Bioceramics 24

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Authors: Toshimitsu Tanaka, Tomohiko Yoshioka, Toshiyuki Ikoma, Tomoya Kuwayama, Tatsuhiko Higaki, M. Tanaka
Abstract: The purposes of this study were to fabricate porous carbonate-substituted apatite (CAp) ceramics and characterize important properties such as compressive strengths and dissolution rates for an artificial bone. Porous A-type CAp (A-CAp) with uniaxial pores was fabricated by sintering porous hydroxyapatite green bodies at 1000°C in carbon dioxide gas atmosphere. B-type CAp (B-CAp) nanocrystals prepared by a wet method were mixed with an organic binder, freeze-dried and heated at 800°C in air, and AB-type CAp (AB-CAp) was fabricated by heating B-CAp porous bodies including the organic binder at 1100°C in carbon dioxide gas atmosphere. From the morphological observations with a scanning electron microscope, the porous A-CAp had uniaxially oriented pores of 104±33 μm in diameter, while the porous B-CAp and AB-CAp had large pores of 143±48 μm and 181±46 μm in diameter, of which pores were interconnected with small pores of 31±11 μm and 45±17 μm in diameter. The dissolution rates of CAps were apparently larger than that of HAp; the calcium concentrations increased in the order of AB-CAp > B-CAp > A-CAp > HAp. This is mainly attributed to carbonate content although it could be partialy depended on the different porous structures and diameters.
Authors: Sajad Farhangdoust, Sayed Mahmood Rabiee, Ali Zamanian, Mana Yasaei, Mina Khorami, Masoud Hafezi-Ardakani
Abstract: Scaffolds have to meet exacting physical, chemical, and biological criteria to function successfully, and those criteria vary with the type of tissue being repaired. In the present work, slurry with different initial content of 7.5-22.5 vol% HA prepared from calcinated hydroxyapatite. The prepared slurries freeze casted unidirectionally with the different cooling rate of 2-14°C/min with intervals of 3°C/min from the ambient temperature. Then, green bodies freeze-dried for 72h following with sintering at temperatures of 1350°C. The results showed that compressive strength goes up with cooling rate and initial content. Total porosity has a range of 66-88% while has a compressive strength of ~0.4-18 MPa. Porosity size has a value of 20-200 μm by initial content and cooling rate. Based on strength and porosity, the specimen with initial content and cooling rate of 15 vol% and 5°C/min, respectively, chose to be the optimum. This specimen has a compressive strength and porosity size of 5.26 MPa and 88 μm, respectively. The compressive strength value of the mentioned lamellar HA scaffolds was in the range of the values reported for human proximal tibia.
Authors: Tram Nguyen Xuan Thanh, Michito Maruta, Kanji Tsuru, Alireza Valanezhad, Shigeki Matsuya, Ishikawa Kunio
Abstract: We have previously reported that calcite foam that had interconnected porous structure could be prepared by ceramic foam method and it transformed to carbonate apatite (CO3Ap). In the ceramic foam method, polyurethane sponge was used as a template. The polyurethane sponge was immersed in the ceramics slurry, and the strut of the polyurethane foam was covered by ceramic powder. After that it was dried and sintered at high temperature. Calcite foams produced by this approach were comprised of a three-dimensional (3D) interconnected porous structure that facilitated cell penetration. However, all foams have a common limitation: the inherent lack of mechanical strength associated with high porosity. Therefore, in this study, an inverse ceramic foam method was studied; multi polyurethane coating method using polyurethane foam as a template. In this study, the compressive strength was improved by an inverse replication allowed for decreasing porosity while at the same time maintaining the interconnectivity. The burnable synthetic resin coating layer was introduced onto struts of polyurethane foam to make the triangular struts become more round and thick, consequently producing large round capillary within the foam structure fulfilling the requirement for osteoblast colonization. In particular, polyurethane foam was dipped orderly into two monomers, followed by centrifugation to remove excess liquids inside foam. After resin curing, a layer of synthetic resin was coated strut of foam. Calcium hydroxide Ca (OH)2 slurry was then infiltrated into resin coated-polyurethane foam. By firing at 600°C in O2-CO2 stream, polyurethane template was burnt off and Ca (OH)2 was converted into calcite. Negative replicated calcite foam was fabricated and characterized micro-structurally with interconnectivity and improved mechanical strength. The results obtained in this study suggested that this method dramatically improved the mechanical strength of the calcite foam without sacrificing the interconnected structure, and this means that the calcite foam obtained in this method could be precursors for the 3D interconnected porous CO3Ap foam.
Authors: Kemal Sariibrahimoglu, Joop G.C. Wolke, Sander C.G. Leeuwenburgh, John A. Jansen
Abstract: Calcium phosphate cements (CPCs) can be a suitable scaffold material for bone tissue engineering because of their osteoconductivity and perfect fit with the surrounding tissue when injected in situ. However, the main disadvantage of hydroxyapatite (HA) forming CPC is its slow degradation rate, which hinders complete bone regeneration. A new approach is to use hydraulic apatite cement with mainly α/β-tricalciumphosphate (TCP) instead of α-TCP. After hydrolysis the α/β-TCP transforms in a partially non-absorbable HA and a completely resorbable β-TCP phase. Therefore, α-TCP material was thermally treated at several temperatures and times resulting in different α/β-TCP ratios. In this experiment, we developed and evaluated injectable biphasic calcium phosphate cements (BCPC) in vitro. Biphasic α/β-TCP powder was produced by heating α-TCP ranging from 1000-11250°C. Setting time and compressive strength of the CPCs were analyzed after soaking in PBS for 6 weeks. Results demonstrated that the phase composition can be controlled by the sintering temperature. Heat treatment of α-TCP, resulted in 100%, 75% and 25% of α-to β-TCP transformation, respectively. Incorporation of these sintered BCP powder into the cement formulation increased the setting time of the CPC paste. Compressive strength decreased with increasing β-TCP content. In this study, biphasic CPCs were produced and characterized in vitro. This injectable biphasic CPC presented comparable properties to an apatitic CPC.
Authors: Toshiisa Konishi, Minori Mizumoto, Michiyo Honda, Mamoru Aizawa
Abstract: We have previously developed hydroxyapatite (HAp) cement based on the chelate-setting mechanism of sodium inositol hexaphosphate (IP6), in which HAp powder was prepared by surface-modification with IP6 after ball-milling of the HAp powder (conventional process). Meanwhile, we have recently established novel powder preparation process (modified process). In the present study, the adsorption behavior of IP6 on the surface of HAp at both the processes was circumstantially examined to clarify the chelating mechanism of IP6. The adsorbed amount of IP6 increased with the IP6 concentration in both the processes; however, the adsorbed amount of IP6 at the modified process was lower than that at the conventional process. X-ray photoelectron spectroscopic study revealed that the IP6 adsorbed on the surface of HAp powders. The degree in dispersion of the HAp particles at the modified process was higher than that at conventional process. Furthermore, the elution of IP6 from the powders prepared at the novel process was lower than that of the powders at the conventional process.
Authors: Toshiisa Konishi, Shuhei Takahashi, Minori Mizumoto, Michiyo Honda, Koki Kida, Yukiko Horiguchi, Kazuya Oribe, Ken Ishii, Hikaru Morisue, Yoshiaki Toyama, Morio Matsumoto, Mamoru Aizawa
Abstract: We have developed novel calcium-phosphate cements (CPCs) based on the chelate-setting mechanism of inositol phosphate (IP6) using hydroxyapatite (HAp), β-tricalcium phosphate (β-TCP) and α-TCP as starting materials. These cements (IP6-HAp, IP6-β-TCP and IP6-α-TCP cements) have different bioresorbability due to the chemical composition of starting materials. In the present study, biocompatibility and bioresorbability of the above three cements and commercially available cement (Biopex®-R) was histologically evaluated in vivo using rabbit model for 4, 8, and 24 weeks, in addition to their dissolution in vitro. The dissolution of these cements increased in the order of IP6-HAp, IP6-β-TCP and IP6-α-TCP cements. The newly-formed bones were directly in contact with both the IP6-HAp and Biopex®-R cement specimens. As for the IP6-β-TCP and IP6-α-TCP cements, newly-formed bones were formed time-dependently slightly apart from the cement specimens. Resorption rate for Biopex®-R, IP6-HAp, IP6-β-TCP, and IP6-α-TCP cements after 24 weeks implantation were of 7.2, 5.0, 13.7, and 16.2%, respectively, compared to original cements. The present chelate-setting cements with different bioresorbability are promising candidates for application as the novel CPCs.
Authors: Michiyo Honda, Toshiisa Konishi, Minori Mizumoto, Mamoru Aizawa
Abstract: Cancers frequently metastasize to bone, where it leads to secondary tumor growth, and osteolytic bone degradation. Bone metastases are often associated with fractures and severe pain resulting in decreased quality of life. Accordingly, effective therapies to inhibit the development or progression of bone metastases will have important clinical benefits. Bone cement, one of the powerful tools as bone substitutes, is used to fill the resection voids. The aim of this study was to develop a local drug delivery system using HAp cement as a carrier of chemotherapeutic agents. In the present study, we have fabricated chelate-setting apatite cements (IP6-HAp cements) using HAp particles surface-modified with inositol hexaphosphate (IP6) and evaluated their anti-tumor effect. Human osteosarcoma (HOS) cultured on IP6-HAp cements (over 3000 ppm IP6) resulted in inhibition of cell growth. DNA microarray analysis indicated changes in the expression of apoptosis-related genes on IP6-HAp cement surface-modified with 5000 ppm IP6 compared with HAp cement, suggesting activation of apoptosis machinery by IP6-HAp cement. To clarify the mechanism of anti-tumor effect of IP6-HAp cement, the properties of cement were investigated. The release kinetics of IP6 from IP6-HAp cement showed that the level of released IP6 was insufficient to induce anti-tumor activity. These results led us to consider that locally high concentration of IP6 which was released from cement acts on the cells directly as anti-tumor agent and induces the apoptosis. Consequently, IP6-HAp cement might gain the anti-tumor effect and act as a carrier for local drug delivery system.
Authors: Tetsuya Inayama, Hidenori Konishi, Mamoru Aizawa, Nobuyuki Kanzawa
Abstract: Chelate-setting apatite cement is a novel biomaterial developed as a bone substitute. We previously reported a chelate-setting apatite cement, IP6-HAp, which exhibits anti-tumor activity via apoptotic cell death. However, our preliminary data showed that excess IP6 arrests osteoblast growth. We found that a high, transient amount of IP6 was released from the cement. We therefore hypothesized that a high performance cement specific for tumor cells can be developed by controlling the release of IP6 from the cement. To validate this, we used a murine calvarial osteoblast cell line (MC3T3-E1) and a human osteosarcoma cell line (HOS). Culturing HOS or MC3T3-E1 in medium containing various concentrations of IP6 more effectively arrested the growth of HOS than that of MC3T3-E1. Although the proliferation of osteoblasts was suppressed at early growth stages in response to the release of IP6 from the cements, there was no difference in the number of cells after a prolonged culture period. In contrast, osteosarcoma cell growth remained suppressed even after a prolonged culture period. To better understand why these two cell types respond differently to IP6, we investigated cell viability by measuring the ratio of living and dead cells. Our findings suggest that this novel bone graft cement will find unique uses due the different sensitivity of tumor cells and osteoblasts towards IP6.
Authors: Yusuke Nakashima, Michiyo Honda, Toshiisa Konishi, Minori Mizumoto, Mamoru Aizawa
Abstract: In our previous study, silicon-containing hydroxyapatite (Si-HAp) powder was prepared via an aqueous precipitation reaction. The Si-HAp powders were synthesized with desired Si contents (0, 0.4, 0.8, 1.6, and 2.4 mass%) as a nominal composition. Another previous study in our group demonstrated surface-modification of HAp powder with inositol phosphate (IP6) enhanced the compressive strength of apatite cement. Thus, to fabricate the cements with higher bioactivity, the above Si-HAp powders were surface-modified with IP6 (IP6-Si-HAp). The IP6-Si-HAp cements with various Si contents were fabricated by mixing with pure water at the powder/liquid ratio of 1/0.4 [w/v]. In order to clarify biocompatibility of the IP6-Si-HAP cements in the present work, MC3T3-E1 cells as a model of osteoblast were seeded on the cement specimens. As for the numbers of cells cultured on the IP6-Si-HAp cements, the substitution of lower levels of Si into HAp lattice did not greatly influence the cell proliferation. However, the substitution of Si amount over 0.8 mass% enhanced the cell proliferation. Especially, the IP6-Si-HAp cement with the Si content of 2.4 mass% showed excellent cell proliferation among examined specimens. Therefore, to fabricate the cements with higher bioactivity, it is necessary to control the amount of Si in the IP6-Si-HAp cements. The usage of these IP6-Si-HAp cements may make it possible to fabricate the cements with higher bioactivity, compare to conventional pure HAp cements.
Authors: Hitomi Ohsugi, Yuri Habuto, Michiyo Honda, Mamoru Aizawa, Nobuyuki Kanzawa
Abstract: We have developed a chelate-setting apatite cement. Synthesized hydroxyapatite (HAp) powders surface-modified with inositol hexaphosphate (IP6-HAp powder) were set by chelate-bonding with inositol hexaphosphate (IP6). Our aim is to fabricate IP6-HAp cement with anti-bacterial activity by adding lactoferrin (LF). It is known that LF has both anti-bacterial and osteoinductive activity. Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli were used to examine the effect of LF on biofilm formation and localization of living and dead cells. In addition, the cell viability of MC3T3-E1 osteoblastic cells was determined. Our results show that the anti-bacterial activity of LF is not due to a bactericidal effect but to the inhibition of bacterial adhesion to surfaces. Furthermore, LF cement did not affect cell proliferation. Thus, LF cement is a candidate for bifunctional biomaterials having both anti-bacterial and osteo-conductive activity.

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