Fibroblasts can condense a hydrated collagen lattice to a tissue-like structure 1/28th the area of the starting gel in 24 hr. The rate of the process can be regulated by varying the protein content of the lattice, the cell number, or the con- centration of an inhibitor such as Colcemid. Fibroblasts of high population doubling level propagated in vitro, which have left the cell cycle, can carry out the contraction at least as efficiently as cycling cells. The potential uses of the system as an immu- nologically tolerated "tissue" for wound hea ing and as a model for studying fibroblast function are discussed.

Production of a tissue-like structure by contraction of collagen lattices by human fibroblasts of different proliferative

Transepithelial/transendothelial electrical resistance (TEER) is a widely accepted quantitative technique to measure the integrity of tight junction dynamics in cell culture models of endothelial and epithelial monolayers. TEER values are strong indicators of the integrity of the cellular barriers before they are evaluated for transport of drugs or chemicals. TEER measurements can be performed in real time without cell damage and generally are based on measuring ohmic resistance or measuring impedance across a wide spectrum of frequencies. The measurements for various cell types have been reported with commercially available measurement systems and also with custom-built microfluidic implementations. Some of the barrier models that have been widely characterized using TEER include the blood–brain barrier (BBB), gastrointestinal (GI) tract, and pulmonary models. Variations in these values can arise due to factors such as temperature, medium formulation, and passage number of cells. The aim of this article ...

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TEER Measurement Techniques for In Vitro Barrier Model Systems

Lymphatic metastasis contributes to mortality from solid tumors. Whether metastasizing cancer cells reach lymph nodes via intratumor lymphatic vessels is unknown. Here, we examine functional lymphatics associated with mouse tumors expressing normal or elevated levels of vascular endothelial growth factor-C (VEGF-C), a molecule that stimulates lymphangiogenesis. Although VEGF-C overexpression increased lymphatic surface area in the tumor margin and lymphatic metastasis, these tumors contained no functional lymphatics, as assessed by four independent functional assays and immunohistochemical staining. These findings suggest that the functional lymphatics in the tumor margin alone are sufficient for lymphatic metastasis and should be targeted therapeutically.

https://science.sciencemag.org/content/sci/296/5574/1883.full.pdf

Lymphatic metastasis in the absence of functional intratumor lymphatics.

In salt-sensitive hypertension, the accumulation of Na(+) in tissue has been presumed to be accompanied by a commensurate retention of water to maintain the isotonicity of body fluids We show here that a high-salt diet (HSD) in rats leads to interstitial hypertonic Na(+) accumulation in skin, resulting in increased density and hyperplasia of the lymphcapillary network The mechanisms underlying these effects on lymphatics involve activation of tonicity-responsive enhancer binding protein (TonEBP) in mononuclear phagocyte system (MPS) cells infiltrating the interstitium of the skin TonEBP binds the promoter of the gene encoding vascular endothelial growth factor-C (VEGF-C, encoded by Vegfc) and causes VEGF-C secretion by macrophages MPS cell depletion or VEGF-C trapping by soluble VEGF receptor-3 blocks VEGF-C signaling, augments interstitial hypertonic volume retention, decreases endothelial nitric oxide synthase expression and elevates blood pressure in response to HSD Our data show that TonEBP-VEGF-C signaling in MPS cells is a major determinant of extracellular volume and blood pressure homeostasis and identify VEGFC as an osmosensitive, hypertonicity-driven gene intimately involved in salt-induced hypertension

Macrophages regulate salt-dependent volume and blood pressure by a vascular endothelial growth factor-C–dependent buffering mechanism

Recirculation of fluid and cells through lymphatic vessels plays a key role in normal tissue homeostasis, inflammatory diseases, and cancer. Despite recent advances in understanding lymphatic function (Alitalo, K., T. Tammela, and T.V. Petrova. 2005. Nature. 438:946–953), the cellular features responsible for entry of fluid and cells into lymphatics are incompletely understood. We report the presence of novel junctions between endothelial cells of initial lymphatics at likely sites of fluid entry. Overlapping flaps at borders of oak leaf–shaped endothelial cells of initial lymphatics lacked junctions at the tip but were anchored on the sides by discontinuous button-like junctions (buttons) that differed from conventional, continuous, zipper-like junctions (zippers) in collecting lymphatics and blood vessels. However, both buttons and zippers were composed of vascular endothelial cadherin (VE-cadherin) and tight junction–associated proteins, including occludin, claudin-5, zonula occludens–1, junctional adhesion molecule–A, and endothelial cell–selective adhesion molecule. In C57BL/6 mice, VE-cadherin was required for maintenance of junctional integrity, but platelet/endothelial cell adhesion molecule–1 was not. Growing tips of lymphatic sprouts had zippers, not buttons, suggesting that buttons are specialized junctions rather than immature ones. Our findings suggest that fluid enters throughout initial lymphatics via openings between buttons, which open and close without disrupting junctional integrity, but most leukocytes enter the proximal half of initial lymphatics.

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Functionally specialized junctions between endothelial cells of lymphatic vessels

The differentiation between discocytic and stomatocytic red blood cell (RBC) shape using conventional microscopic imaging and image analysis tools is still on a very poor level. A procedure to differentiate the degree of stomatocytic shape changes was developed. We obtained multiple microscopic images of the same RBCs settled on a human albumin coated cover slip. The images were acquired when the microscope objective was subsequently focused through the cell layer. At equidistant horizontal planes (z-axis) below, within, and above the microscopic focal plane the light intensity distribution was considered. Using a model based on light refraction, we calculated the intensity distribution of the planes which are out of focus. Using this tool we are able to differentiate RBC shapes precisely. On the other hand, using this model from and the light intensity distributions of different focal planes, we are able to reconstruct the shape of one single RBC located in the optical axis of the microscope.© (1995) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.

Shape quantification of single red blood cells based on their scattering patterns from microscopic images

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.

Mechano-Pharmacological Testing of L-Type Ca2+ Channel Modulators via Human Vascular Celldrum Model.

Mechanotransduction-bioreactor For Tissue Engineering Of A Ureter Prosthesis

Autonomous robotic systems for penetrating thick ice shells with simultaneous collecting of scientific data are very promising devices in both terrestrial (glacier, climate research) and extra-terrestrial applications. Technical challenges in development of such systems are numerous and include 3D-navigation, an appropriate energy source, motion control, etc. Not less important is the problem of forward contamination of the pristine glacial environments with microorganisms and biomolecules from the surface of the probe. This study was devoted to establishing a laboratory model for microbial contamination of a newly-constructed ice-melting probe called IceMole and to analyse the viability and amount of the contaminating microorganisms as a function of distance. The used bacterial strains were Bacillus subtilis (ATCC 6051) and Escherichia coli (ATCC 11775). The main objective was development of an efficient and reliable in-situ decontamination method of the melting probe. Therefore, several chemical substances were tested in respect of their efficacy to eliminate bacteria on the surface of the melting probe at low temperature (0 - 5°C) and at continuous dilution by melted water. Our study has shown that at least 99.9% decontamination of the IceMole can be successfully achieved by the injection of 30% (v/v) hydrogen peroxide and 3% (v/v) sodium hypochlorite into the drilling site. We were able to reproduce this result in both time-dependent and depth-dependent experiments. The sufficient amount of 30% (v/v) H 2 O 2 or 3% (v/v) NaClO has been found to be approximately 18 μL per cm² of the probe’s surface.

Feasibility of an In-Situ Microbial Decontamination of an Ice-Melting Probe

Kardiopulmonare Bypassoperationen (KPB) verursachen Reaktionen des Immunsystems verbunden mit einem erhöhten Gefäßwiderstand. Bei KPB unter milden Temperaturen steigt die Prostaglandin E2konzentration (PGE2) im Blut schneller als unter Hypothermie bei 28°C, verbunden mit einem niedrigeren systemischen Gefäßwiderstand [1]. Neben den vasodilatativen Effekten von PGE2 könnten temperaturabhängige Unterschiede in der Pseudopodienbildung und rückbildung polymorphonuklearer Leukozyten (PMNs) zum erhöhten systemischen Gefäßwiderstand unter hypothermen KPB beitragen. Die Temperaturabhängigkeit der Bildung bzw. Rückbildung der Pseudopodien war Gegenstand dieser Studie. Als Modell für die Aktivierung der PMNs benutzten wir das Chemoattraktant N-formyl-methionyl-leucyl-phenylalanine (fMLP). Die Zellen bilden als Antwort Pseudopodien in Richtung des Konzentrationsgradienten des fMLP [2].

Gerhard M. Artmann

Papers

Shape quantification of single red blood cells based on their scattering patterns from microscopic images

Mechano-Pharmacological Testing of L-Type Ca2+ Channel Modulators via Human Vascular Celldrum Model.

Mechanotransduction-bioreactor For Tissue Engineering Of A Ureter Prosthesis

Feasibility of an In-Situ Microbial Decontamination of an Ice-Melting Probe

Temperaturabhängigkeit der pseudopodienbildung und rückbildung von granulozyten