METHODOLOGY: Sprague-Dawley rats were divided into 5 groups of 33 each. Group 1 was administered intravitreally with PBS and group 2 was similarly injected with NMDA (160 nmol). Groups 3, 4 and 5 were injected with TAU (320 nmol) 24 hours before (pre-treatment), in combination (co-treatment) and 24 hours after (post-treatment) NMDA exposure respectively. Seven days after injection, rats were sacrificed; eyes were enucleated, fixed and processed for morphometric analysis, TUNEL and caspase-3 staining. Optic nerve morphology assessment was done using toluidine blue staining. The estimation of BDNF, pro/anti-apoptotic factors (Bax/Bcl-2) and caspase-3 activity in retina was done using ELISA technique.
RESULTS: Severe degenerative changes were observed in retinae after intravitreal NMDA exposure. The retinal morphology in the TAU pre-treated group appeared more similar to the control retinae and demonstrated a higher number of nuclei than the NMDA group both per 100 μm length (by 1.5-fold, p
METHODS: Sprague Dawley rats, 180-250 g in weight were divided into four groups. Groups 1, 2, 3 and 4 were intravitreally administered with vehicle and NMDA at the doses 80, 160 and 320 nmol respectively. Seven days after injection, rats were euthanized, and their eyes were taken for optic nerve toluidine blue and retinal hematoxylin and eosin stainings. The TUNEL assay was done for detecting apoptotic cells.
RESULTS: All groups treated with NMDA showed significantly reduced ganglion cell layer (GCL) thickness within inner retina, as compared to control group. Group NMDA 160 nmol showed a significantly greater GCL thickness than the group NMDA 320 nmol. Administration of NMDA also resulted in a dose-dependent decrease in the number of nuclei both per 100 µm GCL length and per 100 µm2 of GCL. Intravitreal NMDA injection caused dose-dependent damage to the optic nerve. The degeneration of nerve fibres with increased clearing of cytoplasm was observed more prominently as the NMDA dose increased. In accordance with the results of retinal morphometry analysis and optic nerve grading, TUNEL staining demonstrated NMDA-induced excitotoxic retinal injury in a dose-dependent manner.
CONCLUSION: Our results demonstrate dose-dependent effects of NMDA on retinal and optic nerve morphology in rats that may be attributed to differences in the severity of excitotoxicity and oxidative stress. Our results also suggest that care should be taken while making dose selections experimentally so that the choice might best uphold study objectives.
MATERIALS AND METHODS: Five groups of rats were intravitreally administered with vehicle or Aβ(1-40) in doses of 1.0, 2.5, 5 and 10 nmol. Animals were sacrificed and eyes were enucleated at weeks 1, 2 and 4 post-injection. The retinae were subjected to morphometric analysis and TUNEL staining. Optic nerve sections were stained with toluidine blue and were graded for neurodegenerative effects. The estimation of BDNF and markers of oxidative stress in retina were done using ELISA technique.
RESULTS AND CONCLUSIONS: It was observed that intravitreal Aβ(1-40) causes significant retinal and optic nerve damage up to day 14 post-injection and there was increasing damage with increase in dose. However, on day 30 post-injection both the retinal and optic nerve morphology showed a trend towards normalization. The observations made for retinal cell apoptosis, retinal glutathione, superoxide dismutase activity and BDNF were in accordance with those of morphological changes with deterioration till day 14 and recovery by day 30 post-injection. The findings of this study may provide a guide for selection of appropriate experimental conditions for future studies.
METHODS: Sprague Dawley rats were intravitreally injected with ET1. MgAT and TAU were administered as pre-, co-, or posttreatment. Subsequently, the expression of NOS isoforms was detected in retina by immunohistochemistry, retinal nitrotyrosine level was estimated using ELISA, and retinal cell apoptosis was detected by TUNEL staining.
RESULTS: Intravitreal ET1 caused a significant increase in the expressions of nNOS and iNOS while eNOS expression was significantly reduced compared to vehicle treated group. Administration of both MgAT and TAU restored the altered levels of NOS isoform expression, reduced retinal nitrosative stress and retinal cell apoptosis. The effect of MgAT, however, was greater than that of TAU alone.
CONCLUSIONS: MgAT and TAU prevent ET1-induced retinal cell apoptosis by reducing retinal nitrosative stress in Sprague Dawley rats. Addition of TAU to Mg seems to enhance the efficacy of TAU compared to when given alone. Moreover, the pretreatment with MgAT/TAU showed higher efficacy compared to co- or posttreatment.
METHODS: We investigated the ocular permeation of topical tazocin after single drop application in normal rabbit eyes by estimating piperacillin and tazobactam concentrations in cornea, aqueous, and vitreous using a validated LC-MS/MS method. Furthermore, we determined the efficacy of repeated dose administration of tazocin against experimentally induced P. aeruginosa keratitis in rabbits in comparison to moxifloxacin. To determine the efficacy, clinical examination, histopathological examination, and estimation of bacterial load and inflammatory cytokines in cornea were done.
RESULTS: Significant corneal concentration of piperacillin and tazobactam was detected in normal rabbit corneas after single dose treatment with tazocin. In rabbits with Pseudomonas-induced keratitis, topical tazocin caused significant clinical and histopathological improvement. This improvement was associated with reduction in corneal bacterial load and inflammatory cytokines. Compared to moxifloxacin 0.5%, tazocin treated group showed greater clinical response which was associated with higher interleukin (IL)-1β, lower tumor necrosis factor (TNF)-α, a comparable level of IL-8, greater reduction in corneal bacterial load, and lesser inflammatory cell infiltration.
CONCLUSION: Tazocin showed good ocular penetration and was effective in treatment of Pseudomonas induced keratitis in rabbits.
Methods: Excitotoxic retinal injury was induced with intravitreal injection of NMDA in Sprague-Dawley rats. All treatments were given as pre-, co-, and post-treatment with NMDA. Seven days post-injection, the retinas were processed for measurement of the expression of NOS isoforms using immunostaining and enzyme-linked immunosorbent assay (ELISA), retinal 3-NT content using ELISA, retinal histopathological changes using hematoxylin and eosin (H&E) staining, and retinal cell apoptosis using terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) staining.
Results: As observed on immunohistochemistry, the treatment with NMDA caused a 4.53-fold increase in retinal nNOS expression compared to the PBS-treated rats (p<0.001). Among the MgAT-treated groups, only the pretreatment group showed significantly lower nNOS expression than the NMDA-treated group with a 2.00-fold reduction (p<0.001). Among the TAU-treated groups, the pre- and cotreatment groups showed 1.84- and 1.71-fold reduction in nNOS expression compared to the NMDA-treated group (p<0.001), respectively, but remained higher compared to the PBS-treated group (p<0.01). Similarly, iNOS expression in the NMDA-treated group was significantly greater than that for the PBS-treated group (2.68-fold; p<0.001). All MgAT treatment groups showed significantly lower iNOS expression than the NMDA-treated groups (3.58-, 1.51-, and 1.65-folds, respectively). However, in the MgAT co- and post-treatment groups, iNOS expression was significantly greater than in the PBS-treated group (1.77- and 1.62-folds, respectively). Pretreatment with MgAT caused 1.77-fold lower iNOS expression compared to pretreatment with TAU (p<0.05). In contrast, eNOS expression was 1.63-fold higher in the PBS-treated group than in the NMDA-treated group (p<0.001). Among all treatment groups, only pretreatment with MgAT caused restoration of retinal eNOS expression with a 1.39-fold difference from the NMDA-treated group (p<0.05). eNOS expression in the MgAT pretreatment group was also 1.34-fold higher than in the TAU pretreatment group (p<0.05). The retinal NOS expression as measured with ELISA was in accordance with that estimated with immunohistochemistry. Accordingly, among the MgAT treatment groups, only the pretreated group showed 1.47-fold lower retinal 3-NT than the NMDA-treated group, and the difference was significant (p<0.001). The H&E-stained retinal sections in all treatment groups showed statistically significantly greater numbers of retinal cell nuclei than the NMDA-treated group in the inner retina. However, the ganglion cell layer thickness in the TAU pretreatment group remained 1.23-fold lower than that in the MgAT pretreatment group (p<0.05). In line with this observation, the number of apoptotic cells as observed after TUNEL staining was 1.69-fold higher after pretreatment with TAU compared to pretreatment with MgAT (p<0.01).
Conclusions: MgAT and TAU, particularly with pretreatment, reduce retinal cell apoptosis by reducing retinal nitrosative stress. Pretreatment with MgAT caused greater improvement in NMDA-induced changes in iNOS and eNOS expression and retinal 3-NT levels than pretreatment with TAU. The greater reduction in retinal nitrosative stress after pretreatment with MgAT was associated with lower retinal cell apoptosis and greater preservation of the ganglion cell layer thickness compared to pretreatment with TAU.