Activation of caspase‐8 and Erk‐1/2 in domes regulates cell death induced by confluence in MDCK cells

Chang, Yung-Heng; Lin, Hsi Hui; Wang, Yang Kao; Chiu, Wen Tai; Su, Hsiao Wen; Tang, Ming Jer

doi:10.1002/jcp.20926

Cited by 8 publications

(5 citation statements)

References 38 publications

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“…Thus, an increased expression of NHE3, a Na + /H + exchanger in intestinal and renal brush border membrane that is activated by the transcription factor Stat3, was observed in MDCK domeforming cells [24]. In vitro studies also showed that dome formation is associated with increased levels of intracellular cAMP [25] and that caspase-8 and p-Erk are also expressed in cells forming domes [11]. Our present results demonstrate that in MDCK cells, derived from distal tubule epithelia, exposure to flow conditions strongly affects the dynamic nature of cell dome formation, with complete disappearance of cell domes within 6 hours after beginning of monolayer perfusion.…”

Section: Discussionmentioning

confidence: 86%

“…These structural alterations likely derive from active fluid reabsorption at luminal level and basolateral secretion, with consequent cell detachment from the substratum and reorganization of junctional contacts. These events induce and sustain focal elevation of groups of cells along the monolayer, and are associated with cell differentiation and expression of pro-apoptotic proteins [11]. Cell dome formation is also involved in tubular cell differentiation and morphogenesis [12].…”

Section: Introductionmentioning

confidence: 99%

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Shear Stress Reverses Dome Formation in Confluent Renal Tubular Cells

Cattaneo

Condorelli

Terrinoni³

et al. 2011

Cell Physiol Biochem

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Background/Aims. It has been shown that MDCK cells, a cell line derived from canine renal tubules, develop cell domes due to fluid pumped under cell monolayer and focal detachment from the adhesion surface. In vitro studies have shown that primary cilia of kidney tubular epithelial cells act as mechanosensors, increasing intracellular calcium within seconds upon changes in fluid shear stress (SS) on cell membrane. We then studied the effect of prolonged SS exposure on cell dome formation in confluent MDCK cell monolayers. Methods. A parallel plate flow chamber was used to apply laminar SS at 2 dynes/cm² to confluent cell monolayers for 6 hours. Control MDCK cell monolayers were maintained in static condition. The effects of Ca²⁺ blockade and cell deciliation on SS exposure were also investigated. Results. Seven days after reaching confluence, static cultures developed liquid filled domes, elevating from culture plate. Exposure to SS induced almost complete disappearance of cell domes (0.4±0.8 vs. 11.4±2.8 domes/mm², p < 0.01, n=14). SS induced dome disappearance took place within minutes to hours, as shown by time-lapse videomicroscopy. Exposure to SS importantly affected cell cytoskeleton altering actin stress fibers expression and organization, and the distribution of tight junction protein ZO-1. Dome disappearance induced by flow was completely prevented in the presence of EGTA or after cell deciliation. Conclusions. These data indicate that kidney tubular cells are sensitive to apical flow and that these effects are mediated by primary cilia by regulation of Ca²⁺ entry in to the cell. SS induced Ca²⁺ entry provokes contraction of cortical actin ring that tenses cell-cell borders and decreases basal stress fibers. These processes may increase paracellular permeability and decrease basal adhesion making dome disappear. Elucidation of the effects of apical fluid flow on tubular cell function may open new insights on the pathophysiology of kidney diseases associated with cilia dysfunction.

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Section: Discussionmentioning

confidence: 86%

Section: Introductionmentioning

confidence: 99%

Shear Stress Reverses Dome Formation in Confluent Renal Tubular Cells

Cattaneo

Condorelli

Terrinoni³

et al. 2011

Cell Physiol Biochem

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“…Epithelial barriers are known to undergo dynamic growth, differentiation and morphogenic processes to replace terminally differentiated cells which are thought to comprise locally formed domes. 11,84,85 Domes and valleys in the epithelial monolayer were observed after the cells reached confluence (around one week of co-culture), and these morphological features persisted during the remaining culture weeks. Visualization of both intact domes (arrows) and a subsequent barrier defect are representative of features observed after three weeks in co-culture (Fig.…”

Section: Resultsmentioning

confidence: 99%

Local remodeling of synthetic extracellular matrix microenvironments by co-cultured endometrial epithelial and stromal cells enables long-term dynamic physiological function

Cook

Hill

Guo

et al. 2017

Integrative Biology

101

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Mucosal barrier tissues, comprising a layer of tightly-bonded epithelial cells in intimate molecular communication with an underlying matrix-rich stroma containing fibroblasts and immune cells, are prominent targets for drugs against infection, chronic inflammation, and other disease processes. Although human in vitro models of such barriers are needed for mechanistic studies and drug development, differences in extracellular matrix (ECM) needs of epithelial and stromal cells hinder efforts to create such models. Here, using the endometrium as an example mucosal barrier, we describe a synthetic, modular ECM hydrogel suitable for 3D functional co-culture, featuring components that can be remodeled by cells and that respond dynamically to sequester local cell-secreted ECM characteristic of each cell type. The synthetic hydrogel combines peptides with off-the-shelf reagents and is thus accessible to cell biology labs. Specifically, we first identified a single peptide as suitable for initial attachment of both endometrial epithelial and stromal cells using a 2D semi-empirical screen. Then, using a co-culture system of epithelial cells cultured on top of gel-encapsulated stromal cells, we show that inclusion of ECM-binding peptides in the hydrogel, along with the integrin-binding peptide, leads to enhanced accumulation of basement membrane beneath the epithelial layer and more fibrillar collagen matrix assembly by stromal cells over two weeks in culture. Importantly, endometrial co-cultures composed of either cell lines or primary cells displayed hormone-mediated differentiation as assessed by morphological changes and secretory protein production. A multiplex analysis of apical cytokine and growth factor secretion comparing cell lines and primary cells revealed strikingly different patterns, underscoring the importance of using primary cell models in analysis of cell-cell communication networks. In summary, we define a “one-size-fits-all” synthetic ECM that enables long-term, physiologically responsive co-cultures of epithelial and stromal cells in a mucosal barrier format.

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“…3A and B and 4). Since the phosphorylation of ERK is a major pathway triggered by FGF-2 (42), we examined the effect of FGF-P on this pathway. The results (Fig.…”

Section: Discussionmentioning

confidence: 99%

Fibroblast Growth Factor-Peptide Improves Barrier Function and Proliferation in Human Keratinocytes After Radiation

Zhang¹,

Tian²,

Yin³

et al. 2011

International Journal of Radiation Oncology*Biology*Physics

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Purpose Epidermal keratinocytes, which can be severely damaged after ionizing radiation (IR), are rapid turnover cells that function as a barrier protecting the host from pathogenic invasion and fluid loss. We tested fibroblast growth factor-peptide (FGF-P), a small peptide derived from the receptor-binding domain of FGF-2, as a potential mitigator of radiation effects via proliferation and the barrier function of keratinocytes. Methods and Materials Keratinocytes isolated from neonatal foreskin were grown on transwells. After 0, 5, or 10-Gy, the cells were treated with a vehicle or FGF-P. The permeability of IR cells was assessed through transepithelial electrical resistance (TEER) and a paracellular tracer flux of fluorescein isothiocyanate-conjugated bovine serum albumin (FITC-BSA) with Ussing chambers. The cell proliferation was measured with yellow tetrazolium salt (MTT) and tritiated thymidine (3H-TdR) assays. The phosphorylation of extracellular signal-regulated kinases (ERK) was measured in an enzyme-linked immunosorbent (ELISA)-like assay, and the proteins related to tight junctions (TJ) and adherens junctions (AJ) were examined with Western blotting. We used a mouse model to assess the ability of FGF-P to promote the healing of skin β-burns created with a strontium applicator. Results 1) FGF-P reduced the permeability of irradiated keratinocytes, as evidenced by increased TEER and decreased diffusion of FITC-BSA both associated with the regulation of different proteins and levels of TJ and AJ; 2) FGF-P enhanced the proliferation of irradiated keratinocytes, as evidenced by increased MTT activity and 3H-TdR incorporation, which was associated with activation of the ERK pathway; and 3) FGF-P promoted the healing of skin β-burns. Conclusions FGF-P enhances the barrier function, including up-regulation of TJ proteins, increases proliferation of human keratinocytes, and accelerates the healing of skin β-burns. FGF-P is a promising mitigator that improves the proliferation and barrier function of keratinocytes after IR.

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Activation of caspase‐8 and Erk‐1/2 in domes regulates cell death induced by confluence in MDCK cells

Cited by 8 publications

References 38 publications

Shear Stress Reverses Dome Formation in Confluent Renal Tubular Cells

Shear Stress Reverses Dome Formation in Confluent Renal Tubular Cells

Local remodeling of synthetic extracellular matrix microenvironments by co-cultured endometrial epithelial and stromal cells enables long-term dynamic physiological function

Fibroblast Growth Factor-Peptide Improves Barrier Function and Proliferation in Human Keratinocytes After Radiation

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