An in vivo method for the biological evaluation of metal implants

The peritoneal cavity of the rat was used as an implantation site in order to study the quantitative, cellular response and the qualitative, histopathological response to three metals (Ag, Sn, Cu). The effects of the metals on the cells were correlated with the cellular concentrations of the metal as determined by chemical analysis. Small variations in the cell population and a minimal foreign body reaction was observed with an implanted control material (silicone polymer). Large increases in the number of cells and an intense foreign body reaction was observed with Cu implants. Decreases in the number of cells were seen with Sn and Ag implants, but only Sn elicited a foreign body reaction. Implantation of Ag failed to elicit a foreign body reaction. Significant concentrations of all three metals were detected in the retrieved cells.     

Mechanical basis for bone retention around dental implants

This study, analytically, through finite element analysis, predicts the minimization of crestal bone stress resulting from implant collar surface treatment. A tapered dental implant design with (LL) and without (control, C) laser microgrooving surface treatment are evaluated. The LL implant has the same tapered body design and thread surface treatment as the C implant, but has a 2-mm wide collar that has been laser micromachined with 8 and 12 microm grooves in the lower 1.5 mm to enhance tissue attachment. In vivo animal and human studies previously demonstrated decreased crestal bone loss with the LL implant. Axial and side loading with two different collar/bone interfaces (nonbonded and bonded, to simulate the C and LL surfaces, respectively) are considered. For 80 N side load, the maximum crestal bone distortional stress around C is 91.9 MPa, while the maximum crestal bone stress around LL, 22.6 MPa, is significantly lower. Finite element analysis suggests that stress overload may be responsible for the loss of crestal bone. Attaching bone to the collar with LL is predicted to diminish this effect, benefiting crestal bone retention.     

Evaluation of the effect of adipose tissue-derived stem cells on the quality of bone healing around implants

Purpose/Aim: This study evaluates the efficacy of grafted adipose-derived stem cells (ADSCs) on blade-type implants in improving osseointegration in rat femurs using a low-densitybone model. Materials and Methods: After isolating and expanding ADSCs, twice-passaged cells were seeded on blade-type implants on culture plates. Osteogenic induction of grafted cells began after attaching cells to the prepared titanium surfaces and it continued for 4 days. The scaffolds were then implanted in the femurs of Wistar rats. Osteogenic differentiation of these cells was confirmed using polymerase chain reaction (PCR) and alizarin red staining ofthe mineralized extracellular matrix. After 8 weeks, histological and histomorphometric evaluations of undecalcified resin sections (bone–implant contact [BIC] % and bone mineral index [BMI]) were performed using light microscopy and scanning electron microscopy. Results: Alizarin red staining in conjunction with gene expression results confirmed osteogenic differentiation. Histomorphometric assessment using scanning electron microscopy demonstrated improved BIC% and BMI near the treated surface compared with the untreated surface. Conclusions: The complex of differentiated grafted ADSCs and extracellular matrix and the macrodesign and microdesign of the implant can improve osseointegration in low-density bone.     

Collagen I-coated titanium surfaces: mesenchymal cell adhesion and in vivo evaluation in trabecular bone implants

The goal of the study was the evaluation of the effect of modification of titanium implants by acrylic acid surface grafting-collagen I coupling. Tests were performed on titanium samples treated by galvanostatic anodization to create a porous surface topography. Surface characterization by X-ray photoelectron spectroscopy (XPS) and scanning electron microscopy (SEM) confirms the biochemical modification of the surface and shows a surface topography characterized by pores mostly below 1 mum diameter. In vitro evaluation involving human mesenchymal cells shows enhanced cell growth on collagen coated surfaces as compared to titanium ones. Four weeks in vivo evaluation of implants in rabbit femur trabecular bone shows improvements of bone-to-implant contact, while improvement of bone ingrowth is slightly not significant (p = 0.056), when compared to the control. Overall, these data indicate that integration in trabecular, or cancellous, bone can be enhanced by the surface collagen layer, confirming previous findings obtained by modification of machined surfaces by the same approach in cortical bone implants.     

RGD-coated titanium implants stimulate increased bone formation in vivo

Numerous studies have demonstrated that peptide modified surfaces influence short- and long-term cell responses such as attachment, shape and function in vitro. These responses are mediated via cell receptors known as integrins which bind specifically to short peptide sequences from larger proteins. Integrins transduce information to the nucleus through several cytoplasmic signalling pathways. Little is known, however, about the ability of peptide-coated surfaces to influence cell responses in vivo. The present study was designed to evaluate the quality and quantity of the new bone formed in response to titanium rods surface-coated with the peptide sequence Arg-Gly-Asp-Cys (RGDC) using gold-thiol chemistry and implanted in rat femurs. Histomorphometric analysis of cross-sections perpendicular to the implant long axis showed a significantly thicker shell of new bone formed around RGD-modified versus plain implants at 2 weeks (26.2 +/- 1.9 vs. 20.5 +/- 2.9 microm; P < 0.01). A significant increase in bone thickness for RGD implants was also observed at 4 weeks while bone surrounding controls did not change significantly in thickness (32.7 +/- 4.6 vs. 22.6 +/- 4.0 microm; P < 0.02). Mechanical pull-out testing conducted at 4 weeks revealed the average interfacial shear strength of peptide modified rods was 38% greater than control rods although this difference was not statistically significant. These pilot data suggest that an RGDC peptide coating may enhance titanium rod osseointegration in the rat femur. Long-term studies and evaluation of other peptides in larger animal models are warranted.     

Comparative bone healing near eroding polylactide-polyglycolide implants of differing crystallinity in rabbit tibial bone chambers

Eroding poly(DL-lactide-co-glycolide) (PDLLG) washers and poly(L-lactide-co-glycolide) (PLLG) threads were observed chronically in vivo following loading in a bone chamber tibial implant (BCI). Images were recorded using intravital microscopy of the implanted rabbit. Erosion and bone healing, as represented by angiogenesis and osteogenesis, was determined from changes in projected area of observed polymer, vessels and bone, respectively. Erosion rates of the two polymers were significantly different. Healing adjacent to both polymers differed significantly from controls. Healing response to each polymer was also different, with the faster eroding PDLLG causing more deviation from normal osteogenesis and angiogenesis than did PLLG. It was speculated that the faster eroding polymer released macrophage-stimulating fragments earlier in the healing process, thus altering the normal macrophage-endothelial cell interaction which in turn affected angiogenesis-linked components of osteogenesis.     

Influence of RGD-loaded titanium implants on bone formation in vivo

Little is known about the ability of peptide-coated surfaces to influence cell responses in vivo. Many studies have demonstrated that peptide-modified surfaces influence cell responses in vitro. Integrins, which bind specifically short peptide sequences, are responsible for these cell responses. In this way, information can be transmitted to the nucleus through several cytoplasmic signaling pathways. The peptide sequence Arg-Gly-Asp (RGD peptide) plays an important role in osteoblast adhesion. The present study was designed to investigate new bone formation in a porous titanium (Ti) fiber mesh implant, which was coated with cyclic RGD peptide. The RGD-Ti implants were inserted into the cranium of a rabbit and were compared with porous titanium fiber mesh disks without RGD sequence (Ti) and with an open control defect. Histologic and histomorphometric examinations were performed 2, 4, and 8 weeks postoperatively. A significant increase in bone formation, or bone ingrowth, was seen in the RGD-Ti group compared with the Ti group after 4 and 8 weeks. All control defects stayed open in all three periods. It was concluded that the use of cyclic RGD peptide in combination with titanium fiber mesh has a positive effect on bone formation in vivo in a rabbit animal model.     

Recurrent bone regeneration in titanium implants. Experimental model for determining the healing capacity of bone using quantitative microradiography

An experimental technique for quantitative assessment of bone repair was tested. The recurrent regeneration capacity of cortical bone was analyzed in consecutive 3 wk periods, using an osseointegrated titanium implant, the Bone Harvest Chamber (BHC), in the proximal tibial metaphysis of the rabbit. The BHC is a divisible implant penetrated by a canal into which newly formed bone tissue will grow during a 3 wk healing period. The newly formed bone tissue may easily be collected (harvested) without the animal being killed. After 3 wk, bone tissue can again be harvested, in principle, indefinitely. Intact harvested specimens were quantified by microradiography-videodensitometry, yielding a total specimen mass in mg aluminium equivalent. This unit correlated very well to a specimen dry weight (r = 0.996) and an average mineralized bone density (r = 0.937). The specimens were also examined by correlative histology. Three weeks after implant insertion, the chambers had become integrated in the bone tissue but the average bone mass varied widely, influenced by the surgical insertion trauma. Six weeks after insertion, the greatest average bone mass was found, indicating an intense ongoing osseointegration. The amount of bone regenerated at later harvests was fairly equal, indicating a stabilization of the implant bed to the repeat stimulus, i.e. harvesting. Bone regeneration differed significantly between animals, but also intraindividual variations, i.e. different amounts of bone formed in the same chamber, were observed.     

Composite implants for orthopedic applications: in vivo evaluation of candidate resins.

Three resins which include poly (methyl methacrylate), surgical Simplex P, and ultrahigh molecular weight polyethylene were selected to be evaluated as candidate systems for a polymer based composite for hard tissue prosthesis. Characterization of the mechanical behavior of these polymers in different environments including in vitro and in vivo storage was accomplished. As a result, conclusions were made as to which material maintained the least amount of mechanical variance as influenced by environmental effects. In vivo studies of implanted materials provided for a study of soft tissue response to each material. Conclusions were then developed as to the varying degrees of tissue reaction initiated by each material, and which resin generated the greatest tissue response with respect to the others.     

生物可吸收材料在骨折内固定治疗的临床研究

目的评价生物可吸收材料在骨折内固定治疗中的作用。方法对713例不同部位骨折采用生物可吸收材料进行内固定。结果随访3个月~4年,平均18个月,骨折全部临床愈合;功能评估优良率96.5%,并发症发生率4.3%,均为异物反应。结论生物可吸收材料是治疗骨折的有效方法,正确选择适应症和手术操作是治疗成功的重要因素。     

In vivo evaluation of calcium polyphosphate for bone regeneration

Current problems associated with bone allografts include risk of disease transmission, limited availability, and cost. Synthetic scaffolds have been proposed as substitute graft materials to address these issues. Calcium polyphosphate is a novel synthetic scaffold material that has shown good mechanical properties and biocompatibility. Here, we evaluated calcium polyphosphate in terms of its ability to support cell proliferation and differentiation in?vivo. Calcium polyphosphate, morsellized cancellous bone, and hydroxyapatite/tricalcium phosphate particles were seeded with marrow stromal cells and implanted subcutaneously in the back of NOD/Scid mice. At 7, 14, and 28 days the samples were harvested and the proliferation characteristics and gene expression were analyzed. All tested graft materials had similar proliferation characteristics and gene expression. The subcutaneous environment had a stronger impact on the proliferation and differentiation of the cells than the scaffold material itself. However, it was shown that calcium polyphosphate is superior to hydroxyapatite/tricalcium phosphate and bone in its ability to support cell survival in?vivo. The study confirmed that calcium polyphosphate has potential for replacing morsellized cancellous bone as a graft material for bone regeneration.     

A new method of coloring ground bone implants for light microscopy

The material exploration for implants close in also histological study of bone-proofs after implantation in test animals. For light microscopical utilization of implant-bone-polishings was developed a new method of colouring what shows a better distinction between types of tissue around the implant and a time economizing of 3 h.     

Increased bone remodelling around titanium implants coated with chondroitin sulfate in ovariectomized rats

Coating titanium implants with artificial extracellular matrices based on collagen and chondroitin sulfate (CS) has been shown to enhance bone remodelling and de novo bone formation in vivo. The aim of this study was to evaluate the effect of estrogen deficiency and hormone replacement therapy (HRT) on the osseointegration of CS-modified Ti implants. 30 adult female, ovariectomized Wistar rats were fed either with an ethinyl-estradiol-rich diet (E) to simulate a clinical relevant HRT or with a genistein-rich diet (G) to test an alternative therapy based on nutritionally relevant phytoestrogens. Controls (C) received an estrogen-free diet. Uncoated titanium pins (Ti) or pins coated with type-I collagen and CS (Ti/CS) were inserted 8 weeks after ovarectomy into the tibia. Specimens were retrieved 28 days after implantation. Both the amount of newly formed bone and the affinity index (P < 0.05) were moderately higher around Ti/CS implants as compared to uncoated Ti. The highest values were measured in the G-Ti/CS and E-Ti/CS groups, the lowest values for the E-Ti and G-Ti controls. Quantitative synchrotron radiation micro-computed tomography (SRμCT) revealed the highest increase in total bone formation around G-Ti/CS as compared to C-Ti (P < 0.01). The effects with respect to direct bone apposition were less pronounced with SRμCT. Using scanning nanoindentation, both the indentation modulus and the hardness of the newly formed bone were highest in the E-Ti/CS, G-Ti/CS and G-Ti groups as compared to C-Ti (P < 0.05). Coatings with collagen and CS appear to improve both the quantity and quality of bone formed around Ti implants in ovarectomized rats. A simultaneous ethinyl estradiol- and genistein-rich diet seems to enhance these effects.     

Primary cilia act as mechanosensors during bone healing around an implant

The primary cilium is an organelle that senses cues in a cell's local environment. Some of these cues constitute molecular signals; here, we investigate the extent to which primary cilia can also sense mechanical stimuli. We used a conditional approach to delete Kif3a in pre-osteoblasts and then employed a motion device that generated a spatial distribution of strain around an intra-osseous implant positioned in the mouse tibia. We correlated interfacial strain fields with cell behaviors ranging from proliferation through all stages of osteogenic differentiation. We found that peri-implant cells in the Col1Cre;Kif3a^fl/fl mice were unable to proliferate in response to a mechanical stimulus, failed to deposit and then orient collagen fibers to the strain fields caused by implant displacement, and failed to differentiate into bone-forming osteoblasts. Collectively, these data demonstrate that the lack of a functioning primary cilium blunts the normal response of a cell to a defined mechanical stimulus. The ability to manipulate the genetic background of peri-implant cells within the context of a whole, living tissue provides a rare opportunity to explore mechanotransduction from a multi-scale perspective.     

Collagen type I increases bone remodelling around hydroxyapatite implants in the rat tibia

The early interface reaction of cancellous bone to a nanocrystalline hydroxyapatite (HA) cement containing 3 wt% collagen type I (HA/Coll) with a setting under physiological temperature and pH was observed using immunohistochemical techniques. Pure HA served as a control. Cylinders with a diameter of 2 mm were implanted into the proximal tibia of 72 adult Wistar rats. Histological sections of 6 animals were prepared after 1, 2, 4, 6, 14 and 28 days. First, osteoblast-like cells as well as a marked reaction for osteonectin, osteopontin and its ligand CD44 were observed as early as 2 days after implantation at the interface around HA/Coll implants. Further, reactivity for ED1 and cathepsin D, both markers for phagocytotic cells, appeared earlier and stronger around HA/Coll. In cell counts, a significantly higher average number of ED1- and cathepsin D-positive phagocytotic cells was observed around the HA/Coll implants on days 6 (p < 0.01), 14 and 28 (p < 0.05). The number of osteopontin-positive cells was significantly higher around HA/Coll implants at days 6 and 14 (p < 0.05). Two weeks after the implantation, first islands of newly formed woven bone were observed around the HA/Coll implant, but not around the control implant. The amount of direct bone contact after 28 days averaged 28% around pure HA and 51% around HA/Coll implants (p < 0.05). While both implants displayed a good osteoconductivity, a higher bone remodelling activity was observed around collagen-containing HA implants compared to pure HA implants. It appears that the addition of collagen to HA implants can enhance both phagocytotic and osteogenic processes. This may result in an earlier acceptance and better osseointegration of the HA/Coll implants into the surrounding tissue.     

The interface zone of inorganic implantsIn vivo: Titanium implants in bone

The interface zone between titanium implants and bone is considered at the macroscopic, microscopic, and molecular levels. A high rate of successful dental implants of pure titanium is associated with a very close apposition of the bone to the titanium surface, called osseointegration. At the macroscopic level, osseointegration allows efficient stress transfer from the implant to the bone without abrasion or progressive movement that can take place if a fibrous layer intervenes. At the microscopic level, surface roughness and porosity provide interlocking of the implant and bone tissue which grows into direct contact with titanium. Sections studied in the electron microscope show that calcified tissue can be identified within 50 Å of the implant surface. The interface zone includes a tightly adherent titanium oxide layer on the surface of the implant which may be similar to a ceramic material in relation to tissue response. The five year success rate of 90% in 2895 implants in clinical trials since 1965 is associated with the favorable behavior of bone tissue at the interface zone with pure titanium.     

Polymers for healing caps on titanium implants

Healing caps are used during the healing period after abutment connection on titanium implants of the Branemark type. These healing caps consist of a screw embedded in polyamide-6 (PA-6) which is considerably weakened during this healing period. The properties of PA-6 and poly(4-methyl-1-pentene) (PMP) were studied. Storage in water at 37 degrees C led to a marked reduction in the flexural modulus of PA-6, but only a minor decrease for PMP. PA-6 showed a marked increase in volume due to water uptake, whereas PMP showed no significant change in volume. The improved properties of PMP compared to PA-6 when used for healing caps were demonstrated. The results of an agar overlay cytotoxicity test indicated that both PMP and PA-6 were nontoxic under laboratory controlled conditions.     

Novel thermosensitive chitosan hydrogels: in vivo evaluation

Chitosan is an attractive biopolymer for the preparation of hydrogels. Its unique combination of biocompatibility, biodegradability, bioadhesivity, and tissue-promoting abilities allows pharmaceutical applications. We investigated novel thermosensitive hydrogels based on chitosan homogeneously reacetylated to a deacetylation degree of about 50%, combined with selected polyols or polyoses such as trehalose, a nontoxic polysaccharide. The latter, a gel-inducing and lyoprotective agent enabled the formulation to be lyophilized and rehydrated without affecting the thermosensitive behavior. This made possible long-term storage and promoted its use in a clinical setup. The thermally induced sol-gel transition allowed injectability and in situ setting. Rheological characterization revealed that storage moduli could be increased by one decade by increasing the chitosan concentration from 1.4 to 2.2% (w/w). Evaluation in vivo provided evidence of in situ implant formation in subcutaneous tissue of Sprague-Dawley rats and permanence for up to 3 months. Histopathological analysis demonstrated a mild, chronic, inflammatory reaction that disappeared with the complete absorption of the gel implant over a few months period. Such in situ forming hydrogels could be advantageous for specific applications in drug delivery and tissue engineering.