Showing papers on "Cellular differentiation published in 1974"

Origin, differentiation and renewal of the four main epithelial cell types in the mouse small intestine. V. Unitarian Theory of the origin of the four epithelial cell types.

Abstract: The previous articles of this series provided presumptive evidence that the four main differentiated cell types in the epithelium of the mouse small intestine: villus columnar, mucous, entero-endocrine, and Paneth cells, originate from the same precursor, the crypt-base columnar cell. In the present work, direct evidence was obtained in support of this view. It was first found that crypt-base columnar cells phagocytose non-viable cells in their vicinity, with the result that a large phagosome appears in the cytoplasm. Such phagosomes were then used as markers to follow the evolution of crypt-base columnar cells. In normal control animals, a rare crypt-base columnar cell includes a large phagosome containing Paneth cell remnants. By six hours after injection of two μCi 3H-thymidine per g body weight, a fair number of crypt-base columnar cells include a different type of phagosome containing labeled nucleus and granulefree cytoplasm, which is attributed to phagocytosis of a labeled crypt-base columnar cell killed by beta-radiation from the incorporated 3H-thymidine. By 12 hours after 3H-thymidine injection, phagosomes have appeared in partly differentiated mid-crypt columnar cells and oligomucous cells; by 18–24 hours, in fully differentiated columnar cells and in Paneth cells; and by 30 hours, in an entero-endocrine cell. Since phagosomes are first found in crypt-base columnar cells and only later in the four differentiated cell types, it is concluded that crypt-base columnar cells transform into cells of these four types and, therefore, behave as the stem cells of the epithelium. The finding of rare epithelial cells containing two different types of secretory material (either mucous globules and entero-endocrine granules, or mucous globules and Paneth cell granules) confirms that the stem cells are multipotential. These findings support the Unitarian Theory of epithelial cell formation in the small intestine.

Stromal cells responsible for transferring the microenvironment of the hemopoietic tissues. Cloning in vitro and retransplantation in vivo.

Abstract: SUMMARYStromsil precursors ran bo detected in populations of homopoietic cells by their ability to form fibroblast colonies (clones) in monolayer cultures. The stable colony-forming efficiency is reached when the initial density of explanted bone marrow or spleen cells is not less than 104 cells/cm2

A Restriction Point for Control of Normal Animal Cell Proliferation

Abstract: This paper provides evidence that normal animal cells possess a unique regulatory mechanism to shift them between proliferative and quiescent states. Cells cease to increase in number under a diversity of suboptimal nutritional conditions, whereas a uniformity of metabolic changes follows these nutritional shifts. Evidence is given here that cells are put into the same quiescent state by each of these diverse blocks to proliferation and that cells escape at the same point in G(1) of the cell cycle when nutrition is restored. The name restriction point is proposed for the specific time in the cell cycle at which this critical release event occurs. The restriction point control is proposed to permit normal cells to retain viability by a shift to a minimal metabolism upon differentiation in vivo and in vitro when conditions are suboptimal for growth. Malignant cells are proposed to have lost their restriction point control. Hence, under very adverse conditions, as in the presence of antitumor agents, they stop randomly in their division cycle and die.

Neoplastic differentiation: characteristics of cell lines derived from a murine teratocarcinoma.

Abstract: Monolayer cultures of a mouse teratocarcinoma were established in vitro. These cultures contained embryonal carcinoma, the malignant stem cell, and its differentiated progeny: parietal yolk sac, neuroepithelial, and mesenchymal cells. Tissues such as squamous epithelium, cartilage, striated muscle, neuroepithelium, and glands were produced from embryonal carcinoma that was maintained under conditions of long term culture. Frequent subcultivation with pancreatin allowed the establishment of cell lines of embryonal carcinoma which have been maintained for more than 18 months in vitro and continue to produce differentiated cells under specific culture conditions. Chromosomally these lines of embryonal carcinoma have a stem line of 39 chromosomes. Two lines of parietal yolk sac cells have been established which produce basement membrane, are not tumorigenic, and chromosomally are hypotetraploid. This system may yield information concerning neoplastic differentiation and its possible use in therapy for cancer.

Origin, differentiation and renewal of the four main epithelial cell types in the mouse small intestine III. Entero‐endocrine cells

Abstract: The origin, differentiation and renewal of entero-endocrine cells was examined in the duodenum, jejunum and ileum of the mouse using light and electron microscopic radioautography after a single injection or continuous infusion of 3H-thymidine. When s-collidine buffered glutaraldehyde was used for fixation prior to electron microscopic study, all granules in all entero-endocrine cells were spheroidal and, therefore, their shape could not be used to classify the cells into subgroups, as done after fixation in phosphate buffered aldehyde. Thus, the cells were all considered as belonging in a single family. In the light microscope, mitosis is not observed in the entero-endocrine cells identified by iron hematoxylin staining. However, under the electron microscope, a few cells that contain a small number of characteristic granules, some filament bundles and many free ribosomes are in division or are labeled one hour after an injection of 3H-thymidine. These cells are interpreted as young entero-endocrine cells. They are located in the crypt base. They resemble crypt-base columnar cells and are, therefore, suspected of arising from them. Differentiation may be examined by following the fate of the young enteroendocrine cells which are labeled by 3H-thymidine. Within the crypts, these cells acquire a gradually increasing number of granules while losing the ability to divide. The few granules initially present usually have a particulate content and may include a small dense core; but, as differentiation proceeds and granules accumulate, their content is mostly dense and homogeneous. The differentiation of precursor cells into mature entero-endocrine cells takes about two days. Meanwhile, in the same manner as columnar and mucous cells, enteroendocrine cells migrate up the crypt, reach the villus and are lost at the extrusion zone. The turnover time of entero-endocrine cells is estimated to be 3.9 days in duodenum and 4.0 days in jejunum.

Clonal selection, attenuation and differentiation in an in vitro model of hyperplasia.

Abstract: Observations of the growth kinetics and morphologies of clones and subclones of diploid human skin fibroblast cultures lead to the working hypothesis that these cells, presumably like their counterparts in healing wounds, constitute a differentiating system. There is attenuation of the growth of serial clones, with continual selection for more vigorous stem cells. The latter segregate daughter cells of varying growth potential, including a class of cells which may be regarded as terminally differentiating; we propose that such cells may be histiocytes or macrophages. These studies a) demonstrate extensive epigenetic heterogeneity in fibroblast cultures, b) suggest that hyperplastic foci may be monoclonal or oligoclonal, c) rule out a simple biologic clock mechanism as an explanation of clonal senescence, d) suggest a new approach to the analysis of various inborn errors of metabolism, such as Werner's Syndrome.

Cell cycle kinetics and development of Hydra attenuata. III. Nerve and nematocyte differentiation.

Abstract: The differentiation of nerve cells and nematocytes in Hydra attenuata has been investigated by labelling interstitial cell precursors with [3H]thymidine and following by autoradiography the appearance of labelled, newly differentiated cells. Nematocyte differentiation occurs only in the gastric region where labelled nematoblasts appear 12 h and labelled nematocytes 72-96 h after addition of [3H]thymidine. Labelled nerves appear in hypostome, gastric region, and basal disk about 18 h after addition of [3H]thymidine. The lag in the appearance of labelled cells includes cell division of the precursor as well as differentiation since nerves and nematocytes have 2n postmitotic nuclear DNA content. A cell flow model is proposed for interstitial cells and their differentiated products. Stem cells occur as single interstitial cells or in pairs. Per cell generation about 60 % of the daughter cells of stem cell divisions remain stem cells and about 40 % differentiate nerves and nematocytes. Nerves differentiate directly from stem cells in about 1 day. Nematocyte differentiation requires 5-7 days including proliferation of a cluster of 4, 8, 16 or 32 interstitial cells and differentiation of a nematocyst capsule in each cell. The numbers of interstitial cells and nematoblasts predicted by the cell flow model from the rates of nerve differentiation (900 nerves/day/ hydra), nematocyte differentiation (1760 nematocyte nests/day/hydra) and stem cell proliferation (stem cell cycle = 24 h), agree with the numbers of these cells observed in hydra. The number of stem cells per hydra is 3000-6000 depending on assumptions about the time of determination. The ratio of nematocyte to nerve differentiation averaged over the whole hydra is 3:1. In the hypostome and basal disk interstitial cell differentiation occurs exclusively to nerve cells while in the gastric region the ratio of nematocyte to nerve differentiation is about 7:1.

Cell cycle kinetics and development of hydra attenuata

Abstract: The differentiation of nerve cells and nematocytes in Hydra attenuata has been investigated by labelling interstitial cell precursors with PHJthymidine and following by autoradiography the appearance of labelled, newly differentiated cells. Nematocyte differentiation occurs only in the gastric region where labelled nematoblasts appear 12 h and labelled nematocytes 72—96 h after addition of 2H thymidine. Labelled nerves appear in hypostome, gastric region, and basal disk about 18 h after addition of 3H thymidine. The lag in the appearance of labelled cells includes cell division of the precursor as well as differentiation since nerves and nematocytes have 2JI postmitotic nuclear DNA content. A cell flow model is proposed for interstitial cells and their differentiated products. Stem cells occur as single interstitial cells or in pairs. Per cell generation about 60 % of the daughter cells of stem cell divisions remain stem cells and about 40 % differentiate nerves and nematocytes. Nerves differentiate directly from stem cells in about 1 day. Nematocyte differentiation requires 5-7 days including proliferation of a cluster of 4, 8, 16 or 32 interstitial cells and differentiation of a nematocyst capsule in each cell. The numbers of interstitial cells and nematoblasts predicted by the cell flow model from the rates of nerve differentiation (900 nerves/day/ hydra), nematocyte differentiation (1760 nematocyte nests/day/hydia) and stem cell proliferation (stem cell cycle = 24 h), agree with the numbers of these cells observed in hydra. The number of stem cells per hydra is 3000-6000 depending on assumptions about the time of determination. The ratio of nematocyte to nerve differentiation averaged over the whole hydra is 3:1. In the hypostome and basal disk interstitial cell differentiation occurs exclusively to nerve cells while in the gastric region the ratio of nematocyte to nerve differentiation is about 7:1.

Different blocks in the differentiation of myeloid leukemic cells.

Abstract: Some clones of mouse myeloid leukemic cells (D+) can be induced to undergo cell differentiation to mature macrophages and granulocytes, and other clones (D-) could not be induced to differentiate to mature cells. Normal mature macrophages and granulocytes have surface receptors that form rosettes with erythrocytes coated with specific immunoglobulin or immunoglobulin-complement. The D+ clones were induced to form receptors by prednisolone, cytosine-arabinoside, 5-iododeoxyuridine, actinomycin D, or serum from mice injected with endotoxin. All these compounds thus induced a common change in the cell surface membrane. The induction of receptors required protein synthesis, and receptors were formed before the appearance of mature cells. There were two types of D- clones. One type was induced by these compounds to form receptors, although with a lower inducibility than D+ clones; in the other type there was no induction of receptors. The results indicate that there are different blocks in the differentiation of myeloid leukemic cells. Some leukemic cells (IR+D+) can be induced to form receptors and to differentiate to mature cells; others (IR+D-) can form receptors but not mature cells; and a third type (IR-D-) could not be induced to form receptors or mature cells.

Neoplasms, differentiations and mutations.

Abstract: Evidence has been presented to support the concept that malignant tumors are postembryonic differentiations superimposed upon the process of tissue maintenance and renewal. Malignant stem cells are derived from normal stem cells. They have a capacity for proliferation and differentiation that operates at a different level of control than the normal. Even so, malignant stem cells are responsive to enviornmental control, suggesting that it may be possible to direct their differentiation or at least to control their ability to replicate. A tumor is a caricature of normal tissue and appears undifferentiated because of the preponderance of undifferentiated proliferating stem cells in relationship to the number of cells that have differentiated and become benign.

Some aspects of the development of the notochord in mouse embryos

Abstract: Prenotochordal cells are derived by proliferation from the pluripotential ectoderm in the Hensen9s node area, and they migrate in the cephalic direction to be incorporated into the roof of the archenteron. Later, after separation from the archenteron, the notochord probably includes some endodermal cells. Topographically it occupies an intermediate position between the ectoderm and endoderm. In this way it assumes a mesodermal-like character. After separation from the archenteron, the differentiation of the organ and its cells follows a craniocaudal gradient. At the time of separation the basal lamina, originally covering the dorsal side of both the prenotochordal and endodermal cells, remains continuous between the endoderm and the notochord. Its presence from the earliest stages of morphogenesis of the notochord presumably indicates that it plays an important formative role. The cells of Hensen9s node contain numerous microtubules, centrioles and cilia. In later stages fibrillogenesis in the prenotochordal and notochordal cells and excortication (ecdysis) occur. The intracellular fibrillar material in the notochordal cells may first form the basal lamina and later the perichordal sheath. In 12 to 13-day-old embryos, during excortication of the fibrillar material, the basal lamina undergoes disruption and eventually disappears. From the 10th day onwards, the outer plasma membrane shows an active endocytosis, by the formation of micropinocytic vesicles. Vacuolization occurs in the later stages of morphogenesis and it is suggested that this is due to dilation of endoplasmic reticulum and mitochondria. There is no evidence of a syncytial stage in the development of the notochord in mouse embryos.

Induction of T-cell differentiation in vitro by thymin, a purified polypeptide hormone of the thymus.

Abstract: Thymin, a purified polypeptide isolated from bovine thymus, was shown to induce the expression of differentiation antigens characteristic of thymocytes [TL and Thy-1 (θ)] when incubated in vitro with mouse bone marrow or spleen cells This induction occurred in 5-10% of the cells from bone marrow after a 2-hr incubation with subnanogram concentrations of thymin The induced cells expressed more TL and Thy-1 (θ) antigens than average normal thymocytes

Growth, Degrowth, and Irreversible Cell Differentiation in Aurelia aurita

Abstract: Growth patterns of the Scyphomedusa Aurelia aurita from Tomales Bay. California, were examined in the field and in the laboratory. Manipulation of growth patterns demonstrated that degrowth and regrowth are not constrained by initial ievelopmental stage. Although initial degrowth of certain tissues is allometric (e.g., gonads regress in 5 to 8 days; bell diameter decreases more rapidly at first than do the oral arms), thereafter regression appears identical to, but reversed from normal growth. Regrowth patterns are normal. Sexual maturation in the sea does not always alter subsequent capacity for degrowth or regrowth to sexual maturity in the laboratory, because reproductive and somatic tissues do not always degenerate after spawning. Gonadal tissue can be renewed and maintained in a ripe condition in the laboratory apparently indefinitely. Sexual maturation is a size-dependent phenomenon, not an agespecific developmental event. Spermatogenesis, once initiated, proceeds irrespective of outside events. Labeled spermatogonial cells can continue to differentiate to form sperm even though the gonad containing those cells, and the animal itself, show rapid degrowth. The importance of this decoupling of developmental events is discussed. The experimental importance of animals with flexible life cycles is emphasized.

Induction, Detection and Characterization of Cell Differentiation Mutants in Drosophila

Abstract: Mitotic recombination has been used as a tool for detecting cell differentiation mutants in clones of heterozygous individuals. With this method, previously mutagenized (ethyl methanesulfonate) genomes can be screened for mutants, induced anywhere in the genome, in a first generation, and irrespectively of being lethal themselves or present ina lethal chromosome. Induced mitotic recombination was used again in order to give the chromo-some arm location and recombinational locus.

Cell population changes during acinus formation in the postnatal rat submandibular gland

Abstract: The postnatal differentiation of acini in the submandibular glands of 2–42 day old rats given 3H-thymidine was studied by using radioautographs prepared from Epon-embedded, PAS and iron hematoxylin stained sections. The changes in morphology, population size and proliferative activity of various cell types in the gland were analyzed. At two days of age, rudimentary secretory units, designated as terminal tubules, were located at the end of the duct system and consisted of three cell types: (1) terminal tubule cells (30.7%) with darkly-stained granules, (2) proacinar cells (23.6%) with large, lightly-stained granules, and (3) acinar cells (1.6%) with PAS-positive granules. The proacinar cells, which underwent mitosis, disappeared within the first two weeks of life. The terminal tubule cells increased in number between 2 and 14 days of age, but became less numerous thereafter and disappeared by six weeks. Concomitantly, the number of acinar cells increased linearly with age and at a much greater rate than that of intercalated duct cells. Yet the rate of proliferation of acinar cells was comparable to that of intercalated duct cells. The overall proliferative activity in the gland decreased with age, and was inversely correlated with the relative frequency of acinar cells in the gland. On the basis of above data, it is postulated that, during the formation of acini from terminal tubules, acinar cells have a dual origin: they arise from proacinar cells during the first one to two weeks and from terminal tubule cells between two and six weeks of age.

Differentiation of B cells: sequential appearance of responsiveness to polyclonal activators.

Abstract: The responses to the polyclonal B-cell activators dextran sulphate (DxS), lipo polysaccharide (LPS), and purified protein derivative from tuberculin (PPD) were followed along the differentiation process of fetal liver cells to mature B cells in irradiated hosts It was shown that these cells sequentially gained responsiveness to DxS. LPS, and PPD, in that order. It was also shown that the result of activation depends only on the differentiation degree of the target cells at the turn they are activated. Thus, more primitive cells can only divide, and most of the DxS-. LPS-, or PPD-induced blasts are lg-negative soon after the cells become responsive to each ligand. The response of more differentiated cells is characterized also by high-rate antibody synthesis. These results provide a unique possibility of using polyclonal B-cell activators as well-defined functional markers for sub-populations of B cells and shed a new light on the problem of immune B-cell triggering

Regeneration of rat tracheal epithelium after mechanical injury. I. The relationship between mitotic activity and cellular differentiation.

Abstract: SummaryThe time of appearance of cytologic specialization in cells of regenerating rat tracheal epithelium was compared to the time of maximal mitotic response. Trauma sufficient to lethally injure most cells reaching the surface but sparing most basal cells resulted in a peak of mitotic activity from the 26th to the 30th hour following injury. The cells in the hyperplastic regenerating epithelium at the peak of mitosis had high nuclear-cytoplasmic ratios with little cytoplasmic or surface specialization. Ciliogenesis and mucinogen formation were not. seen in most cells until the 60th hr after the injury and there was little cell division at that time or later. Since cessation of mitosis does not follow or even coincide with the replenishment of the ciliated or goblet cell populations, the data does not support the hypothesis that control of cell division resides in the elaboration or release of a suppressor substance by the mature cells.

Nerve Growth Factor in Rat Glioma Cells

Abstract: We have shown that rat glioma tumors contain a protein that crossreacts with antibody against mouse 2.5S nerve growth factor prepared in rabbit in microcomplement fixation assays and has analogous isoelectric points to all the hybrids of 2.5S nerve growth factor (a dimer). Partially purified protein preparations from gliomas cause chick dorsal root ganglia to extend neurites in the nerve growth factor assay and cause morphological differentiation of mouse neuroblastoma cells. We conclude that the rat glioma protein is homologous to mouse salivary nerve growth factor and suggest the possibility that glial nerve growth factor plays a role in neuronal development and regeneration.

Erythropoietic Differentiation in Colonies of Cells Transformed by Friend Virus

Abstract: Cells transformed by Friend virus in liquid suspension culture respond to low concentrations of dimethylsulfoxide by initiating hemoglobin synthesis. The kinetics of appearance of such differentiated erythroid cells is consistent with either the induction of differentiation in a uniformly susceptible population of transformed cells or selection for the growth of a distinct erythropoietic subpopulation. The dimethylsulfoxide response of individual colonies of cells transformed by Friend virus grown in semisolid medium was studied in order to distinguish between these alternatives. The data do not support a selective effect of dimethylsulfoxide on the growth of a unique erythropoietic subpopulation; they indicate, rather, that the 745A strain of cells transformed by Friend virus consists of a relatively uniform population of dimethylsulfoxide-sensitive erythropoietic cells.

An ultrastructural study of sex differentiation in the teleost Oryzias latipes

Abstract: Electron-microscopic observations on sex differentiation during normal development in the medaka, Oryzias latipes , are presented in this report. Primordial germ cells in embryos 6–10 days after fertilization are clearly distinguishable from somatic cells by the presence in the cytoplasm of the former of germinal dense bodies, closely associated with large aggregations of mitochondria. The gonadal primordium is composed of the primordial germ cells and the enveloping somatic cells; no special ultrastructural relationships have been detected between these two cell types. The sex of an embryo cannot be decided by means of the electron microscope. The newly formed ovary is distinguishable in newly hatched fry of about 5 mm total body length. In the ovary, a layer of cells forming the ovarian wall encloses the ovarian matrix, consisting of oogonia and their surrounding follicle cells. There are often observable desmosomes between two neighbouring follicle cells. Although marked rearrangement of somatic cells takes place during sex differentiation of the gonad, few visible changes are observed in the ultrastructure of the primordial germ cells during their transition to oogonia. The transition from the oogonium to the oocyte is, however, characterized by a distinctive change in the nucleus, associated with the onset of meiosis. In the young ovary of 5-day-old fry, ooctye chromatin is visibly organized into electron-dense axial elements at the leptotene stage of meiotic prophase. But in the ovary of 10-day-old fry (about 6 mm body length), many oocytes at zygotene or pachytene stages are found, with synaptonemal complex configurations essentially the same as those described in numerous other meiosing plant and animal cells. In contrast, the testis of newly hatched male fry remains in an undifferentiated state, with the somatic cells simply enveloping the spermatogonia. No cells with the ultrastructural characteristics of Leydig cells (as observed in adult testis) can be distinguished in the young ovary or in the undifferentiated testis 10 days after hatching. Cells with the ultrastructural characteristics of adult Leydig cells are detectable for the first time in the matrix of the testis of 25-day-old young, about 8 mm in total length. When proliferation of spermatogonia has occurred, to form the typical testis of 45-day-old (about 15 mm), young, these cells appear in the interstitial region of the testis. Observations from the present study indicate that sex differentiation of germ cells and somatic cells in the gonad precedes the differentiation of steroid-secreting cells. Therefore, the hypothesis that sex hormones are natural sex-inducers is not supported by the present results. The possibility is emphasized that some intracellular mechanisms may be involved in the natural course of sex differentiation of the germ cells and the gonad in this fish.