Salvatore Desantis

In lizards, the longitudinal canal conveys the sperm from the rete testis to the ductuli efferentes, which connect to the cranial end of the epididymal duct. Due to its reduced extension, the longitudinal canal has been largely ignored compared with the ductuli efferentes and the ductus epididymis. The objective of this study was to provide information on the structure and function of the longitudinal canal in the lizard Podarcis siculus by using light and transmission electron microscopy. For this purpose, testes and epididymides from adult male P. siculus lizards captured during the reproductive period were used. Light microscopy showed that the longitudinal canal was lined by a simple squamous or low cuboidal epithelium containing apical periodic acid-Schiff (PAS)-positive granules. Transmission electron microscopy revealed two types of epithelial cells: ciliated and non-ciliated. The non-ciliated cells showed electron-dense granules in the supranuclear and apical regions, suggesting probable secretory activity. These findings demonstrate, for the first time, that the longitudinal canal of the lizard Podarcis siculus is not merely a passageway for the sperm but may play a role in the composition of the medium in which these gametes are found during their flow towards the epididymis.

Vitamin D (VD) is required for testis activity being a key regulator of calcium and testosterone, both essential for spermatogenesis and sperm maturation. VD is activated by a metabolizing machinery of enzymes (CYP2R1, CYP27A1, CYP27B1) and acts throughout its receptor (VDR). Cryptorchidism is a developmental defect caused by the failed scrotal descend of the testicle and can compromise fertility depending on whether one or both testes are retained. Here, we reported the expression and localization of VDR- and VD-activating enzymes in the testis of normal and unilateral cryptorchid equines obtained by Real-Time PCR, Western blotting, and immunohistochemistry. Results demonstrated that cryptorchidism affected gene and protein expression. Leydig cells expressed the highest level of VDR, CYP2R1, CYP27A1, and CYP27B1, whilst CYP27A1 and CYP27B1 were more abundant in spermatids from normal testes compared to scrotal testes of unilateral cryptorchid animals. No different cellular localization of the VD-metabolizing machinery was observed between cryptorchid and scrotal testicles, although some variation in staining intensity was observed. These results provide insights for further studies, required for clarifying VD’s role in equine reproduction.

The white‐eared opossum, Didelphis albiventris , is an opportunistic and omnivorous marsupial, whose diet ranges from wild fruits to eggs and birds. Salivary glycoproteins play a key role in the protection of the oral cavity and the formation of the food bolus. Despite the importance of salivary glycoproteins, their detailed investigation in the white‐eared is lacking. This study investigated the morphology and glycan composition of the mandibular salivary glands of the white‐eared opossum for the first time. Histological and histochemical investigations were conducted on tissue fragments fixed with 4% PBS‐buffered paraformaldehyde and embedded in Paraplast. The pattern of glycoproteins was investigated using traditional histochemical methods (PAS, Alcian Blue pH 2.5, and High‐Iron Diamine staining) and lectin histochemistry. The glandular parenchyma consisted of acinar secretory units and a duct system characterized by abundant striated ducts. Secretory acini secrete neutral glycans and non‐sulfated acid glycans. Mannosylated N‐linked glycans terminating in α2,6‐sialic acid and fucose are expressed in the secretory acini, containing intraluminal α2,3‐sialylated O‐linked glycans. The epithelial lining of the striated and interlobular ducts also shows O‐linked glycans with terminal Galβ1, 3GalNAc, and α GalNAc residues. Finally, the epithelium and lumen of interlobular ducts are enriched with additional GalNAc‐terminated O‐linked glycans with the appearance of lactosaminated glycans and the disappearance of α2,3‐sialylated glycans. These results suggest that the saliva produced by the mandibular gland of the white‐eared opossum consists of a species‐specific pattern of glycoproteins, to whose composition the ductal system also contributes. The observed glycan composition is probably related to the diet of the white‐eared opossum and its adaptations to the environment and food availability. These results indicate that the mandibular salivary gland of the white‐eared opossum Didelphis albiventris has specific histological and molecular characteristics compared to other marsupial species, suggesting that diet and habitat, but not the taxonomic group, influence the mandibular gland features.

The inability to distinguish males from females in young birds is a major obstacle for pair formation in reintroduction–restocking programs and commercial–amateur breeding. Several techniques are employed to address this problem, but not all of them are suitable for juvenile subjects. Among the various tests applied for sex determination, polymerase chain reaction (PCR) is one of the genetic tools that seems to be most effective (rapid, not invasive and cheaper). In this study, DNA was extracted from down and feathers to make the procedure less stressful for nestlings. The DNA was amplified by PCR, and the amplicon was subjected to the restriction endonucleases procedure when the gender was not clearly identified by PCR alone. One hundred and fifty-three avian species were sexed using this procedure, including 27 for the first time. In all the nestlings and juveniles tested, sex was correctly identified; in fact, all pairs that reached sexual maturity during this study gave offspring.

Extracellular vesicles (EVs) are important mediators of cell–cell communication thanks to their ability to transfer their bioactive cargo, thus regulating a variety of physiological contexts. EVs derived from amniotic mesenchymal/stromal cells (eAMC-EVs) are internalized by equine endometrial cells (eECs) with positive effects on regenerative medicine treatments. As the cellular uptake of EVs is influenced by the glycan profile of both EVs and target cells, this study is focused on the role of surface glycans in the uptake of eAMC-EVs by recipient eECs. Equine ECs were obtained by enzymatic digestion of uteri from healthy mares. Equine AMC-EVs were isolated from amniotic cell cultures according to a standardized protocol. The glycan pattern was studied using a panel of lectins in combination with fucosidase and neuraminidase treatment. Both eECs and eAMC-EVs expressed N-linked high mannose glycans, as well as fucosylated and sialylated glycans. All these glycans were involved in the uptake of eAMC-EVs by eECs. The internalization of eAMC-EVs was strongly reduced after cleavage of α1,2-linked fucose and α2,3/α2,6-linked sialic acids. These results demonstrate that surface glycans are involved in the internalization of eAMC-EVs by eECs and that fucosylated and sialylated glycans are highly relevant in the transfer of bioactive molecules with effects on regenerative medicine treatments.

Phenolic antioxidant intake is encouraged to prevent oxidative damage, and antioxidant synergy is considered an advantage in adding polyphenols from varied plants. This study investigated the antioxidant and synergistic interactions among olive leaf (OL), bay laurel (BL), and rosemary (RL) leaf powder mixture (LPM: OL + BL + RL), using in vitro chemical tests [TPC, ORAC, TEAC-ABTS, FRAP; combination index (CI)], and in vivo validation on blood oxidative status, metabolic profile, and intestinal histomorphology in laying hens. The in vitro study indicated a whole higher antioxidant capacity for the LPM than respective single/double-leave combinations. The LPM CI value (IC50, 0.60) indicated a synergistic effect compared to the binary combinations. Thus, the LPM was validated in vivo through dietary supplementation on sixty Lohmann Brown hens (30 weeks old), reared in an indoor–outdoor rearing system divided. The hens were allocated into two experimental groups (n. 30): basal control diet group; and diet supplemented group with 6 g/kg feed of LPM) containing OL, BL, and RL (respectively, at 65.7%:18.9%:15.4%), for 60 days. The LPM improved (p < 0.05) the oxidative status (TAS, FRAP; ROMs, TBARs) and vitamin E level, metabolic and immunological profiles, and it induced region-specific changes in the morphology and carbohydrate composition of mucins along intestinal tracts of the animals. These findings could provide a valuable strategy for identifying synergistic combinations in functional feed formulations for laying hens.

The oviductal fimbria is the first extraovarian anatomical structure that the cumulus–oocyte complex (COC) encounters, and is sensitive to sex hormone changes. The morphology, glycan pattern, expression of heat shock proteins (HSPs), estradiol receptor (ER), and progesterone receptor (PR) were investigated in the oviductal fimbria epithelium of the baboon (Papio hamadryas) during the menstrual cycle. The morphology was investigated by light and scanning electron microscopy; the glycopattern was characterized using conventional and lectin histochemistry; HSPs (60, −70, −90), ER, and PR were localized immunohistochemically. Well-differentiated ciliated and nonciliated cells were present only during the preovulatory phase. The nonciliated cells contained small apical protrusions and thin microvilli. During the preovulatory phase (1) the luminal surface of the fimbria displayed acidic glycans, complex N-glycans containing fucose, and oligolactosamine residues; (2) nonciliated cells expressed HSP60 and HSP90 in the apical blebs, HSP70 in the nucleus and cytoplasm, as well as nuclear ERα and PR; (3) ciliated cells showed HSP70 in the nucleus, cytoplasm, and cilia that also expressed HSP90 and PR. These results are related to the function of the fimbria where the early COC–oviduct crosstalk occurs and may represent a benchmark for translational studies of other primates.

Aquaporins (AQPs) are important for water transport in the gastrointestinal tract. Changes in their expression and/or localization could cause in disorders and be used as therapeutic targets. Aquaporin‐4 (AQP4) is expressed predominantly on the basolateral membrane of the parietal cells in the corpus of the murine gastric glands. Although the secretion of gastric juice is not affected in AQP4‐deficient knockout, we evaluated by light microscopy whether the lack of AQP4 affects the glycopatterns of secreting gastric cells. Wild type (WT) and AQP4‐deficient knockout mice (KO) were fed a standard diet ad libitum before sacrifice. Segments of stomach corpus were collected, fixed in buffered formalin, and embedded in paraffin wax. Sections, 5‐μm thick, were analyzed by histochemical methods (Periodic acid‐Schiff, Alcian Blue pH 2.5), and binding of lectins specific to GalNAc (SBA, DBA), Gal (PNA) GlcNAc (WGA, GSAII) mannose and/or glucose (ConA), and fucose (UEA‐I, AAA, LTA). Immunohistochemical methods such as anti‐Muc6 for neck cells and anti‐ β‐ H⁺/K⁺‐ATPase for parietal cells were also performed. Compared to WT mice, in the mucous cells of KO lower amounts of glycans with galactosyl/galactosaminylated, glycosyl/glycosaminylated, and fucosylated residues were observed; lower fucosylation resulted also in the parietal cells. The observed differences of KO in respect to WT could lead to severer pathological conditions.Research Highlights: Glycopatterns in gastric glands were compared between wild type (WT) and AQP4‐deficient knockout (KO) mice by histochemical and lectin‐binding methods. In the mucous cells of KO lower amounts of glycans with galactosyl/galactosaminylated, glycosyl/glycosaminylated and fucosylated residues were observed. In the parietal cells lower fucosylation also resulted. AQP4‐deficiency affects glycosylation and could result in altered functionality and pathological conditions.