Fig 2: Activation of SARM1 results in NAD depletion and mediates photoreceptor cell death.(A) Schematic diagram of the SARM1 plasmid constructs used for transfection. (B, C, D) MTS metabolic assay, (C) LDH cytotoxicity assay and (D) bright-field images (10× magnification) of 661W cells transfected with 1 μg/ml of empty vector (EV), full-length SARM1 (FL), and dN190-SARM1 (dN190) for 24 h (*P ≤ 0.05, ***P ≤ 0.001 by t test, n = 3). (E) Graphical representation of retinal explant preparation. (F) NADH fluorescent lifetime imaging (FLIM) quantification in WT and sarm1−/− retinal explants treated with 50 μM CCCP for 24 h, showing the relative fold values when normalized to vehicle-treated WT and sarm1−/− retinal explants, respectively (**P ≤ 0.01 by t test, n = 4). (G) Colour-coded FLIM images of the retinal explants from WT and sarm1−/− retinal explants treated with 50 μM CCCP for 24 h.

Fig 3: Optical coherence tomography (OCT) analysis is comparable between SARM1-deficient and WT mice.Linked to Fig 4. (A, B) OCT images taken in vivo from WT and sarm1−/− mice at 6 and 12 wk of age with (B) quantification of the ONL to outer segment (OS) width (marked with red line in the OCT images) using ImageJ (n = 4 mice per group).

Fig 4: Loss of SARM1 preserves photoreceptor number and length in the NaIO3 model of retinal degeneration, following severe RPE degradation. Representative images of paraffin embedded retinal sections from WT and Sarm1–/– mice 7-days post administration of NaIO3 stained with haematoxylin & eosin (H&E) (A). Quantification of the number of nuclei per ONL row (B), number of nuclei in the ONL in an area of height 450 pixels in the centre of the ONL in each image (C), and ONL area (D) (*P ≤ 0.05, **P ≤ 0.01, n = 4 WT NaIO3 mice, n = 4 Sarm1–/– NaIO3 mice). Optical coherence tomography (OCT) images taken in vivo from C57BL/6J and Sarm1–/– mice 7-days post administration of NaIO3 (50 mg/kg) (E). Quantification of the distance of the ONL to the outer segments (OS) (marked with red line on OCT images) InSight (*P ≤ 0.05, by Student’s unpaired t test, n = 4 WT saline mice, n = 3 Sarm1–/– saline mice, n = 4 WT NaIO3 mice, n = 4 Sarm1–/– mice) (F).

Fig 5: SARM1 is expressed in the photoreceptor cells of the retina.(A) Data extracted from Human proteome map showing relative expression of SARM1, Rhodopsin (RHO), and RPE65 in different tissues. (B) qRT-PCR analysis of SARM1 transcript levels in the neural retina and RPE/choroid of wild-type C57BL/6J mice (**P ≤ 0.01, by t test, n = 5 mice). (C) Schematic of a wild-type and rho−/− retina over time. (D) Haemotoxylin and eosin staining of paraffin-embedded sections from wild-type and rho−/− mice on the C57BL/6J background at 12 wk of age (40× magnification). (E) qRT-PCR analysis of SARM1 transcript levels in the neural retina of wild-type and rho−/− mice on the C57BL/6J background at 12 wk of age (***P ≤ 0.001, by t test, n = 6 mice). (F, G) Western blot analysis and (G) quantification of SARM1 expression in neural retina of wild-type n = 5, rho−/− n = 5, and sarm1−/− n = 3 mice (***P ≤ 0.001, by t test).