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Therapeutic effect of RIPK1 inhibitor in testicular ischemia-reperfusion
32nd Annual EAU Congress, 24-28 March 2017, London, United Kingdom
577
Therapeutic effect of RIPK1 inhibitor in testicular ischemia-reperfusion Eur Urol Suppl 2017; 16(3);e1003
Ohira S.1, Hara R.1, Tone S.2, Nagai A.1 1
Kawasaki Medical School, Dept. of Urology, Kurashiki City, Japan, 2Graduate School of Tokyo Denki University, Dept. of Life Science and Engineering, Hatoyama-Cho, Japan INTRODUCTION & OBJECTIVES: Testicular torsion makes testis ischemic. Surgery is immediately needed to reestablish blood flow. Even if the surgery succeed, testicular atrophy is often appear and lead to spermatic dysfunction. However, the etiology is still controversial. Recently, it was reported that loss of the formation in any tissue after ischemia-reperfusion (IR) was involved in necroptosis, one of programmed cell death series. Necrostatin-1 (Nec-1) blocks both necroptosis and indoleamine 2,3-dioxygenase (IDO). In our previous research, we elucidated that IDO inhibitor decrease inflammation. We investigated pathophysiological change of testicular IR researching histological and biochemical phase. Subsequently to the analysis, we tried to inhibit Receptor-interacting protein kinase 1 (RIPK1) and clarify a function of necroptosis in testicular IR. MATERIAL & METHODS: Twelve weeks old ICR male mice were used in this study. Unilateral testicular artery was clamped. Declamping three hours later, testicular blood flow were resumed. After the procedure, bilateral testes were removed in time dependent manner (at day 1, 3, 5 and 7). Histological and biochemical change were evaluated by immunostaining and ELISA method. Semen analysis from the epididymal cauda were evaluated by computer aided sperm analysis. In the following research, Nec-1 4 µg/g was administrated (iv) after declamping. Histological, biochemical and semen analysis were evaluated. Sham surgery was performed as control. Their experiments were duplicated at least. RESULTS: Invasion of lymphocyte-predominant inflammatory cells accumulated at day 3, 5 and destruction of seminiferous structure were observed at day 5, 7. Necroptosis using RIPK staining was abundantly expressed. In semen analysis, significant decreased spermatic concentration was observed at day 5 and 7 compared to control (p<0.05). Significant decreased spermatic motility was observed at day 1, 3, 5 and 7. Interestingly, in contralateral testes, significant decreased spermatic motility was observed at day 5 and 7. Some candidates were picked up as molecular marker. Significant increased Eselectin, marker of leukocyte-endothelial cell adhesion molecule, was observed at day 1. Significant increased IL-6, marker of inflammation, was observed at day 3. Significant increased 8-OHdG, marker of oxidative stress, was observed at day 5. Interestingly, significant increased E-selectin, IL-6 and 8-OHdG were observed in contralateral. Same results were introduced in immunohistological staining. After treatment of Nec-1, testicular structure were maintained and little invasion of inflammatory cells were observed. Significant decreased necroptosis were statistically observed. Significant decreased Eselectin, IL-6 and 8-OHdG were also observed. Importantly, spermatic motility also maintained in bilateral testes.
Eur Urol Suppl 2017; 16(3);e1003
32nd Annual EAU Congress, 24-28 March 2017, London, United Kingdom
577
Therapeutic effect of RIPK1 inhibitor in testicular ischemia-reperfusion Eur Urol Suppl 2017; 16(3);e1004
CONCLUSIONS: Oxidative stress via inflammation and necroptosis should induce spermatogenetic dysfunction in IR testis. First, prominent inflammation was occurred in testicular IR model and not only expression of cytokines were increased, but also in contralateral testes. Subsequently, oxidative stress expressed and cell death appeared. Therefore, their change of bilateral testis would be one of pathophysiology with ischemic testis. To inhibit RIPK1 would be contribute to protection of testicular tissue and spermatogenesis in IR model.
Eur Urol Suppl 2017; 16(3);e1004 Powered by TCPDF (www.tcpdf.org)
577
Therapeutic effect of RIPK1 inhibitor in testicular ischemia-reperfusion Eur Urol Suppl 2017; 16(3);e1003
Ohira S.1, Hara R.1, Tone S.2, Nagai A.1 1
Kawasaki Medical School, Dept. of Urology, Kurashiki City, Japan, 2Graduate School of Tokyo Denki University, Dept. of Life Science and Engineering, Hatoyama-Cho, Japan INTRODUCTION & OBJECTIVES: Testicular torsion makes testis ischemic. Surgery is immediately needed to reestablish blood flow. Even if the surgery succeed, testicular atrophy is often appear and lead to spermatic dysfunction. However, the etiology is still controversial. Recently, it was reported that loss of the formation in any tissue after ischemia-reperfusion (IR) was involved in necroptosis, one of programmed cell death series. Necrostatin-1 (Nec-1) blocks both necroptosis and indoleamine 2,3-dioxygenase (IDO). In our previous research, we elucidated that IDO inhibitor decrease inflammation. We investigated pathophysiological change of testicular IR researching histological and biochemical phase. Subsequently to the analysis, we tried to inhibit Receptor-interacting protein kinase 1 (RIPK1) and clarify a function of necroptosis in testicular IR. MATERIAL & METHODS: Twelve weeks old ICR male mice were used in this study. Unilateral testicular artery was clamped. Declamping three hours later, testicular blood flow were resumed. After the procedure, bilateral testes were removed in time dependent manner (at day 1, 3, 5 and 7). Histological and biochemical change were evaluated by immunostaining and ELISA method. Semen analysis from the epididymal cauda were evaluated by computer aided sperm analysis. In the following research, Nec-1 4 µg/g was administrated (iv) after declamping. Histological, biochemical and semen analysis were evaluated. Sham surgery was performed as control. Their experiments were duplicated at least. RESULTS: Invasion of lymphocyte-predominant inflammatory cells accumulated at day 3, 5 and destruction of seminiferous structure were observed at day 5, 7. Necroptosis using RIPK staining was abundantly expressed. In semen analysis, significant decreased spermatic concentration was observed at day 5 and 7 compared to control (p<0.05). Significant decreased spermatic motility was observed at day 1, 3, 5 and 7. Interestingly, in contralateral testes, significant decreased spermatic motility was observed at day 5 and 7. Some candidates were picked up as molecular marker. Significant increased Eselectin, marker of leukocyte-endothelial cell adhesion molecule, was observed at day 1. Significant increased IL-6, marker of inflammation, was observed at day 3. Significant increased 8-OHdG, marker of oxidative stress, was observed at day 5. Interestingly, significant increased E-selectin, IL-6 and 8-OHdG were observed in contralateral. Same results were introduced in immunohistological staining. After treatment of Nec-1, testicular structure were maintained and little invasion of inflammatory cells were observed. Significant decreased necroptosis were statistically observed. Significant decreased Eselectin, IL-6 and 8-OHdG were also observed. Importantly, spermatic motility also maintained in bilateral testes.
Eur Urol Suppl 2017; 16(3);e1003
32nd Annual EAU Congress, 24-28 March 2017, London, United Kingdom
577
Therapeutic effect of RIPK1 inhibitor in testicular ischemia-reperfusion Eur Urol Suppl 2017; 16(3);e1004
CONCLUSIONS: Oxidative stress via inflammation and necroptosis should induce spermatogenetic dysfunction in IR testis. First, prominent inflammation was occurred in testicular IR model and not only expression of cytokines were increased, but also in contralateral testes. Subsequently, oxidative stress expressed and cell death appeared. Therefore, their change of bilateral testis would be one of pathophysiology with ischemic testis. To inhibit RIPK1 would be contribute to protection of testicular tissue and spermatogenesis in IR model.
Eur Urol Suppl 2017; 16(3);e1004 Powered by TCPDF (www.tcpdf.org)