Molecular Physiology and Pathophysiology of Tight Junctions III. Tight junction regulation by intracellular messengers: differences in response within and between epithelia

2000 ◽  
Vol 279 (4) ◽  
pp. G660-G665 ◽  
Author(s):  
Jurgen Karczewski ◽  
Jack Groot
Keyword(s):  
Tight Junction ◽  
Tight Junctions ◽  
Imaging Techniques ◽  
Effector Proteins ◽  
Molecular Physiology ◽  
Intracellular Messengers ◽  
Tight Junction Regulation ◽  
Tight Junction Permeability ◽  
Permeability Studies ◽  
And Function

Tight junction permeability differs with the type of permeants, their size, and their charge. Selective changes in permeability do occur, and they illustrate the diversity in functional reactions of tight junctions. This suggests that special structures in the tight junctions are involved. More and more structural components of the tight junctions are becoming known. The divergence in behavior of native tissue and filter-grown epithelial monolayers with respect to the effects of intracellular messengers offers the possibility to relate structure and function. In addition to the tools for conventional permeability studies, probes have become available to detect changes in activation of intracellular effector proteins such as the protein kinase C isotypes, and with in situ imaging techniques the way is open for a functional approach in the study of tight junctions.

Physiological regulation of epithelial tight junctions is associated with myosin light-chain phosphorylation

1997 ◽  
Vol 273 (4) ◽  
pp. C1378-C1385 ◽  
Author(s):  
Jerrold R. Turner ◽  
Brian K. Rill ◽  
Susan L. Carlson ◽  
Denise Carnes ◽  
Rachel Kerner ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Tight Junction ◽  
Tight Junctions ◽  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Kinase Inhibitors ◽  
Tight Junction Regulation ◽  
Tight Junction Permeability ◽  
Mlc Phosphorylation

Tight junctions serve as the rate-limiting barrier to passive movement of hydrophilic solutes across intestinal epithelia. After activation of Na+-glucose cotransport, the permeability of intestinal tight junctions is increased. Because previous analyses of this physiological tight junction regulation have been restricted to intact mucosae, dissection of the mechanisms underlying this process has been limited. To characterize this process, we have developed a reductionist model consisting of Caco-2 intestinal epithelial cells transfected with the intestinal Na+-glucose cotransporter, SGLT1. Monolayers of SGLT1 transfectants demonstrate physiological Na+-glucose cotransport. Activation of SGLT1 results in a 22 ± 5% fall in transepithelial resistance (TER) ( P< 0.001). Similarly, inactivation of SGLT1 by addition of phloridzin increases TER by 24 ± 2% ( P < 0.001). The increased tight junction permeability is size selective, with increased flux of small nutrient-sized molecules, e.g., mannitol, but not of larger molecules, e.g., inulin. SGLT1-dependent increases in tight junction permeability are inhibited by myosin light-chain kinase inhibitors (20 μM ML-7 or 40 μM ML-9), suggesting that myosin regulatory light-chain (MLC) phosphorylation is involved in tight junction regulation. Analysis of MLC phosphorylation showed a 2.08-fold increase after activation of SGLT1 ( P< 0.01), which was inhibited by ML-9 ( P < 0.01). Thus monolayers incubated with glucose and myosin light-chain kinase inhibitors are comparable to monolayers incubated with phloridzin. ML-9 also inhibits SGLT1-mediated tight junction regulation in small intestinal mucosa ( P < 0.01). These data demonstrate that epithelial cells are the mediators of physiological tight junction regulation subsequent to SGLT1 activation. The intimate relationship between tight junction regulation and MLC phosphorylation suggests that a critical step in regulation of epithelial tight junction permeability may be myosin ATPase-mediated contraction of the perijunctional actomyosin ring and subsequent physical tension on the tight junction.

Transepithelial resistance can be regulated by the intestinal brush-border Na+/H+ exchanger NHE3

2000 ◽  
Vol 279 (6) ◽  
pp. C1918-C1924 ◽  
Author(s):  
Jerrold R. Turner ◽  
Eric D. Black ◽  
Jeff Ward ◽  
Chung-Ming Tse ◽  
Frederick A. Uchwat ◽  
...  
Keyword(s):  
Tight Junction ◽  
Myosin Ii ◽  
Transepithelial Resistance ◽  
Cell Swelling ◽  
Intestinal Brush Border ◽  
Tight Junction Regulation ◽  
Cytoplasmic Acidification ◽  
Tight Junction Permeability ◽  
Dimethyl Amiloride ◽  
Mlc Phosphorylation

Initiation of intestinal Na+-glucose cotransport results in transient cell swelling and sustained increases in tight junction permeability. Since Na+/H+ exchange has been implicated in volume regulation after physiological cell swelling, we hypothesized that Na+/H+ exchange might also be required for Na+-glucose cotransport-dependent tight junction regulation. In Caco-2 monolayers with active Na+-glucose cotransport, inhibition of Na+/H+ exchange with 200 μM 5-( N, N-dimethyl)- amiloride induced 36 ± 2% increases in transepithelial resistance (TER). Evaluation using multiple Na+/H+ exchange inhibitors showed that inhibition of the Na+/H+ exchanger 3 (NHE3) isoform was most closely related to TER increases. TER increases due to NHE3 inhibition were related to cytoplasmic acidification because cytoplasmic alkalinization with 5 mM NH4Cl prevented both cytoplasmic acidification and TER increases. However, NHE3 inhibition did not affect TER when Na+-glucose cotransport was inhibited. Myosin II regulatory light chain (MLC) phosphorylation decreased up to 43 ± 5% after inhibition of Na+/H+ exchange, similar to previous studies that associate decreased MLC phosphorylation with increased TER after inhibition of Na+-glucose cotransport. However, NHE3 inhibitors did not diminish Na+-glucose cotransport. These data demonstrate that inhibition of NHE3 results in decreased MLC phosphorylation and increased TER and suggest that NHE3 may participate in the signaling pathway of Na+-glucose cotransport-dependent tight junction regulation.

Tight junction regulation through vesicle trafficking: bringing cells together

2014 ◽  
Vol 42 (1) ◽  
pp. 195-200 ◽  
Author(s):  
Sarah J. Fletcher ◽  
Joshua Z. Rappoport
Keyword(s):  
Steady State ◽  
Tight Junction ◽  
Tight Junctions ◽  
Vesicle Trafficking ◽  
Present Review ◽  
Physiological Processes ◽  
Endosomal Recycling ◽  
Tight Junction Regulation ◽  
Epithelial Layers

Epithelial layers are integral for many physiological processes and are maintained by intercellular adhesive structures. During disease, these structures can disassemble, leading to breakdown of epithelia. TJs (tight junctions) are one type of intercellular adhesion. Loss of TJs has been linked to the pathogenesis of many diseases. The present review focuses on the role of vesicle trafficking in regulation of TJs, in particular trafficking of the TJ protein occludin. We examine how endocytosis and endosomal recycling modulate occludin localization under steady-state conditions and during stimulated TJ disassembly.

ZO-1 maintains its spatial distribution but dissociates from junctional fibrils during tight junction regulation

1993 ◽  
Vol 264 (5) ◽  
pp. C1096-C1101 ◽  
Author(s):  
J. L. Madara ◽  
S. Carlson ◽  
J. M. Anderson
Keyword(s):  
Spatial Distribution ◽  
Tight Junction ◽  
Tight Junctions ◽  
Plasma Membranes ◽  
Freeze Fracture ◽  
Cell Junctions ◽  
Absorptive Cell ◽  
Physiological Regulation ◽  
Functional Relationships ◽  
Tight Junction Regulation

Tight junctions restrict diffusion of hydrophilic solutes through the paracellular pathways of columnar epithelia. It is now apparent that the barrier function of tight junctions is physiologically regulated. Current models of the tight junction envisage junctional subunits consisting of extracellular "kisses" between plasma membranes of adjacent cells, intramembrane components represented by freeze-fracture fibrils, and cytoplasmic elements of the cytoskeleton. Insights into functional relationships between these various components of tight junctions should be provided by mapping component interrelationships in states of altered junctional permeability. Here we define the spatial distribution of ZO-1 during a state of physiological regulation of intestinal absorptive cell tight junctions. Enhanced permeation of absorptive cell junctions in response to activation of apical membrane Na(+)-solute cotransporters does not lead to redistribution of the ZO-1 pool, as judged from quantitative ultrastructural immunolocalization studies employing two different ZO-1 antibodies. Surprisingly, ZO-1, which normally localizes under junctional kisses/fibrils, focally persists at sites where junctional kisses/fibrils are cleared. These findings suggest that 1) spatial redistribution of ZO-1 does not contribute to physiological regulation of junctions elicited by activation of Na(+)-solute cotransport and 2) ZO-1 and junctional fibrils may spatially dissociate during such regulated states.

Phosphorylation of occludin correlates with occludin localization and function at the tight junction

1997 ◽  
Vol 273 (6) ◽  
pp. C1859-C1867 ◽  
Author(s):  
Vivian Wong
Keyword(s):  
Molecular Weight ◽  
Tight Junction ◽  
Tight Junctions ◽  
Growth Medium ◽  
Mdck Cells ◽  
Low Calcium ◽  
Phosphatase Treatment ◽  
And Function ◽  
Triton X ◽  
Mdck I

Multiple forms of occludin were found in Madin-Darby canine kidney (MDCK) cells. In the absence of cell-to-cell contacts, achieved by incubating cells in low-calcium growth medium, a cluster of lower-molecular-weight (LMW) occludin bands (∼65,000–68,000) was present in both MDCK I and II cells. On formation of tight junctions, achieved by changing the low-calcium growth medium to normal-calcium growth medium, a cluster of higher-molecular-weight (HMW) bands (∼72,000–75,000 for MDCK I cells and ∼70,000–73,000 for MDCK II cells) was also expressed. The HMW occludin bands could be eliminated by phosphatase treatment. Therefore, the HMW forms of occludin appeared to be the hyperphosphorylated product of the LMW forms. These HMW forms were Triton X-100 insoluble, which correlated with their localization at the tight junctions. Furthermore, depletion of tight junction-localized occludin by an occludin extracellular domian peptide (20) correlated with a decrease in the HMW forms of occludin. In conclusion, phosphorylation of occludin may be a mechanism by which occludin localization and function are regulated.

scholarly journals Protein kinase Cζ phosphorylates occludin and promotes assembly of epithelial tight junctions

2011 ◽  
Vol 437 (2) ◽  
pp. 289-299 ◽  
Author(s):  
Suneet Jain ◽  
Takuya Suzuki ◽  
Ankur Seth ◽  
Geetha Samak ◽  
Radhakrishna Rao
Keyword(s):  
Protein Kinase ◽  
Tight Junction ◽  
Tight Junctions ◽  
Intercellular Junctions ◽  
Cell Monolayers ◽  
Terminal Domain ◽  
Tight Junction Regulation ◽  
Epithelial Tight Junctions ◽  
Zona Occludens

Protein kinases play an important role in the regulation of epithelial tight junctions. In the present study, we investigated the role of PKCζ (protein kinase Cζ) in tight junction regulation in Caco-2 and MDCK (Madin–Darby canine kidney) cell monolayers. Inhibition of PKCζ by a specific PKCζ pseudosubstrate peptide results in redistribution of occludin and ZO-1 (zona occludens 1) from the intercellular junctions and disruption of barrier function without affecting cell viability. Reduced expression of PKCζ by antisense oligonucleotide or shRNA (short hairpin RNA) also results in compromised tight junction integrity. Inhibition or knockdown of PKCζ delays calcium-induced assembly of tight junctions. Tight junction disruption by PKCζ pseudosubstrate is associated with the dephosphorylation of occludin and ZO-1 on serine and threonine residues. PKCζ directly binds to the C-terminal domain of occludin and phosphorylates it on threonine residues. Thr403, Thr404, Thr424 and Thr438 in the occludin C-terminal domain are the predominant sites of PKCζ-dependent phosphorylation. A T424A or T438A mutation in full-length occludin delays its assembly into the tight junctions. Inhibition of PKCζ also induces redistribution of occludin and ZO-1 from the tight junctions and dissociates these proteins from the detergent-insoluble fractions in mouse ileum. The present study demonstrates that PKCζ phosphorylates occludin on specific threonine residues and promotes assembly of epithelial tight junctions.

When Capillary Permeability Increases

Physiology ◽  
1987 ◽  
Vol 2 (1) ◽  
pp. 16-18
Author(s):  
C Crone
Keyword(s):  
Endothelial Cells ◽  
Tight Junction ◽  
Tight Junctions ◽  
Cell Shape ◽  
Capillary Permeability ◽  
Signal Molecule ◽  
Small Pore ◽  
Myosin Filaments ◽  
Tight Junction Permeability

Capillary permeability is usually ascribed to a number of "small" pores. In this article the small-pore pathway is identified as interruptions in the tight junctions between endothelial cells. Modulation of capillary permeability involves control of tight junction permeability with cytosolic Ca2+ as the important signal molecule. Actin and myosin filaments are probably involved in the process, which may implicate subtle changes in cell shape.

Copper treatment alters the permeability of tight junctions in cultured human intestinal Caco-2 cells

1999 ◽  
Vol 277 (6) ◽  
pp. G1138-G1148 ◽  
Author(s):  
Simonetta Ferruzza ◽  
Maria-Laura Scarino ◽  
Giuseppe Rotilio ◽  
Maria Rosa Ciriolo ◽  
Paolo Santaroni ◽  
...  
Keyword(s):  
Tight Junction ◽  
Tight Junctions ◽  
Electrical Resistance ◽  
Transepithelial Electrical Resistance ◽  
Complete Medium ◽  
Zonula Occludens ◽  
Intracellular Effect ◽  
Tight Junction Permeability ◽  
Zonula Occludens 1 ◽  
Effect Of Copper

The effects of copper on tight-junction permeability were investigated in human intestinal Caco-2 cells, monitoring transepithelial electrical resistance and transepithelial passage of mannitol. Apical treatment of Caco-2 cells with 10–100 μM CuCl2(up to 3 h) produced a time- and concentration-dependent increase in tight-junction permeability, reversible after 24 h in complete medium in the absence of added copper. These effects were not observed in cells treated with copper complexed to l-histidine [Cu(His)2]. The copper-induced increase in tight-junction permeability was affected by the pH of the apical medium, as was the apical uptake of64CuCl2, both exhibiting a maximum at pH 6.0. Treatment with CuCl2produced a concentration-dependent reduction in the staining of F actin but not of the junctional proteins zonula occludens-1, occludin, and E-cadherin and produced ultrastructural alterations to microvilli and tight junctions that were not observed after treatment with up to 200 μM Cu(His)2for 3 h. Overall, these data point to an intracellular effect of copper on tight junctions, mediated by perturbations of the F actin cytoskeleton.

scholarly journals Localization of Dysfunctional Tight Junctions inSalmonella enterica Serovar Typhimurium-Infected Epithelial Layers

2000 ◽  
Vol 68 (12) ◽  
pp. 7202-7208 ◽  
Author(s):  
Mark A. Jepson ◽  
Hélène B. Schlecht ◽  
Carla B. Collares-Buzato
Keyword(s):  
Protein Kinase ◽  
Tight Junction ◽  
Tight Junctions ◽  
Diffusion Barrier ◽  
Barrier Function ◽  
Kinase Inhibitor ◽  
Serovar Typhimurium ◽  
Tight Junction Permeability ◽  
Epithelial Layers ◽  
Fence Function

ABSTRACT Infection of polarized MDCK epithelial layers by Salmonella enterica serovar Typhimurium is accompanied by increased tight junction permeability and by contraction of perijunctional actinomyosin. We localized dysfunctional tight junctions in serovar Typhimurium-infected MDCK layers by imaging apical-basolateral intramembrane diffusion of fluorescent lipid and found that loss of the apical-basolateral diffusion barrier (tight junction fence function) was most marked in areas of prominent perijunctional contraction. The protein kinase inhibitor staurosporine prevented perijunctional contraction but did not reverse the effects of serovar Typhimurium on tight junction barrier function. Hence, perijunctional contraction is not required for Salmonella-induced tight junction dysfunction and this epithelial response to infection may be multifactorial.

scholarly journals Elevated plasma cortisol reduces permeability of mammary tight junctions in the lactating bovine mammary epithelium

1998 ◽  
Vol 159 (1) ◽  
pp. 173-178 ◽  
Author(s):  
K Stelwagen ◽  
DC van Espen ◽  
GA Verkerk ◽  
HA McFadden ◽  
VC Farr
Keyword(s):  
Tight Junction ◽  
Tight Junctions ◽  
Plasma Cortisol ◽  
Control Period ◽  
Mammary Epithelium ◽  
Elevated Plasma ◽  
Glucose Levels ◽  
Tight Junction Permeability ◽  
Jersey Cows

Induction of tight junction permeability in the mammary epithelium decreases milk secretion, and in cows tight junctions become leaky after 17 h of milk accumulation.In vitro studies demonstrate the importance of glucocorticoids for the formation and maintenance of tight junctions. In this study we examined whether cortisol can prevent mammary tight junction permeability in the lactating gland in vivo, and inhibit the associated milk loss, using our milk-accumulation model to challenge tight junction patency. Following a 4-day control period Jersey cows were subjected to a 24-h period in which they were milked twice at 0700 and 1500 h (TM;n=6), once at 0700 h (OM;n=7), or once and treated with ACTH (40 IU per 2 h, starting after 14 h of milk accumulation) to increase endogenous cortisol levels (OM+ACTH;n=7). Frequent blood samples for cortisol, lactose and glucose analyses were taken via indwelling jugular catheters. ACTH treatment resulted in a sustained elevation of systemic cortisol concentrations. Plasma lactose, an indicator of tight junction leakiness, was not changed in TM cows, but began to increase rapidly at 17 h of milk accumulation in OM cows. Treatment with ACTH prevented the increase in plasma lactose, although levels were slightly, but not significantly, higher than in TM cows, indicating that elevated plasma cortisol reduced mammary tight junction leakiness. Milk yield was reduced by 12% in both once-milked groups, despite cortisol preventing tight junction leakiness. However, the milk loss in the latter group may not be related to leaky tight junctions, but be due to a reduction in milk precursor uptake by the mammary gland. Consistent with this notion was a 34% increase in plasma glucose levels in OM+ACTH cows only.

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