A. D. van der Meer

Microfluidic Technology in Vascular Research

Vascular cell biology is an area of research with great biomedical relevance. Vascular dysfunction is involved in major diseases such as atherosclerosis, diabetes, and cancer. However, when studying vascular cell biology in the laboratory, it is difficult to mimic the dynamic, three-dimensional microenvironment that is found in vivo. Microfluidic technology offers unique possibilities to overcome this difficulty. In this review, an overview of the recent applications of microfluidic technology in the field of vascular biological research will be given. Examples of how microfluidics can be used to generate shear stresses, growth factor gradients, cocultures, and migration assays will be provided. The use of microfluidic devices in studying three-dimensional models of vascular tissue will be discussed. It is concluded that microfluidic technology offers great possibilities to systematically study vascular cell biology with setups that more closely mimic the in vivo situation than those that are generated with conventional methods.

Analyzing shear stress-induced alignment of actin filaments in endothelial cells with a microfluidic assay

The physiology of vascular endothelial cells is strongly affected by fluid shear stress on their surface. In this study, a microfluidic assay was employed to analyze the alignment of actin filaments in endothelial cells in response to shear stress. When cells were cultured in microfluidic channels and subjected to shear stress, the alignment of filaments in the channel direction was significantly higher than in static cultures. By adding inhibitory drugs, the roles of several signaling proteins in the process of alignment were determined. Thus, it is shown how microfluidic technology can be employed to provide a mechanistic insight into cell physiology.

The uptake and distribution of cadmium in tomato plants as affected by ethylenediaminetetraacetic acid and 2,4-dinitrophenol

The uptake and distribution of cadmium in tomato plants (Lycopersicon esculentum, Mill, cv. Tiny Tim) were examined with and without the presence of ethylenediaminetetraacetic acid (EDTA) as chelating agent and 2,4-dinitrophenol (DNP) as metabolic inhibitor. Eight-week-old intact and derooted tomato seedlings were used in hydroculture experiments with cadmium applied as (115)Cd(NO(3))(2) in a range of concentrations. Measurements of the (115)Cd content of roots, stems and leaves were carried out by gamma-ray spectroscopy. The data showed that applications of both EDTA and DNP resulted in reduced total Cd accumulation in the plants, but relatively enhanced Cd transport into the above-ground plant parts. The Cd mobility in the transport channels in the shoots was increased by EDTA in both intact and derooted plants. Application of DNP leads to increased relative Cd import to leaves in derooted plants, but a reduced import into leaves of intact plants. These results suggest that Cd-complexes are formed in root cells before root-to-shoot transport. Furthermore, initial Cd uptake may be associated with adsorption on the negative charges of the cell walls of the root system. The high Cd mobility in shoots, in experiments with intact plants and Cd-EDTA application, indicates the possibility of simultaneous uptake of Cd and EDTA, possibly as a Cd-EDTA complex.

Flow cytometric analysis of the uptake of low-density lipoprotein by endothelial cells in microfluidic channels

Acceptance of microfluidic technology in everyday laboratory practice by biologists is still low. One of the reasons for this is that the technology combines poorly with standard cell biological and biochemical analysis tools. Flow cytometry is an example of a conventional analytical tool that is considered to be incompatible with microfluidic technology and its inherent small sample sizes. In this study, it is shown that properly designed microfluidic devices contain cell populations that are large enough to be analyzed by flow cytometry. To illustrate this, the uptake of fluorescent human low‐density lipoprotein (LDL) by human endothelial cells that were cultured in a microfluidic channel was analyzed. It was found that the uptake of LDL by the cells increased linearly over time. Moreover, the uptake decreased when cells were pretreated with fluid shear stress inside the microfluidic devices. This study shows that microfluidic technology can be combined with conventional flow cytometry, while retaining the advantages of working with microfluidics such as low reagent use and dynamic cell culture conditions. This approach of combining microfluidic technology with conventional laboratory tools may contribute to greater acceptance of microfluidic devices in biological research.

Shear stress induces a transient and VEGFR-2-dependent decrease in the motion of injected particles in endothelial cells

Vascular endothelial cells form the inner lining of all blood vessels and play a central role in vessel physiology and disease. Endothelial cells are highly responsive to the mechanical stimulus of fluid shear stress that is exerted by blood flowing over their surface. In this study, the immediate micromechanical response of endothelial cells to physiological shear stress was characterized by tracking of ballistically injected, sub-micron, fluorescent particles. It was found that the mean squared displacement (MSD) of the particles decreases by a factor 1.5 within 10 min after the onset of shear stress. This decrease in particle motion is transient, since the MSD returns to control values within 15-30 min after the onset of shear. The immediate micromechanical stiffening is dependent on activation of the vascular endothelial growth factor receptor (VEGFR)-2, because inhibition of the receptor abrogates the micromechanical response. This work shows that the cytoskeleton is actively involved in the acute, functional response of endothelial cells to shear stress.

Shear-sensitive nanocapsule drug release for site-specific inhibition of occlusive thrombus formation

Essentials Vessel stenosis due to large thrombus formation increases local shear 1‐2 orders of magnitude. High shear at stenotic sites was exploited to trigger eptifibatide release from nanocapsules. Local delivery of eptifibatide prevented vessel occlusion without increased tail bleeding times. Local nanocapsule delivery of eptifibatide may be safer than systemic antiplatelet therapies.

Microfabricated tuneable and transferable porous PDMS membranes for Organs-on-Chips

We present a novel and highly reproducible process to fabricate transferable porous PDMS membranes for PDMS-based Organs-on-Chips (OOCs) using microelectromechanical systems (MEMS) fabrication technologies. Porous PDMS membranes with pore sizes down to 2.0 μm in diameter and a wide porosity range (2–65%) can be fabricated. To overcome issues normally faced when using replica moulding and extend the applicability to most OOCs and improve their scalability and reproducibility, the process includes a sacrificial layer to easily transfer the membranes from a silicon carrier to any PDMS-based OOC. The highly reliable fabrication and transfer method does not need of manual handling to define the pore features (size, distribution), allowing very thin (<10 μm) functional membranes to be transferred at chip level with a high success rate (85%). The viability of cell culturing on the porous membranes was assessed by culturing two different cell types on transferred membranes in two different OOCs. Human umbilical endothelial cells (HUVEC) and MDA-MB-231 (MDA) cells were successfully cultured confirming the viability of cell culturing and the biocompatibility of the membranes. The results demonstrate the potential of controlling the porous membrane features to study cell mechanisms such as transmigrations, monolayer formation, and barrier function. The high control over the membrane characteristics might consequently allow to intentionally trigger or prevent certain cellular responses or mechanisms when studying human physiology and pathology using OOCs.

Separation of nuclear isomers for cancer therapeutic radionuclides based on nuclear decay after-effects

177Lu has sprung as a promising radionuclide for targeted therapy. The low soft tissue penetration of its β− emission results in very efficient energy deposition in small-size tumours. Because of this, 177Lu is used in the treatment of neuroendocrine tumours and is also clinically approved for prostate cancer therapy. In this work, we report a separation method that achieves the challenging separation of the physically and chemically identical nuclear isomers, 177mLu and 177Lu. The separation method combines the nuclear after-effects of the nuclear decay, the use of a very stable chemical complex and a chromatographic separation. Based on this separation concept, a new type of radionuclide generator has been devised, in which the parent and the daughter radionuclides are the same elements. The 177mLu/177Lu radionuclide generator provides a new production route for the therapeutic radionuclide 177Lu and can bring significant growth in the research and development of 177Lu based pharmaceuticals.