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Kurulgan Demirci, Eylem ; Demirci, Taylan ; Linder, Peter ; Trzewik, Jürgen ; Gierkowski, Jessica Ricarda ; Gossmann, Matthias ; Kayser, Peter ; Porst, Dariusz ; Digel, Ilya ; Artmann, Gerhard ; Temiz Artmann, Aysegül
All cells generate contractile tension. This strain is crucial for mechanically controlling the cell shape, function and survival. In this study, the CellDrum technology quantifying cell's (the cellular) mechanical tension on a pico-scale was used to investigate the effect of lipopolysaccharide (LPS) on human aortic endothelial cell (HAoEC) tension. The LPS effect during gram-negative sepsis on endothelial cells is cell contraction causing endothelium permeability increase. The aim was to finding out whether recombinant activated protein C (rhAPC) would reverse the endothelial cell response in an in-vitro sepsis model. In this study, the established in-vitro sepsis model was confirmed by interleukin 6 (IL-6) levels at the proteomic and genomic levels by ELISA, real time-PCR and reactive oxygen species (ROS) activation by florescence staining. The thrombin cellular contraction effect on endothelial cells was used as a positive control when the CellDrum technology was applied. Additionally, the Ras homolog gene family, member A (RhoA) mRNA expression level was checked by real time-PCR to support contractile tension results. According to contractile tension results, the mechanical predominance of actin stress fibers was a reason of the increased endothelial contractile tension leading to enhanced endothelium contractility and thus permeability enhancement. The originality of this data supports firstly the basic measurement principles of the CellDrum technology and secondly that rhAPC has a beneficial effect on sepsis influenced cellular tension. The technology presented here is promising for future high-throughput cellular tension analysis that will help identify pathological contractile tension responses of cells and prove further cell in-vitro models.
Background
Minor changes in protein structure induced by small organic and inorganic molecules can result in significant metabolic effects. The effects can be even more profound if the molecular players are chemically active and present in the cell in considerable amounts. The aim of our study was to investigate effects of a nitric oxide donor (spermine NONOate), ATP and sodium/potassium environment on the dynamics of thermal unfolding of human hemoglobin (Hb). The effect of these molecules was examined by means of circular dichroism spectrometry (CD) in the temperature range between 25°C and 70°C. The alpha-helical content of buffered hemoglobin samples (0.1 mg/ml) was estimated via ellipticity change measurements at a heating rate of 1°C/min.
Results
Major results were:
1) spermine NONOate persistently decreased the hemoglobin unfolding temperature T u irrespectively of the Na + /K + environment,
2) ATP instead increased the unfolding temperature by 3°C in both sodium-based and potassium-based buffers and
3) mutual effects of ATP and NO were strongly influenced by particular buffer ionic compositions. Moreover, the presence of potassium facilitated a partial unfolding of alpha-helical structures even at room temperature.
Conclusion
The obtained data might shed more light on molecular mechanisms and biophysics involved in the regulation of protein activity by small solutes in the cell.
Bassam, Rasha ; Artmann, Gerhard ; Hescheler, Jürgen ; Graef, T. ; Temiz Artmann, Aysegül ; Porst, Dariusz ; Linder, Peter ; Kayser, Peter ; Arinkin, Vladimir ; Gossmann, Matthias ; Digel, Ilya
The importance of the availability of stored blood or blood cells, respectively, for urgent transfusion cannot be overestimated. Nowadays, blood storage becomes even more important since blood products are used for epidemiological studies, bio-technical research or banked for transfusion purposes. Thus blood samples must not only be processed, stored, and shipped to preserve their efficacy and safety, but also all parameters of storage must be recorded and reported for Quality Assurance. Therefore, blood banks and clinical research facilities are seeking more accurate, automated means for blood storage and blood processing.
Kurulgan Demirci, Eylem ; Linder, Peter ; Demirci, Taylan ; Trzewik, Jürgen ; Digel, Ilya ; Artmann, Gerhard ; Temiz Artmann, Aysegül
Mechano-pharmacological testing of L-Type Ca²⁺ channel modulators via human vascular celldrum model
(2020)
Bayer, Robin ; Temiz Artmann, Aysegül ; Digel, Ilya ; Falkenstein, Julia ; Artmann, Gerhard ; Creutz, Till ; Hescheler, Jürgen
Background/Aims: This study aimed to establish a precise and well-defined working model, assessing pharmaceutical effects on vascular smooth muscle cell monolayer in-vitro. It describes various analysis techniques to determine the most suitable to measure the biomechanical impact of vasoactive agents by using CellDrum technology. Methods: The so-called CellDrum technology was applied to analyse the biomechanical properties of confluent human aorta muscle cells (haSMC) in monolayer. The cell generated tensions deviations in the range of a few N/m² are evaluated by the CellDrum technology. This study focuses on the dilative and contractive effects of L-type Ca²⁺ channel agonists and antagonists, respectively. We analyzed the effects of Bay K8644, nifedipine and verapamil. Three different measurement modes were developed and applied to determine the most appropriate analysis technique for the study purpose. These three operation modes are called, particular time mode" (PTM), "long term mode" (LTM) and "real-time mode" (RTM). Results: It was possible to quantify the biomechanical response of haSMCs to the addition of vasoactive agents using CellDrum technology. Due to the supplementation of 100nM Bay K8644, the tension increased approximately 10.6% from initial tension maximum, whereas, the treatment with nifedipine and verapamil caused a significant decrease in cellular tension: 10nM nifedipine decreased the biomechanical stress around 6,5% and 50nM verapamil by 2,8%, compared to the initial tension maximum. Additionally, all tested measurement modes provide similar results while focusing on different analysis parameters. Conclusion: The CellDrum technology allows highly sensitive biomechanical stress measurements of cultured haSMC monolayers. The mechanical stress responses evoked by the application of vasoactive calcium channel modulators were quantified functionally (N/m²). All tested operation modes resulted in equal findings, whereas each mode features operation-related data analysis.
Akimbekov, Nuraly S. ; Mansurov, Zulkhair ; Jandosov, J. ; Digel, Ilya ; Gossmann, Matthias ; Artmann, Gerhard ; Temiz Artmann, Aysegül ; Zhubanova, Azhar A.
The carbonized rice husk (CRH) was evaluated for its wound healing activity in rats using excision models. In this study, the influences of CRH on wound healing in rat skin in vivo and cellular behavior of human dermal fibroblasts in vitro were investigated. The obtained results showed that the CRH treatment promoted wound epithelization in rats and exhibited moderate inhibition of cell proliferation in vitro. CRH with lanolin oil treated wounds were found to epithelize faster as compared to controls.
Recently, the SHARP Corporation, Japan, has developed the world’s first "Plasma Cluster Ions (PCI)" air purification technology using plasma discharge to generate cluster ions. The new plasma cluster device releases positive and negative ions into the air, which are able to decompose and deactivate harmful airborne substances by chemical reactions. Because cluster ions consist of positive and negative ions that normally exist in the natural world, they are completely harmless and safe to humans. The amount of ozone generated by cluster ions is less than 0.01 ppm, which is significantly less than the 0.05-ppm standard for industrial operations and consumer electronics. This amount, thus, has no harming effects whatsoever on the human body. But particular properties and chemical processes in PCI treatment are still under study. It has been shown that PCI in most cases show strongly pronounced irreversible killing effects in respect of airborne microflora due to free-radical induced reactions and can be considered as a potent technology to disinfect both home, medical and industrial appliances.