Osteoblastic MC3T3-E1 Cells Research Articles

Zinc-doped hydroxyapatite loaded chitosan gelatin nanocomposite scaffolds as a promising platform for bone regeneration

The advancement in the arena of bone tissue engineering persuades us to develop novel nanocomposite scaffolds in order to improve antibacterial, osteogenic, and angiogenic properties that show resemblance to natural bone extracellular matrix. Here, we focused on the development of novel zinc-doped hydroxyapatite (ZnHAP) nanoparticles (1, 2 and 3 wt%; size: 50-60 nm) incorporated chitosan-gelatin (CG) nanocomposite scaffold, with an interconnected porous structure. The addition of ZnHAP nanoparticles decreases the pore size (∼30 µm) of the CG scaffolds. It was observed that with the increase in the concentration of ZnHAP nanoparticles (3 wt%) in CG scaffolds, the swelling ratio (1760% ± 2.0%), porosity (71% ± 0.98%) and degradation rate (35%) decreased, whereas mechanical property (1 MPa) increased, which was better as compared to control (CG) samples. Similarly, the high deposition of apatite crystals especially CG-ZnHAP3nanocomposite scaffold revealed the excellent osteoconductive potential among all other scaffolds. MC3T3-E1 osteoblastic cells seeded with CG-ZnHAP nanocomposite scaffolds depicted better cell adhesion, proliferation and differentiation to osteogenic lineages. Finally, the chorioallantoic membrane (CAM) assay revealed better angiogenesis of ZnHAP nanoparticles (3 wt%) loaded CG scaffolds supporting vascularization after 7th day incubation in the CAM area. Overall, the results showed that the CG-ZnHAP3nanocomposite scaffold could be a potential candidate for bone defect repair.

Overexpression of RGPR-p117 reveals anticancer effects by regulating multiple signaling pathways in bone metastatic human breast cancer MDA-MB-231 cells.

The role of RGPR-p117, a transcription factor, which binds to the TTGGC motif in the promoter region of the regucalcin gene, in cell regulation remains to be investigated. This study elucidated whether RGPR-p117 regulates the activity of triple-negative human breast cancer MDA-MB-231 cells in vitro. The wild-type and RGPR-p117-overexpressing cancer cells were cultured in DMEM supplemented with fetal bovine serum. RGPR-p117 overexpression suppressed colony formation and growth of cancer cells. Stimulatory effects of epidermal growth factor on cell growth were blocked by RGPR-p117 overexpression. Wild-type cell proliferation was repressed by cell cycle and intracellular signaling inhibitors. These effects were not potentiated in transfectants. Overexpressed RGPR-p117 protected cancer cells against apoptosis inducers. Mechanistic results showed that RGPR-p117 overexpression decreased the expression of Ras, PI3-kinase, Akt, mitogen-activated protein kinase, and mTOR, which are involved in cell growth, while it elevated the levels of the cancer cell suppressor p53, Rb, p21, and regucalcin. Overexpression of RGPR-p117 suppressed cancer cell migration and adhesion. Interestingly, osteoblastic MC3T3-E1 cells or macrophage RAW264.7 cells involved in the bone microenvironment were impaired by coculture with MDA-MB-231 cells. The effects of cancer cells were blocked by transfection. Coculture with conditioned medium obtained from breast cancer cells repressed proliferation and enhanced the death of osteoblastic cells and macrophages. A TNF-α signaling inhibitor blocked these effects. Thus, overexpressed RGPR-p117 was found to suppress the activity of breast cancer cells by regulating various signaling processes, providing new insight into cellular signaling regulation.

Collagen peptides alleviate estrogen deficiency-induced osteoporosis by enhancing osteoblast differentiation and mineralization.

Osteoporosis is a systemic skeletal disorder characterized by decreased bone mass and impaired bone microarchitecture because of an imbalance between bone resorption and formation. Existing pharmacological treatments often have significant side effects and mainly focus on inhibiting bone resorption. Other than inhibiting osteoclast-mediated bone resorption, the present study also investigates the potential role of sheepskin collagen peptide (SSCP) in bone formation by promoting osteoblast proliferation, differentiation and mineralization. SSCP improved bone mineral density in ovariectomized mice by improving bone volume, trabecular thickness and trabecular number. Histological analysis and tartrate-resistant acid phosphatase (TRAP) staining revealed denser trabeculae and decreased osteoclast activity, accompanied by a normalized receptor activator of nuclear factor kappa-B ligand/osteoprotegerin ratio and reduced serum TRAP levels. SSCP promotes the proliferation, differentiation and mineralization of MC3T3-E1 osteoblast cells by upregulating osteogenic markers such as bone morphogenetic protein (BMO)-2, runt-related transcription factor (RUNX)-2 and β-catenin. SSCP enhanced bone formation and suppressed bone resorption by activating the WNT/β-catenin and BMP/Smad signaling pathways. SSCP offers a dual modulatory approach to bone health, addressing both bone formation and resorption. Its activation of key osteogenic pathways and improvement in bone structural integrity highlight its therapeutic potential for managing osteoporosis and enhancing skeletal health. By activating key osteogenic pathways and normalizing bone metabolism markers, SSCP presents a promising therapeutic candidate for osteoporosis and other bone-related conditions. Further clinical studies are needed to confirm these findings and explore its potential applications. © 2024 Society of Chemical Industry.

Study on the antioxidant and antiosteoporotic activities of the oyster peptides prepared by ultrasound-assisted enzymatic hydrolysis.

In this study, the effects of ultrasound-assisted enzymatic hydrolysis on the production of antioxidant and antiosteoporotic peptides derived from oysters were investigated. Results showed that ultrasound-assisted enzymatic hydrolysis significantly enhanced the peptide content, free radical scavenging ability, and ferric reducing antioxidant power of total oyster protein hydrolysate (TOPH), with optimal results achieved at 200W (TOPH-200). Correspondingly, ultrasound treatment at 200W increased the exposure of hydrophobic regions, reduced α-helix content, and facilitated the generation of small molecular weight peptides in TOPH. In an H2O2-induced oxidative damage model of osteoblastic MC3T3-E1 cells, TOPH-200 significantly attenuated intracellular reactive oxygen species and improved mitochondrial membrane potential. Importantly, TOPH-200 effectively enhanced osteogenic cell proliferation, differentiation, and mineralization in H2O2-treated MC3T3-E1 cells. Additionally, two novel peptides, DSQLAPFRF and HFNPRL, were screened from the TOPH-200 using PeptideRanker and molecular docking. Further cell experiments indicated that both peptides exhibited potent antioxidant and antiosteoporotic activities in oxidatively damaged MC3T3-E1 cells. In summary, mild ultrasound-assisted enzymatic hydrolysis proved effective in producing bioactive peptides from oysters, and these newly identified peptides exhibit potential for osteoporosis prevention.

Tailoring magnesium-doped coatings for improving surface and biological properties of titanium-based dental implants

Physicochemical modifications of biomaterials have been proposed to overcome bone integration impairment and microbial infections. The magnesium (Mg) incorporation on dental implant surfaces has shown positive results in bone-to-implant contact and in the reduction of microbial colonization. Here, we explored the potential of using different Mg precursors to synthesize coatings via plasma electrolytic oxidation (PEO) on commercially pure titanium (cpTi), aiming to optimize the surface and biological properties. For this, we investigated Mg acetate and Mg nitrate precursors in different concentrations (0.04 M and 0.12 M), using calcium (Ca) and phosphorus (P) as the base electrolyte for all groups. Coatings with only the CaP base electrolyte were used as the control group. The surfaces were characterized by confocal laser scanning microscopy, scanning electron microscopy, film thickness measurement, profilometry, wettability, X-ray diffraction, energy dispersive spectroscopy, X-ray photoelectron spectroscopy, electrochemical behavior, and ion release. For biological analyses, the adhesion (2 h) of Streptococcus sanguinis was evaluated, as well as MC3T3-E1 osteoblastic cells proliferation at 1 and 3 days, and mineralization of calcium phosphates after 28 days. PEO treatment using different Mg precursors promoted physicochemical modifications of cpTi. The experimental groups MgN 0.04 and MgN 0.12 exhibited higher surface roughness and wettability compared to the other surfaces. Regardless of the Mg precursor, the higher the ion concentration in the electrolyte solution, the higher the Mg atomic concentration on the surfaces. Concerning the electrochemical behavior, the results indicated that the incorporation of Mg in the coatings may enhance the electrochemical performance. Mg treated surfaces did not promote greater bacterial adherence when compared to the control. MgAc 0.04 and MgAc 0.12 coatings displayed improved MC3T3-E1 pre-osteoblastic cells proliferation at day 3 compared to other groups. The hydroxyapatite formation on MgAc 0.12 surfaces was higher than in the other groups. Our data indicate that Mg precursor selection positively influences physicochemical and biological properties of coatings. Specifically, MgAc 0.12 surfaces showed the most promising surface features with greater cell proliferation, without affecting microbial colonization, being an excellent candidate for surface treatment of titanium-based dental implants.

Application of hydrostatic pressure up-regulates sost gene expression in osteocytic spheroids.

In this study, we developed a hydrostatic pressurizing chamber capable of applying hydrostatic pressure to osteocytic spheroids derived from mouse osteoblastic MC3T3-E1 cells. Our results demonstrate that a 4-hour exposure to 200kPa of hydrostatic pressure did not alter the apparent morphology of the spheroids. However, gene expression analysis revealed a significant up-regulation of Sost, marker of late-stage osteocyte differentiation.

Osteoprotegerin secretion and its inhibition by RANKL in osteoblastic cells visualized using bioluminescence imaging

Bone remodeling is regulated by the interaction between receptor activator of nuclear factor kappa-B ligand (RANKL) and its receptor RANK on osteoblasts and osteoclasts, respectively. Osteoprotegerin (OPG) is secreted from osteoblasts and inhibits osteoclast differentiation by acting as a decoy receptor for RANKL. Despite its importance, the mechanism underlying the secretion of OPG remains poorly understood. Here, we applied a method of video-rate bioluminescence imaging using a fusion protein with Gaussia luciferase (GLase) and visualized the secretion of OPG from living mouse osteoblastic MC3T3-E1 cells. The bioluminescence imaging revealed that the secretion of OPG fused to GLase (OPG-GLase) occurred frequently and widely across the cell surface. Notably, co-expression of RANKL significantly reduced the secretion of OPG-GLase, indicating an inhibitory role of RANKL on OPG secretion within cells. Further imaging and biochemical analyses using deletion mutants of OPG and RANKL, as well as RANKL mutants that cause autosomal recessive osteopetrosis, demonstrated the essential role of protein-protein interaction between OPG and RANKL in the inhibition of OPG secretion. Treatment with proteasome inhibitors resulted in increased levels of OPG in both culture medium and cell lysates. However, the fold-increase of OPG was similar regardless of the presence or absence of RANKL, suggesting that the regulation of OPG secretion by RANKL is independent of proteasome activity. This report visualized the secretion of OPG from living cells and provided evidence for a novel intracellular inhibitory effect of RANKL on OPG secretion.

Fabrication and in vitro biological properties of hydroxyapatite-sodium potassium niobate-barium titanate piezoelectric bioceramics

The utilization of piezoelectric materials in bone implants is appealing due to the inherent piezoelectric property of natural bone. The intrinsic electrical characteristics of piezoelectric biomaterials enhance antibacterial activities, biocompatibility, and bioactivity properties. This study delves into investigating the antibacterial properties, biocompatibility, and bioactivity of three-component biocomposites: sodium potassium niobate (KNN)-barium titanate (BT)-hydroxyapatite (HA). The combination of sodium potassium niobate and barium titanate, possessing suitable piezoelectric properties, with hydroxyapatite, known for its favorable biological properties, enhances the requisite properties for a bone implant. Among the various compositions studied, the combination comprising 70 wt% of the piezoelectric component (KNN-BT) and 30 wt% hydroxyapatite, labeled as 30HKB, emerged as the most optimal blend in terms of density, morphotropic phase boundary, and other biological tests conducted. Following polarization, the antibacterial efficacy of 30HKB against S. aureus bacteria cells increased by 61 %. Furthermore, the growth and adhesion of MC3T3-E1 osteoblast cells suggest enhanced biocompatibility of the 30HKB composite attributed to surface polarization. The surface charges generated by polarization facilitated the absorption of Ca2+ ions, as well as the interaction of HPO4 - and OH- ions with the precipitated Ca2+ ions, leading to the formation of the CaP layer. Hence, polarized piezoelectric ceramics exhibit heightened bioactivity compared to their non-polarized counterparts.

O-GlcNAcylation regulates osteoblast differentiation through the morphological changes in mitochondria, cytoskeleton, and endoplasmic reticulum.

To explore the potential mechanisms which O-linked-N-acetylglucosaminylation (O-GlcNAcylation) regulates osteogenesis, a publicly RNA-seq dataset was re-analyzed with literature-mining and showed the primary targets of O-GlcNAcylation in osteoblasts are mitochondria/cytoskeleton. Although the O-GlcNAcylation-regulated mitochondria/cytoskeleton has been extensively studied, its specific role during osteogenesis remains unclear. To address this, we knocked out Ogt (Ogt-KO) in MC3T3-E1 osteoblastic cells. Then, significantly reduced osteoblast differentiation, motility, proliferation, mitochondria-endoplasmic reticulum (Mito-ER) coupling, volume of ER, nuclear tubulins, and oxygen metabolism were observed in Ogt-KO cells. Through artificial intelligence (AI)-predicted cellular structures, the time-lapse live cells imaging with reactive-oxygen-species/hypoxia staining showed that lower cell proliferation and altered oxygen metabolism in the Ogt-KO cells were correlated with the Mito-ER coupling. Bioinformatics analysis, combined with correlated mRNA and protein expression, suggested that Ezh2 and its downstream targets (Opa1, Gsk3a, Wnt3a, Hif1a, and Hspa9) may be involved in O-GlcNAcylation-regulated Mito-ER coupling, ultimately impacting osteoblast differentiation. In conclusion, our findings indicate that O-GlcNAcylation-regulated osteoblast differentiation is linked to morphological changes in mitochondria, cytoskeleton, and ER, with Ezh2 potentially playing a crucial role.

A biodegradable Fe–0.6Se alloy with superior strength and effective antibacterial and antitumor capabilities for orthopedic applications

Iron–selenium (Fe–Se) alloys have potential as attractive biodegradable bone–implant materials, given the antitumor properties of Se in cancer prevention and therapy. However, the fabrication of Fe–Se alloys is challenging due to the volatility of elemental Se and the significantly different melting points of Se and Fe. In this study, we successfully fabricated Fe–xSe (x = 0.2, 0.4, 0.6, 0.8, and 1 wt.%) alloys using suction casting, with FeSe compounds as the Se source. The microstructures, tensile properties, corrosion behavior, biocompatibility, antibacterial ability, and antitumor properties of the Fe–Se alloys were evaluated. The microstructures of the Fe–Se alloys were composed of α–Fe and FeSe phases. Among the Fe–Se alloys, Fe–0.6Se showed the best combination of tensile properties, with a yield strength of 1096.5 ± 7.2 MPa, an ultimate tensile strength of 1271.6 ± 6.3 MPa, and a fracture strain of 15.6 ± 3.3 %, and a degradation rate of 56.9 ± 0.4 μm/year. Moreover, the Fe–0.6Se alloy showed superb antibacterial ability against S. aureus, antitumor activity against 143B osteosarcoma cells, and osteogenicity and biocompatibility toward pre–osteoblast MC3T3–E1 cells. In summary, adding 0.2–1.0 wt.% Se to Fe does not affect the growth of healthy cells but effectively inhibits the growth and reproduction of tumor cells, and the Fe–0.6Se alloy is promising for orthopedic applications owing to its unique combination of mechanical and biofunctional properties. Statement of significanceThis work reports on Fe-xSe (x = 0.2, 0.4, 0.6, 0.8, and 1 wt.%) alloys fabricated using suction casting. The microstructures of the Fe–Se alloys were composed of α-Fe and FeSe phases. Among the Fe–Se alloys, the Fe-0.6Se showed the best combination of tensile properties, with a yield strength of 1058.6 ± 3.9 MPa, an ultimate tensile strength of 1134.1 ± 2.9 MPa, and a fracture strain of 16.8 ± 1.5 %, and a degradation rate of 56.9 ± 0.4 μm/year. Moreover, the Fe-0.6Se alloy showed superb antibacterial ability against S. aureus, antitumor activity against 143B osteosarcoma cells, and significant osteogenic ability and biocompatibility toward pre-osteoblast MC3T3-E1 cells. In summary, the Fe-0.6Se alloy is promising for orthopedic applications owing to its unique combination of mechanical and biofunctional properties.

7050 Effect of Spironolactone On Reducing Sugar-Induced Cellular Damage In MC3T3-E1 Osteoblastic Cells

Abstract Disclosure: S. Yun: None. H. Sang: None. S. Park: None. K. Suh: None. H. Kim: None. S. Chin: None. Background: In diabetes, prolonged hyperglycemia causes oxidative stress, resulting in pancreatic beta cells' dysfunction and diabetic complications. Oxidative stress can inhibit osteoblast differentiation and induce dysfunction and apoptosis. Spironolactone is an aldosterone antagonist used to treat hypertension and heart failure. In this study, we observed whether spironolactone can reduce osteoblast cytotoxicity and differentiation caused by oxidative stress induced by 2-deoxy-d-ribose (dRib). Methods: MC3T3-E1 cells were treated with spironolactone (0-100 uM) in the presence of 15 mM dRib. The cytotoxicity (lactate dehydrogenase, LDH) and differentiation (collagen content, alkaline phosphatase [ALP] activity, and mineralization) of osteoblasts, inflammatory cytokines (tumor necrosis factor-α [TNF-α], interleukin [IL]-6), oxidative stress (reactive oxygen species [ROS], mitochondrial superoxide), endoplasmic reticulum (ER) stress (activating transcription factor 6 [ATF-6], inositol-requiring 1 [IRE1]), mitochondrial function (mitochondrial membrane potential [MMP], adenosine triphosphate [ATP]), glyoxalase I activity, and glutathione (GSH) were measured. Results: Pretreatment of MC3T3-E1 osteoblastic cells with spironolactone prevented the dRib-induced cytotoxicity of osteoblasts and promoted differentiation of osteoblasts suppressed by dRib. Spironolactone significantly reduced dRib-induced inflammatory cytokines, ROS, mitochondrial superoxide, and ER stress. dRib-induced mitochondrial dysfunction was significantly improved by treatment with spironolactone. dRib is detoxified by the glyoxalase system. Pretreatment with spironolactone also increased the level of reduced GSH and the activity of glyoxalase I in dRib. Conclusion: Spironolactone could reduce dRib-induced cytotoxicity and promote differentiation in MC3T3-E1 osteoblastic cells by reducing oxidative stress and inflammatory cytokines, increasing mitochondrial biogenesis, and detoxifying dRib. These results suggest that spironolactone can be useful in the prevention and treatment of diabetic bone disease. Presentation: 6/1/2024

Bioactive matrix scaffold of oxidized sacchachitin via green catalyst of siliacat tempo for enhancing bone regeneration

Bone regeneration remains a critical challenge in orthopedic medicine. Extracellular matrix proteins play important roles in bionic mineralization and osteogenesis. Sacchachitin nanofibers (SCNFs) oxidized via TEMPO have shown great potential as biomaterial scaffolds due to the introduction of carboxylic groups that mimic the natural extracellular matrix. However, TEMPO is harmful to the environment, which calls for greener and sustainable alternatives. Therefore, this study explored the green fabrication of oxidized-SCNFs through SiliaCat-TEMPO (STEMPO) as a renewable catalyst with various amount of oxidation agents to evaluate their bone regeneration ability. We developed an optimal STEMPO oxidation process, enabling efficient removal of most STEMPO particles through centrifugation with minimal residue. The oxidation efficiency of recovered STEMPO particles remained comparable to fresh ones. STEMPO-oxidized SCNFs, oxidized at increasing NaClO levels, showed an increase in carboxylate content, from 0.1360 mmol/g to 3.2415 mmol/g. Additionally, STEMPO-oxidized SCNFs exhibited excellent biocompatibility with MC3T3-E1 osteoblastic cells, facilitating calcium precipitation, cell migration, and osteogenic differentiation. Mixed composites of β-tricalcium phosphate (BCP) and STEMPO-oxidized SCNFs with higher oxidation levels further enhanced osteogenic differentiation. In vivo studies using a rat femur defect model showed that SCNFs at the highest oxidation level (ST100SC) significantly promoted bone regeneration to full recovery, achieving a bone volume to total tissue volume (BV/TV) ratio of 75.6 %, compared to 48.3 % in the control. These results demonstrated that STEMPO-oxidized SCNFs with higher oxidation levels can be used as a bioactive matrix scaffold to achieve full recovery of bone regeneration.

Gallein increases prostaglandin F2α‑induced osteoprotegerin and IL‑6 secretion in osteoblasts.

Gallein is a known Gβγ subunit inhibitor, but its function in bone metabolism, especially in osteoblasts, and its molecular mechanism remains to be elucidated. Osteoprotegerin (OPG), which is secreted from osteoblasts, binds to nuclear factor kB receptor activator (RANK) ligand (RANKL) as a decoy receptor, prevents RANKL-RANK binding, and inhibits bone resorption. IL-6 is not only a bone resorption factor but also as a bone metabolism regulator. Prostaglandin F2α (PGF2α) promotes p44/p42 MAPK, p38 MAPK and stress-activated protein kinase/JNK phosphorylation in osteoblast-like MC3T3-E1 cells. In MC3T3-E1 cells, activated p44/p42 and p38 MAPKs promote IL-6 secretion and activated p44/p42 and p38 MAPKs and JNK promote OPG secretion. The present study aimed to investigate the effect and mechanism of gallein on PGF2α-induced OPG and IL-6 secretion using an osteoblastic MC3T3-E1 cell line. It was found that gallein significantly increased PGF2α-induced OPG and IL-6 secretion in the MC3T3-E1 cell. By contrast, fluorescein, which is a gallein-like compound that does not bind to Gβγ, did not affect PGF2α-induced OPG and IL-6 secretion. Gallein significantly improved the PGF2α-induced OPG and IL-6 mRNA expression levels. Gallein did not affect the PGF2α-activated phosphorylation of p44/p42 and p38 MAPKs and JNK. Gallein increased PGF2α-induced OPG and IL-6 secretion in osteoblasts, indicating that gallein may regulate bone remodeling via OPG/IL-6 in bone metabolism.

Mubong-derived Nanovesicles Regulate Differentiation and Mineralization in Osteoblastic MC3T3-E1 Cells

Mubong-derived Nanovesicles Regulate Differentiation and Mineralization in Osteoblastic MC3T3-E1 Cells

Suppression of bone resorption in ovariectomized mice using estrogen-immobilized polyphosphodiesters

Estrogens play a crucial role in bone remodeling. In this study, we synthesized two types of bone-targeted polyphosphodiesters (PPDEs): poly(diethyl sodium phosphate) (PEP•Na) and β-estradiol 17-valerate terminated PEP•Na (E2V-PEP•Na), and determined the inhibitory effect of the terminal estrogen on ovariectomy-induced bone resorption. We first assessed the impact of these polymers on the differentiation of osteoblasts and osteoclasts in vitro. Both PPDEs effectively promoted the differentiation of mouse osteoblastic MC3T3-E1 cells, with E2V-PEP•Na significantly enhancing the mineralization of these cells. Conversely, the density of osteoclasts formed from bone marrow mononuclear cells (BMNCs) significantly decreased when exposed to E2V-PEP•Na. Additionally, the formation of resorption pits on calcium phosphate-coated plates was reduced after treating BMNCs with both PPDEs. We then investigated the role of these polymers in treating osteoporosis using ovariectomized (OVX) mice. PEP•Na or E2V-PEP•Na was separately injected into the tail vein of the mice three times at 2-week intervals, starting 1 day after OVX treatment. Two weeks after the final injection, we observed that E2V-PEP•Na was distributed in bone tissue in a dose-dependent manner. The injection of E2V-PEP•Na significantly reduced bone resorption, and all bone parameters of OVX mice treated with E2V-PEP•Na were superior to those of sham mice. In contrast, PEP•Na did not affect bone mineral metabolism. In conclusion, our study is the first to demonstrate that E2V-PEP•Na can inhibit bone resorption in vivo, offering a promising new polymeric drug for preventing osteoporosis.

Biological activity of tantalum oxide coating doped with calcium and strontium by micro-arc oxidation under osteoimmunomodulation

Tantalum is widely used in clinics due to its excellent properties. However, its high density and elastic modulus constrain its application as a large implant. The new porous tantalum material addresses the issue of incompatibility between the implant and the human body. Nevertheless, it still exhibits biological inertia and limited bone-bonding ability; therefore, it is necessary to modify its surface. At present, some surface modification methods have inconsistent results in vitro and in vivo. To solve this problem, this study described the preparation of a ceramic coating with a "trabeculae-like" structure on the surface of pure tantalum using micro-arc oxidation (MAO). Calcium and strontium ions were introduced by adding acetate to the electrolyte. The effects of incorporating bioactive ions Ca2+ and Sr2+ into the electrolyte on the coating's morphology, coating thickness, phase composition, roughness, hydrophilicity, and the activity of MC3T3-E1 osteoblast cells under osteoimmunomodulation were investigated. The results demonstrated that the introduction of bioactive ions did not change the porous morphology of the coating; instead, it led to an increase in coating thickness, enhancement of surface roughness and hydrophilicity, as well as promotion of osteoblast spreading, proliferation, and ALP activity. Moreover, it regulated the phenotypic polarization of RAW264.7 macrophages to augment osteoblast proliferation, spreading, and ALP activity, implying that the coating has superior osteogenic potential under osteoimmunomodulation.

Effect of rare earth elements Ce and Yb on the in vitro properties of biodegradable Zn alloys

Biodegradable pure Zn and zinc alloys have received much attention in recent years, but the mechanical properties, corrosion behavior and biocompatibility of the previous reported pure zinc and zinc alloys do not fully meet the clinical requirements. Alloying and deformation technology are widely used in improving the properties of biodegradable pure Zn and zinc alloys. In this work, we have developed two new types of binary zinc alloys with the two rare earth elements (REE) cerium (Ce) and ytterbium (Yb). The effects of Ce and Yb on the microstructure, mechanical properties, corrosion behavior, in vitro biocompatibility of biodegradable Zn-xCe and Zn-xYb (x = 0.1, 0.2, 0.5, 1.0 wt %) were systematically investigated. The ultimate tensile strength, yield strength and elongation of Zn-xCe and Zn-xYb alloys are significantly improved compared with pure Zn. The Zn-xCe and Zn-xYb alloys have superior antibacterial properties The osteoblast MC3T3-E1 cells exhibited excellent cell activity with polygonal or fusiform extension and adhesion cultured in the series of Zn-xCe and Zn-xYb alloys’ extract solutions, indicating the good in vitro biocompatibility of Zn-xCe and Zn-xYb alloys. This work demonstrated that the Zn-xCe and Zn-xYb binary alloys have great potential as new biodegradable metals for future biomedical implants and devices.

Yunnan Baiyao Might Mitigate Periodontitis Bone Destruction by Inhibiting Autophagy and Promoting Osteoblast Differentiation in vivo, ex vivo and in vitro.

Periodontitis is an inflammatory disease that eventually destroys tooth-supporting tissue. Yunnan Baiyao (YNBY), a traditional Chinese medicine compound with haemostatic and anti-inflammatory properties has shown therapeutic potential in several diseases. Our previous study revealed that YNBY suppressed osteoclast differentiation in periodontitis. The purpose of this study is to investigate the influences of YNBY on osteoblasts and explore its potential mechanisms. A rat periodontitis model was established by ligation of maxillary second molars. After the end of modelling, histopathological observation by hematoxylin-eosin (HE) staining and Masson trichrome staining, detection of bone resorption by Micro-CT scanning, detection of osteoclasts by tartrate-resistant acid phosphatase (TRAP) staining, expression of osteocalcin (OCN) and microtubule-associated protein 1 light chain 3 (LC3) by immunohistochemistry. Lipopolysaccharides was used to irritate MC3T3-E1 osteoblastic cells and ex vivo calvarial organ as an in vitro model of inflammation. CCK-8 assay was performed to examine the toxicity of YNBY to MC3T3-E1 osteoblastic cells. Osteogenesis was assessed with alizarin red staining, immunofluorescence staining, Western blot and immunohistochemical staining. Transmission electron microscopy, fluorescent double staining, Western blot and immunohistochemical staining were employed to detect autophagy. Histological and micro-CT analyses revealed that YNBY gavage reduced bone loss caused by experimental periodontitis and upregulated osteogenic proteins in vivo. YNBY attenuated the production of autophagy-related proteins in periodontitis rats. Additionally, YNBY promoted osteogenesis by inhibiting inflammation-induced autophagy in vitro. Furthermore, YNBY suppressed LPS-mediated bone resorption and promoted the production of osteoblast-related proteins in inflamed calvarial tissues ex vivo. This study demonstrated, through in vivo, in vitro and ex vivo experiments, that YNBY promoted osteoblast differentiation by suppressing autophagy, which markedly alleviated bone destruction caused by periodontitis.

Caspase-9 Is a Positive Regulator of Osteoblastic Cell Migration Identified by diaPASEF Proteomics.

Caspase-9 is traditionally considered the initiator caspase of the intrinsic apoptotic pathway. In the past decade, however, other functions beyond initiation/execution of cell death have been described including cell type-dependent regulation of proliferation, differentiation/maturation, mitochondrial, and endosomal/lysosomal homeostasis. As previous studies revealed nonapoptotic functions of caspases in osteogenesis and bone homeostasis, this study was performed to identify proteins and pathways deregulated by knockout of caspase-9 in mouse MC3T3-E1 osteoblasts. Data-independent acquisition-parallel accumulation serial fragmentation (diaPASEF) proteomics was used to compare protein profiles of control and caspase-9 knockout cells. A total of 7669 protein groups were quantified, and 283 upregulated/141 downregulated protein groups were associated with the caspase-9 knockout phenotype. The deregulated proteins were mainly enriched for those associated with cell migration and motility and DNA replication/repair. Altered migration was confirmed in MC3T3-E1 cells with the genetic and pharmacological inhibition of caspase-9. ABHD2, an established regulator of cell migration, was identified as a possible substrate of caspase-9. We conclude that caspase-9 acts as a modulator of osteoblastic MC3T3-E1 cell migration and, therefore, may be involved in bone remodeling and fracture repair.

Scratch resistance and in-vitro biocompatibility of plasma-sprayed baghdadite coatings reinforced with carbon nanotubes

Hydroxyapatite (HA, Ca10(PO4)6(OH)2) coatings, commonly used for metallic implants, have limitations such as excessive brittleness, low fracture toughness, inadequate wear resistance, and slow osseointegration properties. In contrast, baghdadite (BAG, Ca3ZrSi2O9), a calcium zirconium silicate-based bioceramic, exhibits outstanding biological properties, to promote cell proliferation and differentiation of human osteoblasts along with adequate mechanical strength. In this study, plasma-sprayed HA, BAG, and carbon nanotubes (CNT, 1 wt%, and 2 wt%) reinforced BAG coatings are deposited on titanium (Ti) substrate to enhance its mechanical as well as biological properties. The microhardness values of CNT-reinforced BAG coatings increase due to a decrease in porosity and the retention of CNT inside the BAG matrix. Progressive loading scratch tests are performed, revealing a reduction in wear volume loss from 15.051 mm3 to 12.574 mm3 and scratch rate from 43.262 mm3N−1 m−1 to 36.172 mm3N−1 m−1 with the addition of 2 wt% CNT in BAG coatings. The scratch hardness test results demonstrated that introducing CNT into BAG coating significantly enhances its performance, showing a remarkable 38.7 % improvement. In-vitro cell culture studies indicate excellent cell adhesion, growth, and proliferation of MC3T3-E1 osteoblast cells on the surfaces of the CNT-reinforced BAG coatings.