Journal: eLife

Article Title: Sperm motility in mice with oligo-astheno-teratozoospermia restored by in vivo injection and electroporation of naked mRNA

doi: 10.7554/eLife.94514

Figure Lengend Snippet: ( A ) A solution containing 0.05% Fast Green and 1% fluorescent i-particles NIRFiP-180 was prepared, 10 µl was injected into the seminiferous tubules of adult males, through the rete testes and its efferent channels. Injection was performed at constant pressure under a binocular microscope. The progression of filling of the seminiferous tubules was monitored thanks to the Fast Green. ( B ) The testes were only filled to 2/3 capacity in order to prevent damage to the tissue. ( C ) Representative distribution of fluorescent i-particles NIRFiP-180 in a whole cross-section of an injected testis. Nuclei were counterstained with DAPI (blue emission) to reveal tubules. ( D ) Enlargement of a seminiferous tubule showing particles localized inside the lumens of the tubules. Scales bars: 1 mm and 500 µm.

Article Snippet: The optically cleared mouse testes were imaged using a ‘ConfoBright’ system which is a unique adaptive optics confocal microscope (Nikon A1R MP, Nikon Europe B.V., The Netherlands) equipped with a deformable mirror module (AOS-micro, AlpAO, Montbonnot, France) to correct geometrical aberrations.

Techniques: Injection, Microscopy

Journal: eLife

Article Title: Sperm motility in mice with oligo-astheno-teratozoospermia restored by in vivo injection and electroporation of naked mRNA

doi: 10.7554/eLife.94514

Figure Lengend Snippet: Testes were injected/electroporated with GFP- mRNA on day 0. On day 1, whole testes were fixed and subjected to optical clearing. ( A ) Testes were observed before and after optical clearing on a binocular microscope. The right image shows the transparency of the testis after complete clearing, revealing the blue mesh throughout the organ. ( B ) The 3D internal structure of a cleared testis was reconstructed from the lightsheet microscopy images. The reconstruction was possible only for a half testis due to optical issues. Two opposing faces of the same testis are presented, allowing the distribution of GFP fluorescence throughout the seminiferous tubules to be measured. Pink fluorescence corresponds to the autofluorescence of interstitial cells located around the seminiferous tubules. Scale bars A: 1 mm and B: 500 µm.

Article Snippet: The optically cleared mouse testes were imaged using a ‘ConfoBright’ system which is a unique adaptive optics confocal microscope (Nikon A1R MP, Nikon Europe B.V., The Netherlands) equipped with a deformable mirror module (AOS-micro, AlpAO, Montbonnot, France) to correct geometrical aberrations.

Techniques: Injection, Microscopy, Fluorescence

Journal: eLife

Article Title: Sperm motility in mice with oligo-astheno-teratozoospermia restored by in vivo injection and electroporation of naked mRNA

doi: 10.7554/eLife.94514

Figure Lengend Snippet: Adult WT mouse testes were injected with a solution containing Armc2 -mRNA and eGFP- mRNA. After injection, the testes were electroporated and mice were euthanized two weeks later. ( A ) Whole testis under white and blue lights on a fluorescence microscope. ( A1 ) Control testes not injected/electroporated. ( A2 ) Testes injected with Armc2 -mRNA and eGFP- mRNA. eGFP- mRNA was co-injected to follow the transfection efficiency. ( B ) Ratio of injected/electroporated testis weights to control testis weights at several time points post-injection (3-, 6-, 10-, 15-, 21-, 28-, and 35-day post-surgery). n = 1 mouse per time.

Article Snippet: The optically cleared mouse testes were imaged using a ‘ConfoBright’ system which is a unique adaptive optics confocal microscope (Nikon A1R MP, Nikon Europe B.V., The Netherlands) equipped with a deformable mirror module (AOS-micro, AlpAO, Montbonnot, France) to correct geometrical aberrations.

Techniques: Injection, Fluorescence, Microscopy, Control, Transfection