Patterns of cell division, cell death and chondrogenesis in cultured aggregates of normal and talpid3 mutant chick limb mesenchyme cells

Development ◽  
1972 ◽  
Vol 27 (1) ◽  
pp. 245-260
Author(s):  
D. A. Ede ◽  
O. P. Flint
Keyword(s):  
Cell Death ◽  
Cell Division ◽  
Limb Bud ◽  
Intercellular Matrix ◽  
Blue Stain ◽  
Special Pattern ◽  
Mesenchyme Cells ◽  
Division Cell ◽  
Made In

Aggregates were prepared from dissociated mesenchyme cells obtained from normal and talpid mutant chick limb buds at stage 26 and were maintained for 4 days in culture. They were shown by autoradiographic techniques to consist initially of populations of unifoimly dedifferentiated cells within which chondrogenesis was initiated between 1 and 2 days, leading to the formation of areas of precartilage in the interior of the aggregates. Measurements of cell population density, cell death and cell division were made in precartilage and non-cartilage regions on sections prepared from normal and mutant aggregates fixed at 1-day intervals and were related to the pattern of chondrogenesis. Non-cartilage areas consisted of cells surrounding the precartilage areas and extended to the surface of the aggregate; these cells showed no special pattern or histochemical reaction. Precartilage areas consisted of one or more “;condensations”, comprising cells arranged in concentric rings around a central cell or group of cells, characterized by uptake of [35S]sulphate and taking up alcian blue stain in the intercellular matrix. Chondrogenesis was initiated al the condensation foci and spread centrifugally. Condensations were arranged in a simple pattern, roughly equidistantly from each other and never at the surface of the aggregate. The shape and arrangement of the cells comprising them suggested that they were formed by a process of aggregation towards the condensation foci. The relation of these observations to events in the intact limb bud developing in vivo is discussed.

Reduction of the rate of outgrowth, cell density, and cell division following removal of the apical ectodermal ridge of the chick limb-bud

Development ◽  
1977 ◽  
Vol 40 (1) ◽  
pp. 1-21
Author(s):  
Dennis Summerbell
Keyword(s):  
Cell Proliferation ◽  
Cell Death ◽  
Cell Division ◽  
Cell Density ◽  
Apical Ectodermal Ridge ◽  
Limb Bud ◽  
Short Term ◽  
Density Dependent ◽  
Wing Bud

Removal of the apical ectodermal ridge causes a reduction in the rate of outgrowth of the wing-bud and the loss of distal parts. More specifically it causes a short-term increase in cell density and cell death and a decrease in the rate of cell proliferation. The evidence supports the hypothesis of density-dependent control of cell division and suggests that there may also be a mechanism regulating skeletal length at the time of differentiation. An informal model is presented to explain the observations.

scholarly journals In Vivo Analysis of Cell Division, Cell Growth, and Differentiation at the Shoot Apical Meristem in Arabidopsis

2003 ◽  
Vol 16 (1) ◽  
pp. 74-87 ◽  
Author(s):  
Olivier Grandjean ◽  
Teva Vernoux ◽  
Patrick Laufs ◽  
Katia Belcram ◽  
Yuki Mizukami ◽  
...  
Keyword(s):  
Cell Division ◽  
Cell Growth ◽  
Shoot Apical Meristem ◽  
Apical Meristem ◽  
Cell Growth And Differentiation ◽  
In Vivo Analysis ◽  
Division Cell

scholarly journals Fibroblast Growth Factor 4 Directs Gap Junction Expression in the Mesenchyme of the Vertebrate Limb Bud

1997 ◽  
Vol 138 (5) ◽  
pp. 1125-1137 ◽  
Author(s):  
H. Makarenkova ◽  
D.L. Becker ◽  
C. Tickle ◽  
A.E. Warner
Keyword(s):  
Growth Factor ◽  
Gap Junctions ◽  
Gap Junction ◽  
Fibroblast Growth Factor ◽  
Connexin 43 ◽  
Limb Bud ◽  
Functional Coupling ◽  
Fibroblast Growth ◽  
Mesenchyme Cells

Pattern in the developing limb depends on signaling by polarizing region mesenchyme cells, which are located at the posterior margin of the bud tip. Here we address the underlying cellular mechanisms. We show in the intact bud that connexin 43 (Cx43) and Cx32 gap junctions are at higher density between distal posterior mesenchyme cells at the tip of the bud than between either distal anterior or proximal mesenchyme cells. These gradients disappear when the apical ectodermal ridge (AER) is removed. Fibroblast growth factor 4 (FGF4) produced by posterior AER cells controls signaling by polarizing cells. We find that FGF4 doubles gap junction density and substantially improves functional coupling between cultured posterior mesenchyme cells. FGF4 has no effect on cultured anterior mesenchyme, suggesting that any effects of FGF4 on responding anterior mesenchyme cells are not mediated by a change in gap junction density or functional communication through gap junctions. In condensing mesenchyme cells, connexin expression is not affected by FGF4. We show that posterior mesenchyme cells maintained in FGF4 under conditions that increase functional coupling maintain polarizing activity at in vivo levels. Without FGF4, polarizing activity is reduced and the signaling mechanism changes. We conclude that FGF4 regulation of cell–cell communication and polarizing signaling are intimately connected.

scholarly journals Preclinical small molecule WEHI-7326 overcomes drug resistance and elicits response in patient-derived xenograft models of human treatment-refractory tumors

2021 ◽  
Vol 12 (3) ◽  
Author(s):  
Christoph Grohmann ◽  
Francesca Walker ◽  
Mark Devlin ◽  
Meng-Xiao Luo ◽  
Anderly C. Chüeh ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Cell Death ◽  
Cell Division ◽  
Side Effects ◽  
Small Molecule ◽  
Standard Of Care ◽  
Wide Range ◽  
Anti Cancer ◽  
Treatment Refractory

AbstractTargeting cell division by chemotherapy is a highly effective strategy to treat a wide range of cancers. However, there are limitations of many standard-of-care chemotherapies: undesirable drug toxicity, side-effects, resistance and high cost. New small molecules which kill a wide range of cancer subtypes, with good therapeutic window in vivo, have the potential to complement the current arsenal of anti-cancer agents and deliver improved safety profiles for cancer patients. We describe results with a new anti-cancer small molecule, WEHI-7326, which causes cell cycle arrest in G2/M, cell death in vitro, and displays efficacious anti-tumor activity in vivo. WEHI-7326 induces cell death in a broad range of cancer cell lines, including taxane-resistant cells, and inhibits growth of human colon, brain, lung, prostate and breast tumors in mice xenografts. Importantly, the compound elicits tumor responses as a single agent in patient-derived xenografts of clinically aggressive, treatment-refractory neuroblastoma, breast, lung and ovarian cancer. In combination with standard-of-care, WEHI-7326 induces a remarkable complete response in a mouse model of high-risk neuroblastoma. WEHI-7326 is mechanistically distinct from known microtubule-targeting agents and blocks cells early in mitosis to inhibit cell division, ultimately leading to apoptotic cell death. The compound is simple to produce and possesses favorable pharmacokinetic and toxicity profiles in rodents. It represents a novel class of anti-cancer therapeutics with excellent potential for further development due to the ease of synthesis, simple formulation, moderate side effects and potent in vivo activity. WEHI-7326 has the potential to complement current frontline anti-cancer drugs and to overcome drug resistance in a wide range of cancers.

scholarly journals Synergistic CDK control pathways maintain cell size homeostasis

2020 ◽  
Author(s):  
James O. Patterson ◽  
Souradeep Basu ◽  
Paul Rees ◽  
Paul Nurse
Keyword(s):  
Cell Division ◽  
Cell Size ◽  
High Throughput ◽  
Small Cells ◽  
Control Network ◽  
Size Information ◽  
Division Cell ◽  
Diploid Cells ◽  
High Cyclin

AbstractTo coordinate cell size with cell division, cell size must be computed by the cyclin-CDK control network to trigger division appropriately. Here we dissect determinants of cyclin-CDK activity using a novel high-throughput single-cell in vivo system. We show that inhibitory phosphorylation of CDK encodes cell size information and works synergistically with PP2A to prevent division in smaller cells. However, even in the absence of all canonical regulators of cyclin-CDK, small cells with high cyclin-CDK levels are restricted from dividing. We find that diploid cells of equivalent size to haploid cells exhibit lower CDK activity in response to equal cyclin-CDK enzyme concentrations, suggesting that CDK activity is reduced by DNA concentration. Thus, multiple pathways directly regulate cyclin-CDK activity to maintain robust cell size homeostasis.

Incorporation of 5-bromo-2'-deoxyuridine into mesenchymal limb-bud cells destined to die: relationship to polydactyly induction in rats

Development ◽  
1982 ◽  
Vol 72 (1) ◽  
pp. 125-141
Author(s):  
L. David Wise ◽  
William J. Scott
Keyword(s):  
Cell Death ◽  
Limb Bud ◽  
Optimal Time ◽  
Dead Cell ◽  
Temporal Response ◽  
Pregnant Rats ◽  
Near Term ◽  
Drug Incorporation ◽  
Cell Fragments

The thymidine analogue, 5-bromo-2'-deoxyuridine (BUdR), given at the proper dose and time to pregnant rats produces preaxial hindlimb polydactyly in a high proportion of near term foetuses. The lack of physiological cell death in an area of preaxial mesenchyme known as the foyer primaire preaxial (fpp) is thought to be important in the pathogenesis of this defect. This study addresses the question of whether BUdR's well-known antidifferentiative effects, which are due in some way to drug incorporation into DNA, are operative in this in vivo system. The dose and temporal response of BUdR for the induction of preaxial polydactyly inversely parallels the frequency of embryonic hindlimbs with an fpp. Incorporation of BUdR into degenerative fragments within the fpp of these treated limbs is demonstrated with indirect immunofiuorescence using an antibody to bromouridine. Hindlimbs exposed to a threshold dose of BUdR at the optimal time for producing polydactyly have incorporated the drug into degenerative fragments within the fpp. This suggests that a higher, teratogenic dose of BUdR might likewise be incorporated. The resulting higher level of incorporation presumably alters the normal course of terminal differentiation for these cells originally destined to die. Teratogenic doses of BUdR injected at later than the optimal time are also incorporated into dead cell fragments within the fpp, suggesting that presumptive dead cells have additional rounds of DNA synthesis which are BUdR-insensitive. Approximately 12 h prior to overt death presumptive fpp cells no longer incorporate the drug. Results reported support the hypothesis that incorporation of teratogenic levels of BUdR prevent cell death in the fpp. The extra cells are thought to contribute directly or indirectly to the added digit. Contrary to other views, it is suggested that BUdR-induced teratogenesis can be a result of the drug's antidifferentiative effects on specific, ‘sensitive’, populations of cells.

scholarly journals The Relations between Cell Division and Cell Type Specification in Floral and Vegetative Meristems of Tomato and Arabidopsis

1996 ◽  
Author(s):  
Eliezer Lifschitz ◽  
Elliot Meyerowitz
Keyword(s):  
Cell Proliferation ◽  
Cell Division ◽  
Cell Fate ◽  
Regulatory Gene ◽  
Mads Box ◽  
Mads Box Genes ◽  
Threonine Deaminase ◽  
Division Cell ◽  
Floral Meristems

Meristems were the central issue of our project. Genes that are required for cell division, cell elongation, cell proliferation and cell fate were studied in the tomato system. The analysis of the dUTPase and threonine deaminase genes, along with the dissection of their regulatory regions is completed, while that of the RNR2 and PPO genes is at an advanced stage. All these genes were isolated in our laboratory. In addition, 8 different MADS box genes were studied in transgenic plants and their genetic relevances discovered. We have also shown that a given MADS box gene can modify the polarity of cell division without affecting the fate of the organ. In vivo interaction between two MADS box genes was demonstrated and the functional dependency of the tomato agamous gene on the TM5 gene product established. We have exploited the Knotted1 meristematic gene in conjunction with tomato leaf meristematic genes to show that simple and compound leaves and, for that matter, sepals and compound leaves, are formed by two different developmental programs. In this context we have also isolated and characterized the tomato Knotted1 gene (TKnl) and studied its expression pattern. A new program in which eight different meristematic genes in tomato will be studied emerged as a result of these studies. In essence, we have shown that it is possible to study and manipulate plant developmental systems using reverse genetic techniques and have provided a wealth of new molecular tools to interested colleagues working with tomato. Similarly, genes responsible for cell division, cell proliferation and cell fate were studied in Arabidopsis floral meristems. Among these genes are the TSO1, TSO2, HANABA TARANU and UNUSUAL FLORAL ORGANS genes, each affecting in its own way the number of pattern of cell divisions, and cell fate, in developing Arabodopsis flowers. In addition, new methods have been established for the assessment of the function of regulatory gene action in the different clonal layers of developing floral meristems.

An RNA Interference Screen Reveals Fate Determinants Implicated in Asymmetric Cell Division as Potential Regulators of Hematopoietic Stem Cell Self-Renewal

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2462-2462
Author(s):  
Kristin J Hope ◽  
Sonia Cellot ◽  
Stephen Ting ◽  
Guy Sauvageau
Keyword(s):  
Cell Death ◽  
Stem Cell ◽  
Cell Division ◽  
Rna Interference ◽  
Cell Fate ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Fate Determinants ◽  
Cell Fate Determinants

Abstract During periods of extensive regeneration of the hematopoietic system, hematopoietic stem cells (HSC) undergo largely symmetrical self-renewal divisions, necessary to rapidly replenish the stem cell pool. Under homeostasis, however, it is likely that HSC rely more on asymmetric self-renewal divisions to retain an appropriate number of HSC while still enabling sufficient production of mature blood cells. The unequal partitioning of intrinsic fate determinants underlies the process of asymmetric stem cell division in lower organisms including Drosophila and C. elegans. The tumor suppressive function of specific determinants has been demonstrated in studies where mutation of fate determinants shown to be inhibitory to the self-renewal of one of the two daughter cells generated upon Drosophila neuroblast division, drives exclusive symmetrical stem cell divisions ultimately leading to the formation of larval brain tumors. As HSCs can not yet be definitively prospectively identified, it has been difficult to determine whether a similar segregation of such cell fate determinants underlies the asymmetric/symmetric self-renewal of these cells or whether deregulation of these determinants could also generate hematopoietic malignancies by inducing constitutive symmetric self-renewal divisions. We addressed these questions through a functional genetics approach taking advantage of systematic RNA interference to interrogate the function of polarity factors and cell fate determinants representing candidate HSC self-renewal regulators. From a list of 72 of such factors identified in the literature, 32 murine homologs were selected based on their differentially high level of expression in HSC-enriched populations. For each candidate we generated 3 unique short hairpin RNA (shRNA) encoding retroviral constructs also carrying EGFP for the purposes of following transduced cells. In a primary screen equal numbers of HSC-enriched Lin-CD150+CD48− cells were infected with the library in an arrayed 96-well format yielding an average gene transfer of 60.0 ± 3.2%. The in vivo reconstituting potential was then evaluated in a CRU assay such that identical proportions of each well were transplanted in duplicate. An average of 37.6 ± 5.1% long-term donor reconstitution was demonstrated by luciferase shRNA transduced controls. Directly following infection, the EGFP+ fraction of a portion of each well was separated by FACS to facilitate qRT-PCR determination of knockdown efficiency. Immunophenotypes, cell viability and morphology of well contents cultured an additional 7 days were also assessed. The percent of EGFP− and EGFP+ donor cell contribution was determined by flow cytometric evaluation of peripheral blood samples taken every 4 weeks for a period of 16 weeks. Genes for which shRNA vectors altered late transplant EGFP levels below or above defined thresholds were considered as hits. At present we have identified 4 genes for which shRNA-mediated depletion negatively affects repopulation but does not induce indiscriminate cell death in culture and 1 gene that may act as a self-renewal inhibitor. In one example, two shRNAs directed against the candidate EB3 showed a dramatic loss of EGFP+ cells in vivo. EB3, a member of the microtubule plus-end binding protein family, has previously described roles in the search-and-capture mechanism of spindle positioning. Interestingly, EB1, a closely related family member is also critical in directing the symmetrical as opposed to asymmetrical divisions of primitive neuroepithelial cells in Drosophila. Validation of all identified hits as well as further evaluation of their function through cell cycle, cell death and homing studies is ongoing.

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