Endoplasmic Reticulum Stress (ER Stress) and Unfolded Protein Response (UPR) Occur in a Rat Varicocele Testis Model

Abstract

Using a surgically induced varicocele rat model, we show here strong evidence that the misfolded/unfolded protein response that is part of the stress response of the endoplasmic reticulum (ER) is activated in the varicocele testis (VCL), leading to the induction of apoptosis. To support this hypothesis, it is observed that the spliced variant of the X-box protein 1 (XBP1s), resulting from the activation of the inositol-requiring enzyme 1 (IRE1) membrane sensor, is significantly more represented in VCL testicular extracts. The activation of the IRE1/XBP1s pathway is also supported by the observation that the VCL testes show an increase phosphorylation of the c-Jun-kinase (JNK) known to be one intermediate of this pathway and an increased level of caspase-3, the terminal apoptotic effector, partly explaining the apoptotic status of the VCL testis.

Deferasirox, an Iron‐Chelating Agent, Improves Testicular Morphometric and Sperm Functional Parameters in a Rat Model of Varicocele

“…Similar to observations made in 2-month VCL animals, the very first mediator of the ER/UPR stress pathway, the testis chaperone Bip/HSPA5/GRP78, was not elevated in 4-month VCL animals. In contrast to activation of the late ER/UPR phase pathway IRE-1/XBP1s/pJUNK observed in the testis after 2 months of VCL [ 15 ], none of the 3 classical ER/UPR pathways were found to be activated in the testis after 4 months of VCL (supplementary figures 1 and 2 ). Only the mitochondrial proapoptotic signals appeared to be elevated as evidenced by increased accumulation of Bim mRNA and a reduction in Bcl-2 mRNA (supplementary figures 1 and 2 ).…”

“…In order to analyze the effect of DFX on the VCL-induced testis ER/UPR response, we first had to compare the 2-month and 4-month post-VCL induction observations with regard to testicular and spermatozoa parameters. This was necessary because although we had previously reported the ER/UPR status in rat testis 2 months postsurgery [ 15 ], we had no information concerning the ER/UPR status 4 months following VCL induction. Since VCL is a chronic condition with progressive oxidative stress leading to cumulative testicular destruction and germ cell apoptosis, we questioned whether the kinetics of the testis ER/UPR response might be different at this later time point.…”

“…Since these conflicting results could be attributed to the dose and duration of DFX treatment administered to the VCL animals used in this study [ 39 – 41 ], we decided to conduct a deeper investigation into the homeostasis of the VCL-DFX-treated testis. We recently reported that the endoplasmic reticulum/unfolded protein response (ER/UPR response), in which oxidative stress is a major player, is likely to be involved in the VCL testis [ 15 ]. Therefore, this response was monitored in the testes of the different groups of animals used in the present study.…”

“…After 5 min, 60 μ l of nigrosine dye was added. Smears were prepared with 20 μ l of the preparation [ 15 ]. Two hundred spermatozoa on each slide were randomly evaluated on an optical microscope (CX31 OLYMPUS, magnification ×100), and the total percentage of abnormal forms was determined.…”

“…In brief, total proteins were extracted from the testes after homogenization in a lysis buffer [ 15 ]. The protein concentration was evaluated by Bradford's test, and 30 μ g protein/sample was evaluated by 12% SDS-PAGE and then transferred to polyvinylidene difluoride (PVDF) membranes by wet transfer.…”