bluuebunny/crossref_metadata_2025_split huggingface dataset:
The in vivo and ultrastructural effects of injection of lysophosphatidyl choline into myelinated peripheral nerve fibres of the adult mouse
Authors: Hall Susan M. , Gregson N. A. 
Published: April 2021

ABSTRACT
The action of phospholipase A and lysophosphatidyl choline (LPC) on mature, myelinated peripheral nerve fibres has been studied in vivo and electron microscopically, following sub- perineurial injection of these substances. Within 30 min, demyelination was observed in vivo along cylindrico-conical segments, spreading from Schmidt-Lanterman incisures and nodes of Ranvier. By 96 h, all traces of the myelin sheath had disappeared from the area of the lesion, and had been replaced by debris-laden cells lying in chains parallel to one another and the long axis of the fibre. During the next few weeks these cells gradually disappeared, and numerous finely myelinated axons, running between, and in continuity with, the normal fibres proximal and distal to the lesion were observed. If lower concentrations of LPC were used the number of fibres involved decreased, although the demyelinative changes followed the same time-course.
Ultrastructurally, demyelination involved progressive disruption and removal of the lamellar sheath, observed initially as a splitting of the intraperiod line within 30 min. Subsequent breakdown resulted in the formation of strands of 4-6 nm repeat material which was further degraded through quintuple- and triple-layered lamellar units to foam-like systems of disorganized lamellar fragments. The Schwann cell and axons appeared to be undamaged by phospholipase A and LPC, and retained their normal impermeability to exogenous ferritin.
The significance of the demyelinating capacity of LPC in vivo is discussed in terms of its known action on myelin in vitro, the rapidity and apparent specificity of its action demonstrated in this study, and its potential involvement in pathological demyelination.
Pachytene spermatocyte and round spermatid binding to sertoli cells in vitro
Authors: Enders George C. , Millette Clarke F. 
Published: April 2021

ABSTRACT
Spermatogenic cells differentiate in vivo while in continuous contact with the Sertoli cell. During differentiation, the spermatogenic cells and Sertoli cells form a number of morphologically distinct stage-specific adhesions. We describe an in vitro assay system for studying the adhesion of spermatogenic cells to Sertoli cell monolayers. Mixed populations of spermatogenic cells or enriched fractions of pachytene spermatocytes and round spermatids were labelled with the vital dye, fluorescein diacetate, prior to their addition to Sertoli cell monolayers so that the adhesion of viable spermatogenic cells could be quantified. Using this assay system, the number of pachytene spermatocyte and round spermatid binding sites on the Sertoli cell monolayer were similar, but the kinetics of binding were different. Pachytene spermatocytes were able to inhibit significantly round spermatid binding, while round spermatids did not significantly inhibit pachytene spermatocyte binding. After coculture for 24–48 h, spermatocytes form junctional structures with Sertoli cells that are similar to desmosome-like junctions. These results suggest that pachytene spermatocytes and round spermatids bind to Sertoli cells by different mechanisms.
The somatic function of the micronucleus in sexual reproduction of Paramecium tetraurelia: Initiation of oral membranelie assembly
Authors: Chau M. F. , Ng Stephen F. 
Published: April 2021

ABSTRACT
In a previous study, cell lines possessing defective micronuclei, generated by laser-microbeam irradiation, gave rise to cells lacking both oral apparatus and micronuclear derivatives after autogamy. It was concluded that astomy arose as a result of degeneration of all of the meiotic products of the micronuclei after meiotic telophase II, instead of leaving one product for subsequent nuclear reorganization. The present study consolidates this conclusion by employing 15 micronucleus-defective cell lines; these were generated by laser-irradiation of the micronucleus, treatment of the cells with cis-dichloro-diammineplatinum (II), and conjugation between diploids and amicronucleates to produce haploids. A good correlation between the presence of pregametic, gametic and zygotic nuclei and the initiation of oral membranelie assembly in stomatogenesis was demonstrated in 17 cases of autogamy. Therefore, postmeiotic micronuclear activities up to the zygotic nucleus stage, in particular in the gametic stage, are crucial for the initiation of oral membranelie assembly, while premeiotic micronuclear activities are insufficient. Micronuclear genic factors are also likely to be involved in the determination of the fate of the meiotic products.
A morphologically distinct candidate for an epithelial stem cell in mouse mammary gland
Authors: Smith Gilbert H. , Medina Daniel 
Published: April 2021

ABSTRACT
Transplantation studies demonstrate that an epithelial stem cell component must exist in the mouse mammary gland throughout life. Samples taken from any portion of the mammary gland at any age and at any developmental stage, including full functional differentiation, give rise to mammary epithelial outgrowths with complete developmental capacity. Cytological examination of mouse mammary gland explants revealed the presence of morphologically distinct cells distributed sporadically among the mammary epithelium, whose behaviour in vivo and in vitro suggested that they might represent a latent epithelial stem cell population. These pale-staining cells possessed large spherical nuclei, a clear cytoplasm and a round smooth-contoured shape. Electron microscopy confirmed their pale-staining characteristics and revealed a cytoplasm sparsely populated with organellar structures, such as mitochondria and endoplasmic reticulum. Their epithelial genealogy was demonstrated by the presence of terminal bars and tight junctions formed with their epithelial cell neighbours. In vivo, these cells were found among mammary epithelial cell populations in 16-day-old embryos onward in both ductal or lobular structures during all stages of pregnancy, lactation and involution. In explant cultures, these cells did not undertake a secretory morphology in the presence of lactogenic hormones, although occasionally they became immunologically positive for casein. They did not incorporate [3H]-thymidine into their nuclei under any of the experimental conditions used; however, they appeared to undergo mitosis within 4 h regardless of the presence or absence of hormone(s). At 24 h increased numbers of pale cells were found in pairs or in groups. At 72 h in the presence of IFPrl (medium containing insulin, hydrocortisone and prolactin), the pairs and groups of pale cells observed at 24 h were not found. Instead, individual pale cells were seen among groups of cytologically and functionally differentiated secretory epithelial cells. When lactogenic hormones were not present, groups of pale cells were still present in the explants at 72 h. These findings suggest that the pale cells are arrested at Gz phase of the cell cycle and that they give rise by mitosis to daughter cells capable of differentiating in the presence of lactogenic stimuli. Inhibition of DNA synthesis in the explants did not alter these cellular events.
A Dictyostelium mutant with severe defects in α-actinin: its characterization using cDNA probes and monoclonal antibodies
Authors: Schleicher Michael , Noegel Angelika , Schwarz Tobias , Raff Eva Wall , Brink Marijke , Faix Jan , Geriscii Glinther , Isenberg Gerhard 
Published: April 2021

ABSTRACT
Cells of a Dictyostelium discoideum mutant de­ficient in binding a monoclonal antibody to α·actinin have previously been shown to grow and develop similarly to the wild type and to exert unimpaired chemotaxis as well as patching and capping of membrane proteins. Here we show that the normal 3.0 kb message for α-actinin is replaced in the mutant by two RNA species of approximately 3.1 and 2.8 kb. The 3-1 kb RNA was recognized by DNA fragments from all parts of the coding region, while the 2.8 kb RNA hybrid­ized to all but a 3'-terminal fragment. Proteins synthesized in the mutant were analysed using four monoclonal antibodies that in the wild type specifically recognize the 95×l03Mr polypeptide of α-actinin. Cleavage mapping indicated that the binding sites of these antibodies are distributed over a region comprising more than half of the α- actinin polypeptide chain. In the mutant, three of the antibodies faintly labelled two polypeptides of 95×l03Mr and 88×l03Mr; the fourth antibody, which binds closest to one end of the polypeptide chain, faintly labelled the 95×l03Mr polypeptide only. The 88×l03Mr polypeptide most probably lacks the C-terminal portion of α-actinin.
The binding of an antibody that recognized both polypeptides was quantified by a radio- immuno competition assay using wild-type α- actinin as a reference. In a mutant cell extract containing total soluble proteins the antibody binding activity was decreased to 1.1% when compared with wild-type extract. After their par­tial purification and SDS–polyacrylamide gel electrophoresis the mutant 95 × 103Mr and 88 × 103Mr polypeptides were barely detectable as Coomassie Blue-stained bands, indicating that in the mutant not only certain epitopes of α-actinin were altered but the entire molecule is almost completely lacking. When the fitness of mutant cells relative to wild type was determined during growth in nutrient medium, a slight disadvantage for the mutant was indicated, by finding selection coefficients between 0.03 and 0.05.
Cultured epithelial cells derived from human foetal pancreas as a model for the study of cystic fibrosis: Further analyses on the origins and nature of the cell types
Authors: Harris Ann , Coleman Lindsay 
Published: April 2021

ABSTRACT
The establishment of a tissue-culture system for epithelial cells derived from human foetal pancreas has recently been reported. Further analyses have now been made on these cells in vitro, together with parallel investigation of the distribution of different cell types within the intact foetal pancreas. Results support the view that the cultured cells are ductal in origin and nature. Pancreatic epithelial cell cultures have also been established from foetuses with cystic fibrosis.
Differences in cytoplasmic ph and the sensitivity to acid load between prespore cells and prestalk cells of Dictyostelium
Authors: Inouye Kei 
Published: April 2021

ABSTRACT
The relationship between cytoplasmic pH (pH)) and prestalk/prespore differentiation in the cellular slime mould Dictyostelium discoideum was investigated by: (1) comparing the pHi of prestalk and prespore cells, and (2) examining the effects on pH; of weak acids, weak bases and proton-pump inhibitors. These substances are known to affect the spore-to-stalk ratio. The pHi of dissociated slug cells was measured by microfluorometry using a pH-sensitive fluorescence dye, carboxyfluorescein.
There was a slight but significant difference in pHi between prestalk cells (7’38 ± 0·09) and prespore cells (7·49 ± 0·03), and prestalk cells were more sensitive to acid load than prespore cells. In addition, weak acids and proton-pump inhibitors, which promote prestalk differentiation, significantly lowered pHi whereas weak bases, which induce prespore differentiation, elevated pHi. However, differentiation inducing factor (DIF), an endogenous inducer of stalk differentiation, caused only a slight and transient decrease in pHi. These results suggest two alternative possibilities: (1) a small and transient decrease in pHi is sufficient to induce prestalk differentiation, or (2) there is a mediator other than pHi that links the action of DIF and the pathway choice.
Antibodies to rat sertoli cell secretory proteins recognize antigenic sites in acrosome and tail of developing spermatids and sperm
Authors: Kierszenbaum Abraham L. , Ueda Hiroshi , Ping Liu , Abdullah Munir , Tres Laura L. 
Published: April 2021

ABSTRACT
Polyclonal antisera raised against polypeptide components of antigenically related rat Sertoli cell secretory protein S70 and S45–S35 heterodimeric protein were used to identify antigenic sites in rat testis during spermatogenesis. Immunofluorescence, immunoperoxidase and immunogold electron-microscopic methods have shown that all antisera recognize immunoreactive sites in the acrosome of developing spermatids. Apical cytoplasmic regions of Sertoli cells in close association with bundles of developing spermatids displayed an immunoreactive product. The principal piece of the developing tail of maturing spermatids immunostained with antisera to Sertoli cell secretory S45–S35 heterodimeric protein, displayed characteristic apical-to-distal immunoreactive gradient patterns before acquiring uniform immunoreactivity at completion of maturation. Immunogold electron microscopy demonstrated that outer dense fibres were the predominant immunoreactive site. Results of this work, together with previous immunoblotting and immunofluorescent data, demonstrate that both Sertoli cell secretory proteins and components of the acrosome and tail of developing spermatids and epididymal sperm share antigenic homology.
In vitro modulation of endothelial fenestrae: Opposing effects of retinoic acid and transforming grow h factor β
Authors: Lombardi T. , Montesano R. , Furie M. B. , Silverstein S. C. , Orci L. 
Published: April 2021

ABSTRACT
Cultured endothelial cells isolated from fenestrated capillaries express many properties characteristic of their in vivo differentiated phenotype, including the formation of a limited number of fenestrae. In this study, we have investigated whether physiological factors that control cell differentiation might regulate the surface density of fenestrae in capillary endothelial cells. We have found that treatment of the cultures with retinoic acid (10μM) induces a more than threefold increase in the surface density of endothelial fenestrae, whereastransforming growth factor β (TGFβ) (2 ng ml−1) causes a sevenfold decrease in the surface density of these structures. These results show that the expression of endothelial fenestrae is susceptible to bidirectional modulation by physiological signals, and suggest that retinoids and TGFβ may participate in the regulation of fenestral density of capillary endothelium in vivo.
Transferrin receptors on the surfaces of retinal pigment epithelial cells are associated with the cytoskeleton
Authors: Hunt Richard C. , Dewey Art , Davis Alberta A. 
Published: April 2021

ABSTRACT
Retinal pigment epithelial cells, derived from human donor eyes, have been grown in culture as monolayers on membrane filters or plastic surfaces and shown to possess transferrin receptors with a monomeric molecular mass of 93000. These receptors internalize 12SI-labelled transferrin and recycle it to the surrounding medium in a similar manner to other cell types. Scatchard analyses show that there are about 100 000 high-affinity receptors on the surface of each cell and most of these receptors are associated with the cytoskeleton. In total cell extracts, there are additional low-affinity binding sites that do not appear to be strongly associated with the cytoskeleton. The apparent interaction of transferrin receptors with the cytoskeleton was confirmed in two ways: first, using 200 kV electron microscopy for stereo analyses, skeleton-associated transferrin receptors were detected by a monoclonal anti-receptor antibody and a colloidal gold-conjugated second antibody after Triton X-100 extraction of pigment epithelial cells grown directly on laminin-coated gold grids; and, second, when cell surface receptors were labelled with radioiodinated transferrin and then incubated for various periods of time, the labelled transferrin was observed to move from a Triton X-100-insoluble fraction (a putative cytoskeletal compartment) to a Triton-soluble compartment that was not associated with the cytoskeleton. Using either horseradish peroxidase or colloidal gold-labelled transferrin, it has been shown that basolateral and apical surface-located receptors participate in receptor-mediated endocytosis via clathrin-coated pits, endosomes and tubular structures. Initially, transferrin internalized from the apical surface is observed in small endosomes that often appear to be embedded in an apical layer of microfilaments. From these peripheral regions of the cells, the labelled receptors move to larger endosomes and multivesicular bodies deeper in the cytoplasm. These structures have no apparent association with cytoskeletal elements.
Tandem scanning reflected-light microscopy of cell-substratum adhesions and stress fibres in swiss 3T3 cells
Authors: Paddock Stephen W. 
Published: April 2021

ABSTRACT
This paper describes two applications of the tandem scanning reflected-light microscope (TSM) for the observation of the structure of individual cells growing in tissue culture. First, the TSM is used as an alternative to interference reflection microscopy (IRM) or total internal reflection aqueous fluorescence microscopy (TIRAF) to observe cell-substratum adhesions in unstained living cells growing on a glass coverslip. Second, the TSM is used to produce improved images of cellular structures in 3T3 cells stained with various protein dyes including Napthol Blue Black (NBB) and Coomassie Brilliant Blue (CBB). More specifically, close contacts and focal contacts are resolved in living 3T3 cells, and features of the nucleus, the cytoskeleton and extracellular matrix are resolved in both NBB- and CBB-stained cells. The focal contacts and associated stress fibres are clearly imaged in NBB-stained cells. The TSM is an improvement over conventional incident light microscopy because the confocal image excludes information from out-of-focus regions of the cytoplasm, and, unlike the laserbased confocal microscope, the actual colour of the specimen is viewed directly with the TSM in almost real-time.
Cross-linking of microtubules by microtubule-associated proteins (MAPs) from the brine shrimp, Artemia
Authors: Campbell Elizabeth J. , Mackinlay Sara Ann , Macrae Thomas H. 
Published: April 2021

ABSTRACT
Microtubules induced with taxol to assemble in cell-free extracts of the brine shrimp, Artemia, are cross-linked by microtubule-associated proteins (MAPs). When the MAPs, extracted from taxol-stabilized microtubules with lM-NaCl are co-assembled with purified Artemia or mammalian neural tubulin, reconstitution of cross-linking between microtubules occurs. The most prominent non-tubulin protein associated with reconstituted cross-linked microtubules has a molecular weight of 49000 but we cannot yet exclude the possibility that other proteins may be responsible for the crosslinking. Cross-linkers are separated by varying distances while cross-linked microtubules, pre pared under different conditions, are 6·9-7·7nm apart. Cross-linking of microtubules by MAPs occurs whether MAPs are added to assembling tubulin or to microtubules, and it is not disrupted by ATP. The MAPs are heat-sensitive and do not stabilize microtubules to cold. Immunological characterization of Artemia MAPs on Western blots indicates that Artemia lack MAP 1, MAP 2 and tau. Our results clearly demonstrate that Artemia contain novel MAPs with the ability to cross-link microtubules from phylogenetically disparate organisms in an ATP-independent manner.
Synthesis and phosphorylation of cytoskeletal proteins during the in vitro biogenesis of MDCK cell monolayers
Authors: Benítez-King G. , Cazares F. , Meza I. 
Published: April 2021

ABSTRACT
In vitro formation of a functional MDCK cell monolayer requires the dynamic participation of the cytoskeleton. Cell shape, contacts and polarity, as well as transepithelial electric resistance (TER), are actively modified during this differentiation process. We studied the distribution and rearrangement of cytokeratin, vimentin and actin filaments that occur in the monolayer concomitant with the synthesis and phosphorylation of these proteins. Cells cultured for short time in suspension, subconfluent or early confluent cells, showed striking differences in cell shape and arrangement of their cytoskeletal filaments. Subconfluent and early confluent cells synthesized proteins at high levels. In contrast, suspended cells maintained lower rates of protein synthesis. Labeled amino acid uptake was very similar in all these culture conditions. Gel electrophoresis analyses of the synthesized proteins showed increases in the synthesis of actin, vimentin and specific cytokeratins in subconfluent and early confluent cells. On the other hand a decrease in total protein, actin, vimentin and cytokeratin synthesis was observed in cells kept in suspension for 24 h or in 78-h or 174-h older confluent cultures. These cultures also showed a decrease in the uptake of labeled amino acid. Cytokeratin and vimentin phosphorylation rates were also modified during the in vitro formation of a monolayer. In suspended cells, cytokeratins were phosphorylated and three labeled isoelectric variants of the 40, 48 and 58K (K = 103Mr) cytokeratins were present in intermediate filament extracts. In subconfluent and early confluent cells only two isoelectric phosphorylated variants of the 40, 48 and 58K cytokeratins were detected and vimentin was also phosphorylated. In confluent monolayers cultured 78 and 174 h, only 40 and 48K cytokeratins remained labeled, while the 58K cytokeratin and vimentin, although present in the cell extracts, were no longer phosphorylated. Our results indicate that concerted changes occur in the distribution of intermediate and actin filaments, and the relative rate of synthesis and phosphorylation of cytoskeletal proteins during formation of an MDCK cell functional monolayer.
Reduction of G1 phase duration and enhancement of c-myc gene expression in HeLa cells at hypergravity
Authors: Kumei Yasuhiro , Nakajima Tohru , Sato Atsushige , Kamata Nobuyuki , Enomoto Shoji 
Published: April 2021

ABSTRACT
We have found that hypergravity stimulates the proliferation of HeLa cells through reduction of the Gt phase duration, concomitant with enhancement of c-myc gene expression. HeLa cells were grown in monolayer in culture flasks that were centrifuged to generate a constant 18, 35 or 70g at 37°C for up to 4 days. The cell proliferation was enhanced at 18, 35 and 70g, most notably at 35 g. Cell cycle analyses with [3H] thymidine (TdR)-colcemid treatment showed that the cell generation time in the 35 g culture was reduced by 17% as compared to the control, which was attributed to a 26% reduction of the Gi phase duration. No differences were observed in the duration of the S, G2 and M phases or in the [3H]TdR incorporation per S phase cell between the 35g culture and the control. The induction of c-myc gene expression was investigated by
RNA blot hybridization during a 15–360 min exposure of cells to 18, 35 and 70g. Elevated levels of c-myc mRNA were observed after a 15-min exposure, and maintained after a 360-min exposure at all hypergravities examined. The highest induction rate of c-myc mRNA was 3·8-fold higher than the control after a 120-min exposure to 35 g. The 35g condition was the most effective hypergravity for stimulating both cell proliferation and c-myc gene expression. Our study suggests that the appropriate level of hypergravity stimulates HeLa cell proliferation by reducing the G1 phase duration without affecting DNA synthesis rate, mediated through induction of c-myc gene expression.