Abstract
Diallyl disulfide (DADS), a major organosulfur compound derived from garlic, has various biological properties, including anti-cancer effects. However, the protective mechanism of DADS against radiation-induced mouse testis cell apoptosis has not been elucidated. In this study, the magnitude of radiation effects evoked by carbon ion irradiation was marked by morphology changes, significant rise in apoptotic cells, activation expression of p53, up regulation the ratio of pro-apoptotic Tap73/anti-apoptotic ΔNp73, as well as alterations of crucial mediator of the mitochondrial pathway. Interestingly, pretreatment with DADS attenuated carbon ion irradiation-induced morphology damages and apoptotic cells. Additionally, DADS elevated radiation-induced p53 and p21 expression, suggesting that p53 might be involved in the inhibition of cell cycle progression through up regulation of p21. Furthermore, administration with DADS prevented radiation-induced Tap73/ΔNp73 expression and consequently down regulated Bax/Bcl-2 ratio, cytochrome c release and caspase-3 expression, indicating that the balance between Tap73 and ΔNp73 had potential to activate p53 responsive genes. Thus, our results showed that radio protection effect of DADS on mouse testis is mediated by blocking apoptosis through changing the ratio of Tap73/ΔNp73 via mitochondrial pathway, suggesting that DADS could be used as a potential radio protection agent for the testis against heavy-ion radiation.
Highlights
We showed that Diallyl disulfide (DADS) supplementation was better able to ameliorate radiation-induced morphological damage and apoptosis in mouse testis through regulating the ratio of Tap73/Δ Np73 via mitochondrial pathway, but not p53
In an attempt to find the molecular mechanism underlying DADS-reduced apoptosis, we investigated the role of p53 in mouse testis treated by DADS before and exposed to irradiation (Fig. 3)
There are approximate 2.2, 2.3 and 3.2-fold inductions in the p21 expression in irradiation group pretreated with 10, 20 and 40 mg/kg DADS, respectively, in comparison with irradiation group. These results suggested that p53 might be involved in the inhibition of cell cycle progression through up regulation of p21 but does not appear to be critical for DADS-reduced apoptosis in mouse testis exposed to carbon ion irradiation
Summary
For histological analysis tissue specimens were fixed for 24 h in buffered formaldehyde solution (3.7% in PBS) at room temperature, dehydrated by graded ethanol and embedded in paraffin. The membranes were incubated with secondary antibodies conjugated with horseradish peroxidase (1:5000 dilution for anti-rabbit and 1:10 000 dilution for anti-mouse antibody in 5% nonfat dry milk in TBS) and streptavidin-horseradish peroxidase (1:2500) for 1 h at room temperature. Fixed sections were soaked in 3% H2O2 and incubated for 30 min; 1% Triton X-100 for 30 min; blocked for 25 min at room temperature by the drop wise addition of 5% BSA, primary antibody (1:500) 4 °C overnight; incubated donkey anti-rabbit IgG (1:100) secondary antibody Texas red and FITC (Invitrogen Life Technologies, Inc, CA, USA) at 37 °C for 1 h; the above steps were repeated with 0. A P value < 0 .05 was selected as a criterion for a statistically significant difference
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