Saccharin induces morphological changes and enhances prolactin production in GH4C1 cells

Summary The artificial sweetener saccharin inhibits binding of epidermal growth factor (EGF) to cultured rat pituitary tumor cells (GH₄C₁ cells). Saccharin also causes morphological alterations in these cells, resulting in pronounced elongation, stretching, and firmer attachment of cells to the culture dishes. These alterations in cell shape are similar to those observed after treatment of GH₄C₁ cells with EGF and with thyrotropin-releasing hormone (TRH), both of which enhance prolactin (PRL) production in these cells. After assaying for PRL in saccharin-treated cultures, it was observed that this sweetener is also capable of stimulating PRL production two-to sixfold in a dose-dependent manner. Enhancement of PRL production can be observed at 0.5 mM saccharin, yet this is 10 times less than the saccharin concentration required to alter cell shape. These effects of saccharin on cell morphology and on PRL production are reversible in GH₄C₁ cell cultures. When added to cultures along with maximal concentrations of EGF or TRH, the effects of saccharin on PRL production are additive, suggesting that the actions of saccharin are mediated by a somewhat different pathway from that of the peptide hormones. Pulse labeling studies indicate that the enhancement of PRL production is highly specific inasmuch as saccharin was found to decrease the overall rate of protein synthesis in these cells. Saccharin also causes a decrease in the rate of DNA synthesis under these treatment conditions. Mitomycin C, which similarly inhibited DNA synthesis, had no effect on cell morphology or PRL production. This investigation was supported by a Faculty Research Grant from Wheaton College     

Regulation of Na/K/Cl cotransport in vascular smooth muscle cells

The regulation of Na/K/Cl cotransport was investigated in vascular smooth muscle cells. That a Na/K/Cl cotransport system exists was established by the finding that the ouabain insensitive K influx was sensitive to the "loop" diuretic bumetanide. Furthermore, bumetanide sensitive K influx was dependent upon the presence of both Na and Cl in the extracellular milieu. Bumetanide sensitive K influx was inhibited by agents which elevate cellular cyclic AMP levels, and to a lesser extent by agents which elevate cellular cyclic GMP levels. When serum, EGF or TPA was added, bumetanide sensitive K influx was enhanced. These results suggest that vascular smooth muscle cells have a ouabain insensitive, bumetanide sensitive Na/K/Cl cotransport system which is stimulated by serum, EGF or TPA and inhibited by cAMP or cGMP.     

Calcium compartments and fluxes are affected by the src gene product of Rat-1 cells transformed by temperature-sensitive Rous sarcoma virus

Total cellular calcium content (determined by atomic absorption spectrometry) of Rat-1 cells transformed by temperature-sensitive Rous sarcoma virus decreases with cell density, but is found not significantly different at permissive and at non-permissive temperature. Kinetic analysis of 45Ca efflux from preloaded cells exhibits three separable pools of exchangeable calcium. The ratio of pool size of the fast-exchanging Ca-compartment (bound to cell surface) to pool size of the intermediate Ca-compartment (cytoplasmic) was found to decrease from 2.5 to 1.3 upon shift from non-permissive to permissive temperature. The slowly exchanging Ca-pool (presumably mitochondrial) did not change significantly upon temperature shift. These and further data demonstrate a close correlation between distribution of cellular Ca among different cellular compartments and characteristics of cellular proliferation, both attributable to the function(s) of a single oncogene.     

Expression of epidermal growth factor receptor and associated glycoprotein on cultured human brain tumor cells

The expression of epidermal growth factor (EGF-R) in normal glial and glioma cells grown in culture was examined by using several independent assays. Immunoprecipitation with the monoclonal antibody R1 of extracts from metabolically labeled glial and glioma cells revealed a protein of Mr approximately 170,000, with a migration in sodium dodecyl sulfate-polyacrylamide gels identical to the EGR-R of A431 epidermal carcinoma cells. Furthermore, in the majority of glioma extracts, a protein of Mr approximately 190,000 was specifically immunoprecipitated by this antibody. Similar results were obtained by immunoblotting with a second antibody directed against a synthetic peptide in the sequence of the v-erb-B oncogene. In cell lines expressing both proteins, each was specifically phosphorylated on tyrosine in immune complex kinase assays. The majority of glioma cells bound between 40,000 to 80,000 125I-labeled epidermal growth factor molecules per cell. These results suggest that the expression of EGF-R is common in cultured human glioma cells. In addition, a structurally related protein, is expressed in some of these cells.     

Correlation between infection by Rhizobium leguminosarum and lectin on the surface of Pisum sativum L. roots

The lectin on the surface of 4- and 5-dold pea roots was located by the use of indirect immunofluorescence. Specific antibodies raised in rabbits against pea seed isolectin 2, which crossreact with root lectins, were used as primary immunoglobulins and were visualized with fluorescein- or tetramethylrhodamine-isothiocyanate-labeled goat antirabbit immunoglobulin G. Lectin was observed on the tips of newly formed, growing root hairs and on epidermal cells located just below the young hairs. On both types of cells, lectin was concentrated in dense small patches rather than uniformly distributed. Lectin-positive young hairs were grouped opposite the (proto)xylematic poles. Older but still-elongating root hairs presented only traces of lectin or none at all. A similar pattern of distribution was found in different pea cultivars, as well as in a supernodulating and a non-nodulating pea mutant. Growth in a nitrate concentration which inhibits nodulation did not affect lectin distribution on the surface of pea roots of this age. We tested whether or not the root zones where lectin was observed were susceptible to infection by Rhizobium leguminosarum. When low inoculum doses (consisting of less than 10⁶ bacteria·ml^-1) were placed next to lectin-positive epidermal cells and on newly formed root hairs, nodules on the primary roots were formed in 73% and 90% of the plants, respectively. Only a few plants showed primary root nodulation when the inoculum was placed on the root zone where lectin was scarce or absent. These results show that lectin is present at those sites on the pea root that are susceptible to infection by the bacterial symbiont.Abbreviations FITC fluorescein isothiocyanate - TRIC tetramethylrhodamine isothiocyanate     

The cytoskeleton of Cobaea seed hairs:

The cell wall of Cobaea scandens seed hairs developed in a characteristic sequence, with the deposition of a cellulose thread onto a pectic swelling layer was the final event. The cellulose thread was intracellularly accompanied by a band of 10–18 microtubules. During the formation of the swelling layer the microtubules were homogeneously distributed; they ran circumferentially normal to the cell axis. When cellulose-thread formation started, the microtubules became arranged in a helical band. The density of the microtubules varied during the different phases of development. The highest density was observed before cellulosethread formation and ranged from 6–15 m·m^-2. The length of the microtubules, 20–30 m, was determined by direct measurements, as well as estimated from the total microtubular length in a given area and the counted free ends. With the indirect immunofluorescence technique the microtubules of the band stained inhomogeneously. Those which were located at the edges of the band fluoresced more intensely than those of the central part. Attempts to visualize actin filaments in the hair cells with rhodaminyl-conjugated phalloidin resulted in a homogeneous staining of the area of the microtubular band, indicating that actin filaments may be present in this region. Though, in thin sections and dry-cleaved cells, filamentous structures were observed between the microtubules, caution is expressed that the observed fluorescence was, indeed, due to actin filaments. The role of the filamentous structures is discussed with respect to formation and maintenance of the microtubular band. Microtubules apparently did not cross coated pits which were visualized in the plasma membrane through the dry-cleaving technique.Abbreviations IFT indirect immunofluorescence technique - RP rhodaminyl-conjugated phalloidin - SEM scanning electron microscopy     

High-frequency1H NMR studies of the effects of growth factors and phorbol esters on normal syrian hamster diploid fibroblast cells

The nuclear magnetic resonance (NMR) parameters, spin-lattice (T₁), and spin-spin (T₂) relaxation time, are usually longer for neoplastic cells than for normal cells of the same cell type. This has generally been true at low NMR frequencies (≤100 MHz) when comparisons have been made between normal and neoplastic cells that have both spent a short time in culture. We have previously demonstrated that although the T₁ values of paired normal and neoplastic Syrian hamster (SH) fibroblastic cells in culture are not significantly different when measured at 300 MHz, the 300 MHz T₂ values for the neoplastic cells are smaller than those of the normal cells. (Xin et al. (1986),Cell Biophysics 8, 213.) Since treatment of normal diploid cells with polypeptide growth factors or tumor promoters frequently results in reversible expression of neoplasia-associated phenotypes, T₁ and T₂ were obtained at 300 MHz for treated and untreated SH cells to see if these compounds could also produce smaller 300 MHz T₂ values. Secondary culture SH fetal fibroblast cells were treated with epidermal growth factor (EGF), fibroblast growth factor (FGF), phorbol-12,13-didecanoate (PDD) and 4-α-phorbol-12,13-didecanoate (4αPDD). Treatment with either growth factor resulted in smaller T₂ values, but a statistically significant decrease was not observed for PDD or 4αPDD. The observed reductions in T₂ values were correlated with the morphological and growth-stimulatory effects of these compounds on the cells.     

X射线对根尖细胞分裂和分化的作用

用30—70GyX射线照射小麦幼苗(浸种后5天)后发现根毛区到根尖的距离缩短,根毛变密,根毛长度为对照的2—3倍。照射后2—3天就可看到该现象。根毛着生处到根尖的距离随剂量增加而减少,甚至根尖全为根毛所复盖,另外还看到有分叉的根毛。根尖纵切片表明根尖分生区随剂量增加而缩小,分生区后接着就出现输导组织。玉米和黄瓜也有类似现象。这现象说明根细胞的分裂过程对射线很敏感,而分化过程则相当耐辐射,细胞分裂停止后立即转向细胞分化过程。     

Influence of cytosolic pH changes on the organisation of the endoplasmic reticulum in epidermal cells of onion bulb scales: Acidification by loading with weak organic acids

Summary The anastomosing ER system of epidermal cells of onion bulb scales is composed of three modifications: lamellar and tubular elements, located in the cell periphery, and long tubular stands located deeper in the cytoplasm. Cytoplasmic acidification of epidermal cells by loading with weak organic acids like acetic or propionic acid causes the decay of the lamellar elements and the disappearance of long tubular strands. Organelle movement is also inhibited. The effects depend on the pH of the incubation medium and on the administered acid concentration, and are characterized by a distinct lag phase of about 7 min. The induced ER changes are transient with adaptation starting after about 50min. Buffer components alone have little influence on the cellular ER organization within a pH-range of 4.0–8.0. However, the pH of the medium strongly affects the time course of the effects as well as recovery after omitting the administered acid. Both modulation and recovery occur more rapidly at neutral or slightly alkaline pH. Actin filaments, which play a major role in ER organization and organelle movement, are not affected by cytosolic acidification.Dedicated to the memory of Professor Oswald Kiermayer