Enoxacin-loaded Poly (lactic-co-glycolic acid) Coating on Porous Magnesium Scaffold as a Drug Delivery System: Antibacterial Properties and Inhibition of Osteoclastic Bone Resorption

doi:10.1016/j.jmst.2016.07.013

Abstract

Abstract:

Implant-associated infection remains a difficult medical problem in orthopedic surgery. Therefore, the development of multifunctional bone implants for treating infection and regenerating lost bone tissue, which may be a result of infection, is important. In the present study, we report the fabrication of enoxacin-loaded poly (lactic-co-glycolic acid) (PLGA) coating on porous magnesium scaffold (Enox-PLGA-Mg) which combine the favorable properties of magnesium, the antibacterial property and the effect of inhibition of osteoclastic bone resorption of enoxacin. The drug loaded PLGA coating of Mg scaffold enables higher drug loading efficiency (52%-56%) than non-coating enoxacin loaded Mg scaffold (Enox-Mg) (4%-5%). Enox-PLGA-Mg exhibits sustained drug release for more than 14 days, and this controlled release of enoxacin significantly inhibits bacterial adhesion and prevented biofilm formation by Staphylococcus epidermidis (ATCC35984) and Staphylococcus aureus (ATCC25923). Biocompatibility tests with Balb/c mouse embryo fibroblasts (Balb/c 3T3 cells) indicate that PLGA-Mg has better biocompatibility than Mg. Finally, we also demonstrate that Enox-PLGA-Mg extract potently inhibited osteoclast formation in vitro. Therefore, Enox-PLGA-Mg has the potential to be used as a multifunctional controlled drug delivery system bone scaffolds to prevent and/or treat orthopedic peri-implant infections.

Key words: Porous magnesium scaffold, Poly (lactic-co-glycolic acid) (PLGA), Drug delivery system, Bactericidal activity, Enoxacin, Osteolysis

Li Yang,Liu Xuqiang,Tan Lili,Ren Ling,Wan Peng,Hao Yongqiang,Qu Xinhua,Yang Ke,Dai Kerong. Enoxacin-loaded Poly (lactic-co-glycolic acid) Coating on Porous Magnesium Scaffold as a Drug Delivery System: Antibacterial Properties and Inhibition of Osteoclastic Bone Resorption[J]. J. Mater. Sci. Technol., 2016, 32(9): 865-873.

Figures/Tables 7

Fig. 1. FESEM images for (a) Mg, (b) Enox-Mg, (c) PLGA-Mg and (d) Enox-PLGA-Mg.

Fig. 2. (A) Drug loading efficiency of Enox-Mg and Enox-PLGA-Mg in enoxacin solution with different original concentrations: (a) 1?mg/mL, (b) 2?mg/mL, and (c) 3?mg/ML. (B) The cumulative amount of enoxacin released from Enox-Mg and Enox-PLGA-Mg in 1?mL of PBS. Significant difference (**p?<?0.01).

Fig. 3. (a) Representative images of viable bacteria grown on different samples after 12?h of culture. The number of viable bacteria of ATCC35984 (b) and ATCC25923 (c) on different TSA samples after 24?h of culture were counted and normalized against the number of cells on Mg samples. Significant difference: **p?<?0.01, #p?<?0.01.

Fig. 4. (a) CLSM images of bacteria adhered to the samples after 8?h of culture. Bacteria are stained with green fluorescent SYTO 9 and red fluorescent propidium iodide, which cause living cells to appear green and dead cells to appear red under CLSM. Absorption of crystal violet was used as an indicator of biofilm formation by ATCC35984 (b) and ATCC25923 (c) after 2, 5 and 8?h of culture. The relative absorption of crystal violet was normalized to the 2-h control (mean ± SD, **p?<?0.01, #p?<?0.01).

Fig. 5. Attachment and proliferation of the 3T3 cells on the different surfaces of the samples tested. (a) Adhesion measured by CCK-8 assay. (b) The relative proliferation rate of the 3T3 cells. The OD values at Day 4 and Day 7 were normalized to Day 1. (mean ± SD, **p?<?0.01).

Fig. 6. (a) Bone marrow-derived macrophages (BMMs) were treated with PLGA-Mg and Enox-PLGA-Mg extracts followed by 30?ng/mL M-CSF and 50?ng/mL RANKL for 5-7 days. Cells were fixed with 4% paraformaldehyde and subjected to tartrate-resistant acid phosphatase (TRAP) staining. (b) The number and (c) area of TRAP-positive cells were counted. (mean ± SD, *p?<?0.05, **p?<?0.01).

Fig. 7. Enox-PLGA-Mg extract inhibited RANKL-induced osteoclast-specific gene expression. Once mature osteoclasts were observed, total RNA was extracted and osteoclast-specific gene expression (including TRAP, cathepsin K, CTR, c-fos, and NFATC1) was measured using real-time PCR. Results were normalized to the expression of GAPDH. All experiments were performed at least three times (*p?<?0.05; **p?<?0.01).

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