Fig 1.
Real-time PCR primers used in the real-time qPCR assays.
The forward and reverse primer sequences were listed in this form. HIF-1α, hypoxia inducible factor-1α; AIF, apoptosis-inducing factor.
Fig 2.
Comparison of body weight and absolute testis weight between the control and asthmatic mice. Parameters we observed including initial body weight, final body weight, body weight change and absolute testis weight.
* indicates a significant difference (P<0.05), compared with the control group.
Fig 3.
Comparison of sperm count and motility between the control and asthmatic mice.
*indicates a significant difference (P<0.05), compared with the control group. **indicates a very significant difference (P<0.01), compared with the control group.
Fig 4.
Apoptosis is enhanced in the asthmatic mouse testis.
(A) A DNA ladder assay was conducted for the 10 testis samples from control and asthma group (left panel). Quantification of the bands with lower molecular weight (about 150bp) in the left panel was done by using Image J software (right panel). (B) Enzymatic activity of caspase-3/7 was measured for the testis homogenates using Caspase-Glo-3/7 assay kit, and the relative ratio was expressed in the bar graph. *indicates a significant difference (P<0.05), compared with the control group. **indicates a very significant difference (P<0.01), compared with the control group.
Fig 5.
Histological changes in the testis.
The transverse sections of the testis were stained with hematoxylin and eosin (HE). (A) Control group; (B) Asthma group.
Fig 6.
Immunohistochemisty assay for the testis. The transverse sections of the testis were stained for cleaved caspase-3.
(A) Control group; (B) Asthma group.
Fig 7.
Relative mRNA level of several apoptosis-related genes.
(A) Equal amounts of RNA from 5 control or experiment mice were mixed well and split into two groups before cDNA synthesis. (B) Equal amounts of RNA samples from the 10 mouse testes were aliquot individually and then subjected to cDNA synthesis respectively. Transcriptional expression of indicated genes was measured by real-time qPCR. Data were presented as means ± SD (n = 5). ** indicates a very significant difference (P<0.01), compared with the control group.
Fig 8.
Western blot analysis of the apoptosis related proteins.
(A) Equal amounts of the protein homogenate from the testis tissue were immunoblotted with the indicated antibodies. (B) Quantification of the protein expression level in (A) was performed using Image J software.
Fig 9.
Asthma activates the proteinase caspase-9 and enhances the cleavage of PARP in the mouse testis.
(A) Enzymatic activity of caspase-9 in the testis homogenates was measured by Caspase-Glo-9 assay kit, and the relative ratio was expressed in the bar graph. (B) Western blot analysis of the PARP (full-length and cleaved isoform). The relative level of cleaved PARP was determined using Image J software as demonstrated in the middle panel. In the right panel, the ratio of cleaved PARP to full-length PARP was shown based on the intensity calculated by Image J software. Data were expressed as means ± SD. ** indicates a very significant difference (P<0.01), compared with the control group.