Tissue response to microfibers of different polymers: polyester,polyethylene, polylactic acid,and polyurethane

Tissue response to single polymer microfibers of polyester (PET), polyethylene (PE), poly(L-lactic acid) (PLA), and polyurethane (PU) was assessed using a rat subcutaneous model. Fibers of diameters ranging from 1 to 15 microm were aligned parallel to each other on polycarbonate frames and implanted in the subcutaneous dorsum in the subscapular region. After 5 weeks of implantation, fibrous capsule thickness was significantly less for fibers of diameters 1-5 than for those of 11-15 microm for all polymers tested. For PET and PU, 75.0 and 71.4% respectively of the 1-5 microm fibers had no capsule, while for PE and PLA only 45.5 and 56.3% respectively had no capsule. For 1-5 microm fibers, PE had significantly thicker capsules than PET and PU. Reducing fiber diameters from 6-10 to 1-5 microm induced a greater reduction in capsule thickness than changing polymers among PET, PE, and PLA. PU showed the least encapsulation of all polymers, demonstrating significantly thinner capsules than PET, PE, and PLA for 6-10 and 11-15 microm fibers.     

Relative influence of polymer fiber diameter and surface charge on fibrous capsule thickness and vessel density for single-fiber implants

Single polypropylene microfibers plasma-coated with polymers of different surface charge [N,N-dimethylaminoethyl methacrylate (NN) (positive charge), methacrylic acid (MA) (negative charge), and hexafluoropropylene (HF) (neutral)] were implanted in the subcutaneous dorsum of Sprague-Dawley rats for 5-week intervals. Thee groups of fiber diameters were used: (I) 1.0 to 5.9 microm; (II) 6.0 to 10.9 microm; and (III) 11.0 to 15.9 microm. Fibrous capsule thickness and blood-vessel density (number of vessels within 100 microm of the fiber) were assessed in tissue sections in the planes of microfiber cross-sections. Results from a multifactorial analysis of variance demonstrated statistically significant main effects (p < 0.05) for microfiber diameter but not for surface-charge coating. The mean differences in capsule thickness among the microfiber diameter groups were: between groups II and I: 5.4 microm; between groups III and I: 10.2 microm; and between groups III and II: 4.7 microm. The mean differences in capsule thickness among surface-charge coatings were: between MA and NN: 0.7 microm; between MA and HF: 1.4 microm; and between NN and HF: 0.7 microm. Many of the 1.0 to 5.9 microm-in-diameter fibers had no capsule and no sign of a foreign-body reaction. For the vessel density analysis, neither microfiber diameter nor surface-charge coating had a statistically significant effect. Thus the geometric feature of microfiber diameter was more important than was surface charge relative to fibrous capsule formation but not relative to local vessel density. This ranking of the relative influence of design features in relation to tissue response provides useful information for prioritization in biomaterial design.     

Polymer microfiber mechanical properties: a system for assessment and investigation of the link with fibrous capsule formation

A novel microtensile testing instrument was developed to assess the mechanical properties of small-diameter polyethylene, polyurethane, and polyester microfibers. The instrument had a root-mean-square error of 2.96 microN for force measurement and 1.91 microm for displacement measurement. Microfibers ranging in diameter from 1.0 to 10.9 microm were strained at 2 mm/s in the device, and the slopes of their stress-strain curves (material moduli) were determined. Correlations between material modulus and previously published data on fibrous capsule presence and thickness for implanted polyethylene, polyurethane, and polyester microfibers were investigated. Results for the 1.0-5.9-microm microfiber diameter range showed that neither the percentage of unencapsulated fibers nor the capsule thickness correlated well with modulus. Correlation coefficients were 0.04 and 0.09, respectively. However, for the 6.0-10.9 microm diameter range the correlations were strong, 1.00 for both percentage of unencapsulated fibers and capsule thickness. It is suggested that the results reflect the greater attachment and mechanical interaction of cells with microfibers for the 6.0-10.9 microm-diameter range than for the 1.0-5.9 microm-diameter range.     

Fibrous encapsulation of single polymer microfibers depends on their vertical dimension in subcutaneous tissue

The purpose of this research was to investigate possible explanations for why small-diameter microfiber implants do not experience encapsulation in subcutaneous tissue as do large-diameter fiber implants. Single polypropylene microfibers of approximately rectangular cross-section with rounded edges were twisted about their longitudinal axes and affixed at their ends to polycarbonate frames. The frames were implanted in rat subcutaneous dorsum for a 5-week period, then removed and processed for light microscopy analysis. Fibrous capsule presence/absence and thickness around the implants were assessed, and their relationships to geometric features of the fibers investigated. A logistic regression analysis between presence/absence of a fibrous capsule and geometric features of interest demonstrated strong predictive ability (92.4% correct predictions) for implant height and a well-defined threshold separating the presence and absence of a fibrous capsule at 5.9 microm (p < 0.001). Implant height was defined as the vertical distance between the most superficial and deepest level of the implant. This 5.9-microm threshold value of implant height is comparable to the 6.0-microm diameter threshold for capsule presence/absence in fibers of circular cross-section [Sanders et al. J Biomed Mater Res 2000; 52(1):231-237]. Fiber major axis length, minor axis length, aspect ratio, surface area per unit length, implant width, and implant angle did not show similar predictive ability or a well-defined threshold separating the presence and absence of a fibrous capsule. It is reasoned that for fibers greater than the threshold height of 5.9 microm, separation of collagen fibers in the extracellular matrix creates dead space regions adjacent to the fibers that attract inflammatory cells and stimulate fibrous capsule formation.     

Fall in the number of intracardiac neurons in aging rats

The neurons of whole cardiac atria were stained using a NADH-diaphorase technique in young adult (3 months old) (GI) and aging rats (20 months old) (GII). Light microscopy revealed differences in the appearance of the neurons in the two groups. In GI, most ganglia contained 50-100 neurons while in GII, most ganglia usually contained 20 neurons. The mean total number of neurons in the atria of GII was 245+/-31, i.e. only 23% of the mean value in GI (1086+/-203). The mean size of the ganglionic neurons (area of maximum cell profile) was 702 microm2 in GI and 1065 microm2 in GII. Histological sections of the ganglia revealed that a capsule of collagen fibers sheaths each ganglion in both groups. In GII, the density of collagen fibers increases in the capsule and in the septa within the ganglia; yellow or red, type I collagen fibers predominate in this group. No elastic fibers were present in the cardiac ganglia of either group. It is suggested that in aging rats, structural changes and reorganization of the remnant neurons accompany neuron reduction.     

In vitro degradation of porous poly(L-lactic acid) foams

This study investigated the in vitro degradation of porous poly(L-lactic acid) (PLLA) foams during a 46-week period in pH 7.4 phosphate-buffered saline at 37 degrees C. Four types of PLLA foams were fabricated using a solvent-casting, particulate-leaching technique. The three types had initial salt weight fraction of 70, 80, and 90%, and a salt particle size of 106-150 microm, while the fourth type had 90% initial weight fraction of salt in the size range 0-53 microm. The porosities of the resulting foams were 0.67, 0.79, 0.91, and 0.84, respectively. The corresponding median pore diameters were 33, 52, 91, and 34 microm. The macroscopic degradation of PLLA foams was independent of pore morphology with insignificant variation in foam weight, thickness, pore distribution, compressive creep behavior, and morphology during degradation. However, decrease in melting temperature and slight increase in crystallinity were observed at the end of degradation. The foam half-lives based on the weight average molecular weight were 11.6+/-0.7 (70%, 106-150 microm), 15.8+/-1.2 (80%, 106-150 microm), 21.5+/-1.5 (90%, 106-150 microm), and 43.0+/-2.7 (90%, 0-53 microm) weeks. The thicker pore walls of foams prepared with 70 or 80% salt weight fraction as compared to those with 90% salt weight fraction contributed to an autocatalytic effect resulting in faster foam degradation. Also, the increased pore surface/volume ratio of foams prepared with salt in the range 0-53 microm enhanced the release of degradation products thus diminishing the autocatalytic effect and resulting in slower foam degradation compared to those with salt in the range 106-150 microm. Formation and release of crystalline PLLA particulates occurred for foams fabricated with 90% salt weight fraction at early stages of degradation. These results suggest that the degradation rate of porous foams can be engineered by varying the pore wall thickness and pore surface/volume ratio.     

Small fiber diameter fibro-porous meshes: tissue response sensitivity to fiber spacing

The purpose of this research was to determine if fiber spacing for small fiber diameter fibro-porous meshes affected tissue response in vivo. Disk-shaped polyurethane meshes, with mean fiber diameters of 7.6 microm and fiber spacing between 6 and 68 microm, were implanted in rat subcutaneous dorsum for 5-week intervals and then prepared for light microscopy and morphological analysis. Results showed that implants with 12- to 68-microm spacing had no histologically apparent fibrous capsule around the perimeter, a result different from that for 6-microm spacing samples that had a capsule around a mean of 34.2% of the perimeter. For the 12- to 68-microm spacing range, a mean of 21.0% of individual fibers within the meshes were encapsulated. Qualitatively, it appeared that larger fibers were encapsulated more frequently than smaller ones. When nodeless or baggy meshes were implanted, cells tended to cluster three or more fibers into groups and then encapsulate each group. Over the 6- to 68-microm spacing range, cell nuclei volume fraction within the meshes increased from the 6- to the 29-microm spacing (p = 0.000) and then decreased from the 29- to the 68-microm spacing (p = 0.015). There was a trend of an increase in local vessel volume fraction with spacing over the 6- to 68-microm range, though the relationship was weak. The results indicate that the reason for the lack of encapsulation of small-fiber fibro-porous meshes is not exclusively a pore boundary explanation, as is proposed for small-pore porous meshes.     

Development of mammalian cell-enclosing subsieve-size agarose capsules (<100 microm) for cell therapy

Agarose capsules were prepared using a droplet breakup method in a coflowing stream. Subsieve-size capsules 76+/-9 microm in diameter were obtained by extruding 4 wt% agarose solution from a needle (300 microm inner diameter) at a velocity of 1.2 cm/s into an ambient liquid paraffin flow of 20.8 cm/s. Increasing the flow rate of the liquid paraffin and decreasing that of the agarose solution resulted in a decreased resultant capsule diameter. Reduction in diameter from several hundred micrometers to subsieve-size (<100 microm) enhanced molecular exchange and mechanical stability. Measurements based on the percentage of intact mitochondria in the cells demonstrated that the viability of the enclosed cells was independent of capsule diameter. No significant difference was observed between the viabilities of cells enclosed in capsules with diameters of 79+/-8 and 351+/-41 microm (p=0.43). Compared with cells seeded in a tissue culture dish, the cells enclosed in the subsieve-size capsules showed 89.2% viability.     

Histological biocompatibility of new, non-absorbable glaucoma deep sclerectomy implant

We performed this study to compare the intrascleral biocompatibility of three materials: non-absorbable hydrogel contact lens polymer, non-absorbable silicone rubber, and absorbable cross-linked sodium hyaluronate. Intrascleral implantation of three different materials was performed in 13 healthy, pigmented rabbits. Implants of methacrylic hydrogel, silicone rubber, and cross-linked sodium hyaluronate were implanted in 10, 8, and 8 eyes, respectively. The animals were euthanized at 7, 30, 180, and 360 days post implantation. The eyes were enucleated and immediately fixed in 10% buffered formalin. Semithin sections were cut and stained with hematoxylin-eosin. Light microscope analysis of the specimens was performed. The least severe inflammatory reaction was observed with cross-linked sodium hyaluronate implants. The number of inflammatory cells in proximity to methacrylic hydrogel and silicone implants at all periods of follow up was similar. The thickest fibrous capsule was observed with silicone implants (average, 28.38 +/- 11.17 microm). This area was thinner with methacrylic hydrogel implants (average, 14.90 +/- 5.57 microm) and was thinnest around sodium hyaluronate implants (average, 7.21 +/- 2.33 microm). For each type of implant, the wall on the conjunctival side of the fibrous capsule was significantly thicker than the wall on the choiroidal side. The space between the implant, scleral flap, and bed was filled soon after surgery with connective tissue rich in vessels. In our study, cross-linked sodium hyaluronate had the highest intrascleral biocompatibility. Although the inflammatory responses of the sclera to methacrylic hydrogel and silicone rubber were similar in nature, a thicker fibrous capsule was generated around silicone implants.     

Structural characterization and cell response evaluation of electrospun PCL membranes: micrometric versus submicrometric fibers

Electrospinning is a valuable technique to fabricate fibrous scaffolds for tissue engineering. The typical nonwoven architecture allows cell adhesion and proliferation, and supports diffusion of nutrients and waste products. Poly(epsilon-caprolactone) (PCL) electrospun membranes were produced starting from 14% w/v solutions in (a) mixture 1:1 tetrahydrofuran and N,N-dimethylformamide and (b) chloroform. Matrices made up of randomly arranged uniform fibers free of beads were obtained. The average fiber diameters were (a) 0.8 +/- 0.2 microm and (b) 3.6 +/- 0.8 microm. PCL matrices showed the following tensile mechanical properties: tensile modulus (a) 5.0 +/- 0.7 MPa (b) 6.4 +/- 0.2 MPa, yield stress (a) 0.55 +/- 0.06 MPa (b) 0.43 +/- 0.02 MPa, and ultimate tensile stress (a) 1.7 +/- 0.2 MPa and (b) 0.8 +/- 0.1 MPa. The ultimate strain ranged between 300% and 400%. Cytotoxicity of electrospun membranes was continuously evaluated by means of electric cell-substrate impedance sensing technique using human umbilical vein endothelial cells (HUVEC). PCL matrices resulted free of toxic amounts of contaminants and/or process by-products. In vitro studies performed by culturing HUVEC on micrometric and submicrometric fibrous mats showed that both structures supported cell adhesion and spreading. However, cells cultured on the micrometric network showed higher vitality and improved interaction with the polymeric fibers, suggesting an increased ability to promote cell colonization.     

S-100-positive nerve fibers in hepatocellular carcinoma and intrahepatic cholangiocarcinoma: an immunohistochemical study

The aim of the present study was to determine whether or not liver carcinomas are innervated, since there have been no previous reports on the distribution of nerve fibers in human hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC). We investigated nerve fibers by immunohistochemical and morphometric methods in 63 cases of HCC and 28 cases of ICC. An antibody to S-100 protein was used to visualize nerve fibers. In HCC, S-100-positive nerve fibers were absent in the tumoral region, including the sinusoids, and tumoral fibrous septa, while in the capsule of HCC some S100-positive nerve fibers (density: 0.08 +/- 0.03/mm2) were present in contact with vasculatures. In ICC, a few nerve fibers were noted in the tumoral stroma (density: 0.02 +/- 0.01/mm2). No nerve fibers were seen in the neovasculized vessels (tumor vessels) in both HCC and ICC. In invasive regions of HCC and ICC, there were S-100-positive nerve fibers in pre-existing residual portal tracts (density: HCC, 0.11 +/- 0.03/mm2; ICC, 0.13 +/- 0.04/mm2). In non-tumor regions of HCC and ICC, there were many S100-positive nerve fibers (density: 0.41 +/- 0.13/mm2) in portal tracts and, to a much lesser degree, in the sinusoids. These results suggest that tumor cells and vasculatures in HCC and ICC are rarely influenced by nerve fibers.     

Histomorphometric analysis of poly-L/D-lactide 96/4 sutures in the gastrocnemius tendon of rabbits

BACKGROUND: Common Achilles tendon ruptures are not usually fixed by bioabsorbable sutures due to limitations in their strength retention properties. Modern technology has made it possible to develop bioabsorbable sutures with prolonged strength retention. Aims: To evaluate histologically tissue reactions of poly-L/D-lactide (PLDLA) sutures implanted in Achilles tendon of rabbits. MATERIAL AND METHODS: Fifteen rabbits were evaluated at 2, 6 and 12 weeks postoperatively, with five rabbits in each follow-up group. PLDLA monofilament sutures were implanted into the medial gastrocnemius tendon. Polyglyconate monofilament sutures with similar diameter (Maxon 4-0, Cyanamid of Great Britain Ltd., Gosport, UK) were implanted in the contralateral gastrocnemius tendon. The histology was studied in hard-resin embedded samples. The thickness of the formed fibrous tissue capsule was determined histomorphometrically. RESULTS: PLDLA led to formation of significantly thinner fibrous tissue capsule than Maxon sutures of the same diameter. Median thickness (PLDLA vs. Maxon) at two weeks was 5.26 vs.13.22 microm, at six weeks 11.66 vs. 80.97 microm, and at 12 weeks 10.63 vs. 17.59 microm (p<0.01). CONCLUSIONS: During the 12 week follow-up period, PLDLA sutures implanted intratendineously formed thinner fibrous capsule than Maxon sutures of the same diameter. The suture materials were not totally absorbed by 12 weeks.     

Temporal and spatial distribution of macrophage phenotype markers in the foreign body response to glutaraldehyde-crosslinked gelatin hydrogels

Currently, it is not well understood how changes in biomaterial properties affect the foreign body response (FBR) or macrophage behavior. Because failed attempts at biomaterial degradation by macrophages have been linked to frustrated phagocytosis, a defining feature of the FBR, we hypothesized that increased hydrogel crosslinking density (and decreased degradability) would exacerbate the FBR. Gelatin hydrogels were crosslinked with glutaraldehyde (0.05, 0.1, and 0.3%) and implanted subcutaneously in C57BL/6 mice over the course of 3 weeks. Interestingly, changes in hydrogel crosslinking did not affect the thickness of the fibrous capsule surrounding the hydrogels, expression of the pan-macrophage marker F480, expression of three macrophage phenotype markers (iNOS, Arg1, CD163), or expression of the myofibroblast marker aSMA, determined using semi-quantitative immunohistochemical analysis. With respect to temporal changes, the level of expression of the M1 marker (iNOS) remained relatively constant throughout the study, while the M2 markers Arg1 and CD163 increased over time. Expression of these M2 markers was highly correlated with fibrous capsule thickness. Differences in spatial distribution of staining also were noted, with the strongest staining for iNOS at the hydrogel surface and increasing expression of the myofibroblast marker aSMA toward the outer edge of the fibrous capsule. These results confirm previous reports that macrophages in the FBR exhibit characteristics of both M1 and M2 phenotypes. Understanding the effects (or lack of effects) of biomaterial properties on the FBR and macrophage phenotype may aid in the rational design of biomaterials to integrate with surrounding tissue.     

Comparison of histometric data obtained by optical coherence tomography and routine histology

There is a lack of systematic investigations comparing optical coherence tomography (OCT) with histology. OCT assessments were performed on the upper back of 16 healthy subjects. Epidermis thickness (ET) was assessed using three methods: first, peak-to-valley analysis of the A-scan (ET-OCT-V); second, manual measurements in the OCT images (ET-OCT-M); third, light microscopic determination using routine histology (ET-Histo). The relationship between the different methods was assessed by means of the Pearson correlation procedure and Bland and Altman plots. We observed a strong correlation between ET-Histo (79.4+/-21.9 microm) and ET-OCT-V (79.2+/-15.5 microm, r=0.77) and ET-OCT-M (82.9+/-15.8 microm, r=0.75), respectively. Bland and Altman plots revealed a bias of -0.19 microm (95% limits of agreement: -27.94 microm to 27.56 microm) for ET-OCT-V versus ET-Histo and a bias of 3.44 microm (95% limits of agreement: -24.9 microm to 31.78 microm) for ET-OCT-M versus ET-Histo. Despite the strong correlation and low bias observed, the 95% limits of agreement demonstrated an unsatisfactory numerical agreement between the two OCT methods and routine histology indicating that these methods cannot be employed interchangeably. Regarding practicability, precision, and indication spectrum, ET-OCT-V and ET-OCT-M are of different clinical value.     

In vitro and in vivo degradation of porous poly(DL-lactic-co-glycolic acid) foams

This study investigated the in vitro degradation of porous poly(DL-lactic-co-glycolic acid) (PLGA) foams during a 20-week period in pH 7.4 phosphate-buffered saline (PBS) at 37 degrees C and their in vivo degradation following implantation in rat mesentery for up to 8 weeks. Three types of PLGA 85 : 15 and three types of 50 : 50 foams were fabricated using a solvent-casting, particulate-leaching technique. The two types had initial salt weight fraction of 80 and 90%, and a salt particle size of 106-150 microm, while the third type had 90% initial weight fraction of salt in the size range 0-53 microm. The porosities of the resulting foams were 0.82, 0.89, and 0.85 for PLGA 85 : 15, and 0.73, 0.87, and 0.84 for PLGA 50 : 50 foams, respectively. The corresponding median pore diameters were 30, 50, and 17 microm for PLGA 85: 15, and 19, 17, and 17 microm for PLGA 50 : 50. The in vitro and in vivo degradation kinetics of PLGA 85: 15 foams were independent of pore morphology with insignificant variation in foam weight, thickness, pore distribution, compressive creep behavior, and morphology during degradation. The in vitro foam half-lives based on the weight average molecular weight were 11.1 +/- 1.8 (80%, 106-150 microm), 12.0 +/- 2.0 (90%, 106-150 microm), and 11.6 +/- 1.3 (90%, 0-53 microm) weeks, similar to the corresponding values of 9.4 +/- 2.2, 14.3 +/- 1.5, and 13.7 +/- 3.3 weeks for in vivo degradation. In contrast, all PLGA 50 : 50 foams exhibited significant change in foam weight, water absorption, and pore distribution after 6-8 weeks of incubation with PBS. The in vitro foam half-lives were 3.3 +/- 0.3 (80%, 106-150 microm), 3.0 +/- 0.3 (90%, 106-150 microm), and 3.2 +/- 0.1 (90%, 0-53 microm) weeks, and the corresponding in vivo half-lives were 1.9 micro 0.1, 2.2 +/- 0.2, and 2.4 +/- 0.2 weeks. The significantly shorter half-lives of PLGA 50: 50 compared to 85: 15 foams indicated their faster degradation both in vitro and in vivo. In addition, PLGA 50: 50 foams exhibited significantly faster degradation in vivo as compared to in vitro conditions due to an autocatalytic effect of the accumulated acidic degradation products in the medium surrounding the implants. These results suggest that the polymer composition and environmental conditions have significant effects on the degradation rate of porous PLGA foams.     

Fibro-porous meshes made from polyurethane micro-fibers: effects of surface charge on tissue response

The purpose of this research was to evaluate the influence of surface charge on fibrous encapsulation, cell nuclei density, and vessel ingrowth into small-fiber, fibro-porous, biomaterial meshes. Meshes electrospun from polyurethane with mean fiber diameters of 5.8 microm and mean fiber spacing of 64.9 microm were plasma coated with films of different relative surface charge: Hexafluoropropylene (HF) (neutral), N,N-dimethylaminoethyl methacrylate (NN) (positive charge), and methacrylic acid (MA) (negative charge). Samples were implanted in rat subcutaneous dorsum for 5 weeks then fibrous capsule presence around the implants, cell nuclei density, and vessel number were assessed. Results showed that within the resolution of the histological analysis methods used, no implant experienced fibrous encapsulation. There was no significant difference between cell nuclei density and coating for the four groups: uncoated, HF-coated, NN-coated, and MA-coated. HF-coated and NN-coated samples had lower vessel numbers than uncoated samples (p = 0.055 and 0.032, respectively). MA-coated samples had vessel numbers not significantly different from uncoated polyurethane (slightly negatively charged) samples (p = 0.879). The results suggest that negatively charged surfaces may facilitate vessel ingrowth into fibro-porous mesh biomaterials.     

Effect of fiber diameter on spreading, proliferation, and differentiation of osteoblastic cells on electrospun poly(lactic acid) substrates

Electrospinning is a promising method to construct fused-fiber biomaterial scaffolds for tissue engineering applications, but the efficacy of this approach depends on how substrate topography affects cell function. Previously, it has been shown that linear, parallel raised features with length scales of 0.5-2 microm direct cell orientation through the phenomenon of contact guidance, and enhance phenotypic markers of osteoblastic differentiation. To determine how the linear, random raised features produced by electrospinning affect proliferation and differentiation of osteoprogenitor cells, poly(lactic acid) and poly(ethylene glycol)-poly(lactic acid) diblock copolymers were electrospun with mean fiber diameters of 0.14-2.1 microm onto rigid supports. MC3T3-E1 osteoprogenitor cells cultured on fiber surfaces in the absence of osteogenic factors exhibited a lower cell density after 7 and 14 days of culture than cells cultured on spin-coated surfaces, but cell density increased with fiber diameter. However, in the presence of osteogenic factors (2 mM beta-glycerophosphate, 0.13 mM L-ascorbate-2-phosphate), cell density after 7 and 14 days of culture on fiber surfaces was comparable to or exceeded spin-coated controls, and alkaline phosphatase activity after 14 days was comparable. Examination of cell morphology revealed that cells grown on fibers had smaller projected areas than those on planar surfaces. However, cells attached to electrospun substrates of 2.1 microm diameter fibers exhibited a higher cell aspect ratio than cells on smooth surfaces. These studies show that topographical factors designed into biomaterial scaffolds can regulate spreading, orientation, and proliferation of osteoblastic cells.     

Osteointegration of femoral stem prostheses with a bilayered calcium phosphate coating

Our purpose was to evaluate the osteointegration of bilayered calcium phosphate (CaP)-coated femoral hip stems in a canine model. A first layer of hydroxyapatite (HA) 20 microm thick and a superficial layer of Biphasic Calcium Phosphate (BCP) 30 microm thick were plasma-sprayed on to the proximal region of sandblasted Ti6Al4V prostheses. Bilayered CaP-coated and non-coated canine femoral stems were implanted bilaterally under general anesthesia in 6 adult female Beagle dogs. After 6 and 12 months, a significant degradation of the bilayered coating occurred with a remainder of 33.1+/-12.4 and 23.6+/-9.2 microm in thickness, respectively. Lamellar bone apposition was observed on bilayered coated implants while fibrous tissue encapsulation was observed on non-coated femoral stems. The bone-implant contacts (BIC) were 91+/-3% and 81+/-8% for coated and 7+/-8% and 8+/-12% for non-coated implants, at 6 and 12 months, respectively. Our study supports the concept of a direct relationship between the biodegradation of CaP coating and the enhanced osteointegration of titanium prostheses. A bilayered CaP coating might therefore enhance bone apposition in the early stages because of the superior bioactivity of the BCP layer while the more stable HA layer might sustain bone bonding over long periods.     

Structural, mechanical and in vitro characterization of individually structured Ti-6Al-4V produced by direct laser forming

Direct laser forming (DLF) is a rapid prototyping technique which enables prompt modelling of metal parts with high bulk density on the base of individual three-dimensional data, including computer tomography models of anatomical structures. In our project, we tested DLF-produced material on the basis of the titanium alloy Ti-6Al-4V for its applicability as hard tissue biomaterial. To this end, we investigated mechanical and structural properties of DLF-Ti-6Al-4V. While the tensile and yield strengths of untreated DLF alloy ranged beyond 1000 MPa, a breaking elongation of 6.5+/-0.6% was determined for this material. After an additional post-DLF annealing treatment, this parameter was increased two-fold to 13.0+/-0.6%, while tensile and yield strengths were reduced by approx. 8%. A Young's modulus of 118.000+/-2.300 MPa was determined for post-DLF annealed Ti-6Al-4V. All data gained from tensile testing of post-DLF annealed Ti-6Al-4V matched American Society of Testing and Materials (ASTM) specifications for the usage of this alloy as medical material. Rotating bending tests revealed that the fatigue profile of post-DLF annealed Ti-6Al-4V was comparable to casted/hot isostatic pressed alloy. We characterized the structure of non-finished DLF-Ti-6Al-4V by scanning electron microscopy and observed a surface-associated layer of particles, which was removable by sandblasting as a finishing step. We manufactured porous specimens with nominal pore diameters of 500, 700 and 1000 microm. The diameters were reduced by the used DLF processing by approx. 300 microm. In an in vitro investigation, we cultured human osteoblasts on non-porous and porous blasted DLF-Ti-6Al-4V specimens to study morphology, vitality, proliferation and differentiation of the cells. The cells spreaded and proliferated on DLF-Ti-6Al-4V over a culture time of 14 days. On porous specimens, osteoblasts grew along the rims of the pores and formed circle-shaped structures, as visualized by live/dead staining as well as scanning electron microscopy. Overall, the DLF-Ti-6Al-4V approach proved to be efficient and could be further advanced in the field of hard tissue biomaterials.