High-Scale 3D-Bioprinting Platform for the Automated Production of Vascularized Organs-on-a-Chip.

3D bioprinting possesses the potential to revolutionize contemporary methodologies for fabricating tissue models employed in pharmaceutical research and experimental investigations. This is enhanced by combining bioprinting with advanced organs-on-a-chip (OOCs), which includes a complex arrangement of multiple cell types representing organ-specific cells, connective tissue, and vasculature. However, both OOCs and bioprinting so far demand a high degree of manual intervention, thereby impeding efficiency and inhibiting scalability to meet technological requirements. Through the combination of drop-on-demand bioprinting with robotic handling of microfluidic chips, a print procedure is achieved that is proficient in managing three distinct tissue models on a chip within only a minute, as well as capable of consecutively processing numerous OOCs without manual intervention. This process rests upon the development of a post-printing sealable microfluidic chip, that is compatible with different types of 3D-bioprinters and easily connected to a perfusion system. The capabilities of the automized bioprint process are showcased through the creation of a multicellular and vascularized liver carcinoma model on the chip. The process achieves full vascularization and stable microvascular network formation over 14 days of culture time, with pronounced spheroidal cell growth and albumin secretion of HepG2 serving as a representative cell model.

Reversible Host-Guest Crosslinks in Supramolecular Hydrogels for On-Demand Mechanical Stimulation of Human Mesenchymal Stem Cells.

Stem cells are regulated not only by biochemical signals but also by biophysical properties of extracellular matrix (ECM). The ECM is constantly monitored and remodeled because the fate of stem cells can be misdirected when the mechanical interaction between cells and ECM is imbalanced. A well-defined ECM model for bone marrow-derived human mesenchymal stem cells (hMSCs) based on supramolecular hydrogels containing reversible host-guest crosslinks is fabricated. The stiffness (Young's modulus E) of the hydrogels can be switched reversibly by altering the concentration of non-cytotoxic, free guest molecules dissolved in the culture medium. Fine-adjustment of substrate stiffness enables the authors to determine the critical stiffness level E* at which hMSCs turn the mechano-sensory machinery on or off. Next, the substrate stiffness across E* is switched and the dynamic adaptation characteristics such as morphology, traction force, and YAP/TAZ signaling of hMSCs are monitored. These data demonstrate the instantaneous switching of traction force, which is followed by YAP/TAZ signaling and morphological adaptation. Periodical switching of the substrate stiffness across E* proves that frequent applications of mechanical stimuli drastically suppress hMSC proliferation. Mechanical stimulation across E* level using dynamic hydrogels is a promising strategy for the on-demand control of hMSC transcription and proliferation.

Wnt/ß-catenin signaling induces axial elasticity patterns of Hydra extracellular matrix.

The extracellular matrix (ECM) plays crucial roles in animal development and diseases. Here, we report that Wnt/ß-catenin signaling induces the ECM remodeling during Hydra axis formation. We determined the micro- and nanoscopic arrangement of fibrillar type I collagen along Hydra's body axis using high-resolution microscopy and X-ray scattering. Elasticity mapping of the ECM ex vivo revealed distinctive elasticity patterns along the body axis. A proteomic analysis of the ECM showed that these elasticity patterns correlate with a gradient-like distribution of metalloproteases along the body axis. Activation of the Wnt/ß-catenin pathway in wild-type and transgenic animals alters these patterns toward low ECM elasticity patterns. This suggests a mechanism whereby high protease activity under control of Wnt/ß-catenin signaling causes remodeling and softening of the ECM. This Wnt-dependent spatiotemporal coordination of biochemical and biomechanical cues in ECM formation was likely a central evolutionary innovation for animal tissue morphogenesis.

Iron-Catalyzed Wacker-type Oxidation of Olefins at Room Temperature with 1,3-Diketones or Neocuproine as Ligands*.

Herein, we describe a convenient and general method for the oxidation of olefins to ketones using either tris(dibenzoylmethanato)iron(III) [Fe(dbm)3 ] or a combination of iron(II) chloride and neocuproine (2,9-dimethyl-1,10-phenanthroline) as catalysts and phenylsilane (PhSiH3 ) as additive. All reactions proceed efficiently at room temperature using air as sole oxidant. This transformation has been applied to a variety of substrates, is operationally simple, proceeds under mild reaction conditions, and shows a high functional-group tolerance. The ketones are formed smoothly in up to 97 % yield and with 100 % regioselectivity, while the corresponding alcohols were observed as by-products. Labeling experiments showed that an incorporated hydrogen atom originates from the phenylsilane. The oxygen atom of the ketone as well as of the alcohol derives from the ambient atmosphere.

New Class of Crosslinker-Free Nanofiber Biomaterials from Hydra Nematocyst Proteins.

Nematocysts, the stinging organelles of cnidarians, have remarkable mechanical properties. Hydra nematocyst capsules undergo volume changes of 50% during their explosive exocytosis and withstand osmotic pressures of beyond 100 bar. Recently, two novel protein components building up the nematocyst capsule wall in Hydra were identified. The cnidarian proline-rich protein 1 (CPP-1) characterized by a "rigid" polyproline motif and the elastic Cnidoin possessing a silk-like domain were shown to be part of the capsule structure via short cysteine-rich domains that spontaneously crosslink the proteins via disulfide bonds. In this study, recombinant Cnidoin and CPP-1 are expressed in E. coli and the elastic modulus of spontaneously crosslinked bulk proteins is compared with that of isolated nematocysts. For the fabrication of uniform protein nanofibers by electrospinning, the preparative conditions are systematically optimized. Both fibers remain stable even after rigorous washing and immersion into bulk water owing to the simultaneous crosslinking of cysteine-rich domains. This makes our nanofibers clearly different from other protein nanofibers that are not stable without chemical crosslinkers. Following the quantitative assessment of mechanical properties, the potential of Cnidoin and CPP-1 nanofibers is examined towards the maintenance of human mesenchymal stem cells.

The Impact of α-Defensin Test in Diagnosing Periprosthetic Infection After Total Ankle Arthroplasty.

Periprosthetic joint infection (PJI) after total ankle arthroplasty (TAA) is a serious complication, and a reliable diagnostic test to identify PJI is needed. The purpose of this study was to investigate the use of synovial α-defensin levels in identifying PJI of the ankle. Data from 33 patients were retrospectively collected between September 2015 and May 2018. Patients who had pain or suspected loosening after TAA and who had undergone joint aspiration were included in the study. Aspiration was performed in a semisterile theatre. Synovial fluid was processed in descending order for microbiological cultures, α-defensin, leukocyte esterase strip test, and cell count. A periprosthetic infection was defined by Musculoskeletal Infection Society criteria. The sensitivity, specificity, and overall accuracy were calculated, and based on a receiver operating characteristic curve, the quality of the α-defensin test was determined. The calculated area under the curve was 0.97 ± 0.32. Two of 33 patients fulfilled the 2014 Musculoskeletal Infection Society criteria and were scheduled for septic revision arthroplasty. Sensitivity, specificity, and overall accuracy of the α-defensin test were 100% (95% confidence interval [CI], 15.8% to 100%), 93.5% (95% CI, 78.6% to 99.2%), and 93.9% (95% CI, 79.8% to 99.3%), respectively. The positive predictive value was 50% (95% CI, 20.7% to 79.3%), and the negative predictive value was 100%. The α-defensin test seems to be the best available synovial test to detect a late-onset PJI after total ankle arthroplasty. Further prospective studies with a larger number of patients are required.

Dynamic Contact Guidance of Myoblasts by Feature Size and Reversible Switching of Substrate Topography: Orchestration of Cell Shape, Orientation, and Nematic Ordering of Actin Cytoskeletons.

Biological cells in tissues alter their shapes, positions, and orientations in response to dynamic changes in their physical microenvironments. Here, we investigated the dynamic response of myoblast cells by fabricating substrates displaying microwrinkles that can reversibly change their direction within 60 s by axial compression and relaxation. To quantitatively assess the collective order of cells, we introduced the nematic order parameter of cells that takes not only the distribution of cell-wrinkle angles but also the degree of cell elongation into account. On the subcellular level, we also calculated the nematic order parameter of actin cytoskeletons that takes the rearrangement of actin filaments into consideration. The results obtained on substrates with different wrinkle wavelengths implied the presence of a characteristic wavelength beyond which the order parameters of both cells and actin cytoskeletons level off. Immunofluorescence labeling of vinculin showed that the focal adhesions were all concentrated on the peaks of wrinkles when the wavelength is below the characteristic value. On the other hand, we found focal adhesions on both the peaks and the troughs of wrinkles when the wavelength exceeds the characteristic level. The emergence of collective ordering of cytoskeletons and the adaptation of cell shapes and orientations were monitored by live cell imaging after the seeding of cells from suspensions. After the cells had reached the steady state, the orientation of wrinkles was abruptly changed by 90°. The dynamic response of myoblasts to the drastic change in surface topography was monitored, demonstrating the coordination of the shape and orientation of cells and the nematic ordering of actin cytoskeletons. The "dynamic" substrates established in this study can be used as a powerful tool in mechanobiology that helps us understand how cytoskeletons, cells, and cell ensembles respond to dynamic contact guidance cues.

Author Correction: Dynamic Mechano-Regulation of Myoblast Cells on Supramolecular Hydrogels Cross-Linked by Reversible Host-Guest Interactions.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

Dynamic Mechano-Regulation of Myoblast Cells on Supramolecular Hydrogels Cross-Linked by Reversible Host-Guest Interactions.

A new class of supramolecular hydrogels, cross-linked by host-guest interactions between ß-cyclodextrin (ßCD) and adamantane, were designed for the dynamic regulation of cell-substrate interactions. The initial substrate elasticity can be optimized by selecting the molar fraction of host- and guest monomers for the target cells. Moreover, owing to the reversible nature of host-guest interactions, the magnitude of softening and stiffening of the substrate can be modulated by varying the concentrations of free, competing host molecules (ßCD) in solutions. By changing the substrate elasticity at a desired time point, it is possible to switch the micromechanical environments of cells. We demonstrated that the Young's modulus of our "host-guest gels", 4-11 kPa, lies in an optimal range not only for static (ex situ) but also for dynamic (in situ) regulation of cell morphology and cytoskeletal ordering of myoblasts. Compared to other stimulus-responsive materials that can either change the elasticity only in one direction or rely on less biocompatible stimuli such as UV light and temperature change, our supramolecular hydrogel enables to reversibly apply mechanical cues to various cell types in vitro without interfering cell viability.