Osteoblast Cell Culture Research Articles

A salmon DNA scaffold promotes osteogenesis through activation of sodium-dependent phosphate cotransporters

We previously reported the promotion of bone regeneration in calvarial defects of both normal and ovariectomy-induced osteoporotic rats, with the use of biodegradable DNA/protamine scaffold. However, the method by which this DNA-containing scaffold promotes bone formation is still not understood. We hypothesize that the salmon DNA, from which this scaffold is derived, has an osteoinductive effect on pre-osteoblasts and osteoblasts. We examined the effects of salmon DNA on osteoblastic differentiation and calcification in MC3T3-E1 cells, mouse osteoblasts, in vitro and bone regeneration in a calvarial defect model of aged mouse in vivo. The salmon DNA fragments (300 bps) upregulated the expression of the osteogenic markers, such as alkaline phosphatase, Runx2, and osterix (Osx) in MC3T3E1 cells compared with incubation with osteogenic induction medium alone. Measurement of phosphate ion concentrations in cultures showed that the DNA scaffold degraded phosphate ions were released to the cell cultures. Interestingly, we found that the inclusion of DNA in osteoblastic cell cultures upregulated the expression of sodium-dependent phosphate (NaPi) cotransporters, SLC20A1 and SLC34A2, in MC3T3-E1 cells in a time dependent manner. Furthermore, the inclusion of DNA in cell cultures increased the transcellular permeability of phosphate. Conversely, the incubation of phosphonoformic acid, an inhibitor of NaPi cotransporters, attenuated the DNA-induced expression and activation of SLC20A1 and SLC34A2 in MC3T3-E1 cells, resulting in suppression of the osteogenic markers. The implantation of a salmon DNA scaffold disk promoted bone regeneration using calvarial defect models in 30-week-old mice. Our results indicate that the phosphate released from salmon DNA upregulated the expression and activation of NaPi cotransporters, resulting in the promotion of bone regeneration.

Orthotopic Osteogenecity Enhanced by a Porous Gelatin Sponge in a Critical‐Sized Rat Calvaria Defect

The gelatin (Gel) powders, derived from acidic and basic extractions of porcine dermis (referred to as AE and BE), were processed for the porous sponge preparation. The disks, which were less than or greater than 500 μm in diameter [small (S) and large (L) pores, respectively] in both extractions and had an interconnected structure respectively, were implanted in critical-sized defects (CSD) of rat calvaria for 4 and 8 weeks to analyze the bone repair capability. Only the AE-S disk induced bone formation (over 60%) histomorphometrically in the CSD after 8 weeks, although the collagen orientation of the regenerated bone was still immature. Osteoblastic cell culture until 14 days did not substantiate marked superiority of AE-S disk regarding the proliferation and the differentiation, although the initial attachment was enhanced on AE-S disk than BE-L disk. The results provide the findings that a Gel sponge with a specific porous structure is capable of inducing orthotopic bone formation in vivo environment.

Effects of surface treatments on Y-TZP phase stability, microstructure and osteoblast cell response

The effects of hydrothermal and phosphoric acid treatments on the tetragonal ZrO2 stability and surface microstructure were determined for 3mol% yttria- partially stabilized zirconia (Y-TZP) and Al2O3 doped Y-TZP (Y-TZPA). Y-TZP slip cast compacts were sintered at 1300–1500°C to achieve fully dense samples. The increase in the sintering temperature enhanced the hydrothermal degradation of Y-TZP. Although the average grain size of Y-TZPA sintered at 1500°C was lower than that of Y-TZP, no significant difference in the ageing behavior between these two ceramics was found. For Y-TZP sintered at 1500°C, the volume fraction of monoclinic ZrO2 induced by acid treatment was higher than that obtained after 10h of hydrothermal treatment. The Al2O3 doping of Y-TZP improved the acid degradation resistance. Osteoblast cell culture studies on the Y-TZP surface confirmed its biocompatibility. The Y-TZP surface supported a higher cell proliferation compared to Y-TZPA. The less degradation on the 1 day acid-treated Y-TZPA surface stimulated cell proliferation, whereas a longer acid treatment reduced cell survival.

Niobium-Doped Hydroxyapatite Bioceramics: Synthesis, Characterization and In Vitro Cytocompatibility.

Doping calcium phosphates with ionic species can play an important role in biological responses promoting alkaline phosphatase activity, and, therefore inducing the generation of new bone. Thus, in this study, the synthesis of niobium-doped hydroxyapatite (Nb-HA) nanosize particles obtained by the precipitation process in aqueous media followed by thermal treatment is presented. The bioceramics were extensively characterized by X-ray diffraction, wavelength dispersive X-ray fluorescence spectrometry, Fourier transform infrared spectroscopy, scanning electron microscopy/energy dispersive X-ray spectroscopy analysis, transmission electron microscopy, atomic force microscopy and thermal analysis regarding their chemical composition, structure and morphology. The results showed that the precipitate dried at 110 °C was composed of amorphous calcium phosphate and HA, with polidisperse particles ranging from micro to nano dimensions. After the thermal treatment at 900 °C, the bioceramic system evolved predominantly to HA crystalline phase, with evident features of particle sintering and reduction of surface area. Moreover, the addition of 10 mol% of niobium salt precursor during the synthesis indicated the complete incorporation of the Nb(V) species in the HA crystals with detectable changes in the original lattice parameters. Furthermore, the incorporation of Nb ions caused a significant refinement on the average particle size of HA. Finally, the preliminary cytocompatibility response of the biomaterials was accessed by human osteoblast cell culture using MTT and resazurin assays, which demonstrated no cytotoxicity of the Nb-alloyed hydroxyapatite. Thus, these findings seem promising for developing innovative Nb-doped calcium phosphates as artificial biomaterials for potential use in bone replacements and repair.

Indoxyl sulfate exacerbates low bone turnover induced by parathyroidectomy in young adult rats

Low-turnover bone disease is one of the bone abnormalities observed in patients with chronic kidney disease (CKD) and is recognized to be associated with low serum parathyroid hormone (PTH) level and skeletal resistance to PTH. Indoxyl sulfate (IS) is a representative uremic toxin that accumulates in the blood as renal dysfunction progresses in CKD patients. A recent in vitro study using an osteoblastic cell culture system suggests that IS has an important role in the pathogenesis of low bone turnover through induction of skeletal resistance to PTH. However, the effects of IS on the progression of low bone turnover have not been elucidated. In the present study, we produced rats with low bone turnover by performing parathyroidectomy (PTX) and fed these rats a diet containing indole, a precursor of IS, to elevate blood IS level from indole metabolism. Bone metabolism was evaluated by measuring histomorphometric parameters of secondary spongiosa of the femur. Histomorphometric analyses revealed significant decreases in both bone formation–related parameters and bone resorption–related parameters in PTX rats. In indole-treated PTX rats, further decreases in bone formation–related parameters were observed. In addition, serum alkaline phosphatase activity, a bone formation marker, and bone mineral density of the tibia tended to decrease in indole-treated PTX rats. These findings strongly suggest that IS exacerbates low bone turnover through inhibition of bone formation by mechanisms unrelated to skeletal resistance to PTH.

In vitro evaluation of the effects of risedronate associated with cobalamin on osteoblastic cells

OBJECTIVE: To evaluate the action of risedronate and cobalamin, and effects when associated, when administered osteoblastic cells. METHODS: The MC3T3 cells were cultivate in the media α-MEM and α-MEM supplemented with mineralizing factors, ascorbic acid and disodium α-glyicerophosphate, and treated with risedronate, cobalamin, and risedronate associated with cobalamin in a concentration of 10-3 M. The cell proliferation and formation of calcium and phosphate nodules were evaluated at 24 hours, 48 hours, 72 hours, 5 days and 7 days via a Neubauer Chamer count, Alizarin and the von Kossa reaction. RESULTS: The results showed that the growth curve for cell proliferation and the formation of mineral nodules was similar for both cultures analyzed. CONCLUSION: The conclusion was reached that using risedronate, cobalamin and both drugs in combination on osteoblastic cell cultures does not cause alterations to their growth or in the formation of calcium and phosphate nodules.

Validation of an in vitro 3D bone culture model with perfused and mechanically stressed ceramic scaffold.

An engineered three dimensional (3D) in vitro cell culture system was designed with the goal of inducing and controlling in vitro osteogenesis in a reproducible manner under conditions more similar to the in vivo bone microenvironment than traditional two-dimensional (2D) models. This bioreactor allows efficient mechanical loading and perfusion of an original cubic calcium phosphate bioceramic of highly controlled composition and structure. This bioceramic comprises an internal portion containing homogeneously interconnected macropores surrounded by a dense layer, which minimises fluid flow bypass around the scaffold. This dense and flat layer permits the application of a homogeneous loading on the bioceramic while also enhancing its mechanical strength. Numerical modelling of constraints shows that the system provides direct mechanical stimulation of cells within the scaffold. Experimental results establish that under perfusion at a steady flow of 2 µL/min, corresponding to 3 ≤ Medium velocity ≤ 23 µm/s, mouse calvarial cells grow and differentiate as osteoblasts in a reproducible manner, and lay down a mineralised matrix. Moreover, cells respond to mechanical loading by increasing C-fos expression, which demonstrates the effective mechanical stimulation of the culture within the scaffold. In summary, we provide a "proof-of-concept" for osteoblastic cell culture in a controlled 3D culture system under perfusion and mechanical loading. This model will be a tool to analyse bone cell functions in vivo, and will provide a bench testing system for the clinical assessment of bioactive bone-targeting molecules under load.

Does Intrawound Application of Vancomycin Influence Bone Healing in Spinal Surgery?

Introduction Surgical site infections represent a major complication of spinal surgery. The application of lyophilized vancomycin into the wound is reported to significantly decrease infection rates. As concentrations applied locally can exceed the minimal bacterial inhibitory concentration for more than a 1000-fold, toxic side effects on local tissue may be possible. Materials and Methods Primary osteoblast cell cultures were generated from bone tissue samples of 10 patients. Samples were incubated in absence or presence of 3, 6, or 12 mg/cm2 vancomycin according to a planned phase I clinical trial protocol. Changes in pH, osteoblast migration, proliferation, and viability were analyzed. Alkaline phosphatase and mineralization patterns were studied. Results The application of more than 3 mg/cm2 vancomycin induced a decline of pH toward the acidic range. The migration potential of osteoblasts was decreased from 100% (control samples) to zero (12 mg/cm2 vancomycin) in a dose-dependent manner. Cell proliferation was significantly inhibited at dosages above 3 mg/cm2. Significant cell death was observed if the dosage applied exceeded 6 mg/cm2. The synthesis of alkaline phosphatase was markedly reduced in all dosages applied and calcium deposition was significantly decreased in dosages above 3 mg/cm2. Conclusion As bone remodeling requires the immigration, proliferation, and differentiation of osteoblasts at the fusion site, high dosages of intrawound vancomycin might interfere with regenerative processes and increase the risk of nonunion. To allow an appropriate balance of infection risk and the risk of nonunion, the minimal local concentration required should be determined by controlled in vivo studies.

Polypropylene Biocomposites with Boron Nitride and Nanohydroxyapatite Reinforcements.

In this study, we develop binary polypropylene (PP) composites with hexagonal boron nitride (hBN) nanoplatelets and ternary hybrids reinforced with hBN and nanohydroxyapatite (nHA). Filler hybridization is a sound approach to make novel nanocomposites with useful biological and mechanical properties. Tensile test, osteoblastic cell culture and dimethyl thiazolyl diphenyl tetrazolium (MTT) assay were employed to investigate the mechanical performance, bioactivity and biocompatibility of binary PP/hBN and ternary PP/hBN-nHA composites. The purpose is to prepare biocomposite nanomaterials with good mechanical properties and biocompatibility for replacing conventional polymer composites reinforced with large hydroxyapatite microparticles at a high loading of 40 vol%. Tensile test reveals that the elastic modulus of PP composites increases, while tensile elongation decreases with increasing hBN content. Hybridization of hBN with nHA further enhances elastic modulus of PP. The cell culture and MTT assay show that osteoblastic cells attach and proliferate on binary PP/hBN and ternary PP/hBN-20%nHA nanocomposites.

Local delivery of the cationic steroid antibiotic CSA-90 enables osseous union in a rat open fracture model of Staphylococcus aureus infection.

Treatment of infected open fractures remains a major clinical challenge. In this study, we investigated the novel broad-spectrum antibiotic CSA-90 (cationic steroid antibiotic-90) as an antimicrobial agent. CSA-90 was screened in an osteoblast cell culture model for effects on differentiation and mineralization. Local delivery of CSA-90 was then tested alone and in combination with recombinant human bone morphogenetic protein-2 (rhBMP-2) in a mouse ectopic bone formation model (n=40 mice) and in a rat open fracture model inoculated with pathogenic Staphylococcus aureus (n=84 rats). CSA-90 enhanced matrix mineralization in cultured osteoblasts and increased rhBMP-2-induced bone formation in vivo. All animals in which an open fracture had been inoculated with Staphylococcus aureus and not treated with local CSA-90, including those treated with rhBMP-2, had to be culled prior to the experimental end point (six weeks) because of localized osteolysis and deterioration of overall health, whereas CSA-90 prevented establishment of infection in all open fractures in which it was used (p≤0.012). Increased union rates were seen for the fractures treated with rhBMP-2 or with the combination of rhBMP-2 and CSA-90 compared with that observed for the fractures treated with CSA-90 alone (p=0.04). CSA-90 can promote osteogenesis and be used for prevention of Staphylococcus aureus infection in preclinical models. Local delivery of CSA-90 represents a novel strategy for prevention of infection and may have specific benefits in the context of orthopaedic injuries.

Rosmarinic acid exerts an antiosteoporotic effect in the RANKL-induced mouse model of bone loss by promotion of osteoblastic differentiation and inhibition of osteoclastic differentiation.

Bone homeostasis is ensured by the balance between bone formation and resorption. Thus, control of the recruitment, proliferation, and differentiation of bone cells is essential to maintain bone mass. The aim of this study was to elucidate the effects of rosmarinic acid as a potential therapeutic agent on bone metabolism using bone cells and a mouse model. Rosmarinic acid increased alkaline phosphatase activity and induced mineralization in osteoblasts. Addition of rosmarinic acid to cultures of calvarial osteoblastic cells prepared from T-cell factor/β-catenin TOP-GAL mutant mice strongly induced the expression of LacZ and promoted stabilization of β-catenin in the cytoplasm of ST2 cells, suggesting that rosmarinic acid affects the canonical Wnt signaling pathway. Moreover, rosmarinic acid inhibited not only osteoclast formation in cocultures of mouse bone marrow cells and osteoblasts, but also receptor activator of nuclear factor kappa-B ligand (RANKL)-induced osteoclastic differentiation in bone marrow-derived macrophages. RANKL-induced p38 mitogen-activated protein kinase and the expression of nuclear factor of activated T cell, c-Jun, and c-Fos were inhibited by rosmarinic acid in bone marrow macrophages. Finally, we confirmed that rosmarinic acid improved bone mass in a soluble RANKL-induced bone loss mouse model. Rosmarinic acid has dual regulatory effects on bone metabolism and may control the bone functions by controlling osteoblastic and osteoclastic differentiation.

Scaffold Architecture and Properties for Osteoblasts Cell Culture: An Optimization Model and Application by Genetic Algorithm

ABSTRACTIn the developing of scaffolds for cell culture, a large number of architectures with different combinations of properties should be tested to determine the best. This can be costly in time, money and materials. In this paper we have proposed an optimization model that aims to maximize the growth of osteoblasts on polymeric scaffolds by regulating their properties and architecture. Based on the optimization model it was implemented a genetic algorithm to calculate the architecture and properties of the scaffolds. The fiber diameter, pore diameter, porosity, Young's modulus and contact angle of the scaffolds were calculated through four electrospinning parameters: voltage (kV), concentration (% w/v), flow rate (ml/h) and distance (cm). A fitness value was assigned to each scaffold and the highest one was chosen as the best condition for osteoblast growth. The preliminary results obtained by the Genetic Algorithm were consistent with the tendencies reported experimentally in other studies. Also, the methodology established here can be easily adapted to different types of polymers and cells. Finally, the optimization model can be applied not only by means of heuristic method, like a Genetic Algorithm, but also by exact methods.

Cationic Nanogel-mediated Runx2 and Osterix siRNA Delivery Decreases Mineralization in MC3T3 Cells.

Heterotopic ossification (HO) may occur after musculoskeletal trauma, traumatic brain injury, and total joint arthroplasty. As such, HO is a compelling clinical concern in both military and civilian medicine. A possible etiology of HO involves dysregulated signals in the bone morphogenetic protein osteogenic cascade. Contemporary treatment options for HO (ie, nonsteroidal antiinflammatory drugs and radiation therapy) have adverse effects associated with their use and are not biologically engineered to abrogate the molecular mechanisms that govern osteogenic differentiation. We hypothesized that (1) nanogel-mediated short interfering RNA (siRNA) delivery against Runt-related transcription factor 2 (Runx2) and osterix (Osx) genes will decrease messenger RNA expression; (2) inhibit activity of the osteogenic marker alkaline phosphatase (ALP); and (3) inhibit hydroxyapatite (HA) deposition in osteoblast cell cultures. Nanogel nanostructured polymers delivered siRNA in 48-hour treatment cycles against master osteogenic regulators, Runx2 and Osx, in murine calvarial preosteoblasts (MC3T3-E1.4) stimulated for osteogenic differentiation by recombinant human bone morphogenetic protein (rhBMP-2). The efficacy of RNA interference (RNAi) therapeutics was determined by quantitation of messenger RNA knockdown (by quantitative reverse transcription-polymerase chain reaction), downstream protein knockdown (determined ALP enzymatic activity assay), and HA deposition (determined by OsteoImage™ assay). Gene expression assays demonstrated that nanogel-based RNAi treatments at 1:1 and 5:1 nanogel:short interfering RNA weight ratios reduced Runx2 expression by 48.59%±19.53% (p<0.001) and 43.22%±18.01% (both p<0.001). The same 1:1 and 5:1 treatments against both Runx2 and Osx reduced expression of Osx by 51.65%±10.85% and 47.65%±9.80% (both p<0.001). Moreover, repeated 48-hour RNAi treatment cycles against Runx2 and Osx rhBMP-2 administration reduced ALP activity after 4 and 7days. ALP reductions after 4days in culture by nanogel 5:1 and 10:1 RNAi treatments were 32.4%±12.0% and 33.6%±13.8% (both p<0.001). After 7days in culture, nanogel 1:1 and 5:1 RNAi treatments produced 35.9%±14.0% and 47.7%±3.2% reductions in ALP activity. Osteoblast mineralization data after 21days suggested that nanogel 1:1, 5:1, and 10:1 RNAi treatments decreased mineralization (ie, HA deposition) from cultures treated only with rhBMP-2 (p<0.001). However, despite RNAi attack on Runx2 and Osx, HA deposition levels remained greater than non-rhBMP-2-treated cell cultures. Although mRNA and protein knockdown were confirmed as a result of RNAi treatments against Runx2 and Osx, complete elimination of mineralization processes was not achieved. RNAi targeting mid- and late-stage osteoblast differentiation markers such as ALP, osteocalcin, osteopontin, and bone sialoprotein) may produce the desired RNAi-nanogel nanostructured polymer HO prophylaxis. Successful HO prophylaxis should target and silence osteogenic markers critical for heterotopic bone formation processes. The identification of such markers, beyond RUNX2 and OSX, may enhance the effectiveness of RNAi prophylaxes for HO.

Human oral cells' response to different endodontic restorative materials: an in vitro study.

IntroductionThe aim of this study was to compare the biological interaction of human osteoblasts and cells of the human periodontal ligament (PDL) with different endodontic restorative material as Mineral Trioxide Aggregate (MTA), Biodentine, amalgam and composite over a time period of 20 days.Materials and methodsHuman PDL cells and osteoblasts were harvested, cultured and according to standardized protocols. The cell populations were characterized with the corresponding surface markers following standardized procedures. The specimens were produced with special regard to constant dimensions and volume in the different groups. Cell attachment and proliferation were evaluated morphologically after Richardson staining and cell count was performed after 1d, 8d, 13d and 20d. All experiments were performed in triplets. The results were statistically analyzed using the ANOVA- and Tukey-test (p < 0.05).ResultsMorphological analysis proved good proliferation and cell attachment in both cements. A remarkable result was the organized spreading and parallel alignment of the PDL cells in contact with MTA and especially Biodentine (cells maturing in a second cell layer crossway to the first one). From 8d onward Biodentine showed the highest quantity of PDL cells (p < 0.05). Biodentine and MTA resulted in a significantly higher cell density in osteoblast and PDL cell culture. The other groups showed a lower PDL cell density from 8d and a lower osteoblast cell density from 13d when compared to control and cement samples (p < 0.05).ConclusionsMTA and Biodentine showed a good biocompatibility in contact with the human osteoblasts and cells of the periodontal ligament. Regarding cell survival and proliferation particularly of PDL cells Biodentine showed good results and can be considered as a well-tolerated bioactive endodontic material.Electronic supplementary materialThe online version of this article (doi:10.1186/s13005-014-0055-4) contains supplementary material, which is available to authorized users.

Extracellular matrix mineralization in murine MC3T3-E1 osteoblast cultures: An ultrastructural, compositional and comparative analysis with mouse bone

Bone cell culture systems are essential tools for the study of the molecular mechanisms regulating extracellular matrix mineralization. MC3T3-E1 osteoblast cell cultures are the most commonly used in vitro model of bone matrix mineralization. Despite the widespread use of this cell line to study biomineralization, there is as yet no systematic characterization of the mineral phase produced in these cultures. Here we provide a comprehensive, multi-technique biophysical characterization of this cell culture mineral and extracellular matrix, and compare it to mouse bone and synthetic apatite mineral standards, to determine the suitability of MC3T3-E1 cultures for biomineralization studies. Elemental compositional analysis by energy-dispersive X-ray spectroscopy (EDS) showed calcium and phosphorus, and trace amounts of sodium and magnesium, in both biological samples. X-ray diffraction (XRD) on resin-embedded intact cultures demonstrated that similar to 1-month-old mouse bone, apatite crystals grew with preferential orientations along the (100), (101) and (111) mineral planes indicative of guided biogenic growth as opposed to dystrophic calcification. XRD of crystals isolated from the cultures revealed that the mineral phase was poorly crystalline hydroxyapatite with 10 to 20nm-sized nanocrystallites. Consistent with the XRD observations, electron diffraction patterns indicated that culture mineral had low crystallinity typical of biological apatites. Fourier-transform infrared spectroscopy (FTIR) confirmed apatitic carbonate and phosphate within the biological samples. With all techniques utilized, cell culture mineral and mouse bone mineral were remarkably similar. Scanning (SEM) and transmission (TEM) electron microscopy showed that the cultures had a dense fibrillar collagen matrix with small, 100nm-sized, collagen fibril-associated mineralization foci which coalesced to form larger mineral aggregates, and where mineralized sites showed the accumulation of the mineral-binding protein osteopontin. Light microscopy, confocal microscopy and three-dimensional reconstructions showed that some cells had dendritic processes and became embedded within the mineral in an osteocyte-like manner. In conclusion, we have documented characteristics of the mineral and matrix phases of MC3T3-E1 osteoblast cultures, and have determined that the structural and compositional properties of the mineral are highly similar to that of mouse bone.

Ethanolic extract of Coelogyne cristata Lindley (Orchidaceae) and its compound coelogin promote osteoprotective activity in ovariectomized estrogen deficient mice

Coelogyne cristata Lindley (CC) family Orchidaceae is an Indian medicinal plant used for the treatment of fractured bones in folk-tradition of Kumaon region, Uttarakhand, India. In continuation of our drug discovery program, feeding of ethanolic extract to ovariectomized estrogen deficient mice led to significant restoration of trabecular micro architecture in both femoral and tibial bones, better bone quality and also devoid of any uterine estrogenicity. Subsequently, coelogin, a pure compound was isolated from ethyl acetate fraction of C. cristata and evaluated in in vitro osteoblast cell cultures. Treatment of coelogin to osteoblasts led to enhanced ALP activity (a marker of osteoblast differentiation), mineral nodule formation and mRNA levels of osteogenic markers like BMP-2, Type 1 Collagen and RUNX-2. Based on these results, we propose that ethanolic extract of C. cristata and its pure compound coelogin have potential in the management of post menopausal osteoporosis.

Combined effects of infliximab and methotrexate on rheumatoid arthritis osteoblastic cell metabolism.

The goal of this study is to investigate the in vitro effects of two disease-modifying anti-rheumatic drugs, largely used in the treatment of rheumatoid arthritis (RA), [infliximab (IFX) and methotrexate (MTX)], in RA primary osteoblast cell cultures. MTX inhibited proliferation and metabolic activity in RA osteoblasts was able to increase apoptosis. Conversely, IFX increased the proliferation, osteocalcin production and the alkaline phosphatase activity. Interestingly, IFX appeared to antagonise the negative effect exerted by MTX. Both drugs significantly reduced the IL-6 production in osteoblasts when used alone, and the combination of the two agents resulted in a significant additional reduction of IL-6 synthesis, with an apparent additive effect. The present study suggests that MTX exerts negative direct effects on bone metabolism in RA patients, but the combined treatment with anti-TNF-α can be beneficial for the interaction of MTX with bone cells.

β-Cryptoxanthin rich paprika extract prevents ovariectomy induced bone loss in Wistar rats

Recent studies show that high intake of carotenoids is helpful in preventing bone loss in post-menopausal women. Carotenoids like β-cryptoxanthin increases calcium and phosphorus deposition in cultured osteoblasts. β-Cryptoxanthin is also reported to have anabolic property in cultured osteoblast cells. It can also inhibit bone resorption process. Paprika and its dietary products are rich in carotenoids like β-cryptoxanthin. Carotenoids from the paprika oleoresin are also reported to have very good bioavailability in humans. The present study was carried out to investigate the effect of β-cryptoxanthin rich paprika extract on animal model of ovariectomy induced bone loss. Treatment with the test drug was given for 6 weeks. The β-cryptoxanthin rich paprika extract significantly reduced urinary excretion of pyridinium crosslinks, indicating its probable anti-resorptive property. The mechanical strength and density of cancellous bones are significantly improved. Thus the paprika extract may have potential role as a sustainable nutritional approach to improving bone health in post-menopausal condition.

Experimental in vitro infection of rat osteoblasts with measles virus stimulates osteogenic differentiation

In this work we characterized the infection of a primary culture of rat osteoblastic lineage cells (OBCs) with measles virus (MeV) and the effect of infection on cell differentiation and maturation. Infection of OBCs with MeV led to high titers of infectivity released early after infection. Also, analysis of mRNAs corresponding to osteogenic differentiation markers like alkaline phosphatase (ALP), bone sialo-protein (BSP) and bone morphogenetic proteins (BMPs) 1-4-5-7 in OBCs revealed higher values (2–75-fold of increment) for infected cells in comparison with uninfected controls. Differentiation of OBCs in osteogenic medium prior to infection influenced the level of stimulation induced by MeV. Furthermore, treatment of OBCs with Ly294002, a PI3K/AKT inhibitor, increased viral titers, whereas treatment with 10μM or 100μM ATPγS diminished MeV multiplication. In addition, increments of osteogenic differentiation markers induced by MeV infection were not modified either by treatment with Ly294002 or ATPγS. These data provide the first evidence demonstrating that MeV can infect osteoblasts in vitro leading to osteoblastic differentiation, a key feature in bone pathogenic processes like otosclerosis.

Erratum to: Strontium ranelate changes the composition and crystal structure of the biological bone-like apatite produced in osteoblast cell cultures

Erratum to: Strontium ranelate changes the composition and crystal structure of the biological bone-like apatite produced in osteoblast cell cultures