Arsenic Trioxide Decreases Lymphangiogenesis by Inducing Apoptotic Pathways and Inhibition of Important Endothelial Cell Receptors.

Tumor-induced lymphangiogenesis is strongly associated with the formation of tumor metastasis. Therefore, the regulation of lymphangiogenesis offers a promising target in cancer therapy. Arsenic trioxide (ATO) is highly effective in the treatment of patients with acute promyelocytic leukemia (APL). As ATO mediates anti-angiogenic effects on endothelial and tumor cells, we aimed to explore the impact of ATO on lymphangiogenesis in human lymphatic endothelial cells (LEC). The BrdU assay and flow cytometry analysis were used to evaluate the influence of ATO on the proliferation and cell cycle distribution of LECs. The lymphatic suppression effects of ATO were investigated in vitro using the lymphatic tube formation assay. The effects of ATO on apoptosis, mitochondrial membrane potential and endothelial cell receptors were investigated by Western blotting, ELISA, flow cytometry and qRT-PCR. The treatment of LECs with ATO attenuated cell proliferation, blocked tube formation and induced subG0/G1 arrest in LECs, thus suggesting enhanced apoptosis. Although subG0/G1 arrest was accompanied by the upregulation of p21 and p53, ATO treatment did not lead to visible cell cycle arrest in LECs. In addition, ATO caused apoptosis via the release of cytochrome c from mitochondria, activating caspases 3, 8 and 9; downregulating the anti-apoptotic proteins survivin, XIAP and cIAP-2; and upregulating the pro-apoptotic protein Fas. Furthermore, we observed that ATO inhibited the VEGF-induced proliferation of LECs, indicating that pro-survival VEGF/VEGFR signaling was affected by ATO treatment. Finally, we found that ATO inhibited the expression of the important endothelial cell receptors VEGFR-2, VEGFR-3, Tie-2 and Lyve-1. In conclusion, we demonstrate that ATO inhibits lymphangiogenesis by activating apoptotic pathways and inhibiting important endothelial cell receptors, which suggests that this drug should be further evaluated in the treatment of tumor-associated lymphangiogenesis.

Evaluation of 2-methoxyestradiol serum levels as a potential prognostic marker in malignant melanoma.

Experimental findings indicated that 2-methoxyestradiol (2-ME), an endogenous metabolite of 17ß-estradiol, may exhibit anti-tumorigenic properties in various types of tumour, such as melanoma and endometrial carcinoma. In patients with endometrial cancer, the serum levels of 2-ME are decreased compared with those in healthy controls, and this finding has been associated with a poor outcome. The aim of the present study was to examine whether the serum levels of 2-ME are decreased in patients with melanoma, and whether this decrease may be correlated with disease stage and, therefore, serve as a prognostic indicator. ELISA was used to detect serum levels of 2-ME in patients with stage I-IV malignant melanoma (MM). A cohort of 78 patients with MM was analysed, along with 25 healthy controls, among whom 15 were women in the second trimester of pregnancy (positive control). As expected, significantly elevated levels of serum 2-ME were observed in pregnant control patients compared with those in patients with MM and healthy controls. There was no observed correlation between 2-ME serum levels in patients with MM and disease stage, tumour thickness, lactate dehydrogenase or S100 calcium-binding protein B levels. In addition, the 2-ME levels of patients with MM did not differ significantly from those of normal healthy controls. Overall, the findings of the present study indicated that the 2-ME serum levels in patients with MM were not decreased, and there was no correlation with early- or advanced-stage disease. Therefore, in contrast to published results on endometrial cancer, endogenous serum 2-ME levels in MM were not found to be correlated with tumour stage and did not appear to be a suitable prognostic factor in MM.

Assessment of Melanogenesis in a Pigmented Human Tissue-Cultured Skin Equivalent.

BACKGROUND: Organotypic tissue-cultured skin equivalents are used for a broad range of applications either as possible substitute for animal tests or for transplantation in patient-centered care. AIMS: In this study, we implemented melanocytes in a tissue-cultured full-thickness skin equivalent, consisting of epidermis and dermis. The versatility of this skin-like model with respect to pigmentation and morphological criteria was tested. MATERIALS AND METHODS: Pigmented skin equivalents were morphologically characterized, and melanogenesis was evaluated after treatment with kojic acid - a tyrosinase inhibitor and forskolin - a well-known activator of the cyclic adenosine 3,5-monophosphate pathway. Pigmentation was measured either by determination of the extinction at 400 nm after melanin extraction with KOH correlated to a melanin standard curve or by reflectance colorimetric analysis, monitoring reflectance of 660 nm and 880 nm emitting diodes. RESULTS: The morphological analysis revealed characteristic epidermal stratification with melanocytes located at the basal layer. Stimulation with forskolin increased the pigmentation, whereas treatment with kojic acid caused bleaching. CONCLUSION: The present study demonstrates that the herein-introduced organotypic tissue-cultured skin equivalent is comparable to the normal human skin and its versatility in tests regarding skin pigmentation. Therefore, this model might help understand diseases with dysfunctional pigmentation such as melasma, vitiligo, and postinflammatory hyperpigmentation.

Normal and keloid fibroblasts are differentially influenced by IFN-γ and triamcinolone as well as by their combination.

Impaired wound healing as well as imbalanced cell proliferation and extracellular matrix synthesis and degeneration can cause aberrant scarring. The most severe impacts of such scarring on patients' lives are stigmatization and physical restriction. Although, a broad variety of combinatorial approaches with, e.g., glucocorticoids, chemotherapeutics, and immunomodulators are used, there is still a high recurrence rate of keloids. The aim of this study was to investigate which influence interferon γ (IFN-γ, 1.000-10.000 IU/mL) and/or triamcinolone acetonide (TA, 1 µg/mL) have on proliferation, cell viability, collagen type I synthesis, and cytokine secretion in healthy and keloid fibroblasts. It was shown that mono-treatment with IFN-γ or TA for 2 days induced a severe reduction of the proliferative potential in both cell species. The combinatory treatment (IFN-γ plus TA) of keloid fibroblasts enhanced the anti-proliferative effect of the mono-treatments, whereas no additional anti-proliferative effect was observed in normal fibroblasts. Furthermore, we observed that the combinatory treatment regimen reduced the expression of α-smooth muscle actin (α-SMA), an actin isotype contributing to cell-generated mechanical tension, in keloid fibroblasts. In normal fibroblasts, α-SMA was reduced by the mono-treatment with IFN-γ as well as by the combinatory treatment. The analysis of collagen-type I synthesis revealed that TA did not reduce collagen type I synthesis in normal fibroblasts but in keloid fibroblasts. IFN-γ reduced in both cell species the collagen type I synthesis. The combination of TA and IFN-γ intensified the previously observed collagen type I synthesis reduction in keloid fibroblasts. The herein presented data suggest the combinatory application of IFN-γ and TA as a promising therapy concept for keloids.

Fatty acid receptor GPR120: a novel marker for human melanoma.

The correlation between ultraviolet radiation of the skin and melanoma incidence in humans is well established. Interestingly, epidemiologic data suggest also a correlation to an increased BMI pointing to metabolic trigger factors in melanoma pathogenesis. To substantiate this connection, we studied the expression of G-protein-coupled receptor 120 (GPR120), a receptor sensitive to unsaturated long-chain free fatty acids in melanoma tissues. One-hundred fourteen tissue sections histologically confirmed as nevi (n=32), primary melanoma (n=39), and melanoma metastasis (n=43) were immunohistochemically stained against GPR120. The staining was evaluated by three trained dermatopathologists and independently scored. Compared with nevi, primary melanoma and melanoma metastasis showed significantly higher levels of GPR120 staining. Only three out of 32 nevi showed strong GPR120 expression [median immunoreactivity-scoring system (IRS) score: 1, range: 0-10], whereas in primary melanomas 14 out of 39 were highly GPR120-positive (median IRS score: 7, range: 0-12) and in melanoma metastasis 27 out of 43 were highly GPR120-positive (median IRS score: 9, range: 0-12). GPR120 expression and tumor thickness (mm) show a statistically significant correlation in primary melanoma (P=0.011). Moreover, GPR120-positive staining was found throughout the epidermis and in sebaceous and sweat glands, which is yet not described. This study identified GPR120 as a novel marker for melanoma, indicating that melanoma cells are sensitive to free fatty acids. It is tempting to speculate that pharmacologically interfering with GPR120 signaling might improve melanoma therapy.

Histone deacetylase inhibitors interfere with angiogenesis by decreasing endothelial VEGFR-2 protein half-life in part via a VE-cadherin-dependent mechanism.

Recent evidence suggests that histone deacetylase inhibitors (HDACi) may mediate part of their antitumor effects by interfering with tumor angiogenesis. As signalling via the vascular endothelial growth factor receptor-2 (VEGFR-2) pathway is critical for angiogenic responses during tumor progression, we explored whether established antitumor effects of HDACi are partly mediated through diminished endothelial VEGFR-2 expression. We therefore examined the potential impact of three different HDACi, trichostatin A (TSA), sodium butyrate (But) and valproic acid (VPA), on VEGFR-2 protein expression. TSA, VPA and But significantly inhibit VEGFR-2 protein expression in endothelial cells. Pertinent to these data, VEGFR-2 protein half-life is shown to be decreased in response to HDACi. Recently, it could be demonstrated that expression of VE-cadherin influences VEGFR-2 protein half-life. In our experiments, VEGFR-2 downregulation was accompanied by HDACi-induced VE-cadherin suppression. Interestingly, siRNA-mediated knockdown of VE-cadherin led to a pronounced loss of VEGFR-2 expression on the protein as well as on the mRNA level, implicating that VE-cadherin not only influences VEGFR-2 protein half-life but also the transcriptional level. To further distinguish which of the eight different histone deacetylases are responsible for the regulation of VEGFR-2 expression, specific HDAC genes were silenced by transfecting respective siRNAs. These studies revealed that HDACs 1, 4, 5 and 6 are preferentially involved in VEGFR-2 expression. Therefore, these results provide an explanation for the anti-angiogenic action of HDAC inhibitors via a VE-cadherin, HDAC 1 and HDACs 4-6-mediated suppression of VEGFR-2 expression and might be of importance in the development of new anti-angiogenic drugs.

The histone deacetylase inhibitor trichostatin a decreases lymphangiogenesis by inducing apoptosis and cell cycle arrest via p21-dependent pathways.

BACKGROUND: The formation of new lymphatic vessels provides an additional route for tumour cells to metastasize. Therefore, inhibiting lymphangiogenesis represents an interesting target in cancer therapy. First evidence suggests that histone deacetylase inhibitors (HDACi) may mediate part of their antitumor effects by interfering with lymphangiogenesis. However, the underlying mechanisms of HDACi induced anti-lymphangiogenic properties are not fully investigated so far and in part remain unknown. METHODS: Human lymphatic endothelial cells (LEC) were cultured in vitro and treated with or without HDACi. Effects of HDACi on proliferation and cell cycle progress were analysed by BrdU assay and flow cytometry. Apoptosis was measured by quantifying mono- and oligonucleosomes in the cytoplasmic fraction of cell lysates. In vitro lymphangiogenesis was investigated using the Matrigel short term lymphangiogenesis assay. The effects of TSA on cell cycle regulatory proteins and apoptosis-related proteins were examined by western blotting, immunofluorescence staining and semi-quantitative RT-PCR. Protein- and mRNA half-life of p21 were analysed by western blotting and quantitative RT-PCR. The activity of the p21 promoter was determined using a dual luciferase assay and DNA-binding activity of Sp1/3 was investigated using EMSA. Furthermore, siRNA assays were performed to analyse the role of p21 and p53 on TSA-mediated anti-lymphangiogenic effects. RESULTS: We found that HDACi inhibited cell proliferation and that the pan-HDACi TSA induced G0/G1 arrest in LEC. Cell cycle arrest was accompanied by up-regulation of p21, p27 and p53. Additionally, we observed that p21 protein accumulated in cellular nuclei after treatment with TSA. Moreover, we found that p21 mRNA was significantly up-regulated by TSA, while the protein and mRNA half-life remained largely unaffected. The promoter activity of p21 was enhanced by TSA indicating a transcriptional mechanism. Subsequent EMSA analyses showed increased constitutive Sp1/3-dependent DNA binding in response to HDACi. We demonstrated that p53 was not required for TSA induced p21 expression and growth inhibition of LECs. Interestingly, siRNA-mediated p21 depletion almost completely reversed the anti-proliferative effects of TSA in LEC. In addition, TSA induced apoptosis by cytochrome c release contributed to activating caspases-9, -7 and -3 and downregulating the anti-apoptotic proteins cIAP-1 and -2. CONCLUSIONS: In conclusion, we demonstrate that TSA - a pan-HDACi - has distinct anti-lymphangiogenic effects in primary human lymphatic endothelial cells by activating intrinsic apoptotic pathway and cell cycle arrest via p21-dependent pathways.

Peroxisome proliferator-activated receptor (PPAR) α and δ activators induce ICAM-1 expression in quiescent non stimulated endothelial cells.

BACKGROUND: Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors that are implicated in the regulation of lipid and glucose homeostasis. PPAR agonists have been shown to control inflammatory processes, in part by inhibiting the expression of distinct proinflammatory genes such as vascular cell adhesion molecule-1 (VCAM-1), IL-8, and intercellular adhesion molecule-1 (ICAM-1). ICAM-1 is an important endothelial membrane receptor that facilitates the transmigration of leukocytes across the endothelium. To date, the influence of PPARα and δ activators on the expression of ICAM-1 in non-induced, quiescent endothelial cells has been unclear. Therefore, we examined the effects of various PPARα and δ agonists on the expression of ICAM-1 in non-stimulated primary human endothelial cells. RESULTS: We found that PPARα and PPARδ agonists significantly induced ICAM-1 surface, intracellular protein, and mRNA expression in a time and concentration-dependent manner. The PPARδ induced ICAM-1 expression could be paralleled with a significantly increased T-cell adherence to the endothelial cells whereas PPARα failed to do so. Transcriptional activity studies using an ICAM-1 reporter gene constructs revealed that PPARδ, but not PPARα agonists induced gene expression by stimulating ICAM-1 promoter activity via an Sp1 transcription factor binding site and inhibit the binding of the transcription factors Sp1 and Sp3. Furthermore, we performed mRNA stability assays and found that PPARα and PPARδ agonists increased ICAM-1 mRNA stability. CONCLUSION: Therefore, our data provide the first evidence that PPARα and PPARδ agonists induce ICAM-1 expression in non-stimulated endothelial cells via transcriptional and posttranscriptional mechanisms.

Dimethylfumarate inhibits melanoma cell proliferation via p21 and p53 induction and bcl-2 and cyclin B1 downregulation.

Recent evidence suggests that dimethylfumarate (DMF), known as a highly potent anti-psoriatic agent, might have anti-tumorigenic properties in melanoma. It has recently been demonstrated that DMF inhibits melanoma proliferation by apoptosis and cell cycle inhibition and therefore inhibits melanoma metastasis. Nonetheless, the underlying mechanisms remain to be evaluated. To elucidate the effects of DMF on melanoma cell lines (A375, SK-Mel), we first performed cytotoxicity assays. No significant lactatedehydogenase (LDH) release could be found. In further analysis, we showed that DMF suppresses melanoma cell proliferation in a concentration-dependent manner. To examine whether these effects are conveyed by apoptotic mechanisms, we studied the amount of apoptotic nucleosomes and caspase 3/7 activity using ELISA analysis. Significant apoptosis was induced by DMF in both cell lines, and this could be paralleled with bcl-2 downregulation and PARP-1 cleavage. We also performed cell cycle analysis and found that DMF induced concentration-dependent arrests of G0/G1 as well as G2/M. To examine the underlying mechanisms of cell cycle arrest, we analyzed the expression profiles of important cell cycle regulator proteins such as p53, p21, cyclins A, B1, and D1, and CDKs 3, 4, and 6. Interestingly, DMF induced p53 and p21 yet inhibited cyclin B1 expression in a concentration-dependent manner. Other cell cycle regulators were not influenced by DMF. The knockdown of DMF induced p53 via siRNA led to significantly reduced apoptosis but had no influence on cell cycle arrest. We examined the adhesion of melanoma cells on lymphendothelial cells during DMF treatment and found a significant reduction in interaction. These data provide evidence that DMF inhibits melanoma proliferation by reinduction of important cell cycle inhibitors leading to a concentration-dependent G0/G1 or G2/M cell cycle arrest and induction of apoptosis via downregulation of bcl-2 and induction of p53 and PARP-1 cleavage. Hence, DMF might be an interesting agent in the treatment of melanoma and is worth further investigation in vivo.

Cutaneous involvement as a rare first sign of systemic mantle cell lymphoma: A case report and review of the literature.

Mantle cell lymphoma (MCL) is a unique type of B-cell non-Hodgkin's lymphoma, which very rarely exhibits skin involvement. We herein describe the case of a 55-year-old woman, who initially presented with a nodular mass of the right infraorbital region. On histological analysis of the subcutaneous tissue, a diffuse neoplastic cell infiltration was identified, composed of medium-sized lymphoid cells with irregular nuclei, which was diagnosed as MCL. The tumor cells were positive for CD5, CD20, CD79a, cyclin D1 and sex-determining region Y-box 11, but negative for CD10 and CD23. Our patient received six cycles of R-CHOP chemotherapy and intrathecal methotrexate as central nervous system prophylaxis. However, the patient relapsed 1 year later and was treated with two cycles of R-DHAP and one cycle of intrathecal methotrexate. After achieving partial remission, the patient was consolidated with peripheral blood stem cell transplantation using the BEAM conditioning regime. While prior case studies suggest that skin invasion by MCL is associated with a poor prognosis, our patient remains alive almost 4 years after the initial presentation. Skin involvement as a first sign of systemic MCL is very rare and must be considered.

Dimethylfumarate effectively inhibits lymphangiogenesis via p21 induction and G1 cell cycle arrest.

Different pathologies, such as lymphoedema, cancer or psoriasis, are associated with abnormal lymphatic vessel formation. Therefore, influencing lymphangiogenesis is an interesting target. Recent evidence suggests that dimethylfumarate (DMF), an antipsoriatic agent, might have antitumorigenic and antilymphangiogenic properties. To prove this assumption, we performed proliferation and functional assays with primary human dermal lymphendothelial cells (DLEC). We could demonstrated that DMF suppresses DLEC proliferation and formation of capillary-like structures. Underlying apoptotic mechanisms could be ruled out. Cell cycle analysis demonstrated a pronounced G1-arrest. Further evaluations revealed increases in p21 expression. In addition, DMF suppressed Cyclin D1 and Cyclin A expression in a concentration-dependent manner. p21 knockdown experiments demonstrated a p21-dependent mechanism of regulation. Further analysis showed an increased p21 mRNA expression after DMF treatment. This transcriptional regulation was enforced by post-transcriptional and post-translational mechanisms. In addition, we could demonstrate that the combination of a proteasomal inhibitor and DMF superinduced the p21 expression. Hence, DMF is a new antilymphangiogenic compound and might be used in various illnesses associated with increased lymphangiogenesis.

Dimethylfumarate protects against TNF-α-induced secretion of inflammatory cytokines in human endothelial cells.

BACKGROUND: Inflammation, angiogenesis and oxidative stress have been implicated in the pathogenesis of various vascular diseases. Recent evidence suggests that dimethylfumarate (DMF), an antiposriatic and anti-multiple sclerosis agent, possesses anti-inflammatory, anti-oxidative and anti-angiogenic properties. Here, we analyze the influence of DMF on TNF-α-induced expression of the important pro-inflammatory and pro-atherogenic chemokine MCP-1 and investigate the underlying mechanisms of this expression. FINDINGS: We analyzed constitutive and TNF-α-induced expression of MCP-1 in human umbilical vascular endothelial cells (HUVEC) +/- DMF treatment via enzyme-linkes immunosorbent assay (ELISA). DMF significantly inhibited the protein expression levels in a time- and concentration-dependent manner. Furthermore, MCP-1 mRNA expression was also reduced in response to DMF, as demonstrated by RT-PCR. Thus, the regulation occurs at the transcriptional level. Interestingly, DMF prolonged the TNF-α-induced p38 and JNK phosphorylation in HUVEC, as demonstrated by Western blot analysis; however, the p38 and JNK inhibitor SB203580 did not affect the DMF-conveyed suppression of TNF-α-induced MCP-1 expression. DMF suppressed the TNF-α-induced nuclear translocation and phosphorylation (Serine 536) of p65 in these cells. These results were additionally approved by p65 luciferase promoter assays. Furthermore, we found that DMF slightly inhibited the early degradation of IκBα. In addition, we verified our results using other important inflammatory cytokines such as CCL-5, PDGF-BB, GM-CSF and IL-6. CONCLUSION: DMF suppresses various TNF-α-induced pro-inflammatory and pro-atherogenic cytokines/chemokines in human endothelial cells. This action is regulated by reduced p65 activity and nuclear translocation, which can be explained in part by the reduced early degradation of IκBα and more important the reduced phosphorylation of p65 at Serine 536. These effects were independent of the p38, PI3K and p42/44 signaling pathways. As a result, DMF might be suitable for treating patients with vascular diseases.

The Infraorbital Artery in Fetuses: Clinical Relevance in Perforator Flap Surgery.

INTRODUCTION: The reconstruction of soft tissue detects in mid facial region are highly demanding. Most challenging region are nasal alla. For full thickness nasal alla defects most authors use nasolabial flap based on facial/angular arcade, but for recidivans tumors the infraorbital perforator flap is a good solution. AIM: The aim of our research was to analyze the number and the course of the infraorbital artery terminal branches. MATERIAL AND METHODS: Material was 60 fetal hemifacial specimens of different gestational ages. Fetuses were fixed in 10% formalin and arterial blood vessels were injected with Micropaque solution (barium sulfate). Samples were further processed by Spalteholz technique, their images captured with digital camera and analyzed. Infraorbital artery was constant artery and had 2 to 4 terminal branches supplying infraorbital region. The majority of its terminal branches were characterized with descending course. Reach anatomical network of infraorbital artery made anastomoses with facial artery. CONCLUSION: Perforator flap based on infraorbital artery had well defined vascular supply with numerous soft tissue branches, which qualify this flap as safe solution for nasal reconstruction.

Repeated administration of inhibitors for ion pumps reduce markedly tumor growth in vivo.

INTRODUCTION: Measurements of extracellular pH show that the micro environment of malignant tumors is more acidic than that of normal cells, whereas pH does not differ appreciable in normal and malignant cells. The acid micro environment of tumors is created by the secretion of tumor factors and ATP hydrolysis in hypoxic tumor tissue. In order to survive in a low pH-environment tumor cells develop regulatory mechanisms which keep their intracellular pH stable. Two of the most important systems are the Na+/H+ ion pump and the Na-dependent HCO3-/Cl- pump of stilbenian derivatives. MATERIAL AND METHODS: Experiments were carried out on DBA mice of both sexes at the age of 4 month. Laboratory animals were grown in our institute and supplied with food and aqua ad libitum. RESULTS: After termination of the experiments the mean tumor diameter in the control group was 12.4 +/- 0.8mm, in group A it was 6.9 +/- 0.6mm, and in group B we measured 6.6 +/- 3.1mm. At the final day the tumor size in treated animals was twice as small as in the control group. In addition we observed the rate of survival. In the control group only 18% of the animals were still alive at day 18. Considering the rate of survival a statistically significant difference between treated and untreated animals was observed. The survival of tumor cells is dependent on the function of these ion pumps which keep their intracellular pH values constant in the setting of an acid extracellular environment. CONCLUSION: The activity of the ion pump is especially important at the beginning of cell division and in cell proliferation. Our in vivo experiments demonstrate that prolonged administration of intratumoral ion pump inhibitors suppresses tumor growth as well as enhances survival of tumor-bearing animals. Research of inhibitors of ion pumps and their action in tumor growth opens new perspectives into pathophysiology of malignant tumors and may create new therapeutic options.

Single stage turn in perforator infraorbital artery island flap for nasal ala reconstruction.

BACKGROUND: The reconstruction of fullthickness nasal alla defects is challenging procedure. Use of local flaps is acceptable approach. Flap based on infraorbital artery could be used for primary reconstruction of nasal ala defects. METHODS: The prospective study include consecutive series of 15 patients with advanced skin carcinoma of the nasal ala and medial cheek staged T4 by TNM, in whom the turn in infraorbital flap was used. The patient characteristics, type of carcinoma and complications were analyzed. RESULTS: The turn in infraorbital flap was used mostly in male patients (80%), mean age 64 years. The basal cell skin carcinoma was found in 60%. Skin layer was skin grafted. All flaps survived, but in one case a partial wound dehiscence in one partial skin graft loss was found, and in two patients partial nasal obstruction occurred. These three complications were solved as secondary procedures under local anesthesia. CONCLUSION: Full-thickness defect of the nasal ala can be properly reconstructed using flap based on infraorbital artery providing exceptional esthetic and functional results, as single stage procedure.

Ligand activation of peroxisome proliferator-activated receptor delta suppresses cathepsin B expression in human endothelial cells in a posttranslational manner.

Peroxisome proliferator-activated receptor (PPAR) delta agonists are known to have distinct anti-inflammatory and antitumor effects; though, the knowledge regarding their mode of action has thus far been limited. Different cathepsins have been shown to be upregulated in a broad range of pathological events, such as rheumatoid arthritis, psoriasis, atherosclerosis and diverse tumor entities, for example melanoma. Recent work demonstrated that cathepsin B in particular is an important pro-angiogenic protease in various pathological conditions. We therefore analysed whether cathepsins are a valid target for PPARδ agonists. This study reveals an inhibitory effect of two commonly used PPARδ agonists, GW501516 and L-165,041, on the protein expression and enzyme activity of cathepsin B in human endothelial cells. In contrast, no inhibitory effects were observed on cathepsin L and cathepsin D protein expression after treatment with PPARδ agonists. Furthermore, the results substantiate that PPARδ activators mediate their inhibitory action in a PPARδ-dependent manner and that the underlying regulatory mechanism is not based on a transcriptional but rather on a posttranslational mode of action, via the reduction in the cathepsin B protein half-life. Mechanisms conveying the suppressive effect by 5'-alternative splicing, a 3'-UTR-dependent way or by miRNA could be excluded. The data of this study explore cathepsin B as a new valid target for PPARδ agonists in endothelial cells. The results bolster other studies demonstrating PPARδ agonists as anti-inflammatory and anticarcinogenic agents and thus might have the potential to help to develop new pharmaceutical drugs.

AP1-dependent repression of TGFα-mediated MMP9 upregulation by PPARδ agonists in keratinocytes.

Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors that function mainly in the regulation of glucose and lipid homeostasis. PPAR agonists have been shown to control inflammation by inhibition of distinct proinflammatory genes. Aberrant activation of the epidermal growth factor receptor and/or overexpression of its ligand, transforming growth factor-α (TGFα), are key features of both neoplastic and inflammatory hyperproliferative epithelia. Matrix metalloproteinase 9 (MMP9) belongs to the set of genes that are effectively induced by TGFα in keratinocytes. Induction of MMP9 expression is strongly linked to regenerative skin repair mechanisms, inflammatory skin diseases and tumor metastasis. We explored whether the known anti-inflammatory effects of PPARδ ligands involve inhibiting the TGFα-mediated upregulation of MMP9. The PPARδ agonists potently inhibited TGFα-induced MMP9 expression in human keratinocytes. This inhibition was observed at both the protein and mRNA levels. Transcriptional activation studies with deletion constructs of a reporter gene revealed that PPARδ agonists mediate their inhibitory effects via an AP1-binding site. Electromobility shift assay analysis indicated that MMP9 gene expression is inhibited by repressing site-dependent DNA binding and transactivation by c-fos. In conclusion, our data provide the first evidence that MMP9 expression induced by TGFα is a valid target of PPARδ ligands in keratinocytes.