Histiocytic Characteristics And Functional Significance In The Oviduct Of Soft-Shelleturtle Trionyx Sinensis

The soft-shelled turtle, trionyx sinensis, serves as one of reptile, which has a special status in phylogenetic tree. Soft-shelled turtle, Trionyx sinensis was rich in nutrition, from the economical and pharmacological point of view, it aroused more and more investigators’ interests in breeding. The reptilian oviduct was a fascinating organ and it had multiple functions, not only the channel for fertlized eggs to pass, secretes egg white protein, synthesizes the egg shell fibre material, but also the organ for sperm storage. In this paper, we choose soft-shelled turtle as a model animal and explore the detail of organizational structure and cell morphology of oviduct. We also study biological properties of sperm storage from the histology and cytology perspective. Aim to provide theoretical basis on studies applied physiology of reproductive system in female reptiles, but also provide morphological clues for mechanism of sperm storage in female reptiles.Experiment I Oviduct Microstructure and Ultrastructure of the Soft-Shelled Turtle Trionyx sinensis, during Ovulation periodWe used light microscopy and electron microscopy to study the oviduct of soft-shelled turtle Trionyx sinensis. The result showed it was composed of five distinct regions. In the ovulation period, each region showed variation in microstructure, ultrastructure of oviductal epithelium, glands as well as secretory granules (SGs) also showed some difference. Infundibulum was thin and lined with simple epithelium with a few SGs. The intercellular canaliculi between the epithelial cells were usually relatively long and very labyrinthine. Infundibulum was observed non-glandular in lamina propria. Uterine tube was lined by high columnar epithelium which included ciliated cells and secretory cells. The prominent feature of ciliated cells was presence of irregular organelles including concentric circle type rough endoplasmic reticulum (RER) and mitochondria with large vacuoles in one pole, a few homogeneous electron-dense SGs were found in this cell. Secretory cells and gland cells showed more SGs with clearly varying in size, shape, and electron density. Occasionally, some transparent granules were found among the SGs. Isthmus, a short, non-glandular region showed the SGs concentrated mostly in the supranuclear region. Many sperms remained in aggregates in the lumen. Sometimes, the sperms or parts of sperm were found embedded directly in the cilia or secretory cells, and the embedded portions of sperm cells generally appeared normal in structure. In uterus, the SGs in the epithelium showed non-uniform electron densities and their matrix was highly complex. The gland in this region also showed various SGs, especially, concentric lamellae. Anterior uterus was demonstrated form sperm storage tubules (SSTs) which showed amount of sperm in the lumen. Most of the times, secretory cells appeared in pyramidal shape in the vagina. A few SGs from epithelium and glands were similar to those in uterus. Myoepithelial cells were found surrounding the gland along the oviduct, large number of mast cell, fibroblasts and collagen fibers also showed in the loose connective tissue. Obviously, the complex oviduct’s construction is relevant to its functions:albumen secretion, eggshell production, and sperm storage during ovulation period.Experiment Ⅱ Micro-Environment of Oviductal Sperm Storage Structure in the Soft-Shelled Turtle Trionyx sinensisWe investigated the structural features of sperm storage in the oviduct of female soft-shelled turtle Trionyx sinensis with the light microscope and transmission electron microscope. The result showed sperms were distributed from posterior uterine tube to uterus. However, the quantity of sperm and structure of each region showed marked difference. Most of the sperms were concentrated in the lumen of isthmus and sperm storage tubules (SSTs), sperms or part of sperms were embedded into the epithelial cells or entered between the cilia, and the embedded portions of sperm generally appeared normal in structure. However, the lumen of posterior uterus and posterior uterine tube showed only few free sperms. The epithelium contained ciliated, secretory cells along these areas, but the secretory granules (SGs) in the different regions showed various differences. In posterior uterine tube, the epithelium of mucosa consisted of columnar secretory and ciliated cells. Ciliated cells only showed few secretory granules (SGs). Secretory cells showed more SGs in cytoplasm, the size, shape, and electron density of these granules varied markedly. The tubular glands in lamina propria also contained various SGs similar to SGs of secretory cell in this region. The isthmus was a short, nonglandular region, The epithelium arranged tightly and folded as labyrinthine. The SGs in epithelium were electron-dense, mostly concentrated to the supranuclear region. The anterior uterus showed to form sperm storage tubules (SSTs). The epithelium of uterus and SSTs showed branching SGs with irregular dense cores. The gland in this region also contained numerous membrane-bound SGs with various internal ultrastructure. Complex structures and cell of these regions may be important for sperm storage and providing nutrition.ExperimentⅢ Enzyme histochemistry studies on the oviduct of Soft-shelled turtle Trionyx sinensis during breeding seasonWe used tissue staining method to observe the distribution of alkaline phosphatase (ALP), acid phosphatase (ACP), carbonic anhydrase (CA), ATPase, catalase (CAT) in the oviduct of soft-shelled turtle Trionyx sinensis during the breeding period The results showed that:these enzymes were observed in the oviduct of soft-shelled turtle Trionyx sinensis during breeding period, but the distribution and strength of reactions in each region were different. ALP mainly distributed in the wall of vascular cytoplasm of glands, stronger positive reaction showed in the uterus and the site of sperm storage. ACP mainly distributed in the epithelium and glands. CA appeared in cilia lining epithelium and glands from lamina propria, while the place of sperm storage showed weak response to these areas. CAT showed positive results in glands from different regions, especially, the gland from sperm storage tube (SST). Strong positive reaction of ATPase were observed in epithelial cilia and gland from lamina propria in different regions. These results suggest these enzymes have functions on egg formation, transportation and sperm storage.ExperimentlVBiochemical Studies on the oviduct of soft-shelled turtle Trionyx sinensisWe Used immunohistochemical methods adding light microscopy and electron microscopy to study the distribution and characteristics of oviduct mast cells (MC), androgen receptor (AR), Cajal interstitial cells (ICC) oviduct of soft-shelled turtle Trionyx sinensis. The results showed that:MC mainly distributed in the loose connective tissue, two or three MC were along the distribution of vascular or lymphatic vessels. Ultrastructure shows:MC’s cytoplasm contains a large number of high electron density secretory granules. During the breeding season, high concentrations of AR occurred mainly in the the top of epithelial ciliated cells and the glands in the lamina propria, suggesting that AR may be relate with epithelial thickening, glandular hypertrophy, and a large number of secretory granules formation in reproductive stage. High concentrations of AR also appeared gland cells nucleus in the sperm storage tubules and the lumen surface of isthmus, suggesting that AR may also be related with the sperm storage. ICC exist in the oviduct funnel Department, the Ministry of the uterus and vagina department. In the infundibular, ICC distributed in the muscle, showing branch-like. Compared with muscle cells,ICC showed low electron density. Ultrastructural features were observed that the nucleus large and curved, with3-4long protruding interacted with muscle cell through the gap junctions, the junction showed high electron density. Cytoplasm containing few organelles,Occasionally,cell membrane surface were concave, and many membranous vesicles distributed along the cell membrane in the cytoplasm. The ICC also showed in the uterus and vagina and were mainly located in the connective tissue of lamina propria and around blood vessels in muscle layer. The ICC in these regions showed longer protruding. The ultrastructure of ICC were similar to ICC in infundibular. These cells or receptors may be have functions on tissues seasonal changes, sperm storage and migration in the oviduct of Soft-shelled turtle Trionyx sinensis.