Low-Temperature Calcium Phosphate Ceramics Can Modulate Monocytes and Macrophages Inflammatory Response In Vitro.

Creating bioactive materials for bone tissue regeneration and augmentation remains a pertinent challenge. One of the most promising and rapidly advancing approaches involves the use of low-temperature ceramics that closely mimic the natural composition of the extracellular matrix of native bone tissue, such as Hydroxyapatite (HAp) and its phase precursors (Dicalcium Phosphate Dihydrate-DCPD, Octacalcium Phosphate-OCP, etc.). However, despite significant scientific interest, the current knowledge and understanding remain limited regarding the impact of these ceramics not only on reparative histogenesis processes but also on the immunostimulation and initiation of local aseptic inflammation leading to material rejection. Using the stable cell models of monocyte-like (THP-1ATRA) and macrophage-like (THP-1PMA) cells under the conditions of LPS-induced model inflammation in vitro, the influence of DCPD, OCP, and HAp on cell viability, ROS and intracellular NO production, phagocytosis, and the secretion of pro-inflammatory cytokines was assessed. The results demonstrate that all investigated ceramic particles exhibit biological activity toward human macrophage and monocyte cells in vitro, potentially providing conditions necessary for bone tissue restoration/regeneration in the peri-implant environment in vivo. Among the studied ceramics, DCPD appears to be the most preferable for implantation in patients with latent inflammation or unpredictable immune status, as this ceramic had the most favorable overall impact on the investigated cellular models.

Composite Remineralization of Bone-Collagen Matrices by Low-Temperature Ceramics and Serum Albumin: A New Approach to the Creation of Highly Effective Osteoplastic Materials.

This study examined the effectiveness of coating demineralized bone matrix (DBM) with amorphous calcium phosphate (DBM + CaP), as well as a composite of DBM, calcium phosphate, and serum albumin (DBM + CaP + BSA). The intact structure of DBM promotes the transformation of amorphous calcium phosphate (CaP) into dicalcium phosphate dihydrate (DCPD) with a characteristic plate shape and particle size of 5-35 µm. The inclusion of BSA in the coating resulted in a better and more uniform distribution of CaP on the surface of DBM trabeculae. MG63 cells showed that both the obtained forms of CaP and its complex with BSA did not exhibit cytotoxicity up to a concentration of 10 mg/mL in vitro. Ectopic (subcutaneous) implantation in rats revealed pronounced biocompatibility, as well as strong osteoconductive, osteoinductive, and osteogenic effects for both DBM + CaP and DBM + CaP + BSA, but more pronounced effects for DBM + CaP + BSA. In addition, for the DBM + CaP + BSA samples, there was a pronounced full physiological intrafibrillar biomineralization and proangiogenic effect with the formation of bone-morrow-like niches, accompanied by pronounced processes of intramedullary hematopoiesis, indicating a powerful osteogenic effect of this composite.

Biomimetic Remineralized Three-Dimensional Collagen Bone Matrices with an Enhanced Osteostimulating Effect.

Bone grafts with a high potential for osseointegration, capable of providing a complete and effective regeneration of bone tissue, remain an urgent and unresolved issue. The presented work proposes an approach to develop composite biomimetic bone material for reconstructive surgery by deposition (remineralization) on the surface of high-purity, demineralized bone collagen matrix calcium phosphate layers. Histological and elemental analysis have shown reproduction of the bone tissue matrix architectonics, and a high-purity degree of the obtained collagen scaffolds; the cell culture and confocal microscopy have demonstrated a high biocompatibility of the materials obtained. Adsorption spectroscopy, scanning electron microscopy, microcomputed tomography (microCT) and infrared spectroscopy, and X-ray diffraction have proven the efficiency of the deposition of calcium phosphates on the surface of bone collagen scaffolds. Cell culture and confocal microscopy methods have shown high biocompatibility of both demineralized and remineralized bone matrices. In the model of heterotopic implantation in rats, at the term of seven weeks, an intensive intratrabecular infiltration of calcium phosphate precipitates, and a pronounced synthetic activity of osteoblast remodeling and rebuilding implanted materials, were revealed in remineralized bone collagen matrices in contrast to demineralized ones. Thus, remineralization of highly purified demineralized bone matrices significantly enhanced their osteostimulating ability. The data obtained are of interest for the creation of new highly effective osteoplastic materials for bone tissue regeneration and augmentation.

Octacalcium Phosphate for Bone Tissue Engineering: Synthesis, Modification, and In Vitro Biocompatibility Assessment.

Octacalcium phosphate (OCP, Ca8H2(PO4)6·5H2O) is known to be a possible precursor of biological hydroxyapatite formation of organic bone tissue. OCP has higher biocompatibility and osseointegration rate compared to other calcium phosphates. In this work, the synthesis of low-temperature calcium phosphate compounds and substituted forms of those at physiological temperatures is shown. Strontium is used to improve bioactive properties of the material. Strontium was inserted into the OCP structure by ionic substitution in solutions. The processes of phase formation of low-temperature OCP with theoretical substitution of strontium for calcium up to 50 at.% in conditions close to physiological, i.e., temperature 35-37 °C and normal pressure, were described. The effect of strontium substitution range on changes in the crystal lattice of materials, the microstructural features, surface morphology and biological properties in vitro has been established. The results of the study indicate the effectiveness of using strontium in OCP for improving biocompatibility of OCP based composite materials intended for bone repair.

In Vitro Study of Octacalcium Phosphate Behavior in Different Model Solutions.

Octacalcium phosphate (OCP), a new-generation bone substitute material, is a considered precursor of the biological bone apatite. The two-layered structure of OCP contains the apatitic and hydrated layers and is intensively involved in ion-exchange surface reactions, which results in OCP hydrolysis to hydroxyapatite and adsorption of ions or molecular groups presented in the environment. During various in vitro procedures, such as biomaterial solubility, additive release studies, or the functionalization technique, several model solutions are applied. The composition of the environmental solution affects the degree and rate of OCP hydrolysis, its surface reactivity, and further in vitro and in vivo properties. The performed study was aimed to track the structural changes of OCP-based materials while treating in the most popular model solutions of pH values 7.2-7.4: simulated body fluid (SBF), Dulbecco's phosphate-buffered saline (DPBS), supersaturated calcification solution (SCS), normal saline (NS), and Dulbecco's modified Eagle's medium (DMEM). Various degrees of OCP hydrolysis and/or precipitate formation were achieved through soaking initial OCP granules in the model solutions. Detailed data of X-ray diffraction, Fourier-transform infrared spectroscopy, atomic emission spectrometry with inductively coupled plasma, and scanning electron microscopy are presented. Cultivation of osteosarcoma cells was implemented on OCP pre-treated in DMEM for 1-28 days. It was shown that NS mostly degraded the OCP structure. DPBS slightly changed the OCP structure during the first treatment term, and during further terms, the crystals got thinner and OCP hydrolysis took place. Treatment in SBF and SCS caused the precipitate formation along with OCP hydrolysis, with a larger contribution of SCS solution to precipitation. Pre-treating in DMEM enhanced the cytocompatibility of materials. As a result, on performing the in vitro procedures, careful selection of the contact solution should be made to avoid the changes in materials structure and properties and get adequate results.

3D Printed Gene-activated Octacalcium Phosphate Implants for Large Bone Defects Engineering.

The aim of the study was the development of three-dimensional (3D) printed gene-activated implants based on octacalcium phosphate (OCP) and plasmid DNA encoding VEGFA. The first objective of the present work involved design and fabrication of gene-activated bone substitutes based on the OCP and plasmid DNA with VEGFA gene using 3D printing approach of ceramic constructs, providing the control of its architectonics compliance to the initial digital models. X-ray diffraction, scanning electron microscopy (SEM), Fourier transform infrared spectroscopy, and compressive strength analyses were applied to investigate the chemical composition, microstructure, and mechanical properties of the experimental samples. The biodegradation rate and the efficacy of plasmid DNA delivery in vivo were assessed during standard tests with subcutaneous implantation to rodents in the next stage. The final part of the study involved substitution of segmental tibia and mandibular defects in adult pigs with 3D printed gene-activated implants. Biodegradation, osteointegration, and effectiveness of a reparative osteogenesis were evaluated with computerized tomography, SEM, and a histological examination. The combination of gene therapy and 3D printed implants manifested the significant clinical potential for effective bone regeneration in large/critical size defect cases.

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.

3D Printing of Octacalcium Phosphate Bone Substitutes.

Biocompatible calcium phosphate ceramic grafts are able of supporting new bone formation in appropriate environment. The major limitation of these materials usage for medical implants is the absence of accessible methods for their patient-specific fabrication. 3D printing methodology is an excellent approach to overcome the limitation supporting effective and fast fabrication of individual complex bone substitutes. Here, we proposed a relatively simple route for 3D printing of octacalcium phosphates (OCP) in complexly shaped structures by the combination of inkjet printing with post-treatment methodology. The printed OCP blocks were further implanted in the developed cranial bone defect followed by histological evaluation. The obtained result confirmed the potential of the developed OCP bone substitutes, which allowed 2.5-time reducing of defect's diameter at 6.5 months in a region where native bone repair is extremely inefficient.

Synergistic extraction of calcium and strontium into nitrobenzene by using hydrogen dicarbollylcobaltate and n,n´-dimethyl-n,n´-diphenyl-2,6-dipicolinamide.

Extraction of microamounts of calcium and strontium by a nitrobenzene solution of hydrogen dicarbollylcobaltate (H+B-) in the presence of N,N´-dimethyl-N,N´-diphenyl-2,6-dipicolinamide (MePhDPA, L) has been investigated. The equilibrium data have been explained assuming that the species HL+, HL+2, ML2+2 and ML2+3 (M2+ = Ca2+, Sr2+) are extracted into the organic phase. The values of extraction and stability constants of the cationic complex species in nitrobenzene saturated with water have been determined.

Influence of transcranial magnetic stimulation on spike-wave discharges in a genetic model of absence epilepsy.

Transcranial magnetic stimulation (TMS) impulses, (0.5 Hz, 3 impulses) were presented at threshold intensity to male WAG/Rij rats. One group received stimuli, which involved motor responses of hindlimbs, rats of the second group received sham stimulation. Electrocorticograms (ECoG) were recorded before and up to 2 hr from the moment of transcranial magnetic stimulation. It was established that such stimulation engendered a reduction of spike-wave discharge (SWD) bursts duration. This effect was most pronounced in 30 min from the moment of cessation of stimulation, when a decrease of 31.4% was noted in comparison with sham-stimulated control group. The number of bursts of spike-wave discharges was reduced, but did not reach significant difference when compared both with pre-stimulative base-line level and with sham-stimulated control rats. Bursts of spike-wave discharges restored up to pre-stimulative level in 90-150 minutes from the moment of cessation of transcranial stimulation. It can be concluded that transcranical magnetic stimulation possessed an ability to engender short-time suppression of bursts of spike-wave discharges in WAG/Rij rats.

Are there different mechanisms of synchronization in the course of spike-wave discharges (SWDs) burst development in WAG/Rij rats?

In WAG/Rij rats the pair linear correlation r was calculated for bipolar recordings in fronto-temporal, fronto-occipital and occipito-temporal zones of both hemispheres as well as in paleocerebellar cortex (culmen). It was shown that development of SWD bursts resulted in interhemispheric decreases of correlation between the right occipito-temporal cortical region on one side, and left fronto-temporal on the contralateral side. Towards the end of SWD, we found an increased interhemispheric correlation between left fronto-temporal and right fronto-occipital cortical zones, as well as, between both fronto-temporal zones. Paleocerebellum correlates at a weak to moderate level during different periods of SWD burst generation.

Complexes of the proton and its hydrates with carbamoylphosphine oxide in wet dichloroethane solutions.

To better understand the complex equilibria involved in the UNEX process for acidic solvent extraction of radionuclides, the interaction of a carbamoylphosphine oxide ligand (L) with the proton of hydrated chlorinated cobalt(III)dicarbollide acid, H[Co(C2B9H8Cl3)2], has been studied in wet 1,2-dichlorethane (DCE) solution using IR and NMR (13C and 31P) spectroscopy. The formation of two groups of complexes has been determined. The first group contains three complexes with 1:1 composition of acid to ligand (Scheme 1). The second group of complexes has 1:2 composition in the equilibrium, shown in Scheme 2. Within each group, the complexes differ in composition only by the number of incorporated water molecules. The equilibria (Schemes 1 and 2) are both very sensitive to the content of self-associated water in solution and are driven by its concentration, which is unsteady and depends on the solution preparation history. The simultaneous presence of both anhydrous (I, II) and hydrated (III, IIIa, IV) proton solvates indicates that the enthalpies of carbamoylphosphine oxide complex formation with H+, H3O+, and H5O2(+) are very close to each other.