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PubMed2000

Xenopus laevis gelatinase B (Xmmp-9): development, regeneration, and wound healing.

It has been argued that matrix metalloproteinases play important roles in cellular differentiation and regeneration in certain systems. While studying changes in gene expression associated with the phenomena of cornea/lens transdifferentiation ("lens regeneration"), which takes place in the larva of Xenopus laevis, we identified the Xenopus gelatinase B gene. The open reading frame is homologous to other gelatinase B genes identified in other species and encodes all of the domains characteristic of this protein. Xenopus gelatinase B (Xmmp-9) is first expressed during early tail-bud stages in a subset of mesodermal cells scattered throughout the body. Expression is also seen in the peripheral tissues of the developing liver diverticulum, the hindgut/cloaca, and the paired caudal vein, and its dorsal branch in the larval tail. Given the significant role of matrix metalloproteinases in degrading components of the extracellular matrix, Xmmp-9 expression may be important in the morphogenesis of these structures. Xmmp-9 expression was also examined during the processes of cornea/lens transdifferentiation, epithelial wound healing, and limb regeneration in Xenopus larvae. Although Xmmp-9 is expressed very early during cornea/lens transdifferentiation, expression is restricted to the site of the peripheral wound created by removal of the original lens, which triggers transdifferentiation. Expression was not found in the central, uninjured area of the cornea where transdifferentiation takes place. Therefore, Xmmp-9 does not appear to play an important role in cornea/lens transdifferentiation. Xmmp-9 expression is associated with other epithelial wounds, indicating that gelatinase B is expressed in the general context of wound healing in Xenopus. Finally, Xmmp-9 is expressed in the ectoderm and mesoderm at the tip of the amputated limb, very early during limb regeneration, where it is argued to play a role in this process.

Amino Acid Sequence↗
PubMed1997

Lens regeneration in larval Xenopus laevis: experimental analysis of the decline in the regenerative capacity during development.

In Xenopus laevis, the capacity to regenerate a new lens from the outer cornea gradually decreases between stages 50 and 58, is almost negligible during the metamorphic climax, and disappears after metamorphosis. The factors responsible for lens transdifferentiation of the outer cornea are produced by the neural retina and are located in the vitreous chamber. This decrease in the regenerative capacity may be due to: (1) a reduction of the inductive power of the retina, (2) a reduction of lens-forming competence of the outer cornea, (3) an inhibition of the lens transdifferentiation process, (4) a combination of these causes. In order to test these hypotheses, fragments of outer cornea or of outer and inner corneas joined together were isolated from early larvae, late larvae and froglets, and implanted into the eye of host larvae during the premetamorphosis or the metamorphic climax. Results from implants of outer cornea into the vitreous chamber showed that the drop in lens regeneration capacity during the metamorphic climax is not due to a decrease in the inductive power of the retinal factor and that the gradual decrease in the regenerative capacity observed between stages 50 and 58 is not related to a substantial diminution in the capacity of outer cornea cells to transdifferentiate into lens fibers. Results from implants of outer and inner corneas joined together showed that in these implants the lens transdifferentiation of the outer cornea was partially inhibited. These findings indicate that the decrease in lens regeneration is mainly due to an inhibition of the lens transdifferentiation process of the outer cornea by the inner cornea. However, even implants of cornea (multilayered epithelium and substantia propria) excised from metamorphosed animals were able to form lens fibers, although to a lesser percentage than that obtained after implantation of fragments of larval outer and inner corneas. Thus, the lens-forming competence in the corneal epithelium is still present to a certain degree even when lens regeneration capacity is lost. Several observations suggest that in the lentectomized eye of late larvae and froglets the mechanical inhibition of lens transdifferentiation process exerted by the inner cornea (or the substantia propria), due to the rapid formation of a connective barrier against the spreading of the retinal factor toward the outer cornea, has a decisive role in maintaining the phenotypic stability of the outer cornea.

Animals↗
PubMed2005

Dysfunction of releasing adrenaline in asthma by nerve growth factor.

In early embryo period, adrenal medulla development is parallel with adrenergic nerves. For this reason, adrenal medullary cells may be provided with potential ability of transdifferentiation toward nerve cells. In vitro, it was well testified that if adrenal medullary cells are treated with nerve growth factor, then the cells will be transdifferentiated toward sympathetic neurons characterized by morphous, biochemistry and physiological functions. As a result, adrenal medullary cells after being transdifferentiated may be impaired in the functions of endocrine secretion, and the composition and level of adrenal medullary hormone are changed. The above mentioned may be regarded as the reflection of cellular redundancy, besides the transdifferentiation, multiple organelle copies, cellular plasticity and so on are all reflections of cellular redundancy. According to cellular redundancy, in addition to their regular and specific functions, some cells(types) are of other additional functions which will be activated in special state such as diseases, external interference and so on. The theory of cellular redundancy is seemed to unpuzzle the dysfunction of releasing adrenaline in asthma. Compared with the normal control group, the study showed that the levels of adrenaline in the asthmatic patients were not increased when patients encountered the strong stimuli. Subsequently, overexpression of nerve growth factor was also detected in asthmatic patients and animals. However, accompanied with alleviation of airway inflammation, these phenomena were changed when asthmatic models were treated with antibody of nerve growth factor. What makes these phenomena linked? The existence of functional redundancy of medullary cells may be the most important factor. This hypothesis suggests that overexpression of nerve growth factor activate functional redundancy of adrenal medullary cells, which makes these cells transdifferentiated toward sympathetic neurons. The transdifferentiation (tendency) may impair the functions of endocrine secretion of adrenal medullary cells. As a result, adrenal medulla cannot release enough adrenaline to relieve bronchoconstriction in asthmatic attack.

Adrenal Medulla↗
PubMed2000

On the track of a human circulating mesenchymal stem cell of neural crest origin.

The neural markers present in the normal circulating monocytoid cells able, in pathological situations, to trans-differentiate into different mesenchymal-type cells, confirm the hypothesis previously raised that these cells derive from the neural crest. In culture, the normal cells display a great plasticity very reminiscent of microglial cells in culture. Almost a quiescent cell in normal individuals, this monocytoid cell shows its division potentialities in pathological situations of fibrosis and cancer (chondrosarcoma) where it is found to spontaneously proliferate. While the normal neofibroblasts are rapidly recognized and destroyed by fibrophagic T-lymphocytes, the pathological cells escape this control and, as a result, they accumulate in vitro giving rise to a tissue sometimes organized as nodules. Although basically the transdifferentiation process is similar in all the pathological situations of fibrosis and cancer studied so far, the end-result phenotype evokes the pathology the patient is suffering from. It evokes osteoblasts in a case of osteomyelosclerosis, chondroïdocytes in a case of chondrosarcoma, myelofibroblasts in a case of fibrosis of lung and kidney in a patient under ciclosporine treatment. Hence, this circulating monocytoid cell is a multipotent cell with great division potentiality. These are characteristics of stem/preprogenitor cells. Since this circulating monocytoid cell also bears the neural markers we called it a monocytoid ectomesenchymal stem/preprogenitor cell. Therefore, the existence of an ectomesenchymal system is discussed here. The circulating monocytoid ectomesenchymal stem/preprogenitor cell might be involved in the normal cicatrisation process while the fibrophagic T lymphocytes might be involved in its termination. Impairment of this controlled mechanism might result in the development of fibrosis and/or cancer such as chondrosarcoma in vivo. Interestingly, at least in vitro, proliferation is restricted to the monocytoid cell before transdifferentiation takes place. In this model, fibrosis and cancer might share some common steps going from the proliferation of the monocytoid cells to their transdifferentiation into mesenchymal-type cells and the accumulation of these transdifferentiated cells in the tissues. Then, cancer might be distinguished from fibrosis by the additional acquisition of the ability to proliferate by the transdifferentiated cells. The monocytoid ectomesenchymal stem/preprogenitor cell might also be involved in brain neurodegenerative diseases characterized by an accumulation of microglia. The circulating monocytoid ectomesenchymal stem/preprogenitor cell appears as a target for gene therapy in pathological situations of fibrosis and/or cancer where it proliferates out of control. If the normal cell can be expanded and if its transdifferentiation can be directed, the circulating monocytoid ectomesenchymal stem/preprogenitor cell may become a useful tool for cellular therapy, in case of failure in wound healing and tissue regeneration.

Animals↗
PubMed2003

Identification of genes differentially expressed in mouse mammary epithelium transformed by an activated beta-catenin.

Beta-catenin is an executor of Wnt signaling and it can control cell fate and specification. Deletion of exon 3 from the endogenous beta-catenin gene in differentiating mammary alveolar epithelium of the mouse results in the generation of an activated protein that lacks amino acids 5-80. This is accompanied by a loss of mammary epithelial differentiation and a transdifferentiation process to squamous metaplasias. To further understand the molecular process of transdifferentiation, the expression of genes in mammary tissue was profiled in the absence and presence of activated of beta-catenin. Microarrays were generated that carry about 8500 cDNA clones with approximately 6000 obtained from mammary tissue. Mutant tissues, which had undergone either partial (TD1) or complete (TD2) squamous transdifferentiation, were compared with wild-type mammary tissue. Four groups of genes were identified. Group 1 contained genes whose expression was induced in both mutant tissues. Groups 2 and 3 contained genes that were active preferentially in TD2 and TD1, respectively. Group 4 contained genes suppressed in both samples. Using this approach, known and unknown genes activated in the transdifferentiation process were identified. A new 20 kDa protein (PANE1) induced upon transdifferentiation was nuclear in nonconfluent cells and cytoplasmic in confluent or dividing cells. Lastly, stabilization of beta-catenin resulted in the retention of differentiated epithelium upon involution and altered activities of several proteases in transdifferentiated mammary epithelium.

Amino Acid Sequence↗
PubMed2002

Characterizing gene expression during lens formation in Xenopus laevis: evaluating the model for embryonic lens induction.

Few directed searches have been undertaken to identify the genes involved in vertebrate lens formation. In the frog Xenopus, the larval cornea can undergo a process of transdifferentiation to form a new lens once the original lens is removed. Based on preliminary evidence, we have shown that this process shares many elements of a common molecular/genetic pathway to that involved in embryonic lens development. A subtracted cDNA library, enriched for genes expressed during cornea-lens transdifferentiation, was prepared. The similarities/identities of specific clones isolated from the subtracted cDNA library define an expression profile of cells undergoing cornea-lens transdifferentiation ("lens regeneration") and corneal wound healing (the latter representing a consequence of the surgery required to trigger transdifferentiation). Screens were undertaken to search for genes expressed during both transdifferentiation and embryonic lens development. Significantly, new genes were recovered that are also expressed during embryonic lens development. The expression of these genes, as well as others known to be expressed during embryonic development in Xenopus, can be correlated with different periods of embryonic lens induction and development, in an attempt to define these events in a molecular context. This information is considered in light of our current working model of embryonic lens induction, in which specific tissue properties and phases of induction have been previously defined in an experimental context. Expression data reveal the existence of further levels of complexity in this process and suggests that individual phases of lens induction and specific tissue properties are not strictly characterized or defined by expression of individual genes.

Animals↗
PubMed2006

Transforming growth factor beta2-induced myofibroblastic differentiation of human retinal pigment epithelial cells: regulation by extracellular matrix proteins and hepatocyte growth factor.

Retinal pigment epithelial (RPE) cells possess the potential to transdifferentiate into myofibroblasts after stimulation with transforming growth factor beta (TGFbeta) and are implicated in the pathogenesis of proliferative vitreoretinopathy. In this study we evaluated how TGFbeta2 and various extracellular matrix (ECM) proteins modulate the transdifferentiation of human fetal retinal pigment epithelial cells (RPE) cells into myofibroblast-like cells. Furthermore, we investigated whether hepatocyte growth factor (HGF) can suppress this transdifferentiation. RPE cells were cultured on ECM coated or uncoated surfaces in the presence or absence of TGFbeta2. HGF was added to certain cultures only once or on a daily basis during the treatment. Transdifferentiation of RPE cells into myofibroblasts was assessed by the quantitation of alpha-smooth muscle actin (alpha-SMA) using immunocytochemistry, flow cytometry, real-time PCR and Western blotting. TGFbeta2 induced a significant increase of alpha-SMA expression in a dose-dependent manner. Compared with growth on uncoated surfaces, RPE cultured on fibronectin (FN)-coated surfaces and stimulated with TGFbeta2 showed a significantly higher alpha-SMA expression than untreated cells. This upregulation of alpha-SMA could be markedly reduced by daily treatment with HGF; however, a single HGF administration did not significantly reduce alpha-SMA. These findings are important for further understanding the interaction of cytokines, RPE cells and their environment in mesenchymal transformation as well as its possible modulation. Continuous or long-term treatment with HGF should be further investigated for its potential to prevent mesenchymal transdifferentiation of RPE cells, and ultimately, PVR in vivo.

Actins↗
PubMedDate not supplied

Can a bone marrow cell contribute to organ regeneration? In vivo analysis using transgenic rats with reporter genes.

Although implantation of multipotent bone marrow-derived stem cells represents an attractive new cell therapy to repair damaged tissues, recent reports have raised serious concerns over the feasibility of using stem cells deriving from the bone marrow to promote cell transdifferentiation. We established transgenic (Tg) rats with reporter genes as specific molecular tags to examine the effect of bone marrow cells (BMCs) on transdifferentiation into tissues/organs. To monitor transdifferentiation events of locally transplanted BMCs into hepatocytes or capillary endothelial cells, a liver injury model and an ischemic hind-limb model were developed in rats. To test the ability of circulating bone marrow-derived cells to give rise to myocytes after skeletal muscle injury, we used a bone marrow cell transplantation model from Tg rats, which showed ubiquitous expression of beta-galactosidase (lacZ), into lethally irradiated non-Tg rats. Our results show that there was little transdifferentiation of BMCs into the targeted cells in these tissue injury models. However, in the ischemic hind-limb model, laser Doppler imaging and histologic analysis showed that both implantation of BMCs and treatment with microspheres incorporating basic fibroblast-like growth factor (bFGF), which enables the release of bFGF at the site of action over a period of time, effectively induced angiogenesis. In conclusion, rat BMCs with specific marker genes could be a useful tool for detecting transdifferentiation events in vivo.

Animals↗
PubMed2005

Tissue interaction between the retinal pigment epithelium and the choroid triggers retinal regeneration of the newt Cynops pyrrhogaster.

Complete retinal regeneration in adult animals occurs only in certain urodele amphibians, in which the retinal pigmented epithelial cells (RPE) undergo transdifferentiation to produce all cell types constituting the neural retina. A similar mechanism also appears to be involved in retinal regeneration in the embryonic stage of some other species, but the nature of this mechanism has not yet been elucidated. The organ culture model of retinal regeneration is a useful experimental system and we previously reported RPE transdifferentiation of the newt under this condition. Here, we show that cultured RPE cells proliferate and differentiate into neurons when cultured with the choroid attached to the RPE, but they did not exhibit any morphological changes when cultured alone following removal of the choroid. This finding indicates that the tissue interactions between the RPE and the choroid are essential for the former to proliferate. This tissue interaction appears to be mediated by diffusible factors, because the choroid could affect RPE cells even when the two tissues were separated by a membrane filter. RPE transdifferentiation under the organotypic culture condition was abolished by a MEK (ERK kinase) inhibitor, U0126, but was partially suppressed by an FGF receptor inhibitor, SU5402, suggesting that FGF signaling pathway has a central role in the transdifferentiation. While IGF-1 alone had no effect on isolated RPE, combination of FGF-2 and IGF-1 stimulated RPE cell transdifferentiation similar to the results obtained in organ-cultured RPE and choroid. RT-PCR revealed that gene expression of both FGF-2 and IGF-1 is up-regulated following removal of the retina. Thus, we show for the first time that the choroid plays an essential role in newt retinal regeneration, opening a new avenue for understanding the molecular mechanisms underlying retinal regeneration.

Animals↗
PubMed2003

Origin and development of the precursor lesions in experimental pancreatic cancer in rats.

Notwithstanding the importance of understanding how pancreatic ductal adenocarcinoma develops, the process remains controversial. A key question is whether the cells of origin of the tubular complexes that constitute precursor lesions are derived from a single cell type or from multiple types. Suggestions that they arise solely from centroacinar cells or ductal cells have been based on inference due to their morphologic appearance in tissue from patients or investigation of limited numbers of tubular complexes in animal models later in the carcinogenic process. The present study establishes clearly that two steps are involved; rapid transdifferentiation to produce tubular complexes followed later by transformation of the component cells. Animals were killed at intervals beginning 1 day after implantation of the carcinogen dimethylbenzanthracene. Transdifferentiation of acinar cells to ductal cells does not require cell division. Transition of lobules to tubular complexes begins by 2 days after implantation of carcinogen. Within 4 days after implantation well-developed tubular complexes are present. Islets participate in the process. Ductal adenocarcinoma is observed by 1 month after implantation of carcinogen. Chymotrypsin and cytokeratin localized by immunocytochemistry indicate acinar and ductal cell characteristics. Acino-ductal transdifferentiation persists in carcinogen-implanted animals, but not in controls implanted with sodium chloride crystals or subjected to sham implantation. The precursor lesions (tubular complexes) are formed by the transdifferentiation of acinar cells and to a lesser extent islet cells, with the incorporation of the duct cells pre-existing in the lobules. Therefore, cells that at one time were acinar cells, islet cells, and duct cells, provide the precursor cells for the ductal adenocarcinoma that develops from tubular complexes. The results raise the question whether the transdifferentiated cells in the tubular complexes of patients with chronic pancreatitis are more susceptible to carcinogenic influences, resulting in the increased rate of pancreatic cancer.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed2005

Metaplasia in the pancreas.

There is currently much interest in the possibility to treat chronic diseases by cell replacement or regenerative therapies. Most of these studies focus on the manipulation of undifferentiated stem cells. However, tissue repair and regeneration can also be achieved by differentiated cells, which, in certain conditions, can even transdifferentiate to other cell types. Such transdifferentiations can lead to tissue metaplasia. The pancreas is an organ wherein metaplasia has been well investigated and for which experimental models have been recently developed allowing to unravel the molecular basis of transdifferentiation. Pancreatic metaplasias studied so far include the conversion of exocrine acinar cells to duct cells, exocrine cells to endocrine islet cells, endocrine cells to duct cells, and acinar cells to hepatocytes. Epitheliomesenchymal transitions have also been described. The available evidence indicates that mature cells can be reprogrammed by specific environmental cues inducing the expression of cell type-specific transcription factors. For example, the glucocorticoid hormone dexamethasone induces pancreatic transdifferentiation to hepatocytes, whereas the combination of epidermal growth factor and leukemia-inhibitory factor induces exocrine-endocrine transdifferentiation in vitro. Further unravelling of the involved signal transduction pathways, transcription factor networks, and chromatin modifications is required to manipulate metaplasia at will and to apply it in tissue repair or regeneration.

Animals↗
PubMed2004

Interleukin-6 inhibits the growth of prostate cancer xenografts in mice by the process of neuroendocrine differentiation.

In vitro, the human prostate cancer (PCA) cell line LNCaP can be permanently transdifferentiated into a quiescent neuroendocrine (NE) phenotype by the cytokine interleukin-6 (IL-6). Recently, we have shown that the growth of prostate cancer cells is significantly suppressed when cocultured with NE cells. In order to explore the inhibitory activity of IL-6 on prostate tumor growth, nude mice bearing xenografts of the PCA cell lines LNCaP and DU-145 (a line that is incapable of NE transdifferentiation by IL-6 in vitro) were treated with IL-6 for 3 weeks, either injected around the tumor or systematically released from implanted minipumps. Both administration forms of IL-6 inhibited the growth of LNCaP xenografts by more than 75% compared to the control group. In contrast, there was no difference in DU-145 tumor growth between IL-6-treated animals and controls. In comparison to control and DU-145 tumors, both IL-6 injected and pump-infused LNCaP tumors exhibited a significant increase in the expression of the NE markers neuron-specific enolase (NSE) and betaIII tubulin. Serum NSE levels were also significantly elevated in both IL-6-treated LNCaP tumor groups when compared to controls. IL-6 treatment resulted in G(0) cell cycle accumulation as evidenced by a loss of Ki-67 expression in > 90% of LNCaP tumor cells. These combined results demonstrate that IL-6-induced NE transdifferentiation of PCA cells has a significant inhibitory effect on tumor growth in mice. Agents, like IL-6, capable of NE transdifferentiation of PCA cells, should be considered as a new therapeutic approach for the treatment of prostate cancer.

Animals↗
PubMed2002

Involvement of phosphoinositide turnover in tracheary element differentiation in Zinnia elegans L. cells.

Mesophyll cells of Zinnia elegans L., cultured in the presence of phytohormones, will transdifferentiate and undergo programmed cell death to become tracheary elements, thick-walled cells of the xylem. This system is a model system for study of plant cell development and differentiation. We report that a high concentration of extracellular Ca(2+) is necessary during the first 6 h of culturing for tracheary elements to form. Extracellular Ca(2+) is still required at later times, but at a much lower concentration. When cells transdifferentiate in adequate Ca(2+), microsomal phospholipase C activity increases and levels of inositol 1,4,5-trisphosphate rise at about hour 4 of culturing. The production of inositol 1,4,5-trisphosphate appears to be important for tracheary element formation, since inhibitors of phospholipase C inhibit both inositol 1,4,5-trisphosphate production and tracheary element formation. Pertussis toxin, an inhibitor of GTP-binding proteins, inhibits transdifferentiation and eliminates inositol 1,4,5-trisphosphate production. Tracheary element formation was not completely abolished by inhibitors that eliminated inositol 1,4,5-trisphosphate production, suggesting the involvement of other pathways in regulating transdifferentiation.

Asteraceae↗
PubMed1986

Enhancement of expression of lens phenotype in cultures of pigmented epithelial cells by hyaluronidase in the presence of phenylthiourea.

Pigmented epithelial cells isolated from 8-9-day-old chick embryos can transdifferentiate into lens-like cells at the terminal period of the third generation of culture. However, efficiency of this transdifferentiation is usually rather low. Phenylthiourea, a potent inhibitor of melanin synthesis, effectively enhances transdifferentiation of pigmented epithelial cells into lens-like cells in vitro. Lentoid bodies began to appear in the multilayered region of primary cultures of pigmented epithelial cells maintained in medium containing phenylthiourea at concentrations between 0.5 and 1.0 mM. Furthermore, the enhancing effect of phenylthiourea can be amplified with testicular hyaluronidase. Under these conditions, pigmented epithelial cells grow vigorously and lose their differentiative properties, efficiently switching their phenotype into lens-like cells some 20 days after initiation of culture in the presence of both substances. Semiquantitative analysis revealed that testicular hyaluronidase amplified the effect of phenylthiourea more than 100-fold. It has been suggested that phenotypic expression of pigmented epithelial cells during transdifferentiation can be regulated by manipulating the microenvironment in which these cells reside.

Animals↗
PubMedDate not supplied

Tissue-specific muscle, neural and liver stem/progenitor cells reside in the bone marrow, respond to an SDF-1 gradient and are mobilized into peripheral blood during stress and tissue injury.

Several reports imply that bone marrow hematopoietic stem cells transdifferentiate into tissue-specific stem cells; however, the possibility of committed tissue-specific stem cells pre-existing in the bone marrow has not been dealt with adequately. We present here an alternative explanation of the so-called phenomenon of stem cell transdifferentiation. First, we postulate that tissue-committed stem/progenitor cells circulate in the peripheral blood and compete for tissue-specific niches. The circulation of these cells plays an important physiological role in maintaining a pool of stem cells in distant parts of the body and the number of these cells in peripheral blood can be increased by the administration of agents similar to those used for mobilization of hematopoietic stem cells. Second, we postulate that bone marrow tissue is a source of various stem-cell chemoattractants and survival factors and provides an environment that chemoattracts tissue-specific circulating stem/progenitor cells. In this context, we envision bone marrow as a "home" or "hide-out place" not only of hematopoietic stem cells but also of already differentiated circulating tissue-specific stem/progenitors. In support of this concept, we report here that mRNA of several early markers for muscle (Myf-5, Myo-D), neural (GFAP, nestin) and liver (CK19, fetoprotein) is detectable in circulating (adherent cell-depleted) peripheral blood mononuclear cells. Moreover, using real-time RT-PCR, we found that the level of expression of these markers increases in the peripheral blood of humans and mice after mobilization by G-CSF. Furthermore, using stromal-derived factor-1 (SDF-1) chemotaxis and real-time RT-PCR analysis, we present evidence that early tissue-specific stem cells reside in normal human and murine bone marrow, express the CXCR4 receptor on their surface and can be highly enriched (in humans and mice) after chemotaxis to SDF-1 gradient. All our experiments were performed on freshly isolated cells to exclude the potential contribution of transdifferentiated hematopoietic stem or mesenchymal cells in the culture. We maintain that any transdifferentiation studies employing populations of bone marrow cells should rule out the possibility that the apparently pure hematopoietic stem cell population could in fact contain pre-existing tissue-specific stem/progenitors.

Animals↗
PubMed2005

Critical role of the Rho-kinase pathway in TGF-beta2-dependent collagen gel contraction by retinal pigment epithelial cells.

Retinal pigment epithelial cells (RPEs) are thought to be one of the main components of fibrous membrane observed in eyes with proliferative vitreo-retinopathy. We investigated the signalling mechanisms of TGF-beta2-dependent collagen gel contraction by RPEs. An in vitro type I collagen gel contraction assay was performed to evaluate the effect of TGF-beta2 on gel contraction. The expression of alpha-smooth muscle actin (alpha-SMA) and the phosphorylation state of myosin light chain (MLC) were analyzed by Western blotting. The involvement of protein kinases such as p44/42 mitogen-activated protein kinase (MAPK), protein kinase C (PKC), p38 MAPK and phosphatidylinositol-3 kinase was investigated. The contribution of Rho-kinase and/or MLC-kinase was also evaluated using respective kinase inhibitors (Y27632, hydroxyfasudil and ML7). Additionally, RPEs were immunostained to examine whether the expression of alpha-SMA detected in our western blotting correlated to the stress fiber formation within the cells. TGF-beta2 caused time (0-5 days)-and dose (0 10 ng ml(-1))-dependent gel contraction associated with overexpression of alpha-SMA and phosphorylation of MLC (p < 0.01, respectively). PKC inhibitor (GF109203X, 5 microM) and p38 MAPK inhibitor (SB203580, 10 microM) significantly attenuated TGF-beta2-elicited gel contraction via partial downregulation of both alpha-SMA expression and MLC phosphorylation (p < 0.01, respectively). The gel contraction was prominently inhibited in the presence of Y27632 (10 microM) or hydroxyfasudil (10 microM) with strong suppression of MLC phosphorylation but had no significant effect on alpha-SMA expression. Treatment with ML7, in contrast, resulted in a marginal inhibition of MLC phosphorylation and gel contraction. Finally, pretreatment of the cells with Y27632 or hydroxyfasudil prevented the formation of stress fiber within the cells. These results indicate that TGF-beta2-dependent myofibroblastic transdifferentiation and MLC phosphorylation by RPEs involve both PKC and p38 MAPK pathways at least in part. Myofibroblastic transdifferentiation of RPEs appears to be independent of the Rho-kinase pathway, and the presence of alpha-SMA does not necessarily reflect the contractile potential of a cell. While Rho-kinase inhibitors are incapable of preventing myofibroblastic transdifferentiation itself, this pathway could be one of the critical targets of cell-mediated contraction of the tissue containing fibrillar collagens by transdifferentiated RPEs.

Actins↗
PubMed1996

Effect of pentoxifylline on the fibrogenic functions of cultured rat liver fat-storing cells and myofibroblasts.

The effects of pentoxifylline (PTX) an analogue of the methylxanthine theobromine, on basic fibrogenic reactions of cultured fat-storing cells and myofibroblasts (MFB), the cell types most important for the excessive production of extracellular matrix components in fibrosing liver, were studied. The proliferation of MFB (i.e., activated, transdifferentiated fat-storing cells) was more dose-dependently inhibited by pentoxifylline than that of unactivated fat-storing cells (ED50 50 microgram/mL). In addition, PTX retarded the transdifferentiation of fat-storing cells into smooth muscle alpha-actin positive MFB, a 50% reduction in actin-positive cells being reached with concentrations of 0.5 mg PTX/mL medium. The transdifferentiation-associated decrease in retinyl palmitate of cultured fat-storing cells was delayed by PTX. The synthesis of [35S] sulfate-labeled glycosaminoglycans (GAG) and total and cellular fibronectin was not significantly reduced by treatment of MFB with PTX up to 1.0 mg/mL. It is concluded that PTX reduces the transdifferentiation of fat-storing cells to MFB and the proliferation of MFB, but leaves the synthesis of extracellular matrix components GAG and fibronectin unaffected. The effect of PTX on the former reactions might account for the reported antifibrogenic properties of this drug in experimental hepatic fibrogenesis.

Adipocytes↗
PubMed1999

Expression of gicerin, a cell adhesion molecule, in the abnormal retina in silver plumage color mutation of Japanese quail (Coturnix japonica).

Silver plumage color mutant (B/B) quail has an abnormal retina characterizing the transdifferentiation of retinal pigment epithelium (RPE) following the retinal separation in the early developmental stage. In the present study; (i) the expression of gicerin, an immunoglobulin-superfamily cell adhesion molecule, was examined in the retina of B/B quail. In the wild-type quail, gicerin protein was enriched in the apical membrane (facing the neural retina, NR) of RPE cells on embryonic day (E) 4 and then appeared also in NR cells from E5. However, in the B/B retina, no gicerin expression was found in the transdifferentiation area of RPE prior to the retinal separation. (ii) In addition to this, microinjection of anti-gicerin polyclonal antibody into the eyeball of wild-type quail on E3 caused the retinal separation and induced the transdifferentiation of RPE into new NR. These observations suggest that the decrease of gicerin expression might participate in the retinal separation and RPE-transdifferentiation in B/B quail.

Animals↗