In vivo cellular evidence of autophagic associated spermiophagy within the principal cells during sperm storage in epididymis of the turtle.

Imran Tarique,Yonghong Shi,Chang Chen,Baitao Ding,Noor Samad Gandahi,Quisheng Chen,Ping Yang,Waseem Ali Vistro

doi:10.18632/aging.103144

Abstract

The epididymis plays a significant role as a quality control organ for long-term sperm storage, maturation, and fertilizing ability and perform filtration function to eliminate abnormal or residual spermatozoa by phagocytosis. However, the role of autophagy in spermiophagy during sperm storage in turtle epididymis still needs to be studied. In this study, we reported in vivo spermiophagy via the cellular evidence of lysosome engulfment and autophagy within the principal cells during sperm storage in the turtle epididymis. Using immunofluorescence, Lysosome associated membrane protein-1 (LAMP1) and microtubule-associate protein light chain 3 (LC3) showed strong immunosignals within the apical cytoplasm of epididymal epithelia during hibernation than non-hibernation. Co-immunolabeling of LAMP1 and LC3 was strong around the phagocytosed spermatozoa in the epididymal epithelia and protein signaling of LAMP1 and LC3 was confirmed by western blotting. During hibernation, ultrastructure showed epididymal principal cells were involved in spermiophagy and characterized by the membrane’s concentric layers around phagocytosed segments of spermatozoa, degenerative changes in the sperm head and lysosome direct attachment, and with the existence of cellular components related to autophagy (autophagosome, autolysosome). In conclusion, spermiophagy occurs by lysosomal engulfment and autophagic activity within the principal cells of the turtle epididymis during sperm storage.

Highlights

Spermatozoa that develop within testicular seminiferous tubules are morphologically complete when released, but immotile and unable to fertilize an oocyte [1]
Higher magnification showed that numerous spermatozoa were in-contact with the epididymal epithelia (Figure 1B), whereas few spermatozoa were observed within the cytoplasm of the epididymal epithelia (Figure 1C)
To investigate the spermiophagy within the epididymal epithelia of the turtle, we used immunofluorescence to determine the expression of Lysosome associated membrane protein-1 (LAMP1), which is a key lysosomal marker

Summary

Introduction

Spermatozoa that develop within testicular seminiferous tubules are morphologically complete when released, but immotile and unable to fertilize an oocyte [1]. The spermatozoa travel via efferentes ductuli into the epididymis for post testicular maturation where they develop the capacity for motility and acquire fertilizing ability [2]. Prior to or during mating, not all spermatozoa are capable of being ejaculated from the reproductive tract due to their weak capability of energetic disorder of mitochondria or DNA damage [10] or immaturity, and these are referred as residual or unejaculated spermatozoa. Such spermatozoa are eliminated by the duct epithelia or luminal macrophages in the male reproductive tract via phagocytosis or absorbing [4]. We hypothesize that during sperm storage in the epididymis of soft-shelled turtle (Pelodiscus sinensis), epithelial cells play their potential role to eliminate spermatozoa via phagocytosis

Methods

Results

Conclusion