Dietary contribution of iron from limestone and dicalcium phosphate for broiler chickens.

Iron is routinely supplemented in broiler feeds aiming to prevent dietary deficiencies. Limestone and phosphates are very rich in Fe; however, its contribution from these sources have not been thoroughly investigated with chickens. The present research was conducted to evaluate live performance and blood parameters of broilers when using limestone and dicalcium phosphate as sources of Fe. A total of 576 one-day-old male Cobb x Cobb 500 were allocated into a total of 72 battery cages, 6 treatments with 12 replication cages of 8 chicks at placement. Chicks were fed diets formulated with corn, soybean meal (SBM) with laboratory grade calcium carbonate and phosphoric acid (having traces of Fe). All chicks were fed a common prestarter without Fe supplementation (analyzed total 58.2 ± 2.4 mg/kg Fe) from placement to 7 d. Allocation of birds to dietary treatments was completely randomized on day 8. Treatments had increasing Fe derived from commercial limestone and dicalcium phosphate (analyzed Fe 7,218 and 4,783 mg/kg, respectively) progressively replacing calcium carbonate and phosphoric acid to provide graded increases in total Fe (analyzed Fe in the feeds were 57.6 ± 2.1, 92.0 ± 2.3, 124.1 ± 2.7, 159.3 ± 3.1, 187.2 ± 3.2, 223.7 ± 3.6 mg/kg, respectively). There were no effects of dietary Fe on live performance, hematocrit, and hemoglobin the end of the study on day 28 (P > 0.05). Increasing dietary Fe from commercial limestone and dicalcium phosphate led to a linear reduction in the percent ileal digestible Fe. However, linear increments in Fe retention, serum ferritin and liver Fe occurred when compared to feeds without Fe derived from limestone and phosphate dicalcium. It is concluded that Fe from limestone and dicalcium phosphate can be partially utilized by broiler chickens. It was estimated that the Fe retained from limestone and dicalcium phosphate is of 1.9%. Broilers fed corn-soy feeds (58.2 mg/kg Fe) do not require supplemental Fe.

Sex-specific Stone-forming Phenotype in Mice During Hypercalciuria/Urine Alkalinization.

Sex differences in kidney stone formation are well known. Females generally have slightly acidic blood and higher urine pH when compared with males, which makes them more vulnerable to calcium stone formation, yet the mechanism is still unclear. We aimed to examine the role of sex in stone formation during hypercalciuria and urine alkalinization through acetazolamide and calcium gluconate supplementation, respectively, for 4 weeks in wild-type (WT) and moderately hypercalciuric [TRPC3 knockout [KO](-/-)] male and female mice. Our goal was to develop calcium phosphate (CaP) and CaP+ calcium oxalate mixed stones in our animal model to understand the underlying sex-based mechanism of calcium nephrolithiasis. Our results from the analyses of mice urine, serum, and kidney tissues show that female mice (WT and KO) produce more urinary CaP crystals, higher [Ca2+], and pH in urine compared to their male counterparts. We identified a sex-based relationship of stone-forming phenotypes (types of stones) in our mice model following urine alkalization/calcium supplementation, and our findings suggest that female mice are more susceptible to CaP stones under those conditions. Calcification and fibrotic and inflammatory markers were elevated in treated female mice compared with their male counterparts, and more so in TRPC3 KO mice compared with their WT counterparts. Together these findings contribute to a mechanistic understanding of sex-influenced CaP and mixed stone formation that can be used as a basis for determining the factors in sex-related clinical studies.

Amelogenin-inspired peptide, calcium phosphate solution, fluoride and their synergistic effect on enamel biomimetic remineralization: an in vitro pH-cycling model.

BACKGROUND: Several methods were introduced for enamel biomimetic remineralization that utilize a biomimetic analogue to interact and absorb bioavailable calcium and phosphate ions and induce crystal nucleation on demineralized enamel. Amelogenin is the most predominant enamel matrix protein that is involved in enamel biomineralization. It plays a major role in developing the enamel's hierarchical microstructure. Therefore, this study was conducted to evaluate the ability of an amelogenin-inspired peptide to promote the remineralization potential of fluoride and a supersaturated calcium phosphate solution in treating artificially induced enamel carious lesions under pH-cycling regimen. METHODS: Fifty enamel slices were prepared with a window (4*4 mm2 ) on the surface. Five samples were set as control healthy enamel and 45 samples were subjected to demineralization for 3 days. Another 5 samples were set as control demineralized enamel and 40 enamel samples were assigned into 8 experimental groups (n=5) (P/I, P/II, P/III, P/AS, NP/I, NP/II, NP/III and NP/AS) according to peptide treatment (peptide P or non-peptide NP) and remineralizing solution used (I; calcium phosphate solution, II; calcium phosphate fluoride solution, III; fluoride solution and AS; artificial saliva). Samples were then subjected to demineralization/remineralization cycles for 9 days. Samples in all experimental groups were evaluated using Raman spectroscopy for mineral content recovery percentage, microhardness and nanoindentation as healthy, demineralized enamel and after pH-cycling. Data were statistically analysed using two-way repeated measures Anova followed by Bonferroni-corrected post hoc test for pairwise multiple comparisons between groups. Statistical significance was set at p= 0.05. Additionally, XRD, FESEM and EDXS were used for crystal orientation, surface morphology and elemental analysis after pH-cycling. RESULTS: Nanocrystals clumped in a directional manner were detected in peptide-treated groups. P/II showed the highest significant mean values in mineral content recovery (63.31%), microhardness (268.81±6.52 VHN), elastic modulus (88.74±2.71 GPa), nanohardness (3.08±0.59 GPa) and the best crystal orientation with I002/I300 (1.87±0.08). CONCLUSION: Despite pH changes, the tested peptide was capable of remineralizing enamel with ordered crystals. Moreover, the supplementary use of calcium phosphate fluoride solution with peptide granted an enhancement in enamel mechanical properties after remineralization.

The effect of photobiomodulation therapy associated with casein phosphopeptide-amorphous calcium phosphate fluoride paste on the treatment of posthome whitening tooth sensitivity and color change: A randomized clinical trial.

OBJECTIVE: This study assessed whether combining photobiomodulation therapy (PBMT) with casein phosphopeptide-amorphous calcium phosphate fluoride (CPP-ACPF) paste can effectively reduce post-home whitening tooth sensitivity (TS) without compromising shade change. METHODS: Fifty participants were selected and assigned to one of four groups: (1) PLACEBO group-received a placebo paste and PBMT simulation; (2) PBMT group-received a placebo paste + PBMT; (3) CPP-ACPF group-received CPP-ACPF paste and PBMT simulation; (4) CPP-ACPF + PBMT group-received both CPP-ACPF paste and PBMT. The participants used whitening trays containing 22% carbamide peroxide for 2 h a day for 21 days. TS was measured daily using a visual analog scale, while shade change was assessed using a spectrophotometer: before bleaching treatment (T0), after the first (T1), second (T2), and third (T3) weeks of treatment, and 30 days (T4) after completing the whitening treatment. RESULTS: Intragroup analysis revealed that the PLACEBO group had the highest increase in sensitivity during the whitening treatment. The CPP-ACPF and PBMT groups showed no significant difference tooth whitening (TW) between weeks regarding aesthetic change. The CPP-ACPF and PBMT group exhibited a significant reduction in TS between the first and third and between the second and third weeks TW, but not between the first and second. Conversely, the PLACEBO group showed a higher sensitivity than the other groups (p < .05). The CPP-ACPF and PBMT groups did not differ from each other. Furthermore, the CPP-ACPF and PBMT group showed a greater decrease in sensitivity than the PLACEBO group at T1, T2, and T3 (p < .01), and was significantly differed from CPP-ACPF and PBMT groups only at T2 and T3. All groups confirmed TW effectiveness. Student's and paired t-test did not reveal any significant difference between groups (p > .05). CONCLUSION: Therefore, PBMT associated with CPP-ACPF paste can reduce TS without compromising the efficacy of TW.

Prevalencia de osteopenia del prematuro antes y después de la implementación de una estrategia temprana de manejo fosfocálcico / Prevalence of osteopenia of prematurity before and after implementing an early strategy with the use of calcium and phosphate

Introducción. Con el uso de la nutrición parenteral agresiva en recién nacidos de muy bajo peso, se detectaron alteraciones del metabolismo fosfocálcico. En 2016 se implementó una estrategia de prevención a través del monitoreo fosfocálcico y su suplementación temprana. El objetivo fue estudiar si esta estrategia disminuye la prevalencia de osteopenia e identificar factores de riesgo asociados. Población y métodos. Estudio cuasiexperimental que comparó la prevalencia de osteopenia entre dos grupos: uno después de implementar la estrategia de monitoreo y suplementación fosfocálcica (01/01/2017-31/12/2019), y otro previo a dicha intervención (01/01/2013-31/12/2015). Resultados. Se incluyeron 226 pacientes: 133 pertenecen al período preintervención y 93 al posintervención. La prevalencia de osteopenia global fue del 26,1 % (IC95% 20,5-32,3) y disminuyó del 29,3 % (IC95% 21,7-37,8) en el período preintervención al 21,5 % (IC95% 13,6-31,2) en el posintervención, sin significancia estadística (p = 0,19). En el análisis multivariado, el puntaje NEOCOSUR de riesgo de muerte al nacer, recibir corticoides posnatales y el período de intervención se asociaron de manera independiente a osteopenia. Haber nacido luego de la intervención disminuyó un 71 % la probabilidad de presentar fosfatasa alcalina >500 UI/L independientemente de las restantes variables incluidas en el modelo. Conclusión. La monitorización y suplementación fosfocálcica precoz constituye un factor protector para el desarrollo de osteopenia en recién nacidos con muy bajo peso al nacer.

Introduction. With the use of aggressive parenteral nutrition in very low birth weight infants, alterations in calcium and phosphate metabolism were detected. In 2016, a prevention strategy was implemented through calcium phosphate monitoring and early supplementation. Our objective was to study whether this strategy reduces the prevalence of osteopenia and to identify associated risk factors. Population and methods. Quasi-experiment comparing the prevalence of osteopenia between two groups: one after implementing the calcium phosphate monitoring and supplementation strategy (01/01/2017­12/31/2019) and another prior to such intervention (01/01/2013­12/31/2015). Results. A total of 226 patients were included: 133 in the pre-intervention period and 93 in the post-intervention period. The overall prevalence of osteopenia was 26.1% (95% CI: 20.5­32.3) and it was reduced from 29.3% (95% CI: 21.7­37.8) in the pre-intervention period to 21.5% (95% CI: 13.6­31.2) in the post-intervention period, with no statistical significance (p = 0.19). In the multivariate analysis, the NEOCOSUR score for risk of death at birth, use of postnatal corticosteroids, and the intervention period were independently associated with osteopenia. Being born after the intervention reduced the probability of alkaline phosphatase > 500 IU/L by 71%, regardless of the other variables included in the model. Conclusion. Calcium phosphate monitoring and early supplementation is a protective factor against the development of osteopenia in very low birth weight infants.

Influence of casein phosphopeptide-amorphous calcium phosphate on the surface topography and composition of nickel-titanium archwires during orthodontic treatment with fixed appliances.

PURPOSE: To investigate the surface topography and nickel content of nickel-titanium (NiTi) archwires exposed to either routine oral hygiene or a prophylactic regimen with casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) during orthodontic treatment. METHODS: This in vivo study involved 40 orthodontic patients with fixed appliances, who were randomly assigned to either a routine oral hygiene group or a CPP-ACP supplementary regimen group. Twenty new NiTi archwires served as controls. All archwires underwent scanning electron microscopy and energy-dispersive spectroscopy to evaluate their surface topography and elemental composition. The nickel content was quantified as a percentage of total weight and the Ni/Ti ratio, and statistical comparisons were made using pairwise tests. RESULTS: Wires exposed to fluoride toothpaste showed signs of pitting corrosion, deep grooves, and corrosion debris. In contrast, wires exposed to supplementary CPP-ACP exhibited smooth surface areas interspersed with microdefects and deposits. Statistically significant differences in nickel content were found between the new and retrieved archwires, as well as between wires exposed to routine oral hygiene and CPP-ACP (P < 0.001). The archwires exposed to CPP-ACP had the lowest nickel content (P < 0.001). CONCLUSION: The use of CPP-ACP holds promise for application as a safe anticariogenic agent with possible protective properties during orthodontic treatment.

Doxycycline-Loaded Calcium Phosphate Nanoparticles with a Pectin Coat Can Ameliorate Lipopolysaccharide-Induced Neuroinflammation Via Enhancing AMPK.

Neuroinflammation occurs in response to different injurious triggers to limit their hazardous effects. However, failure to stop this process can end in multiple neurological diseases. Doxycycline (DX) is a tetracycline, with potential antioxidant and anti-inflammatory properties. The current study tested the effects of free DX, DX-loaded calcium phosphate (DX@CaP), and pectin-coated DX@CaP (Pec/DX@CaP) nanoparticles on the lipopolysaccharide (LPS)-induced neuroinflammation in mice and to identify the role of adenosine monophosphate-activated protein kinase (AMPK) in this effect. The present study was conducted on 48 mice, divided into 6 groups, eight mice each. Group 1 (normal control), Group 2 (blank nanoparticles-treated), Group 3 (LPS (untreated)), Groups 4, 5, and 6 received LPS, then Group 4 received free DX, Group 5 received DX-loaded calcium phosphate nanoparticles (DX@CaP), and Group 6 received DX-loaded calcium phosphate nanoparticles with a pectin coat (Pec/DX@CaP). At the end of the experimentation period, behavioral tests were carried out. Then, mice were sacrificed, and brain tissue was extracted and used for histological examination, and assessment of interleukin-6 positive cells in different brain areas, in addition to biochemical measurement of SOD activity, TLR-4, AMPK and Nrf2. LPS can induce prominent neuroinflammation. Treatment with (Pec/DX@CaP) can reverse most behavioral, histopathological, and biochemical changes caused by LPS. The findings of the current study suggest that (Pec/DX@CaP) exerts a significant reverse of LPS-induced neuroinflammation by enhancing SOD activity, AMPK, and Nrf2 expression, in addition to suppression of TLR-4.

Reduced graphene oxide-reinforced tricalcium phosphate/gelatin/chitosan light-responsive scaffolds for application in bone regeneration.

Bone is a mineralized tissue with the intrinsic capacity for constant remodeling. Rapid prototyping techniques, using biomaterials that mimic the bone native matrix, have been used to develop osteoinductive and osteogenic personalized 3D structures, which can be further combined with drug delivery and phototherapy. Herein, a Fab@Home 3D Plotter printer was used to promote the layer-by-layer deposition of a composite mixture of gelatin, chitosan, tricalcium phosphate, and reduced graphene oxide (rGO). The phototherapeutic potential of the new NIR-responsive 3D_rGO scaffolds was assessed by comparing scaffolds with different rGO concentrations (1, 2, and 4 mg/mL). The data obtained show that the rGO incorporation confers to the scaffolds the capacity to interact with NIR light and induce a hyperthermy effect, with a maximum temperature increase of 16.7 °C after under NIR irradiation (10 min). Also, the increase in the rGO content improved the hydrophilicity and mechanical resistance of the scaffolds, particularly in the 3D_rGO4. Furthermore, the rGO could confer an NIR-triggered antibacterial effect to the 3D scaffolds, without compromising the osteoblasts' proliferation and viability. In general, the obtained data support the development of 3D_rGO for being applied as temporary scaffolds supporting the new bone tissue formation and avoiding the establishment of bacterial infections.

Prevalence of osteopenia of prematurity before and after implementing an early strategy with the use of calcium and phosphate. / Prevalencia de osteopenia del prematuro antes y después de la implementación de una estrategia temprana de manejo fosfocálcico.

Introduction. With the use of aggressive parenteral nutrition in very low birth weight infants, alterations in calcium and phosphate metabolism were detected. In 2016, a prevention strategy was implemented through calcium phosphate monitoring and early supplementation. Our objective was to study whether this strategy reduces the prevalence of osteopenia and to identify associated risk factors. Population and methods. Quasi-experiment comparing the prevalence of osteopenia between two groups: one after implementing the calcium phosphate monitoring and supplementation strategy (01/01/2017-12/31/2019) and another prior to such intervention (01/01/2013-12/31/2015). Results. A total of 226 patients were included: 133 in the pre-intervention period and 93 in the post-intervention period. The overall prevalence of osteopenia was 26.1% (95% CI: 20.5-32.3) and it was reduced from 29.3% (95% CI: 21.7-37.8) in the pre-intervention period to 21.5% (95% CI: 13.6-31.2) in the post-intervention period, with no statistical significance (p = 0.19). In the multivariate analysis, the NEOCOSUR score for risk of death at birth, use of postnatal corticosteroids, and the intervention period were independently associated with osteopenia. Being born after the intervention reduced the probability of alkaline phosphatase > 500 IU/L by 71%, regardless of the other variables included in the model. Conclusion. Calcium phosphate monitoring and early supplementation is a protective factor against the development of osteopenia in very low birth weight infants.

Introducción. Con el uso de la nutrición parenteral agresiva en recién nacidos de muy bajo peso, se detectaron alteraciones del metabolismo fosfocálcico. En 2016 se implementó una estrategia de prevención a través del monitoreo fosfocálcico y su suplementación temprana. El objetivo fue estudiar si esta estrategia disminuye la prevalencia de osteopenia e identificar factores de riesgo asociados. Población y métodos. Estudio cuasiexperimental que comparó la prevalencia de osteopenia entre dos grupos: uno después de implementar la estrategia de monitoreo y suplementación fosfocálcica (01/01/2017-31/12/2019), y otro previo a dicha intervención (01/01/2013-31/12/2015). Resultados. Se incluyeron 226 pacientes: 133 pertenecen al período preintervención y 93 al posintervención. La prevalencia de osteopenia global fue del 26,1 % (IC95% 20,5-32,3) y disminuyó del 29,3 % (IC95% 21,7-37,8) en el período preintervención al 21,5 % (IC95% 13,6-31,2) en el posintervención, sin significancia estadística (p = 0,19). En el análisis multivariado, el puntaje NEOCOSUR de riesgo de muerte al nacer, recibir corticoides posnatales y el período de intervención se asociaron de manera independiente a osteopenia. Haber nacido luego de la intervención disminuyó un 71 % la probabilidad de presentar fosfatasa alcalina >500 UI/L independientemente de las restantes variables incluidas en el modelo. Conclusión. La monitorización y suplementación fosfocálcica precoz constituye un factor protector para el desarrollo de osteopenia en recién nacidos con muy bajo peso al nacer.

The effect of supplementing the calcium phosphate cement containing poloxamer 407 on cellular activities.

Calcium phosphate cement (CPC) is generally used for bone repair and augmentation. Poloxamers are tri-block copolymers that are used as surfactants but have applications in drug and antibiotic delivery. However, their biological effects on bone regeneration systems remain unelucidated. Here, we aimed to understand how supplementing the prototype CPC with poloxamer would impact cellular activity and its function as a bone-grafting material. A novel CPC, modified beta-tricalcium phosphate (mß-TCP) powder, was developed through a planetary ball-milling process using a beta-tricalcium phosphate (ß-TCP). The mß-TCP dissolves rapidly and accelerates hydroxyapatite precipitation; successfully shortening the cement setting time and enhancing the strength. Furthermore, the addition of poloxamer 407 to mß-TCP could reduce the risk of leakage from bone defects and improve fracture toughness while maintaining mechanical properties. In this study, the poloxamer addition effects (0.05 and 0.1 g/mL) on the cellular activities of MC3T3-E1 cells cultured in vitro were investigated. The cell viability of mß-TCP containing poloxamer 407 was similar to that of mß-TCP. All specimens showed effective cell attachment and healthy polygonal extension of the cytoplasm firmly attached to hydroxyapatite (HA) crystals. Therefore, even with the addition of poloxamer to mß-TCP, it does not have a negative effect to osteoblast growth. These data demonstrated that the addition of poloxamer 407 to mß-TCP might be considered a potential therapeutic application for the repair and regeneration of bone defects.

Calcium phosphate nanoparticles conjugated with outer membrane vesicle of Riemerella anatipestifer for vaccine development in ducklings.

Biodegradable calcium phosphate nanoparticles offer a viable substitute for traditional adjuvants such as aluminum in vaccine production. Calcium phosphate nanoparticle adjuvanted with outer membrane vesicle (OMV) of gram negative bacteria may induce efficient immune response in the host. The present study was carried out to evaluate the potential of a mucosal vaccine formulation of calcium phosphate (CAP) nanoparticle using OMV of Riemerella anatipestifer (RA) as antigen against New Duck disease in ducks. The work was initiated with isolation, identification of RA, followed by OMV production and extraction. The CAP-OMV nanoparticle was prepared and characterized. The efficacy of the vaccine formulation and toxicity were studied in ducks. The average OMV yield in terms of protein concentration was found to be 122.33 ± 3.48 mg per liter of BHI broth. In SDS-PAGE, isolated OMVs exhibited presence of 16 distinct protein bands with molecular weight ranging from 142.1 to 12.1 kDa. Seven protein bands of 74.1, 69.3, 55.5, 50.6, 45.6, 25.1 and 13.1 kDa were detected relatively more distinct. The major bands detected in our findings were 42 kDa, 37 kDa and 16 kDa that corresponds to OmpA, OmpH, P6 respectively. The mean zeta size (±SD) and potential of the nanoparticle were 246.20 ± 0.53 nm and -25.60 ± 5.97 respectively. In transmission electron microscopy (TEM), the nanoparticles exhibited an average diameter of 129.80 ± 11.10 nm and displayed spherical morphology. The median protective dose (PD50) of CAP-OMV nanoparticle was 1881.10 µg of protein. Group I ducks received 3762 µg of protein (entrapped protein in CAP-OMV nanoparticle) via intra nasal route and it showed the highest serum IgG and secretory IgA level than other immunized groups. All experimental ducks were challenged with 10 × LD50 on 35 days of post primary immunization. Group I showed 100 % survivability in the challenge study. No gross and biochemical indication of acute or chronic toxicity were recorded. In conclusion, our results suggest that CAP-OMV nanoparticle can be a safe and efficient mucosal vaccine delivery system for RA, eliciting strong immune response in the host.

Leaching of phosphorus from phosphate tailings and extraction of calcium phosphates: Toward comprehensive utilization of tailing resources.

Phosphate tailing is an extremely fine by-product during phosphate ore flotation. Due to the large quantities and relatively higher P2O5 content, the phosphate tailings have been considered as a potential P resource, compared to other P-bearing wastes. Besides, phosphate tailings also contain a large amount of available components, such as Ca, Mg, and Si. To explore a low-cost and efficient process for the utilization of phosphate tailings, the hydrochloric acid leaching-precipitation method was employed to recover phosphorus. The P in phosphate tailings can be selectively dissolved into leaching liquor, followed by the precipitation of calcium phosphates from the leaching liquor through pH adjustment. The results showed that P was predominantly concentrated in fluorapatite and its dissolution ratio increased with the decrease in pH. At pH 1.0, the P dissolution efficiency from phosphate tailings reached 96.3%, along with the majority of Mg and Ca. However, Si was hardly dissolved. It demonstrated that almost all the fluorapatite and dolomite were dissolved while the quartz was difficult to dissolve. Dolomite was more preferentially dissolved than fluorapatite. Increasing temperature contributed to the dissolution of dolomite while suppressing fluorapatite dissolution. The residue containing 87.9% SiO2 (quartz) and only 0.25% P2O5 has the potential as a building material. As the pH increased to 7.0, the collected precipitate consisted of 34.18% P2O5 and 56.10% CaO, which can serve as a source of a slow-released phosphate fertilizer. The highly efficient utilization of phosphate tailings was achieved via this process.

Long-Term Safety of Bone Regeneration Using Autologous Stromal Vascular Fraction and Calcium Phosphate Ceramics: A 10-Year Prospective Cohort Study.

This prospective cohort study aimed to assess long-term safety, dental implant survival, and clinical and radiological outcomes after maxillary sinus floor elevation (MSFE; lateral window technique) using freshly isolated autologous stromal vascular fraction (SVF) combined with calcium phosphate ceramics. All 10 patients previously participating in a phase I trial were included in a 10-year follow-up. They received either ß-tricalcium phosphate (ß-TCP; n = 5) or biphasic calcium phosphate (BCP; n = 5) with SVF-supplementation on one side (study). Bilaterally treated patients (6 of 10; 3 ß-TCP, 3 BCP) received only calcium phosphate on the opposite side (control). Clinical and radiological assessments were performed on 44 dental implants at 1-month pre-MSFE, and 0.5- to 10-year post-MSFE. Implants were placed 6 months post-MSFE. No adverse events or pathology was reported during a 10-year follow-up. Forty-three dental implants (98%) remained functional. Control and study sides showed similar peri-implant soft-tissue quality, sulcus bleeding index, probing depth, plaque index, keratinized mucosa width, as well as marginal bone loss (0-6 mm), graft height loss (0-6 mm), and graft volume reduction. Peri-implantitis was observed around 6 implants (control: 4; study: 2) in 3 patients. This study is the first to demonstrate the 10-year safety of SVF-supplementation in MSFE for jawbone reconstruction. SVF-supplementation showed enhanced bone regeneration in the short term (previous study) and led to no abnormalities clinically and radiologically in the long term.

In situ mapping of biomineral skeletal proteins by molecular recognition imaging with antibody-functionalized AFM tips.

Spatial localizing of skeletal proteins in biogenic minerals remains a challenge in biomineralization research. To address this goal, we developed a novel in situ mapping technique based on molecular recognition measurements via atomic force microscopy (AFM), which requires three steps: (1) the development and purification of a polyclonal antibody elicited against the target protein, (2) its covalent coupling to a silicon nitride AFM tip ('functionalization'), and (3) scanning of an appropriately prepared biomineral surface. We applied this approach to a soluble shell protein - accripin11 - recently identified as a major component of the calcitic prisms of the fan mussel Pinna nobilis [1]. Multiple tests reveal that accripin11 is evenly distributed at the surface of the prisms and also present in the organic sheaths surrounding the calcitic prisms, indicating that this protein is both intra- and inter-crystalline. We observed that the adhesion force in transverse sections is about twice higher than in longitudinal sections, suggesting that accripin11 may exhibit preferred orientation in the biomineral. To our knowledge, this is the first time that a protein is localized by molecular recognition atomic force microscopy with antibody-functionalized tips in a biogenic mineral. The 'pros' and 'cons' of this methodology are discussed in comparison with more 'classical' approaches like immunogold. This technique, which leaves the surface to analyze clean, might prove useful for clinical tests on non-pathological (bone, teeth) or pathological (kidney stone) biomineralizations. Studies using implants with protein-doped calcium phosphate coating can also benefit from this technology. STATEMENT OF SIGNIFICANCE: Our paper deals with an unconventional technical approach for localizing proteins that are occluded in biominerals. This technique relies on the use of molecular recognition atomic force microscopy with antibody-functionalized tips. Although such approach has been employed in other system, this is the very first time that it is developed for biominerals. In comparison to more classical approaches (such as immunogold), AFM microscopy with antibody-functionalized tips allows higher magnification and keeps the scanned surface clean for other biophysical characterizations. Our method has a general scope as it can be applied in human health, for non-pathological (bone, teeth) and pathological (kidney stone) biomineralizations as well as for bone implants coated with protein-doped calcium phosphate.

An insight into the role of the organic acids produced by Enterobacter sp. strain 15S in solubilizing tricalcium phosphate: in situ study on cucumber.

BACKGROUND: The release of organic acids (OAs) is considered the main mechanism used by phosphate-solubilizing bacteria (PSB) to dissolve inorganic phosphate in soil. Nevertheless, little is known about the effect of individual OAs produced by a particular PSB in a soil-plant system. For these reasons, the present work aimed at investigating the effect of Enterobacter sp. strain 15S and the exogenous application of its OAs on (i) the solubilization of tricalcium phosphate (TCP), (ii) plant growth and (iii) P nutrition of cucumber. To this purpose two independent experiments have been performed. RESULTS: In the first experiment, carried out in vitro, the phosphate solubilizing activity of Enterobacter 15S was associated with the release of citric, fumaric, ketoglutaric, malic, and oxalic acids. In the second experiment, cucumber plants were grown in a Leonard jar system consisting of a nutrient solution supplemented with the OAs previously identified in Enterobacter 15S (jar's base) and a substrate supplemented with the insoluble TCP where cucumber plants were grown (jar's top). The use of Enterobacter 15S and its secreted OAs proved to be efficient in the in situ TCP solubilization. In particular, the enhancement of the morpho-physiological traits of P-starved cucumber plants was evident when treated with Enterobacter 15S, oxalate, or citrate. The highest accumulation of P in roots and shoots induced by such treatments further corroborated this hypothesis. CONCLUSION: In our study, the results presented suggest that organic acids released by Enterobacter 15S as well as the bacterium itself can enhance the P-acquisition by cucumber plants.

Eco-friendly soil stabilization method using fish bone as cement material.

The method of soil improvement by calcium phosphate precipitation is a novel, environmentally friendly, and non-toxic technique. Such technology provides advantages over ureolytic induced calcite precipitation (UICP), the most popular and widely used method in the field of geotechnical engineering. In this paper, an investigation of the consolidation of fine and coarse sand samples by enzyme induced calcium phosphate precipitation (EICPP) was carried out. Tuna bones were used as an alternative source of calcium and phosphorus ions, as one of the most popular fish species in Japan and the main source of food industry waste. Unconfined compressive strength (UCS) of the samples after 21 days of daily injection of the solution showed an increase in strength up to 6,05 MPa in fine and up to 4,3 MPa in coarse sand samples. X-ray powder diffraction (XRD), scanning electron microscope (SEM), and energy dispersive X-ray spectroscopy (SEM-EDS) analysis were performed to investigate the nature and type of deposition. Analyses confirmed that deposition is composed of brushite with needle-like crystals in the case of Toyoura sand and flower-like crystals in the case of Mikawa sand. SEM-EDS showed a presence of both, calcium, and phosphorus in the precipitate, indicating the presence of calcium phosphate compounds (CPCs). This study reveals that tuna bones are a rich source of calcium and phosphorus for EICPP, which results in a strengthening of silicate soil up to 3.4-6.05 MPa and is able to reduce ammonia emissions by 85.7 % - 97.5 % compared to UICP.

Prediction of calcium phosphate generation and behaviors of metals during phosphorus recovery using a modified thermodynamic model.

In this study, behaviors of metals and their effects on phosphorus recovery by calcium phosphate were investigated by the laboratory and pilot experiments as well as by the modified thermodynamic model. Batch experimental results indicated that the efficiency of phosphorus recovery decreased with the increase in metal content and more than 80% phosphorus can be recovered with a Ca/P molar ratio of 3.0 and a pH of 9.0 for the supernatant of an anaerobic tank in the A/O process with the influent containing a high metal level. The mixture of amorphous calcium phosphate (ACP) and dicalcium phosphate dihydrate (DCPD) was assumed to be the precipitated product with an experimental time of 30 min. A modified thermodynamic model was developed using ACP and DCPD as the precipitated products, and the correction equations were incorporated to simulate the short-term precipitation of calcium phosphate based on the experimental results. From the perspective of maximizing both the efficiency of phosphorus recovery and the quality or purity of the recovered product, the simulation results showed that a pH of 9.0 and a Ca/P molar ratio of 3.0 were the optimized operational condition for phosphorus recovery by calcium phosphate when the influent metal content was at the level of actual municipal sewage.

BMP-2 (and partially GDF-5) coating significantly accelerates and augments bone formation close to hydroxyapatite/tricalcium-phosphate/brushite implant cylinders for tibial bone defects in senile, osteopenic sheep.

Bilateral defects (diameter 8 mm) in the medial tibial head of senile, osteopenic female sheep (n = 48; 9.63 ± 0.10 years; mean ± SEM) were treated with hydroxyapatite (HA)/beta-tricalcium phosphate (ß-TCP)/dicalcium phosphate dihydrate (DCPD; brushite) cylinders coated with BMP-2 (25 or 250 micrograms) or growth differentiation factor (GDF)-5 (125 or 1250 micrograms; left side); cylinders without BMP served as controls (right side). Three, 6, and 9 months post-operation (n = 6 each group), bone structure and formation were analyzed in vivo by X-ray and ex vivo by osteodensitometry, histomorphometry, and micro-computed tomography (micro-CT) at 3 and 9 months. Semi-quantitative X-ray evaluation showed significantly increasing bone densities around all implant cylinders over time. High-dose BMP-2-coated cylinders (3 and 9 months) and low-dose GDF-5-coated cylinders (3 and 6 months) demonstrated significantly higher densities than controls (dose-dependent for BMP-2 at 3 months). This was confirmed by osteodensitometry at 9 months for high-dose BMP-2-coated cylinders (and selected GDF-5 groups), and was again dose-dependent for BMP-2. Osteoinduction by BMP-2 was most pronounced in the adjacent bone marrow (dynamic histomorphometry/micro-CT). BMP-2 (and partially GDF-5) significantly increased the bone formation in the vicinity of HA/TCP/DCPD cylinders used to fill tibial bone defects in senile osteopenic sheep and may be suitable for surgical therapy of critical size, non-load-bearing bone defects in cases of failed tibial head fracture or defect healing.

Physicochemical properties and cytocompatibility of radiation-resistant and anti-washout calcium phosphate cement by introducing artemisia sphaerocephala krasch gum.

The anti-washout ability of calcium phosphate cement (CPC) determines the effectiveness of CPC in clinical application. The γ-ray irradiation method often used in the sterilization process of CPC products is easy to degrade some commonly polymer anti-washout agent, which greatly reduces its anti-washout performance. Artemisia sphaerocephala Krasch gum (ASKG) has the potential of radiation resistance and anti-washout, but no one has considered its performance as anti-washout agent of CPC and mechanism of radiation resistance and anti-washout so far. In this study, we report the effect of γ-ray on ASKG and the effectiveness of ASKG for enhancing of radiation resistance and anti-washout ability of CPC, the physical, chemical properties and in vitro cell behaviors of ASKG-CPCs were also investigated. The results showed that addition of ASKG before and after irradiation could significantly enhanced the anti-washout performance of CPC, which is differ from conventional anti-washout agents. Meanwhile, ASKG-CPCs had an excellent injectable property and biocompatibility, and low content of irradiated ASKG could promote bone differentiation well. We anticipate that the radiation-resistant and anti-washout ASKG-CPCs have potential application prospect in orthopaedic surgery.

FePSe3 -Nanosheets-Integrated Cryogenic-3D-Printed Multifunctional Calcium Phosphate Scaffolds for Synergistic Therapy of Osteosarcoma.

Clinical treatment of osteosarcoma encounters great challenges of postsurgical tumor recurrence and extensive bone defect. To develop an advanced artificial bone substitute that can achieve synergistic bone regeneration and tumor therapy for osteosarcoma treatment, a multifunctional calcium phosphate composite enabled by incorporation of bioactive FePSe3 -nanosheets within the cryogenic-3D-printed α-tricalcium phosphate scaffold (TCP-FePSe3 ) is explored. The TCP-FePSe3 scaffold exhibits remarkable tumor ablation ability due to the excellent NIR-II (1064 nm) photothermal property of FePSe3 -nanosheets. Moreover, the biodegradable TCP-FePSe3 scaffold can release selenium element to suppress tumor recurrence by activating of the caspase-dependent apoptosis pathway. In a subcutaneous tumor model, it is demonstrated that tumors can be efficiently eradicated via the combination treatment with local photothermal ablation and the antitumor effect of selenium element. Meanwhile, in a rat calvarial bone defect model, the superior angiogenesis and osteogenesis induced by TCP-FePSe3 scaffold have been observed in vivo. The TCP-FePSe3 scaffold possesses improved capability to promote the repair of bone defects via vascularized bone regeneration, which is induced by the bioactive ions of Fe, Ca, and P released during the biodegradation of the implanted scaffolds. The TCP-FePSe3 composite scaffolds fabricated by cryogenic-3D-printing illustrate a distinctive strategy to construct multifunctional platform for osteosarcoma treatment.