Refine
Has Fulltext
- yes (22)
Is part of the Bibliography
- yes (22)
Year of publication
Document Type
- Journal article (13)
- Doctoral Thesis (9)
Keywords
- extracellular matrix (22) (remove)
Institute
- Klinik und Poliklinik für Unfall-, Hand-, Plastische und Wiederherstellungschirurgie (Chirurgische Klinik II) (4)
- Lehrstuhl für Tissue Engineering und Regenerative Medizin (4)
- Abteilung für Funktionswerkstoffe der Medizin und der Zahnheilkunde (3)
- Graduate School of Life Sciences (2)
- Institut für Anatomie und Zellbiologie (2)
- Institut für Humangenetik (2)
- Institut für Pharmazie und Lebensmittelchemie (2)
- Klinik und Poliklinik für Dermatologie, Venerologie und Allergologie (2)
- Theodor-Boveri-Institut für Biowissenschaften (2)
- Augenklinik und Poliklinik (1)
Bei der Behandlung solider Tumoren spielen systemisch verabreichte Chemotherapeutika eine wich- tige Rolle. Allerdings akkumulieren diese Therapeutika besser in normalem Gewebe als in Tumoren. Als Ursache für diesen unzureichenden Transport von Medikamenten in den Tumor wurde bisher vor allem die dysfunktionale Tumorvaskulatur diskutiert. Diese befindet sich in einem chaotischen und unreifen Zustand ohne ausreichende Bedeckung der Gefäße mit stabilisierenden Perizyten. Aus dem Zustand der Vaskulatur resultierend erreichen Medikamente den Tumor nur in geringem Ausmaß und werden dort heterogen verteilt. Als Grund für den Zustand der Vaskulatur wur- de ein großer Überschuss an pro-angiogenetischen Faktoren im Tumor ausgemacht. Durch eine anti-angiogenetische Behandlung konnte in präklinischen Modellen für einen gewissen Zeitraum die Tumorvaskulatur „normalisiert“ werden. Dies zeichnete sich vor allem durch Veränderung von zwei wichtigen Parametern für die Medikamenteneinbringung aus: zum Einen kommt es zu einer Reduktion der Gefäßdichte. Zum Anderen zu einer Reifung der Blutgefäße. In einem Teil von Pati- enten scheint dabei der Effekt der Gefäßverbesserung zu überwiegen und es kann eine verbesserte Perfusion detektiert werden. Mutmaßlich führt dies auch zu einer verbesserten Einbringung von Therapeutika in den Tumor und so zu einer erhöhten Effizienz der Therapie. In einem weiteren Teil der Patienten scheint jedoch der Effekt der Gefäßreduktion zu überwiegen und die detektierte Perfusion im Tumor wird durch die Behandlung verringert.
Das in dieser Arbeit verwendete MT6-Fibrosarkom-Modell reagierte auf eine anti-angiogenetische Therapie nicht mit einer sonst in murinen Modellen beobachteten Wachstumsreduktion. Die- se ermöglichte eine so bisher nicht mögliche Untersuchung der sekundären Effekte einer anti- angiogenetischen Therapie wie die Medikamenteneinbringung in den Tumor. Die Vaskulatur in MT6-Tumoren zeigte dabei nach einer anti-angiogenetischen Vorbehandlung, die erwarteten Merk-male einer „normalisierten“ Vaskulatur wie eine Reduktion der Gefäßdichte bei gleichzeitiger Rei- fung der verbleibenden Gefäße. Dies führte jedoch nicht zu einer verbesserten Effizienz einer subsequenten Chemotherapie. Durch Vergleich mit einem weiteren Tumor-Modell, dem 4T1-Modell für ein metastasierendes Mammakarzinom, konnten signifikante Unterschiede im Gefäßbild beider Modelle ausgeschlossen werden. Durch mikroskopische Methoden konnte dabei beobachtet werden, dass die Diffusion von Medikamenten aus den Blutgefäßen des MT6-Modells im Vergleich zum 4T1-Modell verringert war. Weitere Untersuchungen deuten auf eine Differenz in der Qualität der extrazellulären Matrix der verwendeten Tumor-Modelle. Durch mRNA-Expressionsanalysen konnte die Enzymfamilie der Lysyloxidasen als mögliche Ursache für diesen Diffusionsunterschied identi- fiziert werden. Lysyloxidasen katalysieren vor allem die Quervernetzung von Proteinen der Extra- zellulärmatrix. Im Weiteren konnte gezeigt werden, dass die Quervernetzung von Matrixproteinen durch Lysyloxidasen ursächlich für die Diffusions-Inhibierung kleiner Moleküle wie das Chemo- therapeutikum Doxorubicin sein kann. Durch spezifische Inhibition der Lysyloxidasen mittels des Inhibitors βAPN konnte diese Diffusions-Inhibition sowohl in vitro als auch im MT6-Tumor-Modell nahezu vollständig verhindert werden. Die hohe Aktivität von Lysyloxidasen im MT6-Modell stell- te allerdings kein Alleinstellungsmerkmal dieses Modells dar. In weiteren Untersuchungen konnte gezeigt werden, dass Lysyloxidasen in einer Vielzahl von murinen und humanen Tumorzelllinien überexprimiert wird. Die Inhibition von Lysyloxidasen durch βAPN konnte dabei in allen unter- suchten Modellen die Einbringung von Medikamenten in den Tumor erhöhen und könnte so eine sinnvolle adjuvante Maßnahme zur Verbesserung bestehender Chemotherapien darstellen.
Supplement-free induction of cellular differentiation and polarization solely through the topography of materials is an auspicious strategy but has so far significantly lagged behind the efficiency and intensity of media-supplementation-based protocols. Consistent with the idea that 3D structural motifs in the extracellular matrix possess immunomodulatory capacity as part of the natural healing process, it is found in this study that human-monocyte-derived macrophages show a strong M2a-like prohealing polarization when cultured on type I rat-tail collagen fibers but not on collagen I films. Therefore, it is hypothesized that highly aligned nanofibrils also of synthetic polymers, if packed into larger bundles in 3D topographical biomimetic similarity to native collagen I, would induce a localized macrophage polarization. For the automated fabrication of such bundles in a 3D printing manner, the strategy of “melt electrofibrillation” is pioneered by the integration of flow-directed polymer phase separation into melt electrowriting and subsequent selective dissolution of the matrix polymer postprocessing. This process yields nanofiber bundles with a remarkable structural similarity to native collagen I fibers, particularly for medical-grade poly(ε-caprolactone). These biomimetic fibrillar structures indeed induce a pronounced elongation of human-monocyte-derived macrophages and unprecedentedly trigger their M2-like polarization similar in efficacy as interleukin-4 treatment.
Primary open-angle glaucoma (POAG) is a leading cause of blindness due to chronic degeneration of retinal ganglion cells and their optic nerve axons. It is associated with disturbed regulation of intraocular pressure, elevated intraocular levels of TGF-β2, aberrant extracellular matrix (ECM) deposition and increased outflow resistance in the trabecular meshwork (TM). The mechanisms underlying these changes are not fully understood. Cell-matrix interactions have a decisive role in TM maintenance and it has been suggested that TGF-β-induced inhibition of matrix metalloproteases may drive aberrant ECM deposition in POAG. Invadopodia and podosomes (invadosomes) are distinct sites of cell-matrix interaction and localized matrix-metalloprotease (MMP) activity. Here, we report on the effects of TGF-β2 on invadosomes in human trabecular meshwork cells. Human TM (HTM) cells were derived from donor tissue and pretreated with vehicle or TGF-β2 (2 ng/ml) for 3d. Invadosomes were studied in ECM degradation assays, protein expression and MMP-2 activity were assessed by western blot and zymography and ECM protein transcription was detected by RT-qPCR. HTM cells spontaneously formed podosomes and invadopodia as detected by colocalization of Grb2 or Nck1 to sites of gelatinolysis. Pretreatment with TGF-β2 enhanced invadosomal proteolysis and zymographic MMP-2 activity as well as MMP-2, TIMP-2 and PAI-1 levels in HTM cell culture supernatants. Rho-kinase inhibition by H1152 blocked the effects of TGF-β2. Concomitant transcription of fibronectin and collagens-1, -4 and -6 was increased by TGF-β2 and fibrillar fibronectin deposits were observed in areas of invadosomal ECM remodelling. In contrast to a current hypothesis, our data indicate that TGF-β2 induces an active ECM remodelling process in TM cells, characterized by concurrent increases in localized ECM digestion and ECM expression, rather than a mere buildup of material due to a lack of degradation. Invadosomal cell adhesion and signaling may thus have a role in POAG pathophysiology.
When aiming at cell‐based therapies in osteoarthritis (OA), proinflammatory conditions mediated by cytokines such as IL‐1β need to be considered. In recent studies, the phytoalexin resveratrol (RSV) has exhibited potent anti‐inflammatory properties. However, long‐term effects on 3D cartilaginous constructs under inflammatory conditions with regard to tissue quality, especially extracellular matrix (ECM) composition, have remained unexplored. Therefore, we employed long‐term model cultures for cell‐based therapies in an in vitro OA environment and evaluated effects of RSV. Pellet constructs made from expanded porcine articular chondrocytes were cultured with either IL‐1β (1–10 ng/ml) or RSV (50 μM) alone, or a cotreatment with both agents. Treatments were applied for 14 days, either directly after pellet formation or after a preculture period of 7 days. Culture with IL‐1β (10 ng/ml) decreased pellet size and DNA amount and severely compromised glycosaminoglycan (GAG) and collagen content. Cotreatment with RSV distinctly counteracted the proinflammatory catabolism and led to partial rescue of the ECM composition in both culture systems, with especially strong effects on GAG. Marked MMP13 expression was detected in IL‐1β‐treated pellets, but none upon RSV cotreatment. Expression of collagen type I was increased upon IL‐1β treatment and still observed when adding RSV, whereas collagen type X, indicating hypertrophy, was detected exclusively in pellets treated with RSV alone. In conclusion, RSV can counteract IL‐1β‐mediated degradation and distinctly improve cartilaginous ECM deposition in 3D long‐term inflammatory cultures. Nevertheless, potential hypertrophic effects should be taken into account when considering RSV as cotreatment for articular cartilage repair techniques.
Articular cartilage is an exceptional connective tissue which by a network of fibrillar collagen and glycosaminoglycan (GAG) molecules allows both low- friction articulation and distribution of loads to the subchondral bone (Armiento et al., 2018, Ulrich-Vinther et al., 2003). Because of its very limited ability to self-repair, chondral defects following traumatic injury increase the risk for secondary osteoarthritis (OA) (Muthuri et al., 2011). Still, current OA treatments such as common nonsteroidal anti-inflammatory drugs (NSAIDs) and joint replacement primarily address end-stage symptoms (Tonge et al., 2014). As low-grade inflammation plays a pivotal role in the pathogenesis of OA (Robinson et al., 2016), there is a strong demand for novel therapeutic concepts, such as integrating application of anti-inflammatory agents into cartilage cell- based therapies in order to effectively treat OA affected joints in early disease stages. The polyphenolic phytoalexin resveratrol (RSV), found in the skin of red grapes, berries, and peanuts, has been shown to have effective anti-inflammatory properties (Shen et al., 2012). However, its long-term effects on 3D chondrocyte constructs cultured in an inflammatory environment with regard to tissue quality have remained unexplored so far. Therefore, in this study, pellets made from expanded porcine articular chondrocytes were cultured for 14 days with either the pro-inflammatory cytokine interleukin-1β (IL-1β) (1 - 10 ng/ml) or RSV (50 μM) alone, or a co-treatment with both agents. Constructs treated with chondrocyte medium only served as control. Treatment with IL-1β at 10 ng/ml resulted in a significantly smaller pellet size and reduced DNA content. However, RSV counteracted the IL-1β-induced decrease and significantly enhanced diameter and DNA content. Also, in terms of GAG deposition, treatment with IL-1β at 10 ng/ml resulted in a tremendous depletion of absolute GAG content and GAG/DNA. Again, RSV co-treatment counteracted the inflammatory stimulus and led to a partial recovery of GAG content. Histological analysis utilizing safranin-O staining confirmed these findings. Marked expression of the cartilage-degrading enzyme matrix metalloproteinase 13 (MMP13) was detected in IL-1β-treated pellets, but none upon RSV co- treatment. Moreover, co-treatment of IL-1β-challenged constructs with RSV significantly increased absolute collagen content. However, under non- inflammatory conditions, RSV induced gene expression and protein accumulation of collagen type X, a marker for undesirable hypertrophy. Taken together, in the present thesis, RSV was demonstrated to elicit marked beneficial effects on the extracellular matrix composition of 3D cartilaginous constructs in long-term inflammatory culture in vitro, but also induced hypertrophy under non-inflammatory conditions. Based on these findings, further experiments examining multiple concentrations of RSV under various inflammatory conditions appear desirable concerning potential therapeutic applicability in OA.
The process of tumor invasion requires degradation of extracellular matrix by proteolytic enzymes. Cancer cells form protrusive invadopodia, which produce and release matrix metalloproteinases (MMPs) to degrade the basement membrane thereby enabling metastasis. We investigated the effect of LASP1, a newly identified protein in invadopodia, on expression, secretion and activation of MMPs in invasive breast tumor cell lines.
By analyzing microarray data of in-house generated control and LASP1-depleted MDA-MB-231 breast cancer cells, we observed downregulation of MMP1, -3 and -9 upon LASP1 depletion. This was confirmed by Western blot analysis. Conversely, rescue experiments restored in part MMP expression and secretion. The regulatory effect of LASP1 on MMP expression was also observed in BT-20 breast cancer cells as well as in prostate and bladder cancer cell lines.
In line with bioinformatic FunRich analysis of our data, which mapped a high regulation of transcription factors by LASP1, public microarray data analysis detected a correlation between high LASP1 expression and enhanced c-Fos levels, a protein that is part of the transcription factor AP-1 and known to regulate MMP expression. Compatibly, in luciferase reporter assays, AP-1 showed a decreased transcriptional activity after LASP1 knockdown.
Zymography assays and Western blot analysis revealed an additional promotion of MMP secretion into the extracellular matrix by LASP1, thus, most likely, altering the microenvironment during cancer progression.
The newly identified role of LASP1 in regulating matrix degradation by affecting MMP transcription and secretion elucidated the migratory potential of LASP1 overexpressing aggressive tumor cells in earlier studies.
Hepatic stellate cells (HSCs) are also known as lipocytes, fat-storing cells, perisinusoidal cells, or Ito cells. These liver-specific mesenchymal cells represent about 5% to 8% of all liver cells, playing a key role in maintaining the microenvironment of the hepatic sinusoid. Upon chronic liver injury or in primary culture, these cells become activated and transdifferentiate into a contractile phenotype, i.e., the myofibroblast, capable of producing and secreting large quantities of extracellular matrix compounds. Based on their central role in the initiation and progression of chronic liver diseases, cultured HSCs are valuable in vitro tools to study molecular and cellular aspects of liver diseases. However, the isolation of these cells requires special equipment, trained personnel, and in some cases needs approval from respective authorities. To overcome these limitations, several immortalized HSC lines were established. One of these cell lines is CFSC, which was originally established from cirrhotic rat livers induced by carbon tetrachloride. First introduced in 1991, this cell line and derivatives thereof (i.e., CFSC-2G, CFSC-3H, CFSC-5H, and CFSC-8B) are now used in many laboratories as an established in vitro HSC model. We here describe molecular features that are suitable for cell authentication. Importantly, chromosome banding and multicolor spectral karyotyping (SKY) analysis demonstrate that the CFSC-2G genome has accumulated extensive chromosome rearrangements and most chromosomes exist in multiple copies producing a pseudo-triploid karyotype. Furthermore, our study documents a defined short tandem repeat (STR) profile including 31 species-specific markers, and a list of genes expressed in CFSC-2G established by bulk mRNA next-generation sequencing (NGS).
Non-contact, Label-free Monitoring of Cells and Extracellular Matrix using Raman Spectroscopy
(2012)
Non-destructive, non-contact and label-free technologies to monitor cell and tissue cultures are needed in the field of biomedical research.1-5 However, currently available routine methods require processing steps and alter sample integrity. Raman spectroscopy is a fast method that enables the measurement of biological samples without the need for further processing steps. This laser-based technology detects the inelastic scattering of monochromatic light.6 As every chemical vibration is assigned to a specific Raman band (wavenumber in cm-1), each biological sample features a typical spectral pattern due to their inherent biochemical composition.7-9 Within Raman spectra, the peak intensities correlate with the amount of the present molecular bonds.1 Similarities and differences of the spectral data sets can be detected by employing a multivariate analysis (e.g. principal component analysis (PCA)).10
Here, we perform Raman spectroscopy of living cells and native tissues. Cells are either seeded on glass bottom dishes or kept in suspension under normal cell culture conditions (37 °C, 5% CO2) before measurement. Native tissues are dissected and stored in phosphate buffered saline (PBS) at 4 °C prior measurements. Depending on our experimental set up, we then either focused on the cell nucleus or extracellular matrix (ECM) proteins such as elastin and collagen. For all studies, a minimum of 30 cells or 30 random points of interest within the ECM are measured. Data processing steps included background subtraction and normalization.
Cell culture models are helpful tools to study inflammatory diseases, like rheumatoid arthritis (RA), osteoarthritis (OA), arteriosclerosis or asthma, which are linked to increased matrix metalloproteinase (MMP) activity. Such cell culture models often focus on the secretion of cytokines and growth factors or the direct effects of disease on tissue destruction. Even though the crucial role of MMPs in inflammatory diseases is known, the results of MMP studies are contradictious and the use of MMPs as biomarkers is inconsistent. MMPs play an important role in disease pathology, as they are involved in elastin degradation in the walls of alveoli in chronic obstructive pulmonary disease (COPD), tumor angiogenesis and metastasis and in cartilage and bone degradation in arthropathies. In RA and OA MMPs are secreted by osteocytes, synoviocytes, and by infiltrating immune cells in response to the increased concentration of inflammatory mediators, like growth factors and cytokines. MMPs are zinc and calcium-dependent proteinases and play an important role in physiological and pathological extracellular matrix (ECM) turn over. Their substrate specificity gives them the ability to degrade all major ECM components, like aggrecan, elastin, gelatin, fibronectin and all types of collagen even the triple helix of collagen monomers. The ECM consists of two large three-dimensional cross-linked macromolecule classes: one are fibrous proteins, like collagen and elastin fibers that are responsible for ECM’s structure, tensile strength, resiliency, reversible extensibility, and deformability and the second class is comprised of proteoglycans composed of glycosaminoglycan (GAG) chains covalently attached to protein cores that are multifunctionally involved in signaling pathways and cell interactions. ECM is present within all tissues and organs and changes in ECM structure contribute to pathogenesis, e.g. wounded and fibrotic tissue, COPD or tumours.
This thesis primarily focuses on the development of a diagnostic peptide system, that enables to gain information on MMP activity from ECM by deploying the isobaric mass encoding strategy. The core element of the developed system is an isotopically labelled peptide sequence (mass tag), that is released in response to elevated levels of MMPs and allows multiplexed detection in tandem mass spectrometry (LC-MS/MS). The mass reporters possess a modular structure with different functionalities. C-terminal either a transglutaminase (TG) recognition sequence or a high molecular weight polyethylene glycol (PEG) moiety was attached to immobilize the mass reporters covalently or physically at the injection site. The following matrix metalloproteinase substrate sequence (MSS) is incorporated in two different versions with different sensitivity to MMPs. The MSS were applied in pairs for relative quantification consisting of the cleavable version synthesized with natural L-amino acids and the non-cleavable D-amino acid variant. The mass tag was synthesized with isotopically labelled amino acids and is separated from the MSS by a UV light-sensitive molecule. N-terminal the mass tag is followed by a tobacco etch virus protease (TEV) sensitive sequence, that is responsible to separate the mass tag from the affinity tag, which was either the Strep-tag II sequence or biotin and were added for purification purposes.
Chapter 1 presents a step-by-step protocol on how to design a mass tag family allowing for multiplexed analysis by LC-MS/MS. The multiplexing is achieved by developing an isobar mass tag family with four family members, which are chromatographically indistinguishable, but due to the mass encoding principles they fragment in distinct y-type ions with a mass difference of 1 or 2 Da each in MS2. Furthermore, it is explained how to covalently attach the mass reporter peptides onto ECM by the activated calcium-catalyzed blood coagulation transglutaminase factor XIII (FXIIIa). The lysine of mass reporter’s TG sequence (D-domain of insulin-like growth factor-I (IGF-I)) and a glutamine in fibronectin are covalently crosslinked by FXIIIa and build an isopeptide bond. Elevated levels of MMP release the mass reporters from ECM by recognizing the inter-positioned MSS.
The designed mass reporters were able to monitor enzyme activity in an in vitro setting with cell-derived ECM, which was shown in Chapter 2. The modular structured mass reporters were investigated in a proof of concept study. First, the different modules were characterized in terms of their MMP responsiveness and their sensitivity to TEV protease and UV light. Then the FXIIIa-mediated coupling reaction was detailed and the successful coupling on ECM was visualized by an immunosorbent assay or confocal laser scanning microscopy. Finally, the immobilized mass reporters on ECM were incubated with MMP-9 to investigate their multiplexing ability of MMP activity. The cleaved mass reporter fragments were purified in three steps and mass tags were analyzed as mix of all four in LC-MS/MS.
Chapter 3 describes the change from an immobilizing system as seen in chapter 1 and 2 to a soluble enzyme activity monitoring system that was applied in an osteoarthritic mouse model. Instead of the immobilizing TG sequence the C-terminal MMS was extended with two amino acids where one holds an azide moiety to perform a strain-promoted azide-alkyne cycloaddition to a high molecular weight dibenzocyclooctyne-polyethylene glycol (DBCO-PEG), which was chosen to retain the mass reporters at the injection site. Furthermore, the N-terminal affinity tag was extended with a 2.5 kDa PEG chain to increase the half-life of the mass reporter peptides after MMP release. The systems biocompatibility was proved but its enzyme monitoring ability in an in vivo setting could not be analyzed as samples degraded during shipping resulting from the Chinese customs blocking transport to Germany.
In summary the diagnostic peptide system was developed in two variants. The immobilized version one from chapter 1 and 2 was designed to be covalently attached to ECM by the transglutaminase-mediated cross-linking reaction. In an in vitro setting the functionality of the mass reporter system for the detection of MMP activity was successfully verified. The second variant comprises of a soluble mass reporter system that was tested in an OA mouse model and showed biocompatibility. With these two designed systems this thesis provides a flexible platform based on multiplexed analysis with mass-encoded peptides to characterize cell culture models regarding their MMP activity, to deploy cell-derived ECM as endogenous depot scaffold and to develop a mass tag family that enables simultaneous detection of at least four mass tags.
Zellmigration ist essentiell für die Invasion und Metastasierung maligner Tumore. Neben der Bewegung von Einzelzellen zeigen Tumore sowohl epithe¬lialen als auch mesenchymalen Ursprungs auch kollektive Migration und Invasion multizellulärer Zellverbände, die sich unter Beibehaltung von Zell-Zell-Adhäsionen koordiniert als Gruppe bewegen. Ziel der Arbeit war, primäre humane Melanomexplantate mittels organotypischer Kultur in 3D Kollagenmatrices einzusetzen, um mittels Zeit-raffermikroskopie und experimentellen Blockadestrategien die zellulären und molekularen Grundlagen kollektiver Migration darzustellen, insbesondere die Bedeutung von Zell-Matrix-Interaktionen und Integrinen. In 3D Explantatkulturen bildeten primäre Melanomexplantate reproduzierbar Invasionszonen und sich ablösende und kollektiv wandernde Zellcluster aus. Diese zeichneten sich durch eine ausgeprägte Polarität mit motiler Vorderfront mit zugartig reorientierten Kollagenfasern und nachgezogenem hinteren Teil der Gruppen aus, vergleichbar der Asymmetrie haptokinetisch migrierender Fibroblasten. β1 Integrine zeigten ein heterogenes Verteilungsmuster mit Fokalisierung an Zell-Matrix-Interaktionen vor allem an der Vorderfront und linearer Anordnung entlang der Zell-Zell-Grenzen. Adhäsionsblockierende anti- β1 Integrin-Antikörper bewirkten nahezu vollständige Hemmung der kollektiven Migration, mit Verlust der Zellgruppenpolarität und Migrationspersistenz. Nach Integrinblockade zerfielen Zellverbände infolge Loslösung von Einzelzellen, die sich mittels β1 Integrin-unabhängiger, amöboider Migration durch die Kollagenmatrix bewegten. Der Übergang von β1 Integrin-abhängiger, kollektiver Migration zu amöboider Einzelzellwanderung (kollektiv-amöboide Transition) ist ein Beispiel für die Plastizität von Tumorzellwanderung, die in Anpassung an das Milieu einen Wechsel der Migrationsstrategie erlaubt. Die Plastizität der Tumorzellmigration muss bei der Entwicklung therapeutischer Konzepte, die auf Hemmung von Tumorinvasion und -metastasierung abzielen, berücksichtigt werden.