Iondoped mesoporous bioactive glass nanoparticles for wound healing

Ion-doped mesoporous bioactive glass nanoparticles for wound healing applications Alessandra Baria, Sonia Fiorillia, Carlotta Pontremolia, Joanna Shepherdb, Anthony J. Bullockc, Sheila Mac. Neilc, Chiara Vitale- Brovarone*a a Department of Applied Science and Technology, Politecnico di Torino, Corso Duca degli Abruzzi 24, 10129 Torino, Italy b School of Clinical Dentistry, The University of Sheffield, 19 Claremont Crescent, Sheffield S 10 2 TA, UK Introduction & Aim of the work c Department of Engineering Materials, Kroto Research Institute, University of Sheffield, Broad Lane, Sheffield S 3 7 HQ, UK Bioactive glasses (BGs) have been extensively studied for bone repair. Besides, recent research has also shown their positive effects on wound healing. Among BGs, mesoporous bioactive glasses (MBGs) are receiving increasing attention as tissue-regeneration systems. Their high surface area and accessible pore volume give the possibility of incorporating and releasing therapeutic ions to impart biological properties, such as pro-angiogenic effects (cerium [1], copper [2]) or antibacterial activities (copper) [3]. The aim of this work is to develop mesoporous bioactive glasses doped with cerium and copper in order to obtain a multifunctional platform for tissue regeneration. To evaluate ions antimicrobial potential, MIC and viable count test were run with copper and cerium salts. Materials Salts & Methods Bacteria strains: Antibacterial test: Salts o S. Aureus S 235 (Gram. Ce(NO 3)3 · 6 H 2 O positive) Cu(NO 3)2 · 2. 5 H 2 O o P. Aeruginosa SOM 1 (Gramnegative) Ag. NO 3 o MRSA-Methicillin Resistant Cu. SO 4 Staphylococcus Aureus (Grampositive) o o 2) Viable count test 1) MIC test Nanoparticles Ø Base-catalyzed sol-gel method [4] o Ce US 2% MTG: 85 Si. O 2/ 13 Ca. O/ 2 Ce. O (%mol) o Cu US 2% MTG: 85 Si. O 2/ 13 Ca. O/ 2 Cu. O (%mol) Ø Spray-drying procedure [5] o Ce SD 2% MTG: 85 Si. O 2/ 13 Ca. O/ 2 Ce. O (%mol) o Cu SD 2% MTG: 85 Si. O 2/ 13 Ca. O/ 2 Cu. O (%mol) Nanoparticles - FESEM XRD N 2 adsorption-desorption analysis Release test in Tris-HCl (3 h, 1, 3, 8 days) Viable count Why Ce and Cu? Cerium Copper - Pro-angiogenic - Antibacterial Results Salts Spray dryer samples: 500 -5000 nm MIC test P. Aeruginosa SOM 1 MRSA Ce(NO 3)3 · 6 H 2 O 6. 250 mg/ml 3. 120 mg/ml 6. 250 mg/ml Ce US 2% MTG 5. 7 505 0. 6 Ag. NO 3 0. 078 mg/ml Cu US 2% MTG 4 535 0. 5 Cu. SO 4 1. 560 mg/ml 3. 125 mg/ml Ce SD 2% MTG 8. 7 470 0. 6 Cu(NO 3)2 · 2. 5 H 2 O 3. 125 mg/ml Cu SD 2% MTG 5. 5 189 0. 3 Sample 1. 00 E+08 1. 00 E+07 1. 00 E+05 Ag. NO 3 1. 00 E+04 Cu. SO 4 1. 00 E+03 Cu. NO 3 ctrl Ce. NO 3 1. 00 E+05 1. 00 E+04 1. 00 E+03 600 400 Ag. NO 3 Cu. SO 4 Cu. No 3 1. 00 E+06 1. 00 E+05 1. 00 E+04 0 ctrl 1. 00 E+02 1. 00 E+01 1. 00 E+00 20 30 40 50 2 Theta [degree] 60 70 80 Ag. NO 3 Cu. SO 4 S. 235 Cu. NO 3 1. 00 E+02 10 Ce. NO 3 1. 00 E+00 Conclusion [1] J. P. Garner, P. S. Heppell. , Burns 31 (2005) 539– 547 [2] A. Hoppe et al. , Biomaterials 32 (2011) 2757 -74 [3] C. Wu et al. , Biomaterials 34 (2013) 422 -433 [4] M. Shi et al. , Acta Biomater. 30 (2016) 334– 344 [5] C. Vitale-Brovarone et al. , Key. Eng. Mat 631 (2015) 43 -47 [6] P. G. et al. , Clinical Microbiology Reviews (2001) [7] O. Zeions et al. , Nanotoxicology 3 (2009) 284– 295 100. 00 90. 00 80. 00 70. 00 60. 00 50. 00 40. 00 30. 00 20. 00 10. 00 70. 00 60. 00 50. 00 40. 00 30. 00 20. 00 1 day Cu. SD Ce. SD 3 days Cu. US Ce. US 8 days 0. 00 60. 00 50. 00 40. 00 30. 00 20. 00 10. 00 suspension 70. 00 suspension 1 day Cu. SD Ce. SD 3 days Cu. US 8 days Ce. US From viable count, it is possible to notice that: - Antibacterial effect against Staphylococcus Aureus S. 235 is more evident and still present after 3 days - Survival rate increases after 3 days of soaking against MRSA and SOM 1, suggesting that ions probably created some compounds with other elements present in the medium which are not able to act against these bacteria strains. The Angiogenesis capacity of the nanomatrices will be tested on fertilised chick eggs by Chorionic Allantoic Membrane assay (CAM). On going Antibacterial studies on salts showed that silver and copper have good antimicrobial properties against Gram Positive and Gram negative strains. These ions, incorporated in the mesoporous glass frameworks, obtained by two different synthesis routes, imparted good antibacterial properties. References From the viable count, it is possible to notice that: - Cu salts have the same effect against Gram + and Gram - Ag is more effective against Gram + - Ce showed not satisfying results against all strains considering the values of cfu/ml (higher than the limit of 10^5 CFU/ml regarded as “infection”) [6] MRSA SOM 1 Survival Rate (%) [7] Ce. NO 3 1. 00 E+06 800 Survival Rate (%) [7] 1. 00 E+06 1000 200 MRSA cfu/ml ctrl Intensity [a. u. ] 1. 00 E+09 1. 00 E+08 P. Aeruginosa SOM 1 cfu/ml S 235 cfu/ml 1200 1. 00 E+09 Release test No oxidic form 1400 1. 00 E+10 Pore volume (cc/g) Ce SD 2% MTG 1600 1. 00 E+10 1. 00 E+09 Surface Pore size area - BET (nm) (m 2/g) S. Aureus 235 Viable count 1. 00 E+10 Sol-gel samples: 100 nm Sample Survival rate [%] [7] v Ag salt is more effective compared to copper and cerium salt v The MIC values for Gram + and Gram - are almost the same for Ag and Cu salts v Ce salt seems to be more effective against Gram - Nanoparticles Kroto Research Institute, University of Sheffield, Sheffield The activity leading to this work has received funding from H 2020 NMP-PILOTS-2015 under grant agreement no. 685872 (MOZART). chiara. vitale@polito. it alessandra. bari@polito. it