In Vivo Gene Transfer to Mouse Spermatogenic Cells: RESULTS(4)


A majority of the positive cells observed as dot-like localizations along the seminiferous tubules turned out to be Sertoli cells, which are nonproliferating somatic cells (Fig. 3A). Another pattern of staining was observed as a relatively long linear mass along the tubules, where p-gal-expressing cells were found as a clump of spermatogenic germ cells present from the basal membrane to the inner lumen of the tubule (Fig. 3B), indicating that the CMV-lacZ transgene was incorporated into undifferentiated germ cells at the time of EP transfection. buy ventolin inhalers

Busulfan-treated testes were used in an attempt to confirm that the transgene can be integrated into undifferentiated cells such as spermatogonia and the precursor stem cells. Adequate busulfan treatment turns off spermatogenesis transiently, after which a new spermatogenic cycle restarts from the surviving stem cells. At 2 mo after EP transfection of CMV-lacZ into busulfan-treated testis, p-gal staining was observed along the long axis of seminiferous tubules (Fig. 3C). In this tubule, p-gal-expressing cells were detected in the reinitiated spermatogenic cell layers (Fig. 3D), whose image was in a pattern very similar to the pattern detected at 1 mo after gene transfer to the nontreated testis (Fig. 3B).
Fig3In Vivo Gene Transfer
FIG. 3. Analysis of long-lasting CMV-lacZ transgene expression. Cross section (8 ^m) of the testis at 1 mo after in vivo transfection (A, B, bar = 20 ^m). Busulfan-treated testis and the section (8 ^m) at 2 mo after in vivo transfection (C, D). A) Transgene expression detected in Sertoli cell. B)Transgene expression detected in a clump of spermatogenic cell layers. C) Transgene expression along the axis of the seminiferous tubule. Bar = 500 ^m. D) Cross section of the positive region of C. p-gal-positive cells are found as a colony located within the seminiferous tubule. Bar = 100 ^m.


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