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Pattern Distribution of Connexins in the Ortho- and Parakeratinized Epithelium of the Lingual Mucosa in Birds
Connexins are important proteins involved in cell-to-cell communication and cytodifferentiation during renewal and cornification of the multilayered epithelia. So far, there is a lack of reports on this subject in birds’ structurally different ortho- and parakeratinized epithelium of the tongue. The study aims to describe the distribution and expression profiles of the α-connexins (Cx40 and 43) and β-connexins (Cx26, 30, and 31) in those epithelia in duck, goose, and domestic turkey. Research revealed the presence of the mentioned connexins and the occurrence of interspecies differences. Connexins form gap junctions in the cell membrane or are in the cytoplasm of keratinocytes. Differences in connexin expression were noted between the basal and intermediate layers, which may determine the proliferation of keratinocytes. Cx40, 43, and Cx30 in the gap junction of the keratinocytes of the intermediate layer are related to the synchronization of the cornification process. Because of the exfoliation of cornified plaques, a lack of connexins was observed in the cornified layer of orthokeratinized epithelium. However, in parakeratinized epithelium, connexins were present in the cell membrane of keratinocytes and thus maintained cellular integrity in gradually desquamating cells. The current studies will be useful in further comparative analyses of normal and pathological epithelia of the oral cavity in birds.
Distribution and length of muscle spindles and their 3D visualisation in the medial gastrocnemius of male and female rats
AbstractThe spatial distribution of the medial gastrocnemius muscle spindles of 10 male and 10 female rats was analysed under a light microscope, and for the first time, visualised using a 3D model of the muscle. Serial cross‐sections of the medial gastrocnemius muscles were separated into 10 divisions along with the proximo‐distal axis. The muscle spindles of the rat medial gastrocnemius were predominantly distributed on the proximo‐medial divisions of the muscle. There were no sex‐related differences in the distribution of the studied receptors. A single division contained an average of 2.71 receptors for animals of both sexes. Moreover, the calculated lengths of male and female rat muscle spindles were comparable, and average lengths did not significantly differ (3.30 ± 1.47 mm for male and 3.26 ± 1.32 mm for female rats). Therefore, the present results fill gaps in recent observations concerning similarities in muscle spindle numbers between male and female animals, despite significant differences in muscle mass and size.
Discovery of mammalian collagens I and III within ancient poriferan biopolymer spongin
Abstract Spongin is a fundamental biopolymer that has played a crucial role in the skeletogenesis of keratosan sponges for over 800 million years. This biomaterial had so far remained chemically unidentified and believed to be an enigmatic type of halogenated collagen-keratin-based bioelastomer. Here we show collagen I and III as the main structural components of spongin. Proteomics, 13C solid state NMR and Raman spectroscopy confirm the identity of collagenous domains in spongin with collagen from mammals. Using an HPLC-MS analysis, we found halogenated di- and tri-tyrosines as crosslinking agents in spongin. Using molecular dynamics modeling, we solvated the crystal structures of collagen mimetic peptides for type I and type III collagens in four different systems, including selected brominated crosslinks. The results underscore the complex interplay between the collagen structures and crosslinks, raising intriguing questions about the molecular mechanisms underlying collagen chemistry within spongin as an ancient biocomposite.
Histogenesis of the Uterine Horn in the Domestic Cat (Felis silvestris catus): LM, TEM, and SEM Study
This study employs light microscopy, scanning electron microscopy, and transmission electron microscopy to describe the morphological changes occurring during the development of the domestic cat’s uterine horns, originating from the uterine segments of paramesonephric ducts (uPD). Comprehensive observations conducted on 60 specimens aged 28–63 days post-conception (p.c.) revealed that the formation of the endometrium and myometrium in the uterine horns begins around day 33 p.c., initiated by mesenchymal differentiation. During endometrial development, fibroblasts align first in perpendicular and then in oblique columns. The subdivision of the lamina propria into basal and functional layers becomes evident shortly before birth, with the functional layer remaining flat until the end of the prenatal period. The endometrial epithelium transforms from a simple columnar to a pseudostratified structure, undulating by day 63 p.c. Myometrial formation commences with the differentiation of myoblasts, which are arranged in a circular pattern. By the end of gestation, these myoblasts differentiate into smooth muscle cells, organizing into distinct inner circular and outer longitudinal sublayers. Although the fundamental layered architecture of the uterine wall is established before birth, its full maturation—including gland formation, epithelial transformation, and further development of the myometrium—continues postnatally.
Alpha-Keratin, Keratin-Associated Proteins and Transglutaminase 1 Are Present in the Ortho- and Parakeratinized Epithelium of the Avian Tongue
The lingual mucosa in birds is covered with two specific types of multilayered epithelia, i.e., the para- and orthokeratinized epithelium, that differ structurally and functionally. Comprehensive information on proteins synthesized in keratinocyte during their cytodifferentiation in subsequent layers of multilayered epithelia in birds concerns only the epidermis and are missing the epithelia of the lingual mucosa. The aim of the present study was to perform an immunohistochemical (IHC) and molecular analysis (WB) of bird-specific alpha-keratin, keratin-associated proteins (KAPs), namely filaggrin and loricrin, as well as transglutaminase 1 in the para- and orthokeratinized epithelium covering the tongue in the domestic duck, goose, and turkey. The results reveal the presence of alpha-keratin and KAPs in both epithelia, which is a sign of the cornification process. In contrast to the epidermis, the main KAPs involved in the cornification process of the lingual epithelia in birds is loricrin. Stronger expression with KAPs and transglutaminase 1 in the orthokeratinized epithelium than in the parakeratinized epithelium may determine the formation of a more efficient protective mechanical barrier. The presence of alpha-keratin, KAPs, and transglutaminase 1 epitopes characteristic of epidermal cornification in both types of the lingual epithelia may prove that they are of ectodermal origin.
The three-dimensional analysis of gustatory papillae and its taste buds on the tongue of the wild-living hare (Lepus europaeus), European rabbit (Oryctolagus cuniculus), and domestic rabbit (Oryctolagus cuniculus f. domestica)
Prenatal development of the intramural vascular system of the uterine tube in the European shorthair cat
Context:Early vascular development during the prenatal period is essential for the proper formation of all organs, including the reproductive system.Aims: This study aimed to characterize the development of the intramural vascular system in the domestic cat's uterine tube, with emphasis on changes associated with wall layer formation.Methods: Uterine tubes from 38 domestic cat fetuses (36-63 days post-conception) were examined using light microscopy, vascular corrosion casts, and scanning electron microscopy.Key Results:In early stages, the vascular system of the rudimentary paramesonephric duct (uPD) is simple, comprising longitudinal vessels and a superficial vascular network. Initially, all vessels resemble capillaries. By days 42-43 p.c., vascular differentiation into arterioles and venules becomes evident, progressing to arteries and veins with distinct surface relief visible on corrosion casts by late gestation. Around day 50 p.c., as the layers develop in the uterine tube wall, a subepithelial capillary network emerges, connecting with the superficial vascular system. At term, the uterine artery becomes tortuous and branches above the isthmus; veins accompany the arteries. The mature intramural vascular system includes circumferential arteries and veins, as well as subepithelial and subserosal capillary networks.Conclusions:A basic vascular architecture supporting tissue growth develops prenatally, laying the foundation for postnatal expansion linked to mucosal and muscular layer maturation.Implications:This first-time analysis of feline uterine tube prenatal angioarchitecture provides novel insights into reproductive system histogenesis in a species both common in human environments and important in veterinary science.
Morphogenesis of the uterine tube in the European shorthair cat in the prenatal and postnatal period: Light microscopy and three-dimensional reconstruction study
Three-dimensional reconstruction of gustatory papillae and its taste buds in short-hair cats (Felis Catus domestica, felidae, Carnivora)
Effects of Tcte1 knockout on energy chain transportation and spermatogenesis: implications for male infertility
Abstract STUDY QUESTION Is the Tcte1 mutation causative for male infertility? SUMMARY ANSWER Our collected data underline the complex and devastating effect of the single-gene mutation on the testicular molecular network, leading to male reproductive failure. WHAT IS KNOWN ALREADY Recent data have revealed mutations in genes related to axonemal dynein arms as causative for morphology and motility abnormalities in spermatozoa of infertile males, including dysplasia of fibrous sheath (DFS) and multiple morphological abnormalities in the sperm flagella (MMAF). The nexin–dynein regulatory complex (N-DRC) coordinates the dynein arm activity and is built from the DRC1–DRC7 proteins. DRC5 (TCTE1), one of the N-DRC elements, has already been reported as a candidate for abnormal sperm flagella beating; however, only in a restricted manner with no clear explanation of respective observations. STUDY DESIGN, SIZE, DURATION Using the CRISPR/Cas9 genome editing technique, a mouse Tcte1 gene knockout line was created on the basis of the C57Bl/6J strain. The mouse reproductive potential, semen characteristics, testicular gene expression levels, sperm ATP, and testis apoptosis level measurements were then assessed, followed by visualization of N-DRC proteins in sperm, and protein modeling in silico. Also, a pilot genomic sequencing study of samples from human infertile males (n = 248) was applied for screening of TCTE1 variants. PARTICIPANTS/MATERIALS, SETTING, METHODS To check the reproductive potential of KO mice, adult animals were crossed for delivery of three litters per caged pair, but for no longer than for 6 months, in various combinations of zygosity. All experiments were performed for wild-type (WT, control group), heterozygous Tcte1+/− and homozygous Tcte1−/− male mice. Gross anatomy was performed on testis and epididymis samples, followed by semen analysis. Sequencing of RNA (RNAseq; Illumina) was done for mice testis tissues. STRING interactions were checked for protein–protein interactions, based on changed expression levels of corresponding genes identified in the mouse testis RNAseq experiments. Immunofluorescence in situ staining was performed to detect the N-DRC complex proteins: Tcte1 (Drc5), Drc7, Fbxl13 (Drc6), and Eps8l1 (Drc3) in mouse spermatozoa. To determine the amount of ATP in spermatozoa, the luminescence level was measured. In addition, immunofluorescence in situ staining was performed to check the level of apoptosis via caspase 3 visualization on mouse testis samples. DNA from whole blood samples of infertile males (n = 137 with non-obstructive azoospermia or cryptozoospermia, n = 111 samples with a spectrum of oligoasthenoteratozoospermia, including n = 47 with asthenozoospermia) was extracted to perform genomic sequencing (WGS, WES, or Sanger). Protein prediction modeling of human-identified variants and the exon 3 structure deleted in the mouse knockout was also performed. MAIN RESULTS AND THE ROLE OF CHANCE No progeny at all was found for the homozygous males which were revealed to have oligoasthenoteratozoospermia, while heterozygous animals were fertile but manifested oligozoospermia, suggesting haploinsufficiency. RNA-sequencing of the testicular tissue showed the influence of Tcte1 mutations on the expression pattern of 21 genes responsible for mitochondrial ATP processing or linked with apoptosis or spermatogenesis. In Tcte1−/− males, the protein was revealed in only residual amounts in the sperm head nucleus and was not transported to the sperm flagella, as were other N-DRC components. Decreased ATP levels (2.4-fold lower) were found in the spermatozoa of homozygous mice, together with disturbed tail:midpiece ratios, leading to abnormal sperm tail beating. Casp3-positive signals (indicating apoptosis) were observed in spermatogonia only, at a similar level in all three mouse genotypes. Mutation screening of human infertile males revealed one novel and five ultra-rare heterogeneous variants (predicted as disease-causing) in 6.05% of the patients studied. Protein prediction modeling of identified variants revealed changes in the protein surface charge potential, leading to disruption in helix flexibility or its dynamics, thus suggesting disrupted interactions of TCTE1 with its binding partners located within the axoneme. LARGE SCALE DATA All data generated or analyzed during this study are included in this published article and its supplementary information files. RNAseq data are available in the GEO database (https://www.ncbi.nlm.nih.gov/geo/) under the accession number GSE207805. The results described in the publication are based on whole-genome or exome sequencing data which includes sensitive information in the form of patient-specific germline variants. Information regarding such variants must not be shared publicly following European Union legislation, therefore access to raw data that support the findings of this study are available from the corresponding author upon reasonable request. LIMITATIONS, REASONS FOR CAUTION In the study, the in vitro fertilization performance of sperm from homozygous male mice was not checked. WIDER IMPLICATIONS OF THE FINDINGS This study contains novel and comprehensive data concerning the role of TCTE1 in male infertility. The TCTE1 gene is the next one that should be added to the ‘male infertility list’ because of its crucial role in spermatogenesis and proper sperm functioning. STUDY FUNDING/COMPETING INTEREST(S) This work was supported by National Science Centre in Poland, grants no.: 2015/17/B/NZ2/01157 and 2020/37/B/NZ5/00549 (to M.K.), 2017/26/D/NZ5/00789 (to A.M.), and HD096723, GM127569-03, NIH SAP #4100085736 PA DoH (to A.N.Y.). The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.
Three-dimensional characteristic of fungiform papillae and its taste buds in European bison (Bison bonasus), cattle (Bos taurus), and Bison bonasus hybrid
Abstract Background Our recent macro- and scanning electron microscopic study of tongue conducted on domesticated cattle, wild living European bison, and Bison bonasus hybrid revealed various spatial arrangement and number of gustatory and mechanical papillae between parental species and their hybrid. Furthermore, scanning electron microscopy analysis of gustatory papillae indicated the variable distribution of fungiform papillae (Fu) over the surface of the tongue, which could be significant in differentiated taste perception during feeding in studied wild living and domesticated husbandry ruminants. To specify the detailed microstructure of Fu papillae with connective tissue cores (CTC) and intraepithelial taste buds system, the first time the three-dimensional computer-aided analysis of serial histoslides resulted in the rendering of 3D reconstructions of Fu papillae. Results The comparative analysis of 3D models Fu papillae conducted in six areas of lingual mucosa of each tongue revealed information about, microstructural diversity of Fu papillae in studied ruminants. The estimation of number and density of Fu papillae on tongues, rate of protrusion of papillae over mucosa, and a number of taste buds per papilla allowed to state the ventral surface of the lingual apex and posterolateral surfaces of the lingual torus as regions important in taste perception, as in the preselection of taken food, as well in the analysis of food during rumination, respectively. On the 3D models were observed three structural types of CTC of different distribution on the tongue in studied species. The quantitative data of the number of taste buds on Fu papillae have regional functional differences in the taste system important in feeding and veterinary practice. Moreover, our analysis determined specific features in examined hybrid and showed similarities of some studied features with cattle, i.e., maternal species. Conclusions The 3D reconstruction method used for the first time in the field of study of the lingual papillae and taste buds system can be considered as an innovative and effective tool in assessing of the microstructures of Fu papillae, and it could be suitable for further studies of taste system structures in normal and pathological condition.