of 23.8% (20/84) was found; 25% (16/64) were FAE nonresponders. We observed 42 (84/50%) patients with G 9STM1*0 homozygous alleles and 42 (84/50%) patients with one or two active GSTM1 alleles. The Ile/Ile GSTP1 genotype was observed in 37 (84/44%), the Ile/Val GSTP1 genotype in 38 (84/45.2%) patients and the Val/Val GSTP1 genotype in nine (84/10.7%) patients. There was no significant (P>0.05) association between the GST genotypes assessed and the frequency FAE responder status, except for the Val/Val GSTP1 polymorphism, which was a significant (overall model fit; P=0.0012) predictor for nonresponders with an odds ratio of 43.4 (95% confidence interval: 4.2-511.1). The coefficient of regression was 3.9, with a SE of 1.2 as assessed by logistic regression analysis (P=0.0017). The Val/Val GSTP1 polymorphism predicts nonresponders in FAE treatment of psoriasis patients and may therefore serve as a biomarker that enables a laboratory-based pretreatment selection of patients.


Subject(s)
Fumarates/therapeutic use , Glutathione S-Transferase pi/genetics , Psoriasis/drug therapy , Psoriasis/genetics , Valine/metabolism , Adult , Aged , Female , Humans , Male , Middle Aged , Pharmacogenomic Variants , Severity of Illness Index , Treatment Outcome
7.
Viruses ; 16(9)2024 Sep 15.
Article in English | MEDLINE | ID: mdl-39339947

ABSTRACT

Immune responses of the epithelia of the upper respiratory tract are likely crucial in early inhibition of the viral replication and finally clearance of SARS-CoV-2. We aimed to compare the expression profiles of antimicrobial peptides/proteins (AMPs) and related cytokines observed in the nasopharynx of SARS-CoV-2-infected patients and non-infected controls and to assess the associations between these parameters and COVID-19 patients' outcomes. We included 45 subjects who had tested positive for SARS-CoV-2 and 22 control subjects who had tested negative for SARS-CoV-2. Biomaterial for SARS-CoV-2 detection, as well as gene and protein expression studies, was obtained from all subjects using nasopharyngeal swabs which were performed a maximum of 7 days before inclusion in the study. Univariable and multivariable statistics were performed. When compared to the controls, the mRNA expression levels of human ß-defensin 1 (hBD-1), LL-37, and trappin-2 were significantly higher in specimens of nasopharyngeal swabs from COVID-19 patients. Protein expression of hBD-1 was also increased in the COVID-19 group. mRNA expression levels of interferon-É£ (IFN-É£), tumor necrosis factor- ɑ (TNF-ɑ), and interleukin-6 (IL-6) measured in SARS-CoV-2-infected patients were significantly higher than those observed in the controls, which could also be confirmed in the protein levels of IFN-É£ and IL-6. A significant correlation between mRNA and protein levels could be observed only for IL-6. Univariable analysis revealed that low IFN-É£ mRNA levels were associated with severe/fatal outcomes. The occurrence of COVID-19 pneumonia was significantly associated with lower expression levels of IL-6 mRNA, IFN-É£ mRNA, and TNF-ɑ mRNA. Concerning the severe/fatal outcomes, the multivariable logistic regression model revealed that none of the aforementioned parameters remained significant in the model. However, the logistic regression model revealed that higher TNF-ɑ mRNA expression was a significant independent predictor of absence of pneumonia [odds ratio: 0.35 (95% CI 0.14 to 0.88, p = 0.024)]. In conclusion, nasopharyngeal expression of AMPs (hBD-1, LL-37, and trappin-2) and cytokines (IL-6, IFN-É£, and TNF-ɑ) is upregulated in response to early SARS-CoV-2 infection, indicating that these AMPs and cytokines play a role in the local host defense against the virus. Upregulated nasopharyngeal TNF-ɑ mRNA expression during the early phase of SARS-CoV-2 infection was a significant independent predictor of the absence of COVID-19 pneumonia. Hence, high TNF-ɑ mRNA expression in the nasopharynx appears to be a protective factor for lung complications in COVID-19 patients.


Subject(s)
Antimicrobial Peptides , COVID-19 , Cytokines , Nasopharynx , SARS-CoV-2 , Humans , COVID-19/immunology , COVID-19/virology , COVID-19/metabolism , Nasopharynx/virology , Nasopharynx/metabolism , Female , Male , Middle Aged , SARS-CoV-2/immunology , Cytokines/metabolism , Cytokines/genetics , Adult , Antimicrobial Peptides/genetics , Antimicrobial Peptides/metabolism , Aged
8.
Viruses ; 16(5)2024 05 20.
Article in English | MEDLINE | ID: mdl-38793691

ABSTRACT

In patients with COVID-19, broad panels of immune checkpoint molecules (ICPMs) and the purinergic signaling have not been studied in parallel. We aimed to perform in-depth immunophenotyping of major cell subsets present in human peripheral blood of COVID-19 patients and controls using PD1, TIM3, LAG3, TIGIT, and CD200R, as well as CD39, as markers for the purinergic signaling pathway. We studied 76 COVID-19 patients and 12 healthy controls using peripheral blood mononuclear cells on flow cytometry. Univariable and multivariable statistics were performed. All ICPMs studied were significantly overexpressed on different cell subsets of COVID-19 patients when compared with healthy controls. Elevated lactate dehydrogenase; C-reactive protein; age; and high expression of CD45+, CD39+CD45+, TIM3+CD39+CD4+CD45+, and TIM3+CD39+CD8+CD3+CD4+ cells were significantly associated with severe COVID-19. On multivariable analysis, however, only high expression of CD39+CD45+ (OR 51.4, 95% CI 1.5 to 1763) and TIM3+CD39+CD4+CD3+CD45+ (OR 22.6, 95% CI 1.8 to 277) cells was an independent predictor for severe COVID-19. In conclusion, numerous ICPMs are overexpressed in COVID-19 patients when compared with healthy controls, suggesting a pathophysiological role of these molecules in SARS-CoV-2 infection. However, only TIM3 in co-expression with CD39 remained as a significant independent prognostic ICPM on multivariable analysis. The flow cytometric evaluation of TIM3+CD39+CD4+CD3+CD45+, as well as CD39+CD45+, is a powerful tool for the prognostication of COVID-19 patients on hospital admission.


Subject(s)
Apyrase , COVID-19 , Adult , Aged , Aged, 80 and over , Female , Humans , Male , Middle Aged , Antigens, CD/blood , COVID-19/blood , COVID-19/diagnosis , COVID-19/mortality , Flow Cytometry , Immune Checkpoint Proteins/genetics , Immune Checkpoint Proteins/metabolism , Immunophenotyping , Leukocytes, Mononuclear/immunology , Prognosis , Prospective Studies , SARS-CoV-2/immunology , Severity of Illness Index
9.
Cell Tissue Res ; 351(1): 85-98, 2013 Jan.
Article in English | MEDLINE | ID: mdl-23111773

ABSTRACT

Perturbations in microRNA (miRNA) expression profiles have been reported for cutaneous malignant melanoma (CMM) predominantly when examined in cell lines. Despite the rapidly growing number of newly discovered human miRNA sequences, the availability of up-to-date miRNA expression profiles for clinical samples of primary cutaneous malignant melanoma (PCMM), cutaneous malignant melanoma metastases (CMMM), and benign melanocytic nevi (BMN) is limited. Specimens excised from the center of tumors (lesional) from patients with PCMM (n=9), CMMM (n=4), or BMN (n=8) were obtained during surgery. An exploratory microarray analysis was performed by miRNA expression profiling based on Agilent platform screening for 1205 human miRNAs. The results from the microarray analysis were validated by TaqMan quantitative real-time polymerase chain reaction. In addition to several miRNAs previously known to be associated with CMM, 19 unidentified miRNA candidates were found to be dysregulated in CMM patient samples. Among the 19 novel miRNA candidates, the genes hsa-miR-22, hsa-miR-130b, hsa-miR-146b-5p, hsa-miR-223, hsa-miR-301a, hsa-miR-484, hsa-miR-663, hsa-miR-720, hsa-miR-1260, hsa-miR-1274a, hsa-miR-1274b, hsa-miR-3663-3p, hsa-miR-4281, and hsa-miR-4286 were upregulated, and the genes hsa-miR-24-1*, hsa-miR-26a, hsa-miR-4291, hsa-miR-4317, and hsa-miR-4324 were downregulated. The results of this study partially confirm previous CMM miRNA profiling studies identifying miRNAs that are dysregulated in CMM. However, we report several novel miRNA candidates in CMM tumors; these miRNA sequences require further validation and functional analysis to evaluate whether they play a role in the pathogenesis of CMM.


Subject(s)
Gene Expression Profiling , Melanoma/genetics , Melanoma/pathology , MicroRNAs/genetics , Nevus, Pigmented/genetics , Oligonucleotide Array Sequence Analysis , Skin Neoplasms/genetics , Adolescent , Aged , Aged, 80 and over , Child , Cluster Analysis , Data Mining , Female , Gene Expression Regulation, Neoplastic , Humans , Male , MicroRNAs/metabolism , Middle Aged , Nevus, Pigmented/pathology , Real-Time Polymerase Chain Reaction , Reproducibility of Results , Reverse Transcriptase Polymerase Chain Reaction , Skin Neoplasms/pathology , Young Adult , Melanoma, Cutaneous Malignant
10.
Cancers (Basel) ; 15(2)2023 Jan 16.
Article in English | MEDLINE | ID: mdl-36672496

ABSTRACT

Merkel cell carcinoma (MCC) is a rare, difficult-to-treat skin cancer once immunotherapy has failed. MCC is associated either with the clonal integration of the Merkel cell polyomavirus (MCPyV) or mutagenic UV-radiation. Fumaric acid esters, including dimethyl fumarate (DMF), have been shown to inhibit cell growth in cutaneous melanoma and lymphoma. We aimed to explore the effects of DMF on MCPyV-negative MCC cell lines. Three MCC cell lines (MCC13, MCC14.2, and MCC26) were treated with different doses of DMF. The cytotoxic effects and cell proliferation were assessed by the MTT cytotoxicity assay and BrdU proliferation assay at different time points. A significant reduction in cell viability and proliferation were demonstrated for all the cell lines used, with DMF proving to be effective.

11.
Mol Carcinog ; 51(11): 916-22, 2012 Nov.
Article in English | MEDLINE | ID: mdl-22025453

ABSTRACT

The microprocessor complex mediates intranuclear biogenesis of precursor microRNAs from the primary microRNA transcript. Extranuclear, mature microRNAs are incorporated into the RNA-induced silencing complex (RISC) before interaction with complementary target mRNA leads to transcriptional repression or cleavage. In this study, we investigated the expression profiles of the microprocessor complex subunit DiGeorge syndrome critical region gene 8 (DGCR8) and the RISC components argonaute-1 (AGO1), argonaute-2 (AGO2), as well as double-stranded RNA-binding proteins PACT, TARBP1, and TARBP2 in epithelial skin cancer and its premalignant stage. Patients with premalignant actinic keratoses (AK, n = 6), basal cell carcinomas (BCC, n = 15), and squamous cell carcinomas (SCC, n = 7) were included in the study. Punch biopsies were harvested from the center of the tumors (lesional), from healthy skin sites (intraindividual controls), and from healthy skin sites in a healthy control group (n = 16; interindividual control). The DGCR8, AGO1, AGO2, PACT, TARBP1, and TARBP2 mRNA expression levels were detected by quantitative real-time reverse transcriptase polymerase chain reaction. The DGCR8, AGO1, AGO2, PACT, and TARBP1 expression levels were significantly higher in the AK, BCC, and SCC groups than the healthy controls (P < 0.05). There was no significant difference in the TARBP2 expression levels between groups (P > 0.05). This study indicates that major components of the miRNA pathway, such as the microprocessor complex and RISC, are dysregulated in epithelial skin cancer.


Subject(s)
Argonaute Proteins/genetics , Carcinoma/genetics , Keratosis, Actinic/genetics , Nuclear Proteins/genetics , Proteins/genetics , RNA-Binding Proteins/genetics , Skin Neoplasms/genetics , Aged , Carcinoma, Basal Cell/genetics , Carcinoma, Squamous Cell/genetics , Female , Gene Expression Regulation, Neoplastic , Humans , Male , MicroRNAs/genetics , RNA-Induced Silencing Complex/genetics , Skin/metabolism
12.
Cell Tissue Res ; 350(1): 119-26, 2012 Oct.
Article in English | MEDLINE | ID: mdl-22706980

ABSTRACT

Although several studies have shown a dysregulation of microRNA (miRNA) expression profiles in cutaneous melanoma, there has been little research on the miRNA machinery itself. In this study, we investigated the mRNA expression profiles of different miRNA machinery components in primary cutaneous malignant melanoma (PCMM), cutaneous malignant melanoma metastases (CMMM) and benign melanocytic nevi (BMN). Patients with PCMM (n = 7), CMMM (n = 6) and BMN (n = 7) were included in the study. Punch biopsies were harvested from the centers of tumors (lesional) and from BMN (control). In contrast to previous reports exploring specific clusters of miRNAs in PCMM, the present study investigates mRNA expression levels of Dicer, Drosha, Exp5, DGCR8 and the RISC components PACT, argonaute-1, argonaute-2, TARBP1, TARBP2, MTDH and SND1, which were detected by TaqMan real-time reverse transcription polymerase chain reaction (RT-PCR). Argonaute-1, TARBP2 and SND1 expression levels were significantly higher in BMN compared to PCMM (p < 0.05). TARBP2 expression levels were significantly higher in CMMM compared to PCMM (p < 0.05). SND1 expression levels were significantly higher in CMMM compared to PCMM and BMN (p < 0.05). Dicer, Drosha, DGCR8, Exp5, argonaute-2, PACT, TARBP1 and MTDH expression levels showed no significant differences within groups (p > 0.05). The results of this study show that the miRNA machinery components argonaute-1, TARBP2 and SND1 are dysregulated in PCMM and CMMM compared to BMN and may play a role in the process of malignant transformation.


Subject(s)
Melanoma/genetics , Melanoma/pathology , MicroRNAs/metabolism , Nevus, Pigmented/genetics , Skin Neoplasms/genetics , Skin Neoplasms/pathology , Adolescent , Adult , Aged , Aged, 80 and over , Child , Female , Gene Expression Regulation, Neoplastic , Humans , Male , MicroRNAs/genetics , Middle Aged , Neoplasm Metastasis , Neoplasm Proteins/genetics , Neoplasm Proteins/metabolism , Nevus, Pigmented/pathology , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain Reaction , Melanoma, Cutaneous Malignant
13.
Eur J Dermatol ; 22(3): 319-23, 2012.
Article in English | MEDLINE | ID: mdl-22562806

ABSTRACT

There is increasing evidence that cytokines as well as chemokines are important players in the pathogenesis of lupus erythematosus (LE). We aimed to compare cytokine and chemokine profiles in different types of cutaneous LE. We investigated lesional mRNA and protein expression of various cytokines and chemokines in patients with chronic discoid LE (CDLE, n=15), subacute cutaneous LE (SCLE, n=11), and lupus erythematosus tumidus (LET, n=21). TNF-α, INF-γ, TGF-ß, IL-6, IL-10, IL-12p40, CXCL9, and CXCL10 mRNA expression were significantly increased in SCLE when compared to CDLE. Moreover, LET also showed significantly increased mRNA expression of TNF-α, TGF-ß, IL-10, IL-12p40 and CXCL9, as compared to CDLE. In all LE subtypes, CXCL9 and CXCL10 mRNA expression significantly correlated with INF-γ mRNA expression, as indicated by r-values ranging from 0.71 - 0.87. Immunohistochemistry for TNF-α, INF-γ, and IL-10 gave support to our RT-PCR results. In conclusion, our results suggest that T helper 1, as well as T helper 2 cytokines are differentially expressed in CDLE, SCLE, and LET. Compared to CDLE, the highest cytokine and chemokine ligand profiles are found in SCLE followed by LET. Our correlation studies also support the importance of an IFN-driven inflammation in cutaneous LE.


Subject(s)
Cytokines/physiology , Gene Expression Profiling , Lupus Erythematosus, Cutaneous/genetics , Adult , Aged , Aged, 80 and over , Chemokines/genetics , Chemokines/physiology , Chronic Disease , Cytokines/genetics , Humans , Immunohistochemistry , Interleukin-10/metabolism , Lupus Erythematosus, Cutaneous/physiopathology , Lupus Erythematosus, Discoid/genetics , Lupus Erythematosus, Discoid/metabolism , Male , Middle Aged , Prospective Studies , Real-Time Polymerase Chain Reaction , Tumor Necrosis Factor-alpha/metabolism
14.
J Am Acad Dermatol ; 65(1): 125-33, 2011 Jul.
Article in English | MEDLINE | ID: mdl-21353331

ABSTRACT

BACKGROUND: Antimicrobial peptides (AMPs) are small effector molecules of the innate immune system with well-known antimicrobial activity. Skin infections rarely occur in patients with cutaneous lupus erythematosus (CLE), and AMP expression in CLE has not been previously evaluated. OBJECTIVES: We aimed to determine the expression of several important AMPs in 3 different subtypes of CLE. METHODS: Skin lesions were analyzed for the gene and protein expression of human ß-defensin (hBD)-1, -2, and -3; RNase-7; the cathelicidin LL-37; and psoriasin (S100A7) using real-time reverse transcriptase polymerase chain reaction and immunohistochemistry. RESULTS: Skin biopsy specimens of 96 study participants including 47 patients with CLE (15 patients with discoid lupus erythematosus [LE], 11 patients with subacute CLE, and 21 patients with LE tumidus), 34 patients with psoriasis, and 15 healthy control subjects were evaluated in this study. HBD-2, hBD-3, LL-37, and psoriasin were significantly more highly expressed in CLE as compared with healthy controls, and most AMPs were significantly more highly induced in subacute CLE as compared with discoid LE and LE tumidus. AMP gene expression paralleled well with AMP protein expression in CLE and controls. Subacute CLE and discoid LE showed a similar correlation of AMP gene expression (significant correlations between hBD-1 and RNase-7, hBD-2 and hBD-3, hBD-2 and psoriasin, and hBD-3 and psoriasin). LIMITATIONS: The relatively small number of samples and the lack of analysis of the lesional bacterial colonization are a limitation. CONCLUSIONS: Several AMPs are increased in CLE at both gene and protein levels. This could explain the low prevalence of skin infections in CLE. It remains to be elucidated whether AMPs play a pathogenic role in CLE.


Subject(s)
Antimicrobial Cationic Peptides/genetics , Cathelicidins/genetics , Lupus Erythematosus, Cutaneous/genetics , S100 Proteins/genetics , Adult , Aged , Aged, 80 and over , Antimicrobial Cationic Peptides/immunology , Case-Control Studies , Female , Gene Expression Regulation , Humans , Lupus Erythematosus, Cutaneous/immunology , Lupus Erythematosus, Cutaneous/physiopathology , Male , Middle Aged , RNA/analysis , Reference Values , Reverse Transcriptase Polymerase Chain Reaction , S100 Calcium Binding Protein A7 , S100 Proteins/metabolism , Sensitivity and Specificity , Severity of Illness Index , Young Adult , beta-Defensins/genetics
16.
Cancers (Basel) ; 13(9)2021 Apr 26.
Article in English | MEDLINE | ID: mdl-33925915

ABSTRACT

To date, the skin remains the most common cancer site among Caucasians in the western world. The complex, layered structure of human skin harbors a heterogenous population of specialized cells. Each cell type residing in the skin potentially gives rise to a variety of cancers, including non-melanoma skin cancer, sarcoma, and cutaneous melanoma. Cutaneous melanoma is known to exacerbate and metastasize if not detected at an early stage, with mutant melanomas tending to acquire treatment resistance over time. The intricacy of melanoma thus necessitates diverse and patient-centered targeted treatment options. In addition to classical treatment through surgical intervention and radio- or chemotherapy, several systemic and intratumoral immunomodulators, pharmacological agents (e.g., targeted therapies), and oncolytic viruses are trialed or have been recently approved. Moreover, utilizing combinations of immune checkpoint blockade with targeted, oncolytic, or anti-angiogenic approaches for patients with advanced disease progression are promising approaches currently under pre-clinical and clinical investigation. In this review, we summarize the current 'state-of-the-art' as well as discuss emerging agents and regimens in cutaneous melanoma treatment.

17.
Curr Oncol ; 28(5): 3316-3322, 2021 08 27.
Article in English | MEDLINE | ID: mdl-34590599

ABSTRACT

There exist relatively sparse and conflicting data on high-level microsatellite instability (MSI-H) and deficient mismatch repair (dMMR) in cutaneous malignancies. We aimed to determine the expression profiles of MMR proteins (MSH2, MSH6, MLH1, and PMS2) in different progression stages of cutaneous squamous cell carcinoma (cSCC, 102 patients in total) by immunohistochemistry, and search for MSI-H in patients with low-level MMR or dMMR using multiplex-PCR. Low-level MMR protein expression was observed in five patients: One patient with primary cSCC < 2 mm thickness and low-level MLH1, three patients with primary cSCC > 6 mm (including one with low-level MSH2, as well as MSH6 expression, and two with low-level PMS2), and one patient with a cSCC metastasis showing low-level MSH2 as well as MSH6. Intergroup protein expression analysis revealed that MLH1 and MSH2 expression in actinic keratosis was significantly decreased when compared to Bowen's disease, cSCC < 2 mm, cSCC > 6 mm, and cSCC metastasis. In cases with low-level MMR, we performed MSI-H tests revealing three cases with MSI-H and one with low-level MSI-L. We found low-level MMR expression in a small subset of patients with invasive or metastatic cSCC. Hence, loss of MMR expression may be associated with tumour progression in a small subgroup of patients with non-melanoma skin cancer.


Subject(s)
Carcinoma, Squamous Cell , Skin Neoplasms , Carcinoma, Squamous Cell/genetics , DNA Mismatch Repair/genetics , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Humans , Microsatellite Instability , Mismatch Repair Endonuclease PMS2/genetics , Mismatch Repair Endonuclease PMS2/metabolism , MutL Protein Homolog 1/genetics , MutL Protein Homolog 1/metabolism , MutS Homolog 2 Protein/genetics , MutS Homolog 2 Protein/metabolism , Skin Neoplasms/genetics
18.
J Dermatol ; 48(1): 64-74, 2021 Jan.
Article in English | MEDLINE | ID: mdl-33180347

ABSTRACT

The relevance of Hedgehog signaling in Merkel cell carcinoma has only been addressed by a few studies with conflicting results. Thus, we aimed to establish the expression of Hedgehog signaling molecules in Merkel cell carcinoma to characterize causes of aberrant expression and to correlate these findings with the clinical course of the patients. Immunohistochemistry was performed for Sonic, Indian, Patched 1 (PTCH1) and Smoothened on patients' tumor tissue. Respective mRNA expression was analyzed in 10 Merkel cell carcinoma cell lines using quantitative real-time polymerase chain reaction. PTCH1 sequencing and DNA methylation microarray analyses were carried out on tumor tissues as well as cell lines. PTCH1 immunoreactivity in Merkel cell carcinoma was similar to that of basal cell carcinomas, which both significantly differed from PTCH1 immunoreactivity in healthy skin. Most PTCH1 mutations found were synonymous or without known functional impact. However, on average, the promoter regions of both PTCH1 were hypomethylated independently from PTCH1 gene expression or Merkel cell polyomavirus status. PTCH1 and GLI1/2/3 genes were differently expressed in different cell lines; notably, there was a significant correlation between GLI2 and PTCH1 mRNA expression. Similar to PTCH1 protein expression in patient tissues, PTCH1 gene expression in Merkel cell carcinoma cell lines is highly variable, but due to the similar methylation pattern across Merkel cell carcinoma cell lines, effects other than methylation seem to be the reason for the differential expression and PTCH1 appears to be upregulated by GLI as a classical Hedgehog target gene.


Subject(s)
Carcinoma, Merkel Cell , Patched-1 Receptor/genetics , Skin Neoplasms , Carcinoma, Merkel Cell/genetics , Hedgehog Proteins , Humans , Skin Neoplasms/genetics , Transcription Factors , Zinc Finger Protein GLI1
19.
Cancers (Basel) ; 13(11)2021 May 21.
Article in English | MEDLINE | ID: mdl-34063983

ABSTRACT

We aimed to assess for the first time the mismatch repair (MMR) protein expression in Merkel cell carcinoma (MCC). Immunohistochemistry was performed for MLH1, MSH2, MSH6, and PMS2 on patients' tumor tissue (n = 56), including neighbored healthy control tissue. In cases with low-level MMR expression (<10th percentile), we performed multiplex PCR in combination with high-resolution capillary electrophoresis in order to confirm microsatellite instability (MSI). Microscopic evaluation revealed a high median expression for all MMR proteins studied (91.6-96.3%). However, six patients (56/10.7%) had low-level MLH1 expression, six (55/10.9%) had low-level MSH2 expression, five (56/8.9%) had low-level MSH6 expression, and six (54/11.1%) had low-level PMS2 expression. Together, we observed nine (56/16.1%) patients who had low-level MMR expression of at least one protein. Of the patients with low-level MMR expression, MSI evaluation was possible in five cases, revealing one case with high-level MSI. In all MMR proteins assessed, low-level expression was significantly (p = 0.0004 to p < 0.0001) associated with a negative Merkel cell polyomavirus (MCPyV) status. However, the expression profiles of the MMR proteins did not correlate with clinical outcome measures such as disease relapse or death (p > 0.05). MCC appears to be a malignancy characterized by low-level MMR rather than completely deficient MMR in a subset of cases, predominantly affecting MCPyV-negative tumors. Future studies will establish whether this subset of MCC patients respond better to immune checkpoint inhibitor therapy.

20.
Int J Cardiol ; 344: 160-169, 2021 Dec 01.
Article in English | MEDLINE | ID: mdl-34517018

ABSTRACT

Hypertrophic cardiomyopathy (HCM) is a complex myocardial disorder with no well-established disease-modifying therapy so far. Our study aimed to investigate how autophagy, oxidative stress, inflammation, stress signalling pathways, and apoptosis are hallmark of HCM and their contribution to the cardiac dysfunction. Demembranated cardiomyocytes from patients with HCM display increased titin-based stiffness (Fpassive), which was corrected upon antioxidant treatment. Titin as a main determinant of Fpassive was S-glutathionylated and highly ubiquitinated in HCM patients. This was associated with a shift in the balance of reduced and oxidized forms of glutathione (GSH and GSSG, respectively). Both heat shock proteins (HSP27 and α-ß crystalline) were upregulated and S-glutathionylated in HCM. Administration of HSPs in vitro significantly reduced HCM cardiomyocyte stiffness. High levels of the phosphorylated monomeric superoxide anion-generating endothelial nitric oxide synthase (eNOS), decreased nitric oxide (NO) bioavailability, decreased soluble guanylyl cyclase (sGC) activity, and high levels of 3-nitrotyrosine were observed in HCM. Many regulators of signal transduction pathways that are involved in autophagy, apoptosis, cardiac contractility, and growth including the mitogen-activated protein kinase (MAPK), protein kinase B (AKT), glycogen synthase kinase 3ß (GSK-3ß), mammalian target of rapamycin (mTOR), forkhead box O transcription factor (FOXO), c-Jun N-terminal protein kinase (JNK), and extracellular-signal-regulated kinase (ERK1/2) were modified in HCM. The apoptotic factors cathepsin, procaspase 3, procaspase 9 and caspase 12, but not caspase 9, were elevated in HCM hearts and associated with increased proinflammatory cytokines (Interleukin 6 (IL-6), interleukin 18 (IL-18), intercellular cell adhesion molecule-1 (ICAM1), vascular cell adhesion molecule-1 (VCAM1), the Toll-like receptors 2 (TLR2) and the Toll-like receptors 4 (TLR4)) and oxidative stress (3-nitrotyrosine and hydrogen peroxide (H2O2)). Here we reveal stress signalling and impaired PQS as potential mechanisms underlying the HCM phenotype. Our data suggest that reducing oxidative stress can be a viable therapeutic approach to attenuating the severity of cardiac dysfunction in heart failure and potentially in HCM and prevent its progression.


Subject(s)
Cardiomyopathy, Hypertrophic , Hydrogen Peroxide , Apoptosis , Extracellular Signal-Regulated MAP Kinases , Humans , Oxidative Stress
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