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Type I and Type II mechanisms of antimicrobial photodynamic therapy: An in vitro study on gram-negative and gram-positive bacteria

Huang, Liyi and Xuan, Yi and Koide, Yuichiro and Zhiyentayev, Timur and Tanaka, Masamitsu and Hamblin, Michael R. (2012) Type I and Type II mechanisms of antimicrobial photodynamic therapy: An in vitro study on gram-negative and gram-positive bacteria. Lasers in Surgery and Medicine, 44 (6). pp. 490-499. ISSN 0196-8092.

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Background and Objectives: Antimicrobial photodynamic therapy (APDT) employs a non-toxic photosensitizer (PS) and visible light, which in the presence of oxygen produce reactive oxygen species (ROS), such as singlet oxygen (^(1)O_2, produced via Type II mechanism) and hydroxyl radical (HO., produced via Type I mechanism). This study examined the relative contributions of ^(1)O_2 and HO. to APDT killing of Gram-positive and Gram-negative bacteria. Study Design/Materials and Methods: Fluorescence probes, 3′-(p-hydroxyphenyl)-fluorescein (HPF) and singlet oxygen sensor green reagent (SOSG) were used to determine HO. and ^(1)O_2 produced by illumination of two PS: tris-cationic-buckminsterfullerene (BB6) and a conjugate between polyethylenimine and chlorin(e6) (PEI–ce6). Dimethylthiourea is a HO. scavenger, while sodium azide (NaN_3) is a quencher of ^(1)O_2. Both APDT and killing by Fenton reaction (chemical generation of HO.) were carried out on Gram-positive bacteria (Staphylococcus aureus and Enterococcus faecalis) and Gram-negative bacteria (Escherichia coli, Proteus mirabilis, and Pseudomonas aeruginosa). Results: Conjugate PEI-ce6 mainly produced ^(1)O_2 (quenched by NaN_3), while BB6 produced HO. in addition to ^(1)O_2 when NaN_3 potentiated probe activation. NaN_3 also potentiated HPF activation by Fenton reagent. All bacteria were killed by Fenton reagent but Gram-positive bacteria needed a higher concentration than Gram-negatives. NaN3 potentiated Fenton-mediated killing of all bacteria. The ratio of APDT killing between Gram-positive and Gram-negative bacteria was 2 or 4:1 for BB6 and 25:1 for conjugate PEI-ce6. There was a NaN_3 dose-dependent inhibition of APDT killing using both PEI-ce6 and BB6 against Gram-negative bacteria while NaN_3 almost failed to inhibit killing of Gram-positive bacteria. Conclusion: Azidyl radicals may be formed from NaN_3 and HO.. It may be that Gram-negative bacteria are more susceptible to HO. while Gram-positive bacteria are more susceptible to ^(1)O_2. The differences in NaN_3 inhibition may reflect differences in the extent of PS binding to bacteria (microenvironment) or differences in penetration of NaN_3 into cell walls of bacteria.

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Additional Information:© 2012 Wiley Periodicals, Inc. Issue published online: 12 July 2012; Article first published online: 3 July 2012; Manuscript Accepted: 29 May 2012. This study was supported by NIH (R01A1050875 to M.R.H.) and US Air Force MFEL Program (FA9550-04-1-0079). These data were presented at ASLMS meeting in 2010 by Dr Huang who received a travel grant from ASLMS. This contribution satisfies the condition of the travel grant that a manuscript should be submitted to LSM within 1 year after the annual conference. Conflict of Interest Disclosures: All authors have completed and submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest and none were reported.
Funding AgencyGrant Number
US Air Force MFEL ProgramFA9550-04-1-0079
Subject Keywords:antimicrobial photodynamic inactivation; singlet oxygen; hydroxyl radical; sodium azide; gram-positive and gram-negative bacteria; antimicrobial photodynamic therapy; polyethylenimine chlorin(e6) conjugate; tris-cationic fullerene
Issue or Number:6
Record Number:CaltechAUTHORS:20120814-102121124
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Official Citation:Huang, L., Xuan, Y., Koide, Y., Zhiyentayev, T., Tanaka, M. and Hamblin, M. R. (2012), Type I and Type II mechanisms of antimicrobial photodynamic therapy: An in vitro study on gram-negative and gram-positive bacteria. Lasers Surg. Med., 44: 490–499. doi: 10.1002/lsm.22045
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:33161
Deposited By: Jason Perez
Deposited On:14 Aug 2012 18:44
Last Modified:03 Oct 2019 04:07

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