Expression and functional involvement of N-cadherin in embryonic limb chondrogenesis

Development ◽  
1994 ◽  
Vol 120 (1) ◽  
pp. 177-187 ◽  
Author(s):  
S.A. Oberlender ◽  
R.S. Tuan
Keyword(s):  
Pattern Formation ◽  
Cell Aggregation ◽  
Mesenchymal Cells ◽  
Limb Bud ◽  
Time Period ◽  
Functional Involvement ◽  
Embryonic Limb ◽  
Cellular Condensation

Cell adhesion molecules have been shown to be important mediators of morphogenesis and pattern formation. In this study, we have shown that N-cadherin is expressed in a specific spatiotemporal manner in the developing limb bud during chondrogenesis in vivo and in cultured limb mesenchyme in vitro. The time period of maximal expression of N-cadherin corresponds to the period of active cellular condensation, an event believed to be a necessary prerequisite for chondrogenic differentiation. To directly assess the functional involvement of N-cadherin in cellular condensation, we have examined the effects of perturbing N-cadherin activity on both cell aggregation and chondrogenesis using NCD-2, a rat monoclonal antibody directed against the binding region of N-cadherin. Non-immune rat IgG was used as a control. Our results show that functional N-cadherin is necessary for chondrogenesis to proceed both in vivo and in vitro. Limb mesenchymal cells exhibited characteristic Ca(2+)-dependent cell aggregation in suspension, which was inhibited in the presence of exogenous NCD-2. In micromass cultures of limb mesenchymal cells, NCD-2 inhibited overt chondrogenesis in a dose-dependent manner. Furthermore, NCD-2 inhibition of chondrogenesis in micromass cultures was time-dependent, suggesting that N-cadherin is crucially involved during the latter half of the first 24 hours of culture, a time period most likely corresponding to active cellular condensation. NCD-2 also significantly influenced limb development when injected into embryonic limb buds in vivo. In addition to significant inhibition of chondrogenesis and developmental delays, gross developmental deformities and perturbation of overall pattern formation were also observed. Taken together, these results demonstrate that N-cadherin is functionally required in mediating the cell-cell interactions among mesenchymal cells important for chondrogenesis in micromass culture in vitro and in the intact limb bud in vivo.

FGF-4 maintains polarizing activity of posterior limb bud cells in vivo and in vitro

Development ◽  
1993 ◽  
Vol 119 (1) ◽  
pp. 199-206 ◽  
Author(s):  
A. Vogel ◽  
C. Tickle
Keyword(s):  
Posterior Margin ◽  
Anterior Margin ◽  
Marked Decrease ◽  
Mesenchymal Cells ◽  
Apical Ectodermal Ridge ◽  
Limb Bud ◽  
Posterior Limb ◽  
Vertebrate Limb

The polarizing region is a major signalling tissue involved in patterning the tissues of the vertebrate limb. The polarizing region is located at the posterior margin of the limb bud and can be recognized by its ability to induce additional digits when grafted to the anterior margin of a chick limb bud. The signal from the polarizing region operates at the tip of the bud in the progress zone, a zone of undifferentiated mesenchymal cells, maintained by interactions with the apical ectodermal ridge. A number of observations have pointed to a link between the apical ectodermal ridge and signalling by the polarizing region. To test this possibility, we removed the posterior apical ectodermal ridge of chick wing buds and assayed posterior mesenchyme for polarizing activity. When the apical ectodermal ridge is removed, there is a marked decrease in polarizing activity of posterior cells. The posterior apical ectodermal ridge is known to express FGF-4 and we show that the decrease in polarizing activity of posterior cells of wing buds that normally follows ridge removal can be prevented by implanting a FGF-4-soaked bead. Furthermore, we show that both ectoderm and FGF-4 maintain polarizing activity of limb bud cells in culture.

Vitamin A alters the internal viscosity of fragments of limb-bud mesenchyme

Development ◽  
1980 ◽  
Vol 59 (1) ◽  
pp. 325-339
Author(s):  
T. E. Kwasigroch ◽  
D. M. Kochhar
Keyword(s):  
Retinoic Acid ◽  
Vitamin A ◽  
Mesenchymal Cells ◽  
Limb Bud ◽  
Mouse Limb ◽  
Vitamin A Acid ◽  
Internal Viscosity ◽  
Fusion Experiments

Two techniques were used to examine the effect of vitamin A compounds (vitamin A acid = retinoic acid and vitamin A acetate) upon the relative strengths of adhesion among mouse limb-bud mesenchymal cells. Treatment with retinoic acid in vivo and with vitamin A acetate in vitro reduced the rate at which the fragments of mesenchyme rounded-up when cultured on a non-adhesive substratum, but these compounds did not alter the behavior of tissues tested in fragment-fusion experiments. These conflicting results indicate that the two tests measure different activities of cells and suggest that treatment with vitamin A alters the property(ies) of cells which regulate the internal viscosity of tissues.

scholarly journals Effects of pomegranate extracts on cartilage, bone and mesenchymal cells of mouse fetuses

2011 ◽  
Vol 107 (5) ◽  
pp. 683-690 ◽  
Author(s):  
Malihezaman Monsefi ◽  
Fatemeh Parvin ◽  
Tahereh Talaei-Khozani
Keyword(s):  
Mesenchymal Cells ◽  
Limb Bud ◽  
Pomegranate Juice ◽  
Alizarin Red ◽  
Cell Proliferation And Differentiation ◽  
Pregnant Mice ◽  
Prevention Of Bone Loss ◽  
Mouse Fetuses

Pomegranate is a rich source of polyphenols, which are believed to be responsible for the oestrogenic activities of extracts of this fruit in mice. One of these potential activities is the prevention of bone loss. The objectives of the present study were to determine the effects of pomegranate extract on chondrogenesis and osteogenesis in mouse embryos in vivo and limb bud cultures in vitro. A total of fifty pregnant Balb/c mice were given vehicle, pomegranate juice extract (PJE), pomegranate husk extract (PHE) or a mixture of husk and juice extract (PME). Their embryos were stained with alizarin red S and alcian blue, and the length of the femur, tibia and their ossification zones were measured on day 19 of gestation. Bone Ca content in pregnant mice was also measured. Mice treated with PJE showed an increase in bone Ca content. Dietary supplementation with all extracts significantly increased embryo femur length and osteogenesis index. Mesenchymal cells from fetal limb buds were cultured and exposed to 10, 100, 1000 and 10 000 μg/ml of PJE, PHE or PME. The number of viable cells was greater in cultures exposed to the extracts than in control cultures. The number of cartilage nodules and their diameters were greater in extract-treated cell cultures, a finding which reflected increased cell proliferation and differentiation rates. In conclusion, the findings of the present study suggest that pomegranate is able to enhance bone formation.

Pattern formation in dissociated limb bud mesenchyme in vitro and in vivo

1998 ◽  
Vol 6 (4) ◽  
pp. S-398-S-402 ◽  
Author(s):  
Hiroyuki Ide ◽  
Hitoshi Yokoyama ◽  
Tetsuya Endo ◽  
Minoru Omi ◽  
Koji Tamura ◽  
...  
Keyword(s):  
Pattern Formation ◽  
Limb Bud

scholarly journals Experimental studies on the differentiation of embryonic tissues growing in vivo and in vitro .—I. The development of the undifferentiated limb-bud (a) when subcutaneously grafted into the post-embryonic chick and (b) when cultivated in vitro

1926 ◽  
Vol 99 (698) ◽  
pp. 340-366 ◽  
Keyword(s):  
Preliminary Investigation ◽  
Experimental Studies ◽  
Limb Bud ◽  
Posterior Limb ◽  
Embryonic Tissues ◽  
Embryonic Limb ◽  
Almost All ◽  
Normal Standards

One method by which the problem of the differentiation of animal tissues may be approached is by studying the behaviour of simple embryonic tissues when growing in an abnormal environment, such as that produced by grafting into atypical situations in vivo or by cultivation in vitro . It is along these lines that the investigations of the present writers are being conducted. The work so far completed, the results of which are recorded in the present communication, consists of a study of the development of the undifferentiated, embryonic limb-bud of the fowl when grafted subcutaneously into a postembryonic chick and when cultivated vitro. A preliminary investigation of the histogenesis of cartilage and bone in the limbs of the embryonic fowl was carried out by one of the writers (Fell, 1925) in order to provide normal standards with which to compare the experimental material. Rous (1910, 1911), Fichera (1909) and many others have successfully grafted foœtal and embryonic tissues into young and adult animals, usually in connection with the study of tumour growth ; a bibliography and summary of the earlier work is given in Fichera’s paper. Almost all the work on the development of grafts of the undifferentiated limb-buds has been carried out on the embryonic Amphibia by Braus, Harrison (1907, 1918, 1921), Detweiler (1918, 1925), Nicholas (1924) and others. Spurting (1923) describes a case of accidental but successful autotransplantation of the posterior limb-bud in a fowl embryo. Murray and Huxley (1925) record two experiments in which part of the limb-bud of a four-days’ embryo was successfully grafted on to the chorioallantoic membranes ; in one case “ a highly differentiated and very easily recognizable femur ” showing early ossification was found after 5 days’ growth.

MESENCHYMAL TNFR1 EXPRESSION PRESERVES THE COLONIC EPITHELIAL STEM CELL NICHE

2021 ◽  
Vol 27 (Supplement_1) ◽  
pp. S7-S8
Author(s):  
Safina Gadeock ◽  
Cambrian Liu ◽  
Brent Polk
Keyword(s):  
Stem Cell ◽  
Stem Cell Niche ◽  
Mesenchymal Cells ◽  
Epithelial Stem Cells ◽  
Epithelial Stem Cell ◽  
Long Term Treatment ◽  
Lgr5 Expression ◽  
Cell Niche

Abstract Tumor necrosis factor (TNF) is a highly expressed cytokine in inflammatory bowel disease (IBD). Although TNF can induce colonic epithelial dysfunction and apoptosis, recent studies suggest that TNF signalling promotes epithelial wound repair and stem cell function. Here we investigated the role of TNF receptor 1 (TNFR1) in mediating TNF’s effects on colonic epithelial stem cells, integral to mucosal healing in colitis. We demonstrate that Tnfr1-/- mice exhibit loss in Lgr5 expression (-52%, p<0.02; N=6) compared to wildtype (WT) controls. However, the opposite result was found in vitro, wherein murine Tnfr1-/- colonoids demonstrated a significant increase in Lgr5 expression (66%, p<0.007; N=6) compared to WT colonoids. Similarly, human colonoids treated with an anti-TNFR1 antibody also demonstrated an increase in Lgr5 expression, relative to IgG controls. To resolve the contradiction in the in vivo versus in vitro environment, we hypothesized that mesenchymal TNFR1 expression regulates the epithelial stem cell niche. To determine the relationships between these cell types, we co-cultured WT or Tnfr1-/- colonoids with WT or Tnfr1-/- colonic myofibroblasts (CMFs). We found that epithelial Lgr5 expression was significantly higher (by 52%, p<0.05; N=3) when co-cultured with WT compared to TNFR1-/- myofibroblasts. The loss of TNFR1 expression in vivo increases the number of αSMA+ mesenchymal cells by nearly 56% (N=6) but considerably reduces the pericryptal PDGFRα+ cells, suggesting modifications in mesenchymal populations that contribute to the epithelial stem cell niche. Functionally, primary Tnfr1-/--CMFs displayed PI3k (p<0.001; N=3) and MAPK (p<0.01; N=3)-dependent increases in migration, proliferation, and differentiation, but RNA profiling demonstrated by diminished levels of stem cell niche factors, Rspo3 (-80%, p<0.0001; N=6) and Wnt2b (-63%, p<0.008; N=6) compared to WT-CMFs. Supplementation with 50ng recombinant Rspo3 for 5 d to Lgr5-GFP organoids co-cultured with TNFR1-/--CMFs restored Lgr5 expression to wildtype levels. Therefore, TNFR1-mediated TNF signalling in mesenchymal cells promotes their ability to support an epithelial stem cell niche. These results should motivate future studies of the stem cell niche in the context of long-term treatment with anti-TNF therapies.

scholarly journals Experimental studies on the differentiation of embryonic tissues growing in vivo and in vitro .—II. The development of the isolated early embryonic eye of the fowl when cultivated in vitro

1926 ◽  
Vol 100 (703) ◽  
pp. 273-283 ◽  
Keyword(s):  
Medical Research ◽  
Research Council ◽  
Medical Research Council ◽  
Experimental Studies ◽  
Limb Bud ◽  
Progressive Development ◽  
Embryonic Tissues ◽  
Self Differentiation

In a previous communication (Strangeways and Fell, 1926) it was shown that if the undifferentiated limb-bud of the embryonic Fowl was cultivated in vitro , it underwent a considerable amount of progressive development. This capacity for independent development in vitro possessed by an isolated organ has been further investigated, and for these later experiments the writers have employed the early embryonic eye, a structure endowed with more complex potentialities than the limb-bud. As a result of these experiments it was found that the eyes of young Fowl embryos possess, in a remarkable degree, the faculty for self-differentiation in vitro and for “organotypic” growth as defined by Maximow (1925). The previous work on organotypic growth in vitro has already been briefly outlined in the writers’ earlier paper and need not be discussed here. The expenses connected with the experiments described in this communication were met by the Medical Research Council, to whom the writers desire to express their thanks.

scholarly journals Adhesion molecules during somitogenesis in the avian embryo.

1987 ◽  
Vol 104 (5) ◽  
pp. 1361-1374 ◽  
Author(s):  
J L Duband ◽  
S Dufour ◽  
K Hatta ◽  
M Takeichi ◽  
G M Edelman ◽  
...  
Keyword(s):  
Adhesion Molecules ◽  
Tissue Remodeling ◽  
Cell Aggregation ◽  
Avian Embryo ◽  
Primary Role ◽  
Calcium Dependent ◽  
Tissue Dissociation ◽  
Spatio Temporal

In avian embryos, somites constitute the morphological unit of the metameric pattern. Somites are epithelia formed from a mesenchyme, the segmental plate, and are subsequently reorganized into dermatome, myotome, and sclerotome. In this study, we used somitogenesis as a basis to examine tissue remodeling during early vertebrate morphogenesis. Particular emphasis was put on the distribution and possible complementary roles of adhesion-promoting molecules, neural cell adhesion molecule (N-CAM), N-cadherin, fibronectin, and laminin. Both segmental plate and somitic cells exhibited in vitro calcium-dependent and calcium-independent systems of cell aggregation that could be inhibited respectively by anti-N-cadherin and anti-N-CAM antibodies. In vivo, the spatio-temporal expression of N-cadherin was closely associated with both the formation and local disruption of the somites. In contrast, changes in the prevalence of N-CAM did not strictly accompany the remodeling of the somitic epithelium into dermamyotome and sclerotome. It was also observed that fibronectin and laminin were reorganized secondarily in the extracellular spaces after CAM-mediated contacts were modulated. In an in vitro culture system of somites, N-cadherin was lost on individual cells released from somite explants and was reexpressed when these cells reached confluence and established intercellular contacts. In an assay of tissue dissociation in vitro, antibodies to N-cadherin or medium devoid of calcium strongly and reversibly dissociated explants of segmental plates and somites. Antibodies to N-CAM exhibited a smaller disrupting effect only on segmental plate explants. In contrast, antibodies to fibronectin and laminin did not perturb the cohesion of cells within the explants. These results emphasize the possible role of cell surface modulation of CAMs during the formation and remodeling of some transient embryonic epithelia. It is suggested that N-cadherin plays a major role in the control of tissue remodeling, a process in which N-CAM is also involved but to a lesser extent. The substratum adhesion molecules, fibronectin and laminin, do not appear to play a primary role in the regulation of these processes but may participate in cell positioning and in the stabilization of the epithelial structures.

Detection of Circulating Immune Complexes in the Sera of Rabbits with Experimental Syphilis: Possible Role in Immunoregulation

1980 ◽  
Vol 29 (2) ◽  
pp. 575-582
Author(s):  
Robert E. Baughn ◽  
Kenneth S. K. Tung ◽  
Daniel M. Musher
Keyword(s):  
Immune Complexes ◽  
Proteolytic Enzymes ◽  
Suppressor Cell ◽  
Suppressive Effect ◽  
Circulating Immune Complexes ◽  
Time Period ◽  
Infected Cells ◽  
Disseminated Disease

The in vivo and in vitro immunoglobulin G plaque-forming cell responses to sheep erythrocytes (SRBC) are nearly obliterated during disseminated syphilitic infection (3 to 8 weeks post-intravenous injection) in rabbits. Splenic and lymph node cells obtained from infected rabbits during this time period were capable of suppressing the normal in vitro responses of uninfected, SRBC-primed cells. Cell-free washings of cells from infected animals were also suppressive. This finding coupled with the fact that treatment of infected cells with proteolytic enzymes abrogated the suppressive effect constitute arguments against involvement of a specific suppressor cell population. The incidence of elevated levels of circulating immune complexes in the sera of rabbits with disseminated disease was also significantly different from that of uninfected controls or infected rabbits before the onset or after the regression of lesions. When added to cultures of lymphocytes from uninfected, SRBC-sensitized rabbits, sera containing complexes caused dose-related suppression of the in vitro immunoglobulin responses. Unlike immune complexes, no correlation was found between the presence of mucopolysaccharide materials and the stage of infection or the ability of serum to suppress the immunoglobulin responses to SRBC.

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