1. Comparison of two methods of in vivo gene transfer by electroporation
Hiroki Kubota, M.D., Yutaro Hayashi, M.D., Yasue Kubota, M.D., Kevin Coward, Ph.D., John Parrington, Ph.D. 
Fertility and Sterility 
Volume 83, Issue 4, Pages (April 2005)
DOI: /j.fertnstert
Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions

2. FIGURE 1
(a) β-Galactosidase activity (X-gal staining) used for detection of LacZ transgene expression after a period of 2 weeks following electroporation in group D (direct injection). The transgene was introduced into both germ cells (G) and somatic cells (S). (b) β-Galactosidase activity (X-gal staining) in group R (rete testis injection) 8 weeks postelectroporation; by this time point, β-galactosidase activity could not be detected in group D. Original magnification 200×.
Fertility and Sterility  , DOI: ( /j.fertnstert )
Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions

3. FIGURE 2
(a) Reverse transcription-polymerase chain reaction (RT-PCR) analysis of LacZ construct expression in group D (direct injection). Lane 1, marker; lane 2, LacZ plasmid DNA (positive control); lane 3, eight weeks after electroporation; lane 4, six weeks after electroporation; lane 5, four weeks after electroporation; lane 6, two weeks after electroporation; lane 7, one week after electroporation. (b) RT-PCR analysis of LacZ construct expression in group R (rete testis injection) (lanes 1–7: same as for group D in a). The transgene was detected until 4 weeks after electroporation in group D but was still detectable at 8 weeks postelectroporation in group R.
Fertility and Sterility  , DOI: ( /j.fertnstert )
Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions

4. FIGURE 3
(a) Histologic section showing seminiferous tubules 2 weeks after electroporation in group D (direct injection); degeneration of spermatogenesis was noted. (b) Histologic section showing seminiferous tubules 8 weeks after electroporation in group R (rete testis injection); spermatogenesis disorder appeared to have been alleviated. (c) Mean Johnsen scores. The Johnsen score of group D was significantly lower than that of group R at 2 weeks postelectroporation and at each subsequent time point thereafter, suggesting that the testes of group D exhibited more severe damage that persisted even 8 weeks after electroporation. *Significant difference between group D and group R; #significant difference between group D and group C; †significant difference between group R and group C; P<.05.)
Fertility and Sterility  , DOI: ( /j.fertnstert )
Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions

5. FIGURE 4
(a–d) In situ detection of testicular cell apoptosis in treated animals using TUNEL analysis (original magnification 200×). There were remarkable increases in the number of apoptotic germ cells two weeks after electroporation in group D (a; direct injection) and group R (b; rete testis injection), but very few apoptotic cells in group C (c; control). Even 8 weeks after electroporation, a considerable number of apoptotic cells were observed in group D (d). Apoptotic cells (black arrows) are shown in both germ cells and somatic cells. (e) Changes in apoptosis index (AI) in electroporated testis. Compared to controls (group C), the number of apoptotic cells increased remarkably in both electroporated groups (D and R) just one week after electroporation. Although testes showed signs of recovery soon after, especially in group R (rete testis injection), the AI of group D (direct injection) remained at high levels even after 8 weeks post electroporation. *Significant difference between group D and group R; #significant difference between group D and group C; †significant difference between group R and group C; P<0.05.)
Fertility and Sterility  , DOI: ( /j.fertnstert )
Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions

6. FIGURE 4
(a–d) In situ detection of testicular cell apoptosis in treated animals using TUNEL analysis (original magnification 200×). There were remarkable increases in the number of apoptotic germ cells two weeks after electroporation in group D (a; direct injection) and group R (b; rete testis injection), but very few apoptotic cells in group C (c; control). Even 8 weeks after electroporation, a considerable number of apoptotic cells were observed in group D (d). Apoptotic cells (black arrows) are shown in both germ cells and somatic cells. (e) Changes in apoptosis index (AI) in electroporated testis. Compared to controls (group C), the number of apoptotic cells increased remarkably in both electroporated groups (D and R) just one week after electroporation. Although testes showed signs of recovery soon after, especially in group R (rete testis injection), the AI of group D (direct injection) remained at high levels even after 8 weeks post electroporation. *Significant difference between group D and group R; #significant difference between group D and group C; †significant difference between group R and group C; P<0.05.)
Fertility and Sterility  , DOI: ( /j.fertnstert )
Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions

7. FIGURE 5
Average sperm motility after electroporation. There were no significant differences (P>0.05) between any of the three groups in terms of sperm motility.
Fertility and Sterility  , DOI: ( /j.fertnstert )
Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions