Analysis Tools
reproductive system
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• 8- to 12-week-old females crossed with wild-type B6 males produce a normal number of offspring with no difference in average litter size relative to wild-type or heterozygous controls
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• number of germ cells is significantly decreased in a portion of seminiferous tubules; this is more pronounced at 30 weeks than at 15 weeks of age
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• after 90 min of treatment with 10 uM apoptosis inducer ABT-737 (a BH3 mimetic) and staining with Cy5-annexin V, testicular germ cells from 3-week-old mice fail to show induction of phosphatidylserine (PtdSer) exposure, whereas >80% of similarly treated heterozygous germ cells show PtdSer exposure after 60 min
• when germ cells are purified by density gradient centrifugation, treated with 5 uM ABT-737 and stained with Fluor 488-annexin V and propidium iodide (PI), PtdSer-positive cells are rarely detected on the cell surface, and, when detected, cells are PI-positive, indicating dead cells that have already undergone secondary necrosis
• however, caspase 3 is normally activated upon 10 uM ABT-737 treatment and cleavage of PARP1 [poly (ADP-ribose) polymerase family, member 1, a substrate of caspase 3] is noted at a level similar to that seen with heterozygous male germ cells
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• at 2 to 4 weeks of age, 67% of DAPI-positive cells in the testes stain positive by TUNEL versus only ~2% in heterozygous testes
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• seminiferous tubules appear abnormal and contain aggregated cells and Sertoli cells with many vacuoles
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• Sertoli cells with many large vacuoles are observed at 15 and 30 weeks of age
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• at 8 weeks of age, aggregated vimentin-positive and Wilms tumor 1 homolog (WT1)-positive Sertoli cells are noted in the lumen of seminiferous tubules
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small testis
(
J:293092
)
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• at 10-weeks of age, testes are significantly smaller than those of wild-type males
• however, neither inflammatory cells nor inflammatory cytokines are observed in the testes
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• average weight of the testes is normal until 5 weeks of age but significantly lower than that of heterozygous males at 10, 15 and 30 weeks of age
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• at 14 to 17 weeks of age, cauda epididymal sperm counts are less than 25% of those in heterozygous males
• however, sperm morphology is similar to that of heterozygous sperm
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• whereas most dead cells in heterozygous control testes are inside Sertoli cells and their nuclei are swollen and degenerated, dead cells are abnormally found outside Sertoli cells and contain condensed nuclei, indicating the presence of unengulfed apoptotic cells
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• at 30 weeks of age, cauda epididymides exhibit more severe reductions in sperm number than at 15 weeks and contain numerous amorphous eosinophilic materials, likely representing debris of dead cells or residual bodies released from maturing sperm
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• in 2 out of 12 cases, average litter size obtained with males is extremely low (0.7) relative to that obtained with heterozygous males (12.6)
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• 10- to 12-week-old males crossed with highly prolific 9-week-old wild-type ICR females produce no offspring in 10 out of 12 challenges
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cellular
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• number of germ cells is significantly decreased in a portion of seminiferous tubules; this is more pronounced at 30 weeks than at 15 weeks of age
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• at 14 to 17 weeks of age, cauda epididymal sperm counts are less than 25% of those in heterozygous males
• however, sperm morphology is similar to that of heterozygous sperm
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• after 90 min of treatment with 10 uM apoptosis inducer ABT-737 (a BH3 mimetic) and staining with Cy5-annexin V, testicular germ cells from 3-week-old mice fail to show induction of phosphatidylserine (PtdSer) exposure, whereas >80% of similarly treated heterozygous germ cells show PtdSer exposure after 60 min
• when germ cells are purified by density gradient centrifugation, treated with 5 uM ABT-737 and stained with Fluor 488-annexin V and propidium iodide (PI), PtdSer-positive cells are rarely detected on the cell surface, and, when detected, cells are PI-positive, indicating dead cells that have already undergone secondary necrosis
• however, caspase 3 is normally activated upon 10 uM ABT-737 treatment and cleavage of PARP1 [poly (ADP-ribose) polymerase family, member 1, a substrate of caspase 3] is noted at a level similar to that seen with heterozygous male germ cells
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• at 2 to 4 weeks of age, 67% of DAPI-positive cells in the testes stain positive by TUNEL versus only ~2% in heterozygous testes
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endocrine/exocrine glands
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• seminiferous tubules appear abnormal and contain aggregated cells and Sertoli cells with many vacuoles
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• Sertoli cells with many large vacuoles are observed at 15 and 30 weeks of age
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• at 8 weeks of age, aggregated vimentin-positive and Wilms tumor 1 homolog (WT1)-positive Sertoli cells are noted in the lumen of seminiferous tubules
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small testis
(
J:293092
)
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• at 10-weeks of age, testes are significantly smaller than those of wild-type males
• however, neither inflammatory cells nor inflammatory cytokines are observed in the testes
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• average weight of the testes is normal until 5 weeks of age but significantly lower than that of heterozygous males at 10, 15 and 30 weeks of age
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