Seasonal and Ageing-Depending Changes of Aquaporins 1 and 9 Expression in the Genital Tract of Buffalo Bulls (Bubalus bubalis).

The presence of Aquaporins 1 (AQP1) and 9 (AQP9), integral membrane water channels that facilitate rapid passive movement of water and solutes, was immunohistochemically detected in the excurrent ducts collected from sexually mature buffalo bulls of proven fertility during the mating (late autumn-winter) and non-mating (late spring to the beginning of autumn) seasons. Furthermore, the research was performed also on the epididymal cauda of a senile buffalo bull with inactive testis. Aquaporins 1 and 9 were immunolocalized at distinct levels. In the efferent ducts, AQP1 immunoreactivity was strongly evidenced at the apical surface of the non-ciliated cells and weakly along the basal membrane of the epithelial cells. The latter reactivity disappeared during the non-mating season. No AQP1 immunoreactivity was detected in the epithelium of epididymis and vas deferens, whereas AQP1 was expressed in the smooth muscle layer of the vas deferens. Aquaporin 1 was present in the blood vessels and in small nerve bundles all along the genital tract. The supranuclear zone of the epididymal principal cells was AQP9 immunoreactive, limited to the corpus and cauda regions, and vas deferens. The samples collected in the two reproductive seasons showed a weaker AQP9 immunoreactivity during the non-mating season. A typical AQP9 immunoreactivity was noticed in the old buffalo examined. The tested AQP molecules showed a different expression pattern in comparison with laboratory mammals, primates, equine, dog and cat. In addition, seasonal differences were noticed which are possibly useful in regard to the comprehension of the morphophysiology of reproduction in the bubaline species, which are still a matter of debate.

A phase II study of bevacizumab in combination with vinorelbine and trastuzumab in HER2-positive metastatic breast cancer.

We aimed to evaluate the efficacy and feasibility of combining trastuzumab/vinorelbine with bevacizumab in patients with first-or second-line HER2-positive, metastatic breast cancer (MBC). Eligible patients had HER2-positive measureable MBC, with no more than one prior line of chemotherapy, and were treated with trastuzumab (4 mg/kg × 2 mg/kg weekly thereafter), vinorelbine (25 mg/m(2) weekly), and bevacizumab (10 mg/kg every 2 weeks). Co-primary endpoints were (a) the proportion of patients alive and progression-free at 1 year and (b) safety profile/feasibility. Feasibility was defined as a rate of grade 3/4 non-hematologic toxicity attributable to protocol-based therapy <20 %. Twenty-nine patients were enrolled (n = 22 first-line, n = 7 second-line). Median age was 48 years (range 37-68). The median number of cycles received was 8 (1-23) and median duration on treatment was 7.4 months (range 1-22). The study was closed early due to higher-than-expected rates of grade 3/4 non-hematologic toxicities, with 50 events in 20 patients. A total of six patients (21 %) were taken off study for treatment-related toxicity. Most common treatment-related toxicities included fatigue (n = 7), febrile neutropenia (n = 4), and headache (n = 3). At 1 year, 8/22 first-line (36 %) and 2/7 second-line (29 %) patients were alive and progression-free. Median PFS was 9.9 months and 7.8 months in the first- and second-line cohorts, respectively. Objective responses were observed in 16/22 (73 %) and 5/7 (71 %) patients in the first- and second-line settings. Although the combination of vinorelbine, trastuzumab, and bevacizumab showed notable activity in HER2-positive MBC, the proportion of first-line patients alive and progression-free at 1 year was deemed unlikely to reach the pre-defined threshold for declaring success. Additionally, unacceptable toxicity was observed, at rates greater than previously reported with vinorelbine/trastuzumab or vinorelbine/bevacizumab doublet combinations.

The effect of neuromuscular blockade on mask ventilation.

We wished to test the hypothesis that neuromuscular blockade facilitates mask ventilation. In order reliably and reproducibly to assess the efficiency of mask ventilation, we developed a novel grading scale (Warters scale), based on attempts to generate a standardised tidal volume. Following induction of general anaesthesia, a blinded anaesthesia provider assessed mask ventilation in 90 patients using our novel grading scale. The non-blinded anaesthesiologist then randomly administered rocuronium or normal saline. After 2 min, mask ventilation was reassessed by the blinded practitioner. Rocuronium significantly improved ventilation scores on the Warters scale (mean (SD) 2.3 (1.6) vs 1.2 (0.9), p<0.001). In a subgroup of patients with a baseline Warters scale value of >3 (i.e. difficult to mask ventilate; n=14), the ventilation scores also showed significant improvement (4.2 (1.2) vs 1.9 (1.0), p=0.0002). Saline administration had no effect on ventilation scores. Our data indicate that neuromuscular blockade facilitates mask ventilation. We discuss the implications of this finding for unexpected difficult airway management and for the practice of confirming adequate mask ventilation before the administration of neuromuscular blockade.

Ramifications of protease-based liquefaction of camel semen on physical, kinematic and surface glyco-pattern of cryopreserved spermatozoa.

The efficiency of incorporating different proteases in the diluent for reducing camel semen viscosity, and subsequent ramifications on morpho-functional and glycan surface properties of cryopreserved spermatozoa were investigated. Ejaculates (n = 48) were collected from three adult camels, Camelus dromedarius, during the breeding season (January - March). A portion of each raw ejaculate was evaluated for sperm physical and morphological traits, whereas the other portion was divided into three aliquots assigned for the following liquefaction treatments: control (untreated), 0.1 mg/mL papain or 5 U/mL bromelain. All samples were diluted with Tris-lactose diluent containing the anti-enzyme E-64 to neutralize both proteases before being processed for cryopreservation. Post-thaw physical and kinematic properties of spermatozoa were analyzed using a computer-assisted sperm analysis (CASA) system. The sperm surface glycocalyx pattern was evaluated with a panel of 14 fluorescent lectins. Although bromelain was more effective in elimination of semen viscosity, there was a negative correlation between bromelain supplementation and values for the variables: normal sperm, intact acrosome and intact sperm cell membrane. Bromelain supplementation, compared to papain-treated and control samples, was positively correlated with secondary sperm abnormalities, increased straight-line velocity (VSL, µm/s) and straightness (%) of spermatozoa. Results from the glycan analysis indicated that both proteases did not affect the N-linked glycan content of the entire sperm surface, whereas the treatment with proteases induced little change in N-acetylgalactosamine and fucose terminating glycans in the tail region of the sperm. Functional studies are needed to evaluate the sperm fertility rates of bromelain- and papain-treated semen for application in camel assisted reproductive technologies.

Brain morphology and immunohistochemical localization of the gonadotropin-releasing hormone in the bluefin tuna, Thunnus thynnus.

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 GnRH-immunoreactive neuronal structures is discussed.

Differential surface glycoprofile of buffalo bull spermatozoa during mating and non-mating periods.

The buffalo has a seasonal reproduction activity with mating and non-mating periods occurring from late autumn to winter and from late spring to beginning of autumn, respectively. Sperm glycocalyx plays an important role in reproduction as it is the first interface between sperm and environment. Semen quality is poorer during non-mating periods, so we aimed to evaluate if there were also seasonal differences in the surface glycosylation pattern of mating period spermatozoa (MPS) compared with non-mating period spermatozoa (NMPS). The complexity of carbohydrate structures makes their analysis challenging, and recently the high-throughput microarray approach is now providing a new tool into the evaluation of cell glycosylation status. We adopted a novel procedure in which spermatozoa was spotted on microarray slides, incubated with a panel of 12 biotinylated lectins and Cy3-conjugated streptavidin, and then signal intensity was detected using a microarray scanner. Both MPS and NMPS microarrays reacted with all the lectins and revealed that the expression of (i) O-glycans with NeuNAcα2-3Galß1,3(±NeuNAcα2-6)GalNAc, Galß1,3GalNAc and GalNAcα1,3(l-Fucα1,2)Galß1,3/4GlcNAcß1 was not season dependent; (ii) O-linked glycans terminating with GalNAc, asialo N-linked glycans terminating with Galß1,4GlcNAc, GlcNAc, as well as α1,6 and α1,2-linked fucosylated oligosaccharides was predominant in MPS; (iii) high mannose- and biantennary complex types N-glycans terminating with α2,6 sialic acids and terminal galactose were lower in MPS. Overall, this innovative cell microarray method was able to identify specific glycosylation changes that occur on buffalo bull sperm surface during the mating and non-mating periods.

Immunohistochemical localization of CYP1A, vitellogenin and Zona radiata proteins in the liver of swordfish (Xiphias gladius L.) taken from the Mediterranean Sea, South Atlantic, South Western Indian and Central North Pacific Oceans.

Cytochrome P4501A (CYP1A) monoxygenase, vitellogenin (Vtg) and Zona radiata proteins (Zrp) are frequently used as biomarkers of fish exposure to organic contaminants. In this work, swordfish liver sections obtained from the Mediterranean Sea, the South African coasts (South Atlantic and South Western Indian Oceans) and the Central North Pacific Ocean were immunostained with antisera against CYP1A, Zrp, and Vtg. CYP1A induction was found in hepatocytes, epithelium of the biliary ductus and the endothelium of large blood vessels of fish from the Mediterranean Sea and South African waters, but not from the Pacific Ocean. Zrp and Vtg were immunolocalized in hepatocytes of male swordfish from the Mediterranean Sea and from South African waters. Plasma Dot-Blot analysis, performed in Mediterranean and Pacific specimens, revealed the presence of Zrp and Vtg in males from Mediterranean but not from Pacific. These results confirm previous findings about the potential exposure of Mediterranean swordfish to endocrine, disrupting chemicals and raise questions concerning the possible presence of xenobiotic contaminants off the Southern coasts of South Africa in both the South Atlantic and South Western Indian Oceans.

Differential lectin binding patterns in the oviductal ampulla of the horse during oestrus.

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 Golgi zones, of non-ciliated cells showed mainly O-linked glycans rich in sialic acid residues. The luminal surface of non-ciliated cells showed N-linked oligosaccharides, containing terminal/internal alphaMan/alphaGlc, betaGlcNAc and terminal alphaGal, as well as mucin-type oligosaccharides terminating with a large variety of either neutral saccharides or sialylated sequences. Apical protrusions containing O-linked oligosaccharides with terminal Forssman pentasaccharide, Neu5Ac-Gal beta1,4GlcNAc, Neu5Ac-GalNAc were seen in non-ciliated cells scattered along the epithelium. These findings show the presence of sialoglycoconjugates in the oviductal ampulla epithelium of the mare and the existence of different lectin binding profiles between ciliated and non-ciliated (secretory) cells, as well as the presence of non-ciliated cell sub-types which might determine functional differences along the ampullary epithelium of mare oviduct.

Modifications of carbohydrate residues in the sheep oviductal ampulla after superovulation.

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 Galß1,3GalNAc. 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 GalNAcα1,3(LFucα1,2)Galß1,3/4GlcNAcß1 sequence (DBA) in superovulated ewes. Apical protrusions from control ampullae nonciliated cells showed glycans containing mannose, GlcNac, GalNAc, galactose, and α2,3-linked sialic acid (Con A, KOH-sialidase- Wheat germ agglutnin [WGA], GSA II, SBA, Griffonia simplicifolia agglutinin-isolectin B4 [GSA I-B4], 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 α2,3-linked sialic acid termini (Ulex europaeus agglutinin I [UEA I], GSA I-B4, Ricinus communis agglutinin120 [RCA120], 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.

The effect of antisecretagogues on gastric microflora.

We examined the effect of cimetidine in altering gastric microflora in patients undergoing gastric resection for peptic ulcer disease. Ten patients had administration of cimetidine stopped at least six hours before surgery. Their mean gastric pH was 2.02 +/- 1.51, and 0.4 +/- 0.97 organisms were isolated per patient. Ten patients were maintained on a regimen of cimetidine and their mean gastric pH was 5.69 +/- 1.80, with 3.2 +/- 1.62 organisms isolated per patient. We reached the following conclusions: cimetidine alters the gastric pH of preoperative patients; cimetidine therapy is associated with an increase in gastric flora; stopping administration of cimetidine six hours before surgery reacidifies the stomach, decreases the number of organisms isolated from the stomach, and decreases the risk of septic complications.

Prenatal exposure to low levels of carbon monoxide alters sciatic nerve myelination in rat offspring.

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.

Histochemical analysis of lizard testicular glycoconjugates during the annual spermatogenetic cycle.

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-linked sialylgalactosyl glycans and N-linked glycans terminating in beta-galactosyl residues. During the reproductive period, the acrosomes of spermatozoa expressed O- and N-linked sialylgalactosyl glycans and beta-galactosyl terminal residues on O- and N-linked glycans. This, in the testis of lizard, the two spermatogenesis periods show the emergence of different types of glycosylation.

Glycoconjugates during the annual sexual cycle in lizard epididymal ductuli efferentes: a histochemical study.

The epididymal ductuli efferentes of the lizard Podarcis sicula campestris De Betta are lined with simple, columnar, nonciliated and ciliated cells. The use of lectin histochemistry has provided information about changes of sugars associated with glycoconjugates of epithelial cells and intraluminal spermatozoa which are conveyed from the longitudinal canal to the cranial region of the ductus epididymis. Epithelial cells exhibited residues of alpha-D-mannose, N-acetylglucosamine, and beta-D-galactose-(1-4)-N-acetylglucosamine as revealed with lectins Con A, WGA and RCA120, respectively, throughout the sexual cycle. An increase of RCA120 staining was observed on microvilli of nonciliated cells and in the cytoplasm of ciliated cells during the reproductive period. However, during the following refractory period, when the organ was in regression, there was a decreased staining with Con A on microvilli and the absorbent surface of nonciliated cells, with WGA in nonciliated cells and the cytoplasm of ciliated cells, and with RCA120 on microvilli and the cytoplasm of both cell types. Terminal alpha-D-galactose residues were increasingly stained from autumn up to the reproductive period with BS I-B4 on microvilli, the absorbent surface and cilia, whereas they were entirely lacking during the refractory period. UEA I revealed alpha-L-fucose residues on the absorbent surface of nonciliated cells during the abortive and reproductive periods, increasing in the latter period when cilia also expressed this sugar. Terminal alpha/beta-D-N-acetylgalactosamine was evidenced with SBA on the absorbent surface of nonciliated cells during the reproductive period. The terminal beta-D-galactose-(1-3)-N-acetylgalactosamine dimer was never found with PNA, whereas O- and N-linked sialoglycoconjugates were present only during the reproductive period. The spermatozoa head exhibited N-linked glycans with high-mannose content, and beta-D-galactose-(1-4)-N-acetylglucosamine as well as O- and N-linked sialoglycoconjugates throughout the year. During the reproductive period, oligosaccharides with alpha-D-mannose residues increased, and oligosaccharides with terminal alpha-D-galactose, alpha-L-fucose and sialic acid-N-acetylgalactosamine dimer were also present. Unlike spermatozoa of seminiferous tubules, the spermatozoa head of the lizard epididymal ductuli efferentes exhibited seasonal variability in the lectin binding pattern which may be related to time-dependent changes in the glycoconjugate profiles of epithelial cells.

Immunohistochemical localization of FSH and LH in the pituitary of male ruin lizards (Podarcis sicula campestris De Betta).

The pars distalis from the pituitary gland of adult male ruin lizards (Podarcis sicula campestris De Betta), captured during the reproductive period (May and June), was studied immunohistochemically using specific antibodies against hFSH beta, hLH beta and oLH beta with the avidin-biotin-peroxidase complex (ABC) procedure to determine the localization of both gonadotropins. The immunostaining with anti-hFSH beta and anti-hLH beta allowed identification of morphologically distinct FSH containing cells and LH containing cells, whereas anti-oLH beta serum showed cross-reactivity with cells immunostained with the anti-hFSH beta and anti-hLH beta sera. The gonadotropic cells took up approximatively 10.5% of the area of the pars distalis: 10% was positive for FSH, whereas only 0.5% was positive for LH. The FSH cells were distributed throughout the pars distalis, whereas the LH cells were only located in the rostral region. Double-immunostaining procedure did not reveal cells containing both gonadotropic hormones. The findings indicate that FSH and LH are produced in separate pituitary cells.

Immunohistochemical evidence of seasonal changes of gonadotropes in male ruin lizard (Podarcis sicula campestris De Betta).

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. FSH cells were elongated or pyriform in shape from spring to autumn, whereas they were round or oval during the winter stasis and until the emergence from hibernation. The size and the percentage area occupied by FSH cells showed an annual pattern with two distinct peaks in the reproductive and in the autumnal recrudescence periods. LH cells did not show seasonal changes in shape, being round or oval throughout the reproductive cycle, whereas their size and the area they occupied underwent seasonal variations. The LH cells reached the largest size during the reproductive period and the smallest size during the summer regression. The percentage area occupied by LH cells in the pars distalis peaked at the emergence from hibernation and during the summer refractory period, when FSH cells displayed their lowest values.

Distribution of sialoglycoconjugates in the oviductal isthmus of the horse during anoestrus, oestrus and pregnancy: a lectin histochemistry study.

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 alphamannose (Man) and/or alphaglucose (Glc) during oestrus and pregnancy and a moderate presence of oligosaccharides terminating in alphafucose (Fuc) during oestrus; cilia exhibited a more complex glycoconjugate pattern for the presence of oligosaccharides terminating in N-acetylgalactosamine (GalNAc), GalNAcalpha1,3 GalNAcalpha1,3galactose(Gal)beta1,4Galbeta1,4N-acetylglucosamine(GlcNAc), Fuc, sialic acid (Neu5Ac)-aGalNAc belonging or not to the GalNAca1,3GalNAca1,3 Galb1,4 Galb1, 4GlcNAc sequence, and. alphaGalNAc and Neu5Aca 2,6Gal/GalNAc increased during oestrus. Cilia displayed terminal Galbeta1,3 GalNAc in pregnancy, terminal alphaGal 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 alphaGalNAc, Neu5Aca2,6Gal/GalNAc, Neu5Ac GalNAca 1,3GalNAcalpha1,3Galbeta1,4Galbeta1,4GlcNAc during the investigated stages, as well as GlcNAc in anoestrus and pregnancy. The supra-nuclear zone of non-ciliated cells exhibited oligosaccharides with terminal Galbeta1,4GlcNAc and internal Man during oestrus and pregnancy as well as terminal alphaGal and Fuc in oestrus and Neu5Ac-Galbeta1,3GalNAc in pregnancy. The luminal surface of non-ciliated cells showed glycans terminating with alphaGalNAc and/or Neu5Ac GalNAcalpha1,3 GalNAcalpha1,3Galbeta1,4Galbeta1,4GlcNAc in all specimens, oligosaccharides with terminal Galbeta1,4GlcNAc and internal Man during oestrus and pregnancy, Neu5Ac alpha2,6Gal/GalNAc in anoestrus and oestrus, and glycans terminating with Galbeta1,3GalNAc, Neu5A acalpha2,3 Galbeta1, 4GlcNac, Neu5ac-Galbeta1,3GalNAc, Neu5Ac-Galbeta1,4 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.

Histological and immunohistochemical investigation on ovarian development and plasma estradiol levels in the swordfish (Xiphias gladius L.).

The paper reports a histological and immunohistochemical description of oocyte growth and ultrastructural aspects of zona radiata (ZR) formation as well as the relationship between plasma estradiol-17beta, (E2) levels and ovarian development in swordfish (Xiphias gladius L.) from the Mediterranean Sea. Ovaries were inactive during March to mid April; maturation occurred during late April to June and spawning in June and July. Zona radiata formation starts, as Pas positive material, in oocytes at the lipid stage. In this stage a deposit of electrondense material between oolemma and follicular cells appears. In the cortical alveoli stage and through the early vitellogenic stage, the deposition of a moderately electrondense material occurred on the inner side of the ZR. Finally, in late vitellogenic oocytes a third layer, made of microfibrillar material, appeared. The immunohistochemical analyses revealed that the initial internalisation of hepatic zona radiata proteins (Zrp) in the swordfish oocyte starts before the uptake of vitellogenin (Vtg) and that it is associated with the low previtellogenic E2 plasma levels, while a significant E2 increase in plasma is associated with the beginning of Vtg uptake. This would appear to confirm the hypothesis that the differential and sequential induction of zonagenesis and vitellogenesis may reflect a general feature of teleost oogenesis.

Evidence of a high percentage of intersex in the Mediterranean swordfish (Xiphias gladius L.).

The first evidence of the presence of intersexuality in a wild population of Mediterranean swordfish (Xiphias gladius L.) is reported. Forty of 162 specimens (25%) macroscopically classified as males, showed the presence of female germ cells within the testes. In two specimens grouped previtellogenic oocytes were present; all the other specimens possessed single scattered previtellogenic oocytes. The presence of vitellogenin was demonstrated immunohistochemically in the liver of both intersex and normal males. These findings could be due to the exposure to oestrogen-mimicking substances.

Tackling hospital costs and delays.

Continually improving processes rather than undertaking projects with temporary problem-solving methods achieves total-quality-control services. Nine delay categories used in a 4-year audit determined reasons for service delivery delays on two cardiology units. As a result, patient length of stay and resource consumption decreased and quality of patient care was enhanced.

Morphometric and ultrastructural features of the mare oviduct epithelium during oestrus.

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.