The role of extracellular matrix macromolecules in cancer and diabetic macroangiopathy – with special reference to decorin and hyaluronan

The central role of extracellular matrix (ECM) macromolecules in diseases such as cancer and atherosclerotic vascular diseases including diabetic macroangiopathy is indisputable. Decorin and hyaluronan (HA) represent vital ECM macromolecules in the microenvironment of cells and are centrally involved in human cancer and cardiovascular biology. In cancer, decorin is considered to play a tumor suppressive role. However, there is some discrepancy whether malignant cells express it. Regarding HA, its contribution to the development of atherosclerotic vascular diseases has been well established. Nevertheless, the precise role of HA in arterial narrowing associated with diabetes is not known. The present study focused on two vital ECM macromolecules, namely decorin and HA. First, decorin expression was studied in human tumorigenesis. Furthermore, the effect of adenovirus-mediated decorin transduction on selected cancer cell lines was investigated. The results invariably showed that cancer cells completely lacked decorin expression. The study also demonstrated that transducing cancer cells with decorin adenoviral vector markedly inhibited their malignant behavior. In line with this, a strong induction of decorin expression in normal human embryonic stem cells (hESCs), but not in abnormal hESCs was observed during their differentiation. Secondly, the significance of HA in the development of diabetic macroangiopathy in response to hyperglycemia was evaluated. Results showed that the synthesis of HA by vascular smooth muscle cells was significantly increased in response to high glucose concentration. This increase was associated with the diminished ability of the cells to contract collagen-rich matrix suggesting that HA participates in the disturbed vascular remodeling of diabetic patients. The results of this study support endeavours to develop novel ECM macromolecule -based therapies targeting cancer and cardiovascular diseases.Siirretty Doriast

Simvastatin Modulates Extracellular Matrix Assembly by Displaying an Antifibrotic Activity in Vitro

Statins, competitive inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase, are known to possess properties beyond their cholesterol-lowering effect including anti-inflammatory, antiproliferative and anti-immunomodulatory effects. We examined the effect of simvastatin on extracellular matrix (ECM) assembly by human skin fibroblasts (HSFs) in vitro. Using collagen gel contraction (CGC) assay we showed that simvastatin inhibits contraction of type I collagen-rich gels in a dose-dependent manner. This effect of simvastatin could be overcome by co-incubating the cells with mevalonate. Actin staining revealed that inhibition of CGC by simvastatin is associated with diminished ability of the cells to form aggregates. Using whole human genome Illumina microarray we sought to search for new candidate genes whose expression is regulated by simvastatin during CGC and focused specifically on the genes related to ECM synthesis and remodeling. We found that simvastatin profoundly downregulated gene expression of 27 ECM molecules including proteoglycans decorin and versican, both of which are known to be essential constituents of proper ECM. Expression of these two molecules was further verified by Northern blot analysis. Finally, when simvastatin treated HSFs were activated with TGF-β1, the cell-mediated contraction of collagen gel was restored. Our results indicate that simvastatin markedly alters ECM assembly in vitro possessing an antifibrotic activity.</p

Decorin-mediated oncosuppression – a potential future adjuvant therapy for human epithelial cancers

Currently, the multifaceted role of the extracellular matrix (ECM) in tumorigenesis has been realized. One ECM macromolecule exhibiting potent oncosuppressive actions in tumorigenesis is decorin, the prototype of the small leucine-rich proteoglycan gene family. The actions of decorin include its capability to function as an endogenous pan-receptor tyrosine kinase inhibitor, a regulator of both autophagy and mitophagy, as well as a modulator of the immune system. In this review, we will discuss these topics in more detail. We will also give a summary of preclinical studies exploring the value of decorin-mediated oncosuppression as a potential future adjuvant therapy for epithelial cancers.   </p

Localization of decorin gene expression in normal human breast tissue and in benign and malignant tumors of the human breast

The small extracellular matrix proteoglycan decorin which possesses a potent antitumor activity has been shown to be present in various amounts in the stroma of several tumors including those of the breast. Regarding decorin in breast malignancies the published data are conflicting, i.e., whether breast cancer cells express it or not. Here, we first compared decorin gene expression levels between healthy human breast tissue and selected types of human breast cancer using GeneSapiens databank. Next, we localized decorin mRNA in tissue specimen of normal human breast, intraductal breast papillomas and various histologic types of human breast cancer using in situ hybridization (ISH) with digoxigenin-labeled RNA probes for decorin. We also examined the effect of decorin transduction on the behavior of cultured human breast cancer MCF7 cells. Analysis of GeneSapiens databank revealed that in various human breast cancers decorin expression is significant. However, ISH results clearly demonstrated that human breast cancer cells independently of the type of the cancer do not express decorin mRNA. This was also true for papilloma-forming cells of the human breast. Indeed, decorin gene expression in healthy human breast tissue as well as in benign and malignant tumors of human breast was shown to take place solely in cells of the original stroma. Decorin transduction using decorin adenoviral vector in decorin-negative MCF7 cells resulted in a significant decrease in the proliferation of these cells and changed cell cohesion. Decorin-transduced MCF7 cells also exhibited increased apoptosis. In conclusion, our study shows that in human breast tissue only cells of the original stroma are capable of decorin gene expression. Our study also shows that transduction of decorin in decorin-negative human breast cancer cells markedly modulates the growth pattern of these cells

A novel double nucleotide variant in the ferritin-L iron-responsive element in a Finnish patient with hereditary hyperferritinaemia-cataract syndrome

ABSTRACT.Purpose: To present a novel Finnish double nucleotide variant in the ironresponsiveelement (IRE) of the ferritin L-chain gene (FTL) leading tohyperferritinaemia-cataract syndrome (HHCS).PurposeTo present a novel Finnish double nucleotide variant in the iron-responsive element (IRE) of the ferritin L-chain gene (FTL) leading to hyperferritinaemia-cataract syndrome (HHCS).MethodsGenomic DNA extracted from peripheral blood leucocytes and synthetized with three different primers flanking the IRE in the FTL 5′-untranslated region of the FTL was used in polymerase chain reaction (PCR). Thereafter, Sanger sequencing was performed on the 487-bp and 602-bp PCR amplification products with specific primers to reveal FTL IRE mutations.ResultsA 58-year-old female patient with elevated serum ferritin level (1339 μg/l) was diagnosed with HHCS after extensive workup. Genetic testing identified a novel double point mutation g.48965355G>C (chr19, hg19) and g.48965356G>T (chr19, hg19) in the lower stem region of the IRE canonical structure of the FTL.ConclusionAfter excluding other causes, elevated serum ferritin level in a person with early onset cataract is indicative for HHCS, a genetic disorder caused by mutation in the IRE of the FTL.</p

Decorin Expression and Oncosuppression in Human Embryonic Carcinomas

Human embryonic stem cells in culture can transform into malignant, cancer-like cells exhibiting lesser differentiation. After transplantation, these transformed cells can form highly malignant germ cell tumors. In humans, germ cell tumors often appear at gonadal sites, like in the testis. In this study, we examined the expression of small leucine rich proteoglycans in normal and karyotypically abnormal human embryonic stem cells using a publicly available transcriptome data. We also examined the expression of the small leucine rich proteoglycans in healthy human testis and in different human testicular non-seminoma germ cell tumors using IST Online database. Furthermore, we localized the expression of decorin, the prototype member of the small leucine rich proteoglycans, in samples representing the above testicular tissues, using in situ hybridization and immunohistochemistry. The analysis revealed that the expression of two small leucine rich proteoglycans, namely decorin and lumican, was induced in normal but not in karyotypically abnormal human embryonic stem cells during early cell differentiation. Similarly, in IST Online database the expression of these two small leucine rich proteoglycans was markedly higher in differentiated teratoma tissue than in undifferentiated embryonal carcinoma tissue. In testicular germ cell tumors decorin expression was completely lacking in the areas of undifferentiated malignant cells. The above results collectively suggest that decorin and lumican have a role in human stem cell differentiation and testicular non-seminoma germ cell tumor formation.</p

Distinctive effects of SGLT2 inhibitors on angiogenesis in zebrafish embryos

Sodium glucose cotransporter 2 (SGLT2) inhibitor canagliflozin has been found to increase the risk for lowerlimb amputations in type 2 diabetics about two-fold. Conversely, empagliflozin and dapagliflozin do not display a similar effect. A question arises whether the increased risk for minor amputations is associated only with canagliflozin or whether it is a class effect of SGLT2 inhibitors. Defective angiogenesis has a role in amputations. We compared the effects of empagliflozin, dapagliflozin and canagliflozin on angiogenesis in vivo using zebrafish model, and in vitro using human umbilical vein endothelial cells (HUVECs). The effects of SGLT2 inhibitors on the formation of intersegmental blood vessels (ISVs) of the zebrafish embryos were clarified. Additionally, transcriptome analysis was performed to explore whether putative angiogenesis-associated genes are differentially regulated by SGLT2 inhibitors. The effects of SGLT2 inhibitors on the viability of HUVECs were examined. We noticed that especially empagliflozin and also dapagliflozin significantly accelerated the formation of ISVs of zebrafish embryos. In contrast, canagliflozin was not able to stimulate ISV formation, and at high concentration, it was lethal to the embryos. Transcriptome analysis demonstrated that in empagliflozin-treated embryos compared to canagliflozin-treated embryos seven genes previously shown to contribute to angiogenesis were upregulated, and four downregulated. Canagliflozin at high concentrations, but not empagliflozin or dapagliflozin, decreased the viability of HUVECs and disrupted their capability to sprout. SGLT2 inhibitors differed in their effects on angiogenic processes in zebrafish embryos and on the viability of HUVECs suggesting that the risk of SGLT2 inhibitors for peripheral amputations likely differs

Metformin decreases hyaluronan synthesis by vascular smooth muscle cells

Metformin is the first-line drug in the treatment of type 2 diabetes worldwide based on its effectiveness and cardiovascular safety. Currently metformin is increasingly used during pregnancy in women with gestational diabetes mellitus, even if the long-term effects of metformin on offspring are not exactly known. We have previously shown that high glucose concentration increases hyaluronan (HA) production of cultured human vascular smooth muscle cells (VSMC) via stimulating the expression of hyaluronan synthase 2 (HAS2). This offers a potential mechanism whereby hyperglycemia leads to vascular macroangiopathy. In this study, we examined whether gestational metformin use affects HA content in the aortic wall of mouse offspring in vivo. We also examined the effect of metformin on HA synthesis by cultured human VSMCs in vitro. We found that gestational metformin use significantly decreased HA content in the intima-media of mouse offspring aortas. In accordance with this, the synthesis of HA by VSMCs was also significantly decreased in response to treatment with metformin. This decrease in HA synthesis was shown to be due to the reduction of both the expression of HAS2 and the amount of HAS substrates, particularly UDP-N-acetylglucosamine. As shown here, gestational metformin use is capable to program reduced HA content in the vascular wall of the offspring strongly supporting the idea, that metformin possesses long-term vasculoprotective effects.</p

Decorin Expression in Human Vulva Carcinoma: Oncosuppressive Effect of Decorin cDNA Transduction on Carcinoma Cells

The extracellular matrix proteoglycan decorin is well-known for its oncosuppressive activity. Here, decorin expression was examined in human vulva carcinoma tissue samples and in primary and commercial cell lines representing this malignant disease. Furthermore, the effect of adenovirus-mediated decorin cDNA (Ad-DCN) transduction on the viability, proliferation, and the expression and activity of the epidermal growth factor receptor (ErbB/HER) family members of the cell lines were investigated. Using in situ hybridization and immunohistochemistry for decorin, it was demonstrated that malignant cells in human vulva carcinoma tissues lack decorin expression. This result was true independently on tumor stage, grade or human papillomavirus status. RT-qPCR analyses showed that the human vulva carcinoma cell lines used in this study were also negative for decorin expression. Transduction of the cell lines with Ad-DCN caused a marked reduction in cell viability, while the proliferation of the cells was not affected. Experiments examining potential mechanisms behind the oncosuppressive effect of Ad-DCN transduction revealed that ErbB2/HER2 expression and activity in carcinoma cells were markedly downregulated. In conclusion, the results of this study showed that human vulva carcinoma cells lack decorin expression, and that Ad-DCN transduction of these cells induces oncosuppressive activity in part via downregulation of ErbB2/HER2.</p