Salvatore Desantis - Academia.edu (original) (raw)

Papers by Salvatore Desantis

Annals of Anatomy - Anatomischer Anzeiger, 2014

Reproduction, Fertility and Development, 2014

ABSTRACT Epithelial to mesenchymal transition (EMT) is the process by which epithelial cells dram... more ABSTRACT Epithelial to mesenchymal transition (EMT) is the process by which epithelial cells dramatically alter their shape and motile behaviour as they differentiate into mesenchymal cells. The EMT and the reverse process, termed mesenchymal-epithelial transition, play central roles in embryogenesis. Gastrulation and neural crest formation are processes governed by EMT in amniotes. It is noteworthy that in placental mammals, the epithelial layer of amnion originates from the trophectoderm and it is continuous with the epiblast. On this basis, it is reasonable to speculate that some amniotic epithelial cells may escape the specification that accompanies gastrulation, and may retain some of the characteristics of epiblastic cells, such as pluripotency, behaving as stem cells that are able to preserve intrinsically the ability to transdifferentiate. Because it seems that malignant cells use the same mechanisms during the formation of tumours in vivo, the amniotic epithelial cells (AEC) could represent a good model to study in vitro this phenomenon that we observed to occur spontaneously in our culture conditions. The aim of this study was to characterise the glycoprotein pattern expressed in fresh or cryopreserved equine AEC, mesenchymal (AMC), and transdifferentiated cells by means of lectin histochemistry. AEC and AMC were cultured until passage (P) 3, while transdifferentiated cells at P1(EMT1) and P2 (EMT2). All cell lines were frozen for 1 month at -196°C in liquid nitrogen. The glycoanalysis was performed with a panel of twelve lectins to detect the glycans terminating with sialic acids (MAL II, SNA, PNA after sialidase digestion (K-s), K-s-DBA), galactose (PNA, RCA120, GSA I-B4,), N-acetylgalactosamine (DBA, HPA, SBA), N-acetylglucosamine (GSA II), fucose (UEA I, LTA), or with internal mannose (Con A). After freezing: 1) AEC exhibited decrease of binding sites for DBA, SBA, HPA, GSA II, and disappearance of GSA I-B4 and UEA I binders; 2) AMC displayed increase of SBA reactivity, decrease of K-s-PNA, HPA, GSA II staining, and absence of GSA I-B4 affinity; 3) EMT1 cells showed the appearance of K-s-DBA staining, the increase of K-s-PNA, RCA120, SBA, GSA I-B4, and UEA I reactivity, the decrease of MAL II, SNA, HPA, GSA II binders, and the disappearance of DBA and LTA binding sites; 4) EMT2 cells revealed the increase of K-s-PNA, GSA I-B4, UEA I affinity, the decrease of MAL II, SNA, RCA120, HPA, GSA II binders, and the lack of DBA, SBA, and LTA reactivity. In conclusion, this study demonstrates that the EMT induces changes in cell surface glycan profile of equine amniotic progenitor cells, and for the first time revealed that freezing modifies the lectin binding pattern of these cells. The observed glycan pattern modification may represent one aspect of the spontaneous complex process of EMT.

Histology and histopathology, 2010

Ultrastructural and biochemical features of efferent ducts (EDs) are indicative of an intense abs... more Ultrastructural and biochemical features of efferent ducts (EDs) are indicative of an intense absorptive activity towards the luminal fluid. This function was investigated by 1) the immunohistochemical localization of different aquaporins, integral membrane water channels that facilitate rapid passive movement of water, and 2) the histochemical localization of lectins, known to have specific affinity for glycoconjugate residues. AQP1 was mostly revealed at the apical surface and adluminal cytoplasm of non-ciliated cells and to a minor extent in their lateral plasma membrane, whereas it was absent in ciliated cells. Blood vessels showed AQP1-immunoreactivity, which was present in endothelial cells of venous vessels and capillaries and around the muscular sheath of arteries. AQP9 was expressed in the apical zone of ciliated and non-ciliated cells and in the lateral cell membrane. AQP2 and AQP5 were undetectable. Lectin histochemistry showed that non-ciliated cells contain glycans with...

Histology and histopathology, 2008

The presence of the mu-opioid receptor and the type of glycosylation in the third extra-cellular ... more The presence of the mu-opioid receptor and the type of glycosylation in the third extra-cellular loop of this receptor was investigated in the isthmus of mare oviduct during oestrus by means of immunoblotting and immunohistochemistry combined with enzymatic (N-glycosidase F and O-glycosidase) and chemical (beta-elimination) treatments. Immunoblotting analysis showed that the mu-opioid receptor consists of two peptides with molecular weights of around 65 and 50 kDa. After N-deglycosylation with N-glycosidase F an additional immunoreactive peptide was observed at around 30 KDa. The cleavage of O-glycans by O-glycosidase failed in immunoblotting as well as in immunohistochemistry investigations, revealing that the third extra-cellular loop of the mu-opioid receptor expressed in mare isthmus oviduct contains some modifications of the Galbeta(1-3)GalNAc core binding to serine or threonine. Immunohistochemistry revealed the mu-opioid receptor in the mucosal epithelium, some stromal cells,...

The Journal of Applied Poultry Research, 2008

Ochratoxins are fungal secondary metabolites that contaminate grains, legumes, coffee, dried frui... more Ochratoxins are fungal secondary metabolites that contaminate grains, legumes, coffee, dried fruits, beer, wine, and meat. Ochratoxins are considered powerful nephrotoxins, carcinogens, teratogens, and immunotoxins in rats and likely in humans. In 2006 during a pilot program of Hazard Analysis Critical Control Point in the Apulian region, Southern Italy, ochratoxin A (OA) was detected in commercial feed for laying hens in 2 farms. The OA-contaminated feed had been administered to animals for at least 2 mo. Analysis by HPLC with fluorometric detection of the tissues of 4 layer hens that displayed gross and microscopical lesions identified OA in the kidney (8.7 to 16.9 g/ kg, average 13.65 ± 3.58 g/kg) and liver (3.7 to 5.1 g/kg, average 4.43 ± 0.64 g/kg) but not in the other tissues.

Histology and histopathology, 2007

The carbohydrate expression in the epithelium lining the oesophagus of the toadfish Halobatrachus... more The carbohydrate expression in the epithelium lining the oesophagus of the toadfish Halobatrachus didactylus was studied by means of conventional and lectin histochemistry. The stratified epithelium was constituted by basal cells, polymorphous cells in the intermediate layer, pyramidal and flattened cells in the outer layer and contained two types of large secretory cells: goblet cells and sacciform cells. PAS, Alcian blue pH 2.5 and pH 1.0 stained very strongly the goblet cells, weakly the surface of the other epithelial cells but did not stain the sacciform cells. The goblet cells cytoplasm contained oligosaccharides with terminal Galbeta1,3GalNAc, alpha/betaGalNAc, Galbeta1,4GlcNAc, alphaL-Fuc and internal betaGlcNAc residues (PNA, SBA, RCA120, UEA I, LTA and KOH-sialidase-WGA affinity). Galbeta1,4GlcNAc, alphaL-Fuc and internal betaGlcNAc were also found in the glycocalyx. The sacciform cells expressed sialyloligosaccharides terminating with Neu5Acalpha2,3Galbeta1,4GlcNac, Neu5A...

European journal of histochemistry : EJH

We investigated the oligosaccharide sequence of glycoconjugates, mainly sialoglycoconjugates, in ... more We investigated the oligosaccharide sequence of glycoconjugates, mainly sialoglycoconjugates, in the horse oviductal ampulla during oestrus by means of lectin and pre-lectin methods such as the KOH-neuraminidase procedure to remove sialic acid residues and incubation with N-glycosidase F to cleave N-linked glycans. Ciliated cells displayed N-linked oligosaccharides throughout the cytoplasm. The cilia glycocalyx expressed both N- and O-linked (mucin-type) oligosaccharides, both showing a high variety of terminal sequences. In the most non-ciliated cells, the whole cytoplasm contained N-linked oligosaccharides with terminal alphaGal as well as mucin-type glycans with terminal Forssman pentasaccharides. In a few scattered non-ciliated cells, the whole cytoplasm displayed sialylated N-linked oligosaccharides with terminal Neu5Ac-GalNAc and O-linked glycans terminating with neutral and/or alphaGalNAc, Neu5Ac alpha2,6Gal/GalNAc, Neu5AcGal beta1,3GalNAc. Supra-nuclear granules, probably Go...

The Histochemical journal

The regional difference in the carbohydrate components of the ductus epididymis epithelium of a l... more The regional difference in the carbohydrate components of the ductus epididymis epithelium of a lizard was delineated by means of 13 lectins. Basal cells expressed only N-acetylglucosamine (GlcNAc). Throughout the ductus, the secretory cells showed oligosaccharides with terminal N-acetylneuraminic acid (Neu5Ac)alpha(2,6)galactose (Gal)/N-acetylgalactosamine (GalNAc) and internal mannose (Man) and/or glucose (Glc) in the whole cytoplasm, oligosaccharides terminating in Neu5Ac alpha(2,6)Gal beta(1,3)GalNAc, Neu5Ac alpha(2,6)Gal beta(1,4)GlcNAc, GalNAc, GlcNAc, and fucose (Fuc) in the supra-nuclear zone, and also glycans terminating in Neu5Ac alpha(2,3)Gal beta(1,4)GlcNAc, Neu5Ac alpha(2,6)Gal beta(1,3)GalNAc, Gal beta(1,4)GlcNAc on the luminal surface. In the caput and corpus regions, the supra-nuclear cytoplasm was characterized by terminal Gal beta(1,4)GlcNAc and alphaGalNAc, the luminal surface by alphaGalNAc and Gal. The Golgi zone, showing oligosaccharides with terminal Neu5Ac al...

European journal of histochemistry : EJH, 2000

The pars distalis from the pituitary gland of adult male ruin lizards (Podarcis sicula campestris... more The pars distalis from the pituitary gland of adult male ruin lizards (Podarcis sicula campestris De Betta), captured during the five periods of the annual sexual cycle (emergence from hibernation, reproductive period, summer regression, autumnal recrudescence, winter arrest), was studied immunohistochemically using specific antibodies against hFSHbeta, hLHbeta, oFSHbeta, and oLHbeta with the avidin-biotin complex (ABC) procedure to monitor the seasonal changes in shape, size and percentage area taken up from gonadotropes. FSH containing cells were specifically identified with anti-hFSHbeta and anti-oFSHbeta sera, whereas the LH cells were localized by anti-hLHbeta. The anti-oLHbeta serum showed cross-reactivity with the cells immunostained by the three above antisera (anti-hFSHbeta, anti-oFSHbeta, and anti-hLHbeta). None of the cells contained both gonadotropic hormones as shown by the double-immunostaining procedure. Generally, FSH cells were larger and more numerous than LH cells...

European journal of histochemistry : EJH, 1995

The distribution of glycoconjugates in the testis of lizard Podarcis s. campestris De Betta was i... more The distribution of glycoconjugates in the testis of lizard Podarcis s. campestris De Betta was investigated by HRP-conjugated lectins during the annual spermatogenetic cycle. In addition, pretreatments of sections with neuraminidase and removal of alkali-labile O-linked sequences by beta-elimination allowed the structure of glycoconjugates to be further explored. Leydig cells displayed changes of lectin-binding sites during annual cycle, and during the abortive spermatogenesis period lacked N-linked sialylgalactosyl glycans. Sertoli cells stained by Con A, WGA, RCA120, BS I-B4, showed, except in July, O-linked sialylgalactosyl glycans. Spermatogonia bound Con A and WGA. Spermatocytes bound also BS I-B4, SBA, UEA I, and during spring spermatogenesis, revealed O-linked sialylgalactosyl glycans. The acrosomes of spermatids were also stained by RCA120 and PNA, whereas the heads of spermatozoa did not bind SBA and PNA. During the abortive spermatogenic period, the acrosomes showed O-lin...

Theriogenology, 2014

Epithelium of oviductal ampulla was studied in normal and in superovulated sheep using morphologi... more Epithelium of oviductal ampulla was studied in normal and in superovulated sheep using morphologic analysis and lectin glycohistochemistry. The lining epithelium consisted of two types of cells, ciliated and nonciliated cells. Unlike superovulated samples, the nonciliated cells from control ewes showed apical protrusions indicating an apocrine secretory activity. The ciliated cells showed lectin-binding sites mainly at the level of the cilia which bound all the used lectins except Peanut agglutinin, suggesting the lack of glycans terminating with Galb1,3GalNAc. In superovulated specimens, the ciliated cells with high mannosylated glycans Concanavalin A (Con A) and GlcNAc and GalNac termini Griffonia simplicifolia agglutinin II (GSA II) and Dolicurus biflorus agglutinin (DBA) decreased. The luminal surface of nonciliated cells showed all investigated sugar residues in controls, whereas it was lacking in high mannosylated (Con A) and terminal GalNAca1,3(LFuca1,2) Galb1,3/4GlcNAcb1 sequence (DBA) in superovulated ewes. Apical protrusions from control ampullae nonciliated cells showed glycans containing mannose, GlcNac, GalNAc, galactose, and a2,3-linked sialic acid Con A, KOH-sialidase-Wheat germ agglutnin [WGA], GSA II, SBA, Griffonia simplicifolia agglutinin-isolectin B 4 [GSA I-B 4 ], Maackia amurensis agglutinin II [MAL II]). The supranuclear cytoplasm of nonciliated cells expressed terminal GlcNAc (GSA II) in all specimens, also O-linked glycans (mucin-type glycans) with GalNAc and sialic acid termini (Helix pomatia agglutinin [HPA] and MAL II) in control animals, and also N-linked glycans with fucose, galactose, lactosamine, and a2,3-linked sialic acid termini (Ulex europaeus agglutinin I [UEA I], GSA I-B 4 , Ricinus communis agglutinin 120 [RCA 120 ], and Sambucus nigra agglutinin [SNA] ) in superovulated ewes. These results report for the first time that the superovulation treatment affects the secretory activity and the glycan pattern of the epithelium lining the sheep oviductal ampulla.

European Journal of Histochemistry, 2008

The present study was focused on the morphology of the diencephalic nuclei (likely involved in re... more The present study was focused on the morphology of the diencephalic nuclei (likely involved in reproductive functions) as well as on the distribution of GnRH (gonadotropin-releasing hormone) in the rhinencephalon, telencephalon and the diencephalon of the brain of bluefin tuna (Thunnus thynnus) by means of immunohistochemistry. Bluefin tuna has an encephalization quotient (QE) similar to that of other large pelagic fish. Its brain exhibits well-developed optic tecta and corpus cerebelli. The diencephalic neuron cell bodies involved in reproductive functions are grouped in two main nuclei: the nucleus preopticus-periventricularis and the nucleus lateralis tuberis. The nucleus preopticus-periventricularis consists of the nucleus periventricularis and the nucleus preopticus consisting of a few sparse multipolar neurons in the rostral part and numerous cells closely packed and arranged in several layers in its aboral part. The nucleus lateralis tuberis is located in the ventral-lateral area of the diencephalon and is made up of a number of large multipolar neurones. Four different polyclonal primary antibodies against salmon (s)GnRH, chicken (c)GnRH-II (cGnRH-II 675, cGnRH-II 6) and sea bream (sb)GnRH were employed in the immunohistochemical experiments. No immunoreactive structures were found with anti sbGnRH serum. sGnRH and cGnRH-II antisera revealed immunoreactivity in the perikarya of the olfactory bulbs, preopticus-periventricular nucleus, oculomotor nucleus and midbrain tegmentum. The nucleus lateralis tuberis showed immunostaining only with anti-sGnRH serum. Nerve fibres immunoreactive to cGnRH and sGnRH sera were found in the olfactory bulbs, olfactory nerve and neurohypophysis. The significance of the distribution of the GnRHimmunoreactive neuronal structures is discussed.

European Journal of Histochemistry, 2004

The distribution of sialic acid residues as well as other glycosidic sugars has been investigated... more The distribution of sialic acid residues as well as other glycosidic sugars has been investigated in the horse oviductal isthmus during anoestrus, oestrus and pregnancy by means of lectin and pre-lectin methods. Ciliated cells and non-ciliated (secretory) cells exhibited different lectin binding profiles that were found to change during the investigated stages. Ciliated cells did not show any reactivity in the basal cytoplasm, while the supra-nuclear cytoplasm displayed a few of oligosaccharides with terminal and internal αmannose (Man) and/or αglucose (Glc) during oestrus and pregnancy and a moderate presence of oligosaccharides terminating in αfucose (Fuc) during oestrus; cilia exhibited a more complex glycoconjugate pattern for the presence of oligosaccharides terminating in N-acetylgalactosamine (GalNAc), GalNAcα1,3 GalNAcα1,3galactose(Gal)β1,4Galβ1,4N-acetylglucosamine(GlcNAc), Fuc, sialic acid (Neu5Ac)-αGalNAc belonging or not to the GalNAcα1,3GalNAcα1,3 Galβ1,4 Galβ1, 4GlcNAc sequence, and. αGalNAc and Neu5Acα 2,6Gal/GalNAc increased during oestrus. Cilia displayed terminal Galβ1,3 GalNAc in pregnancy, terminal αGal in anoestrus and pregnancy and terminal or internal D-GlcNAc during anoestrus and pregnancy, respectively. The whole cytoplasm of non-ciliated cells showed oligosaccharides terminating with αGalNAc, Neu5Acα2,6Gal/GalNAc, Neu5Ac GalNAcα 1,3GalNAcα1,3Galβ1,4Galβ1,4GlcNAc during the investigated stages, as well as GlcNAc in anoestrus and pregnancy. The supra-nuclear zone of non-ciliated cells exhibited oligosaccharides with terminal Galβ1,4GlcNAc and internal Man during oestrus and pregnancy as well as terminal αGal and Fuc in oestrus and Neu5Ac-Galβ1,3GalNAc in pregnancy. The luminal surface of non-ciliated cells showed glycans terminating with αGalNAc and/or Neu5Ac GalNAcα1,3 GalNAcα1,3Galβ1,4Galβ1,4GlcNAc in all specimens, oligosaccharides with terminal Galβ1,4GlcNAc and internal Man during oestrus and pregnancy, Neu5Ac α2,6Gal/GalNAc in anoestrus and oestrus, and glycans terminating with Galβ1,3GalNAc, Neu5A acα2,3 Galβ1, 4GlcNac, Neu5ac-Galβ1,3GalNAc, Neu5Ac-Galβ1,4 GlcNAc in pregnancy. These findings show the presence of sialoglycoconjugates in the oviductal isthmus of the mare as well as the existence of great modifications in the glycoconjugates linked to different physiological conditions.

Advances in Decapod Crustacean Research, 2001

Ovaries from mature giant red shrimp Aristaeomorpha foliacea were investigated histochemically an... more Ovaries from mature giant red shrimp Aristaeomorpha foliacea were investigated histochemically and ultrastructurally. Four growing stages of the oocytes were distinguished: premeiosis stage, previtellogenetic stage, early vitellogenic stage and late vitellogenic stage. In addition, occasional resorptive oocytes were found. Oogonia and premeiotic oocytes were found in germinative zones. Previtellogenic and vitellogenic oocytes were localized in maturative zones. As vitellogenesis proceeded,

Life Sciences, 2000

Prenatal exposure to low concentrations of carbon monoxide (CO, 75 and 150 ppm from day 0 to day ... more Prenatal exposure to low concentrations of carbon monoxide (CO, 75 and 150 ppm from day 0 to day 20 of gestation), resulting in maternal blood HbCO concentrations equivalent to those maintained by human cigarette smokers, leads to subtle myelin alterations in the sciatic nerve of male rat offspring. The rapid growth spurt in pup body weight was related to the period of maximal increase in myelin sheath thickness in both control and CO-exposed animals. A significant reduction in myelin sheath thickness of sciatic nerve fibers, paralleled by changes in the frequency distribution, occurred in both 40- and 90-day-old rats exposed in utero to CO (75 and 150 ppm). Myelin deficit observed in 75 and 150 ppm CO-exposed animals showed up only after the major spurt in myelination but not early during development. The subtle myelin alterations observed in CO-exposed offspring were not accompanied by changes in developmental pattern of axon diameters and did not result in a gross impairment of motor activity. These results suggest that the myelination process is selectively targeted by a prenatal exposure model simulating the CO exposure observed in human cigarette smokers.

Theriogenology, 2011

Morphometric, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) inves... more Morphometric, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) investigations have displayed regional differences in the mare oviductal epithelium. The entire mucosa of the oviduct was lined with a pseudostratified epithelium, which consisted of two distinct cell types, ciliated and non-ciliated. Ciliated cells were predominant in the three different segments of the oviduct and their percentage increased from fimbriae to ampulla and significantly decreased in the isthmus. SEM revealed in the infundibulum finger-like mucosal folds, some of them interconnected, in the ampulla numerous and elaborated branched folds of the mucosa, whereas the isthmus displayed a narrow lumen, short and non-branched mucosal folds. In the ampulla and isthmus the majority of non-ciliated cells showed apical blebs provided or not of short microvilli. TEM displayed different ultrastructural features of ciliated and non-ciliated cells along the oviduct. Isthmus ciliated cells presented a more electron-dense cytoplasm than in infundibulum and ampulla cells and its cilia were enclosed in an amorphous matrix. The non-ciliated cells of infundibulum did not contain secretory granules but some apical endocytic vesicles and microvilli coated by a well developed glycocalyx. Non-ciliated cells of ampulla and isthmus contained secretory granules. Apical protrusions of ampulla displayed two types of secretory granules as well as occasional electron-lucent vesicles. Isthmus non-ciliated cells showed either electron-lucent or electron-dense cytoplasm and not all contained apical protrusions. The electron-dense non-ciliated cells displayed microvilli coated with a well developed glycocalyx. Three types of granules were observed in the isthmus non-ciliated cells. The regional differences observed along the epithelium lining the mare oviduct suggest that the epithelium of the each segment is involved in the production of a distinctive microenvironment with a unique biochemical milieu related to its functional role.

Theriogenology, 2009

Equine cumulus-oocyte complexes (COCs) were analyzed by means of 13 lectins to evaluate their gly... more Equine cumulus-oocyte complexes (COCs) were analyzed by means of 13 lectins to evaluate their glycoconjugate patterns and to verify differences between COCs recovered with compact (Cp) and expanded (Exp) cumulus. Cumulus cells showed a similar staining pattern in both Cp and Exp COCs with all lectins used, except for a higher reactivity with SNA and GSA II in Cp COCs and SBA in Exp COCs. The zona pellucida (ZP) showed (1) uniform staining with MAL II, RCA(120), and SBA in both Cp and Exp COCs, (2) trilaminar binding pattern with WGA as well as higher Con A reactivity in the outer region of both types of COCs, (3) uniform staining with PNA only in Exp COCs, (4) uniform and trilaminar binding pattern with SNA in Cp and Exp COCs, respectively, and (5) major reactivity with GSA II in Exp COCs. Ooplasm showed similar staining intensity with Con A, HPA, GSA I-B(4), and WGA in both Cp and Exp COCs, with stronger reactivity to GSA II in Exp COCs, whereas SNA, UEA I, and LTA binding sites were present only in Cp COCs. Oocyte cortical granules of both Cp and Exp COCs reacted with Con A and WGA. These results suggest that, in the mare, viable (Cp) and atretic (Exp) COCs display different glycoconjugate staining pattern, which may account for the different maturation and developmental competence of COCs.

Theriogenology, 2010

Stallion sperm from semen collected in Southern Italy during the breeding (June-July) and non-bre... more Stallion sperm from semen collected in Southern Italy during the breeding (June-July) and non-breeding (December-January) periods were analyzed by means of twelve lectins to evaluate the glycoconjugate pattern and to verify whether there are any seasonal differences in the glycosylation pattern of the sperm glycocalyx. The acrosomal cap showed reactivity for Maackia amurensis (MAL II), Sambucus nigra (SNA), Arachis hypogaea (PNA), Glycine max (SBA), Helix pomatia (HPA), Canavalia ensiformis (Con A) Triticum vulgaris (WGA), and Griffonia simplicifolia isolectin II (GSA II) in breeding and non-breeding ejaculated sperm, suggesting the presence of oligosaccharides terminating with Neu5Ac alpha 2,3Gal beta 1,4GlcNAc, Neu5Ac alpha 2,6Gal/GalNAc, with Gal beta 1,3GalNAc, alpha/beta GalNAc and glycans with terminal/internal alpha Man and GlcNAc. During the non-breeding period, the acrosomal cap expressed oligosaccharides terminating with Gal beta 1,4GlcNAc (Ricinus communis(120) affinity) (RCA(120)) and L-Fuc alpha 1,2Gal beta 1,4GlcNAc beta (Ulex europaeus affinity) (UEA I). The equatorial segment placed between the acrosomal cap and post-acrosomal region did not display glycans terminating with GalNAc, GlcNAc, and alpha L-Fuc. The post-acrosomal region of sperm collected in the breeding and non-breeding periods bound Con A, MAL II, SNA, and SBA, thus showing the presence of N-linked oligosaccharides from high-Man content, terminating with Neu5Ac alpha 2,3Gal beta 1,4GlcNAc, Neu5Ac alpha 2,6Gal/GalNAc and GalNAc. In winter, the post-acrosomal region also expressed oligosaccharides terminating with alpha GalNAc, GlcNAc, and L-Fuc alpha 1,2Gal beta 1,4GlcNAc beta (HPA, GSA II, and UEA I staining). The tail of sperm from semen collected during the breeding and non-breeding periods showed a lectin binding pattern similar to the post-acrosomal region, except for the absence of HPA staining in sperm collected during the winter season. These results indicate that the surface of stallion sperm contains different glycocalyx domains and that the glycosylation pattern undergoes changes during the breeding and non-breeding periods.

Scientia Marina, 2009

toadfish Halobatrachus didactylus gastric mucosa was studied using conventional and lectin histoc... more toadfish Halobatrachus didactylus gastric mucosa was studied using conventional and lectin histochemistry. conventional histochemistry revealed neutral glycoconjugates predominating over acidic ones in the apical zone of both surface epithelial cells and pit cells. the neck cells contained a few neutral glycoconjugates, whereas gastric glands were negative to PaS and aB staining. lectin histochemistry showed different oligosaccharide expression along the columnar cells. the sub-nuclear cytoplasm was stained with rca 120 , SBa, hPa, GSa i-B 4 , GSa ii, uea i, lta, con a, Kohsialidase-WGa. the Golgi zone reacted with rca 120 , DBa, SBa, hPa, Koh-sialidase-WGa, GSa i-B 4 , GSa ii, uea i, lta, Mal ii, Sna, and showed an increase in DBa staining after Koh-sialidase treatment. the granules of the apical zone stained with Pna, uea i, lta and showed increased Pna reactivity after Koh-sialidase treatment. the luminal cell coat reacted with Pna, hPa, con a, Koh-sialidase-WGa, uea i, lta, Mal ii, Sna and Koh-sialidase-Pna. Pit cells showed a minor expression of lectin-binding sites with respect to columnar cells. neck cells linked uea i and lta and gastric glands reacted with Pna, DBa, SBa, hPa, con a, GSa i-B 4 , Koh-sialidase-WGa and Koh-sialidase-DBa. the results suggest that the stomach of the toadfish H. didactylus is characterised by a species-specific glycoconjugate pattern. reSuMen: Expresión glucohistoquímica en el estómago del pez sapo, HalobatracHus didactylus (Schneider, 1801). -Se ha estudiado la expresión de residuos glucídicos de glucoconjugados en la mucosa gástrica del pez sapo, Halobatrachus didactylus usando histoquímica convencional y de lectinas. la histoquímica clásica reveló la presencia de glicoconjugados neutros predominando sobre los ácidos en la zona apical de la superficie de las células epiteliales y de la criptas gástricas. las células del cuello contienen algunas glicoproteínas neutras, mientras las glándulas gástricas han sido negativas al PaS y al azul alcián (aB). la histoquímica de lectinas mostró diferente expresión de oligosacáridos a lo largo de las células columnares. el citoplasma sub-nuclear reaccionó con las lectinas rca, SBa, hPa, GSa, i-B4, GSa ii, uea i, lta, con a, Koh-sialidasa-WGa. la expresión de rca 120 , DBa, SBa, hPa, Koh-sialidasa-WGa, GSa i-B 4 , GSa ii, uea i, lta, Mal ii y Sna se localizó en la zona del Golgi, donde se observó un incremento de la reactividad de la lectina DBa después del tratamiento con Koh. los gránulos de la zona apical reaccionaron con Pna, uea i, lta, mostrando un incremento de la reactividad hacia la lectina Pna después del tratamiento con Koh. el borde de las células del lumen reaccionó con Pna, hPa, cona, Koh-sialidasa-WGa, uea i, lta, Mal ii, Sna, Koh-sialidasa-Pna. los sitios de unión de la expresión de las lectinas-glucoconjugados fue menor en las células del cuello de las criptas gastricas que en las células columnares epiteliales. las células del cuello reaccionaron con la uea i y lta, y las glándulas gástricas con Pna, DBa, SBa, hPa, con a, GSa i-B 4 , Koh-sialidasa-WGa y Koh-sialidasa-DBa. los resultados sugieren que el estómago del pez sapo, Halobatrachus didactylus, se caracteriza por un patrón especie-específico de expresión de glicoconjugados.

Reproduction, 2013

Oviductal environment affects preparation of gametes for fertilization, fertilization itself, and... more Oviductal environment affects preparation of gametes for fertilization, fertilization itself, and subsequent embryonic development. The aim of this study was to evaluate the effect of oviductal fluid and the possible involvement of deleted in malignant brain tumor 1 (DMBT1) on IVF in porcine and equine species that represent divergent IVF models. We first performed IVF after pre-incubation of oocytes with or without oviductal fluid supplemented or not with antibodies directed against DMBT1. We showed that oviductal fluid induces an increase in the monospermic fertilization rate and that this effect is canceled by the addition of antibodies, in both porcine and equine species. Moreover, pre-incubation of oocytes with recombinant DMBT1 induces an increase in the monospermic fertilization rate in the pig, confirming an involvement of DMBT1 in the fertilization process. The presence of DMBT1 in the oviduct at different stages of the estrus cycle was shown by western blot and confirmed by immunohistochemical analysis of ampulla and isthmus regions. The presence of DMBT1 in cumulus-oocyte complexes was shown by western blot analysis, and the localization of DMBT1 in the zona pellucida and cytoplasm of equine and porcine oocytes was observed using immunofluorescence analysis and confocal microscopy. Moreover, we showed an interaction between DMBT1 and porcine spermatozoa using surface plasmon resonance studies. Finally, a bioinformatic and phylogenetic analysis allowed us to identify the DMBT1 protein as well as a DMBT1-like protein in several mammals. Our results strongly suggest an important role of DMBT1 in the process of fertilization.