Assessment of physical/mechanical properties and cytotoxicity of dual-cured resin cements containing Sr-bioactive glass nanoparticles and calcium phosphate.

The aim was to develop dual-cured resin cements containing Sr-bioactive glass nanoparticles (Sr-BGNPs; 5 or 10 wt%) and monocalcium phosphate monohydrate (MCPM; 3 or 6 wt%). Effects of additives on degree of monomer conversion (DC), biaxial flexural strength/modulus, shear bond strength (SBS), mass/volume change, color stability, ion release, and cytotoxicity were examined. Controls included material without reactive fillers and Panavia SA Plus (PV). Experimental cements showed higher DC than PV regardless of light activation (p<0.05). Mean SBS and color stability were comparable between experimental cements and PV. Cell viability upon the exposure to sample extracts of experimental cements was 80%-92%. High additive concentrations led to lower strength and modulus than PV (p<0.05). The additives increased mass change, reduced color stability, and promoted ion release. The experimental resin cements demonstrated acceptable mechanical/chemical properties and cytotoxicity. The additives reduced the strength but provided ion release, a desirable action to prevent recurrent caries.

Corrosion Resistance and Cytocompatibility of Magnesium-Calcium Alloys Modified with Zinc- or Gallium-Doped Calcium Phosphate Coatings.

In orthopedic surgery, metals are preferred to support or treat damaged bones due to their high mechanical strength. However, the necessity for a second surgery for implant removal after healing creates problems. Therefore, biodegradable metals, especially magnesium (Mg), gained importance, although their extreme susceptibility to galvanic corrosion limits their applications. The focus of this study was to control the corrosion of Mg and enhance its biocompatibility. For this purpose, surfaces of magnesium-calcium (MgCa1) alloys were modified with calcium phosphate (CaP) or CaP doped with zinc (Zn) or gallium (Ga) via microarc oxidation. The effects of surface modifications on physical, chemical, and mechanical properties and corrosion resistance of the alloys were studied using surface profilometry, goniometry, scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDS), nanoindentation, and electrochemical impedance spectroscopy (EIS). The coating thickness was about 5-8 µm, with grain sizes of 43.1 nm for CaP coating and 28.2 and 58.1 nm for Zn- and Ga-doped coatings, respectively. According to EIS measurements, the capacitive response (Yc) decreased from 11.29 to 8.72 and 0.15 Ω-1 cm-2 sn upon doping with Zn and Ga, respectively. The Ecorr value, which was -1933 mV for CaP-coated samples, was found significantly electropositive at -275 mV for Ga-doped ones. All samples were cytocompatible according to indirect tests. In vitro culture with Saos-2 cells led to changes in the surface compositions of the alloys. The numbers of cells attached to the Zn-doped (2.6 × 104 cells/cm2) and Ga-doped (6.3 × 104 cells/cm2) coatings were higher than that on the surface of the undoped coating (1.0 × 103 cells/cm2). Decreased corrosivity and enhanced cell affinity of the modified MgCa alloys (CaP coated and Zn and Ga doped, with Ga-doped ones having the greatest positive effect) make them novel and promising candidates as biodegradable metallic implant materials for the treatment of bone damages and other orthopedic applications.

Biocompatibility and Biological Performance Evaluation of Additive-Manufactured Bioabsorbable Iron-Based Porous Suture Anchor in a Rabbit Model.

This study evaluated the biocompatibility and biological performance of novel additive-manufactured bioabsorbable iron-based porous suture anchors (iron_SAs). Two types of bioabsorbable iron_SAs, with double- and triple-helical structures (iron_SA_2_helix and iron_SA_3_helix, respectively), were compared with the synthetic polymer-based bioabsorbable suture anchor (polymer_SAs). An in vitro mechanical test, MTT assay, and scanning electron microscope (SEM) analysis were performed. An in vivo animal study was also performed. The three types of suture anchors were randomly implanted in the outer cortex of the lateral femoral condyle. The ultimate in vitro pullout strength of the iron_SA_3_helix group was significantly higher than the iron_SA_2_helix and polymer_SA groups. The MTT assay findings demonstrated no significant cytotoxicity, and the SEM analysis showed cells attachment on implant surface. The ultimate failure load of the iron_SA_3_helix group was significantly higher than that of the polymer_SA group. The micro-CT analysis indicated the iron_SA_3_helix group showed a higher bone volume fraction (BV/TV) after surgery. Moreover, both iron SAs underwent degradation with time. Iron_SAs with triple-helical threads and a porous structure demonstrated better mechanical strength and high biocompatibility after short-term implantation. The combined advantages of the mechanical superiority of the iron metal and the possibility of absorption after implantation make the iron_SA a suitable candidate for further development.

The Beneficial Role of Filipendula ulmaria Extract in Prevention of Prodepressant Effect and Cognitive Impairment Induced by Nanoparticles of Calcium Phosphates in Rats.

Mineral components of dental composites are used in many medical and dental applications, including preventive, restorative, and regenerative dentistry. To evaluate the behavioural alterations induced by nanosized particles of novel dental composites, by means of depressive level and cognitive functions, experimental groups of rats were chronically administered with nanosized hydroxyapatite (HA), tricalcium phosphate (TCP), and amorphous calcium phosphate (ACP) with or without simultaneous application of Filipendula ulmaria L. (FU) methanolic extract. The significant prodepressant action was observed in groups solely treated with HA and ACP. Besides, prolonged treatment with ACP also resulted in a significant decline in cognitive functions estimated in the novel object recognition test. The adverse impact of calcium phosphates on estimated behavioural functions was accompanied by increased oxidative damage and apoptotic markers in the prefrontal cortex, as well as diminished specific neurotrophin (BDNF) and gabaergic expression. The results of our investigation showed that simultaneous antioxidant supplementation with FU extract prevented calcium phosphate-induced behavioural disturbances, as well as prooxidative and apoptotic actions, with the simultaneous restoration of BDNF and GABA-A receptors in the prefrontal cortex. These findings suggest that FU may be useful in the prevention of prodepressant impact and cognitive decline as early as the manifestation of calcium phosphate-induced neurotoxicity.

Use of mechanistic information to derive chemical-specific adjustment factors - Refinement of risk assessment.

When extrapolating data from animal toxicological studies a default factor (dUF) of 100 is applied to derive a heath based guidance value. The UF takes into account the interspecies differences (ID) and the intraspecies variability (IV). When re-evaluating the safety of phosphates used as food additives nephrocalcinosis was identified as the critical endpoint. The underlying mechanism for nephrocalcinosis was attributed to the precipitation of calcium phosphate in the kidney, depending on its solubility, irrespective of the species and the population. Based on the mechanism, the volume of primary urine, for which the glomerular filtration rate (GFR) was used as a proxy, was considered to be the only parameter relevant for ID and IV. Median value of GFR in rats was 4.0 ml/min/kg bw. In humans it was 1.6 ml/min/kg bw in healthy adults and 0.9 in elderly. These values were calculated from the distribution of the GFR data from 8 studies in rats (n = 191), 16 studies in adults (n = 1540) and 5 studies in elderly (n = 2608). Multiplying the distribution of the ratio rat/healthy humans (ID) with the distribution of the ratio healthy humans/elderly human (IV) resulted in a phosphate specific factor of 4.5 (3.3-6.7) (median; 25th - 75th percentile).

[Effect of psoralen on rat osteoblasts injuries induced by TCP wear particles in vitro and its mechanism].

Objective: To investigate the effect and mechanism of psoralen on calvarial osteoblasts injuries caused by tricalcium phosphate (TCP) wear particles in vitro.Methods: Primary osteoblasts were obtained from the calvaria of neonatal SD rat by the series of digestion and were identified with ALP staining. Calvarial osteoblasts were treated with TCP wear particles for 48 h to establish the in vitro model of osteoblasts injuries. The rat osteoblasts were randomly divided into control group, TCP wear particles (0.1 mg/ml) group, psoralen treated (at the concentrations of 10-7, 10-6, 10-5 mol/L) groups. WST assay and the flow cytometry were used to detect the cell viability of osteoblasts and apoptosis, respectively. Chemical colorimetry was performed to examine ALP activity of osteobalsts. When the osteoblasts were treated for 14 day, mineral nodules formation was observed with alizarin red S staining. Western blot was applied to examine protein expressions of glucose regulated protein78/94(GRP78/94), inositol dependent enzyme 1 alpha (IREα), spliced X-box binding protein 1 (XBP1s) and phosphorylated c-Jun N-terminal kinase (p-JNK) in calvarial osteoblasts. Results: Compared with control group, the cell viability of osteoblasts, ALP activity and mineral nodules formation in TCP group were decreased significantly (P＜0.05), while the percentage of apoptosis and protein expressions of GRP78/94, IRE1α, XBP1 and p-JNK were obviously increased in calvarial osteoblasts (P＜0.05). Compared with TCP group, the injuries of calvarial osteoblasts and cell apoptosis in psoralen treated groups were obviously decreased (P＜0.05), and the expression levels of GRP78/94, IRE1α, XBP1 and p-JNK were down-regulated remarkably (P＜0.05). Conclusion: Psoralen prevents osteoblasts injuries caused by TCP wear particles through IRE1α-XBP1s-JNK signaling pathway activation.

The effect of calcium phosphate and silk fibroin nanofiber tuning on properties of calcium sulfate bone cements.

Calcium sulfate (CS) bone cements have been used as bone substitutes for a long time, but their clinical use is currently limited due to their rapid degradation rate and brittleness. This work aimed to study the effect of α-tricalcium phosphate (α-TCP) and silk fibroin nanofibers (SFF) on CS bone cements. The bone cements were prepared from α-CS hemihydrate (α-CSH), calcium sulfate dihydrate (CSD; as a setting accelerator) and varying α-TCP contents (0%, 5%, 10%, 15%, 20% and 25%), with SFF solution or deionized water as the solidification solution at the same liquid/solid ratio. Scanning electron microscopy, particle size distribution, x-ray diffraction and Fourier transform infrared spectroscopy were used to measure the composition and characterize the properties of the materials. The compressive strength, setting time and weight loss rate of samples were also tested. Cytotoxicity was evaluated by a Cell Counting Kit-8 assay. The results suggest that the tuning of α-TCP and SFF has an important role in determining the compressive strength and degradation rate of CS bone cements, and the properties could be changed by varying the content of α-TCP. Moreover, cell experiments showed no toxicity of the samples towards MC3T3 cells. Thus, the materials prepared from α-CSH, CSD, α-TCP and SFF in this work could provide the basis for research into CS-based bone repair materials.

Soy protein hydrolysate grafted cellulose nanofibrils with bioactive signals for bone repair and regeneration.

TEMPO oxidized cellulose nanofibers (T-CNF) were prepared from cellulose pulp which is extracted from bagasse. Soy protein hydrolysate (SPH) was grafted on T-CNF via amidation of carboxylic groups. Biomineralization was, then, assessed via calcium phosphates (CaP) precipitation in twice-simulated body fluid until formation of a new bioactive material. Protein was efficiently grafted without alteration of morphology and nanofibrils packing as reported by Fourier Transform infrared analysis /X Ray Diffraction /Scanning and Transmission Electron Microscopy / Atomic Force Microscopy. Highly crystalline calcium phosphate deposits - ca. 22.1% - were detected, with a Ca/P ratio equal to 1.63, in agreement with native bone apatite composition. In vitro response of human Mesenchymal Stem Cells confirmed the biocompatibility. No significant differences in terms of cell adhesion were recognized while a significant increase in cell proliferation was detected until 7 days. The presence of calcium phosphates tends to cover the nanofibrillar pattern, inducing the inhibition of cell proliferation and promoting the ex-novo precipitation of mineral phases. All the results suggest a promising use of these biomaterials in the repair and/or the regeneration of hard tissues such as bone.

Mineral deposition promoted by resin-based sealants with different calcium phosphate additions.

The aim of this study was to evaluate the influence of different calcium phosphates (CaPs) on the physical, biological, and remineralizing properties of experimental resin-based sealants (RBSs). Triethylene-glycol dimethacrylate (90wt%) and bisphenol A-glycidyl methacrylate (10wt%) were used to produce resin-based sealants. Hydroxyapatite (SHAp), α-tricalcium phosphate (Sα-TCP) and octacalcium phosphate (SOCP) were added to the sealants in a 10wt% concentration. One group without CaPs was used as the control group (SCG). The degree of conversion (DC) was assessed with Fourier-transformed infrared spectroscopy, whereas cytotoxicity was tested with the HaCaT keratinocyte cell line. The ultimate tensile strength (UTS) was used to assess the mechanical strength of the experimental RBSs. Sealed enamel was used for colorimetric assay. Mineral deposition was assessed with Raman spectroscopy after 7, 14, and 28 days of sample immersion in artificial saliva. Scanning electron microscopy was used to analyze the surface morphology after 28 days of immersion. The addition of 10wt% of fillers significantly reduced the DC of sealants. SOCP groups showed reduced cell viability. Higher UTS was found for Sα-TCP and SHAp. The color analysis showed that SGC and demineralized teeth presented higher mismatches with the sound tissue. Mineral deposition was observed for SHAp and Sα-TCP after 7 days, with increased phosphate content and mineral deposits for SHAp after 28 days. RBS with the addition of 10% HAp promoted increased mineralization in vitro after 28 days, and did not affect cell viability, DC, mechanical properties, or RBS color in the enamel.

ATRQß-001 Vaccine Prevents Experimental Abdominal Aortic Aneurysms.

Background We have developed a peptide vaccine named ATRQß-001, which was proved to retard signal transduction initiated by angiotensin II (Ang II). Ang II was implicated in abdominal aortic aneurysm (AAA) progression, but whether the ATRQß-001 vaccine would prevent AAA is unknown. Methods and Results Ang II-infused ApoE-/- mice and calcium phosphate-induced AAA in C57BL/6 mice were used to verify the efficiency of ATRQß-001 vaccine in AAA. Results demonstrated that the vaccine effectively restrained the aneurysmal dilation and vascular wall destruction of aorta in both animal models, beyond anti-hypertensive effects. In Ang II-induced AAA vascular sections, Immunohistochemical staining showed that the vaccine notably constrained vascular inflammation and vascular smooth muscle cell (VSMC) phenotypic transition, concurrently reduced macrophages infiltration. In cultured VSMC, the anti-ATR-001 antibody inhibited osteopontin secretion induced by Ang II, thereby impeded macrophage migration while co-culture. Furthermore, metalloproteinases and other matrix proteolytic enzymes were also found to be limited by the vaccine in vivo and in vitro. Conclusions ATRQß-001 vaccine prevented AAA initiation and progression in both Ang II and calcium phosphate-induced AAA models. And the beneficial effects were played beyond decrease of blood pressure, which provided a novel and promising method to take precautions against AAA.

In vitro cytotoxicity of calcium phosphate cement reinforced with multiwalled carbon nanotubes.

The in vitro cytotoxicity of both the multiwalled carbon nanotubes (MWCNT) in suspension with culture medium and the tetracalcium phosphate/monetite cement with addition of 0.8 wt% of MWCNTs on fibroblasts and osteoblasts were studied. The cytotoxicity was evaluated by MTS test (formazan) and live/dead staining. No cytotoxicity of MWCNT extract was measured contrary to about 60% reduction in proliferation of fibroblasts in MWCNT suspension as compared with negative control. The several contact cytotoxicity of MWCNT composite cement surfaces on seeded cells was demonstrated by MTS test and live/dead staining of damaged fibroblasts and dead osteoblasts after 72 h of culture. The detailed microstructure analysis showed a significant refinement of the surface texture due to the formation of thin needle-like hydroxyapatite particles on MWCNTs and this effect could be responsible for cytotoxicity of composites.

Citric acid enhances the physical properties, cytocompatibility and osteogenesis of magnesium calcium phosphate cement.

In recent years, the magnesium phosphate cements showed impressive advantages for their setting property, mechanical strength, and resorption rate in laboratory investigation. While it remained a big challenge to develop the magnesium phosphate cements with ideal self-setting properties, sufficient mechanical strength, excellent biocompatibility, and osteoinductivity for clinical application. In our work, we prepared the magnesium calcium phosphate cement (MCPC) using the MgO, KH2P2O4, and Ca(H2PO4)2 particles with the citric acid added. The citric acid was adopted to modify the setting time and compressive strength of the MCPC, which were investigated by the X-ray diffractometer and scanning electron microscopy. The cytocompatibility and osteoinductivity of the modified cements were evaluated by the MC3T3-E1 cells proliferation and morphology, alkaline phosphatase assay, alizarin red staining and western blot assay. The results demonstrated that the citric acid modified MCPC was featured of satisfactory setting time, ideal mechanical strength, good cytocompatibility and osteoinductivity, indicating its potential application for bone regeneration.

The summary of the most important cell-biomaterial interactions that need to be considered during in vitro biocompatibility testing of bone scaffolds for tissue engineering applications.

Tissue engineered products (TEPs), which mean biomaterials containing either cells or growth factors or both cells and growth factors, may be used as an alternative to the autografts taken directly from the bone of the patients. Nevertheless, the use of TEPs needs much more understanding of biointeractions between biomaterials and eukaryotic cells. Despite the possibility of the use of in vitro cellular models for initial evaluation of the host response to the implanted biomaterial, it is observed that most researchers use cell cultures only for the evaluation of cytotoxicity and cell proliferation on the biomaterial surface, and then they proceed to animal models and in vivo testing of bone implants without fully utilizing the scientific potential of in vitro models. In this review, the most important biointeractions between eukaryotic cells and biomaterials were discussed, indicating molecular mechanisms of cell adhesion, proliferation, and biomaterial-induced activation of immune cells. The article also describes types of cellular models which are commonly used for biomaterial testing and highlights the possibilities and drawbacks of in vitro tests for biocompatibility evaluation of novel scaffolds. Finally, the review summarizes recent findings concerning the use of adult mesenchymal stem cells for TEP generation and compares the potential of bone marrow- and adipose tissue-derived stem cells in regenerative medicine applications.

IL (Interleukin)-33 Suppresses Abdominal Aortic Aneurysm by Enhancing Regulatory T-Cell Expansion and Activity.

Objective- Inflammation occurs during the progression of abdominal aortic aneurysm (AAA). IL (interleukin)-33 is a pleiotropic cytokine with multiple immunomodulatory effects, yet its role in AAA remains unknown. Approach and Results- Immunoblot, immunohistochemistry, and immunofluorescent staining revealed increased IL-33 expression in adventitia fibroblasts from mouse AAA lesions. Daily intraperitoneal administration of recombinant IL-33 or transgenic IL-33 expression ameliorated periaorta CaPO4 injury- and aortic elastase exposure-induced AAA in mice, as demonstrated by blunted aortic expansion, reduced aortic wall elastica fragmentation, enhanced AAA lesion collagen deposition, attenuated T-cell and macrophage infiltration, reduced inflammatory cytokine production, skewed M2 macrophage polarization, and reduced lesion MMP (matrix metalloproteinase) expression and cell apoptosis. Flow cytometry analysis, immunostaining, and immunoblot analysis showed that exogenous IL-33 increased CD4+Foxp3+ regulatory T cells in spleens, blood, and aortas in periaorta CaPO4-treated mice. Yet, ST2 deficiency muted these IL-33 activities. Regulatory T cells from IL-33-treated mice also showed significantly stronger activities in suppressing smooth muscle cell inflammatory cytokine and chemokine expression, macrophage MMP expression, and in increasing M2 macrophage polarization than those from vehicle-treated mice. In contrast, IL-33 failed to prevent AAA and lost its beneficial activities in CaPO4-treated mice after selective depletion of regulatory T cells. Conclusions- Together, this study established a role of IL-33 in protecting mice from AAA formation by enhancing ST2-dependent aortic and systemic regulatory T-cell expansion and their immunosuppressive activities.

Physico-Mechanical Properties of HA/TCP Pellets and Their Three-Dimensional Biological Evaluation In Vitro.

The use of bioceramics, especially the combination of hydroxyapatite (HA) and ß-tricalcium phosphate (ß-TCP), as a three-dimensional scaffold in bone engineering is essential because together these elements constitute 60% of the bone content. Different ratios of HA and ß-TCP were previously tested for their ability to produce suitable bioceramic scaffolds, which must be able to withstand high mechanical load. In this study, two ratios of HA/TCP (20:80 and 70:30) were used to create pellets, which then were evaluated in vitro to identify any adverse effects of using the material in bone grafting. Diametral tensile strength (DTS) and density testing was conducted to assess the mechanical strength and porosity of the pellets. The pellets then were tested for their toxicity to normal human fibroblast cells. In the toxicity assay, cells were incubated with the pellets for 3 days. At the end of the experiment, cell morphological changes were assessed, and the absorbance was read using PrestoBlue Cell Viability Reagent™. An inversely proportional relationship between DTS and porosity percentage was detected. Fibroblasts showed normal cell morphology in both treatments, which suggests that the HA/TCP pellets were not toxic. In the osteoblast cell attachment assay, cells were able to attach to the surface of both ratios, but cells were also able to penetrate inside the scaffold of the 70:30 pellets. This finding suggests that the 70:30 ratio had better osteoconduction properties than the 20:80 ratio.

Mechanical and cytotoxicity properties of hybrid ceramics filled polyamide 12 filament feedstock for craniofacial bone reconstruction via fused deposition modelling.

OBJECTIVE: To compare the mechanical and biological properties of newly developed hybrid ceramics filled and unfilled polyamide 12 (PA 12) for craniofacial reconstruction via a fused deposition modelling (FDM) framework. METHODS: 15wt% of zirconia (ZrO2) as well as 30, 35, and 40wt% of beta-tricalcium phosphate (ß-TCP) were compounded with PA 12, followed by the fabrication of filament feedstocks using a single screw extruder. The fabricated filament feedstocks were used to print the impact specimens. The melt flow rate, tensile properties of fabricated filament feedstocks, and 3D printed impact properties of the specimens were assessed using melt flow indexer, universal testing machine, and Izod pendulum tester, respectively. The microstructure of selected filament feedstocks and broken impact specimens were analysed using a field emission scanning electron microscope and universal testing machine. Human periodontal ligament fibroblast cells (HPdLF) were used to evaluate the cytotoxicity of the materials by (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromid) (MTT) assay. RESULTS: Hybrid ceramics filled PA 12 indicated sufficient flowability for FDM 3D printing. The tensile strength of hybrid ceramics filled PA 12 filament feedstocks slightly reduced as compared to unfilled PA 12. However, the tensile modulus and impact strength of hybrid ceramics filled PA 12 increased by 8%-31% and 98%-181%, respectively. A significant increase was also detected in the cell viability of the developed composites at concentrations of 12.5, 25, 50 and 100mg/ml. SIGNIFICANCE: The newly developed hybrid ceramics filled PA 12 filament feedstock with improved properties is suitable for an FDM-based 3D printer, which enables the creation of patient-specific craniofacial implant at a lower cost to serve low-income patients.

Comparative Evaluation of Cytotoxicity and Genotoxicity of Two Bioceramic Sealers on Fibroblast Cell Line: An in vitro Study.

AIM: The purpose of this study was to evaluate and compare the cytotoxicity and genotoxicity of two bioceramic root canal sealers: EndoSequence BC and iRoot SP with zinc oxide eugenol sealers on fibroblast cell line. MATERIALS AND METHODS: The sealers tested were zinc oxide eugenol, EndoSequence BC, and iRoot SP. Each material was mixed according to the manufacturer's instructions and mounted into sterile polyethylene color-coded rings, for cytotoxicity and genotoxicity evaluation. After 48 hours, the set materials were transferred to previously marked wells and cytotoxicity evaluation to L929 murine fibroblast cells was done by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. The percentages of viable cells were then calculated and values were statistically analyzed by Kruskal-Wallis test. The evaluation of genotoxicity of the materials to L929 murine fibroblast cells was carried out by Comet assay. To quantify deoxyribonucleic acid (DNA) damage, the following comet parameters were evaluated in the assay using Comet scoring software: tail length, tail moment, and Olive moment. The values were statistically analyzed using Kruskal-Wallis test with a significance value set to p < 0.05. RESULTS: The results of the study showed that both cytotoxicity and genotoxicity evaluation by MTT assay and Comet assay can be done on L929 murine fibroblast cell line. Among the three tested materials, zinc oxide eugenol showed maximum cytotoxicity to the cells (30.64% viable cells), followed by EndoSequence BC (71.33% viable cells) and iRoot SP (75.11% viable cells). The evaluation of DNA damage by genotoxicity assessment showed iRoot SP to be least genotoxic followed closely by EndoSequence BC. Zinc oxide eugenol was genotoxic and induced more DNA damage on the fibroblast cell line studied. The statistical analyses for both the assays were nonsignificant. CONCLUSION: All the three tested sealers showed varying degrees of cytotoxicity and genotoxicity while using fibro-blast cell line. Zinc oxide eugenol was most toxic in both the assays and iRoot SP showed least toxicity, followed closely by EndoSequence BC.

Investigation of the Genotoxicity of Aluminum Oxide, ß-Tricalcium Phosphate, and Zinc Oxide Nanoparticles In Vitro.

The aim of this study was to investigate the genotoxicity of aluminum oxide (Al2O3), ß-tricalcium phosphate (ß-TCP) (Ca3(PO4)2), and zinc oxide (ZnO) nanoparticles (NPs) that were 4.175, 9.058, and 19.8 nm sized, respectively, on human peripheral blood lymphocytes using micronucleus (MN) and chromosome aberration (CA) techniques. Aluminum oxide and ß-TCP NPs did not show genotoxic effects on human peripheral blood cultures in vitro, even at the highest concentrations; therefore, these materials may be suitable for use as biocompatible materials. It was observed that, even at a very low dose (≥12.5 ppm), ZnO NPs had led to genotoxicity. In addition, at high concentrations (500 ppm and above), ZnO NPs caused mortality of lymphocytes. For these reasons, it was concluded that ZnO NPs are not appropriate for using as a biocompatible biomaterial.

In vitro and in vivo toxicity assessment of alginate/eudragit S 100-enclosed chitosan-calcium phosphate-loaded iron saturated bovine lactoferrin nanocapsules (Fe-bLf NCs).

Lactoferrin has been known to have antimicrobial properties. This research was conducted to investigate the toxicity of Alginate/EUDRAGIT® S 100-enclosed chitosan-calcium phosphate-loaded Fe-bLf nanocapsules (NCs) by in vitro and in vivo assays. Brine shrimp lethality assay showed that the LC50 value of NCs was more than 1mg/mL which indicated that NCs was not toxic to Brine shrimp. However, the LC50 values for the positive control potassium dichromate at 24h is 64.15µg/mL, which was demostrated the toxic effect against the brine shrimp. MTT cytotoxicity assay also revealed that NCs was not toxic against non-cancerous Vero cell line with IC50 values of 536µg/mL. Genotoxicity studies by comet assay on Vero cells revealed that NCs exerted no significant genotoxic at 100µg/mL without tail or shorter comet tail. Allium cepa root assay carried out at 125, 250, 500 and 1000µg/mL for 24h revealed that the NCs was destitute of significant genotoxic effect under experimental conditions. The results show that there is no significant difference (p>0.05) in mitotic index between the deionized water and NCs treated Allium cepa root tip cells. In conclusion, no toxicity was observed in NCs in this study. Therefore, nontoxic NCs has the good potential to develop as a therapeutic agent.

Phagocytosis of environmental or metabolic crystalline particles induces cytotoxicity by triggering necroptosis across a broad range of particle size and shape.

In crystallopathies, crystals or crystalline particles of environmental and metabolic origin deposit within tissues, induce inflammation, injury and cell death and eventually lead to organ-failure. The NLRP3-inflammasome is involved in mediating crystalline particles-induced inflammation, but pathways leading to cell death are still unknown. Here, we have used broad range of intrinsic and extrinsic crystal- or crystalline particle-sizes and shapes, e.g. calcium phosphate, silica, titanium dioxide, cholesterol, calcium oxalate, and monosodium urate. As kidney is commonly affected by crystallopathies, we used human and murine renal tubular cells as a model system. We showed that all of the analysed crystalline particles induce caspase-independent cell death. Deficiency of MLKL, siRNA knockdown of RIPK3, or inhibitors of necroptosis signaling e.g. RIPK-1 inhibitor necrostatin-1s, RIPK3 inhibitor dabrafenib, and MLKL inhibitor necrosulfonamide, partially protected tubular cells from crystalline particles cytotoxicity. Furthermore, we identify phagocytosis of crystalline particles as an upstream event in their cytotoxicity since a phagocytosis inhibitor, cytochalasin D, prevented their cytotoxicity. Taken together, our data confirmed the involvement of necroptosis as one of the pathways leading to cell death in crystallopathies. Our data identified RIPK-1, RIPK3, and MLKL as molecular targets to limit tissue injury and organ failure in crystallopathies.