XIAP as a radioresistance factor and prognostic marker for radiotherapy in human rectal adenocarcinoma.

A differential responsiveness of patients to ionizing radiation is observed after preoperative radiotherapy for rectal adenocarcinoma that might be related, in part, to an apoptosis defect. To establish if proteins of the apoptotic cascades [pro-apoptotic: active caspase 3, 8, and 9 and DIABLO (direct inhibitor of apoptosis-binding protein with low pI); anti-apoptotic: XIAP (X-linked inhibitor of apoptosis)] are involved, we analyzed their profile in radioresistant (SW480) and radiosensitive (SW48) human colorectal cell lines. We demonstrated that, after irradiation, the SW48 cells increased the expression of the pro-apoptotic proteins, whereas the SW480 cells increased the expression of the anti-apoptotic protein XIAP. Moreover, XIAP knockdown in SW480 cells enhanced the basal and radiation-induced apoptotic index; the propensity of the SW480 cells to undergo apoptosis after radiation was higher compared with SW48 cells. In a translational study of 38 patients with rectal carcinoma, we analyzed the apoptotic profile for tumor and noncancerous tissue for each biopsy specimen using IHC. According to their response to preoperative radiotherapy, patients were classified into two groups: responsive and nonresponsive. Although no difference in expression of caspase 3, 8, or 9 was observed in the tumor/normal tissue ratio between responsive and nonresponsive patients, the ratio decreased for DIABLO and increased for XIAP. In conclusion, inhibition of XIAP rescues cellular radiosensitivity and both DIABLO and XIAP might be potential predictive markers of radiation responsiveness in rectal adenocarcinoma.

Chronic crude garlic-feeding modified adult male rat testicular markers: mechanisms of action.

BACKGROUND: Garlic or Allium sativum (As) shows therapeutic effects such as reduction of blood pressure or hypercholesterolemia but side-effects on reproductive functions remain poorly investigated. Because of garlic's chemical complexity, the processing methods and yield in preparations differ in efficacy and safety. In this context, we clarify the mechanisms of action of crushed crude garlic on testicular markers. METHODS: During one month of treatment, 24 male rats were fed 5%, 10% and 15% crude garlic. RESULTS: We showed that crude garlic-feeding induced apoptosis in testicular germ cells (spermatocytes and spermatids). This cell death process was characterized by increased levels of active CASP3 but not CASP6. Expression of the caspase inhibitors BIRC3 and BIRC2 was increased at all doses of As while expression of XIAP and BIRC5 was unchanged. Moreover, expression of the IAP inhibitor DIABLO was increased at doses 10% and 15% of As. The germ cell death process induced by As might be related to a decrease in testosterone production because of the reduced expression of steroidogenic enzymes (Star, Cyp11a, Hsd3b5 and Hsd17b). Evaluation of Sertoli markers showed that TUBB3 and GSTA2 expression was unchanged. In contrast, AMH, RHOX5 and CDKN1B expression was decreased while GATA4 expression was increased. CONCLUSION: In summary, we showed that feeding with crude garlic inhibited Leydig steroidogenic enzyme expression and Sertoli cell markers. These alterations might induce apoptosis in testicular germ cells.

Differential effects of ionizing radiation and platinum-derivative chemotherapy on apoptotic pathways in testicular germ cells.

PURPOSE: The purpose here was to identify whether ionizing radiation and oxaliplatin triggered testicular germ cells apoptosis through different executionary pathways. MATERIALS AND METHODS: Adult male mice are treated with oxaliplatin (0.5 mg/kg, Ox) 4 h before being locally irradiated (0.5 Gy, IR, considered as time 0 h). RESULTS: The number of apoptotic germ cells was significantly higher for IR (p < 0.008), Ox (p < 0.0001) and Ox + IR (p < 0.0001) groups compared to the untreated mice group. Similarly, the different treatments induced an increase of p53 expression. Downstream p53, IR and Ox used different pathways. Indeed, IR increased effector caspase-3 expression in terms of mRNA (p < 0.002), pro-enzyme p < 0.0001) and active (3.7-fold, p < 0.003) protein levels but not the inhibitors of apoptosis proteins (IAP) including cIAP1, cIAP2 and XIAP. In contrast, while oxaliplatin treatment had no apparent effect on caspase-3 expression, it significantly decreased the cIAP1 (p < 0.005), cIAP2 (p < 0.008) and XIAP (p < 0.02) proteins levels. Finally, the combination of both treatments decreased IAP expression but did not modify caspase-3 levels while it increased the AIF (apoptosis-inducing factor) protein levels (5.5-fold, p < 0.003). No modification of AIF levels was observed with OX or IR alone. CONCLUSIONS: Together, these results indicate that the platinum analogue oxaliplatin and the ionizing radiations trigger apoptosis in the testicular germ cells, probably through different pathways.

Status of the executioner step of apoptosis in human with normal spermatogenesis and azoospermia.

OBJECTIVE: To localize the different proteins involved in the executioner step of apoptosis in the human testicular tissue: effector caspases (3, 6, and 7), caspase inhibitors called inhibitor of apoptosis proteins (IAPs, such as XIAP), and IAP inhibitors such as Smac/DIABLO and to investigate XIAP and Smac expression and activation of caspase-3 in azoospermia. DESIGN: Retrospective study. SETTING: The Biology of Reproduction Center of Poissy and Inserm U407, France. PATIENT(S): Twenty-five patients diagnosed as azoospermic and 4 patients with normal testicular histology. INTERVENTION(S): Testicular biopsies for histopathological assessment. MAIN OUTCOME MEASURE(S): Localization of proteins by immunohistochemistry. RESULT(S): Effector caspase-7 seemed to be absent from normal human testes, whereas procaspase-3 and procaspase-6 were detected in both somatic and germ cells. XIAP was mainly expressed in Sertoli cells, whereas its inhibitor Smac was detected in pachytene spermatocytes. On the other hand, although few apoptotic germ cells were detected in biopsies from patients with obstructive azoospermia, increased levels of apoptotic germ cells were detected in spermatogenetic arrest. This increase in apoptotic germ cells was associated with increased levels of active caspase-3 in patients with spermatogenetic arrest, whereas the expression of XIAP and Smac/DIABLO was at similar levels in all groups. CONCLUSION(S): Active caspase-3 might be important in the apoptotic process observed in spermatogenetic arrest.