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.2008 Jan;29(3):337-49.
doi: 10.1016/j.biomaterials.2007.09.029. Epub 2007 Oct 17.

Miscibility and in vitro osteocompatibility of biodegradable blends of poly[(ethyl alanato) (p-phenyl phenoxy) phosphazene] and poly(lactic acid-glycolic acid)

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Miscibility and in vitro osteocompatibility of biodegradable blends of poly[(ethyl alanato) (p-phenyl phenoxy) phosphazene] and poly(lactic acid-glycolic acid)

Meng Deng et al. Biomaterials.2008 Jan.

Abstract

Previously we demonstrated the ability of ethyl glycinato substituted polyphosphazenes to neutralize the acidic degradation products and control the degradation rate of poly(lactic acid-glycolic acid) (PLAGA) by blending. In this study, blends of high strength poly[(50% ethyl alanato) (50% p-phenyl phenoxy) phosphazene] (PNEA(50)PhPh(50)) and 85:15 PLAGA were prepared using a mutual solvent approach. Three different solvents, methylene chloride (MC), chloroform (CF) and tetrahydrofuran (THF) were studied to investigate solvent effects on blend miscibility. Three different blends were then fabricated at various weight ratios namely 25:75 (BLEND25), 50:50 (BLEND50), and 75:25 (BLEND75) using THF as the mutual solvent. The miscibility of the blends was evaluated by scanning electron microscopy (SEM), differential scanning calorimetry (DSC), and Fourier transform infrared spectroscopy (FTIR). Among these, BLEND25 was miscible while BLEND50 and BLEND75 were partially miscible. Furthermore, BLEND25 formed apatite layers on its surface as evidenced in a biomimetic study performed. These novel blends showed cell adhesion and proliferation comparable to PLAGA. However, the PNEA(50)PhPh(50) component in the blends was able to increase the phenotypic expression and mineralized matrix synthesis of the primary rat osteoblasts (PRO) in vitro. Blends of high strength PNEA(50)PhPh(50) and 85:15 PLAGA are promising biomaterials for a variety of musculoskeletal applications.

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Figures

Figure 1
Figure 1
SEM micrographs of BLEND25 films fabricated using three different solvents, where (A, B) methylene chloride, (C, D) chloroform, and (E, F) tetrahydrofuran were presented in two different magnifications of ×1000 and ×5000 respectively. BLEND25 fabricated using THF resulted in a uniform and smooth surface without any visual phase separation.
Figure 2
Figure 2
Differential scanning calorimetry thermograms for BLEND25 films fabricated using three different solvents namely methylene chloride (MC), chloroform (CF) and tetrahydrofuran (THF). Blends fabricated using THF presented a singleTg value ~33.5°C indicating the blend miscibility, while two temperature transitions (indicated by the arrows) were observed for BLEND25 fabricated using MC and CF.
Figure 3
Figure 3
SEM micrographs of (A) BLEND25, (B) BLEND50, and (C) BLEND75 fabricated using THF as solvent at a magnification of ×1000. BLEND25 showed uniform morphology indicating blend miscibility whereas other two blends showed visible phase separation.
Figure 4
Figure 4
FTIR spectrum presenting carbonyl absorption region for PNEA50PhPh50, PLAGA and their blends. Carbonyl stretching frequencies shifted from ~1750 to ~1739 cm−1 indicating intermolecular hydrogen bonding that cause the miscibility of these blends.
Figure 5
Figure 5
SEM micrographs of PNEA50PhPh50 and BLEND25 films incubated in 1.5x SBF at 37°C for 21 days, where (A–C) represented PNEA50PhPh50 after 21 days of incubation at a magnification of ×350, ×60,000 and ×100,000 respectively; (D–F) represented BLEND25 after 21 days of incubation at a magnification of ×1000, ×60,000 and ×100,000 respectively; (G) confirmed the presence of calcium and phosphorus with Ca/P atomic ratio around 1.67.
Figure 6
Figure 6
SEM micrographs of primary rat osteoblasts cultured on the films of BLEND75 at different time points, where (A, B) 7 day, (C, D) 14 day, and (E, F) 21 day were presented at a magnification of ×300 and ×1000 respectively for each pair. PRO showed a normal morphological sequence of proliferation with time and by day 21 completely covered cell multilayers were observed.
Figure 7
Figure 7
SEM micrographs of primary rat osteoblasts cultured on different polymer films for a period of 21 days, where (A) PNEA50PhPh50, (B) BLEND25, (C) BLEND50, and (D) PLAGA surfaces were presented at a magnification of ×1000. Surfaces of all the polymer and blend films were completely covered by PRO multilayers.
Figure 8
Figure 8
Cell proliferation measured by MTS assay. Cell numbers on these blend films at all the time points were comparable to PLAGA. BLEND25 and BLEND50 showed significantly higher cell number than PNEA50PhPh50 at day 21. (*) denotes significant difference (p<0.05) as compared to PNEA50PhPh50 matrices while (**) indicates significant difference (p<0.05) as compared to PLAGA matrices.
Figure 9
Figure 9
Alkaline phosphatase activity for osteoblastic phenotype expression. At all time points, ALP activity was significantly enhanced on PNEA50PhPh50 and BLEND75 surfaces as compared to PLAGA. (*) denotes significant difference (p<0.05) as compared to PNEA50PhPh50 matrices while (**) indicates significant difference (p<0.05) as compared to PLAGA matrices.
Figure 10
Figure 10
Synthesis of mineralized matrix was assessed by Alizarin Red assay. All the blends showed higher amount of calcium deposition as compared to the parent polymers at day 21. (*) denotes significant difference (p<0.05) as compared to PNEA50PhPh50 matrices while (**) indicates significant difference (p<0.05) as compared to PLAGA matrices.
Scheme I
Scheme I
Synthesis of poly[(50% ethyl alanato) (50% p-phenyl phenoxy) phosphazene] (PNEA50PhPh50).
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