Ultrastructural localization of butyrylcholinesterase in epithelial cells of rat intestine.

The present study reports the histochemical detection, at the ultrastructural level, of butyrylcholinesterase (BChE) in the epithelial cells of rat intestine. The enzyme activity was observed in the crypt cells as well as in the mature cells of the villi. Inside the enterocytes, BChE was seen in the reticulum cisternae, Golgi apparatus and lipid droplets. BChE was also detected in the Grünhagen spaces, associated with chylomicrons. In the intercellular spaces, at the level of the upper part of the cryptic epithelium, the enzyme was found from the base to the apex of the cells. The precise localization of BChE in the cellular structures noted suggests that its function could be related to lipid metabolism and/or cell renewal.

[The bovine zona pellucida: differences in macromolecular composition between oocytes, pre-treated with A-23187 or not, and embryos]. / La zone pellucide bovine: différences de composition macromoléculaire entre ovocytes, prétraités ou non à l'A 23187, et embryons.

The SDS-PAGE method was used to determine the composition of isolated bovine zona pellucida (ZP). This egg extracellular coat appears to be characterized by 3 major glycoproteins (ZP1, ZP2, ZP3), with apparent molecular weight (MW) of 80-70 kD, 66-63 kD and 60 kD, respectively, as revealed by 1-dimensional SDS-PAGE. After 2-dimensional SDS-PAGE, the zona pellucida electrophoretic pattern indicates a fourth glycoprotein, thus called ZP4. SDS-PAGE analysis of ZP isolated from oocytes and embryos after transit through the genital tract of A23187 pretreated oocytes allowed the description of modifications in glycoproteinic composition.

Utility of zona pellucida drilling in cases of severe semen alterations in man.

OBJECTIVE: To evaluate the utility of zona drilling for human severe sperm alterations in in vitro fertilization (IVF). DESIGN: A prospective randomized study was conducted according to random assignment of collected oocytes to a control or zona drilling procedure. SETTING: Reproductive Biology Laboratory, University Hospital of Nantes, France. PATIENTS: Twenty infertile couples with severe semen defects and previous IVF failure. INTERVENTIONS: Ultrasound-guided transvaginal follicular aspiration. RESULTS: Fertilization rates were compared to assess the advantages of zona drilling: 93% (80/86) drilled oocytes survived and 18.75% (15) were fertilized, whereas only 3% (3/100) control oocytes were fertilized. The polyspermy rate for fertilized drilled oocytes was high (10/15; 66.6%). The normal fertilization rate after zona drilling remained very low (5/86; 5.8%) and was not statistically different from that of control oocytes (3/100; 3%). One implantation occurred after replacement of five drilled embryos in three patients but resulted in early miscarriage. Replacement of the single control embryo led to a progressive pregnancy and normal male birth. CONCLUSIONS: This study indicates that zona drilling does not improve IVF results in cases of severe semen alterations.

[Evaluation of the probability of the occurrence of pregnancy during oligo-astheno-teratospermia]. / Evaluation de la probabilité de survenue d'une grossesse dans les oligo- asthéno- tératospermies.

Analysis of several sperm counts makes possible the positive diagnosis of oligo-astheno-teratospermia. Attempts to determine their etiology and consequently their treatment, remain negative, in most cases. The physician involved must then try to answer the couple's concern about the prognosis. The prognosis depends on the alterations seen on the sperm count parameters; but there is no threshold figure under which no pregnancy is observed (except for zero), concerning the count, the gradual mobility or the morphology of the spermatozoids. Motility studies, survival tests in cervical secretions, in the female genital apparatus and in culture media or fertilization in vitro tests, improve the prognosis. However, an answer can only be provided if the fertility takes into account the age of the female partner and a study of her fertility as well as the duration of the infertility. Management will consist of a choice between discontinuation of the treatment, test of fertilization in vitro and temporizing, including trials of male treatment and treatment of possibly associated female factors.

Investigations of endocrine cells in the gastrointestinal tract and pancreas during the metamorphosis of an anuran (Alytes obstetricans L.): histochemical detection of APUD cells.

Endocrine cells were detected at premetamorphosis, prometamorphosis, climax, and juvenile stages using an amine-inducing fluorescence technique with or without previous L-3,4-dihydroxyphenylalanine (L-DOPA) treatment. At premetamorphosis, serotonin cells exhibited yellow fluorescence in the gut primary epithelium of the L-DOPA untreated animals. In the treated animals, green fluorescent APUD cells could be seen in addition to the serotonin cells. In the pancreas, numerous clusters of fluorescent APUD cells were observed. At prometamorphosis the number of fluorescent cells increased in the intestinal primary epithelium and, close to the basal membrane, numerous small regenerative buds devoid of fluorescent cells appeared. In the pancreas of L-DOPA-treated animals, two types of APUD cells could be distinguished by their different fluorescence intensities. At the climax stage, the stomach developed and APUD cells were detectable in the gastric glandular buds. The degenerated primary intestinal epithelium was progressively removed in the intestinal lumen. At this stage, the regenerative buds of the secondary epithelium exhibited APUD cells. In the disorganized pancreas, the induced fluorescence decreased strongly. At the juvenile stage, cords of APUD cells displayed a cytoplasmic green fluorescence in the pancreas. In the stomach and intestine, serotonin and APUD cells were numerous.

Immunohistological detection of methionine-enkephalin-like and endorphin-like material in the digestive tract and in the nervous system of the mussel: Mytilus edulis L.

Using the indirect immunofluorescence technique, methionine-enkephaline-like, alpha- and beta-endorphin-like peptides were detected on whole body sections of Mytilus edulis L. Met-enkephalin-like immunoreactivity was localized in the epithelium of the digestive tract, in the hepatopancreas, and in the nervous system. The immunoreactive cell bodies were very abundant in the anterior gastric epithelium, but sparse in the terminal portion of the digestive tract. By their basal processes the immunoreactive cells were in contact with a plexus of immunoreactive cells and fibers located in the connective tissue underlying the digestive epithelium. In the principal hepatopancreatic ducts, isolated cells showing met-enkephalin-like immunoreactivity were detected between the epithelial cells and the basal lamina. A few immunoreactive cells and fibers were observed between the hepatopancreatic tubules. The three pairs of nervous ganglia contained in their cortical layer numerous met-enkephalin-like immunoreactive perikarya. Their central area possessed fluorescent immunoreactive bundles of fibers extending to the commissures, the connectives, and the nerves. Met-enkephalin-like immunoreactive fibers were detected between the smooth muscle cells. At the surface of these smooth muscle cells, immunopositive met-enkephalin-like tapered nervous endings were observed. The alpha- and beta-endorphin antisera produced a positive immunoreaction in some gastric epithelial cells, in some perikarya of the pedal ganglia, and in some nervous fibers. The endorphin-like structures were far less abundant than the met-enkephalin-like structures, but very close to them.

APUD-like cells in the primitive gut of a 27 day old human embryo.

A 27 +/- 1 days old human embryo arising from an ectopic pregnancy was incubated with 3,4, L-dihydroxyphenylalanine (L-DOPA) to investigate whether cells of the gut primordium have at this early stage of development, the ability to take up the amine precursors. We could observe that a few cells exhibiting greenish fluorescence were located in the epithelium wall of the fore gut. Most of them were situated in contact with the basal lamina and possessed a long apical process extending over towards the gut lumen. The uptake of amine precursor, the morphology and the location of these cells strongly suggest that they represent the gut endocrine precursor cells of the human embryo.

Detection of endocrine cells by immunofluorescence method in the gastroenteropancreatic system of the adult eel, glass-eel, and leptocephalic larva (Anguilla anguilla L.).

Five antisera against insulin (Ins), glucagon (Glu), somatostatin (SRIF), met-enkephalin (met-enk), and serotonin (5-HT) were used for immunofluorescence detection of endocrine cells in pancreas and gastrointestinal tract (GIT) of the European eel (Anguilla anguilla L.) at three stages of development (leptocephalic larva, glass-eel, and adult eel). Comparable distribution of endocrine cells was observed for adults and glass-eels. In their pancreatic islets, positive immunoreactions were obtained only for Ins, SRIF, and Glu; this later was also present in the pancreatic ducts. 5-HT cells were present throughout the GIT. SRIF cells were situated mostly in the stomach and less in the intestine. Met-enk cells were abundant in the pyloric cecum, but less frequent in the intestinal mucosa. Glu cells were present only in the intestine. No insulin-immunoreactive cells could be detected in the GIT. The pancreatic islets of leptocephalic larvae exhibited a strong reaction for SRIF, a weak reaction for Glu, and none at all for Ins, met-Enk, or 5-HT. The GIT of these larvae contained numerous met-enk cells, mainly in the foregut. In the fore- and midgut, cells exhibited a weak fluorescence after treatment with Glu antiserum. No positive immunoreactive cells were observed with 5-HT, SRIF, or Ins antisera.

Experimental analysis of the extensive pigmentation in the Silkie fowl embryo: evidence for an environmental regulatory process.

Heterospecific coelomic grafts, associated with the quail-chick marker system, showed that quail embryo melanoblasts exhibit the same invading behavior as Silkie fowl melanoblasts, when they came into contact with Silkie embryo organs. Thus the colonization or noncolonization of the organs of the Silkie fowl embryo by melanoblasts seems to depend on environmental cues.

[Fluorescent detection of APUD-type cells in the digestive tract of the rabbit fetus. Correlative study in electron microscopy]. / Détection en fluorescence de cellules de type APUD dans le tube digestif de foetus de lapin. Etude correlative en microscopie électronique.

Previous studies performed on different species have shown that these cells could be recognized by their morphologic and immuno-histological features. In early stages, these cells are able to take up and decarboxylate amine precursors. Therefore the aim of the present work was to determine if this uptake could be correlated with ultrastructural modifications. A processing technique allowing amine detection and correlative ultrastructural examination was used. Rabbit foetuses 13, 14, 17 and 21 day old were studied. The gastro-intestinal tracts of L-DOPA treated or untreated foetuses were removed in a glutaraldehyde-formaldehyde mixture and embedded in epoxy-resin. Semi-thin sections allowed to locate fluorescent cells in U.V light microscopy; adjacent thin sections were observed in electron microscopy. The first green fluorescent cells appeared in the 13 day old foetuses treated with L-DOPA. By this stage, these cells were very scarce and appeared poorly differentiated in electron microscopy. Between the 15th and the 18th day, the green fluorescent cells contained only small round granules. By the day 19, orange-yellow cells can be observed in L-DOPA treated and untreated foetuses. These cells possessed characteristic enterochromaffin granules. The green fluorescent cells of 21 day old foetuses, treated with L-DOPA, exhibited various fluorescence intensities correlated with the heterogeneity of the secretory granules. Some foetuses of each stage were treated with Falck's technique. This method gave similar results concerning the chronology of fluorescent cell detection.

[Cytochemical study of intestinal endocrine cells in rabbit fetuses during in vitro induced degeneration and differentiation (author's transl)]. / Etude cytochimique des cellules endocrines intestinales foetales au cours de leur dégénérescence et de leur différenciation in vitro.

Ontogenic differentiation of intestinal serotonin cells of 22 days old rabbit fetuses were studied in vitro. After 3 days of organotypic cultivation in solid medium, a great part of epithelial cells became necrotic and were eliminated into the lumen while a first flattened, then cuboïdal, then prismatic new growed epithelium in which FIF indicated some serotonin cells was present. Comparison of pictures obtained on the same slide by different methods was used in order to estimate correspondances between amine storage, argentaffin, argyrophilic and reductive properties. A new phase in serotonin cell differentiation was readily distinguished since young cells successively yeilded FIF, later argyrophilia, later argentaffinity. These datas, at variance with that occurs in vivo where FIF and argyrophilia appeared simultaneously, give a new criteria for differential mechanism studies, disprove the theory of an amine induced argyrophilia and enhance the hypothesis of a hyaloplasmic amine storage in very young cells.

[Coexistence of reactive and nonreactive endocrine cells in the fundal mucosa in the rabbit]. / Coexistence de cellules endocrines captantes et non captantes (non APUD) dans la muqueuse fundique chez le lapin

Whether the same endocrine like cells are reactive towards different staining methods has been examined in normal and L-Dopa treated rabbits for distinction of cell types. Correlation with electron microscopy is proposed. It seems likely that SM cells (poorly reactive with Sevier-Munger technic) include two cell types: cells having around medium size and dense granules; cells having small round dense granules (D1?). X cells may correspond to a well defined group at the electron or photonic microscopic level. Insertion of D cells (according to Solcia and coworkers nomenclature) is not satisfactory but true non Apud cells are described. The signification of Jt (dim yellow) cells remains unknown.

[Ultrastructural identification and immunocytochemical study of somatostatin cells in antral mucosa of the rabbit and the mouse (author's transl)]. / Identification ultrastructurale et étude immunocytochimique des cellules à somatostatine de la muqueuse antrale du lapin et de la souris

In normal and L-Dopa treated rabbits and mice, combined immunochemical methods, photonic histological methods for endocrine cells and ultrastructural methods were used to elucidate ultrastructure and properties of somatostatin cells of the antral mucosa. In normal rabbits, immunoreactive cells giving no fluorescence with Falck's technic, they corresponded neither to serotonin cells nor gastrin cells; they were unreactive with Fontana, Hellerström-Hellmann, Sevier-Munger and Mac Conaill methods but very slightly stained with Grimelius methods. In L-Dopa treated animals somatostatin cells gave formaldehyde induced fluorescence (they were included in GIC cells, thus in Apud group), exhibited a good reaction with Grimelius and Sevier-Munger methods. In order to carry out the alternate semi-thin/thin section procedure (semi-thin sections for immunofluorescence or immunoenzymatic detection and serial thin sections counter-stained for conventional ultrastructure studies), immunological treatment were performed on M.F.F.--glutaraldehyde fixed small fragments of mucosa before inclusion in Epon 812 or, after inclusion, on semi-thin sections. We succeeded in identifying ultrastructurally somatostatin cells. They displayed round or ovoïd shaped secretory granules, and three constant typical structures: numerous microfilaments--light and homogenous granules, often seeming like lipids---granules made up by coarsely filamentous cores surrounded by a large empty halo. Somatostatin cells seemed different of X cells because of their predominant localisation in the antral mucosa (in the rabbit X cells were predominantly in the fundus) and because of the lack of nuclear microfilaments; they also seemed ultrastructuraly different of D1 cells.

[Quantitative study of serotonin-containing cells in fetal rabbit duodenum]. / Etude quantitative des cellules à sérotonine duodénales chez la foetus de lapin

In order to attempt identification in vitro of parameters involved in the cellular differenciation, it was necessary to possess a standard material with identical content and distribution of serotonin cells; these two properties are investigated in this paper. Falck's technic is the only sensitive and specific method in demonstrating serotonin young cells devoid of argentaffinity. From 21, 22, 23 and 25 days old rabbit foetuses, 10 mm length duodenal pieces were treated according to Falck and sections serially cut at 8 microns. Some of these specimens were divided prior to Falck treatment in two pieces 3 mm away from the pylorus and in each piece 200 paraffin sections were cut at 8 microns beginning by adjacent ends and progressing in opposite direction. The number of EC sections in 400 whole transverse sections were determined for each foetus. Great variations were observed from one foetus to another, along the intestine, with weight and age. The Student-Fisher test applied to two consecutive pieces of the same duodenum gathering foetuses in groups of 5 animals did not show significative differences from a group to another. Thus, this material can be employed for comparisons under experimental conditions.