TY - JOUR A1 - Seher, Axel A1 - Nickel, Joachim A1 - Mueller, Thomas D. A1 - Kneitz, Susanne A1 - Gebhardt, Susanne A1 - Meyer ter Vehn, Tobias A1 - Schlunck, Guenther A1 - Sebald, Walter T1 - Gene expression profiling of connective tissue growth factor (CTGF) stimulated primary human tenon fibroblasts reveals an inflammatory and wound healing response in vitro JF - Molecular Vision N2 - Purpose: The biologic relevance of human connective tissue growth factor (hCTGF) for primary human tenon fibroblasts (HTFs) was investigated by RNA expression profiling using affymetrix (TM) oligonucleotide array technology to identify genes that are regulated by hCTGF. Methods: Recombinant hCTGF was expressed in HEK293T cells and purified by affinity and gel chromatography. Specificity and biologic activity of hCTGF was confirmed by biosensor interaction analysis and proliferation assays. For RNA expression profiling HTFs were stimulated with hCTGF for 48h and analyzed using affymetrix (TM) oligonucleotide array technology. Results were validated by real time RT-PCR. Results: hCTGF induces various groups of genes responsible for a wound healing and inflammatory response in HTFs. A new subset of CTGF inducible inflammatory genes was discovered (e.g., chemokine [C-X-C motif] ligand 1 [CXCL1], chemokine [C-X-C motif] ligand 6 [CXCL6], interleukin 6 [IL6], and interleukin 8 [IL8]). We also identified genes that can transmit the known biologic functions initiated by CTGF such as proliferation and extracellular matrix remodelling. Of special interest is a group of genes, e.g., osteoglycin (OGN) and osteomodulin (OMD), which are known to play a key role in osteoblast biology. Conclusions: This study specifies the important role of hCTGF for primary tenon fibroblast function. The RNA expression profile yields new insights into the relevance of hCTGF in influencing biologic processes like wound healing, inflammation, proliferation, and extracellular matrix remodelling in vitro via transcriptional regulation of specific genes. The results suggest that CTGF potentially acts as a modulating factor in inflammatory and wound healing response in fibroblasts of the human eye. KW - Bone morphogenetic protein-2 KW - Smooth-muscle-cells KW - Myofibroblast differentiation KW - TGF-beta KW - CYR61 KW - Proliferation KW - Mechanisms KW - Apoptosis KW - Receptor KW - Cancer Y1 - 2011 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-140189 VL - 17 IS - 08. Okt ER - TY - JOUR A1 - Lagler, Charlotte A1 - El-Mesery, Mohamed A1 - Kübler, Alexander Christian A1 - Müller-Richter, Urs Dietmar Achim A1 - Stühmer, Thorsten A1 - Nickel, Joachim A1 - Müller, Thomas Dieter A1 - Wajant, Harald A1 - Seher, Axel T1 - The anti-myeloma activity of bone morphogenetic protein 2 predominantly relies on the induction of growth arrest and is apoptosis-independent JF - PLoS ONE N2 - Multiple myeloma (MM), a malignancy of the bone marrow, is characterized by a pathological increase in antibody-producing plasma cells and an increase in immunoglobulins (plasmacytosis). In recent years, bone morphogenetic proteins (BMPs) have been reported to be activators of apoptotic cell death in neoplastic B cells in MM. Here, we use bone morphogenetic protein 2 (BMP2) to show that the "apoptotic" effect of BMPs on human neoplastic B cells is dominated by anti-proliferative activities and cell cycle arrest and is apoptosis-independent. The anti-proliferative effect of BMP2 was analysed in the human cell lines KMS12-BM and L363 using WST-1 and a Coulter counter and was confirmed using CytoTox assays with established inhibitors of programmed cell death (zVAD-fmk and necrostatin-1). Furthermore, apoptotic activity was compared in both cell lines employing western blot analysis for caspase 3 and 8 in cells treated with BMP2 and FasL. Additionally, expression profiles of marker genes of different cell death pathways were analysed in both cell lines after stimulation with BMP2 for 48h using an RT-PCR-based array. In our experiments we observed that there was rather no reduction in absolute cell number, but cells stopped proliferating following treatment with BMP2 instead. The time frame (48–72 h) after BMP2 treatment at which a reduction in cell number is detectable is too long to indicate a directly BMP2-triggered apoptosis. Moreover, in comparison to robust apoptosis induced by the approved apoptotic factor FasL, BMP2 only marginally induced cell death. Consistently, neither the known inhibitor of apoptotic cell death zVAD-fmk nor the necroptosis inhibitor necrostatin-1 was able to rescue myeloma cell growth in the presence of BMP2. KW - apoptosis KW - gene expression KW - necrotic cell death KW - multiple myeloma KW - cell metabolism KW - cell cycle and cell division KW - B cells Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-158993 VL - 12 IS - 10 ER - TY - JOUR A1 - Seher, Axel A1 - Lagler, Charlotte A1 - Stühmer, Thorsten A1 - Müller-Richter, Urs Dietmar Achim A1 - Kübler, Alexander Christian A1 - Sebald, Walter A1 - Müller, Thomas Dieter A1 - Nickel, Joachim T1 - Utilizing BMP-2 muteins for treatment of multiple myeloma JF - PLoS ONE N2 - Multiple myeloma (MM) represents a haematological cancer characterized by the pathological hyper proliferation of antibody-producing B-lymphocytes. Patients typically suffer from kidney malfunction and skeletal disorders. In the context of MM, the transforming growth factor β (TGFβ) member Activin A was recently identified as a promoter of both accompanying symptoms. Because studies have shown that bone morphogenetic protein (BMP)-2-mediated activities are counteracted by Activin A, we analysed whether BMP2, which also binds to the Activin A receptors ActRII and ActRIIB but activates the alternative SMAD-1/5/8 pathway, can be used to antagonize Activin A activities, such as in the context of MM. Therefore three BMP2 derivatives were generated with modified binding activities for the type II (ActRIIB) and/or type I receptor (BMPRIA) showing either increased or decreased BMP2 activity. In the context of MM these BMP2 muteins show two functionalities since they act as a) an anti-proliferative/apoptotic agent against neoplastic B-cells, b) as a bone-formation promoting growth factor. The molecular basis of both activities was shown in two different cellular models to clearly rely on the properties of the investigated BMP2 muteins to compete for the binding of Activin A to the Activin type II receptors. The experimental outcome suggests new therapeutic strategies using BMP2 variants in the treatment of MM-related pathologies. KW - multiple myeloma KW - signaling KW - cell proliferation KW - cell binding KW - membrane receptor signaling KW - BMP KW - gene expression KW - B cell receptors KW - B cells Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-158144 VL - 12 IS - 5 ER -