Establishment and Evaluation of an In Vitro System for Biophysical Stimulation of Human Osteoblasts.
ABSTRACT: While several studies investigated the effects of mechanical or electrical stimulation on osseointegration and bone fracture healing, little is known about the molecular and cellular impact of combined biophysical stimulation on peri-implant osseointegration. Therefore, we established an in vitro system, capable of applying shear stress and electric fields simultaneously. Capacitively coupled electric fields were used for electrical stimulation, while roughened Ti6Al4V bodies conducted harmonically oscillating micromotions on collagen scaffolds seeded with human osteoblasts. Different variations of single and combined stimulation were applied for three days, while samples loaded with Ti6Al4V bodies and untreated samples served as control. Metabolic activity, expression of osteogenic markers and bone remodeling markers were investigated. While combined stimulation showed no substantial benefit compared to sole mechanical stimulation, we observed that 25 µm micromotions applied by roughened Ti6Al4V bodies led to a significant increase in gene expression of osteocalcin and tissue inhibitor of metalloprotease 1. Additionally, we found an increase in metabolic activity and expression of bone remodeling markers with reduced procollagen type 1 synthesis after 100 mVRMS electrical stimulation. We were able to trigger specific cellular behaviors using different biophysical stimuli. In future studies, different variations of electrical stimulation will be combined with interfacial micromotions.
Project description:Here we show a new and effective methodology for rapid/controllable porosification of thin-film ceramics, which may be applied in medical devices/electronics and membrane nano-filtration. Dense hydroxyapatite applied to Ti6Al4V by plasma-assisted PVD was electron-beam irradiated to induce flash melting/boiling. Deposited coatings contained amorphous and nano-crystalline/stoichiometric hydroxyapatite (~35?nm). Irradiation (voltages 13-29?kV) led to ablation (up to 45% mass loss) and average/maximum pore areas from (0.07-1.66)/(0.69-92.53) ?m<sup>2</sup>, mimicking the human cortical bone. Vitrification above 1150?°C formed (~62-30?nm) crystallites of ?-Tri Calcium Phosphate. Unique porosification resulted from irradiation-induced sub-surface boiling and limited thermal conductivity of hydroxyapatite, causing material to expand/explode through the more quickly solidified top surface. Commercially applicable, roughened Ti6Al4V exacerbated the heating and boiling explosion phenomenon in certain regions, producing an array of pore sizes. Scaffold-like morphologies were generated by interconnection of micron/sub-micron porosity, showing great potential for facile generation of a biomimetic surface treatment for osseointegration.
Project description:Dental implantation has been the primary method for the treatment of tooth loss, but longer than 3 months healing times are generally required. Because immediate load implants are suitable only for certain categories of implant patients, it has value to develop a novel method to facilitate the implant-bone osseointegration process. Cylindrical titanium implants were implanted in the tooth sockets of beagles, and microelectrode stimulation of the sympathetic nerves in the infraorbital nerve was performed after implantation for 1 week. The authors found that one-sided nerve stimulation was shown to evoke consistent electric potential changes in both sides of the infraorbital nerves. Moreover, after 4 weeks of implantation, more new bone was clearly observed around the implants in the beagles that received electrical stimulation treatment than was observed in the control animals. Furthermore, a higher mineralization density was measured in the new peri-implant bone tissues of the stimulated beagles when compared to controls. These results demonstrate that the simple and safe physical method of microelectrode stimulation to sympathetic nerves can promote the formation of new bone and the osseointegration of implants. This technique is worth promoting and has the potential to reduce the healing time of dental implantation in future clinical cases.
Project description:The insufficient osteogenesis and osseointegration of porous titanium based scaffold limit its further application. Early angiogenesis is important for scaffold survival. It is necessary to develop a multifunctional surface on titanium scaffold with both osteogenic and angiogenic properties. In this study, a biofunctional magnesium coating is deposited on porous Ti6Al4V scaffold. For osseointegration and osteogenesis analysis, in vitro studies reveal that magnesium-coated Ti6Al4V co-culture with MC3T3-E1 cells can improve cell proliferation, adhesion, extracellular matrix (ECM) mineralization and ALP activity compared with bare Ti6Al4V cocultivation. Additionally, MC3T3-E1 cells cultured with magnesium-coated Ti6Al4V show significantly higher osteogenesis-related genes expression. In vivo studies including fluorochrome labeling, micro-computerized tomography and histological examination of magnesium-coated Ti6Al4V scaffold reveal that new bone regeneration is significantly increased in rabbits after implantation. For angiogenesis studies, magnesium-coated Ti6Al4V improve HUVECs proliferation, adhesion, tube formation, wound-healing and Transwell abilities. HUVECs cultured with magnesium-coated Ti6Al4V display significantly higher angiogenesis-related genes (HIF-1? and VEGF) expression. Microangiography analysis reveal that magnesium-coated Ti6Al4V scaffold can significantly enhance the blood vessel formation. This study enlarges the application scope of magnesium and provides an optional choice to the conventional porous Ti6Al4V scaffold with enhanced osteogenesis and angiogenesis for further orthopedic applications.
Project description:This study was aimed to investigate the osseointegration ability of poly(ether ether ketone) (PEEK) implants with modified surface roughness and/or surface chemistry. The roughened surface was prepared by a sandblast method, and the phosphate groups on the substrates were modified by a two-step chemical reaction. The in vitro osteogenic activity of rat mesenchymal stem cells (MSCs) on the developed substrates was assessed by measuring cell proliferation, alkaline phosphatase activity, osteocalcin expression, and bone-like nodule formation. Surface roughening alone did not improve MSC responses. However, phosphorylation of smooth substrates increased cell responses, which were further elevated in combination with surface roughening. Moreover, in a rabbit tibia implantation model, this combined surface modification significantly enhanced the bone-to-implant contact ratio and corresponding bone-to-implant bonding strength at 4 and 8 weeks post-implantation, whereas modification of surface roughness or surface chemistry alone did not. This study demonstrates that combination of surface roughness and chemical modification on PEEK significantly promotes cell responses and osseointegration ability in a synergistic manner both in vitro and in vivo. Therefore, this is a simple and promising technique for improving the poor osseointegration ability of PEEK-based orthopedic/dental implants.
Project description:<h4>Introduction</h4>Revision total knee arthoplasty often requires modular implants to treat bone defects of varying severity. In some cases, it may not be clear which module size and implant combination (e.g. sleeve and stem) should be chosen for a specific defect. When balancing implant stability and osseointegration against stress-shielding, it is important to choose an appropriate implant combination in order to match the given level of bone loss. Therefore, the necessity of stems in less extensive tibial defects and the advantage of different stems (lengths and stiffnesses) in combination with large metaphyseal sleeves on implant fixation and bone flexibility using a modular tibial revision knee system, were analyzed.<h4>Materials and methods</h4>Four different stem combinations for a tibial revision implant (Sigma TC3, DePuy) were compared to an intact bone. Standardized implantation with n = 4 synthetic tibial bones was performed after generating an Anderson Orthopaedic Research Institute (AORI) Type T1 bone defect. Axial torques around the longitudinal stem axis and varus-valgus torques were separately applied to the implant. Micromotions of bone and implant were tracked using a digital image correlation system to calculate relative micromotions at the implant-bone-interface and bone deformation.<h4>Results</h4>Overall, using stems reduced the proximal micromotions of tray and sleeve compared to no stem, while reducing bone deformation proximally at the same time, indicating some potential for proximal stress-shielding compared to no stem. The potential for increased proximal stress-shield due to reduced proximal deformation appeared to be greater when using the longer stems. The location of lowest relative micromotions was also more distal when using long stems as opposed to short stems. A short stem (especially a smaller diameter short stem which still achieves diaphyseal fixation) displayed less potential for stress-shielding, but greater bone deformation distal to the tip of the stem than in the natural model.<h4>Discussion</h4>In the case of tibial revision implants with metaphyseal sleeves in a simple fully contained Type I defect, the absence of a stem provides for more natural bone deformation. However, adding a stem reduces overall relative micromotions, while introducing some risk of proximal stress-shielding due to increased diaphyseal fixation. Increasing stem length intensifies this effect. Short stems offered a balance between reduced micromotions and more proximal bone deformation that reduced the potential for stress-shielding when compared to long stems. A short stem with slightly smaller diameter (simulating a less stiff stem which still has diaphyseal fixation) increased the proximal bone deformation, but also tended to increase the bone deformation even further at the distal stem's tip.<h4>Conclusion</h4>In conclusion, further investigation should be conducted on fully contained Type I defects and the addition of a stem to offer better initial stability, taking into account stem length (i.e. shorter or more flexible stems) to support metaphyseal fixation and allowing bending found in intact bone. In addition, further study into more extensive tibial defects is required to determine if the stability/micromotion trends observed in this study with stems and sleeves in Type I defects still apply in cases of extensive proximal bone loss.
Project description:Osteoporosis is an age-related metabolic disease that results in limited bone regeneration capacity and excessive osteoclast activity. After arthroplasty in patients with osteoporosis, poor interface osseointegration resulting from insufficient bone regeneration ability often leads to catastrophic complications such as prosthesis displacement and loosening and periprosthetic fractures. In this study, we prepared a thermosensitive hydrogel loaded with bone morphogenetic protein-2 (BMP-2) to promote osteogenesis and osteoprotegerin (OPG) to inhibit excessive osteoclast activity. To construct three-dimensional (3D)-printed composite scaffolds for implantation, a hydrogel loaded with drugs was injected into porous Ti6Al4V scaffolds. The 3D-printed composite scaffolds showed good biocompatibility and sustained release of BMP-2 and OPG for more than 20 days. <i>In vitro</i> experiments indicated that composite scaffolds promoted osteogenic differentiation and reduced the osteoclastic activation simultaneously. Remarkably, immunofluorescence staining, micro-CT, histological, and biomechanical tests demonstrated that the sustained release of both BMP-2 and OPG from composite scaffolds significantly improved bone ingrowth and osseointegration in osteoporotic defects. In conclusion, this study demonstrated that the BMP-2- and OPG-loaded 3D-printed composite scaffolds can potentially promote osseointegration for osteoporotic patients after joint replacement.
Project description:A firm connection of the bone-implant-fixation system is of utmost importance for patients with cranial defects. In order to improve the connection reliability, the current research focuses on finding the optimal fixation method, as well as selection of the implant manufacturing methods and the used materials. For the latter, implementation of bioactive materials such as hydroxyapatite or other calcium phosphates has also been considered in the literature. The aim of this study was to investigate the effect of gradual osseointegration on the biomechanical performance of cranial Ti6Al4V implants with a deposited HA coating as the osseointegration agent. This effect was assessed by two different computational approaches using finite element method (FEM) modeling. The values of key input parameters necessary for FEM were obtained from experimental plasma spray deposition of HA layers onto Ti6Al4V samples. Immediately upon implantation, the HA layer at the bone-implant contact area brought only a slight decrease in the values of von Mises stress in the implant and the micro-screws when compared to a non-coated counterpart; importantly, this was without any negative trade-off in other important characteristics. The major benefit of the HA coatings was manifested upon the modeled osseointegration: the results of both approaches confirmed a significant reduction of investigated parameters such as the total implant displacements (reduced from 0.050 mm to 0.012 mm and 0.002 mm while using Approach I and II, respectively) and stresses (reduced from 52 MPa to 10 MPa and 1 MPa) in the implanted components in comparison to non-coated variant. This is a very promising result for potential use of thermally sprayed HA coatings for cranial implants.
Project description:The aim of this study was to compare osseointegration and surface characteristics of zirconia implants made by the powder injection molding (PIM) technique against those made by the conventional milling procedure in rabbit tibiae. Surface characteristics of 2 types of implants were evaluated. Sixteen rabbits received 2 types of external hex implants with similar geometry, either machined zirconia implants or PIM zirconia implants, in the tibiae. Removal torque tests and histomorphometric analyses were performed. The roughness of the PIM zirconia implants was higher than that of machined zirconia implants. The PIM zirconia implants exhibited significantly higher bone-implant contact and removal torque values than the machined zirconia implants (p<0.001). The osseointegration of the PIM zirconia implant is promising, and PIM, using the roughened mold etching technique, can produce substantially rougher surfaces on zirconia implants.
Project description:Non-drug strategies based on biophysical stimulation have been emphasized for the treatment and prevention of musculoskeletal conditions. However, to date, an effective stimulation system for intracorporeal therapies has not been proposed. This is particularly true for active intramedullary implants that aim to optimize osseointegration. The increasing demand for these implants, particularly for hip and knee replacements, has driven the design of innovative stimulation systems that are effective in bone-implant integration. In this paper, a new cosurface-based capacitive system concept is proposed for the design of implantable devices that deliver controllable and personalized electric field stimuli to target tissues. A prototype architecture of this system was constructed for in vitro tests, and its ability to deliver controllable stimuli was numerically analyzed. Successful results were obtained for osteoblastic proliferation and differentiation in the in vitro tests. This work provides, for the first time, a design of a stimulation system that can be embedded in active implantable devices for controllable bone-implant integration and regeneration. The proposed cosurface design holds potential for the implementation of novel and innovative personalized stimulatory therapies based on the delivery of electric fields to bone cells.
Project description:Additive manufactured, porous bone implants have the potential to improve osseointegration and reduce failure rates of orthopaedic devices. Substantially porous implants are increasingly used in a number of orthopaedic applications. HA plasma spraying-a line of sight process-cannot coat the inner surfaces of substantially porous structures, whereas electrochemical deposition of calcium phosphate can fully coat the inner surfaces of porous implants for improved bioactivity, but the osseous response of different types of hydroxyapatite (HA) coatings with ionic substitutions has not been evaluated for implants in the same in vivo model. In this study, laser sintered Ti6Al4V implants with pore sizes of Ø 700 ?m and Ø 1500 ?m were electrochemically coated with HA, silicon-substituted HA (SiHA), and strontium-substituted HA (SrHA), and implanted in ovine femoral condylar defects. Implants were retrieved after 6 weeks and histological and histomorphometric evaluation were compared to electrochemically coated implants with uncoated and HA plasma sprayed controls. The HA, SiHA and SrHA coatings had Ca:P, Ca:(P+Si) and (Ca+Sr):P ratios of 1.53, 1.14 and 1.32 respectively. Electrochemically coated implants significantly promoted bone attachment to the implant surfaces of the inner pores and displayed improved osseointegration compared to uncoated scaffolds for both pore sizes (p<0.001), whereas bone ingrowth was restricted to the surface for HA plasma coated or uncoated implants. Electrochemically coated HA implants achieved the highest osseointegration, followed by SrHA coated implants, and both coatings exhibited significantly more bone growth than plasma sprayed groups (p?0.01 for all 4 cases). SiHA had significantly more osseointegration when compared against the uncoated control, but no significant difference compared with other coatings. There was no significant difference in ingrowth or osseointegration between pore sizes, and the bone-implant-contact was significantly higher in the electrochemical HA than in SiHA or SrHA. These results suggest that osseointegration is insensitive to pore size, whereas surface modification through the presence of an osteoconductive coating plays an important role in improving osseointegration, which may be critically important for extensively porous implants.