Eyedrop Vaccination Induced Systemic and Mucosal Immunity against Influenza Virus in Ferrets.

We investigated eyedrop vaccination (EDV) in pre-clinical development for immunological protection against influenza and for potential side effects involving ocular inflammation and the central nervous system (CNS). Live attenuated influenza EDV, CA07 (H1N1), PZ-4 (H1N2) and Uruguay (H3N2), induced both systemic and mucosal virus-specific antibody responses in ferrets. In addition, EDV resulted in a clinically significant protection against viral challenge, and suppression of viral replication in nasal secretion and lung tissue. Regarding safety, we found that administered EDV flow through the tear duct to reach the base of nasal cavity, and thus do not contact the olfactory bulb. All analyses for potential adverse effects due to EDV, including histological and functional examinations, did not reveal significant side effects. On the basis of these findings, we propose that EDV as effective, while being a safe administration route with minimum local side effects, CNS invasion, or visual function disturbance.

Inactivated Eyedrop Influenza Vaccine Adjuvanted with Poly(I:C) Is Safe and Effective for Inducing Protective Systemic and Mucosal Immunity.

The eye route has been evaluated as an efficient vaccine delivery routes. However, in order to induce sufficient antibody production with inactivated vaccine, testing of the safety and efficacy of the use of inactivated antigen plus adjuvant is needed. Here, we assessed various types of adjuvants in eyedrop as an anti-influenza serum and mucosal Ab production-enhancer in BALB/c mice. Among the adjuvants, poly (I:C) showed as much enhancement in antigen-specific serum IgG and mucosal IgA antibody production as cholera toxin (CT) after vaccinations with trivalent hemagglutinin-subunits or split H1N1 vaccine antigen in mice. Vaccination with split H1N1 eyedrop vaccine antigen plus poly(I:C) showed a similar or slightly lower efficacy in inducing antibody production than intranasal vaccination; the eyedrop vaccine-induced immunity was enough to protect mice from lethal homologous influenza A/California/04/09 (H1N1) virus challenge. Additionally, ocular inoculation with poly(I:C) plus vaccine antigen generated no signs of inflammation within 24 hours: no increases in the mRNA expression levels of inflammatory cytokines nor in the infiltration of mononuclear cells to administration sites. In contrast, CT administration induced increased expression of IL-6 cytokine mRNA and mononuclear cell infiltration in the conjunctiva within 24 hours of vaccination. Moreover, inoculated visualizing materials by eyedrop did not contaminate the surface of the olfactory bulb in mice; meanwhile, intranasally administered materials defiled the surface of the brain. On the basis of these findings, we propose that the use of eyedrop inactivated influenza vaccine plus poly(I:C) is a safe and effective mucosal vaccine strategy for inducing protective anti-influenza immunity.

Protection from hemolytic uremic syndrome by eyedrop vaccination with modified enterohemorrhagic E. coli outer membrane vesicles.

We investigated whether eyedrop vaccination using modified outer membrane vesicles (mOMVs) is effective for protecting against hemolytic uremic syndrome (HUS) caused by enterohemorrhagic E. coli (EHEC) O157:H7 infection. Modified OMVs and waaJ-mOMVs were prepared from cultures of MsbB- and Shiga toxin A subunit (STxA)-deficient EHEC O157:H7 bacteria with or without an additional waaJ mutation. BALB/c mice were immunized by eyedrop mOMVs, waaJ-mOMVs, and mOMVs plus polymyxin B (PMB). Mice were boosted at 2 weeks, and challenged peritoneally with wild-type OMVs (wtOMVs) at 4 weeks. As parameters for evaluation of the OMV-mediated immune protection, serum and mucosal immunoglobulins, body weight change and blood urea nitrogen (BUN)/Creatinin (Cr) were tested, as well as histopathology of renal tissue. In order to confirm the safety of mOMVs for eyedrop use, body weight and ocular histopathological changes were monitored in mice. Modified OMVs having penta-acylated lipid A moiety did not contain STxA subunit proteins but retained non-toxic Shiga toxin B (STxB) subunit. Removal of the polymeric O-antigen of O157 LPS was confirmed in waaJ-mOMVs. The mice group vaccinated with mOMVs elicited greater humoral and mucosal immune responses than did the waaJ-mOMVs and PBS-treated groups. Eyedrop vaccination of mOMVs plus PMB reduced the level of humoral and mucosal immune responses, suggesting that intact O157 LPS antigen can be a critical component for enhancing the immunogenicity of the mOMVs. After challenge, mice vaccinated with mOMVs were protected from a lethal dose of wtOMVs administered intraperitoneally, conversely mice in the PBS control group were not. Collectively, for the first time, EHEC O157-derived mOMV eyedrop vaccine was experimentally evaluated as an efficient and safe means of vaccine development against EHEC O157:H7 infection-associated HUS.

Effect of voriconazole and ultraviolet-A combination therapy compared to voriconazole single treatment on Fusarium solani fungal keratitis.

PURPOSE: To demonstrate that ultraviolet-A (UV-A) and voriconazole combination therapy is more effective than voriconazole single treatment for fungal keratitis. METHODS: The in vitro UV-A (375 nm) fungicidal effect was evaluated on Fusarium solani solutions. Each fungal solution was irradiated with different UV-A irradiation doses. Also, a fungal solution containing voriconazole was also irradiated with UV-A. The in vivo therapeutic effect of UV-A and voriconazole treatment was studied in a rabbit keratitis model. Fungi were injected intrastromally into the cornea of 16 rabbits. Each treatment was initiated 3 days after fungal injection and continued up to 8 days for the following groups: Group 1, control; Group 2, treated with UV-A once a day; Group 3, treated with voriconazole 3 times a day; Group 4, treated with voriconazole 3 times a day and UV-A once a day. On the last day, the sclera-cornea buttons were extracted and microbiological and histological evaluations were performed. RESULTS: The colony-forming units (CFUs) of fungal solutions in culture significantly decreased with UV-A irradiation. The CFUs of fungal solutions containing voriconazole also decreased with UV-A irradiation. In vivo, clinical scores of Group 3 (P=0.03) and Group 4 (P=0.02) 5 days after treatment were significantly lower compared to that of Group 1. The clinical score of Group 4 (P=0.03) 5 days after treatment was significantly lower compared to that of Group 3. The histopathological scores 5 days after treatment were significantly lower in Group 4 compared to those of Group 1 (P<0.01) and Group 3 (P=0.02). Based on our CFU analysis, only Group 4 showed significantly lower CFUs compared to Group 1 (P=0.04). CONCLUSIONS: UV-A and voriconazole combination treatment could be a safe and effective alternative to voriconazole single treatment for fungal keratitis.

The prismatic effect on stereoacuity in intermittent exotropia.

PURPOSE: To evaluate the effect of acrylic refractive prism and Fresnel membrane prism on stereoacuity in intermittent exotropia. MATERIALS AND METHODS: Stereoacuities of fifty-two patients (mean age, 12.4 years; range 6 to 45 years) with intermittent exotropia were measured using the Titmus and TNO stereotests, while they wore prisms of varying power on nonfixating eye or evenly on each eye. RESULTS: Stereoacuities were significantly reduced with increasing prism power for both prisms, ranging from 8 to 25 prism dipotres. The effects on stereoacuity in single acrylic prism and single Fresnel prism were similar, whereas spilt Fresnel prisms reduced stereoacuity more than spilt acrylic prisms. Spilt prisms were found to have much less effect on stereoacuity than single prisms for both acrylic and Fresnel prisms. CONCLUSION: The use of acrylic refractive prism shared evenly on each eye would be optimal method to minimize the reduction of stereoacuity during the prismatic therapy for intermittent exotropia.