Murine male germ cell apoptosis induced by busulfan treatment correlates with loss of c‐kit‐expression in a Fas/FasL‐ and p53‐independent manner

Abstract

Male germ cell apoptosis has been extensively explored in rodents. In contrast, very little is known about the susceptibility of developing germ cells to apoptosis in response to busulfan treatment. Spontaneous apoptosis of germ cells is rarely observed in the adult mouse testis, but under the experimental conditions described here, busulfan-treated mice exhibited a marked increase in apoptosis and a decrease in testis weight. TdT-mediated dUTP-X nicked end labeling analysis indicates that at one week following busulfan treatment, apoptosis was confined mainly to spermatogonia, with lesser effects on spermatocytes. The percentage of apoptosis-positive tubules and the apoptotic cell index increased in a time-dependent manner. An immediate effect was observed in spermatogonia within one week of treatment, and in the following week, secondary effects were observed in spermatocytes. RT-PCR analysis showed that expression of the spermatogonia-specific markers c-kit and Stra 8 was reduced but that Gli I gene expression remained constant, which is indicative of primary apoptosis of differentiating type A spermatogonia. Three and four weeks after busulfan treatment, RAD51 and FasL expression decreased to nearly undetectable levels, indicating that meiotic spematocytes and post-meiotic cells, respectively, were lost. The period of germ cell depletion did not coincide with increased p53 or Fas/FasL expression in the busulfan-treated testis, although p110Rb phosphorylation and PCNA expression were inhibited. These data suggest that increased depletion of male germ cells in the busulfan-treated mouse is mediated by loss of c-kit/SCF signaling but not by p53-or Fas/FasL-dependent mechanisms. Spermatogonial stem cells may be protected from cell death by modulating cell cycle signaling such that E2F-dependent protein expression, which is critical for G1 phase progression, is inhibited.

Developmental arrest of scNT‐derived fetuses by disruption of the developing endometrial gland as a result of impaired trophoblast migration and invasiveness

“…Immunohistochemical staining was conducted as previously described (Choi et al, 2004;Park et al, 2005Park et al, , 2009. Briefly, paraffin sections of placenta and uterus tissue were cleared in Histoclear for approximately 10 min and dehydrated in increasing concentrations of ethanol.…”

“…TdT-Mediated dUTP-X Nicked End Labeling (TUNEL) Assay TUNEL assays were performed as described previously (Choi et al, 2004;Park et al, 2009). Tissues fixed in 4% (w/v) paraformaldehyde in 0.01 M PBS (pH 7.4) were washed with PBS, dehydrated in ethanol (70, 90, and 100%), and embedded in paraffin wax.…”

“…Real-time RT-PCR was performed as described previously (Choi et al, 2004) by using the Taqman model 7700 sequence Detector (ABI, Perkin-Elmer, Waltham, MA). The porcine gene expression levels were normalized to GAPDH gene expression, which was unaffected in the scNT-derived pigs.…”

Primary extragonadal germ cell tumor: A case report on prostate seminoma

“…The alkylating groups take the place of the hydrogen atom leading to mismatch between DNA intrastrand and interstrand cross-linking, chain fractures, termination of replication and loss of protein or physiological activity, resulting in cell apoptosis. The second is the activation and regulation of gene expression changes induced by several signal transduction pathways resulting in interference to cell growth, proliferation and differentiation (17)(18)(19).…”

Implications of Sertoli cell induced germ cell apoptosis to testicular pathology

“…52,53 Bulsufan, a germ cell toxicant initially causes the loss of c-kit/stem cell factor signaling before spermatogonia apoptosis occurs through the intrinsic pathway. 54 Thus, the subtype of the apoptotic germ cell provides evidence for the cell death pathway but not the mechanism of induction.…”