DEVELOPMENT GROWTH & DIFFERENTIATION, Issue 2 2006
Sidhartha S. Tulachan
Epithelial,mesenchymal interactions are crucial for the proper development of many organs, including the pancreas. Within the pancreas, the ducts are thought to harbor stem/progenitor cells, and possibly to give rise to pancreatic ductal carcinoma. Little is known about the mechanism of formation of pancreatic ducts in the embryo. Pancreatic mesenchyme contains numerous soluble factors which help to sustain the growth and differentiation of exocrine and endocrine structures. Here, we report that one such morphoregulatory mesenchymal protein, epimorphin, plays an important role during pancreatic ductal proliferation and differentiation. We found that epimorphin is expressed in pancreatic mesenchyme during early stages of development, and at mesenchymal,epithelial interfaces surrounding the ducts at later stages. Strong upregulation of epimorphin expression was seen during in vitro pancreatic duct differentiation. Similarly, in vitro pancreatic duct formation was inhibited by a neutralizing antibody against epimorphin, whereas addition of recombinant epimorphin partially rescued duct formation. Together, our study demonstrates the role of epimorphin in pancreatic ductal morphogenesis. [source]

Increased bacterial permeation in long-lasting ileoanal pouches

INFLAMMATORY BOWEL DISEASES, Issue 8 2006
Anton J. Kroesen MD
Abstract Background and Aims: Bacterial overgrowth appears to play an important role in the pathogenesis of ileoanal pouches. Therefore, the capability of bacterial permeation and its determinants is of great interest. The aim of this study was to examine bacterial permeation in the ileoanal pouch and to correlate the results with the degree of inflammation, the epithelial resistance, the mucosal transport function, and the age of the ileoanal pouches. Materials and Methods: Biopsies were taken from 54 patients before colectomy (n = 13; preileal pouch-anal anastomosis [IPAA]), and closure of ileostomy (n = 7; deviation), <1 year after closure of ileostomy (n = 8; intact pouch I), >1 year after closure of ileostomy (n = 16; intact pouch II), in the case of pouchitis (n = 11), and in 11 controls. Tissues were mounted in a miniaturized Ussing chamber. Escherichia coli was added to the mucosal side of the Ussing chamber, and the permeation was proven by serosal presence of E. coli. Epithelial and subepithelial resistance was determined by transmural impedance analysis. Active Na+ -glucose cotransport and active Cl, secretion were measured. Specimens were analyzed by fluorescent in situ hybridization with oligonucleotide probes targeting the bacterial 16s ribosomal RNA. The bacteria in and on the tissue were enumerated. Results: Bacterial permeation occurred in 2 of 13 pre-IPAA, 2 of 7 deviations, 0 of 8 intact pouch I, 9 of 16 intact pouch II, 5 of 11 pouchitis specimens, and 0 of 11 ileum controls. The frequency of bacterial permeation in the intact pouch II group is higher than in the intact pouch I group (P < 0.001). Epithelial resistance, mannitol fluxes, electrogenic chloride secretion, sodium-glucose cotransport of the bacterially permeated specimens versus nonpermeated of the intact pouch II group, and the pouchitis group and subepithelial resistance remained unchanged. Intramural bacteria could be detected by fluorescence in situ hybridization mainly in long-lasting pouches, but there was no correlation with bacterial permeation. Conclusions: The long-lasting ileoanal pouch is associated with increased bacterial permeability. This is not correlated with a disturbed function of the pouch mucosa but could be a precursor of pouchitis. [source]

Involvement of vascular endothelial growth factor, CD44 and CD133 in periodontal disease and diabetes: an immunohistochemical study

JOURNAL OF CLINICAL PERIODONTOLOGY, Issue 1 2009
Guendalina Lucarini
Abstract Aim: The aim of this study was to investigate the relationship between expression of angiogenic and regeneration markers and periodontal disease in subjects with/without diabetes mellitus. Material and Methods: Immunohistochemical detection of vascular endothelial growth factor (VEGF), CD44 and CD133 was performed in 16 samples each of (1) healthy gingiva from non-diabetic subjects (controls), (2) gingiva from non-diabetic subjects with periodontitis, (3) gingiva from subjects with type 1 diabetes and periodontitis, (4) gingiva from subjects with type 2 diabetes and periodontitis. Results: Diseased gingivae from patients with diabetes and periodontitis had greater clinical measures of periodontal disease than those with periodontitis only. VEGF expression was significantly enhanced in epithelial and endothelial cells from patients with periodontitis compared with controls (p<0.05). Epithelial CD44 expression was strong in all groups, while CD44 was significantly enhanced (p<0.05) in connective tissue cells from both diabetic groups. Epithelial and endothelial CD133 expression was comparable in all patients except those with type 2 diabetes and periodontitis, where it was not detected. Stromal CD133 expression was significantly lower in patients with type 2 diabetes and periodontitis and was increased in periodontitis patients (p<0.05). Conclusions: The involvement and high expression of VEGF, CD44 and CD133 in periodontal disease may predict a greater regeneration capacity of gingival tissue. [source]

Epithelial,mesenchymal interactions in experimental oral mucosal carcinogenesis

JOURNAL OF ORAL PATHOLOGY & MEDICINE, Issue 7 2001
Alison M. Rich
Abstract: In an effort to come to a better understanding of human oral mucosal carcinogenesis, an animal model was used in which the carcinogen 4-nitroquinoline-1-oxide was applied to rat palatal mucosa for varying periods of time. Histological and histometric analyses showed that there were quantifiable differences in the palatal epithelium to which carcinogen had been applied in comparison with control tissue. Tissue recombination experiments, using various combinations of the palatal mucosa and analysed after recovery from transplantation to hypothymic BALB/c mice, showed that control epithelium recombined with connective tissue from carcinogen-treated mucosa was altered, indicating that the underlying connective tissue modified histomorphological aspects of the epithelium in the later stages of carcinogenesis. [source]

Alcohol Stimulates Activation of Snail, Epidermal Growth Factor Receptor Signaling, and Biomarkers of Epithelial,Mesenchymal Transition in Colon and Breast Cancer Cells

ALCOHOLISM, Issue 1 2010
Christopher B. Forsyth
Background:, Alcohol consumption is associated with the risk of progressive cancers including colon and breast cancer. The mechanisms for the alcohol-induced aggressive behavior of these epithelial cancer cells have not been fully identified. Epithelial,mesenchymal transition (EMT) is a developmental program recently shown to play a role in cancer progression and metastases. We hypothesized that alcohol might promote cancer progression by inducing EMT in cancer cells and tested this hypothesis by assessing alcohol-stimulated changes in phenotypic markers of EMT as well as the EMT transcription factor Snail and its related cell signaling. Methods:, Colon and breast cancer cell lines and a normal intestinal epithelial cell line were tested as well as colonic mucosal biopsy samples from alcoholic subjects. Cells were treated with alcohol and assessed for EMT-related changes using immunofluorescent microscopy, western blotting, reporter assays, RT-PCR, and knockdown of Snail with siRNA. Results:, We show alcohol upregulated the signature EMT phenotypic marker vimentin as well as matrix metalloprotease (MMP)-2, MMP-7, and MMP-9 and cell migration in colon and breast cancer cells,all characteristics of EMT. Alcohol also stimulated nuclear localization of Snail phosphorylated at Ser246, transcription from a Snail reporter plasmid, and Snail mRNA expression by RT-PCR. Snail siRNA knockdown prevented alcohol-stimulated vimentin expression. In vivo, Snail expression was significantly elevated in colonic mucosal biopsies from alcoholics. Also, we found alcohol stimulated activation of epidermal growth factor receptor (EGFR) signaling and an EGFR inhibitor blocked alcohol-induced cell migration and Snail mRNA expression. Conclusions:, Collectively, our data support a novel mechanism for alcohol promoting cancer progression through stimulating the EMT program in cancer cells via an EGFR-Snail mediated pathway. This study reveals new pathways for alcohol-mediated promotion of cancer that could be targeted for therapy or prevention of alcohol-related cancers. [source]

Expression of Etk/Bmx Tyrosine Kinase in Intrahepatic Cholangiocarcinoma

JOURNAL OF SURGICAL ONCOLOGY, Issue 5 2008
Linlang Guo MD
Abstract Background Epithelial and endothelial tyrosine kinase (Etk), also known as bone marrow X kinase (Bmx) plays an important role in the growth, differentiation, apoptosis, proliferation, and tumorigenicity of epithelial cells. The purpose of this study was to investigate the expression of Etk/Bmx in intrahepatic cholangiocarcinoma (ICC) and correlated the expression with clinicopathological parameters. Methods Fifty-seven cases of ICC were immunostained for Etk/Bmx, HCV NS5, PCNA, bcl-2, and NF-,B p65. Results Etk/Bmx expression was present in 19 (33.3%) of 57 ICC specimens and correlated with cell differentiation and survival rate but not closely with tumor size and lymph node metastasis. There was a significant difference of expression of either PCNA-LI or Bcl-2 between Etk/Bmx-positive and -negative cases (P,<,0.05). However, no statistically significant association was found between Etk/Bmx expression and presence of HCV-NS5 or NF-,B p65. Conclusions Our results indicate that Etk/Bmx may be involved in the development of ICC and be a predictor of poor prognosis for ICC. HCV infection and NF-,B appears unrelated to Etk/Bmx expression. J. Surg. Oncol. 2008;97:428,432. © 2008 Wiley-Liss, Inc. [source]

Epithelial to mesenchymal transition in primary sclerosing cholangitis

LIVER INTERNATIONAL, Issue 8 2008
John A. Kirby
[source]

Muco-ciliary differentiation of nasal epithelial cells is decreased after wound healing in vitro

ALLERGY, Issue 8 2009
D. S. Lazard
Background:, Epithelial damage and modifications of cell differentiation are frequent in airway diseases with chronic inflammation, in which transforming growth factor-,1 (TGF-,1) plays an important role. The aim of this study was to evaluate the differentiation of human nasal epithelial cells (HNEC) after wound healing and the potential effects of TGF-,1. Methods:, Basal, mucus, and ciliated cells were characterized by cytokeratin-14, MUC5AC, and ,IV tubulin immunodetection, respectively. Their expression was evaluated in situ in nasal polyps and in an in vitro model of wound healing in primary cultures of HNEC after wound closure, under basal conditions and after TGF-,1 supplementation. Using RT-PCR, the effects of TGF-,1 on MUC5AC and DNAI1 genes, specifically transcribed in mucus and ciliated cells, were evaluated. Results:,In situ, high TGF-,1 expression was associated with low MUC5AC and ,IV tubulin expression. In vitro, under basal conditions, MUC5AC expression remained stable, cytokeratin-14 expression was strong and decreased with time, while ,IV tubulin expression increased. Transforming growth factor-,1 supplementation downregulated MUC5AC and ,IV tubulin expression as well as MUC5AC and DNAI1 transcripts. Conclusion:, After a wound, differentiation into mucus and ciliated cells was possible and partially inhibited in vitro by TGF-,1, a cytokine that may be involved in epithelial remodeling observed in chronic airway diseases. [source]

Epithelial,mesenchymal interactions in keloid pathogenesis modulate vascular endothelial growth factor expression and secretion,

THE JOURNAL OF PATHOLOGY, Issue 1 2007
CT Ong
Abstract Vascular endothelial growth factor (VEGF) plays an important role in angiogenesis during the wound healing process. As epithelial,mesenchymal interactions have been shown to regulate a plethora of genes in wound healing, we hypothesized that these interactions might have a role in modulating VEGF expression and angiogenesis. A two chamber co-culture model was used, wherein normal and keloid keratinocytes and fibroblasts were physically separated by membrane inserts while allowing cytokine diffusion. Cell lysates obtained from keratinocytes co-cultured with fibroblasts demonstrated increased expression of VEGF. An enzyme-linked immunosorbent assay (ELISA) showed significant increase in VEGF expression in co-culture conditioned media compared with controls. Additionally, the conditioned medium from keloid keratinocyte and fibroblast co-cultures increased proliferation and formation of complex three-dimensional capillary-like structures in human umbilical vein endothelial cells, emphasising the importance of epithelial,mesenchymal interactions in the angiogenic process. Immunostaining of keloid tissue localized VEGF in the basal layer of the epidermis and also demonstrated higher blood vessel density than normal skin. Keloid tissue extract also demonstrated increased expression of VEGF compared with normal skin. It is likely that epidermal VEGF exerts significant paracrine control over the dynamics and expression profile of underlying dermal fibroblasts. Addition of the inhibitors WP631, mitoxantrone, and Rapamycin to keloid keratinocyte and fibroblast co-cultures, downregulated secreted VEGF expression in a dose-dependent manner, suggesting therapeutic potential for these compounds in the treatment of keloid scars. Copyright © 2006 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [source]

Smad3 signalling plays an important role in keloid pathogenesis via epithelial,mesenchymal interactions

THE JOURNAL OF PATHOLOGY, Issue 2 2005
TT Phan
Abstract Smad signalling plays important roles in developmental and cancer biology as well as in fibropathogenesis. Its role in keloid biology is not known. Epithelial,mesenchymal interactions, originally described in normal skin, have recently been established to play a significant role in keloid pathogenesis, and demonstrate the important influence of keratinocyte paracrine factor signalling on fibroblast behaviour. The present study investigated the role of downstream Smad cascade induction in this interaction. Normal fibroblasts (NF) and keloid fibroblasts (KF) were co-cultured in serum-free medium with normal keratinocytes (NK) or keloid keratinocytes (KK) for 5 days, after which fibroblast cell lysates were subjected to western blot and immunoprecipitation analysis to quantify the levels of Smad and Smad2/3/4 binding complex. In another set of experiments, wild-type (wt), Smad2-null (Smad2,/,) and Smad3-null (Smad3,/,) mouse embryonic fibroblasts (MEF) were assayed for cell proliferation and collagen production after serum-free co-culture with KK or exposure to conditioned media collected from serum-free KK/KF co-culture. Compared to normal skin, keloids expressed high basal levels of TGF,R1 and TGF,R2, Smad2, 3 and 4 and phospho-Smad2. Upregulation of TGF,R1 and TGF,R2, Smad3 and p-Smad2 was observed in KF co-cultured with KK, together with enhanced Smad3 phosphorylation and Smad2/3/4 binding complex production. When MEF-wt, MEF-Smad2,/, or MEF-Smad3,/, were co-cultured with KK or exposed to KK/KF co-culture conditioned media, enhanced proliferation and collagen production were seen in MEF-wt and MEF-Smad2,/, but not in MEF-Smad3,/, cells. The activation of Smad signalling, importantly that of Smad3, appears to be one facet of the complex epithelial,mesenchymal interactions in keloid pathogenesis, resulting in active KF proliferation and collagen-ECM production in co-culture with KK. This finding suggests the suppression of Smad signalling as a novel approach in keloid therapy. Copyright © 2005 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [source]

Inflammation and Epithelial to Mesenchymal Transition in Lung Transplant Recipients: Role in Dysregulated Epithelial Wound Repair

AMERICAN JOURNAL OF TRANSPLANTATION, Issue 3 2010
L. A. Borthwick
Epithelial to mesenchymal transition (EMT) has been implicated in the pathogenesis of obliterative bronchiolitis (OB) after lung transplant. Although TNF-, accentuates TGF-,1 driven EMT in primary human bronchial epithelial cells (PBECs), we hypothesized that other acute pro-inflammatory cytokines elevated in the airways of patients with OB may also accentuate EMT and contribute to dysregulated epithelial wound repair. PBECs from lung transplant recipients were stimulated with TGF-,1 ± IL-1,, IL-8, TNF-, or activated macrophages in co-culture and EMT assessed. The quality and rate of wound closure in a standardized model of lung epithelial injury was assessed in response to above stimuli. Co-treatment with TGF-,1 + TNF-, or IL-1, significantly accentuates phenotypic and some functional features of EMT compared to TGF-,1 alone. Co-treatment with TGF-,1 + TNF-, or IL-1, accelerates epithelial wound closure however the quality of repair is highly dysregulated. Co-treatment with TGF-,1 + IL-8 has no significant effect on EMT or the speed or quality of wound healing. Activated macrophages dramatically accentuate TGF-,1-driven EMT and cause dysregulated wound repair. Crosstalk between macrophage-derived acute inflammation in the airway and elevated TGF-,1 may favor dysregulated airway epithelial repair and fibrosis in the lung allograft via EMT. [source]

Posttransplant Bronchiolitis Obliterans Syndrome Is Associated with Bronchial Epithelial to Mesenchymal Transition

AMERICAN JOURNAL OF TRANSPLANTATION, Issue 4 2009
S. Hodge
Bronchiolitis obliterans syndrome (BOS) compromises lung transplant outcomes and is characterised by airway epithelial damage and fibrosis. The process whereby the normal epithelial configuration is replaced by fibroblastic scar tissue is poorly understood, but recent studies have implicated epithelial mesenchymal transition (EMT). The primary aim of this study was to assess the utility of flow cytometry in detecting and quantifying EMT in bronchial epithelial cells. Large airway brushings were obtained at 33 bronchoscopies in 16 BOS-free and 6 BOS grade 1,3 patients at 2,120 months posttransplant. Flow cytometry was used to assess expression of the mesenchymal markers ,SMA, S100A4 and ED-A FN and HLA-DR. TGF ,1 and HGF were measured in Bronchoalveolar lavage (BAL). Expression of all three mesenchymal markers was increased in BOS, as was HLA-DR. BAL HGF, but not TGF ,1 was increased in BOS. Longitudinal investigation of one patient revealed a 100% increase in EMT markers concurrent with a 6-fold increase in BAL TGF ,1 and the diagnosis of BOS at 17 months posttransplant. Flow cytometric evaluation of bronchial epithelium may provide a novel and rapid means to assess lung allografts at risk of BOS. [source]

Epithelial to Mesenchymal Transition During Late Deterioration of Human Kidney Transplants: The Role of Tubular Cells in Fibrogenesis

AMERICAN JOURNAL OF TRANSPLANTATION, Issue 6 2005
Attapong Vongwiwatana
The hallmark of failing renal transplants is tubular atrophy and interstitial fibrosis (TA/IF). Injury to tubular epithelial cells (TEC) could contribute to fibrogenesis via epithelial,mesenchymal transition (EMT). We examined the features of EMT in renal transplants that developed TA/IF. Biopsies from 10 allograft kidneys with impaired function and TA/IF and 10 biopsies from transplants with stable function were compared to their implantation biopsies. Relative to implantation biopsies, TEC in TA/IF kidneys showed loss of epithelial markers (E-cadherin, cytokeratin) with altered distribution. Some TEC also showed new cytoplasmic expression of mesenchymal markers vimentin, S100A4, and alpha smooth muscle actin (,-SMA) and collagen synthesis marker heat shock protein (HSP-47), both in deteriorating and atrophic tubules. Double immunostaining showed coexpression of cytokeratin and vimentin, S100A4 and HSP-47, indicating intermediate stages of EMT in TA/IF. These changes were absent or much less in transplants with stable function. EMT features in the TA/IF group correlated with serum creatinine (vimentin, S100A4, HSP-47), history of T-cell-mediated rejection (cytokeratin, S100A4) and proteinuria (cytokeratin). These findings support a model in which the TEC damage induces loss of epithelial features and expression of fibroblast features, as a common pathway of deterioration by either immunologic or nonimmunologic processes. [source]

Epithelial,mesenchymal transition in cancer development and its clinical significance

CANCER SCIENCE, Issue 2 2010
Masaaki Iwatsuki
The epithelial,mesenchymal transition (EMT) plays a critical role in embryonic development. EMT is also involved in cancer progression and metastasis and it is probable that a common molecular mechanism is shared by these processes. Cancer cells undergoing EMT can acquire invasive properties and enter the surrounding stroma, resulting in the creation of a favorable microenvironment for cancer progression and metastasis. Furthermore, the acquisition of EMT features has been associated with chemoresistance which could give rise to recurrence and metastasis after standard chemotherapeutic treatment. Thus, EMT could be closely involved in carcinogenesis, invasion, metastasis, recurrence, and chemoresistance. Research into EMT and its role in cancer pathogenesis has progressed rapidly and it is now hypothesized that novel concepts such as cancer stem cells and microRNA could be involved in EMT. However, the involvement of EMT varies greatly among cancer types, and much remains to be learned. In this review, we present recent findings regarding the involvement of EMT in cancer progression and metastasis and provide a perspective from clinical and translational viewpoints. (Cancer Sci 2009) [source]

Signaling networks guiding epithelial,mesenchymal transitions during embryogenesis and cancer progression

CANCER SCIENCE, Issue 10 2007
Aristidis Moustakas
Epithelial,mesenchymal transition (EMT) describes the differentiation switch between polarized epithelial cells and contractile and motile mesenchymal cells, and facilitates cell movements and generation of new tissue types during embryogenesis. Many secreted polypeptides are implicated in the EMT process and their corresponding intracellular transduction pathways form highly interconnected networks. Transforming growth factor-,, Wnt, Notch and growth factors acting through tyrosine kinase receptors induce EMT and often act in a sequential manner. Such growth factors orchestrate the concerted regulation of an elaborate gene program and a complex protein network, needed for establishment of new mesenchymal phenotypes after disassembly of the main elements of epithelial architecture, such as desmosomes, as well as tight, adherens and gap junctions. EMT of tumor cells occurs during cancer progression and possibly generates cell types of the tumor stroma, such as cancer-associated myofibroblasts. EMT contributes to new tumor cell properties required for invasiveness and vascular intravasation during metastasis. Here we present some of the current mechanisms that mediate the process of EMT and discuss their relevance to cancer progression. (Cancer Sci 2007; 98: 1512,1520) [source]

Application for regenerative medicine of epithelial cell culture-vistas of cultured epithelium

CONGENITAL ANOMALIES, Issue 3 2006
Hajime Inoue
ABSTRACT This review describes culture techniques for the epithelial system as well as trends in the clinical application of cultured keratinocytes in our department and the possibility of applying the techniques to other organs. Cultured epithelium and cultured dermis in particular have considerably preceded regeneration of other organs in the field of regenerative medicine. Since 1988 we have grafted cultured keratinocytes by the Rheinwald-Green modified method in at least 500 patients with large skin defects. As a result of the establishment of a culture technique for individual patients, it is now possible to prepare enough regenerated epithelium to cover the body surface area of as many as 10 adult patients in approximately three weeks after collecting 1 cm2 of skin, and then remaining cultured keratinocytes can be cryo-preserved for two-stage dermatoplasty at another site. This procedure makes it possible to avoid frequent skin collection from the same patient and thereby improves patients' quality of life and activities of daily living. On the other hand, to solve the problem of regenerated epithelium shrinking and problems with graft efficiency on dermis defect lesion, we have developed a proteinase-resistant regenerated dermis by mixing a certain protein with a fibrin scaffold. Recently we also took the initiative in grafting hybrid-type regenerated trachea in an animal experiment by using the epithelial and dermal cell culture technique, and some results of the graft were obtained. [source]

Released nucleotides amplify the cilium-dependent, flow-induced [Ca2+]i response in MDCK cells

ACTA PHYSIOLOGICA, Issue 3 2009
H. A. Praetorius
Abstract Aim:, Changes in perfusate flow produce increases in [Ca2+]i in renal epithelial cells. Cultured renal epithelia require primary cilia to sense subtle changes in flow. In perfused kidney tubules this flow response is caused by nucleotide signalling via P2Y2 receptors. It is, however, not known whether nucleotides are released by mechanical stress applied to renal primary cilia. Here we investigate whether nucleotides are released during the cilium-dependent flow response and contribute to the flow-induced, cilium-dependent [Ca2+]i signal. Methods:, MDCK cells loaded with Fluo-4-AM were observed at 37 °C in semi-open single or closed-double perfusion chambers. Results:, Our data suggest a purinergic component of the cilium-dependent flow-response: (1) ATP scavengers and P2 receptor antagonists reduced (55%) the cilium-dependent flow-response; (2) ATP added at subthreshold concentration sensitized the renal epithelia to flow changes; (3) increases in fluid flow transiently enhanced the ATP concentration in the superfusate (measured by biosensor-cells). To test if nucleotides were released in sufficient quantities to stimulate renal epithelia we used non-confluent MDCK cells without cilia as reporter cells. We confirmed that non-confluent cells do not respond to changes in fluid flow. Placing confluent, ciliated cells upstream in the in-flow path of the non-confluent cells made them responsive to fluid flow changes. This phenomenon was not observed if either non-confluent or de-ciliated confluent cells were placed upstream. The [Ca2+]i -response in the non-confluent cells with ciliated cells upstream was abolished by apyrase and suramin. Conclusion:, This suggests that subtle flow changes sensed by the primary cilium induces nucleotide release, which amplifies the epithelial [Ca2+]i -response. [source]

Effects of normobaric hyperoxia on water content in different organs in rats

ACTA PHYSIOLOGICA, Issue 1 2002
L. E. B. Stuhr
ABSTRACT Pulmonary oxygen toxicity is a dose-dependent effect on alveolar epithelial and endothelial cells resulting in pulmonary oedema. Any concomitant effects on systemic capillary endothelium would be expected to result in capillary leakage and an increase in the tissues' water content. Total tissue water (TTW) in different organs was therefore studied in freely moving rats exposed to 100% O2 at normobaric pressure for 24 or 48 h, and compared to air-breathing control rats. The TTW for the following tissues was measured: Trachea, left bronchus, left lung, left and right ventricle, left kidney, skin (left paw-hindlimb), skin (back of the rat), left brain, left eye and thigh muscle left side. There was a significant increase in TTW of the lung accompanied by pleural effusion after 48 h of oxygen exposure as expected in all exposed animals. There was a small increase in TTW of the paw only, and a small decrease or no change in other tissues after 24 and 48 h of exposure. We conclude that there is no evidence of systemic capillary dysfunction as measured by tissue water content after exposure to hyperoxia in a dosage causing pulmonary oedema. [source]

Actin-dependent motility of melanosomes from fish retinal pigment epithelial (RPE) cells investigated using in vitro motility assays

CYTOSKELETON, Issue 2 2004
E. L. McNeil
Melanosomes (pigment granules) within retinal pigment epithelial (RPE) cells of fish and amphibians undergo massive migrations in response to light conditions to control light flux to the retina. Previous research has shown that melanosome motility within apical projections of dissociated fish RPE cells requires an intact actin cytoskeleton, but the mechanisms and motors involved in melanosome transport in RPE have not been identified. Two in vitro motility assays, the Nitella assay and the sliding filament assay, were used to characterize actin-dependent motor activity of RPE melanosomes. Melanosomes applied to dissected filets of the Characean alga, Nitella, moved along actin cables at a mean rate of 2 ,m/min, similar to the rate of melanosome motility in dissociated, cultured RPE cells. Path lengths of motile melanosomes ranged from 9 to 37 ,m. Melanosome motility in the sliding filament assay was much more variable, ranging from 0.4,33 ,m/min; 70% of velocities ranged from 1,15 ,m/min. Latex beads coated with skeletal muscle myosin II and added to Nitella filets moved in the same direction as RPE melanosomes, indicating that the motility is barbed-end directed. Immunoblotting using antibodies against myosin VIIa and rab27a revealed that both proteins are enriched on melanosome membranes, suggesting that they could play a role in melanosome transport within apical projections of fish RPE. Cell Motil. Cytoskeleton 58:71,82, 2004. © 2004 Wiley-Liss, Inc. [source]

Drosophila multiplexin (Dmp) modulates motor axon pathfinding accuracy

DEVELOPMENT GROWTH & DIFFERENTIATION, Issue 5 2009
Frauke Meyer
Multiplexins are multidomain collagens typically composed of an N-terminal thrombospondin-related domain, an interrupted triple helix and a C-terminal endostatin domain. They feature a clear regulatory function in the development of different tissues, which is chiefly conveyed by the endostatin domain. This domain can be found in proteolytically released monomeric and trimeric versions, and their diverse and opposed effects on the migratory behavior of epithelial and endothelial cell types have been demonstrated in cell culture experiments. The only Drosophila multiplexin displays specific features of both vertebrate multiplexins, collagens XV and XVIII. We characterized the Drosophila multiplexin (dmp) gene and found that three main isoforms are expressed from it, one of which is the monomeric endostatin version. Generation of dmp deletion alleles revealed that Dmp plays a role in motor axon pathfinding, as the mutants exhibit ventral bypass defects of the intersegmental nerve b (ISNb) similar to other motor axon guidance mutants. Transgenic overexpression of monomeric endostatin as well as of full-length Dmp, but not trimeric endostatin, were able to rescue these defects. In contrast, trimeric endostatin increased axon pathfinding accuracy in wild type background. We conclude that Dmp plays a modulating role in motor axon pathfinding and may be part of a buffering system that functions to avoid innervation errors. [source]

Retinoic acid affects craniofacial patterning by changing Fgf8 expression in the pharyngeal ectoderm

DEVELOPMENT GROWTH & DIFFERENTIATION, Issue 9 2008
Makoto Abe
Retinoic acid signaling plays important roles in establishing normal patterning and cellular differentiation during embryonic development. In this study, we show that single administration of retinoic acid at embryonic day 8.5 causes homeotic transformation of the lower jaw into upper jaw-like structures. This homeosis was preceded by downregulation of Fgf8 and Sprouty expression in the proximal domain of the first pharyngeal arch. Downregulation of mesenchymal genes such as Dlx5, Hand2, Tbx1 and Pitx2 was also observed. The oropharynx in retinoic acid-treated embryos was severely constricted. Consistent with this observation, Patched expression in the arch endoderm and mesenchyme was downregulated. Thus, retinoic acid affects the expression of subsets of epithelial and mesenchymal genes, possibly disrupting the regional identity of the pharyngeal arch. [source]

MMP-2 contributes to the development of the mouse ventral prostate by impacting epithelial growth and morphogenesis

DEVELOPMENTAL DYNAMICS, Issue 9 2010
Alexandre Bruni-Cardoso
Abstract Epithelial growth, branching, and canalization are important morphogenetic events of the rodent ventral prostate (VP) that take place during the first postnatal week. In this study, we evaluated the effect of knocking out MMP-2 (MMP-2,/,), by examining developmental and structural aspects of the VP in MMP-2,/, mice. Neonate (day 6) MMP-2,/, mice showed fewer epithelial tips, a lower epithelial cell proliferation rate, and also reticulin fiber accumulation. The VP of adult MMP-2,/, mice showed lower relative weight, smaller epithelial and smooth-muscle cell volume, and a larger amount of thicker reticulin fibers. No differences in cell proliferation or apoptotic index were noted between adult MMP-2,/, and wild-type mice. MMP-9 was found in the adult MMP-2,/,, but not in the wild-type. In conclusion, MMP-2 function is essential for the epithelial morphogenesis of the mouse VP, and expression of MMP-9 is not sufficient for acquisition of the normal adult histology. Developmental Dynamics 239:2386,2392, 2010. © 2010 Wiley-Liss, Inc. [source]

Manipulating gene activity in Wnt1-expressing precursors of neural epithelial and neural crest cells

DEVELOPMENTAL DYNAMICS, Issue 1 2010
Wei Hsu
Abstract Targeted gene disruption or expression often encounters lethality. Conditional approaches, permitting manipulation at desired stages, are required to overcome this problem in order to analyze gene function in later developmental processes. Wnt1 has been shown to be expressed in neural crest precursors at the dorsal midline region. However, its expression was not detected in emigrated neural crest cells, the descendants of Wnt1-expressing precursors. We have developed mouse transgenic systems to manipulate gene activity in the Wnt1-expressing precursors and their derivatives by integrating the tetracycline-dependent activation and Cre-mediated recombination methods. A new Wnt1-rtTA strain, carrying rtTA under control of Wnt1 regulatory elements, has been created for gene manipulation in a spatiotemporal-specific fashion. Together with our previously developed Wnt1-Cre;R26STOPrtTA model, these systems permit conditional gene expression and ablation in pre-migratory and/or post-migratory neural crest cells. This study demonstrated the versatility of our mouse models to achieve gene manipulation in early neural development. Developmental Dynamics 239:338,345, 2010. © 2009 Wiley-Liss, Inc. [source]

Wnt6 expression in epidermis and epithelial tissues during Xenopus organogenesis

DEVELOPMENTAL DYNAMICS, Issue 3 2008
Danielle L. Lavery
Abstract Here, we report the localization within embryonic tissues of xWnt6 protein; together with the temporal and spatial expression of Xenopus laevis Wnt6 mRNA. Wnt6 expression in Xenopus embryos is low until later stages of neurulation, when it is predominantly found in the surface ectoderm. Wnt6 expression increases during early organogenesis in the epidermis overlaying several developing organs, including the eye, heart, and pronephros. At later stages of development, Wnt6 mRNA and protein generally localize in epithelial tissues and specifically within the epithelial tissues of these developing organs. Wnt6 localization correlates closely with sites of both epithelial to mesenchymal transformations and mesenchymal to epithelial transformations. Xenopus Wnt6 sequence and its expression pattern are highly conserved with other vertebrates. Xenopus embryos, therefore, provide an excellent model system for investigating the function of vertebrate Wnt6 in organ development and regulation of tissue architecture. Developmental Dynamics 237:768,779, 2008. © 2008 Wiley-Liss, Inc. [source]

Shh/BMP-4 signaling pathway is essential for intestinal epithelial development during Xenopus larval-to-adult remodeling

DEVELOPMENTAL DYNAMICS, Issue 12 2006
Atsuko Ishizuya-Oka
Abstract During amphibian larval-to-adult intestinal remodeling, progenitor cells of the adult epithelium actively proliferate and differentiate under the control of thyroid hormone (TH) to form the intestinal absorptive epithelium, which is analogous to the mammalian counterpart. We previously found that TH,up-regulated expression of bone morphogenetic protein-4 (BMP-4) spatiotemporally correlates with adult epithelial development in the Xenopus laevis intestine. Here, we aimed to clarify the role of BMP-4 in intestinal remodeling. Our reverse transcriptase-polymerase chain reaction and in situ hybridization analyses indicated that mRNA of BMPR-IA, a type I receptor of BMP-4, is expressed in both the developing connective tissue and progenitor cells of the adult epithelium. More importantly, using organ culture and immunohistochemical procedures, we have shown that BMP-4 not only represses cell proliferation of the connective tissue but promotes differentiation of the intestinal absorptive epithelium. In addition, we found that the connective tissue-specific expression of BMP-4 mRNA is up-regulated by sonic hedgehog (Shh), whose epithelium-specific expression is directly induced by TH. These results strongly suggest that the Shh/BMP-4 signaling pathway plays key roles in the amphibian intestinal remodeling through epithelial,connective tissue interactions. Developmental Dynamics 235:3240,3249, 2006. © 2006 Wiley-Liss, Inc. [source]

Characterization of Bves expression during mouse development using newly generated immunoreagents

DEVELOPMENTAL DYNAMICS, Issue 6 2006
Travis K. Smith
Abstract Bves (blood vessel/epicardial substance) is a transmembrane protein postulated to play a role in cell,cell interaction/adhesion. It was independently isolated by two groups as a gene product highly enriched in the developing heart. Disagreement exists about its expression during development. Most notably, the expression of Bves in non-muscle cells is disputed. Determining the expression profile of Bves is a critical initial step preceding the characterization of protein function in development and in the adult. We have generated new monoclonal antibodies against mouse Bves and used these immunoreagents to elucidate Bves expression in development. As expected, we detect Bves in myocytes of the developing heart throughout development. In addition, skeletal and smooth muscle cells including those of the coronary system express Bves. Finally, specific, but not all, epithelial derivatives of the three germ layers are stained positively with these monoclonal antibodies. Protein expression in cultured epithelial and muscle cell lines corroborate our in vivo findings. Taken together, these results demonstrate the expression of Bves in a wide range of epithelial and muscle cells during mouse embryogenesis and indicate a broad function for this protein in development, and show that these newly generated reagents will be invaluable in further investigation of Bves. Developmental Dynamics 235:1701,1708, 2006. © 2006 Wiley-Liss, Inc. [source]

Skeletal elements in the vertebrate eye and adnexa: Morphological and developmental perspectives

DEVELOPMENTAL DYNAMICS, Issue 5 2006
Tamara A. Franz-Odendaal
Abstract Although poorly appreciated, the vertebrate eye and adnexa are relatively common sites for skeletogenesis. In many taxa, the skeleton contributes to internal reinforcement in addition to the external housing of the eye (e.g., the circumorbital bones and eyelids). Eyeball elements such as scleral cartilage and scleral ossicles are present within a broad diversity of vertebrates, albeit not therian mammals, and have been used as important models for the study of condensations and epithelial,mesenchymal interactions. In contrast, other elements invested within the eye or its close surroundings remain largely unexplored. The onset and mode of development of these skeletal elements are often variable (early versus late; involving chondrogenesis, osteogenesis, or both), and most (if not all) of these elements appear to share a common neural crest origin. This review discusses the development and distribution of the skeletal elements within and associated with the developing eye and comments on homology of the elements where these are questionable. Developmental Dynamics 235:1244,1255, 2006. © 2006 Wiley-Liss, Inc. [source]

In vivo and in vitro analysis of the vasculogenic potential of avian proepicardial and epicardial cells,

DEVELOPMENTAL DYNAMICS, Issue 4 2006
Juan A. Guadix
Abstract Coronary vessel formation is a special case in the context of embryonic vascular development. A major part of the coronary cellular precursors (endothelial, smooth muscle, and fibroblastic cells) derive from the proepicardium and the epicardium in what can be regarded as a late event of angioblastic and smooth muscle cell differentiation. Thus, coronary morphogenesis is dependent on the epithelial,mesenchymal transformation of the proepicardium and the epicardium. In this study, we present several novel observations about the process of coronary vasculogenesis in avian embryos, namely: (1) The proepicardium displays a high vasculogenic potential, both in vivo (as shown by heterotopic transplants) and in vitro, which is modulated by vascular endothelial growth factor (VEGF) and basic fibroblast growth factor signals; (2) Proepicardial and epicardial cells co-express receptors for platelet-derived growth factor-BB and VEGF; (3) Coronary angioblasts (found all through the epicardial, subepicardial, and compact myocardial layers) express the Wilms' tumor associated transcription factor and the retinoic acid-synthesizing enzyme retinaldehyde-dehydrogenase-2, two markers of the coelomic epithelium involved in coronary endothelium development. All these results contribute to the development of our knowledge on the vascular potential of proepicardial/epicardial cells, the existent interrelationships between the differentiating coronary cell lineages, and the molecular mechanisms involved in the regulation of coronary morphogenesis. Developmental Dynamics 235:1014,1026, 2006. © 2006 Wiley-Liss, Inc. [source]

PDGFR-, signaling is critical for tooth cusp and palate morphogenesis

DEVELOPMENTAL DYNAMICS, Issue 1 2005
Xun Xu
Abstract Platelet-derived growth factor receptor alpha (PDGFR-,) and PDGF ligands are key regulators for embryonic development. Although Pdgfr, is spatially expressed in the cranial neural crest (CNC)-derived odontogenic mesenchyme, mice deficient for Pdgfr, are embryonic lethal, making it impossible to investigate the functional significance of PDGF signaling in regulating the fate of CNC cells during tooth morphogenesis. Taking advantage of the kidney capsule assay, we investigated the biological function of PDGF signaling in regulating tooth morphogenesis. Pdgfr, and Pdgfa are specifically and consistently expressed in the CNC-derived odontogenic mesenchyme and the dental epithelium, respectively, throughout all stages of tooth development, suggesting a paracrine function of PDGF signaling in regulating tooth morphogenesis. Highly concentrated expression patterns of Pdgfr, and Pdgfa are associated with the developing dental cusp, suggesting possible functional importance of PDGF signaling in regulating cusp formation. Loss of the Pdgfr, gene does not affect proper odontoblasts proliferation and differentiation in the CNC-derived odontogenic mesenchyme but perturbs the formation of extracellular matrix and the organization of odontoblast cells at the forming cusp area, resulting in dental cusp growth defect. Pdgfr,,/, mice have complete cleft palate. We show that the cleft palate in Pdgfr, mutant mice results from an extracellular matrix defect within the CNC-derived palatal mesenchyme. The midline epithelium of the mutant palatal shelf remains functionally competent to mediate palatal fusion once the palatal shelves are placed in close contact in vitro. Collectively, our data suggests that PDGFR, and PDGFA are critical regulators for the continued epithelial,mesenchymal interaction during tooth and palate morphogenesis. Disruption of PDGFR, signaling disturbs the growth of dental cusp and interferes with the critical extension of palatal shelf during craniofacial development. Developmental Dynamics 232:75,84, 2005. © 2004 Wiley-Liss, Inc. [source]

Ventral otic cell lines as developmental models of auditory epithelial and neural precursors

DEVELOPMENTAL DYNAMICS, Issue 4 2004
G. Lawoko-Kerali
Abstract Conditionally immortal cell lines were established from the ventral otocyst of the Immortomouse at embryonic day 10.5 and selected to represent precursors of auditory sensory neural and epithelial cells. Selection was based upon dissection, tissue-specific markers, and expression of the transcription factor GATA3. Two cell lines expressed GATA3 but possessed intrinsically different genetic programs under differentiating conditions. US/VOT-E36 represented epithelial progenitors with potential to differentiate into sensory and nonsensory epithelial cells. US/VOT-N33 represented migrating neuroblasts. Under differentiating conditions in vitro the cell lines expressed very different gene expression profiles. Expression of several cell- and tissue-specific markers, including the transcription factors Pax2, GATA3, and NeuroD, differed between the cell lines in a pattern consistent with that observed between their counterparts in vivo. We suggest that these and other conditionally immortal cell lines can be used to study transient events in development against different backgrounds of cell competence. Developmental Dynamics 231:801,814, 2004. © 2004 Wiley-Liss, Inc. [source]