Journal: Aging Cell

Article Title: Nicotinamide Riboside Supplementation Alleviates Testicular Aging Induced by Disruption of Qprt‐Dependent NAD + De Novo Synthesis in Mice

doi: 10.1111/acel.70004

Figure Lengend Snippet: Qprt deletion led to a reduction in testicular germ cell numbers with aging in mouse testis. (a) Representative immunofluorescence staining for VASA (red) with DAPI (blue) counterstaining in the testes of WT and Qprt −/− mice at 3, 6, and 9 months of age. Scale bar = 100 μm. (b) Quantification of VASA‐positive cells per tubule at 3, 6, and 9 months of age ( n = 61 convoluted seminiferous tubules from 3 mice per group). (c) Representative TUNEL staining (green) with DAPI counterstaining (blue) in testis sections from WT and Qprt −/− mice at 3, 6, and 9 months of age. Scale bar = 100 μm. (d) The average number of apoptotic cells per seminiferous tubule at 3, 6, and 9 months ( n = 3–4 mice per group; at least 50 seminiferous tubules were counted per mouse in each group). * p < 0.05, ** p < 0.01, *** p < 0.001. Data are presented as means ± SEM.

Article Snippet: Add the primary antibodies of QPRT (1:100, Proteintech) or VASA (1:100, Abcam) to the sections and incubate overnight at 4°C.

Techniques: Immunofluorescence, Staining, TUNEL Assay

Journal: Aging Cell

Article Title: Nicotinamide Riboside Supplementation Alleviates Testicular Aging Induced by Disruption of Qprt‐Dependent NAD + De Novo Synthesis in Mice

doi: 10.1111/acel.70004

Figure Lengend Snippet: NR supplementation increased testicular NAD + levels and decreased spermatogenesis defects in Qprt −/− mice. (a, b) NAD + content in pachytene spermatocytes (a) and testis (b) from 6‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice. ( n = 3 mice per group). (c) Testis weights in 6‐ and 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice ( n = 6 mice per group). (d) Sperm count in 6‐ and 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice ( n = 6 mice per group). (e) Representative hematoxylin and eosin (H&E) staining of testicular sections from 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice, showing seminiferous tubule structure. Scale bar = 100 μm. (f) Measurement of seminiferous tubule diameter in Qprt −/− and NR‐supplemented Qprt −/− mice at 9 months ( n = 65 seminiferous tubules from 3 mice per group). (g) Representative immunofluorescence images of testicular sections from 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice stained for VASA (red) and DAPI (blue). Scale bar = 100 μm. (h) Quantification of VASA‐positive cells per seminiferous tubule in the testes of 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice ( n = 65 seminiferous tubules from 3 mice per group). (i) TUNEL staining (green) with DAPI counterstaining (blue) in testicular sections from 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice. Scale bar = 100 μm. (j) Quantification of apoptotic cells per seminiferous tubule in 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice ( n = 3 mice per group). (k) Immunofluorescence staining of SYCP3 (red) and γH2AX (green) in spermatocytes at different stages of prophase I from Qprt −/− and NR‐supplemented Qprt −/− mice. Scale bar = 10 μm. (l) Percentage of spermatocytes at different prophase I stages in 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice ( n = 3 mice per group). (m) Percentage of γH2AX‐positive pachytene spermatocytes in 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice ( n = 3 mice per group). (n, o) Immunofluorescence staining of SYCP3 (red) and Rad51 (green) in pachytene spermatocytes from 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice, and quantification of Rad51 foci per cell ( n = 46–50 pachytene spermatocytes per group). Scale bar = 10 μm. (p, q) Representative images of immunofluorescence staining of SYCP3 (red) and MLH1 (green) in pachytene spermatocytes from 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice, and quantification of MLH1 foci per cell ( n = 41–42 pachytene spermatocytes per group). Scale bar = 10 μm. (r, s) Representative images of immunofluorescence staining of SYCP3 (red) and RNA pol II (green) in pachytene spermatocytes from 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− mice, with insets showing magnified XY body regions. Relative RNA pol II signal intensity in the XY body ( n = 67 pachytene spermatocytes per group). Scale bar = 10 μm. (t) Relative mRNA expression levels of X‐ and Y‐linked genes in 9‐month‐old Qprt −/− and NR‐supplemented Qprt −/− spermatocytes, as determined by qRT‐PCR ( n = 9 mice per group). * p < 0.05, ** p < 0.01, *** p < 0.001. Data are presented as means ± SEM.

Article Snippet: Add the primary antibodies of QPRT (1:100, Proteintech) or VASA (1:100, Abcam) to the sections and incubate overnight at 4°C.

Techniques: Staining, Immunofluorescence, TUNEL Assay, Expressing, Quantitative RT-PCR