Characterization of tricalcium phosphate solubilization by Stenotrophomonas maltophilia YC isolated from phosphate mines

The phosphate solubilizing characteristics of a strain YC, which was isolated from phosphate mines (Hubei, China), were studied in National Botanical Research Institute’s phosphate (NBRIP) growth medium containing tricalcium phosphate (TCP) as sole phosphorus (P) source. The strain YC is identified as Stenotrophomonas maltophilia (S. maltophilia) based upon the results of morphologic, physiological and biochemical characteristics and 16S rRNA sequences analysis. The results show that the strain S. maltophilia YC can solubilize TCP and release soluble P in NBRIP growth medium. A positive correlation between concentration of soluble P and population of the isolate and a negative correlation between concentration of soluble P and pH in the culture medium are observed from statistical analysis results. Moreover, gluconic acid is detected in the culture medium by HPLC analysis. It indicates that the isolate can release gluconic acid during the solubilizing experiment, which causes acidification of the culture medium and then TCP solubilization. S. maltophilia YC has a maximal TCP solubilizing capability when using maltose as carbon source and ammonium nitrate as nitrogen source, respectively, in NBRIP growth medium.

Biodegradation of Tricalcium Phosphate Ceramics by Osteoclasts

Biodegradation of tricalcium phosphate (TCP) ceramics was observed through mixed culture of osteoclasts and TCP discs in vitro in this study. Osteo-clasts were isolated from newborn SD rat's marrow of long bone and cultured on TCP discs. The culture terminated at the 48th h and 96th h respectively. Under an inverted microscope, the osteoclasts imparted round or oval body with multinu-clear and many thin processes. These cells were positively stained for tartrate-re-sistance acid phosphatase (TRAP). Scanning electron microscope showed that many resorption lacunae on TCP disc surface and their diameters were smaller than 20 μm. Osteoclasts were located in the lacunae. At the 96th h, the resorption lacunae become larger and osteoclasts showed degeneration. It is suggested that osteoclasts possess ability to re-absorb TCP ceramics under in vitro culturing condition.

The Bacterial Inhibitory Ability and In Vivo Drug Release Pattern of a New Drug Delivery System:Ciprofloxacine/tricalcium Phosphate Delivery Capsule

The bacterial inhibitory ability of a new drug delivery system (DDS):Ciprofloxacine/tricalcium phosphate delivery capsule (CTDC), its in vivo drug release pattern, and the influence of ultrasonic irradiation on its drug release were investigated. It was found that CTDC had a strong and sustained inhibitory ability to some common pathogens of bone and joint infections, such as staphylococcus aureus, escherichia coli and pseudomonas aeruginosa. In vivo drug-release study in animals demonstrated a high concentration of ciprofloxacine in the bone tissue surrounding CTDC which was placed in the greater trochanter of the rabbit and continued to release ciprofloxacine for at least 5 weeks and the blood level of ciprofloxacine was low. In vivo study also showed ultrasonic irradiation could increase the amount of ciprofloxacine released from CTDC, which may be an economical, effecient and safe new method to achieve the control of drug release from DDS.

Expression of TGF-β in Region of Bone Defect Repaired by Collagen/Nano-beta-Tricalcium Phosphate Composite Artificial Bone

The distribution and function of transforming growth factor-beta (TGF-β) in the region of bone defect repaired by collagen/nano-beta-tricalcium phosphate composite artificial bone (Co/N-TCP) and the ability of Co/N-TCP recruiting osteoblasts to precipitate the repair of bone defect were investigated Twenty-four domestic rabbits were operated on bilateral cranial bone to create an experimental bone defect of 8 0 mm in diameter through the whole bone On the left, Co/N-TCP was implanted as experimental group, but on the right, Co/TCP was implanted as control group At 2nd, 4th, 8th, 12th week after operation, all animals were sacrificed and the implanted materials with surrounding bone were taken out Immunohistochemical staining was performed for TGF-β assay by avidin-biotin complex method (SABC) Simultaneously, TGF-β was quantitatively analyzed by HPIAS-1000 imaging analysis system The inmmunohistochemical staining for TGF-β revealed that osteoblasts and immature osteocytes highly expressed TGF-β Diffused TGF-β positive staining particles appeared in the mesenchymal and fibrous-tissue There was no significant difference in the TGF-β positive staining between two groups in the medial region to original osseous beds at different time points ( P >0 05) However, in distal original osseous bed of the defected region, the positive expression of TGF-β in the Co/N-TCP group was significantly stronger than in the control group ( P <0 05 or 0 01) The Co/N-TCP has good bioactivities and ability of stimulating and conducting TGF-β to aggregate and precipitate the healing of bone defect

The Experimental Study on Mixed Culture of Osteoblasts and Tricalcium Phosphate Ceramics In Vitro

To study the effects of tricalcium phosphate (TCP) ceramics on osteoblasts, the rat osteoblasts were cultured with the TCP ceramics in vitro . Scanning electron microscopy and the colorimetric methyl thiazol tetrazolium assay showed that the osteoblasts could adhere well to the surface of the ceramics and the culture dish, and the proliferation of the cells was not inhibited. The results demonstrated that TCP ceramics possessed an excellent cytocompatibility with the osteoblasts, and had some promoting effects on proliferation of osteoblasts.

Effect on Hydration and Hardening of Tricalcium Phosphate Bone Cement

The bioactive α-Ca3（ PO4 ）2 bone cement was studied by XRD , SEM and isothermal calorimetric measurements. The results showed that a mixed pattern of TCP and hydroxylapatite were obtained after hydration and hardening. The mechanism of hydration and hardening of the α-Ca3 （ PO4）2 was dissolution-precipitation, （ NH4 ） 112 PO4 was the best set accelerator to the α-Ca3 （ PO4）2 cement, and the HAP powers and the （ NH4）H2 PO4 concentration had a great effect on the hydration rate of a-Ca3（ PO4）2.

Raman Spectroscopy Analysis of Wollastonite/Tricalcium Phosphate Glass-Ceramics after Implantation in Critical Bone Defect in Rats

In this work wollastonite/tricalcium phosphate (W/TCP) glass-ceramics with three W/TCP weight ratios (20/80;60/40 and 80/20) were implanted in rat calvaria and the modifications taking place during implantation were studied by Raman spectroscopy. The experimental glass-ceramics were composed of different contents of βW, αW, βTCP, αTCP, and glassy phases. Materials were implanted for 7-, 15-, 45- and 120-day periods after which the implanted materials were recovered and analyzed by FT-Raman spectroscopy. The results suggested that the αW phase reabsorbs fast during implantation in the glass-ceramics 60/40 and 80/20, whereas βTCP and αTCP glass-ceramic are gradually attenuated and replaced by biological apatite-like bands. In the glass-ceramic 20/80, the bands related to the βTCP phase remained unvaried in all analyzed periods. New bands associated with the deposition of collagenous material appeared during implantation for all 60/40 and 80/20 glass-ceramics experimental groups, but important differences in intensities between both groups. The spectra corresponding to implants of 60/40 glass-ceramic at the 120-day period were very similar to those of the control group (normal cortical bone), with regards to Raman shifts and intensities, as well as in the FWHM value of the 962 cm-1 apatite band (ν1 PO4 in hydroxyapatite), evidencing that apatite deposited at the implant site has the same crystallinity than biological apatite in normal bone mineral. The glass-ceramic 20/80 behaved just as an osteoconductive filling material, while glass-ceramics 60/40 and 80/20 were able to induce deposition of organic matrix mineralized new tissue. The 60/40 glass-ceramic showed the best performance and the most similar Raman spectrum to normal cortical bone.

Ionized jet deposition of antimicrobial and stem cell friendly silver-substituted tricalcium phosphate nanocoatings on titanium alloy

Orthopedic infections pose severe societal and economic burden and interfere with the capability of the implanted devices to integrate in the host bone,thus significantly increasing implants failure rate.To address infection and promote integration,here nanostructured antibacterial and bioactive thin films are proposed,obtained,for the first time,by Ionized Jet Deposition(IJD)of silver-substituted tricalcium phosphate(Ag-TCP)targets on titanium.Coatings morphology,composition and mechanical properties are characterized and proof-of-concept of biocompatibility is shown.Antimicrobial efficacy is investigated against four Gram positive and Gram negative bacterial strains and against C.albicans fungus,by investigating the modifications in planktonic bacterial growth in the absence and presence of silver.Then,for all bacterial strains,the capability of the film to inhibit bacterial adhesion is also tested.Results indicate that IJD permits a fine control over films composition and morphology and deposition of films with suitable mechanical properties.Biological studies show a good efficacy against Escherichia coli,Staphylococcus aureus,Pseudomonas aeruginosa,Enterococcus faecalis and against fungus Candida albicans,with evidences of efficacy against planktonic growth and significant reduction of bacterial cell adhesion.No cytotoxic effects are evidenced for equine adipose tissue derived mesenchymal stem cells(ADMSCs),as no reductions are caused to cells viability and no interference is assessed in cells differentiation towards osteogenic lineage,in the presence of silver.Instead,thanks to nanostructuration and biomimetic composition,tricalcium phosphate(TCP)coatings favor cells viability,also when silver-substituted.These findings show that silver-substituted nanostructured coatings are promising for orthopedic implant applications.

Biocompatibility of biphasic α,β-tricalcium phosphate ceramics in vitro

The biocompatibility of biphasicα,β-tricalcium phosphate ceramics,obtained by annealing a compact preform based onβ-tricalcium phosphate powder,was studied in vitro.It was found that within 10–30 days the adhesion of primary dental pulp stem cells located on the surface of biphasicα,β-tricalcium phosphate ceramics is suppressed.Decrease of the cell number on the surface of biphasicα,β-tricalcium phosphate ceramics,most likely,can be associated with both the pH level(acidic)as a result of hydrolysis of the more soluble phase ofα-tricalcium phosphate and with the nature of surface that changes as a result of the formation and growth of hydroxyapatite crystals.

Repair of the radial defect of rabbit with polyester/tricalcium phosphate scaffolds prepared by rapid prototyping technology

Objective： To evaluate the effects of repairing rabbit radial defects with polyester/tricalcium phosphate scaffolds prepared by rapid prototyping technology loaded with bovine bone morphogenetic protein （ bBMP）, and find new carriers for growth factors. Methods： Polyester/tricalcium phosphate scaffolds prepared by rapid prototyping technology loaded with and without bovine BMP were used to repair the 15 mm radial defect in rabbit. Then the results of radiography, histology, scaffolds degrade rates and bone mineral density （BMD） were appraised to examine the effects at the 12th week. Results ： At the 12th week postoperatively, all defects treated with bBMP were radiographically repaired. No radius implanted polyester/tricalcium phosphate scaffolds without bBMP showed radiographic and histological union. At experimental groups, longitudinal alignment of lamellar structure was observed histologically at the 12th week,indicating that remodeling of regenerated bone was complete in different degree. Of the three experimental groups, the bony regeneration and remodeling of callus in poly lactide-co-glycolide/tricalcium phosphate （PLGA/ TCP） group was the best. The BMD values were beyond 70% of normal value at the 12th week while the PLGA/ TCP scaffolds group was the highest, and no abnormalities were observed in the surrounding soft tissue in all groups. Conclusions - Polyester/tricalcium phosphate scaffolds prepared by rapid prototyping technology loaded with bovine BMP can repair a 15 mm radial defect of rabbit. As for the results, the PLGA/TCP scaffold is ideal and better than poly L-lactide-co-D, L-lactide （ PDLLA/TCP ） scaffold, but the ploy L-lactic acid （PLLA/TCP） is not so good for its low degradation rates.

Bioadaptable bioactive glass-β-tricalcium phosphate scaffolds with TPMS-gyroid structure by stereolithography for bone regeneration

Bone defect repair remains a troubling problem in clinical orthopedics,which involves complex biological processes.Calcium phosphates(CaPs)have been widely used owing to their advantage of biocompatibility.However,single component and traditional fabrication methods cannot meet the requirements of bioadaptability during the tissue repair process.In this work,0%,5%,15%,25%wt%of BG-TCP(bioactive glass-β-tricalcium phosphate)bioresorbable scaffolds with triply-periodic minimal surfaces(TPMS)-gyroid structure were prepared by the stereolithography(SLA)technology.TPMS-gyroid structure provided an accurate mimicry of natural bone tissue,and the incorporation of BG improved the compressive strength ofβ-TCP matrix,matched with the defective bone(2–12 MPa).Rapid but tunable degradation kinetics(compared with pure TCP)of BG enabled the BG-TCP system to sh8ow adaptable biodegradability to new bone generation.In vitro studies have shown that composite scaffolds have better mechanical properties(7.82 MPa),and can released appropriate contents of calcium,phosphorous,and magnesium ions,which promoted the osteogenic differentiation of bone marrow mesenchymal stem cells(BMSCs)and angiogenic ability of endothelial progenitor cells(EPCs).Moreover,the in vivo assessment of rat femoral defect revealed that TPMS-structure-based TCP scaffolds accelerated bone ingrowth to the pores.Moreover,BG-TCP scaffolds,especially 15BG-TCP group,exhibited superior bone regeneration capacity at both 4 and 8 weeks,which achieved an optimal match between the rate of material degradation and tissue regeneration.In summary,this study provides insight into influences of bioactive components(BG)and bionic structures(TPMS)on the physical-chemical properties of materials,cell behavior and tissue regeneration,which offers a promising strategy to design bioadaptive ceramic scaffolds in the clinical treatment of bone defects.

Efficacy of two calcium phosphate pastes on the remineralization of artificial caries: a randomized controlled double-blind in situ study

To test the efficacy of two calcium phosphate pastes compared to that of fluoride toothpaste on remineralizing artificial caries in situ, this study had a double-blind crossover in situ design, involving three experimental phases of 14 days each, with an 8-day washout period between phases. Nine healthy subjects participated in the study. The subjects wore removable palatal appliances mounted with six human enamel slabs with artificial caries lesions, and in each of the experimental phases, used one of the following methods two times/day： group A, brushing with 1.0 g of Colgate Regular Flavor, followed by applying 0.25 g of Tooth Mousse Plus; group B, brushing with 0.25 g of Clinpro Tooth Crbme; and group C, brushing with 1.0 g of Colgate Regular Flavor. After 14 days, the enamel slabs （54 slabs/ group） were embedded in resin, sectioned and examined with a polarized-light microscope, and the lesion areas were quantified using Image-Pro Plus. All experimental groups showed a significant reduction in lesion area compared to the initial lesion area （paired t-test, P〈O.O01）. The mean reduction in lesion area of Groups A, B and C were （0.029__.0.010）, （0.030_＋0.009） and （0.027_＋0.009） mm2, respectively. There were no statistical differences between groups （KruskaI-Wallis test, P〉0.05）. All three groups remineralized the enamel slab lesions, indicating model sensitivity to fluoride. Given the differences in usage amounts and treated regimens, Clinpro Tooth Crbme provided similar benefits to the fluoride toothpaste; however, no additional benefit of Tooth Mousse Plus was observed when used in conjunction with the fluoride toothpaste.

Three-dimensionally Perforated Calcium Phosphate Ceramics

Porous calcium phosphate ceramics were produced by compression molding using a special mold followed by sintering. The porous calcium phosphate ceramics have three-dimensional and penetrated open pores 380-400μm in diacneter spaced at intervals of 200μm. The layers of the linear penetration pores alternately lay perpendicular to pore direction. The porosity was 59%-65% . The Ca/ P molar ratios of the porous calcium phos phate ceramics range from 1.5 to 1.85. A binder cantaining methyl cellulose was most effective for preparing the powder compact among vinyl acetate, polyvinyl alcohol, starch, stearic acid, methyl cellulose and their mixtures . Stainless steel, polystyrene, nylon and bamboo were used as the long columnar dies for the penetrated open pores. When polystyrene, nylon and bamboo were used as the long columnar male dies, the dies were burned oat during the sintering process. Using stainless steel as the male dies with the removal of the dies before heat treatment resulted in a higher level of densification of the calcium phosphate ceramic.

Diversity of endophytic phosphate solubilising fungi associated with Pomatocalpa decipiens(Lindl.)J.J.Smith-an endangered orchid in Barbara forest of Odisha,India

Investigations were done to obtain potential phosphate solubilising strains from endophytic mycoflora isolated from the orchid,Pomatocalpa decipiens.928 endophytic phosphate solubilising fungal isolates were obtained from 2400 leaf segments(0.38%recovery)from rare epiphytic orchid Pomatocalpa decipiens present in the Barbara hills of Odisha(India).A number of isolates belonged to different genera such as Paecilomyces,Curvularia,Aspergillus,Cladosporium,Penicillium,Colletotrichum,while others,which were unidentified were classified as mycelia sterilia.Root sampling done from 25 different sites resulted in isolation of 20 endophytic phosphate solubilising fungal isolates from 300 segments(0.1%recovery).Aspergillus,Paecilomyces,Fusarium,Penicillium,and mycelia sterilia were mostly obtained.The qualitative and quantitative assessments of Phosphate(P)solubilisation were performed using TCP and Rock phosphate as P source for those strains.Aspergillus niger(leaf isolate)showed a maximum of 33.2 and 22.7%solubilisation in presence of TCP and Rock phosphate respectively whereas Aspergillus niger(Root isolate)showed a maximum of about 23.9%and 36.2%solubilisation in presence of TCP and Rock phosphate respectively.

Studies on the Ossification of Compound Transplantation of Porous TCP Ceramics and Bone Marrow

Following the culture of human red marrow with porous TCP ceramics in vitro, a large amount culturing marrow cells adhered well to the surface and inner wall of TCP ceramics and spread well over the dish bottom 4 weeks after cultivation. After the compound transplantation of porous TCP ceramics with red marrow into quadriceps femoris muscle of rabbits, a huge amount of new bone and marrow formed in these sites on and in the ceramics 4 weeks after transplantation. These results showed an excellent biocompatibility between TCP ceramics and marrow and an excellent osteogenic effect in the compound transplantation.The mechanism of ossification and clinical significance about the compound transplantation were discussed.

Reconstruction of Mandibular Defect with Porous β-TCP/BMG Artificial Bone

The purpose of this study is to explore the feasibility of repairing mandibular defect with osteoconductive porous beta-tricalcium phosphate （ β- TCP ） combined with osteoinductive bone matrix gelatin （ BMG ）, which has good physical property and biompatibility and maybe a new type of artificial bone substitute. Bilateral mandible defects sized 1.0 cm ×0. 6 cm×0. 3 cm of fifteen rabbits were implanted with β- TCP （ control group ） and β- TCPI BMG（ experimental group ） respectively. The specimens were harvested at 4^th , 8^th and 12^th weeks post-operation and examined by gross, X-ray, histomorphological and the computer analysis. The volume of newly-formed bone of the experimental side was much much than that of the control side at 4^th , 8^th and 12^th weeks. There is significant difference （p 〈 0.05） between experimental group and control group. Computer-aided X- ray analysis showed that the density of newly-formed bone, compared with the control side, were similar to the surrounding natural bone at 12^th week after operntion. Therefore, β- TCP/ BMG is a better bone substitute than TCP and is a degradable and biocompntible artificial bone substitute material and is osteoinductive and osteoconductive .

Phosphate Solubilizing Ability and Phylogenetic Diversity of Bacteria from P-Rich Soils Around Dianchi Lake Drainage Area of China

The phylogenetic diversity of phosphate solubilizing bacteria （PSB） distributed in P-rich soils in the Dianchi Lake drainage area of China was characterized, and the tricalcium phosphate （TCP） solubilizing activities of isolated PSB were determined. Among 1 328 bacteria isolated from 100 P-rich soil samples, 377 isolates （28.39% of the total） that exhibited TCP solubilization activity were taken as PSB. These PSB showed different abilities to solubilize TCP, with the concentrations of solubilized P in bacterial cultures varying from 33.48 to 69.63 mg L^（-1）. A total of 123 PSB isolates, with relatively high TCP solubilization activity （〉 54.00 mg L^（-1））, were submitted for restriction fragment length polymorphism （RFLP） analysis, which revealed 32 unique RFLP patterns. Based on these patterns, 62 representative isolates, one to three from each RFLP pattern, were selected for 16S rRNA sequencing. Phylogenetic analysis placed the 123 PSB into three bacterial phyla, namely Proteobacteria, Aetinobacteria and Firmicutes. Members of Proteobacteria were the dominant PSB, where 107 isolates represented by 26 RFLP patterns were associated with the genera of Burkholderia, Pseudomonas, Acinetobacter, Enterobacter, Pantoea, Serratia, Klebsiella, Leclercia, Raoultella and Cedeeea. Firmicutes were the subdominant group, in which 13 isolates were affiliated with the genera of Bacillus and Brevibacterium. The remaining 3 isolates were identified as three species of the genus Arthrobacter. This research extends the knowledge on PSB in P-rich soils and broadens the spectrum of PSB for the development of environmentally friendly biophosphate fertilizers.

Dental pulp stem cells and BonelikeVR for bone regeneration in ovine model

Development of synthetic bone substitutes has arisen as a major research interest in the need to find an alternative to autologous bone grafts.Using an ovine model,the present pre-clinical study presents a synthetic bone graft(BonelikeVR)in combination with a cellular system as an alternative for the regeneration of non-critical defects.The association of biomaterials and cell-based therapies is a promising strategy for bone tissue engineering.Mesenchymal stem cells(MSCs)from human dental pulp have demonstrated both in vitro and in vivo to interact with diverse biomaterial systems and promote mineral deposition,aiming at the reconstruction of osseous defects.Moreover,these cells can be found and isolated from many species.Non-critical bone defects were treated with BonelikeVR with or without MSCs obtained from the human dental pulp.Results showed that BonelikeVR and MSCs treated defects showed improved bone regeneration compared with the defects treated with BonelikeVR alone.Also,it was observed that the biomaterial matrix was reabsorbed and gradually replaced by new bone during the healing process.We therefore propose this combination as an efficient binomial strategy that promotes bone growth and vascularization in non-critical bone defects.