scholarly journals Autocrine signals enable chondrocytes to survive in culture.

1994 ◽  
Vol 126 (4) ◽  
pp. 1069-1077 ◽  
Author(s):  
Y Ishizaki ◽  
J F Burne ◽  
M C Raff
Keyword(s):  
Epithelial Cells ◽  
Cell Density ◽  
Mammalian Cells ◽  
Cell Types ◽  
High Density ◽  
Lens Epithelial Cells ◽  
Low Density ◽  
Cell Type ◽  
Adult Rats ◽  
And Cartilage

We recently proposed that most mammalian cells other than blastomeres may be programmed to kill themselves unless continuously signaled by other cells not to. Many observations indicate that some mammalian cells are programmed in this way, but is it the case for most mammalian cells? As it is impractical to test all of the hundreds of types of mammalian cells, we have focused on two tissues--lens and cartilage--which each contain only a single cell type: if there are cells that do not require signals from other cells to avoid programmed cell death (PCD), lens epithelial cells and cartilage cells (chondrocytes) might be expected to be among them. We have previously shown that rat lens epithelial cells can survive in serum-free culture without signals from other cell types but seem to require signals from other lens epithelial cells to survive: without such signals they undergo PCD. We show here that the same is true for rat (and chick) chondrocytes. They can survive for weeks in culture at high cell density in the absence of other cell types, serum, or exogenous proteins or signaling molecules, but they die with the morphological features of apoptosis in these conditions at low cell density. Medium from high density cultures, FCS, or a combination of known growth factors, all support prolonged chondrocyte survival in low density cultures, as long as antioxidants are also present. Moreover, medium from high density chondrocyte cultures promotes the survival of lens epithelial cells in low density cultures and vice versa. Chondrocytes isolated from adult rats behave similarly to those isolated from developing rats. These findings support the hypothesis that most mammalian cells require signals from other cells to avoid PCD, although the signals can sometimes be provided by cells of the same type, at least in tissues that contain only one cell type.

scholarly journals Control of lens epithelial cell survival.

1993 ◽  
Vol 121 (4) ◽  
pp. 899-908 ◽  
Author(s):  
Y Ishizaki ◽  
J T Voyvodic ◽  
J F Burne ◽  
M C Raff
Keyword(s):  
Epithelial Cells ◽  
Cell Types ◽  
Lens Epithelial Cell ◽  
High Density ◽  
Lens Epithelial Cells ◽  
Cell Contact ◽  
Low Density ◽  
First Time ◽  
Cells Cultured ◽  
Do So

We have studied the survival requirements of developing lens epithelial cells to test the hypothesis that most cells are programmed to kill themselves unless they are continuously signaled by other cells not to do so. The lens cells survived for weeks in both explant cultures and high-density dissociated cell cultures in the absence of other cells or added serum or protein, suggesting that they do not require signals from other cell types to survive. When cultured at low density, however, they died by apoptosis, suggesting that they depend on other lens epithelial cells for their survival. Lens epithelial cells cultured at high density in agarose gels also survived for weeks, even though they were not in direct contact with one another, suggesting that they can promote one another's survival in the absence of cell-cell contact. Conditioned medium from high density cultures promoted the survival of cells cultured at low density, suggesting that lens epithelial cells support one another's survival by secreting survival factors. We show for the first time that normal cell death occurs within the anterior epithelium in the mature lens, but this death is strictly confined to the region of the anterior suture.

scholarly journals EEA1, a Tethering Protein of the Early Sorting Endosome, Shows a Polarized Distribution in Hippocampal Neurons, Epithelial Cells, and Fibroblasts

2000 ◽  
Vol 11 (8) ◽  
pp. 2657-2671 ◽  
Author(s):  
Jean M. Wilson ◽  
Meltsje de Hoop ◽  
Natasha Zorzi ◽  
Ban-Hock Toh ◽  
Carlos G. Dotti ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Mammalian Cells ◽  
Hippocampal Neurons ◽  
Cell Types ◽  
Early Endosomes ◽  
Filamentous Material ◽  
Endocytic Vesicles ◽  
Fibroblastic Cells ◽  
Darby Canine Kidney ◽  
Endocytic Pathways

EEA1 is an early endosomal Rab5 effector protein that has been implicated in the docking of incoming endocytic vesicles before fusion with early endosomes. Because of the presence of complex endosomal pathways in polarized and nonpolarized cells, we have examined the distribution of EEA1 in diverse cell types. Ultrastructural analysis demonstrates that EEA1 is present on a subdomain of the early sorting endosome but not on clathrin-coated vesicles, consistent with a role in providing directionality to early endosomal fusion. Furthermore, EEA1 is associated with filamentous material that extends from the cytoplasmic surface of the endosomal domain, which is also consistent with a tethering/docking role for EEA1. In polarized cells (Madin-Darby canine kidney cells and hippocampal neurons), EEA1 is present on a subset of “basolateral-type” endosomal compartments, suggesting that EEA1 regulates specific endocytic pathways. In both epithelial cells and fibroblastic cells, EEA1 and a transfected apical endosomal marker, endotubin, label distinct endosomal populations. Hence, there are at least two distinct sets of early endosomes in polarized and nonpolarized mammalian cells. EEA1 could provide specificity and directionality to fusion events occurring in a subset of these endosomes in polarized and nonpolarized cells.

scholarly journals RhoA localization with caveolin-1 regulates vascular contractions to serotonin

2012 ◽  
Vol 303 (9) ◽  
pp. R959-R967 ◽  
Author(s):  
Daniel W. Nuno ◽  
Sarah K. England ◽  
Kathryn G. Lamping
Keyword(s):  
Lipid Rafts ◽  
Rho Kinase ◽  
Initial Response ◽  
Cell Types ◽  
Smooth Muscle Contraction ◽  
High Density ◽  
Low Density ◽  
Density Fractions ◽  
Caveolin 1 ◽  
Multiple Cell

Vascular smooth muscle contraction occurs following an initial response to an increase in intracellular calcium concentration and a sustained response following increases in the sensitivity of contractile proteins to calcium (calcium sensitization). This latter process is regulated by the rhoA/rho kinase pathway and activated by serotonin. In multiple cell types, signaling molecules compartmentalize within caveolae to regulate their activation. We hypothesized that serotonin differentially compartmentalizes rhoA within caveolar versus noncaveolar lipid rafts to regulate sustained vascular contractions. To test this hypothesis, we measured aortic contractions in response to serotonin in wild-type (WT) and cav-1-deficient mice (cav-1 KO). RhoA-dependent contractions in response to serotonin were markedly augmented in arteries from cav-1 KO mice despite a modest reduction in rhoA expression compared with WT. We found that under basal conditions, rhoA in WT arteries was primarily localized within high-density sucrose gradient fractions but temporally shifted to low-density fractions in response to serotonin. In contrast, rhoA in cav-1 KO arteries was primarily in low-density fractions and shifted to high-density fractions in a similar timeframe as that seen in WT mice. We conclude that localization of rhoA to caveolar versus noncaveolar lipid rafts differentially regulates its activation and contractions to rhoA-dependent agonists with greater activation associated with its localization to noncaveolar rafts. Disruption of rhoA localization within caveolae may contribute to increased activation and enhanced vascular contractions in cardiovascular disease.

Discordant regulatory changes in monocrotaline-induced megalocytosis of lung arterial endothelial and alveolar epithelial cells

2006 ◽  
Vol 290 (6) ◽  
pp. L1216-L1226 ◽  
Author(s):  
Somshuvra Mukhopadhyay ◽  
Pravin B. Sehgal
Keyword(s):  
Cell Cycle ◽  
Epithelial Cells ◽  
Dna Synthesis ◽  
A549 Cells ◽  
Cell Types ◽  
Alveolar Epithelial Cells ◽  
Cell Type ◽  
Alveolar Epithelial ◽  
Unfolded Protein ◽  
Cell Cycle Regulatory Proteins

Monocrotaline (MCT) causes pulmonary hypertension in the rat by a mechanism characterized by megalocytosis (enlarged cells with enlarged endoplasmic reticulum and Golgi and a cell cycle arrest) of pulmonary arterial endothelial (PAEC), arterial smooth muscle, and type II alveolar epithelial cells. In cell culture, although megalocytosis is associated with a block in entry into mitosis in both lung endothelial and epithelial cells, DNA synthesis is stimulated in endothelial but inhibited in epithelial cells. The molecular mechanism(s) for this dichotomy are unclear. While MCTP-treated PAEC and lung epithelial (A549) cells both showed an increase in the “promitogenic” transcription factor STAT3 levels and in the IL-6-induced nuclear pool of PY-STAT3, this was transcriptionally inactive in A549 but not in PAEC cells. This lack of transcriptional activity of STAT3 in A549 cells correlated with the cytoplasmic sequestration of the STAT3 coactivators CBP/p300 and SRC1/NcoA in A549 cells but not in PAEC. Both cell types displayed a Golgi trafficking block, loss of caveolin-1 rafts, and increased nuclear Ire1α, but an incomplete unfolded protein response (UPR) with little change in levels of UPR-induced chaperones including GRP78/BiP. There were discordant alterations in cell cycle regulatory proteins in the two cell types such as increase in levels of both cyclin D1 and p21 simultaneously, but with a decrease in cdc2/cdk1, a kinase required for entry into mitosis. While both cell types showed increased cytoplasmic geminin, the DNA synthesis-initiating protein Cdt1 was predominantly nuclear in PAEC but remained cytoplasmic in A549 cells, consistent with the stimulation of DNA synthesis in the former but an inhibition in the latter cell type. Thus differences in cell type-specific alterations in subcellular trafficking of critical regulatory molecules (such as CBP/p300, SRC1/NcoA, Cdt1) likely account for the dichotomy of the effects of MCTP on DNA synthesis in endothelial and epithelial cells.

scholarly journals Density-dependent Growth Inhibition of Fibroblasts Ectopically Expressing p27kip1

2000 ◽  
Vol 11 (6) ◽  
pp. 2117-2130 ◽  
Author(s):  
Xiaohong Zhang ◽  
Walker Wharton ◽  
Marcia Donovan ◽  
Domenico Coppola ◽  
Rhonda Croxton ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Density ◽  
Ectopic Expression ◽  
High Density ◽  
Quiescent State ◽  
Low Density ◽  
Cdk Inhibitor ◽  
Cyclin Dependent Kinase ◽  
Tumor Virus ◽  
Dependent Growth

The cyclin/cyclin-dependent kinase (cdk) inhibitor p27kip1 is thought to be responsible for the onset and maintenance of the quiescent state. It is possible, however, that cells respond differently to p27kip1 in different conditions, and using a BALB/c-3T3 cell line (termed p27-47) that inducibly expresses high levels of this protein, we show that the effect of p27kip1 on cell cycle traverse is determined by cell density. We found that ectopic expression of p27kip1blocked the proliferation of p27-47 cells at high density but had little effect on the growth of cells at low density whether exponentially cycling or stimulated from quiescence. Regardless of cell density, the activities of cdk4 and cdk2 were markedly repressed by p27kip1 expression, as was the cdk4-dependent dissociation of E2F4/p130 complexes. Infection of cells with SV40, a DNA tumor virus known to abrogate formation of p130- and Rb-containing complexes, allowed dense cultures to proliferate in the presence of supraphysiological amounts of p27kip1 but did not stimulate cell cycle traverse when cultures were cotreated with the potent cdk2 inhibitor roscovitine. Our data suggest that residual levels of cyclin/cdk activity persist in p27kip1-expressing p27-47 cells and are sufficient for the growth of low-density cells and of high-density cells infected with SV40, and that effective disruption of p130 and/or Rb complexes is obligatory for the proliferation of high-density cultures.

scholarly journals ATP Mediates Yeast Cell-Cell Communication

2020 ◽  
Author(s):  
Michael Caplow
Keyword(s):  
Cell Density ◽  
High Cell Density ◽  
Cell Communication ◽  
Growth Stimulation ◽  
High Density ◽  
Mutant Protein ◽  
Chromosome Loss ◽  
Low Density ◽  
High Cell ◽  
In Cells

AbstractYeast secrete ATP in response to glucose, a property with previously unknown functional consequence. In this report, we show that extracellular ATP is a signal for growth of surrounding cells. The ATP signaling behavior was uncovered by finding reduced toxicity of an inducible, dominant-lethal form of alpha tubulin (tub1-828) in cells grown at high, compared to low cell density. Reduced cell death at high cell density resulted because the rate of chromosome loss/cell division was lower (18-fold) in a cultures inoculated with a high density (350,000) compared to a low density (5,000) of cells. The sparing effect of growth at high cell density could be replicated by growing together 3440 cells that express tub1-828, with 2.3 E6 cells that do not express the mutant protein. Toxicity was reduced at high cell density apparently because a secreted signal induces growth, so that the mutant protein is rapidly diluted by synthesis of wild-type α-tubulin. Further, fluorescence-activated cell sorting (FACS) analysis after DNA staining showed that the rate of the G1-G2 transition was faster with cells at high density. ATP replaced the need for high cell density for resistance to tub1-828, and stimulated the transition from G1 to G2 in cells at low density. Cells lacking the enzyme nucleoside diphosphate kinase did not respond to nucleotide stimulation of growth during expression of mutant tubulin, suggesting that NDP kinase has a regulatory role in growth stimulation. This newly discovered quorum sensing response in yeast, mediated by ATP, indicates that yeast decision-making is not entirely autonomous.

scholarly journals Quantitative imaging of intracellular density with ratiometric stimulated Raman scattering microscopy

2021 ◽  
Author(s):  
Benjamin Figueroa ◽  
Fiona Xi Xu ◽  
Ruoqian Hu ◽  
Shuaiqian Men ◽  
Dan Fu
Keyword(s):  
Light Scattering ◽  
Raman Scattering ◽  
Cell Size ◽  
Cell Density ◽  
Cell Function ◽  
Stimulated Raman Scattering ◽  
Cell Types ◽  
Tissue Homeostasis ◽  
Cell Type ◽  
Intact Tissue

AbstractCell size and density impact a wide range of physiological functions, including tissue homeostasis, growth regulation, and osmoregulation. Both are tightly regulated in mammalian cells. In comparison, density variation of a given cell type is much smaller than cell size, indicating that maintenance of cell type-specific density is important for cell function. Despite this importance, little is known about how cell density affects cell function and how it is controlled. Current tools for intracellular cell density measurements are limited either to suspended cells or cells growing on 2D substrates, neither of which recapitulate the physiology of single cells in intact tissue. While optical measurements have the potential to measure cell density in situ and noninvasively, light scattering in multicellular systems prevents direct quantification. Here, we introduce an intracellular density imaging technique based on ratiometric stimulated Raman scattering microscopy (rSRS). It quantifies intracellular drymass density through vibrational imaging of macromolecules. Moreover, water is used as an internal standard to correct for aberration and light scattering. We demonstrate real-time measurement of intracellular density quantification and show that density is tightly regulated across different cell types and can be used to differentiate cell types as well as cell states. We further demonstrate dynamic imaging of density change in response to osmotic challenge as well as intracellular density imaging of a 3D tumor spheroid. Our technique has the potential for imaging intracellular density in intact tissue and understanding density regulation and its role in tissue homeostasis.

Parametric Finite Element Analysis of Physical Stimuli Resulting From Mechanical Stimulation of Tissue Engineered Cartilage

2008 ◽  
Author(s):  
Omotunde M. Babalola ◽  
Lawrence J. Bonassar
Keyword(s):  
Finite Element Analysis ◽  
Cell Types ◽  
Cartilage Tissue ◽  
Low Density ◽  
Cell Type ◽  
Element Analysis ◽  
Physical Stimuli ◽  
Parametric Finite Element Analysis ◽  
Engineered Cartilage ◽  
Stimulation Of

The avascular nature of cartilage results in its limited inability to repair itself upon injury. As a result numerous approaches are being investigated as potential therapies for repair, including tissue engineering strategies. In addition, due to the low density of chondrocytes and the characteristic de-differentiation of the cells when expanded in monolayer [1], other cell types are being investigated as a source for cartilage repair as well. Mesenchymal stem cells (MSCs), which have been shown to differentiate into cells of several lineages including chondrocytes, osteoblasts and adipocytes [2], are being explored as a potential cell type for the regeneration of articular cartilage tissue [3,4].

scholarly journals Single cell RNA sequencing identifies IGFBP5 and QKI as ciliated epithelial cell genes associated with severe COPD

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Xiuying Li ◽  
Guillaume Noell ◽  
Tracy Tabib ◽  
Alyssa D. Gregory ◽  
Humberto E. Trejo Bittar ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Single Cell ◽  
Rna Sequencing ◽  
Cell Types ◽  
Cell Type ◽  
Protein Levels ◽  
Single Cell Rna Sequencing ◽  
Ciliated Epithelial Cells ◽  
Severe Copd

Abstract Background Whole lung tissue transcriptomic profiling studies in chronic obstructive pulmonary disease (COPD) have led to the identification of several genes associated with the severity of airflow limitation and/or the presence of emphysema, however, the cell types driving these gene expression signatures remain unidentified. Methods To determine cell specific transcriptomic changes in severe COPD, we conducted single-cell RNA sequencing (scRNA seq) on n = 29,961 cells from the peripheral lung parenchymal tissue of nonsmoking subjects without underlying lung disease (n = 3) and patients with severe COPD (n = 3). The cell type composition and cell specific gene expression signature was assessed. Gene set enrichment analysis (GSEA) was used to identify the specific cell types contributing to the previously reported transcriptomic signatures. Results T-distributed stochastic neighbor embedding and clustering of scRNA seq data revealed a total of 17 distinct populations. Among them, the populations with more differentially expressed genes in cases vs. controls (log fold change >|0.4| and FDR = 0.05) were: monocytes (n = 1499); macrophages (n = 868) and ciliated epithelial cells (n = 590), respectively. Using GSEA, we found that only ciliated and cytotoxic T cells manifested a trend towards enrichment of the previously reported 127 regional emphysema gene signatures (normalized enrichment score [NES] = 1.28 and = 1.33, FDR = 0.085 and = 0.092 respectively). Among the significantly altered genes present in ciliated epithelial cells of the COPD lungs, QKI and IGFBP5 protein levels were also found to be altered in the COPD lungs. Conclusions scRNA seq is useful for identifying transcriptional changes and possibly individual protein levels that may contribute to the development of emphysema in a cell-type specific manner.

Deficient epithelial-fibroblast heterocellular gap junction communication can be overcome by co-culture with an intermediate cell type but not by E-cadherin transgene expression

1998 ◽  
Vol 111 (23) ◽  
pp. 3529-3539 ◽  
Author(s):  
T.L. Woodward ◽  
M.A. Sia ◽  
O.W. Blaschuk ◽  
J.D. Turner ◽  
D.W. Laird
Keyword(s):  
Epithelial Cells ◽  
Gap Junction ◽  
Cell Lines ◽  
Intercellular Communication ◽  
Cell Types ◽  
Intermediate Cell ◽  
Cell Type ◽  
Gap Junction Intercellular Communication ◽  
Intermediate Cells ◽  
E Cadherin

Epithelial, fibroblast and intermediate cell lines were employed to examine the mechanism(s) essential for heterocellular gap junction intercellular communication in vitro. These cell lines were characterized extensively for cell type based on morphology, intermediate cytoskeletal proteins, cell adhesion molecules and their associated proteins, tight junction proteins as well as functional differentiation. All cell types expressed connexin43 and were dye-coupled in homocellular culture. Epithelial and intermediate cells or fibroblasts and intermediate cells readily assembled heterocellular connexin43-positive gap junction plaques when co-cultured, while gap junction plaques in mixed cultures of epithelial cells and fibroblasts were rare. Dye microinjection studies were used to show that there was little gap junction intercellular communication between epithelial cells and fibroblasts. However, intermediate cells were able to communicate with epithelial cells and, to a lesser extent, fibroblasts and could transfer dye to both epithelial cells and fibroblasts when all three cell types were cultured together. Fibroblasts that were stably transfected with a cDNA encoding E-cadherin had a greater tendency to aggregate and exhibited a more epithelial-like phenotype but heterocellular gap junction intercellular communication with epithelial cells, which endogenously express E-cadherin, was not enhanced. These results suggest that mutual expression of E-cadherin is insufficient to stimulate gap junction formation between epithelial cells and fibroblasts. Moreover, our results also demonstrate that communication gaps between epithelial cells and fibroblasts can be bridged by intermediate cells, a process that may be important in mammary gland development, growth, differentiation and cancer.

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