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Hibernation is a biological status during which hibernating animals acclimatize themselves to reduced energy consumption through extreme but governed decline in self-metabolism. The role of mitochondria (Mt) in metabolic suppression during hibernation has already been elaborated in different organs and species. Nonetheless, the concretely changing process of mitochondrial architecture and the mechanism underlying this transformation during hibernation remains unclear. Herein, the present study was aimed at clarifying the detailed alteration of mitochondrial morphology and its potential role in the Chinese soft-shelled turtle (Pelodiscus sinensis) during different stages of hibernation. Compared with the nonhibernation period, the mitochondrial architecture was changing from round to crescent, and lipid droplet (LD)/Mt interaction was enhanced during hibernation, as observed by transmission electron microscopy (TEM). Further ultrastructural analysis uncovered that mitochondrial fusion was promptly accelerated in the early stage of hibernation, followed by mitochondrial fission in the middle stage, and mitophagy was boosted in the late stage. Moreover, gene and protein expression related to mitochondrial fusion, fission, and mitophagy accorded closely with the mitochondrial ultrastructural changes in different stages of hibernation. Taken together, our results clarified that the transformation of mitochondrial architecture and mitochondrial dynamics are of vital importance in maintaining internal environment homeostasis of Pelodiscus sinensis.
The blood-brain barrier (BBB) is an important internal barrier. Herein, the electron microscope examination of duck BBB was performed during the brain development. Meanwhile, the genes/proteins of tight junctions (TJs) including zonula occludens-1, occludin, and claudin-5 in the duck brain were detected by Q-PCR and immunohistochemistry. The results showed the density of capillaries in the brain gradually increased during the embryonic period. The generation of the BBB and the specialization of its components occurred mainly in the embryonic stage. During this period, the endothelial cells (ECs) became thinner and pinocytic vesicles decreased; the TJs between EC membranes became longer and more electron-dense; the basement membrane surrounding ECs and pericytes gradually thickened; and the astrocyte foot processes appeared to wrap around the vessels. By the day of hatching (P1), the whole set of duck BBB structures was completely assembled and gradually improved in the subsequent growth process. Interestingly, compared with the cerebrum and cerebellum, the maturity level of the midbrain BBB was earlier seen during the embryonic stage. The expression of TJs increased during the embryonic period and remained stable by post-hatching. The study systematically investigated the histochemical and ultrastructural features of duck BBB during development and explored the corresponding relationship between structure and function.
In order to clarify fine structures of the hypothetical meridian conduits of Chinese traditional medicine (CTM) in the skin, the present study used light and transmission electron microscopy to examine fasciae in different vertebrate species. Collagen fiber bundles and layers were arranged in a crisscross pattern, which developed into a special tissue micro-channel (TMC) network, in a manner that was analogs to the proposed skin meridian conduits. It was further revealed that tissue fluid in lateral TMC branches drained into wide longitudinal channels, which were distinctly different from lymphatic capillary. Mast cells, macrophages, and extracellular vesicles such as ectosomes and exosomes were distributed around telocytes (TCs) and their long processes (Telopodes, Tps) within the TMC. Cell junctions between TCs developed, which could enable the communication between contiguous but distant Tps. On the other hand, winding free Tps without cell junctions were also uncovered inside the TMC. Tissue fluid, cell junctions of TCs, mast cells, macrophages, and extracellular vesicles within the TMC corresponded to the circulating “气血” (“Qi-Xue”, i.e., information, message, and energy) of meridian conduits at the cytological level. These results could provide morphological evidence for the hypothesis that “meridians are the conduit for Qi-Xue circulation” in CTM.
This paper proposes a set of novel indices for evaluating the kinematic performance of a 3-RRS (R and S denote revolute and spherical joint respectively, R denotes active joint.), parallel mechanism whose translational and rotational movements are strongly coupled. First, the indices are formulated using the decoupled overall Jacobian matrix, which is developed using coordinate transformation. Then, the influences of the homogeneous dimensionless parameters on these indices are investigated. In addition, the dimension synthesis of the 3-RRS parallel mechanism is carried out by minimizing the mean value of the kinematic performance indices and their standard deviation. The results demonstrate that the established approach facilitates good global kinematic performance of the parallel mechanism.
Telocytes (TCs) are very long, non-neuronal, somatic cells whose function is widely believed to be involved in providing connections between different cells within the body. The cellular characteristics of TCs in various organs have been studied by immunohistochemistry, double immunofluorescence and electron microscopy in different vertebrate species, and here we investigate the proposed properties of these cells in the context of the “meridian” in Chinese Traditional Medicine (CTM). The results show that TCs and their long extensions, telopodes (Tps) develop a complicated network by homo- and heterocellular junctions in the connective tissue throughout the body, which can connect the skin with distant organs. In concept, this is the analogue of ancient meridian maps connecting skin acupoints with the viscera. Various active cells and extracellular vesicles including exosomes move along Tps, which, along with developed mitochondria within the podoms of Tps, may account for the structural evidence for “Qi” (vital energy and signal communication) in CTM. Morphological associations of TCs with the nerve, vascular, endocrine, and immune systems are also compatible with previously proposed meridian theories in CTM. Close relationships exist between TCs and collagen fiber bundles and some structures in skin fascia provide the microanatomical support for acupuncture treatment based on the meridian principle. The dynamicity in the distribution and structure of TCs reflects the plasticity of the meridian at the cellular level. As the same attribute, both the meridian and the TC have been associated with various diseases. Here, we summarize structural analogues between the TC and the meridian, suggesting that TCs have the cytological characteristics of the CTM meridian. We, therefore, hypothesize that TCs are the “essence cells” of the CTM meridian, which can connect and integrate different cells and structures in the connective tissue.
The seminiferous tubule (ST) is the location of spermatogenesis, where mature spermatozoa are produced with the assistance of Sertoli cells. The role of extracellular vesicles in the direct communication between Sertoli-germ cells in the ST is still not fully understood. In this study, we reported multivesicular bodies (MVBs) and their source of CD63-enriched exosomes by light and ultrastructure microscopy during the reproductive phases of turtles. Strong CD63 immunopositivity was detected at the basal region in the early and luminal regions of the ST during late spermatogenesis by immunohistochemistry (IHC), immunofluorescence (IF), and western blot (WB) analysis. Labeling of CD63 was detected in the Sertoli cell cytoplasmic processes that surround the developing germ cells during early spermatogenesis and in the lumen of the ST with elongated spermatids during late spermatogenesis. Furthermore, ultrastructure analysis confirmed the existence of numerous MVBs in the Sertoli cell prolongations that surround the round and primary spermatogonia during acrosome biogenesis and with the embedded heads of spermatids in the cytoplasm of Sertoli cells. Additionally, in spermatids, Chrysanthemum flower centers (CFCs) generated isolated membranes involved in MVBs and autophagosome formation, and their fusion to form amphiosomes was also observed. Additionally, autophagy inhibition by 3-methyladenine (after 24 h) increased CD63 protein signals during late spermatogenesis, as detected by IF and WB. Collectively, our study found MVBs and CD63 rich exosomes within the Sertoli cells and their response to autophagy inhibition in the ST during the spermatogenesis in the turtle.
The present study was designed to investigate the in vivo biological processes of multivesicular bodies (MVBs) and exosomes in mitochondria-rich cells (MRCs), goblet cells (GCs), and absorptive cells (ACs) in turtle intestines during hibernation. The exosome markers, cluster of differentiation 63 (CD63) and tumor susceptibility gene 101 (TSG101), were positively expressed in intestinal villi during turtle hibernation. The distribution and formation processes of MVBs and exosomes in turtle MRCs, GCs, and ACs were further confirmed by transmission electron microscopy. During hibernation, abundantly secreted early endosomes (ees) were localized in the luminal and basal cytoplasm of the MRCs and ACs, and late endosomes (les) were dispersed with the supranuclear parts of the MRCs and ACs. Many “heterogeneous” MVBs were identified throughout the cytoplasm of the MRCs and ACs. Interestingly, the ees, les, and MVBs were detected in the cytoplasm of the GCs during hibernation; however, they were absent during nonhibernation. Furthermore, the exocytosis pathways of exosomes and autophagic vacuoles were observed in the MRCs, GCs, and ACs during hibernation. In addition, the number of different MVBs with intraluminal vesicles (ILVs) and heterogeneous endosome–MVB–exosome complexes was significantly increased in the MRCs, GCs, and ACs during hibernation. All these findings indicate that intestinal epithelial cells potentially perform a role in the secretion of MVBs and exosomes, which are essential for mucosal immunity, during hibernation.
Fine structure observations were performed by means of electron microscopy on oogenesis and vitellogenesis and the special functions of follicular cells in the Chinese soft-shelled turtle (Pelodiseus sinensis). Histological examination of the ovary showed a well developed lacunae system containing fine granules, fibres or gelatiniform materials with one or two germinal beds dispersed on the dorsal surface of the ovarian cortex. The process of oogenesis comprised 10 consecutive phases according to the morphology of the yolk platelets, follicular cells and zona pellucida (ZP). Electron microscopy of vitellogenesis revealed some of the mitochondria gradually being transformed into yolk granules. In the advanced stage of vitellogenesis, large amounts of rough endoplasmic reticula, Golgiosomes and other cell organelles that are involved in synthesis and secretion were observed in follicular cells. The ZP was formed by microvilli, thus increasing the absorptive surface of the oocyte and facilitating transport of nutrients from the follicular epithelium to the ooplasm. This study demonstrated that the ovaries of members of the Testudinidae share more features with Archosaurs than with Squamates, indicating that these features were phylogenetically conserved in the Archosauria. The present observations suggest that the accumulation of yolk materials was controlled by the intrinsic and extrinsic pathways as well as by the activity of follicular cells. These results might also support a sibling relationship of the Testudinidae with the Archosauria and not with all extant reptiles.
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