Phagocytosis of wear debris by osteoblasts affects differentiation and local factor production in a manner dependent on particle composition

Wear debris is considered to be one of the main factors responsible for aseptic loosening of orthopaedic endoprostheses. Whereas the response of cells in the monocytic lineage to foreign materials has been extensively studied, little is known about cells at the bone formation site. In the present study, we examined the hypothesis that the response of osteoblasts to wear debris depends on the chemical composition of the particles. We produced particles from commercially pure titanium (cpTi), Ti-6Al-4V (Ti-A), and cobalt-chrome (CoCr) and obtained ultrahigh molecular weight polyethylene (UHMWPE; GUR 4150) particles from a commercial source. The equivalent circle diameters of the particles were comparable: 1.0 +/- 0.96 microm for UHMWPE; 0.84 +/- 0.12 microm for cpTi; 1.35 +/- 0.09 microm for Ti-A, and 1.21 +/- 0.16 microm for CoCr. Confluent primary human osteoblasts and MG63 osteoblast-like cells were incubated in the presence of particles for 24 h. Harvested cultures were examined by transmission electron microscopy to determine if the cells had phagocytosed the particles. Particles were found intracellularly, primarily in the cytosol, in both the primary osteoblasts and MG63 cells. The chemical composition of the particles inside the cells was confirmed by energy-dispersive X-ray analysis. Morphologically, both cell types had extensive ruffled cell membranes, less-developed endoplasmic reticulum, swollen mitochondria, and vacuolic inclusions compared with untreated cells. CpTi, Ti-A, and CoCr particles were also added to cultures of MG63 cells to assess their effect on proliferation (cell number) and differentiation (alkaline phosphatase activity), and PGE2 production. All three types of particles had effects on the cells. The effect on cell number was dependent on the chemical composition of the particles; Ti-A and CoCr caused a dose-dependent increase, while cpTi particles had a biphasic effect with a maximal increase in cell number observed at the 1:10 dilution. Alkaline phosphatase specific activity was also affected and cpTi was more inhibitory than Ti-A or CoCr. PGE2 production was increased by all particles, but the magnitude of the effect was particle-dependent: CoCr > cpTi > Ti-A. This study demonstrates clearly that human osteoblast-like cells and MG63 cells can phagocytose small UHMWPE, CoCr, Ti-A, and cpTi particles. Phagocytosis of the particles is correlated with changes in morphology, and analysis of MG63 response shows that cell proliferation, differentiation, and prostanoid production are affected. This may have negative effects on bone formation adjacent to an orthopaedic implant and may initiate or contribute to the cellular events that cause aseptic loosening by inhibiting bone formation. The effects on alkaline phosphatase and PGE2 release are dependent on the chemical composition of the particles, suggesting that both the type and concentration of wear debris at an implant site may be important in determining clinical outcome.     

The influence of alumina and ultra-high molecular weight polyethylene particles on osteoblast-osteoclast cooperation

Particle-induced macrophage activation, mainly by UHMWPE wear, has been recognized as the biological mechanism leading to periprosthetic bone resorption, which is responsible for the loosening of the total hip replacements (THR). Ceramic-on-ceramic implants have been advocated as a means of reducing wear products. Many studies investigated the effect of alumina (Al(2)O(3)) particles on monocytes/macrophages, but only limited information are available on their participation to bone turnover. An in vitro model was performed to investigate how Al(2)O(3) and UHMWPE particles may influence the osteoblast-osteoclast interaction: human osteoblasts (HOB) were obtained from trabecular bone, while osteoclasts were derived from peripheral blood mononuclear cells (PBMC) of healthy donors. The amount of IL6, TNF alpha, GM-CSF, and other factors acting on the bone turnover, i.e. the 'receptor activator of NF kappa B' ligand (RANKL) and osteoprotegerin (OPG), was detected in culture medium of particle-challenged HOB (HOB-CM). The Al(2)O(3) and UHMWPE particles did not affect either cell viability or TNF and GM-CSF release, while the increase in IL6 release seemed to be dependent on the particle concentration. UHMWPE increased the release of RANKL from HOB, while OPG and OPG-to-RANKL ratio were significantly inhibited. The ability of HOB-CM to promote osteoclastogenesis was tested via osteoblast/monocyte cooperation: after seven days of culture UHMWPE HOB-CM induced a large amount of multinucleated TRAP-positive giant cells, as well as significantly reduced the amount of IL6, GM-CSF and RANKL in the supernatant. With regard to the inductive effect on the osteoclastogenesis, our results show that the Al(2)O(3) wear debris are less active.     

Ultrahigh molecular weight polyethylene wear debris inhibits osteoprogenitor proliferation and differentiation in vitro

Polyethylene wear debris induces progressive osteolysis by increasing bone degradation and suppressing bone formation. Polyethylene particles inhibit the function of mature osteoblasts, but whether polyethylene particles also interfere with the proliferation and differentiation of osteoprogenitor cells is unknown. In this study, we investigated the effects of ultrahigh molecular weight polyethylene (UHMWPE) particles on the osteogenic activity of primary murine bone marrow osteoprogenitors and MC3T3-E1 preosteoblastic cells in vitro. Submicron-sized UHMWPE particles generated from wear simulator tests were isolated from serum-containing solution by density gradient centrifugation. The particles were coated onto the surface of culture wells at concentrations of 0.038, 0.075, 0.150, 0.300, and 0.600% v/v in a layer of type I collagen matrix. Primary murine bone marrow cells and MC3T3-E1 preosteoblasts were seeded onto the particle-collagen matrix and induced to differentiate in osteogenic medium for 20 days. Exposure of both cell populations to UHMWPE particles resulted in a dose-dependent decrease in mineralization, proliferation, alkaline phosphatase activity, and osteocalcin production when compared with control cells cultured on collagen matrix without particles. Complete suppression of osteogenesis was observed at particle concentrations > or =0.150% v/v. This study demonstrated that UHMWPE particles inhibit the osteogenic activity of osteoprogenitor cells, which may result in reduced periprosthetic bone regeneration and repair.     

Biological glass coating on ceramic materials: in vitro evaluation using primary osteoblast cultures from healthy and osteopenic rat bone

ZrO2 and Al2O3 substrates were successfully coated by a double layer of a silica-based glass named RKKP, using a low-cost firing technique. RKKP is a glass well known for its bioactivity; therefore, a RKKP coating on Al2O3 or ZrO2, allows to combine the excellent mechanical properties of these strong ceramic substrates with its bioactivity. ZrO2 samples were easily coated using a double layer of RKKP by a simple enamelling technique. To accommodate the thermal expansion coefficient mismatch between Al2O3 and RK K P, this substrate was coated using a multilayered composite approach. All of the coatings were characterised from a morphological and compositional point of view, and an extensive biological evaluation was performed using fresh rat osteoblasts. Osteoblast primary cultures were derived from the trabecular bone of femoral condyles harvested from intact (NB) and osteopenic (OB) rats. After characterisation of their phenotype, osteoblasts were seeded on material samples of ZrO2 or Al2O3 coated with RKKP, and cultured for 7 days. Cell proliferation (MTT test) and cell differentiation (alkaline phosphatase activity) were evaluated at the end of the experiment, to assess osteoblast behaviour in the presence of biomaterials and determine if the results were related to the host bone quality. Results of both materials showed a good level of biocompatibility. In particular, MTT significant higher values were detected in NB cultures on ZrO2-RKKP samples; ALP activity significantly increased in NB cultures on Al2O3-RKKP and in OB cultures on both coated samples.     

Polymethylmethacrylate particles inhibit osteoblastic differentiation of bone marrow osteoprogenitor cells

Aseptic implant loosening of total joint replacements often results from particle-mediated bone loss, which may be a combined effect of osteolysis and suppressed bone formation. Bone regeneration in the prosthetic bed depends on the activity of osteoblasts and their differentiation from osteoprogenitors in the bone marrow. This study investigated the effects of polymethylmethacrylate (PMMA) particles on the ability of bone marrow osteoprogenitors to differentiate into osteoblasts in vitro. Murine bone marrow cells challenged with PMMA particles on the first day of differentiation in osteogenic medium showed a dose-dependent decrease in osteoprogenitor proliferation, alkaline phosphatase expression, and mineralization. Undifferentiated bone marrow cells pretreated with PMMA particles in nonosteogenic medium for 5 days also showed a dose-dependent loss in osteogenic potential, which was sustained throughout subsequent growth in particle-free, osteogenic medium. Bone marrow cells challenged with PMMA particles after the fifth day of differentiation in osteogenic medium showed significant reductions in cellular proliferation, but not alkaline phosphatase expression and mineralization, indicating that bone marrow cells were most sensitive to particle treatment during the first 5 days of differentiation. This study demonstrated that PMMA particles inhibit osteoblastic differentiation of bone marrow osteoprogenitor cells, which may contribute to periprosthetic bone loss and implant failure.     

Molecular basis of osteoclastogenesis induced by osteoblasts exposed to wear particles

In this study, we investigate the molecular mechanisms by which human osteoblasts (HOB) challenged with wear debris promote the differentiation of osteoclast precursors. HOB were obtained from trabecular bone and exposed to alumina (Al(2)O(3)) or 'ultra-high molecular weight polyethylene' (UHMWPE) particles for 24h. The supernatant (HOB-CM) was used for the immunoenzymatic detection of receptor activator of NF-kappaB ligand (RANKL) and osteoprotegerin (OPG), as well as for inducing the osteoclast differentiation from peripheral blood mononuclear cells (PBMC). The OPG-to-RANKL ratio was significantly decreased in the conditioned medium of UHMWPE-challenged HOB. Morphological and cytochemical analysis showed that HOB-CM induced by itself the osteoclast formation, but a large amount of multinucleated tartrate-resistant acid phosphatase (TRAP)-positive giant cells were obtained when PBMCs were cultured with 1 microg/mL UHMWPE HOB-CM. The expression of genes involved in osteoclast differentiation and activation was evaluated, i.e. c-fms, RANK, c-src, c-fos, cathepsin-K (CATK), TRAP, and calcitonin R (CTR). The UHMWPE HOB-CM increases c-src expression, suggesting that polyethylene debris favour the paracrine activity of HOB in inducing the pathway involved in osteoclast polarization and adhesion. On the contrary, Al(2)O(3) HOB-CM downregulates c-fos expression, suggesting that the passage from macrophages into the osteoclast lineage is deviated. These results show that Al(2)O(3) wear debris is less active than UHMWPE in inducing osteoclast differentiation. Moreover, they provide new insight into the molecular basis of particle-induced osteoclastogenesis, that is the starting point for planning mode-specific targeting of periprosthetic osteolysis.     

内质网应激介导成骨细胞凋亡在骨溶解骨组织中的作用及机制

背景：磨损微粒能够在体外诱导成骨细胞凋亡,但是发生骨溶解的骨组织中是否也存在成骨细胞的凋亡以及骨组织中的成骨细胞凋亡信号通过何种途径进行传导目前尚不清楚。 目的：分析内质网应激反应在骨溶解骨组织中成骨细胞凋亡和骨溶解发生发展中的作用。 方法：制备磨损微粒诱导骨溶解动物模型。实验分为4组：空白对照组只接受PBS的刺激;磨损微粒组只接受纳米合金粉末悬液的刺激;内质网应激阳性对照组接受纳米合金粉末+毒胡萝卜素的刺激;内质网应激抑制组接受纳米合金粉末悬液及造模后当时、造模后1,2,3和5 d分别腹腔注射4-苯基丁酸。通过甲苯胺蓝染色、苏木精-伊红染色和碱性磷酸酶染色观察骨溶解的病理变化;分析骨溶解颅骨组织中成骨细胞分化成熟情况;Western Blot方法检测骨溶解颅骨组织内内质网应激反应标志蛋白的表达变化;TUNEL和Caspase-3免疫组织化学方法检测骨溶解颅骨组织内成骨细胞的凋亡情况。 结果与结论：磨损微粒能够在体外诱导小鼠颅骨骨溶解的发生、加重炎症细胞的浸润以及抑制成骨细胞分化成熟,同时磨损微粒还可以上调成骨细胞内质网应激反应标志蛋白以及促进骨溶解骨组织中成骨细胞的凋亡。经内质网应激抑制剂(4-苯基丁酸)的治疗后,骨溶解症状明显缓解,骨侵蚀和炎症浸润显著降低,成骨细胞的分化成熟得到改善,凋亡的成骨细胞急剧减少,内质网应激标志蛋白的表达逐渐减弱。表明内质网应激反应参与骨溶解的形成并在骨溶解的发生发展中发挥重要作用。提示内质网应激可作为一种新的治疗靶点,为临床逆转或治疗骨溶解和无菌性松动提供新的思路和方法。 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程全文链接：     

不同直径的钛颗粒负荷对成骨细胞分化和矿化能力的影响

植入假体磨损碎屑颗粒所引起的无菌性松动是假体周围骨形成与骨吸收过程失衡的结果 ,成骨细胞所参与的骨形成代谢受阻在这一病理生理过程中起着重要作用 ,而且不同大小的磨屑颗粒对骨形成影响的机制应该有所不同。为了探讨磨屑颗粒对成骨细胞骨形成能力的影响机制 ,本研究分析了 3种不同直径的钛颗粒负荷对成骨细胞分化成熟和矿化能力的影响。结果显示 ,未经钛颗粒负荷的成骨细胞表现出良好的分化和矿化能力 ;Φ6 .9μm钛颗粒负荷对成骨细胞分化和矿化能力的抑制作用并不显著 ,而 Φ2 .7μm和 Φ0 .9μm钛颗粒负荷 ,尤其是Φ 0 .9μm的抑制作用非常明显 ,且具有一定的时间依赖性。透射电镜观察显示 ,颗粒负荷后成骨细胞的骨形成功能异常与其超微结构改变有关。结合我们以前的工作 ,提示磨屑颗粒直径对骨形成的影响在无菌性松动中发挥至关重要的作用 ,亚微米级颗粒与骨形成受抑有明显的相关性 ,而大直径颗粒可促进骨吸收 ,有关不同颗粒直径对骨形成影响本质的更深入研究必将促进无菌性松动机制的早日阐明。     

Morphological characteristics of total joint arthroplasty-derived ultra-high molecular weight polyethylene (UHMWPE) wear debris that provoke inflammation in a murine model of inflammation

It is recognized that the chronic inflammation in peri-prosthetic tissue that contributes to implant failure frequently is provoked by the presence of wear debris. Some wear debris is inevitable because of the nature of the prosthesis, but not all patients develop severe inflammatory responses. The precise factors that mediate the severity of tissue inflammation to wear debris has yet to be fully defined. Because wear debris retrieved from peri-prosthetic tissue consists of a heterogeneous mixture of materials with various sizes and shapes, this study evaluated the influence of two major physical aspects of ultra-high molecular weight polyethylene (UHMWPE) wear debris (shape and surface texture) using a model of tissue inflammation. UHMWPE debris particulates recovered from 50 peri-prosthetic tissue samples were examined by scanning electron microscopy and categorized into four groups based upon aspect ratio and surface texture of the material. The four groups were defined as: 1) smooth and globular, 2) smooth and fibular, 3) rough and globular, and 4) rough and fibular. Histological analysis and ELISA assays were conducted to evaluate variations in cellular responses and cytokine production between the groups. The strongest expression of tumor necrosis factor alpha and interleukin-1 beta was found in tissues exposed to UHMWPE debris with both a rough surface texture and fibular shape, and this response was significantly elevated over debris particles with a smooth surface texture and globular shape. The data suggest that both shape and texture influence the severity of specific inflammatory responses and that rough debris surface texture exerts a marked effect on adverse tissue responses when combined with particles that have a sharp, elongated shape.     

Dynamic mechanical analysis of provisional resin materials reinforced by metal oxides

In this study, several provisional resin materials were investigated by dynamic mechanical analysis (DMA). The materials were autopolymerized PMMA and PEMA, light cured PMMA. Autopolymerized PMMA has the highest T(g) (131 degrees C) compared to that of the autopolymerized PEMA (102 degrees C) and light cured PMMA (120 degrees C). The storage moduli for autopolymerized PMMA, autopolymerized PEMA and light cured PMMA are 2.9, 1.8 and 2.3 GPa, respectively. The loss moduli of the same resins are 330, 300 and 350 MPa, respectively. Each of these resins were reinforced with 1%, 3% and 5% of each of metal oxides of MgO, ZrO2 and Al2O3 and then studied with DMA. Small changes were observed for dynamic mechanical properties tested. However, the changes are not systematic and noticeable. This is most probably due to smaller size of metal oxides particles compared to that of polymer particle size.     

Analysis of released products from oxidized ultra-high molecular weight polyethylene incubated with hydrogen peroxide and salt solutions

The wear of ultra-high molecular weight polyethylene (UHMWPE) implants generates polymeric and metallic particulate, which can be phagocytosed by human macrophages. The generation of these UHMWPE particles has been attributed to wear mechanisms and oxidation of the material. Many cell/particle studies have focused specifically on investigating particles of virgin materials themselves (i.e. virgin UHMWPE), while in fact, there is a strong likelihood that the oxidation processes encountered by the materials will yield particles with very different surface chemistries. Therefore, it is conceivable that chemical changes in the material would lead to altered cellular responses, as measured in the various cell study models. This paper has focused on the characterization of UHMWPE particulates that have been exposed to various conditions simulating processing steps and some of the oxidative and hydrolytic agents related to inflammatory responses. These include gamma-irradiation, thermal treatment and chemical oxidation by H2O2 and saline solutions. Oxidation of the particles was measured using Fourier transform infrared spectroscopy (FTIR). Degradation products were isolated from the incubation solutions using high-performance liquid chromatography (HPLC). UHMWPE particulates underwent extensive oxidation after gamma-irradiation and thermal treatments. There were marked differences following treatments of film samples taken from bar stock and the virgin particle samples. Polymer-related products, containing alkenes, alkanes and hydroxyl groups, were found in the incubation solutions. The study concluded that future work must consider both the particulates' surface chemistry and the possibility of soluble degradation products when assessing UHMWPE/cellular interactions.     

Inhibition of leukotriene function can modulate particulate-induced changes in bone cell differentiation and activity.

Aseptic loosening remains the major problem facing arthroplasty longevity with particulates from component materials touted as the cause of periprosthetic osteolysis. Proposed mechanisms in aseptic bone loss include: increased resorption, increased differentiation of osteoclasts (and/or macrophages locally), and decreased osteoblastic bone formation. Leukotrienes participate in osteoclastic bone resorption. We investigated inhibiting leukotrienes synthesis, using ICI 230487, to ameliorate the effects of particulates on osteoclast pit formation and also assessed the effects of alendronate, a bisphosphonate, on pit formation. Three particulates were used: ultra high molecular weight polyethylene (UHMWPE), polymethylmethacrylate (PMMA) and hydroxyapatite (HA). Osteoclast resorption was increased with UHMWPE, PMMA, and HA particles. Interventions with alendronate and ICI 230487 reduced particulate-induced osteoclast resorption. Both ICI 230487 and alendronate reduced osteoclast numbers at higher doses. To assess the effect of particulates on osteoclast and macrophage differentiation, mouse bone marrow was cultured and stained for tartrate resistant acid phosphatase colonies (TRAP+, osteoclasts) and nonspecific esterase positive colonies (NSE+, macrophage precursors). Particulates increased both TRAP+ and NSE+ colony formation. These increases were inhibited by ICI 230487. Particulates also inhibited osteoblast function assessed by the development of mineralized nodules and alkaline phosphatase positive (AP+) colony area. ICI 230487 partly protected osteoblast function from this particulate effect. Blockade of leukotriene production may prove a useful therapeutic intervention for particulate-induced aseptic loosening by inhibiting resorptive activity, reducing the pro-inflammatory cell populations induced and recruited by these particulates, as well as ameliorating the negative effects of inflammatory mediators on osteoblast function.Copyright 2001 John Wiley & Sons, Inc.     

Biological reaction to alumina,zirconia, titanium and polyethylene particles implanted onto murine calvaria

Periprosthetic osteolysis is a serious problem that limits long-term survival of total hip arthroplasty. Ceramics have been introduced as a joint surface material to reduce osteolysis due to wear particles. The aim of this study is to investigate the biological reaction of ceramic particles on murine calvarial bone, in comparison with polyethylene and titanium particles. Sixty CL/BL6 mice were divided into five groups according to the materials implanted onto the murine calvariae: control, Al(2)O(3), ZrO(2), high-density polyethylene (HDP) and Ti6Al4V. One week after the implantation, each calvarial tissue was dissected and the release of proinflammatory mediators (IL-1beta, IL-6, TNF-alpha) and bone resorption were assessed. The particles of HDP and Ti6Al4V induced three and two times larger osteolytic lesions than the control, respectively. The levels of IL-1beta and IL-6 were significantly elevated in the medium subcultured with the calvariae of HDP and Ti6Al4V groups. Any particle type did not increase the levels of TNF-alpha. There were no significant differences observed in the levels of proinflammatory mediators or osteolytic area among Al(2)O(3), ZrO(2) and control groups. The inflammatory response and bone resorption induced by ceramic particles were much smaller than those induced by HDP and Ti6Al4V. These biological features suggest the biocompatibility of ceramics as a joint surface material for artificial joints.     

Induction of macrophage apoptosis by ceramic and polyethylene particles in vitro

The purpose of this study was to investigate in vitro the presence of apoptotic cell death after macrophage stimulation with different ceramic (Al2O3 and ZrO2) and high density polyethylene (HDP) particles. We also analyzed the effects of particle size, concentration, and composition. The J774 mouse macrophage cell line was exposed to commercial particles of different sizes (up to 4.5 microm) and concentrations (up to 500 particles per macrophage). Fluorescence microscopy and DNA laddering were used to investigate the presence of apoptosis in cell cultures after 24 h of incubation. Fluorescence microscopy of propidium iodide stained cells showed two characteristic morphological features that occur in apoptotic cells, namely nuclear condensation and heterogeneity of stain uptake. The effect of ceramic particles on apoptotic nuclear morphology was size- and concentration-dependent and reached a plateau above 150 particles per macrophage at 1.3 microm. With regards to composition, we did not find any difference in cell morphology between Al2O3 and ZrO2. Ceramic and HDP particles induced DNA fragmentation into oligonucleosomes as evidenced by DNA laddering, another characteristic of apoptosis. The induction of DNA laddering was size- and concentration-dependent whereas particle composition (Al2O3 vs. ZrO2 and Al2O3 vs. HDP) had no effect. In conclusion, our results demonstrated that ceramic and HDP particles induce macrophage apoptotic cell death in vitro and open doors for possible modulation of debris-induced periprosthetic osteolysis.     

葛根素在体外对成骨细胞增殖和分化的影响

背景：成骨细胞是骨代谢平衡过程中的关键功能细胞,植物雌激素对成骨细胞的增殖和分化有重要影响,葛根素作为植物雌激素的一种,在体外以较大范围浓度对成骨细胞功能的影响仍少见报道。 目的：观察葛根素在体外对大鼠成骨细胞增殖和分化功能的影响。 方法：取新生Wistar大鼠的颅盖骨,对成骨细胞进行分离、培养、纯化及鉴定。将培养的成骨细胞随机分为对照组、10-3~10-10 mol/L不同浓度葛根素组,观察不同浓度葛根素对体外培养的成骨细胞增殖和碱性磷酸酶活性表达的影响。 结果与结论：细胞经葛根素处理后10-5~10-9 mol/L组成骨细胞增殖活性较对照组明显增加(P < 0.05),第3天增殖最快(P < 0.01),第4天开始下降;诱导第4天,各组碱性磷酸酶活性与对照组相比,差异均有显著性意义(P < 0.01),其中以10-6 mol/L组最显著(P < 0.01)。然而葛根素10-3 mol/L组成骨细胞增殖活性、碱性磷酸酶活性表达较对照组均减少(P < 0.05)。提示葛根素对成骨细胞的影响存在剂量依赖性,并且具有双向性,即在低浓度(10-5~10-8 mol/L)下刺激骨形成;在高浓度(10-3~10-4 mol/L)下抑制骨形成。     

人工关节磨损颗粒诱导钛合金植入物松动的实验研究

为比较不同人工关节微小磨损颗粒诱导假体周围骨吸收及假体松动生物效应的差异。方法：本实验采用Ｘ线摄片骨密度灰度值测定以及植入物推出力学强度测试等手段,对Ｔｉ－６Ａｌ－４Ｖ,Ｃｏ－Ｃｒ－Ｍｏ与ＵＨＭＷＰＥ三种微小颗粒诱导的假体周围骨结构改变及假体松动作定量分析。结果统计分析显示,直径２．５微米的Ｔｉ－６Ａｌ－４Ｖ颗粒诱导植入物旁骨吸收与植入物松动的程度明显低于相同直径的Ｃｏ－Ｃｒ－Ｍｏ颗粒或ＵＨＭＷＰ     

Proliferation and differentiation of rat calvarial osteoblasts on type I collagen-coated titanium alloy.

Several attempts have been made to improve osseointegration of titanium alloy as an implant material by modification of its surface. In the present study, proliferation, differentiation, and mineralization of osteoblasts on type I collagen-coated Ti6Al4V were investigated. The activity of alkaline phosphatase and the accumulation of calcium by osteoblasts grown on titanium alloy were significantly higher compared to cells grown on polystyrene. Precoating of the implant surface with type I collagen did not extensively affect proliferation, the activity of alkaline phosphatase, collagen synthesis, calcium accumulation, or the mRNA levels for collagen I alpha1, osteopontin, osteocalcin, MMP-2, and TIMP-2. Maximum collagen synthesis by osteoblasts was observed at day 4 of culture independent of the type of implant material. The specific activity of alkaline phosphatase reached its maximum at day 18 of culture. Accumulation of calcium and elevated mRNA levels for osteocalcin were found at day 22. These results indicate that collagen-coating alone is not sufficient to accelerate differentiation of rat calvarial osteoblasts on Ti6Al4V.     

Alumina and zirconia coated vitallium oral endosteal implants in beagles.

The purpose of this study was to assess the addition of a ceramic coating upon a Vitallium implant to increase the implant's biologic acceptability in the oral environment. The mandibular premolar teeth in 9 adult beagle dogs were removed bilaterally and these areas allowed to heal for 6 weeks. Ceramic coating with either Al2O3 or ZrO2 was carried out by flame spray deposition upon Vitallium anchor implants (9 of each), and the implants placed into the 18 healed premolar areas. Clinical and radiographic evaluation was conducted by 2 independent investigators over a 32 week period. Implants which exhibited mobility greater than II on a scale of 0 to III, at intervals of one-half, were judged unsatisfactory. After 19 weeks, all 9 Al2O3 coated implants and 5 ZrO2 coated implants were rated unsatisfactory. After 32 weeks, 4 ZrO2 coated implants were in situ with 0 or I mobility. Radiographically the width of the peri-implant space increased in direct proportion to both time and mobility. Histologic sections demonstrated encapsulating dense fibrous connective tissue which was oriented parallel to both ZrO2 and Al2O3 implants. Results suggest the zirconia used is a superior ceramic coating to the alumina. Neither seemed to increase biologic acceptability over uncoated Vitallium implants.     

Effect of a CCR1 receptor antagonist on systemic trafficking of MSCs and polyethylene particle-associated bone loss

Particle-associated periprosthetic osteolysis remains a major issue in joint replacement. Ongoing bone loss resulting from wear particle-induced inflammation is accompanied by continued attempts at bone repair. Previously we showed that mesenchymal stem cells (MSCs) are recruited systemically to bone exposed to continuous infusion of ultra high molecular weight polyethylene (UHMWPE) particles. The chemokine-receptor axis that mediates this process is unknown. We tested two hypotheses: (1) the CCR1 receptor mediates the systemic recruitment of MSCs to UHMWPE particles and (2) recruited MSCs are able to differentiate into functional mature osteoblasts and decrease particle-associated bone loss. Nude mice were allocated randomly to four groups. UHMWPE particles were continuously infused into the femoral shaft using a micro-pump. Genetically modified murine wild type reporter MSCs were injected systemically via the left ventricle. Non-invasive imaging was used to assay MSC migration and bone mineral density. Bioluminescence and immunohistochemistry confirmed the chemotaxis of reporter cells and their differentiation into mature osteoblasts in the presence of infused particles. Injection of a CCR1 antagonist decreased reporter cell recruitment to the UHMWPE particle infusion site and increased osteolysis. CCR1 appears to be a critical receptor for chemotaxis of MSCs in the presence of UHMWPE particles. Interference with CCR1 exacerbates particle-induced bone loss.     

Response of human osteoblasts exposed to wear particles generated at the interface of total hip stems and bone cement

Aseptic loosening of total hip replacement is mainly caused by wear particles. Abrasive wear occurs at articulating surfaces or as a consequence of micro-motions at the interface between femoral stem and bone cement. Direct impact of wear particles on osteolysis, the remodeling of the bone stock and a directly affected function of osteoblasts was described. The present study examined the response of human osteoblasts exposed to different wear particles, which were generated in a test device providing oscillating micro-motions at the interface between femoral stem and standard bone cement. Characterization of released particles was performed by quantifying the size distribution and the metal content of the wear debris. Human osteoblasts were incubated with particles obtained from hip stems with different material compositions (Ti-6Al-7Nb and Co-28Cr-6Mo) and rough and smooth surface finishings combined with standard bone cement (Palacos(R) R) containing zirconium oxide particles. Commercially pure titanium particles (cp-Ti) and particulate zirconium oxide (ZrO(2)) were used for comparative analyses. The results revealed significant (p < 0.05) reduction of the cell viability after exposure to higher concentration of metallic particles, particularly from Co-based alloys. In contrast, ZrO(2) alone showed significantly less adverse effects on the cells. When increasing metallic particle concentrations massive inhibition was also observed in the release of cytokines including interleukine-6 (IL-6) and interleukine-8 (IL-8), but the expression of Procollagen I and the cell viability showed the highest reduction after exposure to Co-based alloy particles from rough stems.