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. 2024 Jun 17;14(10):3827–3842. doi: 10.7150/thno.95633

Figure 1.

Figure 1

Co-injection of AAV9 and 3-MA increase the transduction efficiency of spermatogenic cells in vivo. (A) Stereomicroscopic fluorescence imaging of representative testes (postnatal days [PND] 7, 21, and 56) 4 weeks after the microinjection of PBS, AAV9-CMV-RFP, or AAV9-CMV-RFP+3-MA. RFP: red fluorescence protein reporter gene. Scale bar: 1 mm. (B) Immunostaining of testicular sections for RFP (red) derived from testes (PND 7, 21, and 56) microinjected with PBS, AAV9-CMV-RFP, or AAV9-CMV-RFP+3-MA. Nuclei were stained with DAPI (blue). Scale bar: 100 μm. (C) Immunostaining of testicular sections from testes co-injected with AAV9-CMV-RFP and 3-MA for RFP (red), c-KIT (green, top), and DDX4 (green, bottom). Nuclei were stained with DAPI (blue). Scale bar: 50 μm. (D) Flow cytometric analysis of the percentage of RFP+ germ cells in testes 4 weeks after microinjection of PBS, AAV9-CMV-RFP, or AAV9-CMV-RFP+3-MA. Left: representative flow cytometry contour plot; Right: quantification of the percentage of RFP+ germ cells; Data are presented as the mean ± SEM (n = 6). P-values were determined using the two-tailed Student's t-test; **P < 0.01. (E) Stereomicroscopic fluorescence imaging of representative testes co-injected with AAV9-CMV-RFP and 3-MA at the indicated AAV titer. Scale bar: 1 mm. (F) H&E staining of testicular sections from testes co-injected with AAV9-CMV-RFP and 3-MA at the indicated AAV titer. Scale bar: 50 μm.