Bone Laser Patterning to Decipher Cell Organization.

The laser patterning of implant materials for bone tissue engineering purposes has proven to be a promising technique for controlling cell properties such as adhesion or differentiation, resulting in enhanced osteointegration. However, the possibility of patterning the bone tissue side interface to generate microstructure effects has never been investigated. In the present study, three different laser-generated patterns were machined on the bone surface with the aim of identifying the best surface morphology compatible with osteogenic-related cell recolonization. The laser-patterned bone tissue was characterized by scanning electron microscopy and confocal microscopy in order to obtain a comprehensive picture of the bone surface morphology. The cortical bone patterning impact on cell compatibility and cytoskeleton rearrangement on the patterned surfaces was assessed using Stromal Cells from the Apical Papilla (SCAPs). The results indicated that laser machining had no detrimental effect on consecutively seeded cell metabolism. Orientation assays revealed that patterns with larger hatch distances were correlated with higher cell cytoskeletal conformation to the laser-machined patterns. To the best of our knowledge, this study is the first to consider and evaluate bone as a biological interface that can be engineered for improvement. Further investigations should focus on the in vivo implications of this direct patterning.

Proof of concept of intracochlear drug administration by laser-assisted bioprinting in mice.

Transtympanic administration is used clinically for the injection of gentamicin and/or corticosteroids. This atraumatic route is based on passive diffusion through the round window membrane (RWM). The main limitation of this method is related to the clearance through the Eustachian tube, making the concentration of the therapeutic agent at the intracochlear level uncertain and limited. Moreover, this technique remains unsuitable for molecules of high molecular weight or in the case of gene therapies. The purpose was to study a new technique of intracochlear administration in an atraumatic, direct and controlled manner by laser-assisted bioprinting (LAB). LAB was used to deliver dexamethasone phosphate with thermosensitive hydrogel on the mouse RWM. After validation of the regularity and homogeneity of the pattern, the diffusion in vivo of the dexamethasone into the perilymph after LAB has been confirmed by ELISA. Auditory function measurements showed no hearing impairment suggesting that bioprinting does not induce significant cochlear damage. Hence, the present proof of concept study introduces a promising approach for inner ear drug delivery.

In Vivo Application of Silica-Derived Inks for Bone Tissue Engineering: A 10-Year Systematic Review.

As the need for efficient, sustainable, customizable, handy and affordable substitute materials for bone repair is critical, this systematic review aimed to assess the use and outcomes of silica-derived inks to promote in vivo bone regeneration. An algorithmic selection of articles was performed following the PRISMA guidelines and PICO method. After the initial selection, 51 articles were included. Silicon in ink formulations was mostly found to be in either the native material, but associated with a secondary role, or to be a crucial additive element used to dope an existing material. The inks and materials presented here were essentially extrusion-based 3D-printed (80%), and, overall, the most investigated animal model was the rabbit (65%) with a femoral defect (51%). Quality (ARRIVE 2.0) and risk of bias (SYRCLE) assessments outlined that although a large majority of ARRIVE items were "reported", most risks of bias were left "unclear" due to a lack of precise information. Almost all studies, despite a broad range of strategies and formulations, reported their silica-derived material to improve bone regeneration. The rising number of publications over the past few years highlights Si as a leverage element for bone tissue engineering to closely consider in the future.

In vitroandin vivocharacterization of a novel tricalcium silicate-based ink for bone regeneration using laser-assisted bioprinting.

Grafts aside, current strategies employed to overcome bone loss still fail to reproduce native tissue physiology. Among the emerging bioprinting strategies, laser-assisted bioprinting (LAB) offers very high resolution, allowing designing micrometric patterns in a contactless manner, providing a reproducible tool to test ink formulation. To this date, no LAB associated ink succeeded to provide a reproduciblead integrumbone regeneration on a murine calvaria critical size defect model. Using the Conformité Européenne (CE) approved BioRoot RCS® as a mineral addition to a collagen-enriched ink compatible with LAB, the present study describes the process of the development of a solidifying tricalcium silicate-based ink as a new bone repair promoting substrates in a LAB model. This ink formulation was mechanically characterized by rheology to adjust it for LAB. Printed aside stromal cells from apical papilla (SCAPs), this ink demonstrated a great cytocompatibility, with significantin vitropositive impact upon cell motility, and an early osteogenic differentiation response in the absence of another stimulus. Results indicated that thein vivoapplication of this new ink formulation to regenerate critical size bone defect tends to promote the formation of bone volume fraction without affecting the vascularization of the neo-formed tissue. The use of LAB techniques with this ink failed to demonstrate a complete bone repair, whether SCAPs were printed or not of at its direct proximity. The relevance of the properties of this specific ink formulation would therefore rely on the quantity appliedin situas a defect filler rather than its cell modulation properties observedin vitro. For the first time, a tricalcium silicate-based printed ink, based on rheological analysis, was characterizedin vitroandin vivo, giving valuable information to reach complete bone regeneration through formulation updates. This LAB-based process could be generalized to normalize the characterization of candidate ink for bone regeneration.

α-Tocopherol Attenuates Oxidative Phosphorylation of CD34+ Cells, Enhances Their G0 Phase Fraction and Promotes Hematopoietic Stem and Primitive Progenitor Cell Maintenance.

Alpha tocopherol acetate (αTOA) is an analogue of alpha tocopherol (αTOC) that exists in the form of an injectable drug. In the context of the metabolic hypothesis of stem cells, we studied the impact of αTOA on the metabolic energetic profile and functional properties of hematopoietic stem and progenitor cells. In ex vivo experiments performed on cord blood CD34+ cells, we found that αTOA effectively attenuates oxidative phosphorylation without affecting the glycolysis rate. This effect concerns complex I and complex II of the mitochondrial respiratory chain and is related to the relatively late increase (3 days) in ROS (Reactive Oxygen Species). The most interesting effect was the inhibition of Hypoxia-Inducible Factor (HIF)-2α (Hexpression, which is a determinant of the most pronounced biological effect-the accumulation of CD34+ cells in the G0 phase of the cell cycle. In parallel, better maintenance of the primitive stem cell activity was revealed by the expansion seen in secondary cultures (higher production of colony forming cells (CFC) and Severe Combined Immunodeficiency-mice (scid)-repopulating cells (SRC)). While the presence of αTOA enhanced the maintenance of Hematopoietic Stem Cells (HSC) and contained their proliferation ex vivo, whether it could play the same role in vivo remained unknown. Creating αTOC deficiency via a vitamin E-free diet in mice, we found an accelerated proliferation of CFC and an expanded compartment of LSK (lineagenegative Sca-1+cKit+) and SLAM (cells expressing Signaling Lymphocytic Activation Molecule family receptors) bone marrow cell populations whose in vivo repopulating capacity was decreased. These in vivo data are in favor of our hypothesis that αTOC may have a physiological role in the maintenance of stem cells. Taking into account that αTOC also exhibits an effect on proliferative capacity, it may also be relevant for the ex vivo manipulation of hematopoietic stem cells. For this purpose, low non-toxic doses of αTOA should be used.

α-Tocopherol Acetate Attenuates Mitochondrial Oxygen Consumption and Maintains Primitive Cells within Mesenchymal Stromal Cell Population.

We present here the data showing, in standard cultures exposed to atmospheric O2 concentration, that alpha-tocopherol acetate (α-TOA) has a positive impact on primitive cells inside mesenchymal stromal cell (MstroC) population, by maintaining their proliferative capacity. α-TOA decreases the O2 consumption rate of MStroC probably by impacting respiratory chain complex II activity. This action, however, is not associated with a compensatory increase in glycolysis activity, in spite of the fact that the degradation of HIF-1α was decreased in presence of α-TOA. This is in line with a moderate enhancement of mtROS upon α-TOA treatment. However, the absence of glycolysis stimulation implies the inactivity of HIF-1α which might - if it were active - be related to the maintenance of stemness. It should be stressed that α-TOA might act directly on the gene expression as well as the mtROS themselves, which remains to be elucidated. Alpha-tocopherol acetate (α-TOA), a synthetic vitamin E ester, attenuates electron flow through electron transport chain (ETC) which is probably associated with a moderate increase in mtROS in Mesenchymal Stromal Cells. α-TOA action results in enhancement of the proliferative capacity and maintenance of the differentiation potential of the mesenchymal stem and progenitor cells.