Cai L, Hales B F, Robaire B
Department of Pharmacology and Therapeutics, McGill University, Montreal, Quebec, Canada.
Biol Reprod. 1997 Jun;56(6):1490-7. doi: 10.1095/biolreprod56.6.1490.
Treatment with cyclophosphamide, a commonly used anticancer drug, may result in oligozoospermia or azoospermia. The objective of this study was to determine whether exposure of male rats to cyclophosphamide induces apoptosis in male germ cells, and if so, when the peak of apoptosis occurs and at what specific stages of spermatogenesis. The presence of apoptosis was determined by terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) detection in situ and by an increase in DNA fragmentation (DNA ladder). To determine the time course of drug-induced apoptosis, male Sprague-Dawley rats were treated with a single dose (70 mg/kg BW) of cyclophosphamide, and the testes were fixed 0, 4, 8, 12, 18, 24, and 48 h after treatment. To determine the dose response, rats were treated with doses of cyclophosphamide (0, 2, 7, 20, and 70 mg/kg), and the testes were fixed 12 h after treatment. A low spontaneous incidence of apoptosis was observed in controls, in particular in premeiotic germ cells of stages I-IV and XI-XIV of the seminiferous tubules. In cyclophosphamide-exposed rats, the incidence of apoptosis increased progressively at 4 h and 8 h, reached a peak at 12 h (about 3.5-fold above control), and then decreased rapidly to control levels by 48 h. A 70-mg/kg dose of cyclophosphamide induced a significant increase in apoptosis; lower doses did not. Although drug-induced apoptosis occurred in all stages of germ cells, it was most pronounced in spermatogonia and spermatocytes in stages I-IV and XI-XIV. Thus, apoptosis may be involved in the occurrence of oligozoospermia or azoospermia after cyclophosphamide treatment. Apoptosis of damaged premeiotic germ cells may serve a critical role in protecting subsequent generations from the diverse effects of toxicants.
常用抗癌药物环磷酰胺治疗可能导致少精子症或无精子症。本研究的目的是确定雄性大鼠暴露于环磷酰胺是否会诱导雄性生殖细胞凋亡，如果会，凋亡高峰期出现在何时以及在精子发生的哪些特定阶段。通过原位末端脱氧核苷酸转移酶介导的dUTP缺口末端标记（TUNEL）检测和DNA片段化增加（DNA梯状条带）来确定凋亡的存在。为了确定药物诱导凋亡的时间进程，对雄性Sprague-Dawley大鼠单次给予环磷酰胺（70 mg/kg体重），并在治疗后0、4、8、12、18、24和48小时固定睾丸。为了确定剂量反应，对大鼠给予不同剂量的环磷酰胺（0、2、7、20和70 mg/kg），并在治疗后12小时固定睾丸。在对照组中观察到低水平的自发凋亡发生率，特别是在生精小管I-IV期和XI-XIV期的减数分裂前生殖细胞中。在暴露于环磷酰胺的大鼠中，凋亡发生率在4小时和8小时逐渐增加，在12小时达到峰值（比对照组高约3.5倍），然后在48小时迅速降至对照水平。70 mg/kg剂量的环磷酰胺诱导凋亡显著增加；较低剂量则没有。尽管药物诱导的凋亡发生在生殖细胞的所有阶段，但在I-IV期和XI-XIV期的精原细胞和精母细胞中最为明显。因此，凋亡可能参与了环磷酰胺治疗后少精子症或无精子症的发生。受损的减数分裂前生殖细胞的凋亡可能在保护后代免受毒物的多种影响中起关键作用。
