In vitro degradation of polymeric networks of poly(propylene fumarate) and the crosslinking macromer poly(propylene fumarate)-diacrylate

Polymeric networks of poly(propylene fumarate) (PPF) crosslinked with poly(propylene fumarate)-diacrylate (PPF-DA) are currently being investigated as an injectable, biodegradable bone cement. This study examined the effect of crosslinking density, medium pH, and the incorporation of a beta-tricalcium phosphate (beta-TCP) filler on the in vitro degradation of PPF/PPF-DA. Cylindrical specimens were submerged in buffered saline at 37 degrees C and the change in weight, geometry, and compressive mechanical properties were monitored over a 52-week period. All formulations showed an initial increase in modulus and yield strength over the first 12 weeks, achieving maxima of 1307+/-101 and 51+/-3MPa, respectively, for the beta-TCP composite. PPF/PPF-DA networks with the lower crosslinking density demonstrated the greatest degradation with a 17% mass loss. Samples in the lower buffer pH 5.0 compared to physiological pH 7.4 did not show any differences in mass loss, but exhibited a faster decrease in the compressive strength over time. The beta-TCP composites maintained their mechanical properties at the level following their initial increase. These results show that the degradation of PPF/PPF-DA networks can be controlled by the crosslinking density, accelerated at a lower pH, and prolonged with the incorporation of the beta-TCP filler.     

Increased frequency of the rare PstI allele (P2) in a population of CAD patients in Northern Greece

Restriction fragment length polymorphisms (RFLPs) at the apolipoprotein AI-CIII-AIV gene cluster and their association with coronary artery disease (CAD) and lipid levels were studied in a Northern Greek population. Ninety-five patients with CAD and fifty-four normal controls, angio-graphically proven, were included in this study. Using genomic hybridization techniques, three polymorphic restriction sites were identified at this locus: the PstI at the 3' end of the apoAI gene, the SacI at the 3' non-coding region of the apoCIII gene and the PvuII at the intergenic region between the apoCIII-AIV genes. The rare allele (P2) arising from the absence of the PstI restriction site was observed with a significantly higher frequency (p<0.01) in patients compared to normals (0.11 vs 0.02). In contrast, the rare allele for the SacI polymorphic site had a similar distribution among patients and controls (0.12 vs 0.16). The same was observed for the PvuII RFLP (0.04 vs 0.05). Correlation of lipid and apolipoprotein AI levels with the three RFLPs revealed no significant association, although apo AI and HDL were lower in patients with the P2 allele. Thus, in this Greek population, only the PstI polymorphism, among the polymorphic restriction sites examined, appears to be associated with CAD.     

Flow Perfusion Culture of Marrow Stromal Cells Seeded on Porous Biphasic Calcium Phosphate Ceramics

Calcium phosphate ceramics have been widely used for filling bone defects to aid in the regeneration of new bone tissue. Addition of osteogenic cells to porous ceramic scaffolds may accelerate the bone repair process. This study demonstrates the feasibility of culturing marrow stromal cells (MSCs) on porous biphasic calcium phosphate ceramic scaffolds in a flow perfusion bioreactor. The flow of medium through the scaffold porosity benefits cell differentiation by enhancing nutrient transport to the scaffold interior and by providing mechanical stimulation to cells in the form of fluid shear. Primary rat MSCs were seeded onto porous ceramic (60% hydroxyapatite, 40% β-tricalcium phosphate) scaffolds, cultured for up to 16 days in static or flow perfusion conditions, and assessed for osteoblastic differentiation. Cells were distributed throughout the entire scaffold by 16 days of flow perfusion culture whereas they were located only along the scaffold perimeter in static culture. At all culture times, flow perfused constructs demonstrated greater osteoblastic differentiation than statically cultured constructs as evidenced by alkaline phosphatase activity, osteopontin secretion into the culture medium, and histological evaluation. These results demonstrate the feasibility and benefit of culturing cell/ceramic constructs in a flow perfusion bioreactor for bone tissue engineering applications.     

Use of stereolithography to manufacture critical-sized 3D biodegradable scaffolds for bone ingrowth

A novel approach to the manufacture of biodegradable polymeric scaffolds for tissue-engineering utilizing stereolithography (SLA) is presented. SLA is a three-dimensional (3D) printing method that uses an ultraviolet laser to photo-crosslink a liquid polymer substrate. The current generation of SLA devices provide a 3D printing resolution of 0.1 mm. The experiments utilized a biodegradable resin mixture of diethyl fumarate (DEF), poly(propylene fumarate) (PPF), and a photoinitiator, bisacylphosphine oxide (BAPO). The PPF is crosslinked with the use of the SLA's UV laser (325-nm wavelength). An SLA device was retrofitted with a custom fixture build tank enclosing an elevator-driven build table. A 3D prototype model testing the manufacturing control this device provides was created in a computer-aided-design package. The resulting geometric data were used to drive the SLA process, and a DEF/PPF prototype part was successfully manufactured. These scaffolds have application in the tissue engineering of bony substrates.     

Emphysematous lung destruction by cigarette smoke. The effects of latent adenoviral infection on the lung inflammatory response.

This study was designed to test the hypothesis that cigarette smoke-induced inflammation and emphysema are amplified by the presence of latent adenoviral (Ad) infection, and to determine whether this emphysematous process can be reversed by all-trans-retinoic acid (RA) treatment. The results confirm that in guinea pigs, chronic cigarette-smoke exposure caused lesions similar to human centrilobular emphysema. They also show that latent Ad infection combined with cigarette-smoke exposure caused an excess increase in lung volume (P < 0.001), air-space volume (P < 0.001), and lung weight (P < 0.01), and further decrease in surface-to-volume ratio (P < 0.001) compared with smoke exposure alone. RA treatment failed to reverse these emphysematous changes. Analysis of inflammatory response in parenchymal and airway tissue showed that smoking caused an increase of polymorphonuclear leukocytes (PMNs) (P < 0.0002), macrophages (P < 0.001), and CD4 cells (P < 0.0009), and that latent Ad infection independently increased PMNs (P < 0.001), macrophages (P = 0.003), and CD8 cells (P < 0.001). We conclude that latent Ad infection amplifies the emphysematous lung destruction and increases the inflammatory response produced by cigarette-smoke exposure. In this study, the increase in CD4 was associated with cigarette smoke and the increase in CD8 cells with latent Ad infection.     

Development of an injectable,in situ crosslinkable,degradable polymeric carrier for osteogenic cell populations. Part 2. Viability of encapsulated marrow stromal osteoblasts cultured on crosslinking poly(propylene fumarate)

The effect of temporary encapsulation of rat marrow stromal osteoblasts in crosslinked gelatin microparticles on cell viability and proliferation was investigated in this study for microparticles placed on a crosslinking poly(propylene fumarate) (PPF) composite over a 7 day time period. Encapsulated cells were seeded on crosslinking PPF composites at times up to 10 min following initiation of the crosslinking reaction, and also on fully crosslinked PPF composites and tissue culture polystyrene controls, with a cell seeding density of 5.3 x 10(4) cells/cm2. The crosslinked PPF composite exhibited an average gel point of 10.3 min and an average maximum crosslinking temperature of 47.5 degrees C. Cell viability and proliferation were assessed by DNA and 3H-thymidine assays and the results were compared with those for nonencapsulated cells. The results showed that the addition time of cells to a crosslinking PPF composite had a large effect on cell viability and proliferation for both encapsulated and nonencapsulated cells with more surviving cells added at later time points. Most importantly, the temporary encapsulation of cells significantly enhanced cell viability at earlier time points. The data indicate that the presence of gelatin microparticles does not affect the crosslinking of a PPF composite. They further suggest that the temporary encapsulation of cells in crosslinked gelatin microparticles may preserve the viability of cells contained in an actively crosslinking PPF composite used as an injectable polymeric scaffold serving also as a carrier for osteogenic cell populations.     

Effect of fibronectin- and collagen I-coated titanium fiber mesh on proliferation and differentiation of osteogenic cells

The objective of this study was to evaluate the effects of fibronectin and collagen I coatings on titanium fiber mesh on the proliferation and osteogenic differentiation of rat bone marrow cells. Three main treatment groups were investigated in addition to uncoated titanium fiber meshes: meshes coated with fibronectin, meshes coated with collagen I, and meshes coated first with collagen I and then subsequently with fibronectin. Rat bone marrow cells were cultured for 1, 4, 8, and 16 days in plain and coated titanium fiber meshes. In addition, a portion of each of these coating treatment groups was cultured in the presence of antibodies against fibronectin and collagen I integrins. To evaluate cellular proliferation and differentiation, constructs were examined for DNA, osteocalcin, and calcium content and alkaline phosphatase activity. There were no significant effects of the coatings on cellular proliferation as indicated by the DNA quantification analysis. When antibodies against fibronectin and collagen I integrins were used, a significant reduction (p < 0.05) in cell proliferation was observed for the uncoated titanium meshes, meshes coated with collagen, and meshes coated with collagen and fibronectin. The different coatings also did not affect the alkaline phosphatase activity of the cells seeded on the coated meshes. However, the presence of antibodies against fibronectin or collagen I integrins resulted in significantly delayed expression of alkaline phosphatase activity for uncoated titanium meshes, meshes coated with collagen, and meshes coated with collagen and fibronectin. Calcium measurements did not reveal a significant effect of fibronectin or collagen I coating on calcium deposition in the meshes. Also, no difference in calcium content was observed in the uncoated titanium meshes and meshes coated with fibronectin when antibodies against fibronectin or collagen I integrins were present. Meshes coated with both collagen I and fibronectin showed significantly higher calcium content when cultured in the presence of antibodies to collagen and fibronectin integrins. A similar phenomenon was also observed for collagen-coated meshes cultured in the presence of antibodies to fibronectin integrins. No significant differences in osteocalcin content were observed between the treatment groups. However, all groups exposed to antibodies against fibronectin integrins showed a significant decrease in osteocalcin content on day 16. These results show that a fibronectin or collagen I coating does not stimulate the differentiation of rat bone marrow cells seeded in a titanium fiber mesh.     

Skeletal differences between black and white men and their relevance to body composition estimates

Skeletal differences exist between closely matched Black and White women, although it is unknown if similar differences also exist between Black and White men after controlling for age, body weight, and stature. The aim of this study was twofold: to test the hypothesis that Black men have greater bone mass, higher bone mineral density, and longer limbs compared to White men of similar age, weight, and height; and second, to establish if ethnic variation in skeletal characteristics has an impact on the models upon which three widely used methods for estimating total body fat are based. Twenty-four healthy Black men were matched by age (±5 years), height (±3 cm), and weight (±2 kg) to 24 healthy White men. Skeletal characteristics and body composition were studied using anatomical and compartment estimates derived by anthropometry, ³H₂O dilution, hydrodensitometry, whole-body ⁴⁰K counting, and dual photon systems. Black men had greater bone mineral mass (P = 0.007), higher bone density (P = 0.054), longer femurs (P = 0.002), longer anthropometric arm and thigh lengths (P = 0.001 and P = 0.002, respectively), lower spine to femur ratio (P = 0.004), and similar spine length (P = 0.271) compared to White men. Total body fat and fat-free body mass (FFM) were estimated in the men using a four-compartment model. Black and White men had similar total body fat, K (TBK), water (TBW), and FFM. Density of FFM and TBK/FFM were also similar between Black and White men, suggesting that current two-compartment hydrodensitometry and TBK models for estimating fat may not require adjustments for ethnicity. The TBW/FFM ratio, which is the main assumed steady-state relation for the two-compartment TBW method of estimating fat, was modestly increased (P = 0.05) in Black men (x? ± SD, 0.744 ± 0.018) compared to White men (0.732 ± 0.021). These results confirm that Black and White men differ significantly in some skeletal characteristics and these differences have implications in the study of both osteoporosis and human body composition. © 1994 Wiley-Liss, Inc.     

Solute-free water excretion and electrolyte-free water excretion are better terms than solute-free water clearance and electrolyte-free water clearance

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