Showing papers on "Transdifferentiation published in 1992"

Rapid acinar to ductal transdifferentiation in cultured human exocrine pancreas

Abstract: Experiments have been performed to define conditions for the primary culture of human exocrine pancreas, as a first step towards molecular reconstruction experiments of pancreatic neoplasia. Normal human exocrine pancreas was digested using collagenase and dispase and the resulting cellular aggregates were cultured in vitro. The phenotype of the digested pancreatic cells was almost exclusively acinar (amylase-positive, keratin 19 and mucin antigens-negative), yet within 4 days of culture the cells had taken on a ductal phenotype (amylase-negative, keratin 19 and mucin antigens-positive). The kinetics of these observations exclude the possibility of overgrowth of the acinar population by a ductal sub-population, and selective adherence is excluded by examination of those cells that do not adhere, which are representative of the initiating population. We interpret these data as indicating that, under the conditions of culture, the acinar cell phenotype is not stable and can transdifferentiate to a ductal phenotype. Taken together with recent data from transgenic animals, this in vitro observation has possible implications for our view of the pathogenesis of pancreatic neoplasia.

Phenotype‐specific “tissue” transglutaminase regulation in human neuroblastoma cells in response to retinoic acid: Correlation with cell death by apoptosis

Abstract: Neuroblastomas in culture are characterized by the presence of 2 morphologically and biochemically distinct phenotypes (i.e., neural "N-type" and flat substrate-adherent "S-type") which undergo transdifferentiation. Human neuroblastoma SK-N-BE(2) cells differentiate toward a neural phenotype upon retinoic acid (RA) treatment. However, we recently showed that, during the RA treatment, a subset of SK-N-BE(2) cells undergo apoptosis; these cells specifically express a high "tissue" transglutaminase (tTG) level. This study was undertaken to investigate the cellular and molecular basis of the action of retinoic acid on apoptosis in human neuroblastoma cells. As a biochemical marker of the phenomenon we studied the tTG gene expression in the parental line SK-N-BE(2) and in 2 clones which stably express neuroblastic [BE(2)-M17] and substrate-adherent [BE(2)-C] features, respectively. Data showed a differential phenotype-specific regulation of tTG gene expression. In fact, RA treatment enhanced tTG expression and apoptotic index in the flat substrate-adherent variant, whereas, in cells expressing the neural phenotype, very low tTG expression and apoptosis were found. Northern-blotting analysis revealed that the substrate-adherent cells had a basal 3-fold higher level of tTG mRNA. An increase in tTG mRNA major transcript levels (3.7 kb) occurred within a few hours of exposure to RA in both the phenotypic variants. By contrast, tTG protein level was very low in the cell expressing the neuronal phenotype, even after prolonged exposure to RA. Immunohistochemical analysis indicated that tTG protein, in addition to mature apoptotic cells, was specifically localized in the flat substrate-adherent variant both in the wild-type and in the BE(2)-C clone. These findings suggest that the ability to undergo apoptosis in the neuroblastoma cells is associated with the expression of a non-neuronal neuroectodermal (substrate-adherent cells) immature phenotype.

Transdifferentiation in Medusae

Abstract: Publisher Summary This chapter discusses the in vitro transdifferentiation system that is established using isolated medusa tissues of differentiated cells. Transdifferentiation can be defined as a switch (or reprogramming or rechanneling) of cells that have already expressed specific differentiated traits into another cell type distinguished from the original cells by a set of phenotypic characters. In all known cases of true transdifferentiation the newly acquired state is stable and a switch to the original cell type has never been reported. Medusae (jellyfish) belong to Cnidaria , a group of animals that lack a true mesoderm and are essentially classified as bilayered animals. They have two properties that have been of crucial importance for the establishment of an experimental transdifferentiation system: (1) many species are able to regenerate most of their body parts, and (2) the swimming organ (the umbrella or bell) is composed of three tissue layers, of which two consist of only one well-differentiated cell type. The transdifferentiation processes observed in medusa demonstrate that well-differentiated animal cells can retain the ability to change cellular commitment and to form various new cell types that can be organized into a functional regenerate.

Progressive stages of "transdifferentiation" from epidermal to mesenchymal phenotype induced by MyoD1 transfection, 5-aza-2'-deoxycytidine treatment, and selection for reduced cell attachment in the human keratinocyte line HaCaT.

Abstract: The ability of the myogenic determination gene (MyoD1) to convert differentiating human keratinocytes (HaCaT cell-line) to the myogenic pathway and the effect of MyoD1 on the epidermal phenotype was studied in culture and in surface transplants on nude mice MyoD1 transfection induced the synthesis of myosin, desmin, and vimentin without substantially altering the epidermal differentiation properties (morphology, keratin profile) in vitro nor epidermal morphogenesis (formation of a complex stratified squamous epithelium) in surface transplants, demonstrating the stability of the keratinocyte phenotype 5-Aza-CdR treatment of these MyoD1-transfected cells had little effect on the cultured cells but a morphologically unstructured epithelium was formed with no indications of typical cell layers including cornification Since prevention of epidermal strata in transplants was not accompanied by blocked epidermal differentiation markers (keratins K1 and K10, involucrin, and filaggrin), the dissociation of morphogenesis and expression of these markers argues for independently controlled processes A subpopulation of less adhesive cells, isolated from the 5-aza-CdR treated MyoD1-transfectants, had lost most epithelial characteristics in culture (epidermal keratins, desmosomal proteins, and surface-glycoprotein Gp90) and had shifted to a mesenchymal/myogenic phenotype (fibroblastic morphology, transactivation of Myf3 and myogenin, expression of myosin, desmin, vimentin, and Gp130) Moreover, the cells had lost the ability to stratify and remained as a monolayer of flat elongated cells in transplants These subsequent changes from a fully differentiated keratinocyte to a mesenchymal/myogenic phenotype strongly argue for a complex "transdifferentiation" process which occurred in the original monoclonal human epidermal HaCaT cells

Bi-modal differentiation pattern in a new human neuroblastoma cell line in vitro.

Abstract: We have isolated a human neuroblastoma (NB) cell line, HTLA230, from the bone-marrow aspirate of a patient with stage-IV disease. Subcutaneous tumors after inoculation of HTLA230 cells into nude mice were composed of primitive neuroblasts which rarely contained neuro-secretory granules. Cytogenetic studies of the cell line demonstrated 2 distinct populations of cells with common chromosomal markers. Stable sub-clones with a differentiated or undifferentiated cell morphology were isolated, demonstrating phenotypical heterogeneity of the HTLA230 parental cell line. Treatment with retinoic acid (RA) induced extensive neurite outgrowth in the parental cell line and in phenotypically differentiated sub-clones, but rarely in undifferentiated ones. Long-term treatment with RA was not associated with down-modulation of mycN-gene expression, which could be achieved only in cultures treated additionally with aphidicolin, a DNA-synthesis inhibitor, thus eliminating growing NB cells. A RA resistant subclone (CI-5) was isolated from parental HTLA230 cells grown at clonal cell density. Cells originally showed a homogeneously differentiated morphology; however, flat cells (F-cells) appeared with time and were subsequently separately propagated. Transdifferentiation of isolated F-cells into cells with neuron-like (N-cell) morphology was observed. Immunohistochemical analysis demonstrated that F-cells had lost the expression of neuronal markers, including HNK-l and A2B5, and expressed the intermediate filament, vimentin. Furthermore, F-cells showed high incorporation of [methyl-3H] thymidine (3H-TdR) by autoradiography but no mycN protein could be detected, although present in the parental cell line. These results then suggest that the isolated NB cell line and the RA-resistant variant line represent an excellent in vitro model with which the bi-modal differentiation pathway of NB can be analyzed on a molecular biological level.

Analysis of a unique molecule responsible for regeneration and stabilization of differentiated state of tissue cells.

Abstract: In Wolffian regeneration in the newt, a functional lens can be regenerated through cellular transdifferentiation of the pigmented epithelium of the mid-dorsal marginal iris. A novel monoclonal antibody, 2NI-36 mAb, generated in our laboratory has been utilized as a highly useful probe to study newt lens regeneration. The antigen molecule against this 2NI-36 mAb (2NI-36) became temporarily undetectable only at the site of lens regeneration. Moreover, the ventral iris pieces expressed the ability to differentiate a lens when pretreated with this monoclonal antibody and implanted in lentectomized eyes (Eguchi, Cell Differ. Dev. 25, Suppl., 1988). We have investigated the distribution of 2NI-36 in newt tissues. 2NI-36 was not specific to iris pigmented epithelium and distributed in many different kinds of mesodermal tissues, including dermis, blood vessel, mesonephros and so forth. 2NI-36 was also detected in either cell surface or intercellular spaces of cultured pigmented epithelial cells when they organized an epithelial cell sheet. Western blot analysis showed that 2NI-36 had the molecular weight of 50-200kD and was completely digested by trypsin, suggesting that 2NI-36 was a glycoprotein with many carbohydrate chains. It was also revealed by Western blot analysis that all the tissues in which 2NI-36 could be detected expressed this molecule similar to that in the iris epithelium. We expect that 2NI-36 is a glycoprotein expressed by various newt tissues and is functional to stabilize the differentiated state of each tissue cell in the same way as observed in the iris pigmented epithelial cells.

Transdifferentiation of macrophages into fibroblasts as a result of Schistosoma mansoni infection.

Abstract: The possibility of transdifferentiation of macrophages into fibroblasts which could be at the origin of fibrotic tissue in schistosome-infected mice was studied using immunocytochemical techniques. Macrophage cell samples extracted from the peritoneal cavity of schistosome-infected mice were fractionated on a Percoll gradient. The cultures were purified by treatment with a trypsin solution to eliminate any fibroblasts possibly collected along with the macrophages. Immunocytochemical methods were then used to characterize the cells at different points in time. The fibroblastic property of the morphologically transformed cells was confirmed by their positive labeling with the anti-procollagen antibody. However, these cells still possessed the mac-1 and mac-2 antigens which characterize the monomacrophage line.

Immunohistochemical evidence of high concentrations of metallothionein in pancreatic hepatocytes induced by cadmium in rats

Abstract: A recent study from our laboratory has shown that cadmium, a toxic heavy metal, is one of the most effective agents known for inducing hepatocytic transdifferentiation of the rat pancreas. With repeated injections of cadmium, the incidence of rats with pancreatic hepatocytic foci can be as high as 93%. Cadmium is also well known as a very potent inducer of metallothionein, a metal-binding protein that appears to be important in the biologic response to several toxic heavy metals in most tissues, including the pancreas. Therefore, the present study sought to determine if metallothionein was associated with cadmium-induced transdifferentiation of pancreatic cells. Expression of metallothionein was studied immunohistochemically by the peroxidase-antiperoxidase method in tissue sections of the pancreas of rats with pancreatic hepatocytes. High levels of metallothionein were localized primarily within the pancreatic hepatocytes. Surrounding normal pancreatic islet and acinar cells were not immunoreactive. Thus...

Fate of ciliated epidermal cells during early development of Xenopus laevis using whole-mount immunostaining with an antibody against chondroitin 6-sulfate proteoglycan and anti-tubulin: transdifferentiation or metaplasia of amphibian epidermis.

Abstract: Xenopus embryonic epidermis changes its cellular composition during development: the appearance of ciliated epidermal cells before hatching is a remarkable characteristic. In this study, the functional change of ciliated cells to mucus-secreting cells was examined with immunocytochemistry using anti-tubulin and anti-chondroitin 6-sulfate (C6S). Before hatching, most epidermal cells were labeled with anti-C6S in a granular fashion. Immunoelectron microscopy revealed that the anti-C6S-positive structure was the mucus granule. Ciliated epidermal cells lacked anti-C6S staining, but were strongly labeled with anti-tubulin. After hatching, most ciliated cells in the surface of the embryo disappeared. During their disappearance, some ciliated cells exhibited anti-C6S-positive granular labeling. This strongly suggests that the disappearance of ciliated cells is a functional conversion to mucus-secreting cells instead of shedding through cell death.

Role of Heterotypic Interactions Between Endothelial Cells and Parenchymal Cells in Organspecific Differentiation: A Possible Trigger for Vasculogenesis

Abstract: Endothelial cell organ-specificity, diversity and heterogeneity. The endothelial cell (EC) lining of the vascular wall is presumably the largest complex functional organ in the body, in terms of exposed surface area. Among the most striking features of ECs are phenotypic diversity within the vascular tree, functional heterogeneity, and organ-specificity1-3. Details of the cellular and molecular basis for organ-specificity of ECs are not understood 4.5. Early embryonic differentiation of ECs occurs via two distinct mechanisms, namely angiogenesis and vasculogenesis6. Angiogenesis is defined as the formation of new blood vessels by endothelial cell-sprouting from existing blood conduits. In some organs, for example the brain and the kidneys, angiogenesis is the primary form of vascularization. Angiogenic mechanisms are also responsible for the vascularization of tumors. Vasculogenesis indicates the in situ transdifferentiation of endothelial cells from local mesenchymal precursor cells. For example, the initial formation of the dorsal aortae and the vascularization of several organs, such as the liver, lung, and pancreas occur, predominantly, via vasculogenesis8.

A choice between glycogen and δ-crystallin accumulation is made in glial cells and not influenced by overlying neurons

Abstract: Chick-embryo neuroretinal cells convert extensively into lens under low-glucose conditions, but this transdifferentiation process is blocked by high-glucose media. We have previously observed an inverse relationship between the levels of glycogen (a marker of normal retinoglial differentiation) and of delta-crystallin (a lens marker) in such cultures. However, most of the glycogen accumulated under high-glucose conditions is apparently localized in those glial (G) cells underlying clusters of neurons (N cells). We here show that glial-enriched cultures (largely depleted of N cells) both accumulate glycogen and fail to transdifferentiate in high-glucose media. Moreover, glycogen localization in groups of glial cells is unaffected by the absence of N cells. Thus the choice between normal and foreign differentiation pathways is made autonomously within the retinoglial-cell population and is not influenced significantly by the presence or absence of N cells.