Lectin binding in the human umbilical cord from normally grown pregnancies and from pregnancies complicated by intra-uterine growth retardation with absent or reversed diastolic flow.

The content and the distribution of the glycoconjugate sugar residues were investigated in the human umbilical cord from normally grown pregnancies and from pregnancies complicated by intra-uterine growth retardation with absent or reversed flow in the umbilical artery. The gestational age range was 29-41 weeks. For this purpose, a battery of eight horseradish peroxidase conjugated lectins was used (ConA, WGA, PNA, SBA, DBA, LTA, UEA I, and OOA; see text). Our data showed that in both study groups ConA, WGA and PNA (after neuraminidase digestion) and LTA, UEA I, and OOA reacted with Wharton's jelly and vessels of the umbilical cord from the first weeks of gestation considered. SBA reactivity was observed only in the amniotic epithelial cells of the jelly and in the endothelial cells of the vessels. The reactivity with the most part of the lectins decreased or was lacking in the various components of the umbilical cord between 39 and 41 weeks of gestation in the normal group and between 36 and 37 weeks of gestation in the pathological one. Moreover, the intensity of reactivity with ConA, WGA and PNA (after neuraminidase digestion) and LTA, UEA I, and OOA in the normal group and with PNA (before and after neuraminidase digestion) and LTA and OOA in the pathological one was stronger in the peripheral portion of the jelly with respect to the adventitial one. The reactivity with ConA and UEA I was stronger in the vein than in the arteries in both study groups. Some hypotheses are reported on the role played by the sugar residues in the normal and pathological umbilical cords.

The glycoconjugate sugar residues of the sessile and motile cells in the thymus of normal and cyclosporin-A-treated rats: lectin histochemistry.

It is well known that cell surface glycoconjugates play a determinant role in cellular recognition, cell-to-cell adhesion and serve as receptor molecules. T-lymphocytes are in strict contact with the thymic epithelial cells, which control their process of maturation and proliferation. On the other hand the normal maturation of the epithelial cells is believed to be induced by T-lymphocytes. For these reasons we have studied the glycoconjugates saccharidic moieties of the sessile and motile cells in the thymus of normal male albino Wistar rats and their changes following cyclosporin-A treatment, using a battery of seven HRP-lectins. Cytochemical controls were performed for specificity of lectin-sugar reaction. Some sections were pre-treated with neuraminidase prior to staining with HRP-lectins. Our results have demonstrated, in the control rats, a large amount and a variety of terminal and subterminal oligosaccharides within and/or on the epithelial thymic cells and in macrophages. After cyclosporin-A treatment, among the thymic epithelial cells, the subcapsular, paraseptal and perivascular cells showed the loss of some sugar residues, which characterized the same cells in the intact thymus. Some hypotheses are reported on the role played by the glycoconjugate sugar residues in control and cyclosporin-A treated rats.

Lectin histochemistry for detection of the oligosaccharides in the testis of the chick embryo.

The oligosaccharidic content, distribution and changes of the glycoconjugates in the testis of the chick embryo, from the 8th day of incubation to hatching, were studied using a battery of seven HRP-conjugated lectins (DBA, SBA, PNA, ConA, WGA, LTA, and UEA I). Our findings showed that ConA and WGA appeared to characterize the spermatogonia during their differentiation, maturation, migration and meiosis. In the Sertoli cells, a change of localization of staining with ConA and WGA was revealed during the differentiation and maturation of these cells. The basal membrane was characterized by the reactivity with ConA and WGA from the early stages of incubation. ConA, WGA and PNA reacted with the endothelial cells of the testis for the whole period of incubation considered. Moreover, the interstitial cells, since their appearance, showed reactivity with ConA, WGA and PNA at the plasma membrane and the cytoplasm.

Lectin binding in the ovary of the chick embryo and newborn.

The content, distribution and changes of the glycoconjugates sugar residues in the ovaries of chick embryos, from the 8th day of incubation to hatching and in 1-day old chick, were investigated. For this purpose, a battery of seven HRP-conjugated lectins was used (DBA, SBA, PNA, ConA, WGA, LTA and UEA I). Our data showed that SBA was a marker of the most immature oogonia in the ovarian cortex and medulla. The reactivity with ConA appeared to characterize the cells immediately prior to as well as during the meiotic division, as demonstrated by the presence of alpha-D-mannose at the "Balbiani bodies" in the oogonia of the ovarian cortex. The detection of Con A and SBA reactivity corresponded to maturative stages of the early oogonia in different cortical zones of the chick ovary. Our data also revealed that PNA seemed to be a marker of the degenerating oogonia located in the ovarian medulla. Moreover, PNA binding was a characteristic finding in the endothelial cells of the vessels located in the compact portion of the medulla in the left ovary, from the 8th to the 21st day of incubation and after hatching; PNA reactivity was only seen from the 16th day onwards in the endothelial cells of the cortex. During the whole considered period of incubation and after hatching, reactivity with UEAI, LTA and DBA was never detected.

Glycoconjugate saccharidic moieties of the exocrine and endocrine pancreas in the chick embryo, newborn and adult.

A battery of horseradish peroxidase-conjugated lectins (PNA, SBA, DBA, WGA, ConA, LTA and UEAI) was used to study the distribution of glycoconjugate sugar residues in the exocrine and endocrine pancreas of chick embryos, 1-day-old chicks and adult animals. During the period of incubation considered here, the time of appearance and changes in the oligosaccharides were noted in the acinar cells, in the endocrine cells of the "light islets" and in the capillary endothelial cells. In the developing exocrine pancreas, the basolateral surface of the acinar cells reacted with PNA, WGA, LTA and ConA for the entire period of incubation while SBA, WGA, LTA and ConA reactivity was detected in the apical plasma membrane. Zymogen granule membrane reacted with PNA, WGA and LTA from the 12th day onward, while ConA reactivity was detectable from the 7th day onward. Owing to its early appearance, alpha-D-mannose, revealed by ConA lectin, might in some way contribute to the maturation of the zymogen granule. In the 1-day-old chick and in the adult the cellular surface reacted with WGA and ConA. The zymogen granule membrane reacted with PNA, WGA and ConA in the 1-day-old chick. In the adult, WGA and ConA reactivity was observed in the zymogen granule membrane, whereas at this site PNA reactivity was revealed only after neuraminidase digestion. The hypothesis formulated by others that granule membrane lectin reactivity is equal to the apical membrane reactivity, owing to exocytotic processes, is not consistent with our results in the chick embryos, the 1-day-old chick and the adults. The surface and the granules of the islet beta cells in different periods of incubation reacted with WGA, SBA and, after neuraminidase treatment, with PNA. The same reactivity was seen also in 1-day-old chick while in the adult the granules of the beta cells also reacted with ConA. These findings show the achievement of an almost adult-like state of glycosylation of the glycoconjugates in the beta cells at an early stages of development.

Changes in expression of the oligosaccharides in the human fetal skin.

Lectin histochemistry was used to investigate the glycoconjugate saccharidic moieties in the skin of human fetuses ranging in age from the 8th to the 13th week of gestation. For this purpose, seven HRP-conjugated lectins were employed (Con A, WGA, PNA, DBA, SBA, UEAI and LTA). The distribution and changes of the sugar residues of glycoconjugates at the level of the various layers of the fetal epidermis, as well of the underlying mesenchyme were studied. We have shown that UEAI reactivity is a characteristic finding of the cells of the intermediate layer at the 11th and 12th week of gestation. LTA reactivity was detected only at the free border of the peridermal cells from the 10th to the 13th week of gestation. We demonstrated the presence of PNA binding sites at both the basal and peridermal cells from the 8th week of gestation, whereas other authors have stated that the appearance of PNA reactivity coincides with the onset of fetal skin stratification (11th-12th week of gestation). Moreover, some typical features in lectin reactivity at the level of the mesenchymal cells and fibres have been pointed out.

Histochemical study of the epithelial mucins in the gizzard of the chick embryo.

Histochemical methods used to investigate the appearance and the changes of the epithelial mucins produced by the lining and glandular epithelium of the chick embryo gizzard from the 7th day of incubation to hatching. Carboxylated, sulfated and neutral mucins appeared simultaneously at the superficial layer of the lining epithelium (7th day), at cells located in the inner part of the lining epithelium surrounding intercellular spaces (9th day) and at the tubular glands (15th day). During the period of incubation considered, an increase and decrease in the production of the various mucins were observed at the lining epithelium. However, a gradual increase of mucous production was detected at the ephitelial cells surrounding intercellular spaces (from the 9th to the 17th day) and at the tubular glands (from the 15th to the 21st day). Moreover, three different overlapping steps of mucous release were observed during the development of the gizzard. The histochemical characteristics and the trend of production of the mucous material in the gizzard during the embryonic development were different when compared with those found in the proventriculus (Sgambati et al., 1995). These differences are probably related to the different function of the mucins in the two parts of the stomach.

Lectin histochemical study of glycoconjugates in the chick embryo proventriculus.

A lectin histochemical study was performed to investigate the glycoconjugate saccharidic moieties of the lining and glandular epithelium of the chick embryo proventriculus from day 7 to hatching. The sugar residues D-galactose-(beta 1-->3)-N-acetyl-D-galactosamine, N-acetyl-D-glucosamine, alpha/beta-N-acetyl-D-galactosamine and alpha-D-mannose were present at the lining epithelium of the proventriculus from the 7th day onwards; alpha-D-fucosyl residues were never detected. The same sugar residues were detected in the cells of the deep glands from the 7th day; alpha-L-fucose appeared later. The sugar residues appeared in the cells of the superficial glands at different times during the development of the proventriculus. In comparison to that of the deep glands, the secretion of the superficial glands was characterized, by the absence of N-acetylgalactosamine and alpha-D-mannose. From day 20 onwards, the composition of the mucus released by the two types of glands was identical from a qualitative point of view. The histological and histochemical results showed that the superficial glands are well defined morphological structures and not "an artefact of contraction", as claimed by other authors.

Histochemical study of human nasopharyngeal epithelium by horseradish peroxidase conjugated lectins.

This study proposes a simple method for assessing whether non-respiratory epithelia of the nasopharynx (i.e. cuboidal, squamous) represent a modified differentiation from the normal pseudo-stratified lining, or whether they possess common characteristics suggesting they are just in a different morphological stage. Peroxidase-labelled lectins were used, since they detect sugar residues of membrane glycoconjugates which play an important role in cell differentiation. Biopsies were taken from healthy controls and from patients affected by upper airway inflammation of varying degree and duration. While pseudostratified and cuboidal epithelia possess similar histochemical characteristics, squamous epithelium has a different behaviour; in particular it is the only type to show intense reactivity to Lotus tetragonolobus lectin (LTA). This would seem to indicate that squamous epithelium is in a different state of differentiation, possibly suggesting dysplasia. Lectins may be valuable both in morphological studies and in the clinical evaluation of airway epithelia.

Characterization of the glycoconjugate sugar residues in developing chick esophageal epithelium.

The development of the esophagus in the chick embryo is characterized by remarkable morphological changes especially at the level of the epithelium. Using horseradish peroxidase-conjugated lectins (DBA, PNA, SBA, WGA, ConA, LTA, UEAI) we have studied, at the level of the esophagus of chick embryos from the 8th to the 21st day of incubation and of 1- and 2-day-old chicken, the evolution of the saccharidic moieties of glycoconjugates, which precedes and/or is concomitant with the epithelial morphological transformations. We have found differences in content and cellular distribution of oligosaccharides during the histogenetic processes which characterize the lining and glandular epithelium. Before the appearance of cilia and mucus secretion at the bathyprismatic epithelial cells, the sugar residues D-galactose-(beta 1-->3)-N-acetyl-D-galactosamine, beta-N-acetyl-D-galactosamine and alpha-L-fucose were detected only at the luminal cell surface. These oligosaccharides were probably involved in giving rise to the polarization of the esophageal epithelial cells. The esophageal gland mucus was first characterized by the presence of alpha-L-fucose and afterwards also by the presence of D-galactose-(beta 1-->3)-N-acetyl-D-galactosamine, D-glucosamine and sialic acid.

Computerized morphometric analysis of the chick embryo esophagus.

By means of a computerized morphometric system and a semiautomatic digital system (videoplan II), the quantitative aspects of the organogenesis of the chick embryo esophagus were investigated from the 6th to the 15th day of incubation. The area of the total section, of the lumen, of the wall and its components (subserous stratum, muscle layer, lamina propria and epithelium) and the thickness of the epithelium and the muscle layer were measured. The mean area +/- S.E.M. of the values obtained was calculated. The percentage of shrinkage, due to the histological procedures, was calculated for each day. The mean values for each day of incubation were modified on the basis of the shrinkage percentage. Exponential curves and r coefficient were determined to evaluate the growing pattern of the mean area of the esophageal wall components as a function of age. The main results are the following: 1) From the 6th to the 10th day of incubation the areas of all the wall components show an increase. 2) From the 11th day onwards, the wall components undergo a different growth pattern; in fact, the area of the subserous stratum decreases from the 11th to the 13th day, the muscle layer and the lamina propria show a decrease on the 12th day, while the growing pattern of the epithelium area appears irregular from the 12th day onwards. 3) The lamina propria results to be the component of the wall that shows the highest growth rate. Moreover, the esophageal anlage shows different developmental behaviour compared with that of the other intestinal tracts previously examined (ileum, duodenum, colon, caeca) in the same chick embryo.

Identification of sugar residues in human fetal olfactory epithelium using lectin histochemistry.

Lectin-binding histochemistry was used to investigate the distribution and the changes of the glycoconjugate saccharidic moieties in the olfactory epithelium of human fetuses ranging from 8 to 12 weeks of gestation. It was found that the basal cells, the sustentacular cells and the olfactory neurons exhibit differences in oligosaccharide cellular content and distribution. Differences in lectin binding was also demonstrated at the dendrite, cell body and axon of the receptor cells. From the 11th week onwards, Ulex europaeus agglutinin I was found to be a marker of the olfactory neurons.

[Sugar residues of glycoconjugates in the olfactory epithelium of the human fetus: histochemical study using peroxidase-conjugated lectins]. / I residui zuccherini dei glicoconiugati nell'epitelio olfattivo di feti umani: studio istochimico con lectine coniugate a perossidasi.

The oligosaccharide content of glycoconjugates was studied at the olfactory epithelium of human fetuses ranging from 8 to 12 weeks of gestation by mean of peroxidase labelled lectins (DBA, PNA, WGA, SBA, ConA, LTA, UEA I). The main results demonstrated that: 1-The olfactory epithelium (olfactory cells, supporting cells and basal cells) was generally characterized by different amount of a-D-mannose, a-D-galactosamine, a-D-glucose, D-galactose-(beta 1-3)-N-acetyl-galactosamine, sialic acid and a-L-fucose. 2-At the 11th-12th week of gestation the largest amount of sugar residues was detected at the olfactory cells and at some basal cells. 3-At the 12th week of gestation, UEA I may be considered a specific marker of the olfactory cells in different stages of development.

[Study of the glycoconjugates in the mesonephros of the chick embryo using peroxidase-conjugated lectin]. / Studio dei glicoconiugati nel mesonefro di embrione di pollo mediante l'uso di lectine coniugate a perossidasi.

The distribution of the sugar residues in glycoconjugates along the mesonephric nephron of chick embryo from the 4th day of incubation till hatching has been investigated, by means of six different horseradish peroxidase-labelled lectins. ConA and WGA showed an ubiquitous presence of alpha-D-mannose and N-acetyl-D-glucosamine along the nephrons. SBA was found to be a specific marker of the proximal tubule. PNA and LTA reacted only for a short time at some sites during the considered period of incubation. Sialic acid was detected at the glomerulus in the podocytes, capillary wall and, with a lesser extent, in the mesangial cells. Significant changes of the glycosylation pattern of the glycoconjugates during the period of mesonephric activity and the period of involution were seen.

[Distribution of saccharide residues in glycoconjugates of the kidney in Gallus domesticus using peroxidase-conjugated lectins]. / Studio della distribuzione dei residui zuccherini dei glicoconiugati nel rene di Gallus domesticus mediante l'uso di lectine coniugate a perossidasi.

A battery of seven different horseradish-peroxidase labelled lectins (DBA, PNA, SBA, UEA I, WGA, ConA, LTA) was used to study the distribution of sugar residues in the glycoconjugates along the nephron and the collecting duct of the kidney of Gallus domesticus. As far as the glomerular components are concerned, we have demonstrated that the podocytes and, with a lesser extent, the mesangial cells are characterised by the presence of D-mannose, D-galactose-(beta 1- greater than 3)-N-acetyl-D-galactosamine and sialic acid. The glomerular capillary wall shows the presence of the disaccharide D-galactose-(beta 1- greater than 3)-N-acetyl-D-galactosamine and sialic acid. With regards to the tubules, the proximal tubule, the descending limb of the loop of Henle, the connecting tubule and the collecting one, are characterised by N-acetyl-D-galactosamine, (1- greater than 6)-alpha-L-fucose, D-mannose, N-acetyl-D-galactosamine and D-galactose-(beta 1- greater than 3)-N-acetyl-D-glucosamine. The cells of the connecting and collecting ducts show the presence of intracellular sialic acid, found also as component of the mucous secretion. The ascending limb of the loop of Henle and the distal tubule contain only three saccharidic residues, i.e. (1- greater than 6)-alpha-L-fucose, D-mannose and N-acetyl-D-glucosamine. Lectin histochemistry was also useful to define the saccharidic components of the mucus, which is normally present within the connecting and collecting ducts of the kidney of the birds. The cellular variability of the connecting and the collecting ducts is similar to that found in the kidney of some mammals. Such a variability seems to suggest a possible cell specialization along a single kidney tubule.

[The development of the tongue in the chick embryo: observation under a scanning electron microscope]. / Lo sviluppo della lingua nell'embrione di pollo: osservazioni al microscopio elettronico a scansione.

The morphological features of the chick embryo tongue from the 8th day of incubation till hatching and during the early post incubation period have been investigated by means of Scanning Electron Microscope. At the SEM, it is possible to observe that already at the 8th day of incubation, the body and the root are separated by a low smooth-surfaced ridge. In the following days this ridge develops, giving rise to the so-called lingual spines, whose significance is still uncertain. As concerns the evolutive pattern of the superficial layer of the epithelium, in the first days of the considered incubation period the cells appear dome-shaped and have microvilli on their apical surface; afterwards they tend to become more flattened, and the microvilli are replaced by a thick net of microplicae. In the last days of incubation and after hatching desquamative phenomena become evident. The above described evolutive process can be regarded as a common feature of the whole dorsal lingual surface; only few regional differences are to be noted, such as the earlier development of the microplicae on the apex and borders of the tongue. In particular, the microplicae observed at the apex of the tongue show a typical aspect and arrangement; they run regularly parallel to each other. On the lingual dorsal surface taste bud-like structures have never been observed.

The development of the chick embryo gallbladder studied by scanning electron microscope.

The differentiation of the gallbladder mucous membrane in chick embryos has been studied from the 11th day of incubation until hatching (stages 37-46 of Hamburger an Hamilton 1951) with scanning electron microscope. The surface of the mucous membrane, at the 11th day of incubation, appears regularly smooth. From the 12th day onwards, longitudinal folds appear on the surface of the mucous membrane, becoming more and more numerous and complicated in the following days. During the last days of incubation, branching and anastomosing folds are present. At the 11th day of incubation, the epithelial cells lining the gallbladder surface have flattened apices, with short microvilli. From the 12th day onwards, the epithelial cells show dome shaped apices, with long numerous microvilli. From the 15th day of incubation onwards, some particular secretory cells can be detected. The appearance of these cells is probably related to the water-absorbing function of the gallbladder which in the last days of incubation completes its functional development, because of precise digestive requirements of the chick embryo.

[Various aspects of organogenesis of the colon in Gallus domesticus: morphologic observation]. / Alcuni aspetti dell'organogenesi del colon nel Gallus domesticus: osservazioni morfologiche.

The authors have investigated the morphological aspects of the wall components of the developing colon in the chick embryo (Gallus domesticus) from the 7th to th 15th day of incubation. Particular attention has been given to the lumen recanalization, phenomenon which occurs also in other animal species. The most significant results can be summarized as follows: 1) the lumen is recanalized at the 7th day only at the proximal part of the colon (Fig. 1, Tav. 1), while at the distal tract it is still completely filled by an epithelial plug (Fig. 2, Tav. 1). Therefore the recanalization of the lumen takes place cranio-caudad. 2) At the 8th day the process of recanalization of the lumen shows, in the distal part of the colon, well defined modalities. Radially oriented intraepithelial spaces within the epithelium filling the lumen join other semilunar intercellular spaces, which are placed near the central part of the occluded lumen (Fig. 3). By the junction of a couple of radially oriented spaces with one semilunar space, an U-shaped intercellular space derives, which delimits an incoming epithelial fold (Tav. 3). Such a phenomenon is continued also during the 9th and 10th day of incubation (Fig. 6, Tav. 2). 3) At the 11th day the colonic lumen is completely open and, in its distal part, the appearance of the primordial previllous ridges can be observed (Fig. 7). In the proximal tract the previllous ridges develop one day later (Fig. 8). 4) At the 13th day, in the distal part of the colon, the first appearance of crypts occurs (Fig. 10). So, while the process of recanalization of the lumen is cranio-caudad, the formation of previllous ridges and crypts proceeds caudo-cranially. 5) From the 11th day onwards the lamina propria is actively involved in the process of formation of the previllous ridges. Only at the 14th day, in the distal part of the colon anlage, the appearance of the muscularis mucosae is observed (Fig. 11). 6) The muscle layer and the subserous stratum do not show appreciable morphological changes in the course the considered period of incubation.

Histochemical analysis of goblet cell mucins in the chick embryo colon.

The histochemical characteristics of colonic epithelial mucins were investigated in the chick embryo. At the 14th day of incubation it was possible to demonstrate the presence of glycogen. At the 15th day a few epithelial cells showed the presence of neutral and sialylated mucins. On the 16th day, also sulfated secretory material was detectable together with neutral and sialylated mucins in cells with the typical shape of goblet cells. From the 17th day to the 20th day of incubation the two types of acid mucins appeared in some cells to be placed in distinct zones of the supranuclear cytoplasm. At the 21st day, neutral, sialylated and sulfated mucins were all present in the majority of goblet cells, which were found mainly in the epithelium lining the crypts.