Nod cb17 prkdescid j

All animal studies were conducted in accordance with the Guide for the Care and Use of Animals (National Academy Press) and were approved by the University of Pittsburgh Animal Care and Use Committee (Protocol # 20108281). Both male and female mice were used for this study [17 (link),21 (link)]. RPE-specific Akt2 KI were generated as described previously [22 (link)]. The RPE-specific Akt2 KI mice were generated by Cyagen. Briefly, the T2A sequence followed by Akt2 coding sequence (CDS) was inserted between the last exon and the 3′ untranslated region (3′UTR) of the mouse Best1 gene, which in the eye is specifically expressed in the RPE. The Neo cassette flanked with self-deletion anchors (SDA) was inserted in the intron area between exons 11 and 12 of the mouse Best1 for germ cell deletion of the gene. βA3/A1-crystallin conditional (Cryba1 cKO) [17 (link),19 (link),20 (link)] and complete knockout (Cryba1 KO) [16 (link),21 (link)] mice were also generated and maintained as previously described [16 (link),17 (link),19 (link),20 (link),21 (link),22 (link)]. Nonobese diabetic/severe combined immunodeficiency (NOD-SCID) mice (NOD.CB17-Prkdescid/J; 5 weeks old) were purchased from The Jackson Laboratory, USA. All mice used in this study were RD8 negative.

NOD-SCID mice (NOD.CB17-Prkdescid/J, Jackson Laboratories, USA, male, 4–5-weeks-old) were used for the study. A large sample size, n = 10, was taken to nullify any experimental anomaly. Mice were anaesthetized and sub-retinal injections of neutrophils from different experimental groups or recombinant LCN-2 protein were given as described earlier^{73 (link)}. Seven days after treatment, the NOD-SCID mice were anaesthetized by intraperitoneal injection of a ketamine and xylazine mixture and then subjected to Fundus imaging along with OCT analysis using the Bioptigen Envisu R2210 system. OCT images were analyzed on optical sections (100 sections per retina) from each eye ranging from−2.0 to +2.0 mm with respect to the optic nerve head (ONH) using the FIJI-ImageJ (NIH) plugin provided with the instrument along with Diver 2.4 software (Bioptigen). After the experiment, the animals were euthanized with CO₂ gas and the eyes were harvested for further experiments.