Relatively low-dose cyclophosphamide is likely to induce apoptotic cell death in rat thymus through Fas/Fas ligand pathway

Abstract

Cyclophosphamide (CPA) is widely used as an efficiently antineoplastic drug, but also causes immunosuppression as its adverse-side effect. To understand the effect of low- or relative low-dose CPA on the immune system, apoptotic cell death in rat thymus, either exposed to different doses of CPA (0, 2, 7, 20 and 70 mg/kg) for 12 h or exposed to 70 mg/kg for different times (4-48 h), was investigated by DNA fragmentation (DNA ladder) detection and in situ morphological examination using hematoxylin and eosin (H and E) staining. Immunohistochemical staining for Fas protein expression in the thymus of rats exposed to CPA was performed. Results showed that exposure of rats to CPA 0–70 mg/kg for 12 h did not cause significant decrease in the ratio of thymus weight to body weight. However, the ratio of thymus weight to body weight was decreased significantly at 48 h after exposure to 70 mg/kg CPA. Exposure to 20 and 70 mg/kg CPA for 12 h caused a visible DNA ladder in gel electrophoresis. DNA ladder formation was increased progressively in the groups from 8 h to optimal magnitude at 12–24 h and then disappeared at 48 h after 70 mg/kg CPA. This pattern was confirmed by a quantitative evaluation of the apoptotic cells using H and E staining. Expression of Fas protein was enhanced in the thymus of rats exposed to 70 mg/kg CPA for 4–8 h as compared to control rats. These results are different from previous studies on high dose CPA and the induction of the apoptotic cell death in thymus by low or relative low doses of CPA might be a result of Fas/Fas–ligand interactions.