Immunohistochemical study of seminiferous epithelium in adult bovine testis using monoclonal antibodies against Ki-67 protein and proliferating cell nuclear antigen (PCNA)

Abstract

The distribution pattern of proliferating cell nuclear antigen (PCNA) and Ki-67 protein was studied in adult bovine seminiferous epithelium by means of immunohistochemistry using monoclonal antibodies. Tailoring the methodological protocol for each of the two proliferation markers was a necessary prerequisite for obtaining optimal results in tubular sections and whole-mounts. A-, I- and B-spermatogonia displayed PCNA-positive nuclei, except during meta-, ana- and telophases of mitosis. PCNA-negative nuclei in the basal tubular compartment, excluding those from non-cycling Sertoli cells, belonged to the spermatogonia precursor cell line. However, only about 30%, 45% and 47% of the respective A-, I-, B-spermatogonia had positive nuclei after exposure to the MIB-1 antibody directed against the Ki-67 protein. Spermatogonia with MIB-1-negative nuclei represented cells in the G1-phase. Both antibodies reacted intensely with the nuclei of preleptotene primary spermatocytes. PCNA reactivity was also present during leptotene through pachytene. Ki-67 protein expression was absent during leptotene and zygotene but was again encountered during pachytene and meiosis I and II. Anti-PCNA/anti-protein gene product 9.5 double-labelling indicated that the transition from spermatogonia precursor cells into A1-spermatogonia is not strictly synchronized in a given tubular segment, a possible reason for the flexibility in A-spermatogonial propagation seen in bovine seminiferous tubules.

Ubiquitin ligase Rad18Sc localizes to the XY body and to other chromosomal regions that are unpaired and transcriptionally silenced during male meiotic prophase

“…The experiments described herein were performed to obtain insight into possible gametogenic functions of the mouse homolog of RAD18, Rad18 Sc (Hoege et al, 2002;Kannouche et al, 2004). PCNA staining has been reported for mitotically active spermatogonia and for preleptotene spermatocytes, and persistence of a low level of nuclear staining at later stages of the meiotic prophase (zygotene and early-mid pachytene) has been described (Kamel et al, 1997;Wrobel et al, 1996). In mouse testis crosssections, using immunostaining, we did not detect PCNA in late pachytene and diplotene spermatocytes or in the XY body.…”

Disruption of PLZP in Mice Leads to Increased T-Lymphocyte Proliferation, Cytokine Production, and Altered Hematopoietic Stem Cell Homeostasis

“…Morphology of the PLZP lacZ/lacZ testis appeared grossly normal, plus sperm were being generated normally within the tubules suggestive of an intact differentiation pathway of the spermatogonia into spermatids. The spermatogonia within the tubules were intact, as judged by a specific marker for this population, PCNA (40). In addition, both spermatogonia proliferation and apoptosis appeared unaffected in the PLZP lacZ/lacZ testis, as determined by cyclin D1 expression and activated Caspase-3 staining, respectively (6,21).…”

Essential role of Plzf in maintenance of spermatogonial stem cells

“…To further define the progressive loss of spermatogonia and the status of the Sertoli cells (a morphologically distinct cell population that support the spermatogonia), we used proliferating cell nuclear antigen (PCNA) and p27 as distinct markers for the proliferative spermatogonia 10 and Sertoli cells 11 , respectively. Many tubules of Zfp145 -/-mice lacked PCNA-positive cells but retained p27-positive Sertoli cells along the basement membrane (Fig.…”