Abstract
Diabetic retinopathy (DR) is a retinal microvascular disease characterized by inflammatory and angiogenic pathways. In this study, we evaluated NLRP3 inflammasome in a double transgenic mouse model, Akimba (Ins2AkitaxVEGF+/−), which demonstrates hyperglycemia, vascular hyperpermeability and neovascularization seen in the proliferative DR. Retinal structural integrity, vascular leakage and function were examined by fundus photography, fluorescein angiography, optical coherence tomography, retinal flat mounts, laser speckle flowgraphy (LSFG), and electroretinography in Akimba and its parental strains, Akita (Ins2Akita) and Kimba (trVEGF029) mice. Inflammatory mechanisms involving NLRP3 inflammasome were investigated using real time-PCR, immunohistochemistry, ELISA and western blots. We observed an increased vascular leakage, reduced retinal thickness, and function in Akimba retina. Also, Akimba retina depicts decreased relative flow volume measured by LSFG. Most importantly, high levels of IL-1β along with increased NLRP3, ASC, and Caspase-1 at mRNA and protein levels were observed in Akimba retina. However, the in vivo functional role remains undefined. In conclusion, increased activation of macroglia (GFAP), microglia (Iba-1 and OX-42) and perivascular macrophages (F4/80 and CD14) together with pro-inflammatory (IL-1β and IL-6) and pro-angiogenic markers (PECAM-1, ICAM-1, VEGF, Flt-1, and Flk-1), suggested a critical role for NLRP3 inflammasome in the Akimba mouse model depicting advanced stages of DR pathogenesis.
Highlights
Of pro-inflammatory cytokines such as C-reactive protein (CRP), Interleukin-1beta (IL-1β), Interleukin-6 (IL-6) and Tumour Necrosis Factor-alpha (TNF-α) have been demonstrated in Diabetic retinopathy (DR) and PDR8,9
We studied other angiogenic mediators such as platelet endothelial cell adhesion molecular (PECAM)-1 and intracellular adhesion molecule (ICAM)-1, both known to play a critical role in pathological retinal neovascularization
A double transgenic Akimba mouse model to study the advanced stages of DR has been characterized
Summary
Of pro-inflammatory cytokines such as C-reactive protein (CRP), Interleukin-1beta (IL-1β), Interleukin-6 (IL-6) and Tumour Necrosis Factor-alpha (TNF-α) have been demonstrated in DR and PDR8,9. The DAMPs such as oxidized lipoproteins, glycated proteins, uric acid, DNA and RNA from necrotic cells have been suggested to contribute to the pathogenesis of retinal diseases[16,17] This may compromise the first line of defense system embodied by the innate immune system and protected by the blood-retina barrier (BRB) as seen in PDR and DME18. Advanced proliferative and neovascular changes were not observed in these rodents eyes even after 18–24 months of diabetes[25,26] Mouse models such as Akita (Ins2Akita) and Kimba (trVEGF029) have been described in DR studies. The Kimba mouse, which has transient photoreceptor (PhR)-specific overexpression of human VEGF165 (hVEGF), depicts a less destructive and slower form of retinal neovascularization present in the advanced stages of DR28 This model has several vascular abnormalities seen in DR, it lacks hyperglycemic background. Since inflammatory pathways contribute significantly to DR, we designed this study to evaluate the role of the NLRP3 inflammasome in Akimba retina to better understand the disease pathogenesis in the advanced stages of DR
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