Showing papers on "Cellular differentiation published in 1970"

Inhibition of myoblast fusion after one round of dna synthesis in 5-bromodeoxyuridine

Abstract: The thymidine analogue 5-bromodeoxyuridine (BUdR) has a differential effect on the synthesis of tissue-specific products and molecules required for growth and division. Proliferating myogenic cells cultured in BUdR fail to fuse and fail to initiate the synthesis of contractile protein filaments. Conversely, BUdR has but a minor effect on cell viability and reproductive integrity. Low concentrations of BUdR result in an enhancement of cell number relative to the controls; higher concentrations are cytotoxic. Suppression of myogenesis is reversible after at least 10 cell generations of growth in the analogue. Cells that do not synthesize DNA, such as postmitotic myoblasts and myotubes, are not affected by BUdR. Incorporation of BUdR for one round of DNA synthesis was accomplished by first incubating myogenic cells, prior to fusion, in 5-fluorodeoxyuridine (FUdR) to block DNA synthesis and collect cells in the presynthetic phase. The cells were then allowed to synthesize either normal DNA or BU-DNA for one S period by circumventing the FUdR block with BUdR or BUdR plus thymidine (TdR). The cultures were continued in FUdR to prevent dilution of the incorporated analogue by further division. After 3 days, the cultures from the FUdR-BUdR series showed the typical BUdR effect; the cells were excessively flattened and few multinucleated myotubes formed. Cells in the control cultures were of normal morphology, and multinucleated myotubes were present. These results were confirmed in another experiment in which BUdR-3H was added to 2-day cultures in which myotubes were forming. Fusion of thymidine-3H-labeled cells begins at 8 hr after the preceding S phase. In contrast, cells which incorporate BUdR-3H for one S period do not fuse with normal myotubes.

In vitro induction of granulocyte differentiation in hematopoietic cells from leukemic and non-leukemic patients.

Abstract: Human spleen-conditioned medium can induce the formation in vitro of large granulocyte colonies from normal human bone marrow cells. The granulocyte colonies contained cells in various stages of differentiation, from myeloblasts to mature neutrophile granulocytes. Human spleen-conditioned medium also induced colony formation with rodent bone-marrow cells, whereas rodent spleen-conditioned medium induced colony formation with rodent bone marrow but not with human cells. This in vitro system has been used to determine the potentialities for cell differentiation in bone-marrow and peripheral blood cells from patients with a block in granulocyte differentiation in vivo. The cloning efficiency, colony size, and number of mature granulocytes in bone-marrow colonies from patients with congential neutropenia, whose bone marrow contained only 1% mature granulocytes, were not less than in people whose bone marrow had the normal level of about 40% mature granulocytes. The cloning efficiency of peripheral blood cells from patients with acute myeloid leukemia was 350 times higher, with 10 times larger colonies, than the cloning efficiency of peripheral blood cells from normal people. The cytochemical properties and number of mature granulocytes in colonies from the leukemic patients were the same as in colonies from non-leukemic people. The results indicate that a block in cell differentiation in vivo, in these cases with neutropenia and acute myeloid leukemia, was overcome in vitro, in the presence of an inducer in the conditioned medium. In patients with chronic myeloid leukemia, colony formation was induced only in some of the cases. This indicates that there are blast cells with different potentialities for the development of colonies in different patients.

5-Bromodeoxyuridine-Induced Differentiation of a Neuroblastoma

Abstract: 5-Bromodeoxyuridine induces the differentiation of mouse neuroblastoma C1300 to cells that morphologically resemble mature neurons. The induced differentiation can take place in the absence of DNA synthesis. This suggests that the halogenated pyrimidine need not be incorporated into DNA to alter the phenotype of the cell.

Regulation of Acetylcholinesterase in Neuroblastoma Cells

Abstract: The specific activity of mouse neuroblastoma acetylcholinesterase (EC 3.1.1.7) increased 25-fold when the rate of cell division was restricted. The results show that acetylcholinesterase activity is regulated in neuroblastoma cells and that the regulatory mechanism is inversely related to the rate of cell division. Under the same conditions the specific activity of catechol-O-methyl transferase (EC 2.1.1.6) did not change significantly.

Studies on the differentiation of thymus-derived lymphocytes

Abstract: The development pathway from embryonic thymus-stem cell to peripheral thymus-derived lymphocyte has been demonstrated using the alloantigens θ (theta) and TL as surface markers of cell differentiation. On the basis of cytotoxicity tests carried out on CBA.H or A embryo thymus cultured in diffusion chambers and on CBA.H embryo thymus grafts and peripheral lymphocytes derived from them in AKR hosts, it has been concluded that two differentiation stages take place during the maturation of thymus-derived cells, namely a first step from stem cell to thymocyte and a second step from thymocyte to peripheral lymphocyte.

Cellular differentiation in skeletal tissues

Abstract: CONTENTS

Formation of horny cells: The fate of cell organelles and differentiation products in ruminal epithelium

Abstract: Epithelial cells changing from the granular stage of differentiation to the horny stage are more numerous, and reveal sequential events of transformation in finer detail in the rumen epithelium than in other keratinizing epithelia thus far studied in the electron microscope. Studies of such cells indicate that transformation is initiated by the release of hydrolytic enzymes, as evidenced by the appearance of lysosomes. As lysosomes increase in number, the nucleus, ribosomes, mitochondria, Golgi apparatus, and mucous granules are gradually degraded. Furthermore, marked changes occur in permeability of the plasma membrane as voluminous amounts of the lysed cell components pass through and accumulate in the intercellular space in the form of an amorphous mass. Filaments, keratohyalin granules, and the content of the ER (ER-protein) are not lysed, revealing the action of released enzymes to be specific. During transformation, filaments become displaced toward the cell periphery and keratohyalin granules disperse and mix with the ER-protein in the cell center. Subsequently, the keratohyalin-ER-protein complex infiltrates the filament network whereby a fibrous-amorphous cell content is formed. Loss of fluids through the plasma membrane leads to reduction of cell volume and consolidation of the remaining cell content. The deep interdigitations formed between the cells ultimately interlock the outer part of the epithelium into a cohesive and protective stratum corneum.

Transfer RNA and cell differentiation.

Abstract: Publisher Summary This chapter explores that cell differentiation involves drastic changes in cell metabolism. Thus, the synthesis of many proteins is stopped and that of new ones has begun. This type of regulation could be called macroregulation in contrast to microregulation, in which the synthesis of a particular protein, or set of proteins in an operon, is specifically regulated by induction and repression. The chapter deals with the possibility of tRNA involvement as a critical factor in cell differentiation at the macroregulation level. Various mechanisms for macroregulation can be visualized. The translation of mRNA into protein involves various components, such as ribosomes, initiation and termination factors, chain elongation factors, tRNA and aminoacyl-tRNA synthetase. Among these components for code translation, tRNA is the one most likely to be involved in macroregulation because of the unique feature of being multiple for each amino acid and yet ubiquitous for the synthesis of various proteins of the cell. Controls are likely to be operating at both levels. Degeneracy of the code and degeneracy of corresponding adaptors (tRNA) have been established. On the other hand, there is generally a single aminoacyl-tRNA synthetase for each amino acid in bacteria, although, some contradictory results have been reported. In higher organisms, the mitochondria1 system seems to have its own synthetases and tRNA's in addition to cytoplasmic synthetases and tRNA's.

Induction of Glutamine Synthetase in Embryonic Retina: Its Dependence on Cell Interactions

Abstract: A relation between enzyme induction in embryonic cells and cellular organization is indicated by the finding that the levels of glutamine synthetase induced by hydrocortisone in the embryonic neural retina in vitro are dependent on the associations between the retina cells. Intact retina tissue, aggregates of dissociated cells, and cells in monolayer culture showed a decreasing response, in this order, to glutamine synthetase induction. With time of culture, the enzyme activity continued to rise in the intact retina and in cell aggregates, but activity declined in monolayer cultures even though the inducer was continuously present. Dispersed cells cultured in monolayer without the inducer showed after 24 hours a loss of inducibility which could not be reversed by reaggregating such modified cells but could be prevented by maintaining the freshly dispersed cells at a low temperature.

Influence of ionic conditions on cell differentiation and morphogenesis of the cellular slime molds.

Abstract: Effects of various ionic conditions on the development of the cellular slime molds D. discoideum and D. mucoroides were studied. A certain concentration of lithium ions (7 mM) promoted differentiation of the stalk cells and conversely inhibited formation of the spores. The presence of calcium and magnesium ions was needed for Li to manifest its specific effect. A high concentration of Ca (100-120 mM) also facilitated differentiation of the stalk cells. On the other hand, fluoride ions stimulated considerable formation of spores at 15 mM. In the absence of divalent cations, sodium ions inhibited morphogenesis and cell differentiation proportionately with its concentration, and complete inhibition was obtained at 20 mM. The inhibitory effect of Na was nullified by addition of small amounts of Ca. Possible mechanisms by which these ions exert their influences on development of this organism were discussed.

Cellular differentiation in the thymus iii. surface properties of rat thymus and lymph node cells separated on density gradients

Abstract: Young adult rat thymus and lymph node cell subpopulations were obtained by differential flotation on discontinuous BSA density gradients and assayed for properties characteristic of mature thymus-derived lymphocytes. One such subpopulation (C) of thymocytes was enriched in its ability to respond mitotically to a hemiallogeneic MLR stimulus, to localize in the parenchyma of lymph nodes and spleen, and to initiate a GVH reaction in a suitable host. These cells did not respond well to mitotic stimulation by PHA, they were lighter in density than the majority of mature lymph node thymus-derived lymphocytes, and they possessed a thymus-specific antigen (RTA) not present on peripheral lymphoid cells. We conclude that the acquisition of peripheral properties occurs sequentially, during an intrathymic differentiation cycle or shortly after the cells leave the thymus.

Proliferative behavior of differentiating cells in the developing rat parotid gland.

Abstract: Parotid glands of litters of rats at age intervals from 20 days in utero to 100 days were assayed for DNA content and examined by light- and electron-microscopy. The age differences in total DNA and DNA concentration indicated that there was a rapid rate of proliferation of parenchymal cells until 25 postnatal days, after which the rate declined rapidly, and that there was a rapid increase in cell size between 18 and 25 days. These findings were substantiated by histologic observations, such as the presence of numerous mitotic figures until 25 days of age, and the rapid maturation of the acinar cells between 18 and 25 days. These data suggest that the acinar cells of the rat parotid gland comprise an expanding cell population. Light- and electron-microscopic observations consistently indicated that cells with mitotic figures were about as well differentiated as other parenchymal cells at all stages of gland development, including mature acinar cells observed at ages 23 and 25 days. These observations support the view that the division of cells in advanced stages of differentiation may be important in the growth of certain organs and tissues.

CEREBELLAR GRANULE CELLS IN VITRO: A Light and Electron Microscope Study

Abstract: The behavior of granule cells in mature cerebellar cultures derived from newborn mice was studied by light and electron microscopy. Many granule cells remained in the explants as an external granular layer. These cells were differentiated, as evidenced by formation of bundles of parallel fibers and by development of synapses between granule cell axons and Purkinje cell branchlet spines, and between Golgi cell axons and granule cell dendrites. Although the over-all architecture of the cerebellar explants after 18–33 days in vitro was similar to that of the newborn mouse, the evident differentiation of the granule cells suggested that interneuronal relationships resemble those of the mature cerebellum in vivo.

Chapter 2 On the Long-Term Control of Nuclear Activity During Cell Differentiation

Abstract: Publisher Summary The control of gene activity during development has proved very hard to approach experimentally, but certain techniques have been developed that have made fruitful investigation possible. This chapter emphasizes one of this microinjection into living cells. Three general ways have been recognized in which the activity of nuclear genes may be regulated in multicellular organisms: (1) by changes in the composition of chromosomal DNA; (2) by long-term gene repression, in which some genes are committed to inactivity for long periods of time, like the length of a cell cycle; And (3) by the short-term adjustment of gene activity in response to the appropriate changes of intra- and extracellular conditions. It is mainly concerned with regulation of the second kind, which is believed to be characteristic of differentiated cells. The chapter summarizes some of the evidence that has demonstrated the stability of the differentiated state and of nuclear activity in differentiated cells. It considers that the basis of this stability is unlikely to lie in the altered composition of chromosomal DNA. It also draws attention to certain conditions under which apparently stable nuclear expression is changed. One of these is nuclear transplantation that results in a complete “reprogramming” of nuclear activity. The chapter is concerned with the cellular and molecular events which accompany this reprogramming. As the cytoplasmic proteins that enter transplanted nuclei may well be important in determining the nature of their subsequent activity, the chromosomal proteins of normal cells are discussed.

Studies on thymus function. I. Cooperative effect of thymic function and lymphohemopoietic cells in restoration of neonatally thymectomized mice.

Abstract: Immunological restoration of 45-day old, neonatally thymectomized C3Hf mice by treatment with humoral thymic function (thymoma grafts, thymus or thymoma in diffusion chambers) ranges from 0 to 12% and is difficult to acheive. When small numbers (5–20 x 106) of young adult lymphohemopoietic cells, ineffective by themselves, are given in association with humoral thymic function, a cooperative effect is observed and restoration ranges from 30 to 60%. With a particular cell dosage (20 x 106), effectivity for cooperation with thymic function was the following in decreasing order: spleen, lymph nodes, thoracic duct cells, bone marrow, blood leukocytes, thymus, and Peyer's patch cells. Comparable results were obtained using spleen, thymus, and hemopoietic liver from newborn donors in association with thymic function. For similar cell dosages, newborn thymus cells were more effective than adult thymus in their cooperative effect with thymic function. Dispersed thymus cells in association with young adult bone marrow or newborn hemopoietic liver cells showed no synergism for the cooperative effect with thymic function in the present model. Using hemiallogeneic cells (F1 hybrid into parent) it was possible to show that restoration was mediated by proliferative expansion of the injected cells. This was indicated by specific tolerance to tissues of the other parental strain and by cellular chimerism, especially of lymphoid tissues, as indicated by chromosome markers and absence of significant numbers of immunocompetent cells of host origin. A population of paritally differentiated cells of hemopoietic origin, termed postthymic, sensitive to humoral activity of the thymus and present in the lymphohemopoietic tissues of adult and newborn mice is postulated to explain our results. These cells are postthymic and thymus dependent in the sense that they already received thymic influence, probably through traffic, and are incapable of self-renewal in absence of the thymus. Sensitivity to humoral activity of the thymus is characterized by proliferative expansion and/or a differentiative process eventually leading to larger numbers of competent cells.

Studies on antibody-producing cells. I. Ultrastructure of 19S and 7S antibody-producing cells.

Abstract: Antibody-bearing cells of spleen and lymph node of the mouse and rabbit detected by rosette formation with the antigenic red blood cells were collected by micropipet and studied by electron microscopy. More than 300 such cells were examined. In the lymph nodes, rosette-forming cells were all in the lymphocytic and plasmacytic categories. In cells of the mouse spleen, macrophages were also found among the RFC, especially in the later days after immunization. The great majority of the RFC, 70–100%, were of the lymphocytic category. These included small, medium, and large lymphocytes with fine gradations of differentiation, and blast forms with little heterochromatin. The endoplasmic reticulum of these cells occurred in short, very narrow pieces, usually in contact with a mitochondrion. The cells of the plasmacytic category also showed fine gradations from plasmablasts to typical mature plasma cells. Plaque-forming cells of mouse and rabbit were also collected by micropipet. Of 162 such cells, fine gradations were also found throughout the lymphocytic and plasmacytic categories, but in this case the great majority were in the plasmacytic group, and more plasma cells showed amorphous nuclear chromatin. Among antibody-forming cells detected by both reactions, some of the more highly differentiated large lymphocytes contained ER which differed from that in the other large lymphocytes in that the channels were slightly and variably distended, with deposition of some precipitate, and with some tendency to a more nearly parallel orientation of the few channels seen. These were considered transitional cells. Of 10 RFC found in mitosis, all were in the lymphocytic category, in various stages of differentiation, the most advanced of which (in 2 of the 10 cells) was that of the transitional lymphocyte described here. Cells producing plaques facilitated by antisera vs. IgG of the mouse or rabbit (7S) showed the same distribution between cell categories and the same fine gradations as the direct (19S) PFC. Cells producing rosettes which were resistant to lysis in the presence of complement, and were thus presumably producing 7S antibody, showed a distribution similar to that found generally with rosette-forming cells, approximately 80–90% in the lymphocytic category.

Differentiation of epithelial cells in the mouse thymus.

Abstract: The foetal and post-natal development of the mouse thymus was studied with the electron microscope paying particular attention to the differentiation of the epithelial cells. At about 13 days' gestation, the thymus was composed principally of undifferentiated epithelial cells and some lymphoblasts. The latter accumulated rapidly but did not show much evidence of mitotic activity until after the development of differentiated “cortical” epithelial cells which appeared during the 15th day of gestation. Further differentiation of epithelial cells did not occur until near term when medullary “cystic” epithelial cells appeared, and post-natally when small Hassall's corpuscles were developed. Undifferentiated and dividing epithelial cells were seen in the medulla and were present in all postnatal animals examined.

Cellular and humoral aspects of the primary immune response of the toad, Bufo marinus.

Abstract: The primary immune response of the toad, Bufo marinus, to Salmonella adelaide flagella has been analysed in terms of three immunological parameters: (a) the kinetics of appearance and morphology of cells producing specific antibodies; (b) antigen retention in one of the lymphoid organs, the jugular bodies; and (c) the nature of the immunoglobulins produced. It was found that toads responded to the bacterial antigen by producing antibody-forming cells and serum antibody in quantities comparable to those observed in mammals. In the early phase of antibody production, small and medium lymphocytes played an important role in antibody synthesis. In the later stages of the response, large cells which resembled immature plasma cells were the predominant producers of antibody. Studies on the incorporation of tritiated thymidine suggested that the majority of antibody-forming cells observed during the logarithmic phase of the response arose by division. Electron microscopic studies showed that the localization of radioactively-labelled antigen in the jugular bodies was similar to the follicular localization in mammalian lymph nodes, i.e. the antigen was associated with the surfaces of cells, rather than with intracellular sites. Although the toad was found to possess distinct classes of 18S and 7S immunoglobulins, antibody activity to this antigen was present only in the 18S molecules. These results are consistent with the hypothesis that the major patterns of differentiation and proliferation of immunologically competent cells, antigen retention and immunoglobulin structure have emerged by the phylogenetic level of anuran amphibians.

The expression of differentiation by chick embryo thyroid in cell culture. I. Functional and fine structural stability in mass and clonal culture.

Abstract: The stability of glandular epithelial cells has been investigated utilizing the techniques of cell culture. Embryonic chick thyroid was chosen as a representative cell type and methods were developed for the successful clonal culture of thyroid follicular cells. Thyroid cells were found to be morphologically and functionally stable while undergoing rapid division in both dense (monolayer) and dilute (clonal) cell culture. Differentiated features were retained for a minimum of 32 days of primary clonal culture (approximately 17 generations under clonal conditions). During the culture period, the cells retained their epithelial morphology, retained their cytoplasmic rough endoplasmic reticulum, and continued to produce chromatographically detectable thyroid hormones. Hormone production in culture was a specific thyroid characteristic since control cultures of embryonic heart and liver did not contain the hormones.

Electron microscopic observations on cell proliferation and differentiation in the gastric mucosa of the mouse.

Abstract: After an injection of 3H-thymidine, the renewal of epithelial cells in the gastric mucosa of normal adult mice was studied by simple electron microscopy and electron microscopic autoradiography.1. Undifferentiated cells are present in the isthmus as a common matrix for surface mucous cells and mucous neck cells. Their differentiation to chief and parietal cells seems probable but could not be evidenced. Undifferentiated cells are proliferative in the isthmus.2. Immature surface mucous cells have mitotic activity in the isthmus and in deeper parts of the pit. They grow mature as they migrate towards the surface of the mucosa.3. Mucous neck cells, especially in their immature stage, are proliferative in the isthmus and in the neck. They migrate from the isthmus to deep parts of the neck.4. Primitive chief cells are first identified in the upper parts of the glandular base. They have mitotic activity and become mature with downwards migration.5. Parietal cells grow mature with their migration from the isthmus towards the base. Mitotic activity of parietal cells could not be confirmed.6. There were no finding suggesting the transformation from mucous neck cells to chief and parietal cells.7. Ultrastructural changes during the differentiation of each cell type were described.