Calcium phosphate deposition rate, structure and osteoconductivity on electrospun poly(l-lactic acid) matrix using electrodeposition or simulated body fluid incubation

Abstract

Mineralized nanofibrous scaffolds have been proposed as promising scaffolds for bone regeneration due to their ability to mimic both nanoscale architecture and chemical composition of natural bone extracellular matrix. In this study, a novel electrodeposition method was compared with an extensively explored simulated body fluid (SBF) incubation method in terms of the deposition rate, chemical composition and morphology of calcium phosphate formed on electrospun fibrous thin matrices with a fiber diameter in the range ~200-1400 nm prepared using 6, 8, 10 and 12 wt.% poly(l-lactic acid) (PLLA) solutions in a mixture of dichloromethane and acetone (2:1 in volume). The effects of the surface modification using the two mineralization techniques on osteoblastic cell (MC3T3-E1) proliferation and differentiation were also examined. It was found that electrodeposition was two to three orders of magnitude faster than the SBF method in mineralizing the fibrous matrices, reducing the mineralization time from ~2 weeks to 1h to achieve the same amounts of mineralization. The mineralization rate also varied with the fiber diameter but in opposite directions between the two mineralization methods. As a general trend, the increase of fiber diameter resulted in a faster mineralization rate for the electrodeposition method but a slower mineralization rate for the SBF incubation method. Using the electrodeposition method, one can control the chemical composition and morphology of the calcium phosphate by varying the electric deposition potential and electrolyte temperature to tune the mixture of dicalcium phosphate dihydrate and hydroxyapatite (HAp). Using the SBF method, one can only obtain a low crystallinity HAp. The mineralized electrospun PLLA fibrous matrices from either method similarly facilitate the proliferation and osteogenic differentiation of preosteoblastic MC3T3-E1 cells as compared to neat PLLA matrices. Therefore, the electrodeposition method can be utilized as a fast and versatile technique to fabricate mineralized nanofibrous scaffolds for bone tissue engineering.

Keywords: Bone tissue engineering; Calcium phosphate; Electrodeposition; Nanofiber; SBF incubation.

Figure 1. A schematic diagram of experimental setup for fabricating mineralized nanofibers by combining electrospinning and electrodeposition methods

Figure 2

The diameters of electrospun fibers from various PLLA solution concentrations.

Figure 3

Mass increases of matrices electrospun from different PLLA concentrations versus time: (a) Using the electrodeposition method at 3V and 60°C; (b) Using the SBF incubation method in 1.5 × SBF at 37°C.

Figure 3

Mass increases of matrices electrospun from different PLLA concentrations versus time: (a) Using the electrodeposition method at 3V and 60°C; (b) Using the SBF incubation method in 1.5 × SBF at 37°C.

Figure 4

SEM micrographs of mineralized PLLA (10 wt%) matrices. (a) Electrodeposition at 3V, 60°C for 60 min, (b) the magnified image of (a), (c) mineralized in 1.5× SBF for 12 days, (d) The magnified image of (c), (e) Mineralized in 1.5× SBF for 30 days, (f) The magnified image of (e).

Figure 5

XRD patterns of mineralized PLLA matrices (--DCPD, -- HAp).

Figure 6

XPS survey spectra of un-mineralized and mineralized PLLA matrices.

Figure 7

SEM micrographs of MC3T3-E1 cells grown on the mineralized and un-mineralized nanofibrous PLLA matrices after 3 days in culture: (a) Neat PLLA matrices, (b) The magnified image of (a), (c) ED PLLA matrices (3V, 60oC for 60 min), (d) The magnified image of (c), (e) SBF PLLA matrices (SBF incubation for 12 days), (f) The magnified image of (e).

Figure 8

Proliferation of MC3T3-E1 cells on the surface of the mineralized and un-mineralized nanofibrous PLLA matrices after 1, 4 and 10 days in culture. Data were expressed as mean ± SD (n = 3). Significant difference between groups is indicated by * (p < 0.05 compared with the neat PLLA matrix group).

Figure 9

ALP activity of MC3T3-E1 cells cultured on different nanofibrous matrices after 7 and 14 days in culture. Data were expressed as mean ± SD (n = 3). Significant difference between groups is indicated by * (p < 0.05 compared with the neat PLLA matrix).

Figure 10

Schematic drawing of calcium phosphate coating formation on nanofibrous PLLA matrices using different mineralization methods: (a) Electrodeposition, in which relatively large crystals such as flower-like and flake-like deposits can be formed; (b) SBF incubation, where the prepared mineral layer is relatively uniform and mainly composed of lower-crystallinity apatite, each mineralized fiber exhibited a core-shell structure with polymer fiber as the core and apatite as the shell.