Control of cross-linking density in bioinks and integration of nanotechnology

Biofabrication is a new field of research where cells are fabricated together with materials in automated processes to 3D constructs with stratified organization [1]. Lately, the need for a broader variety of bioinks has generated considerable research interest [2]. We have in this context explored physically cross-linked hydrogels [3] as well as thiol-ene clickable artificial systems based on polyglycidols [4]. In order to transfer this to more biologically active bioinks, we have recently prepared and thoroughly characterized thiol-ene clickable gelatin (GelAGE) as bioink. The first part of the lecture will thus compare standard methacrylated gelatin (GelMA) with GelAGE especiall with regards to the formation of high molecular weight polyacrylates in the case of GelMA. Moreover, we show that the controlled network formation of GelAGE enables the system to serve as a platform bioink for several Biofabrication technologies, and we demonstrate this at the examples of extrusion printing and digital light processing. Aside the pure printability, a control of drug loading and release into bioinks is of interest in order to accelerate tissue maturation or for drug testing. This may be achieved by supplementation of bioinks with nanoparticles. Mesoporous silica nanoparticles (MSN) for example can be loaded with drugs and designed to only release their payload after cell internalization [5]. The second part of the talk will concern MSN with diameters of 350 nm as well as gold nanoparticles with a diameter of 30 nm as model systems. Both particle types were prepared with either positive or negative surface charge and formulated into a thiol-ene clickable bioink comprising negatively charged Hyaluronic acid [6]. Rheological experiments show that both particle types can be supplemented in concentrations up to 10 mg/mL without affecting printability. Our data quantitatively shows that electrostatic interactions can be used to control the migration and release behavior of nanoparticles in and from printed hydrogels, and the subsequent uptake by cells. These results display a promising approach towards the local and temporal control of drug vectors in Biofabrication through a combination of bioink development with nanotechnology using a generic principle. References: 1. Groll J, et al. Biofabrication 2016;8: 013001. 2. Jungst T, et al. Chem. Rev. 2016;116:1496–1539 3. Schacht K, et al. Angew. Chem. Int. Ed. 2015;54:2816–2820. 4. Stichler S, et al. Ann. Biomed. Eng. 2017;45:273-285. 5. Bocking D, et al. Nanoscale 2014;6:1490-1498. 6. Jungst, T, et al. Angew. Chem. Int. Ed. 2017, DOI: 10.1002/anie.201700153

Two-step screening process to evaluate printability of inks for extrusion-based bioprinting

One of the key bottlenecks of biofabrication are suitable ink materials. These materials need to meet various requirements and combine good printing properties with adequate cytocompatibility. Many scientists try to overcome this bottleneck by designing novel inks. Their approaches range from multimaterial inks to alternative crosslinking strategies. To encourage efficient application of new strategies, we developed a two-step assessment (Fig. 1) focusing on ink printability with extrusion-based bioprinters. The first step only requires a syringe, a nozzle and the material. We have shown that simple screening based on fiber formation and layer stacking properties was an efficient method to assess printability. In a second step, the materials were transferred to a rheometer and evaluated regarding their shear thinning and post-printing recovery properties. We demonstrated that mathematical modelling of the extrusion process can help gaining deeper understanding of the material behavior and enables better evaluation by estimating the conditions present in the nozzle. The data from the shear viscosity plot was fitted and the shear-rate, extrusion velocity, shear stress and residence time profiles for the conditions present in the nozzle during dispensing were calculated (Fig. 2). This approach enabled calculation of the mean shear rate that was used for the recovery tests. These investigations helped to estimate if a material will be printable with a given printer and how needle diameter changes, among other factors, influence the pressure suitable for printing, considering the velocity limits of the device. Furthermore, we demonstrated how these findings can help design bioinks. Please click Additional Files below to see the full abstract

A comparative analysis of detachment forces and energies in initial and mature cell-material interaction

Single cell force spectroscopy (SCFS) enables data on interaction forces to be acquired during the very early adhesion phase. However, SCFS detachment forces and energies have not been compared so far with the forces and energies after maturation of the cell-material contact on a single cell level and with comparable time resolution. We used FluidFM (R) to physically attach single cells to the cantilever by aspiration through a microfluidic channel, in order to achieve the higher forces required for detaching maturely adhering cells. Combining these two approaches allowed us to compare cell adhesion in the initial and maturation phases of adhesion for two exemplary cell-substrate combinations - L929 fibroblasts on fibronectin and MC3T3 osteoblasts on collagen type I. Uncoated glass substrates were used as a reference. For both cell lines, SCFS measurements after contact times of 5, 15 and 30 s revealed significantly higher maximum detachment forces (MDFs) and energies on glass compared to the protein-coated surfaces in the 0.5-4 nN (1-40 fJ) range. FluidFM (R) measurements after 1, 2 and 3 days of culture revealed a significant absolute increase in the MDFs and detachment energies for both cell lines on protein-coated substrates to values of about 600 nN and 10 pJ. On glass, the MDFs were similar for MC3T3 cells, while they were significantly lower for L929 cells. For both cell types, the differences in detachment energy were significant. These differences underline the importance of investigating early and mature adhesion states to obtain a holistic assessment of the cell-material interactions.The authors are grateful to the DFG (German Research Council) graduate school GRK1505/2 "Welisa" and grant number BA 2479/2-1 for funding the position of P. Wysotzki, as well as the consumables for the experiments. We also acknowledge the ERC (European Research Council), Grant Number 617989 for the financial support given. We are grateful to Dr. W. Baumann (Department of Biophysics, Univ. of Rostock) for helpful discussions

Dr. Ursula Göllner-Scheiding zum 90. Geburtstag.

Groll, Eckhard K., Deckert, Jürgen (2012): Dr. Ursula Göllner-Scheiding zum 90. Geburtstag. Beiträge Zur Entomologie = Contributions to Entomology 62: 259-262, DOI: 10.21248/contrib.entomol.62.2.259-262, URL: http://dx.doi.org/10.21248/contrib.entomol.62.2.259-26

Magnesium Phosphate Cement as Mineral Bone Adhesive

Mineral bone cements were actually not developed for their application as bone-bonding agents, but as bone void fillers. In particular, calcium phosphate cements (CPC) are considered to be unsuitable for that application, particularly under moist conditions. Here, we showed the ex vivo ability of different magnesium phosphate cements (MPC) to adhere on bovine cortical bone substrates. The cements were obtained from a mixture of farringtonite (Mg$_3$(PO$_4$)$_2$) with different amounts of phytic acid (C$_6$H$_{18}$O$_{24}$P$_6$, inositol hexaphosphate, IP6), whereas cement setting occurred by a chelation reaction between Mg$^{2+}$ ions and IP6. We were able to show that cements with 25% IP6 and a powder-to-liquid ratio (PLR) of 2.0 g/mL resulted in shear strengths of 0.81 ± 0.12 MPa on bone even after 7 d storage in aqueous conditions. The samples showed a mixed adhesive–cohesive failure with cement residues on the bone surface as indicated by scanning electron microscopy and energy-dispersive X-ray analysis. The presented material demonstrated appropriate bonding characteristics, which could enable a broadening of the mineral bone cements’ application field to bone adhesive

Improving printability of a thermoresponsive hydrogel biomaterial ink by nanoclay addition

As a promising biofabrication technology, extrusion-based bioprinting has gained significant attention in the last decade and major advances have been made in the development of bioinks. However, suitable synthetic and stimuli-responsive bioinks are underrepresented in this context. In this work, we described a hybrid system of nanoclay Laponite XLG and thermoresponsive block copolymer poly(2-methyl-2-oxazoline)-b-poly(2-n-propyl-2-oxazine) (PMeOx-b-PnPrOzi) as a novel biomaterial ink and discussed its critical properties relevant for extrusion-based bioprinting, including viscoelastic properties and printability. The hybrid hydrogel retains the thermogelling properties but is strengthened by the added clay (over 5 kPa of storage modulus and 240 Pa of yield stress). Importantly, the shear-thinning character is further enhanced, which, in combination with very rapid viscosity recovery (similar to 1 s) and structure recovery (similar to 10 s), is highly beneficial for extrusion-based 3D printing. Accordingly, various 3D patterns could be printed with markedly enhanced resolution and shape fidelity compared to the biomaterial ink without added clay. Graphic abstractPeer reviewe

Исследование электрического взрыва Al проводников в среде He

Электрический взрыв проводника является одним из перспективных методов получения металлических нанопорошков. Дисперсностью частиц, получаемых данным методом, можно управлять либо с помощью начальных условий взрыва, либо изменяя состав и давление газа в котором осуществляется синтез порошка. В настоящее время практически все металлические порошки получают в среде аргона. Осуществление ЭВП в среде гелия, может привести к образованию частиц с меньшим средним размером, что обуславливается меньшей плотностью газа более высоким коэффициентом теплопроводности. В работе приведены исследования электрического взрыва алюминиевых проводников в среде гелия, установлены основные закономерности процесса, а так же влияния газа на средний размер получаемых частиц.One of the promising methods for obtaining metallic nanopowders is electrical explosion of wire. The dispersion of the particles received by this method can be operated or by means of entry conditions of explosion, or changing structure and pressure of gas in which powder synthesis is carried out. Now practically all metal powders receive in the environment of argon. The implementation of EEW in a helium environment can lead to the formation of particles with a smaller average size, which is caused by a lower gas density with a higher thermal conductivity. Researches of electric explosion of aluminum conductors in the environment of helium are given in work, the main consistent patterns of process and also influences of gas on the average size of the received particles are determined

Treatment of Focal Cartilage Defects in Minipigs with Zonal Chondrocyte/Mesenchymal Progenitor Cell Constructs

Despite advances in cartilage repair strategies, treatment of focal chondral lesions remains an important challenge to prevent osteoarthritis. Articular cartilage is organized into several layers and lack of zonal organization of current grafts is held responsible for insufficient biomechanical and biochemical quality of repair-tissue. The aim was to develop a zonal approach for cartilage regeneration to determine whether the outcome can be improved compared to a non-zonal strategy. Hydrogel-filled polycaprolactone (PCL)-constructs with a chondrocyte-seeded upper-layer deemed to induce hyaline cartilage and a mesenchymal stromal cell (MSC)-containing bottom-layer deemed to induce calcified cartilage were compared to chondrocyte-based non-zonal grafts in a minipig model. Grafts showed comparable hardness at implantation and did not cause visible signs of inflammation. After 6 months, X-ray microtomography (_CT)-analysis revealed significant bone-loss in both treatment groups compared to empty controls. PCL-enforcement and some hydrogel-remnants were retained in all defects, but most implants were pressed into the subchondral bone. Despite important heterogeneities, both treatments reached a significantly lower modified O’Driscoll-score compared to empty controls. Thus, PCL may have induced bone-erosion during joint loading and misplacement of grafts in vivo precluding adequate permanent orientation of zones compared to surrounding native cartilage. © 2019 by the authors. Licensee MDPI, Basel, Switzerland