Effect of polymer molecular weight and addition of calcium stearate on response of MG63 osteoblast-like cells to UHMWPE particles.

Periprosthetic osteolysis and implant loosening is associated with the presence of ultrahigh molecular weight polyethylene (UHMWPE) wear debris particles. Osteoblast phenotypic expression in vitro is affected by UHMWPE particles, suggesting that bone formation may also be affected by wear debris. Here we tested the hypothesis that the response of osteoblasts to UHMWPE can be modified by changes in UHMWPE particle chemistry. We used four different commercially available preparations of GUR UHMWPE particles to determine if chemical composition (+/- Ca-stearate) or polymer molecular weight (3.1-4.2 million or 5.4-6.5 million g/mol) modulates osteoblast response. Particles were characterized by size distribution, morphology, and number of particles added to the culture medium. They had an average equivalent circle diameter ranging from 0.46-1.26 microm. MG63 cell response was assessed by measuring cell number, cellular and cell layer alkaline phosphatase, and prostaglandin E2 (PGE2) production. There were dose-dependent effects of the particles on cell response. Cell number and PGE, production were increased, while alkaline phosphatase specific activity was decreased. In addition, there was a marked difference between cultures treated with particles containing Ca-stearate and as a function of polymer molecular weight. Particles of higher molecular weight caused a greater stimulation of proliferation and inhibition of alkaline phosphatase than particles of lower molecular weight. The presence of Castearate exerted a more pronounced depression of osteoblast phenotype as well as a significantly greater increase in PGE2 release by the cells. The present study shows that chemical composition and polymer molecular weight of UHMWPE are capable of modulating osteoblast response to particles. The results suggest that osteoblast differentiation is inhibited by UHMWPE particles, whereas cell proliferation and PGE2 production are stimulated. This may have direct effects on osteoblasts and bone formation, but also paracrine effects on cells of the monocytic lineage inducing bone resorption and promoting inflammation which may lead to aseptic loosening. The present results suggest that the cellular events in aseptic loosening may be modulated or even accelerated by changes in the composition of the UHMWPE used to fabricate implants.     

Ultrahigh molecular weight polyethylene particles have direct effects on proliferation,differentiation, and local factor production of MG63 osteoblast-like cells

Small particles of ultrahigh molecular weight polyethylene stimulate formation of foreign-body granulomas and bone resorption. Bone formation may also be affected by wear debris. To determine if wear debris directly affects osteoblasts, we characterized a commercial preparation of ultrahigh molecular weight polyethylene (GUR4150) particles and examined their effect on MG63 osteoblast-like cells. In aliquots of the culture medium containing ultrahigh molecular weight polyethylene, 79% of the particles were less than 1 μm in diameter, indicating that the cells were exposed to particles of less than 1 μm. MG63 cell response to the particles was measured by assaying cell number. [³H]thymidine incorporation, alkaline phosphatase specific activity, osteocalcin production, [³⁵S]sulfate incorporation, and production of prostaglandin E₂ and transforming growth factor-β. Cell number and [³H]thymidine incorporation were increased in a dose-dependent manner. Alkaline phosphatase specific activity, a marker of cell differentiation for the cultures, was significantly decreased, but osteocalcin production was not affected. [³⁵S]sulfate incorporation, a measure of extracellular matrix production, was reduced. Prostaglandin E₂ release was increased, but transforming growth factor-β production was decreased in a dose-dependent manner. This shows that ultrahigh molecular weight polyethylene particles affect MG63 proliferation, differentiation, extracellular matrix synthesis, and local factor production. These effects were direct and dose dependent. The findings suggest that ultrahigh molecular weight polyethylene wear debris particles with an average size of approximately 1 μm may inhibit bone formation by inhibiting cell differentiation and reducing transforming growth factor-β production and matrix synthesis. In addition, increases in prostaglandin E₂ production may not only affect osteoblasts by an autocrine pathway but may also stimulate the proliferation and activation of cells in the monocytic lineage. These changes favor decreased bone formation and increased bone resorption as occur in osteolysis.     

Nitric oxide and prostaglandin E2 production in response to ultra-high molecular weight polyethylene particles depends on osteoblast maturation state

BACKGROUND: Recent studies have shown that osteoblast-like cells respond directly to ultra-high molecular weight polyethylene particles in culture, suggesting that they may be involved in aseptic loosening of endoprostheses. We tested the hypothesis that the state of cell maturation plays a role in the response of osteogenic cells to ultra-high molecular weight polyethylene particles. METHODS: MG63 cells (immature osteoblast-like cells), OCT-1 cells (mature secretory osteoblast-like cells), and MLO-Y4 cells (osteocyte-like cells) were treated for twenty-four hours with commercial ultra-high molecular weight polyethylene particles with an average diameter of 1 mm. The effect of particle treatment on cell proliferation was assessed by measuring the number of cells, whereas the effects on differentiation and local factor production were assessed by measuring the production of osteocalcin, prostaglandin E2, and nitric oxide. The effect of particles on apoptosis was also evaluated. RESULTS: The addition of ultra-high molecular weight polyethylene particles increased the number of MG63 cells, did not affect the number of OCT-1 cells, and led to a decrease in the number of MLO-Y4 cells. The observed changes in cell number were not due to programmed cell death, as no more than 3% of the cells in cultures treated with the highest concentration of particles were undergoing apoptosis. Osteocalcin production was not affected by the addition of particles. Prostaglandin E2 production was increased in all three types of cultures, but the effect was greatest in OCT-1 cell cultures, as was the absolute amount of prostaglandin E2 produced. Nitric oxide production was unaffected in MG63 cell cultures, but it was stimulated in OCT-1 and MLO-Y4 cell cultures. CONCLUSIONS: The results of the present study support the hypothesis that osteoblast cell maturation state plays an important role in the response to ultra-high molecular weight polyethylene particles and that the terminally differentiated osteocyte may be involved in the bone response to wear debris in vivo.     

Osteolysis in alloarthroplasty of the hip. The role of ultra-high molecular weight polyethylene wear particles

Massive localized osteolysis around artificial joints has been seen more frequently in the past few years. It is still not generally accepted that ultra-high molecular weight polyethylene (UHMWPE) wear particles can induce massive bone resorption, even distant from the joint. This article describes a series of eight soft-top prostheses with large UHMWPE ball heads that contributed to the erosion of surrounding bone. Roentgenographically, all of the cases showed a marked loss of proximal cortical bone, more or less combined with osteolysis, which was distal to the femoral shaft and deep into the acetabulum. In two cases, remodeling and resorption transformed the bone into a tumorlike appearance. Tissue samples from areas of osteolysis as well as from the joint capsule were taken at revision surgery, processed for histology, examined microscopically, and evaluated semiquantitatively. The retrieved devices were also carefully inspected. Large amounts of UHMWPE wear debris were found not only in the joint capsule but also in layers of granulomatous tissue from the acetabulum and femur, whereas metal particles and fragmented polymethylmethacrylate were either completely absent or occurred only in very small amounts. The results of this study demonstrate that UHMWPE wear products alone can cause massive osteolysis by triggering foreign-body granuloma formation at the bone-cement interface; the bone cement may remain fixed in areas beyond the osteolytic transformation.     

The effectiveness of polyethylene versus titanium particles in inducing osteolysis in vivo.

Bearing surface wear and periprosthetic osteolysis due to wear particles are among the most common reasons for joint replacement failure. A murine calvarial model of wear particle-induced osteolysis has been used to identify different biologic factors associated with this problem and to test nonsurgical methods of modulating the host response to particulate debris. This model has utilized titanium particles, however, in clinical practice the most common source of particulate debris is polyethylene particles from bearing surface wear. We now report a calvarial model of wear particle-induced osteolysis based on commercially available polyethylene particles. We found that compared to sham surgery osteoclast recruitment and bone resorption can be induced by introduction of the titanium particles or polyethylene particles. However, bone resorption was significantly higher with polyethylene particles compared to titanium particles (p=0.02). We consider the polyethylene based murine calvarial model of wear particle-induced osteolysis a reliable and clinically relevant tool to understand the host factors and potential pharmacologic interventions that can influence wear debris generated osteolysis. This model might serve as an extension of the well-established titanium based bone resorption model.     

磨损微粒引起的成骨细胞内质网应激反应在骨溶解中的作用及机制研究

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,探讨人工关节松动的原因,为人工关节松动的防治提供新的思路和理论依据。方法：采用小鼠颅骨建立磨损微粒诱导骨溶解的动物模型,随机分成4组,每组7只：组1,空白对照组;组2,磨损微粒TiAl6V4纳米合金粉末（TiNPs）组;组3,内质网应激反应阳性对照（TiNPs+Tg）组;组4,内质网应激反应抑制剂（TiNPs+4-PBA）组。通过甲苯胺蓝染色、HE染色和ALP染色观察骨溶解的病理变化;Western Blotting方法检测骨溶解颅骨组织中内质网应激反应标志蛋白的表达变化;TUNEL和Caspase-3免疫组化方法检测骨溶解颅骨组织内成骨细胞的凋亡情况。结果：磨损微粒TiNPs能够在体外诱导骨溶解的发生、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。在磨损微粒TiNPs的基础上加入内质网应激的抑制剂（4-PBA）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也逐渐减弱。结论：内质网应激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。同时,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。     

Presence of silicon in massive osteolysis after cementless total hip arthroplasty: Case report

Osteolysis associated with artificial joint arthroplasty seems to be the result of particles of wear debris (from ultra-high molecular weight polyethylene (UHMWPE), polymethylmethacrylate (PMMA), and metals), causing a macrophage response, Silicon particles, as residual contaminants embedded in the surface of both textured metal implants and polyethylene sockets, may be a factor in osteolysis. We present the case of a 57-year-old woman who had massive and aggressive osteolysis. The osteolytic lesion was isolated in the greater trochanter region 5 years after she had had primary cementless total hip arthroplasty (Cobalt-chromium alloy). There were no signs of mechanical loosening, but she experienced moderate pain. Under a polarized microscope and scanning electron micrography, a biopsied specimen from the osteolytic lesion revealed conglomerates of UHMWPE particles of various sizes between proliferated synovium-like cells. Quantative energy-dispersive X-ray analysis focused on the conglomerates of UHMPWE particles demonstrated a marked presence of silicon. Although the definitive causative factor for a the osteolysis was regarded as a foreign-body reaction induced by UHMWPE particles, the presence of silicon was interesting in terms of the pathogenesis of osteolysis associated with artificial joint surgery.     

Osteolysis in alloarthroplasty of the hip. The role of bone cement fragmentation

Movement at the interface between bone and cement and fractures of the cement can cause fragmentation of the polymethylmethacrylate (PMMA) bone cement implant. In order to obtain further information about the effect of PMMA fragments on the surrounding tissue and the role of such particles in the development of bone resorption, the authors investigated 17 patients with cemented total hip endoprostheses showing osteolysis and implant loosening in the femoral shaft with (Group B) and without (Group A) involvement of the acetabulum. The roentgenographic follow-up examinations revealed an initially slow and later more rapid extension of the endosteal bone erosions, with a predilection for the tip of the stem, the lesser trochanter, and laterally for the middle of the stem. At revision surgery, tissue samples were taken from the joint capsule and the bone-cement interface, in particular from the osteolysis in the femoral shaft and the acetabulum. The tissue samples were processed for histology, microscopically examined, and semiquantitatively evaluated. The retrieved devices were also carefully inspected. Large foreign-body granulomas were found at the bone-cement interface and in the joint capsule. Histiocytes and foreign-body giant cells stored particles of PMMA and polyethylene, of which fragmented bone cement predominated. Granulomatous tissue invaded bone canals and marrow spaces and induced resorption of the surrounding bone. In four cases in Group A, tissue from the osteolysis contained only fragmented bone cement, demonstrating that PMMA particles alone may be responsible for triggering focal bone resorption. Osteolysis seems to begin at the site where disintegration of bone cement begins. In cases in which polyethylene particles were found in the tissue in addition to fragmented bone cement, wear from the ultrahigh molecular weight polyethylene socket has been increased by entrapment of PMMA particles between the joint surfaces. Thus, fragmentation of bone cement and abrasion of polyethylene enhance each other. Bone cement particles promote polyethylene wear, which in turn promotes granuloma formation, bone resorption, and subsequent bone cement disintegration.     

Polyethylene particles from a hip simulator cause (45)Ca release from cultured bone

Periprosthetic osteolysis is a dominant factor in the success or failure of total hip prostheses. Polyethylene wear debris has been implicated in the process of bone resorption and subsequent implant loosening. The present study is the first to examine the effect of ultra high molecular weight polyethylene (UHMWPE) wear debris produced by a hip simulator on calvarial bone resorption in vitro. (45)Ca release was measured in cultured mouse calvarial bone samples. Although short-term exposure to UHMWPE particles (2 h) decreased (45)Ca release, longer-term exposure for 1-2 days increased release in a dose-dependent manner. After one-day exposure to 7.5 x 10(6) particles per mL, 18% more (45)Ca was released from cultured calvarial bone than from control samples. It was concluded that UHMWPE wear particles either directly or indirectly stimulated osteoclasts to activate bone resorption. Polyethylene wear debris contributes to the osteolytic process at the bone-implant interface.     

Synergistic effect of particles and cyclic pressure on cytokine production in human monocyte/macrophages: proposed role in periprosthetic osteolysis.

Macrophages, activated by particulate wear debris, are important in the process of osteolysis, which occurs during joint implant loosening. We previously found increased levels of interleukin-1beta (IL-1beta), IL-6, and tumor necrosis factor-alpha in cultured macrophages subjected to cyclical pressure of 0.138 MPa, suggesting that cyclic pressure may be another relevant cause of macrophage activation. The current study first investigated the effects of a range of cyclic pressures on cultured macrophages, including an investigation of the time course of cytokine expression. At 0.138 MPa, supernatant levels of TNF-alpha were maximal at 12 h, whereas IL-6 and IL-1beta were maximal at 24 h. All four cyclic pressure levels tested (without particles) resulted in increased production of all three cytokines relative to control. These increases were most marked at 0.069 and 0.035 MPa, and the increase in cytokine production at 0.017 MPa was not statistically significant. Further studies demonstrated that conditioned media from cyclically pressurized macrophages stimulated bone resorption in a neonatal mouse calvarial assay system. There were increased levels of calcium released from calvaria cultured in conditioned media from pressurised monocytes, and an increase in tartate-resistant acid phosphatase-positive osteoclasts was observed microscopically. As particulate wear debris is important in implant loosening, ultra high molecular weight polyethylene particles were also added to the pressurized cell cultures. The experiments compared the effect of atmospheric pressure, cyclic pressure alone, particles alone, and particles and cyclic pressure combined. A combination of ultra high molecular weight polyethylene particles and cyclic pressure at 0.017 MPa resulted in a dramatic synergistic elevation of levels of all three cytokines compared with the levels found with either pressure or particles alone. We propose that monocyte/macrophage activation by cyclic pressure plays a major role in the osteolysis seen in aseptic loosening of implants. The synergistic effect observed between particles and pressure could accelerate implant loosening, and implies that reduction in either cyclic pressure (by improving implant fixation) or wear debris load would reduce osteolysis.     

New insight into the mechanism of hip prosthesis loosening: effect of titanium debris size on osteoblast function.

The incidence of rheumatoid arthritis and osteoarthritis is on the rise due to our expanding elderly population. Total joint arthroplasty is the most successful, prevalent treatment modality for these and other degenerative hip conditions. Despite the wide array of prosthetic devices commercially available, hip prostheses share a common problem with a gradual and then accelerating loss of bone tissue and bone-implant interface integrity, followed by implant instability and loosening. Implant failure is largely the result of inevitable wear of the device and generation of wear debris. To provide information for the development of improved prosthetic wear characteristics, we examined the effects of size-separated titanium particles on bone forming cell populations. We demonstrate unequivocally that particle size is a critical factor in the function, proliferation, and viability of bone-forming osteoblasts in vitro. In addition, we have elucidated the time-dependent distribution of the phagocytosed particles within the osteoblast, indicating an accumulation of particles in the perinuclear area of the affected cells. The report finds that particle size is a critical factor in changes in the bone formation-related functions of osteoblasts exposed to simulate wear debris, and that 1.5-4 microm titanium particles have the greatest effect on osteoblast proliferation and viability in vitro. The size of titanium particles generated through wear of a prosthetic device may be an important consideration in the development of superior implant technology.     

Polyethylene and titanium particles induce osteolysis by similar, lymphocyte-independent, mechanisms.

Periprosthetic osteolysis is a major clinical problem that limits the long-term survival of total joint arthroplasties. Osteolysis is induced by implant-derived wear particles, primarily from the polyethylene bearing surfaces. This study examined two hypotheses. First, that similar mechanisms are responsible for osteolysis induced by polyethylene and titanium particles. Second, that lymphocytes do not play a major role in particle-induced osteolysis. To test these hypotheses, we used the murine calvarial model that we have previously used to examine titanium-induced osteolysis. Polyethylene particles rapidly induced osteolysis in the murine calvaria 5-7 days after implantation. The polyethylene-induced osteolysis was associated with large numbers of osteoclasts as well as the formation of a thick periosteal fibrous tissue layer with numerous macrophages containing phagocytosed polyethylene particles. Polyethylene-induced osteolysis was rapidly repaired and was undetectable by day 21 after implantation. Lymphocytes were noted in the fibrous layer of wild-type mice. However, the amount of osteolysis and cytokine production induced by polyethylene particles was not substantially affected by the lack of lymphocytes in Pfp/Rag2 double knock out mice. All of these findings are similar to our observations of osteolysis induced by titanium particles. These results provide strong support for both of our hypotheses: that similar mechanisms are responsible for osteolysis induced by polyethylene and titanium particles and that lymphocytes do not play a major role in particle-induced osteolysis.     

Analysis of wear and oxidation on retrieved bipolar polyethylene liner

BACKGROUND: For bipolar prostheses, most of the previous studies attributed the occurrence of osteolysis to wear debris generated from the bearing surface. We looked closely into the wear debris and reported on our findings with respect to the oxidation index and the rate of wear in ultra-high molecular weight polyethylene (UHMWPE) inserts retrieved from bipolar prostheses after various spans of time in vivo. METHOD: The inserts were retrieved from the heads of three types of bipolar prosthesis (UH1, UPF1, UPF2). We retrieved 24 bipolar prostheses from 23 patients whose mean implantation period was 10.0 years (2.7-15.4 years). RESULTS: All the retrieved polyethylene had a burnished bearing surface. In all, 92% (22/24) of these inserts had indentation and roughness at the rim and flange, suggesting neck-cup impingement; periprosthetic fracture occurred in the other two inserts. The mean linear wear rate was 0.035 mm per year. The average maximum oxidation index for the inserts with osteolysis was 3.34, and it was was 3.49 for the inserts without osteolysis. We, therefore, could not detect any significant difference between the aforesaid groups of inserts. CONCLUSIONS: The results strongly suggest that most of the polyethylene wear debris was not generated from the bearing surface. Moreover, the wear debris generated from neck-cup impingement may well be the cause of an inflammatory reaction, which in turn has a strong potential to become the primary cause of osteolysis.     

磨损颗粒引发假体周围骨溶解的机制及药物治疗进展

目的对磨损颗粒引发假体周围骨溶解的机制及药物治疗进展进行了综述介绍。方法对国外近期相关文献进行综述。结果及结论人工关节置换是治疗晚期骨关节炎及老年股骨颈骨折的有效方法,能达到解除关节疼痛、重建活动功能及提高生活质量的目的。人工关节无菌性松动是影响人工关节使用寿命和远期疗效的最重要的并发症。磨损颗粒诱发假体周围组织细胞产生一系列生物学反应是导致假体周围骨溶解及假体无菌性松动的重要因素。磨损颗粒刺激假体周围的巨噬细胞、成骨细胞、成纤维细胞等产生多种细胞因子,形成破骨细胞性骨吸收,同时影响成骨细胞的分化及功能,抑制骨形成。药物在预防和治疗磨损颗粒引起的人工关节松动方面能起到积极的作用。     

Identification of nanometre-sized ultra-high molecular weight polyethylene wear particles in samples retrieved in vivo

Nanometre-sized particles of ultra-high molecular weight polyethylene have been identified in the lubricants retrieved from hip simulators. Tissue samples were taken from seven failed Charnley total hip replacements, digested using strong alkali and analysed using high-resolution field emission gun-scanning electron microscopy to determine whether nanometre-sized particles of polyethylene debris were generated in vivo. A randomised method of analysis was used to quantify and characterise all the polyethylene particles isolated. We isolated nanometre-sized particles from the retrieved tissue samples. The smallest identified was 30 nm and the majority were in the 0.1 microm to 0.99 microm size range. Particles in the 1.0 microm to 9.99 microm size range represented the highest proportion of the wear volume of the tissue samples, with 35% to 98% of the total wear volume comprised of particles of this size. The number of nanometre-sized particles isolated from the tissues accounted for only a small proportion of the total wear volume. Further work is required to assess the biological response to nanometre-sized polyethylene particles.     

Migration of polyethylene wear debris in one type of uncemented femoral component with circumferential porous coating: an autopsy study of 5 femurs

This autopsy study analyzed the migration of polyethylene wear debris adjacent to uncemented femoral components with circumferential porous coating. Five femoral specimens retrieved at autopsy from 3 patients were investigated. Two stems were 40% porous coated, 2 were 80% porous coated, and 1 was 100% porous coated. The implants' time in situ ranged from 53 to 132 months (average, 94.8 months). All patients were followed clinically and radiographically until death. Radiographically, 3 of the stems were bone ingrown with proximal bone loss, 1 stem was mainly fibrous encapsulated, and 1 stem was completely fibrous encapsulated. Histologic examination of bone adjacent to the middle and distal sections of the femoral implant revealed no polyethylene wear debris or granulation tissue in any of the specimens. In 2 bone-ingrown cases, a small number of polyethylene particles and small areas of granulation tissue were present at the proximal level. This granulation tissue, however, did not cause major osteolysis. The findings in this study imply that circumferential porous coating of cementless femoral components could prevent distal migration of polyethylene wear debris along the bone-implant interface in both bone-ingrown and fibrous-encapsulated femoral implants.     

Periprosthetic osteolysis: genetics, mechanisms and potential therapeutic interventions

Aseptic loosening and periprosthetic osteolysis occur as a result of the biological response to particulate wear debris and are one of the leading causes of arthroplasty failure. Periprosthetic osteolysis originates from chronic inflammatory responses triggered by implant-derived particulate debris, which cause recruitment of cells, including macrophages, fibroblasts, lymphocytes and osteoclasts. These cells secrete proinflammatory and osteoclastogenic cytokines, exacerbating the inflammatory response. In addition to their direct activation by phagocytosis, there are contributing autocrine and paracrine effects that create a complex milieu within the periprosthetic space, which ultimately governs the development of osteolysis. Chronic cell activation may upset the delicate balance between bone formation and bone resorption leading to periprosthetic osteolysis. This article summarizes the genetic mechanisms underlying periprosthetic loosening and identifies potential therapeutic agents.