bluuebunny/crossref_metadata_2025 huggingface dataset:
In vitro growth characteristics of rat mesothelioma cells in culture
Authors: Jaurand M. C. , Renier A. , Van der Meeren A. , Magne L. , Pinchon M. C. , Bignon J.
Published: February 2012

The study reports morphological growth characteristics and chromosome analysis of neoplastic rat pleural mesothelial cells (RPMC) isolated from a mesothelioma‐bearing rat. The pleural mesothelioma was induced by intrapleural injection of chrysotile fibers. Neoplastic RPMC were cultured by the standard methods used for normal RPMC. Neoplastic RPMC cultures had a population doubling time of 19 hr versus 30 hr for the normal cells. Plating efficiency in liquid medium was almost 100%. Cultures of neoplastic RPMC were anchorage‐independent since 70% of the seeded cells formed colonies after one week; after the second week, colony size was enhanced but colony recovery was not. The serum dependence of neoplastic cells was less than that of the normal cells. 79 out of 100 metaphase cells analyzed had 41 to 43 chromosomes, and the modal number was 42 (38%). A large metacentric chromosome was observed in 77 of the 100 neoplastic metaphase cells analyzed, but not in any normal metaphase cells. In nude mice, the neoplastic RPMC were tumorigenic.
Inositol trisphosphate‐induced Ca2+ release from rat parotid subcellular fractions
Authors: Bonis D. , Giraud F. , Rossignol B.
Published: February 2012

Inositol trisphosphate (IP3) has been shown to induce a release of sequestered calcium in a large (7,000‐48,000 g) microsomal fraction isolated from rat parotid gland. This effect was also observed after the calcium uptake had been stopped by EGTA and thus was not the result of an inhibition of the active calcium transport. IP3‐induced Ca2+ release was also demonstrated in a more purified microsomal fraction (16,000‐48,000 g) apparently free of mitochondria, and in a fraction enriched in rough endoplasmic reticulum markers. These results support the hypothesis that IP3 may be a second messenger for intracellular calcium mobilization during stimulation of the parotid gland by calcium mobilising agonists.
Colchicine‐induced tubular, vesicular and cisternal organelle aggregates in absorptive cells of the small intestine of the rat. II.—Endocytosis studies
Authors: Ellinger A. , Pavelka M.
Published: February 2012

Administration of the antimicrotubular agent colchicine to adult rats (0.5 mg/100 g of body weight for 6 hr) induces formation of extended aggregates of tubular, vesicular, and cisternal organelles in the absorptive cells of the small intestine. The phosphatase reaction pattern (thiamine pyrophosphatase, acid phosphatase, acid trimetaphosphatase) suggests that the majority of them belongs to the lysosomal system (Ellinger and Pavelka, 1984). The present study extends these findings and examines the uptake and fate of intravenously injected horseradish peroxidase (HRP) at the basal and lateral cell surfaces and of intraluminally applied HRP at the apical cell surface. HRP, applied to control animals and animals pretreated with colchicine, was internalized at both apical and basolateral cell surfaces of the absorptive cells, and delivered into endosome‐like vesicles, multivesiculated bodies (mvbs), dense bodies (dbs), and in several instances into Golgi cisternae. Following intraluminal application, evidence was obtained for the transport of HRP across the cell; in contrast, intravenously applied HRP was never detected at the apical cell surface. Colchicine pretreatment did not stop the uptake of HRP, which was rapidly sequestered to the clustered tubules, vesicles, and cisternae, as well as to the mvbs and dbs. After longer intervals, the portion of HRP‐reactive tubules, vesicles, and cisternae within the clusters increased: 60 min after HRP‐administration all of them contained HRP‐activity. These results indicate that the colchicine‐induced clustered organelles are recipients of endocytic materials internalized at the apical as well as at the basolateral cell surface.
Immunological detection of actin in isolated cilia from quail oviduct
Authors: Chailley B. , Bork K. , Gounon P. , Sandoz D.
Published: February 2012

Cilia from quail oviduct were isolated with their membrane. The ultrastructural study revealed a good preservation of cilia in the purified fraction. Electrophoresis on SDS‐PAGE showed a reproducible pattern of ciliary proteins, the major bands being those of tubulins 57 kDa and dyneins above 250 kDa. Among the minor bands, an immunological study was focused on a 43 kDa molecular mass protein, using monospecific antibodies against actin. Presence of actin was then detected by immunoblotting of isolated cilia fractions as well as of demembranated cilia, suggesting that actin is associated with the axoneme. The presence of actin in the cilia was confirmed by immunofluorescence. The cilia were found stained only on the proximal part, suggesting an heterogeneous distribution of actin within the axonemal length.
Long‐term culture of fetal rat hepatocytes in media supplemented with fetal calf‐serum Ultroser SF or Ultroser G
Authors: Yeoh G. , Douglas A. , Brighton V.
Published: February 2012

Fetal hepatocytes cultured in medium supplemented with fetal calf serum (FCS) or Ultroser SF do not maintain production of albumin or transferrin beyond one week of culture. When dexamethasone (10(‐7) M) is present, secretion of albumin and transferrin can be extended to two weeks, however, levels are extremely low. By three weeks, neither plasma protein can be detected in the culture medium in either conditions of culture. In contrast, hepatocytes maintained in medium supplemented with Ultroser G continue to produce albumin and transferrin at high levels for the entire three week period of this study. The morphology of the cultures are different. In FCS and Ultroser SF supplemented medium there are many more fibroblast and epithelial‐like cells and relatively fewer cells which are distinctly hepatocytes when compared with Ultroser G supplemented medium. The level of tyrosine aminotransferase, which is a dexamethasone inducible enzyme, is found to be much higher in Ultroser G cultures, with no further increase demonstrable by addition of dexamethasone. In contrast, dexamethasone induces the enzyme by about eight‐fold in cultures maintained in FCS supplemented medium. Therefore it appears that Ultroser G already contains sufficient steroid activity to maximize the level of tyrosine aminotransferase. A comparison between Ultroser C and SF (steroid‐free) suggests that the mixture of steroid and steroid derivatives in the G formulation must be important in the maintenance of differentiated functions of hepatocytes in culture. However, supplementation of FCS cultures with dexamethasone, which is known to be present in Ultroser G, does not allow hepatocytes to retain their differentiated functions over an extended period. Therefore it is concluded that other components besides dexamethasone must be important.
Resistance of Candida parapsilosis to drugs
Authors: Camougrand N. , Velours G. , Guerin M.
Published: February 2012

Several strains of Candida parapsilosis, isolated independently in our laboratory, had their resistance compared to a series of inhibitors which act either at the level of mitochondrial ribosomes (chloramphenicol, erythromycin, paromomycin) or at the level of mitochondrial respiration and oxidative phosphorylation (oligomycin, antimycin A, diuron, carbonylcyanide m‐chlorophenylhydrazone). Cells were grown on glycerol media supplemented with one of these inhibitors, and it was demonstrated that the resistance of these yeasts to a large spectrum of antibiotics was due to several features: a resistance to oligomycin was found at the permeation level; the resistance to the other drugs was correlated to the relative insensitivity of cytochrome biosynthesis to the drugs; the cells developed, at the same time, two types of alternative pathways: the one branched at the ubiquinone level which drove electrons from Krebs cycle substrates to oxygen, and the other, antimycin A‐insensitive but inhibited by amytal, salicylhydroxamic acid and high cyanide concentrations. This secondary mitochondrial pathway, driving reducing equivalents from cytoplasmic NADH to cytochrome c and then to cytochrome aa3 or to alternate oxidase, allowed the growth of Candida parapsilosis on a non fermentescible medium, supplemented with these drugs.
Flow DNA analysis in human lung cancer: potentiality and limitations due to sampling methods
Authors: Chassevent A. , Berruchon J. , Bertrand G. , Chretien M. F. , Tuchais C. , George P. , Larra F. , Rebel A. , Oury M.
Published: February 2012

Flow DNA analysis was performed on samples from 71 surgically removed lung cancers and from 145 patients undergoing bronchoscopy. Abnormal DNA stem lines, characterized by their DNA index (DI), were frequently observed in operated lung carcinomas (87%). Two or three abnormal DNA stem lines were discovered simultaneously in 10% of the samples. The mean DI of all abnormal tumor stem lines was lowest for rare tumor cell types and highest for adenocarcinomas. Intermediate mean DI values were found for epidermoid and small cell carcinomas, which were among the most proliferative tumors. The high rate of false negative results suggests poor diagnostic reliability of flow DNA analysis on bronchoscopic samples. However, the method appears to provide a promising objective tool capable of evaluating tumor behavior and prognosis.
Applications of flow cytometry to the structural and functional study of immune responsive cells
Authors: Noan G. Merdrignac‐Le , Genetet B.
Published: February 2012

Although flow cytometry is not yet widely used for diagnostic purposes, it provides a powerful tool for investigating structural and functional properties of immune‐responsive cells. By far, the largest application of immunofluorescence is in identifying specific cell surface features thereby allowing the discrimination of lymphocyte subpopulations (phenotyping normal and malignant cells), for analysing complex receptors on neutrophils and monocytes, for detecting the appearance of new surface antigens induced by cell differentiation or activation. Besides this, the technique sets objective standards for appreciating cell activation (changes in RNA content, increased phagocytosis, DNA synthesis, changes in membrane fluidity, pH gradient changes). Computer processing of measurements allows rapid accurate studies of several properties simultaneously.
Flow cytometry, a very useful technique for the characterization of intestinal membrane vesicles
Authors: Gorvel J. P. , Mishal Z.
Published: February 2012

The plasma membrane of enterocytes comprises two structurally and functionally distinct domains. These are the apical brush border, containing digestive hydrolases and glycocalyx, and the basolateral domain, characterized by other specific markers. Using a fast and easy subcellular fractionation, we purified four membrane vesicle fractions from rabbit small intestinal mucosa: brush border, basolateral, rough endoplasmic reticulum and Golgi + smooth endoplasmic reticulum. Using flow cytometry, the fluorescence polarization of diphenylhexatriene was determined in brush border and in basolateral + Golgi + smooth endoplasmic reticulum membrane fractions in order to investigate changes in the membrane fluidity of both fractions and to compare the results obtained with those of spectroscopic techniques. Moreover, it was possible with flow cytometry to detect and quantify basolateral and brush border markers by using polyclonal and monoclonal antibodies. The advantages of flow cytometry in the detection of brush border membrane markers found in small amounts in the basolateral domain are discussed. Finally, flow cytometry holds great promise for the analysis and sorting of subcellular fractions.
Variation of cellular mitochondrial activity in culture: analysis by flow cytometry
Authors: Ratinaud M. H. , Julien R.
Published: February 2012

Cytometric analysis of various cultured cells using fluorescent probes to stain mitochondria, in combination with other methods, has shown that mitochondrial activity is an essential part of cell cycle completion. Among the existing fluorochromes, Rhodamine 123 is most often used for analyses of growth and cellular differentiation, and the action of various compounds. These studies permit a better understanding of the role of the mitochondria in situ and especially of the interactions that occur between the cytoplasm and the nucleus. However, the development of this type of study is limited by the small number of specific fluorochromes available.
Endocytosis in guinea pig seminal vesicle epithelial cells cultivated in chemically defined medium
Authors: Mata L. , Petersen O. W. , Van Deuers B.
Published: February 2012

We have developed a chemically defined monolayer culture system for guinea pig seminal vesicle epithelial cells (SVEP). The cells appeared as a polarized monolayer with apical microvilli, tight junctions and desmosome‐like junctions, and often dilated intercellular spaces. SVEP expressed epithelial‐specific cytokeratins as detected immunocytochemically. Growth was obtained during the first week of culture. In this period, the cells were exposed to unconjugated horseradish peroxidase (HRP), a ricin‐peroxidase conjugate (Ri‐HRP), or cationized ferritin (CF). HRP was endocytosed without binding to the SVEP surface (fluid‐phase endocytosis) and was found mainly in multivesicular endosomes and lysosomes. Ri‐HRP and CF, however, were endocytosed following binding to the cell surface. Initially these markers were present in multivesicular endosomes, but later also in smaller tubular and vesicular endosomes, some Golgi‐associated elements (but not Golgi stacks), and lysosomes. We conclude that our SVEP culture system may be useful in further studies, on e.g. hormonal regulation of endocytosis and other processes of importance for SVEP maintenance and modulation of the seminal fluid in vivo.
Long‐term treatment of tumor‐operated mice with high doses of alpha‐difluoromethylornithine
Authors: de Lustig E. S. , Klein S. , Fuchs A. , Algranati I. D.
Published: February 2012

Mice inoculated subcutaneously with a mammary adenocarcinoma M3cell suspension and submitted 10 to 15 days later to the surgical removal of primary tumors have been chosen in our laboratory as the experimental model to study the effect of several drugs on metastasis spreading. A 90‐day treatment of tumor‐operated mice with high doses of alpha‐difluoromethylornithine showed, in addition to a marked decrease of lung metastases, several side effects including a loss of body weight, a delay of surgical wound healing and a reduced number of megakaryocytes in bone marrow. The measurement of ceruloplasmin in serum has indicated that treated animals contained increased levels of this oxidase activity in spite of the polyamine depletion caused by alpha‐difluoromethylornithine. These results rule out a direct correlation between ceruloplasmin activity and polyamine concentration.
Zymograms of Kurloff cell acid phosphatases. Thin layer isoelectric focusing and native polyacrylamide 4–15% gradient gel electrophoresis
Authors: Landemore G. , Buat M. L. , Izard J.
Published: February 2012

Following previous light and ultrastructural cytochemical reports, this paper presents the first zymograms of Kurloff cell acid phosphatases extracted from highly purified cell suspensions. Using isoelectric focusing, up to 20 isoenzymes were observed both with hexazotised pararosanilin or fast garnet GBC as coupler and naphthol ASB1 phosphate as substrate. Most were tartrate‐labile. After Clostridium‐derived neuraminidase digestion, the highly stained bands observed at pH 4‐5 disappeared and, simultaneously, a few alkaline bands were enhanced. Two main bands of Mr 190,000 and 500,000 were characterized by their acid phosphatase activity after electrophoresis on a 4‐15% gradient native polyacrylamide gel.
Ultrastructural study of epithelial cells and basement membrane. Differentiation of the rat epididymis after prenatal irradiation
Authors: Delongeas J. L. , Leheup B. P. , Gelly J. L. , Grignon G.
Published: February 2012

The effects of prenatal irradiation on the testis are well documented, but less is known about its effects on epididymal differentiation. Pregnant rats were irradiated on the 18th day of gestation. The increase in microfilaments and lipid inclusions in the epithelial cells, in favor of a direct radiation effect, is maximal at birth and disappears thereafter. Narrow cells and clear cells show a normal differentiation pattern. On the other hand, the principal cell maturation is largely altered. The synthesis capacities are decreased based on a reduction in the size of the Golgi apparatus and the smooth endoplasmic reticulum. The aspects of invaginations of the apical plasmalemma, coated vesicles and multivesicular bodies are not modified, suggesting normal absorption functions. The epithelial basement membranes become irregular and thicker than normal, enfolding the basal part of the epithelial cells. The basement membrane proteoglycans, demonstrated by the cationic marker polyethyleneimine, are irregularly distributed in contrast to the normal pattern. These modifications of the principal cells and the basement membrane are more prominent in the proximal epididymis. This suggests a differential maturation dependence of the epithelial cells on the luminal factors, normally secreted by the testis, and likely disturbed by prenatal irradiation which leads to germ cell degeneration, and then to a new balance in the seminiferous epithelium.
Immunolocalization of plakoglobin in endothelial junctions: identification as a special type of Zonulae adhaerentes
Authors: Franke W. W. , Kapprell H. P. , Cowin P.
Published: February 2012

We have characterized the junctions between endothelial cells of diverse blood vessels at the light and electron microscopic level using various antibodies to plakoglobin (polypeptide Mr 83,000) and vinculin. Endothelial cells from fenestrated and non‐fenestrated capillaries to large arteries are connected to each other by extended junctions that are coated on their cytoplasmic face by plaques of loosely matted filamentous material that form a continuous belt system along the cell circumference. These plaques are devoid of desmosome‐specific proteins such as desmoplakin(s) and desmoglein, but contain plakoglobin. Immunofluorescence microscopic reactions of these regions with vinculin antibodies have also been observed, although they are much weaker and less consistent. This composition, together with their association with actin microfilaments, classifies this extended plaque system as Zonulae adhaerentes. Our results also show that such endothelia may be distinguished from truly epithelial cells by the absence of desmosomes and intermediate filaments of the cytokeratin type. The relationship of the various kinds of adhering junctions and the physiological importance of these junctions are discussed.
Internalization of atrial natriuretic factor by AtT‐20 corticotropin‐secreting cells
Authors: Morel G. , Belles‐Isles M. , Heisler S.
Published: February 2012

Because extended exposure of AtT‐20 corticotropin‐secreting cells to atrial natriuretic factor (ANF) results in a desensitization of ANF‐induced cGMP synthesis, we sought to establish whether pretreatment of AtT‐20 cells with the atrial peptide also led to an internalization process. In fact, by coupling an ultrastructural approach to cryoultramicrotomy, ANF‐immunoreactivity was detected at both the plasma membrane level and at intracellular sites in AtT‐20 cells. Internalization was observed within 5 min at which time labelling was observed in the plasma membrane level, in vacuole‐like structures in close proximity to the plasma membrane, in cytoplasmic matrix and sometimes in mitochondria. After 30 min exposure Golgi apparatus, mitochondria and nuclear euchromatin were also labelled. Following 1‐4 hr, labelling in other cell compartments, e.g. lysosomal, was increased, while it was reduced in plasma membranes and vacuole‐like structures. Secretory granules and endoplasmic reticulum were not labelled throughout the time course. Extraction of a intracellular [125I] ANF from AtT‐20 cells following 4 hr incubation suggested that about 90% of the peptide was intact. The data suggest that internalization of ANF may serve to terminate the biological response associated with ANF receptor activation; subcellular distribution of internalized, intact ANF suggests that the peptide may have other, as yet unidentified, intracellular actions.
Monoclonal antibody against a nuclear matrix antigen in proliferating human cells
Authors: Philipova R. N. , Zhelev N. Z. , Todorov I. T. , Hadjiolov A. A.
Published: February 2012

A monoclonal antibody of the IgG2a subclass was isolated from the supernate of a hybridoma line obtained with splenocytes from a mouse immunized with a crude nucleolar fraction of human Namalwa cells. This antibody identifies a single nuclear polypeptide antigen characterized by: (a) presence in proliferating human cell lines and phytohemagglutinin‐stimulated lymphocytes, but absence in resting lymphocytes; (b) appearance in stimulated lymphocytes in parallel with the onset of DNA synthesis; (c) a speckled distribution in the nucleoplasm; (d) tight association with nuclear matrix structures identified by both biochemical and in situ extraction and enzyme treatment procedures; (e) mol wt of 125 kDa and pI 6.5 as determined by immunoprecipitation or immunoblotting of nuclear or nuclear matrix proteins fractionated by gel electrophoresis. The above characteristics identify the p125/6.5 nuclear matrix protein recognized by the isolated monoclonal antibody as belonging to the class of proliferating cell nuclear antigens.
Segregation of mitochondria in the cytoplasm of Xenopus vitellogenic oocytes
Authors: Mignotte F. , Tourte M. , Mounolou J. C.
Published: February 2012

In actively growing vitellogenic oocytes of Xenopus laevis mitochondria segregate into 2 populations. One stays around the nucleus, actively replicates mitochondrial DNA (mtDNA), and builds up most of the stock of the mitochondria in the full‐grown oocyte. The other moves toward the vegetal pole and stops replicating mtDNA early in vitellogenesis. Organelles of this population are components of the germ plasm of the cell.
The calcium antagonist nisoldipine and the calmodulin antagonist W‐7 synergistically inhibit initiation of DNA synthesis in cultured arterial smooth muscle cells
Authors: Thyberg J. , Palmberg L.
Published: February 2012

Cultured arterial smooth muscle cells go through a transition from a contractile to a synthetic phenotype. Morphologically, the transition includes a reduction in size of the myofilament bundles and the formation of an extensive rough endoplasmic reticulum and a large Golgi complex. Functionally, it leads to loss of contractility, onset of cellular proliferation, and secretion of extracellular matrix components. This change in differentiated characteristics under in vitro conditions has attracted attention because of its resemblance to the modification of the smooth muscle cells that occurs in vivo during atherogenesis. Here, transmission electron microscopy and [3H]‐thymidine autoradiography were used to study the role of calcium ions in the control of phenotypic properties and growth of cultivated rat aortic smooth muscle cells. The calcium antagonist nisoldipine was found to lack distinct effect on the structural reorganization of the cells, but showed a moderate prohibitory effect on the start of DNA synthesis early in primary culture. In growth‐arrested secondary cultures, nisoldipine inhibited induction of DNA synthesis by serum or platelet‐derived growth factor (PDGF). The agent's effect was inversely related to the concentration of calcium ions in the extracellular medium and was partially counteracted by the calcium agonist BAY K 8644. In contrast, W‐7, an antagonist of the calcium‐binding protein calmodulin, potentiated the effect of nisoldipine and, at higher concentrations, inhibited induction of DNA synthesis in itself. The results suggest that the mitogenic stimulation of arterial smooth muscle cells involves a flux of calcium ions through the plasma membrane and requires participation of calmodulin.
Morphometric study of the neuromuscular synapses in the adult rat with special reference to the remodelling concept
Authors: Tomas i Ferre J. , Mayayo E. , Fenoll i Brunet R.
Published: February 2012

There is increasing morphologic evidence that neuromuscular synapses are not rigid structures in the mature muscles of adult animals. On the contrary, they may be submitted to a continuous process of remodelling. In silver‐impregnated sternocleidomastoid muscles of the young adult rat, we measured synaptic parameters such as nerve terminal length, the number of branching points of terminal arborization, and muscle fiber diameter, and used a morphometric approach to explore specific questions concerning neuromuscular remodelling. Quantitative data indicate that: (a) The complexity and maturation of the nerve endings in this muscle are very variable and the increase in branching points is not paralleled by an increase in terminal length; (b) Muscle fiber diameter is related only marginally to presynaptic parameters; (c) Accessory ending formation occurs when the original ending does not reach the mean size of endings in singly innervated areas; (d) The complexity of individual endings at dually innervated junctions is smaller than the mean development of singly innervated synapses, indicating the existence of some mutual inhibitory influence between closely spaced endings. Morphometric results suggest a continuous process of synaptic formation in this adult muscle.
New in vitro procedures for experimental studies on the development of 11‐day mouse embryo forelimb buds
Authors: Desbiens X. , Revillion‐Carette F. , Meunier L. , Bart A.
Published: February 2012

A new method has been developed for culturing 11‐day mouse forelimb buds in vitro. In cultures performed with conventional procedures, skeletal pieces frequently appeared distorted and reduced in size. Moreover, forelimb buds explanted from embryos younger than a stage corresponding to 50 pairs of somites developed narrow hand plates devoid of radiated autopods. By contrast, in the new procedure using media supplemented with fetal calf serum and growth factors and enhancing distal feeding with carrier implants of catgut, enlarged pads were obtained that exhibited at least 4 digital rays in buds explanted from embryos with 40‐44 pairs of somites. Compared with conventional procedures, the mean value of DNA content per limb bud was twice as great with use of our improved method. The ability of limb bud cells to proliferate and differentiate when cultured either in classical or in modified conditions, and the importance of the technical procedures, are discussed in the new prospect of in vitro developmental studies.
Apical membrane marker is expressed early in colonic epithelial cells
Authors: Le Bivic A. , Hirn M. , Reggio H.
Published: February 2012

We have identified and characterized a membrane glycoprotein located at the apical plasma membrane of adult human colon epithelial cells, by the use of the monoclonal antibody technique in combination with immunocytochemical and biochemical methods. Analysis of membranes extracted with Triton X‐114 and treated with specific hydrolases indicated that the antigen was an integral membrane glycoprotein. In the colon, the antigen was expressed in differentiated cells and along the entire crypt. It was also expressed at the apical membrane of the crypt cells of the distal ileum. It was not found in the proximal ileum, jejunum, or duodenum. In contrast, the antigen was found in all segments of the intestine of a 24‐week‐old embryo. Furthermore, the antigen had different apparent molecular weights in the adult ileum (200 kDa), adult colon (200 kDa and 301 kDa), and embryo (170 kDa). Therefore, this antigen should prove to be a useful marker to study the appearance of epithelial cell polarity during embryogenesis.
Recognition of xenogeneic erythrocytes: the GalNAc/Gal‐particle receptor of rat liver macrophages mediates or participates in recognition
Authors: Mohr M. , Kolb H. , Kolb‐Bachofen V. , Schlepper‐Schäfer J.
Published: February 2012

We studied mechanisms that mediate recognition of human erythrocytes (HRBC) and sheep erythrocytes (SRBC) by rat liver macrophages. We used an in vitro cell binding assay that allows spontaneous formation of cell contacts. Binding of HRBC to rat macrophages shows the following characteristics: inhibition studies with several monosaccharides and oligosaccharides yield complete inhibition of cell contacts with saccharides, which block the GalNAc/Gal‐particle receptor on rat liver macrophages. We found the inhibition pattern: N‐acetyl‐D‐galactosamine, lactose greater than D‐galactose, D‐fucose greater than L‐fucose much greater than N‐acetyl‐D‐glucosamine. Cell binding is dependent on the presence of calcium ions, but not influenced by heat‐aggregated IgG or gangliosides. The inhibition pattern was the same after treatment of HRBC with neuraminidase. Therefore, binding of HRBC, as well as binding of neuraminidase‐treated HRBC, is mediated by the GalNAc/Gal‐particle receptor. Binding of SRBC is partly inhibited by galactose‐related saccharides. Binding is also partly inhibited by heat‐aggregated IgG, gangliosides, and L‐fucose. Complete inhibition of cell contacts with SRBC is achieved by combination of all inhibitors. We therefore conclude that binding of SRBC is mediated by several different mechanisms, including the GalNAc/Gal‐particle receptor. Binding of neuraminidase‐treated SRBC, however, was found to be completely inhibited by saccharides, which block the GalNAc/Gal‐particle receptor. We conclude that the GalNAc/Gal‐particle receptor mediates or participates in recognition of non‐self structures.
Glycosylation process in gonadotrophs: a quantitative electron microscope autoradiographic study with labeled glucosamine
Authors: Hurbain‐Kosmath I. , Counis R. , Jutisz M.
Published: February 2012

Incorporation of [3H]glucosamine into dispersed anterior pituitary cells was studied by electron microscope autoradiography. Gonadotrophs were examined to determine the intracellular route and kinetic patterns of glycosylation. Studies were performed with cells from; (a) normal adult male rats; (b) rats orchidectomized 3 wk earlier; and (c) orchidectomized rats treated with tunicamycin. Our results show that incorporation of [3H]glucosamine first occurs in the rough endoplasmic reticulum (RER), then proceeds in the Golgi elements (where peripheral carbohydrates are attached). Treatment with tunicamycin results in a decrease in labeling of these 2 organelles. Comparison of the kinetic patterns in normal and castrated male rats shows that the accumulation of labeled glycosylated proteins in granules reaches a plateau within 2 hr post‐pulse in normal rats, and rises during a 6‐hr chase in castrated rats. However, because of the necessity for a rather long 15 min pulse, we cannot exclude the possibility that incorporation of glucosamine during the pulse may occur concomitantly in the RER and the Golgi saccules, to be followed by rapid transfer to the secretory granules.
Binucleate cells in the Ehrlich ascites tumor. Action of 5‐fluorouracil
Authors: Pellicer J. A. , Pertusa J. , Alcober V.
Published: February 2012

Time‐dependent frequency distribution of binucleate cells (BC) was studied in Ehrlich ascites tumor (EAT) growing in mice. In animals that received no further treatment, the number of BC increased slowly from 2.6% to 16.5% of total cells within 8 days. In animals that were treated with different doses of 5‐fluorouracil (FU) we found clearly higher numbers of BC. The number of BC increased with tumor age. The increase observed after treatment was reached more quickly in animals that had received the highest FU dose. The final number of BC was also dependent on the age of the tumor at the time of FU injection.
Transmucosal passage of polyalkylcyanoacrylate nanocapsules as a new drug carrier in the small intestine
Authors: Aprahamian M. , Michel C. , Humbert W. , Devissaguet J. P. , Damge C.
Published: February 2012

The enteral absorption of particles has been investigated in the dog using a colloidal drug carrier, polyalkylcyanoacrylate nanocapsules loaded with an iodized oil (Lipiodol), as a tracer for X‐ray microprobe analysis in a scanning electron microscope. Nanocapsules are spherical capsules, 100 to 200 nm in diameter, with a continuous polymeric wall surrounding a cavity which encapsulates the drug. Administered in the jejunal lumen, Lipiodol nanocapsules improved the absorption of the tracer as indicated by increased concentration of iodine in the plasma of mesenteric blood. In order to follow nanocapsules at the cellular level, all tissue compartments were preserved in a life‐like state by cryofixation and freeze‐drying of intestinal biopsies. Nanocapsules appeared in the intestinal lumen close to the mucus, then in intercellular spaces and defects of the mucosa and finally in the lamina propria and blood capillaries; in this latter compartment, the iodine content was four‐fold higher than after intra‐jejunal administration of Lipiodol emulsion. This complete phenomenon occurred only at the tip of the villi and happened within less than 60 min. We conclude that nanocapsules enhance the rate of absorption of Lipiodol and transport the drug from the intestinal lumen to the vascular compartment using a paracellular pathway. Thus they may be useful as drug carrier for oral administration of many chemicals.
Characterization of a 30,000‐35,000 m.w. monokine involved in the specific immune response: intracellular production, secretion and partial purification
Authors: Souvannavong V. , Adam A.
Published: February 2012

Peritoneal macrophages from normal mice which do not secrete interleukin 1 (IL 1) spontaneously release a factor with an approximate m.w. of 30,000‐35,000. In contrast, in the presence of silica only IL 1 is produced. IL 1 as well as this macrophage‐replacing factor (MRF) can restore the antibody response of macrophage‐depleted spleen cells. IL 1, however, is the only one that has the capacity to increase the proliferation of thymocytes. In every strain of mice studied, including nude mice, we observed a spontaneous release of MRF and a silica‐induced shift to the secretion of IL 1, except in lipopolysaccharide (LPS)‐hyporesponsive C3H/HeJ mice. Macrophages from these mice are unable to secrete MRF spontaneously, or IL 1 when stimulated with silica. The kinetics of the secretion of MRF and IL 1 appear to be similar. Macrophages, regardless of whether they have been stimulated to secrete IL 1, produce an intracellular IL 1‐like activity with an approximate m.w. of 15,000. In contrast, the intracellular PFC‐restoring activity is widely distributed in the 15,000‐60,000 m.w. range; one of these compounds could be related to IL 1 precursor and/or to MRF itself. Chromatofocusing and chromatography on Blue Trisacryl have led to a partial purification and resolution from a possible contamination by IL 1. Purified MRF induces, in conjunction with lymphokines, the differentiation of B cells into antibody‐forming cells.
Effects of electrical stimulation upon post‐hatching development of fibre types in normally innervated fast and slow latissimus dorsii muscles of the chicken
Authors: Khaskiye A. , Renaud D. , Le Douarin G.
Published: February 2012

In chicken, the main characteristic properties of muscle fibre types in slow anterior (ALD) and fast posterior (PLD) latissimus dorsii are acquired during post‐hatching development. At day 4 it becomes possible to distinguish between alpha' and beta' fibre types in ALD muscle. At the same time, mATPase staining and NADH‐TR activity permit recognition of alpha w and alpha R fibres within PLD muscle. During further development, muscle fibre typology progressively changes towards the adult slow and fast type. Chronic stimulation at a slow rhythm (5 Hz) of PLD prevents the change in relative proportions of alpha R and alpha W fibres within the muscle that occurs in normal post‐hatching development and increases the number of beta R fibres. Moreover, oxidative activity is increased in all muscle fibre types following stimulation. In ALD muscle, chronic stimulation at a fast rhythm (40 Hz) results in a decrease in oxidative activity and inhibits the differentiation of alpha' and beta' muscle fibre types. This study demonstrates that in young chicken, the pattern of activity influences the differenciation of fibre types in slow and fast muscles.
Spatial organization of collagen in annelid cuticle: order and defects
Authors: Lepescheux Liên
Published: February 2012

The epidermis of Paralvinella grasslei (Polyhaete, Annelida) is covered by an extracellular matrix, the cuticle, mainly composed as in other annelids of superimposed layers of non‐striated collagen fibrils. The collagen fibrils of annelid cuticle are shown to be composed of parallel and sinuous microfibrils (thin section and freeze‐fracture replicas). The 3‐dimensional organization of collagen is characterized by 2 different types of geometrical order: (a) Fibrils form a quasi‐orthogonal network, whose structure is comparable to that of a “plywood”; (b) Fibrils are helical, and goniometric studies show that microfibrils present a definite order within each fibril, which is termed “cylindrical twist”. These 2 characteristics are those which have recently been evidenced in “blue phases”,i.e., liquid crystals which are closely related to cholesteic liquid crystalline phases. Non‐fluid analogues of cholesteric liquids are widespread among invertebrate cuticles and the presence of blue phase analogues suggests that a self‐assembly mechanism is involved in cuticle morphogenesis, which is derived from that governing blue phase growth. The cuticular network presents local rearrangements of fibrils called “defects”, despite the fact that they are elaborate structures which trigonal and pentagonal singularities. Branched fibrils are regularly observed. We discuss the involvement of these pattern disruptions in the cuticle growth process.
Microcalorimetric studies of hybridoma cells
Authors: Nässberger Lennart , Truedsson Lennart , Monti Mario
Published: February 2012

In the present study, overall metabolism has been estimated in hybridoma cells by microcalorimetric measurement. Heat production rate was found to be 30–50 pW/cell at cell concentrations 0.65t‐4.5×105/ml. High cell concentrations (1×106 cells/ml) caused unstable power‐curves with an initial high peak and a rapid declining phase, whereas low cell concentrations (0.6−4.5×105 cells/ml) produced steady‐state power‐curves. Oxygen consumption was found to range between 1.5−6.1×10−5 mol 02/cell/min, corresponding to about 80% of the total metabolic activity.

The metabolic inhibitors sodium fluoride (50 mM), sodium azide (160 mM) androtenone (0.1 mM) caused a reduction in overall cell metabolism of 60, 55 and 40% respectively.
Isolation of developmentally regulated genes in Drosophila melanogaster
Authors: Aït‐Ahmed Ounissa , Griffin‐Shea Ruth , Vola Christine , Capri Michèle , Thomas‐Cavallin Michèle , Pardo Daniel
Published: February 2012

We used a molecular approach to search for maternally expressed genes in Drosophila melanogaster. The relative merits of differential and competition screens were analyzed in a series of reconstruction experiments using either purified phage plaques or derivative DNA sequences. In the course of this study, we isolated 5 clones whose RNA level varies during early embryogenesis. Three gastrula differential clones, b4, b8 and d3, are present in numerous copies in the genome; clone b4 hybridizes with the copia‐like B104 repetitive sequence described by Scherr et al. [29]. We also isolated 2 maternally‐expressed genes, not previously identified in either classical genetic or similarly molecular‐based screens. These clones, b11 and d6, map at cytogenetic positions 98F and 4F respectively, on the polytene chromosome map.
Changes in a nuclear matrix antigen during the cell cycle: interphase and mitotic cells
Authors: Todorov Ivan Todorov , Philipova Padha Nikolava , Zhelev Nikolai Zhelev , Hadjiolov Asen Asenov
Published: February 2012

We studied the behaviour in interphase and mitotic human cells of a 125 kDa (pI 6.5) antigen, associated with the nuclear matrix and detected in proliferating cells. Indirect immunofluorescence with a specific monoclonal antibody reveals that during interphase in WISH and Namalwa cells, as well as phytohaemagglutinin—stimulated lymphocytes, the antigen displays a speckled distribution in the nucleoplasm of all cells. At early prophase the fluorescence intensity of the coalesced speckles increases markedly. During metaphase and anaphase the antigen gives maximal fluorescence distributed diffusely in the nucleoplasm, while chromosomes remain negative. At anaphase and cytokinesis the antigen is still cytoplasmic, but fluorescence intensity decreases. Two‐dimensional gel electrophoresis and immunoblotting reveal that the p125/6.5 antigen displays a net increase in isolated mitotic cells as compared to interphase cells. These results suggest that the p125/6.5 protein participates in late G2 phase and G2/M transition events preparing the cell for mitosis.
Transferrin secretion and hepatocyte ploidy: analysis at the single cell level using a semi‐automatic image analysis method
Authors: Foucrier Jean , Pechinot Danielle , Rigaut Jean Paul , Feldmann Gérard
Published: February 2012

In a previous work it was shown that transferrin (Tf) secretion is directly related to the membrane surface area of hepatocytes (Péchinot D. et al. [31]). The aim of the present work was to search for a possible relationship between Tf secretion and hepatocytic ploidy using a semi‐automatic image analysis method. A determination of Tf secretion by isolated normal adult hepatocytes was achieved at the single cell level, using a modified reverse hemolytic plaque test. A Feulgen reaction was also performed on these hepatocytes. It allowed the evaluation, for each secreting hepatocyte, of the quantity of Tf secreted and its nuclear characteristics. Discrimination between diploid (2c) and tetraploid (4c and 2c2c) hepatocytes was performed and the amount of Tf secreted by each subpopulation determined. It appeared that a 2‐fold secretion ratio was not found between tetraploid and diploid hepatocytes. These results suggest, as Tf production is not directly proportional to the degree of ploidy of hepatocytes, that some not yet elucidated regulatory mechanisms may act on Tf gene expression.
Regeneration of muscles after cardiotoxin injury I. Cytological aspects
Authors: Couteaux René , Mira Jean‐Ciaude , d'Albis Anne
Published: February 2012

Regeneration of several adult rat and mouse skeletal muscles was studied after degeneration of muscle fibers had been obtained by the selective action of the cardiotoxin of Naja mossambica mossambica venom. Experimental conditions were set up to ensure minimal damage to satellite cells and also the nerves and blood vessels of the original muscles. As in the other types of experimental regeneration, the structure of the regenerated muscle appeared in many respects different from that of the normal muscle. Moreover the neuromuscular junctions of ‘en plaque’ type were transformed to ‘en grappe’ type junctions. May ultrastructural abnormalities often displayed by these junctions might be linked, at least partially, to the persistence in the regenerating muscle of the original synaptic basal lamina sheaths and their inductive properties.
The iron mineralization of Spongia officinalis L. (Porifera, Dictyoceratida) and its relationships with the collagen skeleton
Authors: Vacelet Jean , Verdenal Benoît , Perinet Guy
Published: February 2012

Lepidocrocite (γ FeOOH) granules are frequently found in the spongin fibers of some horny sponges. The granules also contain calcite and a small amount of goethite (α FeOOH). In the genus Spongia, the amount of iron is highly variable according to the habitat and to the age of the fiber. Some indications about the skeletal growth and structure can be inferred from the distribution of the biominerals and the arrangement of the crystallites: i.e. (1) Mineralization is a slow process in comparison with fiber formation; (2) Secondary connective fibers may have a discontinuous growth; (3) The collagen microfibrils display a definite arrangement in the successive spongin layers; (4) Although spongin is a collagen highly resistant in vitro to enzymatic digestion, the sponge is able to perform remodeling of its skeleton through autodigestion of the fibers.
Culture of an epithelial cell line (MDCK) with a serum substitute
Authors: Gutiérrez M.C. , Díaz‐Pontones D.M. , Vancell M.R. López , Beaty G.
Published: February 2012

This paper reports the suitability of culturing a line of dog kidney epithelial cells, MDCK, in the presence of a serum substitute, Ultroser G. Serial subcultivation with this product was possible for at least 10 passages without any change in cell shape and size, saturation density, dome‐forming ability, transepithelial resistance, and growth curve. Adhesion of newly plated cells to plastic was somewhat lower than in fetal calf serum but the trypsin‐harvesting kinetics were essentially the same. However, the membrane ion transport systems was alterd: cell sodium influx was greatly diminished, suggesting a deep change in the amiloride‐sensitive Na+ channels: sodium efflux was highly enhanced (both active and passive).
Biosynthesis of glycosaminoglycans in the human colonic tumor cell line Caco‐2: structural changes occurring with the morphological differentiation of the cells
Authors: Levy Peggy , Robert Agnès , Picard Jacques
Published: February 2012

The human colon cancer cell line Caco‐2 cultured in vitro displayed morphological differentiation which was shown to be a growth‐related event. We have investigated this phenomenon further in relation to the cell surface glycosaminoglycans produced by growing (5‐day, i.e., prior to differentiation) and confluent (9‐day, i.e., after morphological and functional differentiation) cultures. Neosynthesized [35S]glycosaminoglycans were purified on DEAE‐cellulose; at confluency, they were bound more strongly to the column than the corresponding fractions from the growing cells. Analysis of Kav values of heparan sulfate and chondroitin sulfates from growing and confluent cells indicated an increase in chain length of both glycosaminoglycans in morphologically differentiated cells. Heparan sulfate was the main 35S‐labeled glycosaminoglycan of the cell surface of both 5‐day and 9‐day cultures. Paper chromatography of the unsaturated disaccharides obtained by chondroitinase digestion showed that chondroitin sulfate chains were primarily 6‐sulfated in the 2 studied extracts. Heparan sulfate chains were isolated as chondroitinase‐resistant material and treated with nitrous acid. Analysis of N‐ and O‐sulfate group‐related radioactivity showed an increase in the amount of 35S‐label in the form of N‐sulfate groups and an increase in the O‐35S‐sulfation pattern in heparan sulfate from morphologically differentiated cells. Thus, the structural features of both chondroitin sulfates and heparan sulfate were significantly different when the growing cells became morphologically differentiated.
Immunocytochemical detection of ribulose bisphosphate carboxylase in capsicum plastids
Authors: Cheniclet Catherine , Suire Claude , Carde Jean‐Pierre
Published: February 2012

Ribulose bisphosphate carboxylase (RUBPCase) was localized by fluorescence and gold immunocytochemistry in Capsicum fruits. Chloroplasts of the green fruit are heavily labelled. A positive staining is also obtained with chromoplasts of the ripe rad fruit, but gold labelling is fainter. The presence of reactive RuBPCase in chromoplasts is discussed in relation with the absence of ribosomes in these plastids.