Alzheimer's Disease Mouse as a Model of Testis Degeneration

Abstract

Pituitary adenylate cyclase activating polypeptide (PACAP) is a neuropeptide with protective functions in the central nervous system and various peripheral organs. PACAP has the highest expression level in the testes, among the peripheral organs, and has a positive regulative role in spermatogenesis and in sperm motility. In the present study, we explored testicular degenerative alterations in a mouse model of Alzheimer's disease (AD) (B6C3-Tg(APPswe,PSEN1dE9)85Dbo/J) and demonstrated changes in PACAP-regulated signaling pathways. In addition, the effects of increased physical activity of AD (trained AD (TAD)) mice on testis were also followed. Reduced cell number and decreased thickness of basement membrane were detected in AD samples. These changes were compensated by physical activity. Expression of PACAP receptors and canonical signaling elements such as PKA, P-PKA, PP2A significantly decreased in AD mice, and altered Sox transcription factor expression was also detected. Via this signaling mechanism, physical activity compensated the negative effects of AD on the expression of type IV collagen. Our findings suggest that the testes of AD mice can be a good model of testis degeneration. Moreover, it can be an appropriate organ to follow the effects of various interventions such as physical activity on tissue regeneration and signaling alterations.

Keywords: Alzheimer’s disease; Sox9; collagen type IV; physical activity; testis degeneration.

Figure 1

(A) Representative microphotograph of wild-type (WT), Alzheimer’s disease (AD) and trained AD (TAD) convoluted seminiferous tubules stained with hematoxylin-eosin (HE). Original magnification was 40×. Scale bar, 20 µm. (B) Cell number (spermatogonia, spermatocytes, spermatozoa) determination of convoluted seminiferous tubules and Leydig cell in the interstitium. (C) Thickness analysis of testicular basement membrane. Representative data of 5 independent experiments. Asterisks indicate significant (* p < 0.05) difference in thickness of basement membrane compared to WT and (^# p < 0.05) compared to AD.

Figure 2

Representative photos of mRNA (A) and protein (B) expression of pituitary adenylate cyclase activating polypeptide (PACAP) receptors in the testis. Optical density of signals was measured, and results were normalized to the optical density of WT. For panels (A,B), numbers below the signals represent integrated densities of signals determined by ImageJ software. Asterisks indicate significant (* p <0.05) alteration of expression as compared to the WT and (^# p <0.05) compared to AD. Representative data of 5 independent experiments. For RT-PCR reactions and for Western blot, actin was used as controls.

Figure 3

mRNA (A) and protein (B) expression of PACAP signaling in testis. For panels (A,B), numbers below the signals represent integrated densities of signals determined by ImageJ software. Asterisks indicate significant (* p <0.05) alteration of expression as compared to the WT and (^# p <0.05) compared to AD. Representative data of 5 independent experiments. For RT-PCR reactions and for Western blot, actin was used as controls. (C) Immunohistochemistry of P-Sox9 in seminiferous tubules. Magnification was made with 40× objective. Scale bar: 20 µm.

Figure 4

mRNA (A) and protein (B) expression of Col. IV, Col. IX and testatin. For RT-PCR and for Western blot reactions, actin was used as control. Optical density of signals was measured, and results were normalized to the optical density of controls. For panels (A,B), numbers below the signals represent integrated densities of signals determined by ImageJ software. Asterisks indicate significant (* p < 0.05) alteration of expression as compared to the WT and (^# p <0.05) compared to AD. Representative data of 5 independent experiments. For RT-PCR reactions and for Western blot, actin was used as controls. (C) Immunohistochemistry of Col. IV in seminiferous tubules. Magnification was made with 20× objective. Scale bar: 50 µm.