Functional regulation of H+-ATPase-rich cells in zebrafish embryos acclimated to an acidic environment

2009 ◽  
Vol 296 (4) ◽  
pp. C682-C692 ◽  
Author(s):  
Jiun-Lin Horng ◽  
Li-Yih Lin ◽  
Pung-Pung Hwang
Keyword(s):  
Single Cells ◽  
Cell Number ◽  
Alveolar Type ◽  
Zebrafish Embryos ◽  
Ph Homeostasis ◽  
Epithelial Stem Cells ◽  
Acid Secreting ◽  
Functional Regulation ◽  
Cell Acid ◽  
Apical Opening

It is important to maintain internal pH homeostasis in biological systems. In our previous studies, H+-ATPase-rich (HR) cells were found to be responsible for proton secretion in the skin of zebrafish embryos during development. In this study, zebrafish embryos were exposed to acidic and basic waters to investigate the regulation of HR cell acid secretion during pH disturbances. Our results showed that the function of HR cells on the skin of zebrafish embryos can be upregulated in pH 4 water not only by increasing the cell number but also by enlarging the acid-secreting function of single cells. We also identified an “alveolar-type” apical opening under scanning electron microscopy observations of the apical membrane of HR cells, and the density and size of the alveolar type of apical openings were also increased in pH 4 water. p63 and PCNA immunostaining results also showed that additional HR cells in pH 4 water may be differentiated not only from ionocyte precursor cells but also newly proliferating epithelial stem cells.

scholarly journals Decreased expression of Met during differentiation in rat lung

2016 ◽  
Vol 60 (1) ◽  
Author(s):  
T. Kato ◽  
K. Oka ◽  
T. Nakamura ◽  
A. Ito
Keyword(s):  
Stem Cells ◽  
3D Culture ◽  
Immunofluorescence Assay ◽  
Lung Epithelial Cells ◽  
Alveolar Type ◽  
Cell Marker ◽  
Epithelial Stem Cells ◽  
Club Cell ◽  
Adult Mice ◽  
Lung Epithelial

<p>Organ-specific stem cells play key roles in maintaining the epithelial cell layers of lung. Bronchioalveolar stem cells (BASCs) are distal lung epithelial stem cells of adult mice. Alveolar type 2 (AT2) cells have important functions and serve as progenitor cells of alveolar type 1 (AT1) cells to repair the epithelium when they are injured. Hepatocyte growth factor (HGF) elicits mitogenic, morphogenic, and anti-apoptotic effects on lung epithelial cells through tyrosine phosphorylation of Met receptor, and thus is recognized as a pulmotrophic factor. To understand which cells HGF targets in lung, we identified the cells expressing Met by immunofluorescence assay. Met was strongly expressed in BASCs, which expressed an AT2 cell marker, pro-SP-C, and a club cell marker, CCSP. In alveoli, we found higher expression of Met in primary AT2 than in AT1 cells, which was confirmed using primary AT2 cells. We further examined the mitogenic activity of HGF in AT2-cell-derived alveolar-like cysts (ALCs) in 3D culture. Multicellular ALCs expressed Met, and HGF enhanced the ALC production. Taking these findings together, BASCs could also be an important target for HGF, and HGF-Met signaling could function more potent on cells that have greater multipotency in adult lung.</p>

scholarly journals A computational method to aid the design and analysis of single cell RNA-seq experiments for cell type identification

2018 ◽  
Author(s):  
Douglas Abrams ◽  
Parveen Kumar ◽  
R. Krishna Murthy Karuturi ◽  
Joshy George
Keyword(s):  
Experimental Design ◽  
Single Cell ◽  
Single Cells ◽  
Cell Types ◽  
Cell Number ◽  
Fold Change ◽  
Computational Method ◽  
Marker Genes ◽  
Cell Type ◽  
Estimate Sample Size

AbstractBackgroundThe advent of single cell RNA sequencing (scRNA-seq) enabled researchers to study transcriptomic activity within individual cells and identify inherent cell types in the sample. Although numerous computational tools have been developed to analyze single cell transcriptomes, there are no published studies and analytical packages available to guide experimental design and to devise suitable analysis procedure for cell type identification.ResultsWe have developed an empirical methodology to address this important gap in single cell experimental design and analysis into an easy-to-use tool called SCEED (Single Cell Empirical Experimental Design and analysis). With SCEED, user can choose a variety of combinations of tools for analysis, conduct performance analysis of analytical procedures and choose the best procedure, and estimate sample size (number of cells to be profiled) required for a given analytical procedure at varying levels of cell type rarity and other experimental parameters. Using SCEED, we examined 3 single cell algorithms using 48 simulated single cell datasets that were generated for varying number of cell types and their proportions, number of genes expressed per cell, number of marker genes and their fold change, and number of single cells successfully profiled in the experiment.ConclusionsBased on our study, we found that when marker genes are expressed at fold change of 4 or more than the rest of the genes, either Seurat or Simlr algorithm can be used to analyze single cell dataset for any number of single cells isolated (minimum 1000 single cells were tested). However, when marker genes are expected to be only up to fC 2 upregulated, choice of the single cell algorithm is dependent on the number of single cells isolated and proportion of rare cell type to be identified. In conclusion, our work allows the assessment of various single cell methods and also aids in examining the single cell experimental design.

scholarly journals Author Correction: Mapping histone modifications in low cell number and single cells using antibody-guided chromatin tagmentation (ACT-seq)

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Benjamin Carter ◽  
Wai Lim Ku ◽  
Jee Youn Kang ◽  
Gangqing Hu ◽  
Jonathan Perrie ◽  
...  
Keyword(s):  
Histone Modifications ◽  
Single Cells ◽  
Cell Number

scholarly journals Nuclear gene proximity and protein interactions shape transcript covariations in mammalian single cells

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Marcel Tarbier ◽  
Sebastian D. Mackowiak ◽  
João Frade ◽  
Silvina Catuara-Solarz ◽  
Inna Biryukova ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Nuclear Organization ◽  
Single Cells ◽  
Expression Patterns ◽  
Embryonic Stem ◽  
Nuclear Gene ◽  
Biological Properties ◽  
Sequencing Studies ◽  
Genes Encoding ◽  
Functional Regulation

Abstract Single-cell RNA sequencing studies on gene co-expression patterns could yield important regulatory and functional insights, but have so far been limited by the confounding effects of differentiation and cell cycle. We apply a tailored experimental design that eliminates these confounders, and report thousands of intrinsically covarying gene pairs in mouse embryonic stem cells. These covariations form a network with biological properties, outlining known and novel gene interactions. We provide the first evidence that miRNAs naturally induce transcriptome-wide covariations and compare the relative importance of nuclear organization, transcriptional and post-transcriptional regulation in defining covariations. We find that nuclear organization has the greatest impact, and that genes encoding for physically interacting proteins specifically tend to covary, suggesting importance for protein complex formation. Our results lend support to the concept of post-transcriptional RNA operons, but we further present evidence that nuclear proximity of genes may provide substantial functional regulation in mammalian single cells.

Effects of Fetal Tracheal Occlusion (TO) on Alveolar Type 2 Cell Number and Maturation in the Rat

1999 ◽  
Vol 45 (4, Part 2 of 2) ◽  
pp. 307A-307A
Author(s):  
Yoshihiro Kitano ◽  
Daniel Von Allmen ◽  
Alan W Flake ◽  
Annapurna Korimilli ◽  
Susan H Guttentag
Keyword(s):  
Cell Number ◽  
Alveolar Type ◽  
Tracheal Occlusion

scholarly journals Cation Metabolism in Relation to Cell Size in Synchronously Grown Tissue Culture Cell

1967 ◽  
Vol 50 (4) ◽  
pp. 917-932 ◽  
Author(s):  
Chan Jung ◽  
Aser Rothstein
Keyword(s):  
Tissue Culture ◽  
Cell Division ◽  
Exponential Function ◽  
Single Cells ◽  
Cell Number ◽  
Culture Cell ◽  
Tissue Culture Cell ◽  
Content Increase ◽  
Cation Content ◽  
Unidirectional Fluxes

In randomly grown tissue culture cells (mouse leukemic lymphoblast, L5178Y) the number, volume, and Na+ and K+ content increase as an exponential function with a doubling time of 11.3 hr. In synchronously grown cells the volume increase of the population and of single cells follows the same exponential function as in randomly grown cells. In contrast, the cation content fluctuates during a single cell cycle. About 1½ hr after the cell division burst (at the beginning of the S period), a net loss of K+ occurs for a period of about 1 hr amounting to about 20% of the total K. Over the next 5 to 6 hr, the deficit in K+ is eliminated. The Na+ content shows a double fluctuation. It falls during the cell division burst, rises when the K+ content decreases, falls again when K+ content rises, and then increases again before the next cell division burst. The net fluxes of both Na+ and K+ are very small compared to the unidirectional fluxes (less than 5%), thus small changes in the balance of influx and efflux account for the changes in cation content during the growth cycle. Both unidirectional fluxes increase dramatically (by a factor of two) about 2 hr after the cell division burst, and then remain constant until after the next cell division. The pattern of electrolyte regulation during cell division does not follow a simple function such as cell number, cell surface, or cell volume, but must be related to specific internal events in the cell.

Surfactant phospholipids and proteins are increased in fetal sheep with pulmonary hypertension secondary to fetal systemic arteriovenous fistula

2005 ◽  
Vol 288 (3) ◽  
pp. L562-L568 ◽  
Author(s):  
Alexandra Benachi ◽  
Jean-Marie Jouannic ◽  
Anne-Marie Barlier-Mur ◽  
Bernadette Chailley-Heu ◽  
Jacques R. Bourbon
Keyword(s):  
Pulmonary Hypertension ◽  
Arteriovenous Fistula ◽  
Lamellar Body ◽  
State Level ◽  
Surfactant Protein ◽  
Cell Number ◽  
Alveolar Type ◽  
Fetal Sheep ◽  
Surfactant Phospholipids ◽  
Surfactant Material

To determine whether prenatal surfactant storage was altered in a model of systemic arteriovenous fistula (SAVF) with pulmonary hypertension, a fistula was created between the internal jugular vein and the carotid artery in 120-day fetal lambs, and surfactant material was explored at 134 days. Total phospholipids (TPL) and disaturated phosphatidylcholine (DSPC) were increased in whole lung tissue. Phospholipid analysis of isolated lamellar body fraction evidenced a specific increase of surfactant pool size: TPL and DSPC in this fraction were enhanced 1.9 and 2.9 times, respectively, when referred to DNA. Although the steady-state level of transcripts of surfactant protein (SP)-A and SP-B was not found to be changed at the time of death, semiquantitative Western blot analysis revealed elevated SP-A and SP-B protein contents three- and twofold, respectively. These findings indicate markedly enhanced accumulation of surfactant material in the presence of surgically induced prenatal pulmonary hypertension. Although total lung cell number was increased by 26%, SP-B immunolabeling indicated that increased surfactant amount did not result from an increased alveolar type II cell proportion, but rather from an increased rate of storage. Whether similar changes in surfactant are encountered in human neonates with persistent pulmonary hypertension is worthy of investigation.

High-Throughput Production of Single-Cell Microparticles Using an Inkjet Printing Technology

2008 ◽  
Vol 130 (2) ◽  
Author(s):  
Tao Xu ◽  
Helen Kincaid ◽  
Anthony Atala ◽  
James J. Yoo
Keyword(s):  
Single Cell ◽  
High Throughput ◽  
Single Cell Analysis ◽  
Single Cells ◽  
Stem Cell Differentiation ◽  
Cell Number ◽  
High Throughput Drug Screening ◽  
Alginate Concentration ◽  
High Throughput Method ◽  
Normal Cellular Function

In this study, a novel biocompatible and inexpensive method for the rapid production of single-cell based microparticles is described. Using an HP DeskJet 550C printer, alginate microparticles containing one to several insulin-producing cells (beta-TC6) were fabricated by coprinting the cells and sodium alginate suspension into a CaCl2 solution. This method is able to generate microparticles of 30–60μm in diameter at a rate as high as 55,000particles∕s. Cell survival assays showed that more than 89% of printed cells survived the fabrication process. The printed beta-TC6 cells demonstrated continuous insulin secretion over a 6day period, which suggests that the printed cells are able to maintain normal cellular function within the microparticles. We show that the printing conditions, such as cell number, alginate concentration, and ionic strengths of CaCl2, influence cellular distribution and geometry of the printed particles. This study presents a simple and high-throughput method to encapsulate single cells, and this technique may be applied in various research investigations, including single-cell analysis, high-throughput drug screening, and stem cell differentiation at the single-cell level.

scholarly journals Jamming dynamics of stretch-induced surfactant release by alveolar type II cells

2012 ◽  
Vol 112 (5) ◽  
pp. 824-831 ◽  
Author(s):  
Arnab Majumdar ◽  
Stephen P. Arold ◽  
Erzsébet Bartolák-Suki ◽  
Harikrishnan Parameswaran ◽  
Béla Suki
Keyword(s):  
Surface Layer ◽  
Single Cells ◽  
Nonlinear Process ◽  
Alveolar Type ◽  
Type Ii Cells ◽  
Type Ii ◽  
Alveolar Epithelial ◽  
Interfacial Surface ◽  
Elastic Membranes ◽  
Surfactant Transport

Secretion of pulmonary surfactant by alveolar epithelial type II cells is vital for the reduction of interfacial surface tension, thus preventing lung collapse. To study secretion dynamics, rat alveolar epithelial type II cells were cultured on elastic membranes and cyclically stretched. The amounts of phosphatidylcholine, the primary lipid component of surfactant, inside and outside the cells, were measured using radiolabeled choline. During and immediately after stretch, cells secreted less surfactant than unstretched cells; however, stretched cells secreted significantly more surfactant than unstretched cells after an extended lag period. We developed a model based on the hypothesis that stretching leads to jamming of surfactant traffic escaping the cell, similar to vehicular traffic jams. In the model, stretch increases surfactant transport from the interior to the exterior of the cell. This transport is mediated by a surface layer with a finite capacity due to the limited number of fusion pores through which secretion occurs. When the amount of surfactant in the surface layer approaches this capacity, interference among lamellar bodies carrying surfactant reduces the rate of secretion, effectively creating a jam. When the stretch stops, the jam takes an extended time to clear, and subsequently the amount of secreted surfactant increases. We solved the model analytically and show that its dynamics are consistent with experimental observations, implying that surfactant secretion is a fundamentally nonlinear process with memory representing collective behavior at the level of single cells. Our results thus highlight the importance of a jamming dynamics in stretch-induced cellular secretory processes.

Effects of Cadmium on Differentiation and Cell Cycle Progression in Cultured Xenopus Kidney Distal Epithelial (A6) Cells

2007 ◽  
Vol 35 (3) ◽  
pp. 343-348 ◽  
Author(s):  
Henning F. Bjerregaard
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Visual Inspection ◽  
Single Cells ◽  
Flow Cytometric Analysis ◽  
Cell Number ◽  
Cellular Growth ◽  
G1 Phase ◽  
Epithelial Cell Line ◽  
A6 Cells

Cadmium (Cd) is an important industrial and environmental pollutant, and the kidney is the primary organ to be affected. To elucidate the effects of Cd on cell proliferation, an epithelial cell line (A6) originally derived from the distal part of the Xenopus laevis kidney was cultured in media containing 10% fetal bovine serum. The effects of Cd (added as CdCl2) on cellular growth and differentiation from single cells to confluent epithelia were investigated by visual inspection and by measurement of the degree to which living cells covered a unit area. Over a concentration range from 5 to 50μM, Cd did not affect the settling and adherence of single cells to the bottom of the culture well. The addition of 5μM Cd for 4 days did not affect the ability of the A6 cells to develop confluent epithelia, measured as the area covered by adherent living epithelial cells (99 ± 4% of the control value). However, 10μM Cd did effectively inhibit development of confluent epithelia to 13 ± 5% compared to control. Visual inspection of adherent cells exposed to 50μM Cd for 7 days revealed no increase in cell number or in cell death, which indicated the induction of cell cycle arrest. Flow cytometric analysis showed that treatment of cells with Cd (0.4mM) for 24 hours induced a significant increase in the proportion of G1 phase cells from 58.6 ± 3.9 to 80.6 ± 3.7%, and a corresponding reduction in the proportion of cells in both the S and G2 phases from 24.0 ± 3.6 to 13.4 ± 3.3% and 17.2 ± 1.7 to 5.8 ± 2.1%, respectively. This study showed that Cd stopped cell proliferation in a very narrow concentration range, between 5 and 10μM, and cell cycle analysis indicated that Cd arrested the cells in the G1 phase of the cell cycle.

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