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(A) Cartoon representation of <t>Lamp2a</t> knockout mouse generation. (B) Confocal microscopy imaging validating deletion of Lamp2a. Red staining reflects Lamp2a, green staining shows Lamp2b. Scale bar =300 µm (C) Immunoblot of RPE/choroid lysate showing loss of Lamp2a expression in Lamp2a -/- mice compared to <t>wild</t> <t>type</t> (WT) age matched mice. (D) FAF images showing high-intensity autofluorescence spots in WT and Lamp2a -/- mice. Lamp2a deficiency results in an age-dependent increase in high-intensity autofluorescence spots compared with WT mice. (E) Quantification of high-intensity autofluorescence spots (n=8). Statistical analysis was performed using Two-way ANOVA with Dunnett’s multiple comparisons test. ***P < 0.001; ****P < 0.0001 . (F) Retinal B-scans of Lamp2a -/- and WT mice. Significant structural disturbance in retinal layers was seen in 12 months Lamp2a -/- mice as compared to WT aged mice and Lamp2a -/- younger mice (n=6). Scale bar =100 µm (width) and 60 = µm (height). Values are expressed as mean ± SD. (G) Representative scotopic ERG response of 6-7-month-old WT and Lamp2a -/- mice. (H) b-wave quantification indicating decline in scotopic response in Lamp2a -/- mice relative to age-matched wild type (n=6 mice/group). (I) Representative scotopic response of 10-12-month-old WT and Lamp2a -/- mice. (J) B-wave quantification shows significant decline in scotopic response in Lamp2a -/- mice relative to age-matched WT (n=8 mice/group). (K) Representative maximal response of 6-7-month-old WT and Lamp2a -/- mice. (L) b-wave amplitude quantification shows decreased scotopic response in 6-7-month-old Lamp2a -/- mice relative to age-matched WT (n=6 mice/group). (M) Representative maximal response of 10-12 months old WT and Lamp2a -/- mice. (N) b-wave amplitude quantification shows significant decrease in scotopic response in 10-12-month-old Lamp2a -/- mice relative to age-matched wild type (n=8 mice/group). (O) Representative photopic response of 6-7-month-old WT and Lamp2a -/- mice. (P) b-wave amplitude quantification shows photopic response changes in 6-7-month-old Lamp2a -/- mice relative to age-matched WT (n=6 mice/group). (Q) Representative photopic response of 10-12-month-old WT and Lamp2a -/- mice. (R) b-wave amplitude quantification shows pronounced decrease in photopic response in 10-12-month-old Lamp2a -/- mice relative to age-matched WT (n=8 mice/group). Statistical differences were analyzed using an unpaired two-tailed student’s t-test. Level of significance was considered at p value < 0.05 .
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(A) Cartoon representation of <t>Lamp2a</t> knockout mouse generation. (B) Confocal microscopy imaging validating deletion of Lamp2a. Red staining reflects Lamp2a, green staining shows Lamp2b. Scale bar =300 µm (C) Immunoblot of RPE/choroid lysate showing loss of Lamp2a expression in Lamp2a -/- mice compared to <t>wild</t> <t>type</t> (WT) age matched mice. (D) FAF images showing high-intensity autofluorescence spots in WT and Lamp2a -/- mice. Lamp2a deficiency results in an age-dependent increase in high-intensity autofluorescence spots compared with WT mice. (E) Quantification of high-intensity autofluorescence spots (n=8). Statistical analysis was performed using Two-way ANOVA with Dunnett’s multiple comparisons test. ***P < 0.001; ****P < 0.0001 . (F) Retinal B-scans of Lamp2a -/- and WT mice. Significant structural disturbance in retinal layers was seen in 12 months Lamp2a -/- mice as compared to WT aged mice and Lamp2a -/- younger mice (n=6). Scale bar =100 µm (width) and 60 = µm (height). Values are expressed as mean ± SD. (G) Representative scotopic ERG response of 6-7-month-old WT and Lamp2a -/- mice. (H) b-wave quantification indicating decline in scotopic response in Lamp2a -/- mice relative to age-matched wild type (n=6 mice/group). (I) Representative scotopic response of 10-12-month-old WT and Lamp2a -/- mice. (J) B-wave quantification shows significant decline in scotopic response in Lamp2a -/- mice relative to age-matched WT (n=8 mice/group). (K) Representative maximal response of 6-7-month-old WT and Lamp2a -/- mice. (L) b-wave amplitude quantification shows decreased scotopic response in 6-7-month-old Lamp2a -/- mice relative to age-matched WT (n=6 mice/group). (M) Representative maximal response of 10-12 months old WT and Lamp2a -/- mice. (N) b-wave amplitude quantification shows significant decrease in scotopic response in 10-12-month-old Lamp2a -/- mice relative to age-matched wild type (n=8 mice/group). (O) Representative photopic response of 6-7-month-old WT and Lamp2a -/- mice. (P) b-wave amplitude quantification shows photopic response changes in 6-7-month-old Lamp2a -/- mice relative to age-matched WT (n=6 mice/group). (Q) Representative photopic response of 10-12-month-old WT and Lamp2a -/- mice. (R) b-wave amplitude quantification shows pronounced decrease in photopic response in 10-12-month-old Lamp2a -/- mice relative to age-matched WT (n=8 mice/group). Statistical differences were analyzed using an unpaired two-tailed student’s t-test. Level of significance was considered at p value < 0.05 .
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(A) Cartoon representation of Lamp2a knockout mouse generation. (B) Confocal microscopy imaging validating deletion of Lamp2a. Red staining reflects Lamp2a, green staining shows Lamp2b. Scale bar =300 µm (C) Immunoblot of RPE/choroid lysate showing loss of Lamp2a expression in Lamp2a -/- mice compared to wild type (WT) age matched mice. (D) FAF images showing high-intensity autofluorescence spots in WT and Lamp2a -/- mice. Lamp2a deficiency results in an age-dependent increase in high-intensity autofluorescence spots compared with WT mice. (E) Quantification of high-intensity autofluorescence spots (n=8). Statistical analysis was performed using Two-way ANOVA with Dunnett’s multiple comparisons test. ***P < 0.001; ****P < 0.0001 . (F) Retinal B-scans of Lamp2a -/- and WT mice. Significant structural disturbance in retinal layers was seen in 12 months Lamp2a -/- mice as compared to WT aged mice and Lamp2a -/- younger mice (n=6). Scale bar =100 µm (width) and 60 = µm (height). Values are expressed as mean ± SD. (G) Representative scotopic ERG response of 6-7-month-old WT and Lamp2a -/- mice. (H) b-wave quantification indicating decline in scotopic response in Lamp2a -/- mice relative to age-matched wild type (n=6 mice/group). (I) Representative scotopic response of 10-12-month-old WT and Lamp2a -/- mice. (J) B-wave quantification shows significant decline in scotopic response in Lamp2a -/- mice relative to age-matched WT (n=8 mice/group). (K) Representative maximal response of 6-7-month-old WT and Lamp2a -/- mice. (L) b-wave amplitude quantification shows decreased scotopic response in 6-7-month-old Lamp2a -/- mice relative to age-matched WT (n=6 mice/group). (M) Representative maximal response of 10-12 months old WT and Lamp2a -/- mice. (N) b-wave amplitude quantification shows significant decrease in scotopic response in 10-12-month-old Lamp2a -/- mice relative to age-matched wild type (n=8 mice/group). (O) Representative photopic response of 6-7-month-old WT and Lamp2a -/- mice. (P) b-wave amplitude quantification shows photopic response changes in 6-7-month-old Lamp2a -/- mice relative to age-matched WT (n=6 mice/group). (Q) Representative photopic response of 10-12-month-old WT and Lamp2a -/- mice. (R) b-wave amplitude quantification shows pronounced decrease in photopic response in 10-12-month-old Lamp2a -/- mice relative to age-matched WT (n=8 mice/group). Statistical differences were analyzed using an unpaired two-tailed student’s t-test. Level of significance was considered at p value < 0.05 .

Journal: bioRxiv

Article Title: Loss of Lamp2a-dependent chaperone-mediated autophagy drives dry AMD-like retinal pathology in mice and is rescued by BK channel activation

doi: 10.64898/2026.03.19.712761

Figure Lengend Snippet: (A) Cartoon representation of Lamp2a knockout mouse generation. (B) Confocal microscopy imaging validating deletion of Lamp2a. Red staining reflects Lamp2a, green staining shows Lamp2b. Scale bar =300 µm (C) Immunoblot of RPE/choroid lysate showing loss of Lamp2a expression in Lamp2a -/- mice compared to wild type (WT) age matched mice. (D) FAF images showing high-intensity autofluorescence spots in WT and Lamp2a -/- mice. Lamp2a deficiency results in an age-dependent increase in high-intensity autofluorescence spots compared with WT mice. (E) Quantification of high-intensity autofluorescence spots (n=8). Statistical analysis was performed using Two-way ANOVA with Dunnett’s multiple comparisons test. ***P < 0.001; ****P < 0.0001 . (F) Retinal B-scans of Lamp2a -/- and WT mice. Significant structural disturbance in retinal layers was seen in 12 months Lamp2a -/- mice as compared to WT aged mice and Lamp2a -/- younger mice (n=6). Scale bar =100 µm (width) and 60 = µm (height). Values are expressed as mean ± SD. (G) Representative scotopic ERG response of 6-7-month-old WT and Lamp2a -/- mice. (H) b-wave quantification indicating decline in scotopic response in Lamp2a -/- mice relative to age-matched wild type (n=6 mice/group). (I) Representative scotopic response of 10-12-month-old WT and Lamp2a -/- mice. (J) B-wave quantification shows significant decline in scotopic response in Lamp2a -/- mice relative to age-matched WT (n=8 mice/group). (K) Representative maximal response of 6-7-month-old WT and Lamp2a -/- mice. (L) b-wave amplitude quantification shows decreased scotopic response in 6-7-month-old Lamp2a -/- mice relative to age-matched WT (n=6 mice/group). (M) Representative maximal response of 10-12 months old WT and Lamp2a -/- mice. (N) b-wave amplitude quantification shows significant decrease in scotopic response in 10-12-month-old Lamp2a -/- mice relative to age-matched wild type (n=8 mice/group). (O) Representative photopic response of 6-7-month-old WT and Lamp2a -/- mice. (P) b-wave amplitude quantification shows photopic response changes in 6-7-month-old Lamp2a -/- mice relative to age-matched WT (n=6 mice/group). (Q) Representative photopic response of 10-12-month-old WT and Lamp2a -/- mice. (R) b-wave amplitude quantification shows pronounced decrease in photopic response in 10-12-month-old Lamp2a -/- mice relative to age-matched WT (n=8 mice/group). Statistical differences were analyzed using an unpaired two-tailed student’s t-test. Level of significance was considered at p value < 0.05 .

Article Snippet: Adult male Lamp2a knockout mice ( Lamp2a y/- ), female heterozygous Lamp2a +/- and their wild type (WT) littermates ( Lamp2a y/+ ) were obtained from Cyagen (Serial Number KOCMP-16784-Lamp2-B6J-VA; strain C57BL/6J-Lamp2 em1Cya ).

Techniques: Knock-Out, Confocal Microscopy, Imaging, Staining, Western Blot, Expressing, Two Tailed Test