Optoelectronic Strain-Measurement System Demonstrated on Scaled-Down Flywheels.

Monitoring the strain in the rotating flywheel in a kinetic energy storage system is important for safe operation and for the investigation of long-term effects in composite materials like carbon-fiber-reinforced plastics. An optoelectronic strain-measurement system for contactless deformation and position monitoring of a flywheel was investigated. The system consists of multiple optical sensors measuring the local relative in-plane displacement of the flywheel rotor. A special reflective pattern, which is necessary to interact with the sensors, was applied to the surface of the rotor. Combining the measurements from multiple sensors makes it possible to distinguish between the deformation and in-plane displacement of the flywheel. The sensor system was evaluated using a low-speed steel rotor for single-sensor performance investigation as well as a scaled-down high-speed rotor made from PVC plastic. The PVC rotor exhibits more deformation due to centrifugal stresses than a steel or aluminum rotor of the same dimensions, which allows experimental measurements at a smaller flywheel scale as well as a lower rotation speed. Deformation measurements were compared to expected deformation from calculations. The influence of sensor distance was investigated. Deformation and position measurements as well as derived imbalance measurements were demonstrated.

Eddy Current Position Measurement in Harsh Environments: A Temperature Compensation and Calibration Approach.

Eddy current displacement sensors (ECDSs) are widely used for the noncontact position measurement of small displacements (lift-offs). Challenges arise with larger displacements as the sensitivity of the ECDSs decreases. This leads to a more pronounced impact of temperature variations on the inductance and, consequently, an increased position error. Design solutions often rely on multiple coils, suitable coil carrier materials, and compensation measures to address the challenges. This study presents a single-coil ECDS for large displacement ranges in environments with high temperatures and temperature variations. The analysis is based on a sensor model derived from an equivalent circuit model (ECM). We propose design measures for both the sensing coil and the target, focusing on material selection to handle the impact of temperature variations. A key part of improving performance under varying temperatures includes model-based temperature compensation for the inductance of the sensing coil. We introduce a method to calibrate the sensor for large displacements, using a modified coupling coefficient based on field simulation data. Our analysis shows that this single-coil ECDS design maintains a position error of less than 0.2% full-scale for a temperature variation of 100 K for the sensing coil and 110 K for the target.

Excitation of Mechanical Resonances in the Stationary Ring of a Mechanical Seal by a Continuously Operated Electromagnetic Acoustic Transducer.

Acoustic/ultrasonic testing is now a common method in the field of nondestructive testing for detecting material defects or monitoring ongoing mechanical changes in a structure during operation. In many applications, piezoelectric transducers are used to generate mechanical waves inside the specimen. Their actual operating frequency is highly dependent on the dimensions of the transducer. Larger dimensions of the piezoelectric transducer allow for a lower operating frequency. However, these dimensions limit the use of piezoelectric transducers in certain applications where the size of the transducer is restricted due to limited installation space and when low-frequency excitation is required. One application that places these requirements on the transducer is the monitoring of mechanical seals. Here, the transducer must be mounted on the stationary ring of the seal. In this paper, a continuously operated electromagnetic acoustic transducer (EMAT) is presented as an alternative to piezoelectric transducers as a transmitter. The advantage of a EMAT is that it meets the requirements of limited sensor size (sensor area < 10 × 6 mm) and can excite mechanical waves with frequencies below 10 kHz. A structural analysis of the stationary ring shows that the first two mechanical resonances occur around 4 and 5.5 kHz. An experimental study meterologically demonstrates the ability of the EMAT to excite these first two mechanical resonances of the ring. A comparative simulation agrees well with the measurement.

Uncertainty Analysis of an Optoelectronic Strain Measurement System for Flywheel Rotors.

The strain in a fast spinning carbon fiber flywheel rotor is of great interest for condition monitoring, as well as for studying long-term aging effects in the carbon fiber matrix. Optoelectronic strain measurement is a contactless measurement principle where a special reflective pattern is applied to the rotor which is scanned by a stationary optical setup. It does not require any active electronic components on the rotor and is suited for operation in a vacuum. In this paper, the influences of the key parts comprising the optoelectronic strain measurement are analyzed. The influence of each part on the measurement result including the uncertainty is modeled. The total uncertainty, as well as each part's contribution is calculated. This provides a valuable assessment of requirements for component selection, as well as tolerances of mechanical parts and processes to reach a final target measurement uncertainty or to estimate the uncertainty of a given setup. We have shown that the edge quality of the special reflective pattern has the strongest influence, and how to improve it. Considering all influences, it is possible to measure strain with an uncertainty of less than 1% at a rotation speed of 500Hz.

Angiogenic capacity of M1- and M2-polarized macrophages is determined by the levels of TIMP-1 complexed with their secreted proMMP-9.

A proangiogenic function of tissue-infiltrating monocytes/macrophages has long been attributed to their matrix metalloproteinase-9 zymogen (proMMP-9). Herein, we evaluated the capacity of human monocytes, mature M0 macrophages, and M1- and M2-polarized macrophages to induce proMMP-9-mediated angiogenesis. Only M2 macrophages induced angiogenesis at levels comparable with highly angiogenic neutrophils previously shown to release their proMMP-9 in a unique form, free of tissue inhibitor of metalloproteinases-1 (TIMP-1). Macrophage differentiation was accompanied by induction of low-angiogenic, TIMP-1-encumbered proMMP-9. However, polarization toward the M2, but not the M1 phenotype, caused a substantial downregulation of TIMP-1 expression, resulting in production of angiogenic, TIMP-deficient proMMP-9. Correspondingly, the angiogenic potency of M2 proMMP-9 was lost after its complexing with TIMP-1, whereas TIMP-1 silencing in M0/M1 macrophages rendered them both angiogenic. Similar to human cells, murine bone marrow-derived M2 macrophages also shut down their TIMP-1 expression and produced proMMP-9 unencumbered by TIMP-1. Providing proof that angiogenic capacity of murine M2 macrophages depended on their TIMP-free proMMP-9, Mmp9-null M2 macrophages were nonangiogenic, although their TIMP-1 was severely downregulated. Our study provides a unifying molecular mechanism for high angiogenic capacity of TIMP-free proMMP-9 that would be uniquely produced in a pathophysiological microenvironment by influxing neutrophils and/or M2 polarized macrophages.

The VEGF-regulated transcription factor HLX controls the expression of guidance cues and negatively regulates sprouting of endothelial cells.

The HLX gene encoding a diverged homeobox transcription factor has been found to be up-regulated by vascular endothelial growth factor-A (VEGF-A) in endothelial cells. We have now investigated the gene repertoire induced by HLX and its potential biologic function. HLX strongly increased the transcripts for several repulsive cell-guidance proteins including UNC5B, plexin-A1, and semaphorin-3G. In addition, genes for transcriptional repressors such as HES-1 were up-regulated. In line with these findings, adenoviral overexpression of HLX inhibited endothelial cell migration, sprouting, and vessel formation in vitro and in vivo, whereas proliferation was unaffected. This inhibition of sprouting was caused to a significant part by HLX-mediated up-regulation of UNC5B as shown by short hairpin RNA (shRNA)-mediated down-modulation of the respective mRNA. VEGF-A stimulation of endothelial cells induced elevated levels of HLX over longer time periods resulting in especially high up-regulation of UNC5B mRNA as well as an increase in cells displaying UNC5B at their surface. However, induction of HLX was strongly reduced and UNC5B up-regulation completely abrogated when cells were exposed to hypoxic conditions. These data suggest that HLX may function to balance attractive with repulsive vessel guidance by up-regulating UNC5B and to down-modulate sprouting under normoxic conditions.

Tumor-recruited neutrophils and neutrophil TIMP-free MMP-9 regulate coordinately the levels of tumor angiogenesis and efficiency of malignant cell intravasation.

Tumor-associated neutrophils contribute to neovascularization by supplying matrix metalloproteinase-9 (MMP-9), a protease that has been genetically and biochemically linked to induction of angiogenesis. Specific roles of inflammatory neutrophils and their distinct proMMP-9 in the coordinate regulation of tumor angiogenesis and tumor cell dissemination, however, have not been addressed. We demonstrate that the primary tumors formed by highly disseminating variants of human fibrosarcoma and prostate carcinoma recruit elevated levels of infiltrating MMP-9-positive neutrophils and concomitantly exhibit enhanced levels of angiogenesis and intravasation. Specific inhibition of neutrophil influx by interleukin 8 (IL-8) neutralization resulted in the coordinated diminishment of tumor angiogenesis and intravasation, both of which were rescued by purified neutrophil proMMP-9. However, if neutrophil proMMP-9, naturally devoid of tissue inhibitor of metalloproteinases (TIMP), was delivered in complex with TIMP-1 or in a mixture with TIMP-2, the protease failed to rescue the inhibitory effects of anti-IL8 therapy, indicating that the TIMP-free status of proMMP-9 is critical for facilitating tumor angiogenesis and intravasation. Our findings directly link tumor-associated neutrophils and their TIMP-free proMMP-9 with the ability of aggressive tumor cells to induce the formation of new blood vessels that serve as conduits for tumor cell dissemination. Thus, treatment of cancers associated with neutrophil infiltration may benefit from specific targeting of neutrophil MMP-9 at early stages to prevent ensuing tumor angiogenesis and tumor metastasis.

Neutrophil MMP-9 proenzyme, unencumbered by TIMP-1, undergoes efficient activation in vivo and catalytically induces angiogenesis via a basic fibroblast growth factor (FGF-2)/FGFR-2 pathway.

The structural and catalytic requirements for neutrophil MMP-9 proenzyme (proMMP-9) to induce angiogenesis were investigated using a quantitative angiogenesis model based on grafting of collagen onplants onto the chorioallantoic membrane of chick embryos. Both physiological activation of neutrophil proMMP-9 and proteolytic activity of the generated MMP-9 enzyme were critically dependent on the tissue inhibitor of metalloproteinase (TIMP)-free status of the zymogen. The presence of an intact active site and hemopexin domain were required for full angiogenesis-inducing activity of the MMP-9 enzyme. Timed additions of TIMP-1 to the onplants containing TIMP-free neutrophil proMMP-9 indicated that in vivo activation of the zymogen occurred during the first 24 h after grafting. Within the onplant tissue, MMP-9 activation was accompanied by proteolytic modifications of fibrillar collagen and an influx of host proteins, the rate of which depended on the TIMP-free status of the zymogen. By quantifying the levels of host angiogenic factors, we demonstrated that basic fibroblast growth factor (FGF-2) was a major cytokine becoming bioavailable in the onplant tissue undergoing a neutrophil proMMP-9-mediated angiogenic switch. Inhibition of angiogenesis with specific function-blocking antibodies further indicated an involvement of a FGF-2/FGFR-2 pathway in neutrophil proMMP-9-induced angiogenesis. The enhanced angiogenesis catalyzed by neutrophil MMP-9 appears to evoke also a localized, low threshold level vascular endothelial growth factor (VEGF)/VEGFR-2 pathway, likely functioning in the formation and/or stabilization of blood vessels. That neutrophil proMMP-9, unencumbered by TIMP-1, directly mediates FGF-2-dependent angiogenesis was also demonstrated in our quantitative mouse angiogenesis model employing subcutaneous collagen implants, thus implicating the novel TIMP-free MMP-9/FGF-2/FGFR-2 pathway in proMMP-9-induced angiogenesis in a mammalian setting.

The VEGF-induced transcriptional response comprises gene clusters at the crossroad of angiogenesis and inflammation.

VEGF-A is the major trigger of vasculogenesis and physiologic angiogenesis. We have investigated to which extent the gene repertoire induced by VEGF-A in endothelial cells is distinct from that of other growth factors and inflammatory cytokines. Genes upregulated in human umbilical vein endothelial cells treated with VEGF, EGF or IL-1 were compared by microarray analysis and clusters characteristic for individual or combinations of inducers were defined. VEGF-A upregulated in comparison to EGF a five-fold larger gene repertoire, which surprisingly overlapped to 60% with the inflammatory repertoire of IL-1. As shown by real-time RT-PCR for selected genes, VEGF-induction was mostly mediated by VEGF receptor-2 and the capacity of VEGF-A to induce genes in common with IL-1 largely depended on activation of the calcineurin/NFAT pathway, since cyclosporin A inhibited this induction. Another angiogenic growth factor, bFGF, did not share a comparable induction of inflammatory genes, but partially induced a small group of genes in common with VEGF-A, which were not regulated by EGF. Thus, the data display that VEGF-A induces a distinct gene repertoire, which, contrasting with other growth factors such as EGF or bFGF, includes an inherent inflammatory component possibly contributing to the cross-regulation of angiogenesis and inflammation as further indicated by the VEGF-mediated induction of leukocyte adhesion. Furthermore, a small group of genes selectively induced by VEGF-A with potential importance for angiogenesis is defined.

Signal transduction induced in endothelial cells by growth factor receptors involved in angiogenesis.

New vessel formation during development and in the adult is triggered by concerted signals of largely endothelial-specific receptors for ligands of the VEGF, angiopoietin and ephrin families. The signals and genes induced by these receptors operate in the context of additional signals transduced by non-endothelial specific growth factor receptors, inflammatory cytokine receptors as well as adhesion molecules. We summarize here available data on characteristic signaling of the VEGF receptor-2 and the current state of knowledge regarding the additional different receptor tyrosine kinases of the VEGF, Tie and Ephrin receptor families. Furthermore, the potential cross-talk with signals induced by other growth factors and inflammatory cytokines as well as the modulation by VE-cadherin is discussed.

Nuclear factor of activated T cells and early growth response-1 cooperate to mediate tissue factor gene induction by vascular endothelial growth factor in endothelial cells.

Based on the finding that tissue factor belongs to a group of genes upregulated in endothelial cells by VEGF, but not by EGF, we investigated signals selectively triggered by VEGF. Whereas the transcription factor early growth response (EGR)-1, which has previously been shown by us to be essentially involved in tissue factor gene regulation, was similarly induced by both factors, one major difference between VEGF and EGF signaling was the activation of the Ca(++)-mediated calcineurin/nuclear factor of activated T cells (NFAT) pathway by VEGF. Consistent with the importance of this pathway for tissue factor induction, treatment of endothelial cells with the Ca(++) chelator BAPTA-AM, as well as the calcineurin inhibitor cyclosporin A, partially inhibited VEGF-induced tissue factor upregulation. Furthermore, tissue factor reporter gene assays revealed a synergistic cooperation of NFAT and EGR-1 in the induction of the TF promoter, and a physical interaction between the two factors was indicated by co-immunoprecipitation assays. Another gene upregulated by VEGF predominantly via NFAT, which is not induced by EGF, is the DSCR-1 gene. The calcineurin inhibitor DSCR-1 seems to be induced by VEGF in a negative feed-back loop to limit NFAT activation. When we tested adenoviral overexpression of DSCR-1, VEGF-mediated induction of tissue factor mRNA was reduced, and complete suppression could be achieved by a combination of viruses expressing DSCR-1 and NAB2, a corepressor of EGR-1. These findings support that both, NFAT and EGR-1, are required for tissue factor upregulation in response to VEGF.

Signals and genes induced by angiogenic growth factors in comparison to inflammatory cytokines in endothelial cells.

The evaluation of signaling pathways leading to gene induction by VEGF-A and IL-1 in endothelial cells supports the importance of the NF-kappaB pathway for the IL-1-induced gene repertoire, whereas VEGF-A is a strong and preferential trigger of signals via PLC-gamma. This leads (i) via Ca(++) to the activation of calcineurin and NFAT and (ii) via PKC and the MEK/ERK MAPK pathway to the upregulation of EGR-1. Part of the VEGF-triggered gene induction depends on a cooperation of the transcription factors NFAT and EGR-1. Gene activation via PLC-gamma provides VEGF with the potency to induce a wide spectrum of genes including many also upregulated by IL-1. A gene upregulated by VEGF and IL-1 is the DSCR-1 gene, which encodes an inhibitor of calcineurin. DSCR1 is induced by NFAT or NF-kappaB and limits Ca(++) signaling in a negative feed-back loop. Similarly, NAB2, a corepressor of EGR-1, is induced by EGR-1 and limits EGR-1 effects. Adenoviral overexpression of DSCR1 or NAB2 inhibited part of VEGF-induced gene expression and reduced sprouting in angiogenesis models.

Opposing Roles of JNK and p38 in Lymphangiogenesis in Melanoma.

In primary melanoma, the amount of vascular endothelial growth factor C (VEGF-C) expression and lymphangiogenesis predicts the probability of metastasis to sentinel nodes, but conditions boosting VEGF-C expression in melanoma are poorly characterized. By comparative mRNA expression analysis of a set of 22 human melanoma cell lines, we found a striking negative correlation between VEGF-C and microphthalmia-associated transcription factor (MITF) expression, which was confirmed by data mining in GEO databases of human melanoma Affymetrix arrays. Moreover, in human patients, high VEGF-C and low MITF levels in primary melanoma significantly correlated with the chance of metastasis. Pathway analysis disclosed the respective c-Jun N-terminal kinase and p38/mitogen-activated protein kinase activities as being responsible for the inverse regulation of VEGF-C and MITF. Predominant c-Jun N-terminal kinase signaling results in a VEGF-C(low)/MITF(high) phenotype; these melanoma cells are highly proliferative, show low mobility, and are poorly lymphangiogenic. Predominant p38 signaling results in a VEGF-C(high)/MITF(low) phenotype, corresponding to a slowly cycling, highly mobile, lymphangiogenic, and metastatic melanoma. In conclusion, the relative c-Jun N-terminal kinase and p38 activities determine the biological behavior of melanoma. VEGF-C and MITF levels serve as surrogate markers for the respective c-Jun N-terminal kinase and p38 activities and may be used to predict the risk of metastasis in primary melanoma.

A microarray analysis of two distinct lymphatic endothelial cell populations.

We have recently identified lymphatic endothelial cells (LECs) to form two morphologically different populations, exhibiting significantly different surface protein expression levels of podoplanin, a major surface marker for this cell type. In vitro shockwave treatment (IVSWT) of LECs resulted in enrichment of the podoplanin(high) cell population and was accompanied by markedly increased cell proliferation, as well as 2D and 3D migration. Gene expression profiles of these distinct populations were established using Affymetrix microarray analyses. Here we provide additional details about our dataset (NCBI GEO accession number GSE62510) and describe how we analyzed the data to identify differently expressed genes in these two LEC populations.

The transcription factor MEF2C negatively controls angiogenic sprouting of endothelial cells depending on oxygen.

The MADS box transcription factor MEF2C has been detected by us to be upregulated by the angiogenic factors VEGF-A and bFGF in endothelial cells. We have here investigated its potential role for angiogenesis. MEF2C was surprisingly found to strongly inhibit angiogenic sprouting, whereas a dominant negative mutant rather induced sprouting. The factor mainly affected migratory processes of endothelial cells, but not proliferation. In gene profiling experiments we delineated the alpha-2-macroglobulin gene to be highly upregulated by MEF2C. Further data confirmed that MEF2C in endothelial cells indeed induces alpha-2-macroglobulin mRNA as well as the secretion of alpha-2-macroglobulin and that conditioned supernatants of cells overexpressing MEF2C inhibit sprouting. Alpha-2-macroglobulin mediates, at least to a large extent, the inhibitory effects of MEF2C as is shown by knockdown of alpha-2-macroglobulin mRNA by lentiviral shRNA expression which reduces the inhibitory effect. However, under hypoxic conditions the VEGF-A/bFGF-mediated upregulation of MEF2C is reduced and the production of alpha-2-macroglobulin largely abolished. Taken together, this suggests that the MEF2C/alpha-2-macroglobulin axis functions in endothelial cells as a negative feed-back mechanism that adapts sprouting activity to the oxygen concentration thus diminishing inappropriate and excess angiogenesis.

Molecular and cellular effects of in vitro shockwave treatment on lymphatic endothelial cells.

Extracorporeal shockwave treatment was shown to improve orthopaedic diseases and wound healing and to stimulate lymphangiogenesis in vivo. The aim of this study was to investigate in vitro shockwave treatment (IVSWT) effects on lymphatic endothelial cell (LEC) behavior and lymphangiogenesis. We analyzed migration, proliferation, vascular tube forming capability and marker expression changes of LECs after IVSWT compared with HUVECs. Finally, transcriptome- and miRNA analyses were conducted to gain deeper insight into the IVSWT-induced molecular mechanisms in LECs. The results indicate that IVSWT-mediated proliferation changes of LECs are highly energy flux density-dependent and LEC 2D as well as 3D migration was enhanced through IVSWT. IVSWT suppressed HUVEC 3D migration but enhanced vasculogenesis. Furthermore, we identified podoplaninhigh and podoplaninlow cell subpopulations, whose ratios changed upon IVSWT treatment. Transcriptome- and miRNA analyses on these populations showed differences in genes specific for signaling and vascular tissue. Our findings help to understand the cellular and molecular mechanisms underlying shockwave-induced lymphangiogenesis in vivo.