Ischemia-Reperfusion Injury Model
Objective: To assess VEGF-A's neuroprotective effects and dose-dependent reduction in retinal neuron apoptosis using an ischemia-reperfusion injury model
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Equipment2
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Protocol Steps
Ischemia-reperfusion injury induction
Induce ischemic injury to the retina using the ischemia-reperfusion injury model
Note: Model used to assess VEGF-A's neuroprotective effects
View evidence from paper
“we used a model of ischemia-reperfusion injury and found that VEGF-A exposure resulted in a dose-dependent reduction in retinal neuron apoptosis”
VEGF-A exposure at varying doses
Expose retinal tissue to dose-dependent amounts of VEGF-A
Note: Dose-dependent reduction in retinal neuron apoptosis was observed
View evidence from paper
“VEGF-A exposure resulted in a dose-dependent reduction in retinal neuron apoptosis”
Mechanistic assessment of blood flow
Measure volumetric blood flow to the retina to assess contribution to neuroprotection
Note: VEGF-A-induced volumetric blood flow may be partially responsible for neuroprotection
View evidence from paper
“VEGF-A-induced volumetric blood flow to the retina may be partially responsible for the neuroprotection”
Ex vivo retinal culture assessment
Culture retinal tissue ex vivo to demonstrate direct neuroprotective effect independent of blood flow
Note: Direct neuroprotective effect of VEGF-A demonstrated in culture
View evidence from paper
“ex vivo retinal culture demonstrated a direct neuroprotective effect for VEGF-A”
VEGFR2 expression detection
Detect VEGF receptor-2 expression in retinal neuronal cell layers
Note: VEGFR2 expression found in several neuronal cell layers
View evidence from paper
“VEGF receptor-2 (VEGFR2) expression was detected in several neuronal cell layers of the retina”
Functional analysis of VEGFR2
Perform functional analyses to determine VEGFR2 involvement in retinal neuroprotection
Note: VEGFR2 shown to be involved in retinal neuroprotection
View evidence from paper
“functional analyses showed that VEGFR2 was involved in retinal neuroprotection”
Ischemic preconditioning protocol
Apply ischemic preconditioning 24 hours before ischemia-reperfusion injury
Note: Increased VEGF-A levels and substantially decreased apoptotic retinal cells
View evidence from paper
“Ischemic preconditioning 24 hours before ischemia-reperfusion injury increased VEGF-A levels and substantially decreased the number of apoptotic retinal cells”
VEGF-A inhibition during preconditioning
Inhibit VEGF-A function during ischemic preconditioning to assess its role in adaptive response
Note: Protective effect of preconditioning was reversed by VEGF-A inhibition
View evidence from paper
“The protective effect of ischemic preconditioning was reversed after VEGF-A inhibition”
Chronic VEGF-A inhibition in normal animals
Chronically inhibit VEGF-A function in normal adult animals to assess effects on retinal ganglion cells
Note: Significant loss of retinal ganglion cells observed with no effect on vascular parameters
View evidence from paper
“chronic inhibition of VEGF-A function in normal adult animals led to a significant loss of retinal ganglion cells yet had no observable effect on several vascular parameters”