Combined Impact of Magnetic Force and Spaceflight Conditions on Escherichia coli Physiology.

Changes in bacterial physiology caused by the combined action of the magnetic force and microgravity were studied in Escherichia coli grown using a specially developed device aboard the International Space Station. The morphology and metabolism of E. coli grown under spaceflight (SF) or combined spaceflight and magnetic force (SF + MF) conditions were compared with ground cultivated bacteria grown under standard (control) or magnetic force (MF) conditions. SF, SF + MF, and MF conditions provided the up-regulation of Ag43 auto-transporter and cell auto-aggregation. The magnetic force caused visible clustering of non-sedimenting bacteria that formed matrix-containing aggregates under SF + MF and MF conditions. Cell auto-aggregation was accompanied by up-regulation of glyoxylate shunt enzymes and Vitamin B12 transporter BtuB. Under SF and SF + MF but not MF conditions nutrition and oxygen limitations were manifested by the down-regulation of glycolysis and TCA enzymes and the up-regulation of methylglyoxal bypass. Bacteria grown under combined SF + MF conditions demonstrated superior up-regulation of enzymes of the methylglyoxal bypass and down-regulation of glycolysis and TCA enzymes compared to SF conditions, suggesting that the magnetic force strengthened the effects of microgravity on the bacterial metabolism. This strengthening appeared to be due to magnetic force-dependent bacterial clustering within a small volume that reinforced the effects of the microgravity-driven absence of convectional flows.

Heterotypic Multicellular Spheroids as Experimental and Preclinical Models of Sprouting Angiogenesis.

Sprouting angiogenesis is the common response of live tissues to physiological and pathological angiogenic stimuli. Its accurate evaluation is of utmost importance for basic research and practical medicine and pharmacology and requires adequate experimental models. A variety of assays for angiogenesis were developed, none of them perfect. In vitro approaches are generally less physiologically relevant due to the omission of essential components regulating the process. However, only in vitro models can be entirely non-xenogeneic. The limitations of the in vitro angiogenesis assays can be partially overcome using 3D models mimicking tissue O2 and nutrient gradients, the influence of the extracellular matrix (ECM), and enabling cell-cell interactions. Here we present a review of the existing models of sprouting angiogenesis that are based on the use of endothelial cells (ECs) co-cultured with perivascular or other stromal cells. This approach provides an excellent in vitro platform for further decoding of the cellular and molecular mechanisms of sprouting angiogenesis under conditions close to the in vivo conditions, as well as for preclinical drug testing and preclinical research in tissue engineering and regenerative medicine.

Scaffold-free, Label-free, and Nozzle-free Magnetic Levitational Bioassembler for Rapid Formative Biofabrication of 3D Tissues and Organs.

Scaffolding is the conceptual framework of conventional tissue engineering. Over the past decade, scaffold-free approaches as a potential alternative to classic scaffold-based methods have emerged, and scaffold-free magnetic levitational tissue engineering (magnetic force-based tissue engineering [Mag-TE]) is a type of this novel tissue engineering strategy. However, Mag-TE is often based on the use of potentially toxic magnetic nanoparticles. Scaffold-free and label-free magnetic levitational bioassembly do not employ magnetic nanoparticles and thus, the potential toxicity of magnetic nanoparticles can be avoided. In this short review, we describe the conceptual foundation of scaffold-free, label-free, and nozzle-free formative biofabrication using magnetic fields as "scaffields." The design and implementation of "Organ.Aut," the first commercial magnetic levitational bioassembler, and the potential applications of magnetic bioassembler are discussed as well.

Breast cancer organoid model allowed to reveal potentially beneficial combinations of 3,3'-diindolylmethane and chemotherapy drugs.

Epigenetic alterations represent promising therapeutic targets in cancer treatment. Recently it was revealed that small molecules have the potential to act as microRNA silencers. Capacity to bind the discrete stem-looped structure of pre-miR-21 and prevent its maturation opens opportunities to utilize such compounds for the prevention of initiation, progression, and chemoresistance of cancer. Molecular simulations performed earlier identified 3,3'-diindolylmethane (DIM) as a potent microRNA-21 antagonist. However, data on DIM and microRNA-21 interplay is controversial, which may be caused by the limitations of the cell lines.

Magnetic levitational bioassembly of 3D tissue construct in space.

Magnetic levitational bioassembly of three-dimensional (3D) tissue constructs represents a rapidly emerging scaffold- and label-free approach and alternative conceptual advance in tissue engineering. The magnetic bioassembler has been designed, developed, and certified for life space research. To the best of our knowledge, 3D tissue constructs have been biofabricated for the first time in space under microgravity from tissue spheroids consisting of human chondrocytes. Bioassembly and sequential tissue spheroid fusion presented a good agreement with developed predictive mathematical models and computer simulations. Tissue constructs demonstrated good viability and advanced stages of tissue spheroid fusion process. Thus, our data strongly suggest that scaffold-free formative biofabrication using magnetic fields is a feasible alternative to traditional scaffold-based approaches, hinting a new perspective avenue of research that could significantly advance tissue engineering. Magnetic levitational bioassembly in space can also advance space life science and space regenerative medicine.

Biofabrication of a Functional Tubular Construct from Tissue Spheroids Using Magnetoacoustic Levitational Directed Assembly.

In traditional tissue engineering, synthetic or natural scaffolds are usually used as removable temporal support, which involves some biotechnology limitations. The concept of "scaffield" approach utilizing the physical fields instead of biomaterial scaffold has been proposed recently. In particular, a combination of intense magnetic and acoustic fields can enable rapid levitational bioassembly of complex-shaped 3D tissue constructs from tissue spheroids at low concentration of paramagnetic agent (gadolinium salt) in the medium. In the current study, the tissue spheroids from human bladder smooth muscle cells (myospheres) are used as building blocks for assembling the tubular 3D constructs. Levitational assembly is accomplished at low concentrations of gadolinium salts in the high magnetic field at 9.5 T. The biofabricated smooth muscle constructs demonstrate contraction after the addition of vasoconstrictive agent endothelin-1. Thus, hybrid magnetoacoustic levitational bioassembly is considered as a new technology platform in the emerging field of formative biofabrication. This novel technology of scaffold-free, nozzle-free, and label-free bioassembly opens a unique opportunity for rapid biofabrication of 3D tissue and organ constructs with complex geometry.

Fabrication of calcium phosphate 3D scaffolds for bone repair using magnetic levitational assembly.

The calcium phosphate particles can be used as building blocks for fabrication of 3D scaffolds intended for bone tissue engineering. This work presents for the first time a rapid creation of 3D scaffolds using magnetic levitation of calcium phosphate particles. Namely, tricalcium phosphate particles of equal size and certain porosity are used, which undergo the process of recrystallization after magnetic levitational assembly of the scaffold to ensure stitching of the scaffold. Label-free levitational assembly is achieved by using a custom-designed magnetic system in the presence of gadolinium salts, which allows the levitation of calcium phosphate particles. Chemical transformation of tricalcium- to octacalcium phosphate under the condition of magnetic levitation in non-homogeneous magnetic field is also demonstrated. This approach allows obtaining rapidly the octacalcium phosphate phase in the final 3D product, which is biocompatible.

Scaffold-free and label-free biofabrication technology using levitational assembly in a high magnetic field.

The feasibility of magnetic levitational bioassembly of tissue-engineered constructs from living tissue spheroids in the presence of paramagnetic ions (i.e. Gd3+) was recently demonstrated. However, Gd3+ is relatively toxic at concentrations above 50 mM normally used to enable magnetic levitation with NdFeB-permanent magnets. Using a high magnetic field (a 50 mm-bore, 31 T Bitter magnet) at the High Field Magnet Laboratory at Radboud University in Nijmegen, The Netherlands, we performed magnetic levitational assembly of tissue constructs from living spheroids prepared from the SW1353 chondrosarcoma cell line at 0.8 mM Gd3+ containing salt gadobutrol at 19 T magnetic field. The parameters of the levitation process were determined on the basis of polystyrene beads with a 170 µm-diameter. To predict the theoretical possibility of assembly, a zone of stable levitation in the horizontal and vertical areas of cross sections was previously calculated. The construct from tissue spheroids partially fused after 3 h in levitation. The analysis of viability after prolonged exposure (1 h) to strong magnetic fields (up to 30 T) showed the absence of significant cytotoxicity or morphology changes in the tissue spheroids. A high magnetic field works as a temporal and removal support or so-called 'scaffield'. Thus, formative biofabrication of tissue-engineered constructs from tissue spheroids in the high magnetic field is a promising research direction.