Carolyn P. Ojano-Dirain

Mitochondria-targeted antioxidant MitoQ reduces gentamicin-induced ototoxicity.

Hypothesis Oral supplementation with mitoquinone (MitoQ) prevents gentamicin-induced ototoxicity in guinea pigs. Background Antioxidants have been shown to protect against aminoglycoside (AG)-induced ototoxicity. MitoQ, a mitochondria-targeted derivative of the antioxidant ubiquinone, is attached to a lipophilic triphenylphosphonium (TPP) cation, which enables its accumulation inside the mitochondria several hundred-fold over the untargeted antioxidant. MitoQ has improved bioavailability and can reach most tissues and has been used in Parkinson’s disease and hepatitis C human trials, which demonstrated that MitoQ can be safely used in humans. Thus, MitoQ is a promising novel therapeutic approach for protecting against AG-induced ototoxicity. Methods Gentamicin-treated guinea pigs were supplied with water alone (control), decyl-TPP (positive control), or MitoQ-supplemented drinking water. Auditory function was assessed by auditory brainstem response. Cochlear damage was assessed using scanning electron microscopy. Western blotting was performed to evaluate changes in proteins related to apoptosis and oxidative damage in the cochlea. Results Threshold shifts at 4 and 8 kHz at 4 and 7 weeks after gentamicin treatment were smaller in animals treated with MitoQ compared with those in the control- and decyl-TPP–treated animals (p < 0.05). Protein carbonyls and levels of the proapoptotic protein Bak were lower (p < 0.05 and p = 0.008, respectively), whereas the level of the antioxidant enzyme manganese superoxide dismutase was higher (p = 0.01) in the cochlea of MitoQ-treated animals. The expression of 3-nitrotyrosine and Hrk were not different between groups (p > 0.05). Conclusion Oral supplementation with MitoQ attenuated gentamicin-induced cochlear damage and hearing loss in guinea pigs. MitoQ holds promise as a means for protecting against AG ototoxicity.

An animal model of PDH deficiency using AAV8-siRNA vector-mediated knockdown of pyruvate dehydrogenase E1α

We evaluated the feasibility of self-complementary adeno-associated virus (scAAV) vector-mediated knockdown of the pyruvate dehydrogenase complex using small interfering RNAs directed against the E1α subunit gene (PDHA1). AAV serotype 8 was used to stereotaxically deliver scAAV8-si3-PDHA1-Enhanced Green Fluorescent Protein (knockdown) or scAAV8-EGFP (control) vectors into the right striatum and substantia nigra of rats. Rotational asymmetry was employed to quantify abnormal rotation following neurodegeneration in the nigrostriatal system. By 20weeks after surgery, the siRNA-injected rats exhibited higher contralateral rotation during the first 10min following amphetamine administration and lower 90-min total rotations (p≤0.05). Expression of PDC E1α, E1β and E2 subunits in striatum was decreased (p≤0.05) in the siRNA-injected striatum after 14weeks. By week 25, both PDC activity and expression of E1α were lower (p≤0.05) in siRNA-injected striata compared to controls. E1α expression was associated with PDC activity (R(2)=0.48; p=0.006) and modestly associated with counterclockwise rotation (R(2)=0.51;p=0.07). The use of tyrosine-mutant scAAV8 vectors resulted in ~17-fold increase in transduction efficiency of rat striatal neurons in vivo. We conclude that scAAV8-siRNA vector-mediated knockdown of PDC E1α in brain regions typically affected in humans with PDC deficiency results in a reproducible biochemical and clinical phenotype in rats that may be further enhanced with the use of tyrosine-mutant vectors.

Prevention of Gentamicin-Induced Apoptosis With the Mitochondria-Targeted Antioxidant Mitoquinone

Antioxidants have been shown to protect against aminoglycoside‐induced hearing loss. Mitoquinone (MitoQ) is a mitochondria‐targeted derivative of the antioxidant ubiquinone. MitoQ is attached to a lipophilic triphenylphosphonium (TPP) cation, which enables its accumulation inside the mitochondria several hundred‐fold over the untargeted antioxidant. The goals of this study were to determine if MitoQ attenuates gentamicin‐induced activation of caspase‐3/7 activity as a marker of apoptosis and to determine if MitoQ impacts aminoglycoside antimicrobial efficacy.

Biofilm Formation by Mycobacterium abscessus in a Lung Cavity

Biofilm Formation by Mycobacterium abscessus in a Lung Cavity Kevin P. Fennelly, Carolyn Ojano-Dirain, Qingping Yang, Lin Liu, Li Lu, Ann Progulske-Fox, Gary P. Wang, Patrick Antonelli, and Gregory Schultz Department of Medicine, Emerging Pathogens Institute, Department of Otolaryngology, Institute for Wound Research, and Department of Pathology, Immunology and Laboratory Medicine, College of Medicine, and Center for Molecular Microbiology, Department of Oral Biology, College of Dentistry, University of Florida, Gainesville, Florida; and Medical Service, North Florida/South Georgia Veterans Health System, Gainesville, Florida

Microbial flora of cochlear implants by gene pyrosequencing.

Objective To describe the microbial flora associated with cochlear implants (CIs) removed for infectious and noninfectious indications. Study Design Prospective, controlled. Setting Academic, tertiary medical center. Patients All patients undergoing CI removal. Intervention CIs were removed with aseptic technique and processed for microbial identification. Main Outcome Measure CI microbes were identified with routine culture and speciation and 16s deoxyribonucleic acid 454-pyrosequencing. Results All CIs had evidence of microbes. Propionibacterium acnes and Acidovorax facilis were more common on noninfected CIs (p = 0.005, 0.031). Staphylococcus aureus was more common on infected CIs (p = 0.003). The microbial profiles associated with CI infection were different from, but overlapped with those of noninfected CIs. Microbial culture with selective media identified pathogens not identified on pyrosequencing. Conclusion Bacteria are present on all CIs, both with and without evidence of clinical infection, but species differ with clinical status. Empiric therapy for CI infections should include coverage for S. aureus. Gene pyrosequencing and selective culture techniques may yield complementary results that may impact the management of CI infections.

Efficacy of disinfecting solutions in removing biofilms from polyvinyl chloride tracheostomy tubes.

Bacterial biofilms are prevalent in pediatric tracheostomy tubes (TTs) and are not completely cleared by standard cleaning with gauze and household detergents. We aimed to examine the effectiveness of different disinfecting solutions to remove Staphylococcus aureus (SA) and Pseudomonas aerginosa (PA) biofilms from TTs.

Biofilm formation on silicone tympanostomy tubes with polyvinylpyrrolidone coating.

OBJECTIVE To determine whether biofilm formation on silicone tympanostomy tubes (TTs) is prevented by polyvinylpyrrolidone (PVP) coating. DESIGN In vitro microbiologic study. SUBJECTS Silicone TTs with and without a PVP coating. INTERVENTION The TTs were exposed to blood or phosphate-buffered saline and cultured with Pseudomonas aeruginosa or Staphylococcus aureus. After 4 days, antibiotics were added to kill planktonic bacteria. Biofilm formation was assessed by quantitative bacterial counts and scanning electron microscopy. RESULTS Human blood enhanced S aureus biofilm formation on TTs with and without PVP (P < .001). Staphylococcus aureus biofilm formation was similar on TTs with and without PVP coating. Pseudomonas aeruginosa biofilm formation was less on TTs with PVP coating after exposure to phosphate-buffered saline (P = .04), but this difference was not significant after blood exposure (P = .19). CONCLUSIONS Polyvinylpyrrolidone coating of TTs imparts resistance to P aeruginosa biofilm formation. The clinical impact of PVP on TTs may be attenuated by exposure to blood, but this will require study in clinical trials.

Topical antibiotic treatment reduces tympanostomy tube biofilm formation

Single doses of different ototopical antibiotic preparations (OAPs) have been shown to have an unequal reduction of post tympanostomy tube otorrhea (PTTO). Microbial biofilm formation on the tympanostomy tube (TT) has been implicated as one cause of PTTO. The goal of this study was to determine if TT exposure to a single dose of OAP reduces biofilm formation by Pseudomonas aeruginosa.

Effectiveness of pediatric tracheostomy tube cleaning.

OBJECTIVE To determine the effectiveness of typical reprocessing of pediatric tracheostomy tubes (TTs) with detergent-soaked gauze pads. DESIGN In vitro microbiologic study. SETTING Department of Otolaryngology, University of Florida, Gainesville. PARTICIPANTS Polyvinyl chloride and silicone TTs of 3 different manufacturers. INTERVENTION Thirty TTs were cultured with Pseudomonas aeruginosa or Staphylococcus aureus and reprocessed after 4 days. Eighteen additional TTs were exposed to plasma and then cultured with P aeruginosa or S aureus for 7 days. MAIN OUTCOME MEASURES The presence of biofilms was assessed before and after cleaning by quantitative bacterial counts and scanning electron microscopy. RESULTS Bacterial counts that were obtained before cleaning of the tubes did not differ among brands. Reprocessing reduced P aeruginosa and S aureus bacterial counts in the 4-day group (P = .003 and P = .004, respectively), but clean TTs had a mean count of 10(5) colony-forming units/mL. Reprocessing did not significantly reduce S aureus or P aeruginosa bacterial counts in TTs pretreated with plasma and exposed to 7-day culture. CONCLUSIONS P aeruginosa and S aureus biofilms in pediatric TTs are not eradicated by standard cleaning methods. Further research is needed to determine the clinical significance of these findings and whether strategies to prevent biofilm formation or more effective disinfection methods would result in improved clinical outcomes.

Effect of repeated tracheostomy tube reprocessing on biofilm formation

To determine the effect of repeated reprocessing of pediatric tracheostomy tubes (TTs) on biofilm formation.