scholarly journals Role of Defensins in Corneal Epithelial Barrier Function againstPseudomonas aeruginosaTraversal

2010 ◽  
Vol 79 (2) ◽  
pp. 595-605 ◽  
Author(s):  
Danielle K. Augustin ◽  
Susan R. Heimer ◽  
Connie Tam ◽  
Wing Y. Li ◽  
Jeff M. Le Due ◽  
...  
Keyword(s):  
Barrier Function ◽  
Ex Vivo ◽  
Bacterial Colonization ◽  
Corneal Epithelial Cell ◽  
Epithelial Barrier ◽  
Epithelial Barrier Function ◽  
Corneal Epithelial

ABSTRACTStudies have shown that epithelium-expressed antimicrobial peptides (AMPs), e.g., β-defensins, play a role in clearing bacteria from mouse corneas already infected withPseudomonas aeruginosa. Less is known about the role of AMPs in allowing the cornea to resist infection when healthy. We previously reported that contact lens exposure, a major cause ofP. aeruginosakeratitis, can inhibit the upregulation of human β-defensin 2 (hBD-2) by corneal epithelial cells in response toP. aeruginosaantigensin vitro. Here, we studied the role of AMPs in maintaining the corneal epithelial barrier toP. aeruginosapenetration using bothin vitro(human) andin vivo(mouse) experiments. Results showed that preexposing human corneal epithelial multilayers to bacterial antigens in a culture supernatant (known to upregulate AMP expression) reduced epithelial susceptibility toP. aeruginosatraversal up to 6-fold (P< 0.001). Accordingly, small interfering RNA (siRNA) knockdown of any one of four AMPs expressed by human epithelia promotedP. aeruginosatraversal by more than 3-fold (P< 0.001). The combination knockdown of AMPs further enhanced susceptibility to bacterial traversal by ∼8-fold (P< 0.001).In vivoexperiments showed that the loss of murine β-defensin 3 (mBD-3), a murine ortholog of hBD-2, enhanced corneal susceptibility toP. aeruginosa. The uninjured ocular surface of mBD-3−/−mice showed a reduced capacity to clearP. aeruginosa, and their corneal epithelia were more susceptible to bacterial colonization, even when inoculatedex vivoto exclude tear fluid effects. Together, thesein vitroandin vivodata show functional roles for AMPs in normal corneal epithelial cell barrier function againstP. aeruginosa.

scholarly journals The role of secreted Hsp90α in HDM-induced asthmatic airway epithelial barrier dysfunction

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Cuiping Ye ◽  
Chaowen Huang ◽  
Mengchen Zou ◽  
Yahui Hu ◽  
Lishan Luo ◽  
...  
Keyword(s):  
Barrier Function ◽  
Epithelial Barrier ◽  
Airway Epithelial ◽  
Barrier Dysfunction ◽  
Epithelial Barrier Function ◽  
Asthmatic Airway ◽  
Epithelial Barrier Dysfunction

Abstract Background The dysfunction of airway epithelial barrier is closely related to the pathogenesis of asthma. Secreted Hsp90α participates in inflammation and Hsp90 inhibitor protects endothelial dysfunction. In the current study, we aimed to explore the role of secreted Hsp90α in asthmatic airway epithelial barrier function. Methods Male BALB/c mice were sensitized and challenged with HDM to generate asthma model. The 16HBE and Hsp90α-knockdown cells were cultured and treated according to the experiment requirements. Transepithelial Electric Resistance (TEER) and permeability of epithelial layer in vitro, distribution and expression of junction proteins both in vivo and in vitro were used to evaluate the epithelial barrier function. Western Blot was used to evaluate the expression of junction proteins and phosphorylated AKT in cells and lung tissues while ELISA were used to evaluate the Hsp90α expression and cytokines release in the lung homogenate. Results HDM resulted in a dysfunction of airway epithelial barrier both in vivo and in vitro, paralleled with the increased expression and release of Hsp90α. All of which were rescued in Hsp90α-knockdown cells or co-administration of 1G6-D7. Furthermore, either 1G6-D7 or PI3K inhibitor LY294002 suppressed the significant phosphorylation of AKT, which caused by secreted and recombinant Hsp90α, resulting in the restoration of epithelial barrier function. Conclusions Secreted Hsp90α medicates HDM-induced asthmatic airway epithelial barrier dysfunction via PI3K/AKT pathway, indicating that anti-secreted Hsp90α therapy might be a potential treatment to asthma in future.

cis 9, trans 11, but not trans 10, cis 12 CLA isomer, impairs intestinal epithelial barrier function in IPEC-J2 cells and mice through activation of GPR120-[Ca2+]i and the MLCK signaling pathway

2020 ◽  
Vol 11 (4) ◽  
pp. 3657-3667
Author(s):  
Han Su ◽  
Weijie Zhao ◽  
Fenglin Zhang ◽  
Min Song ◽  
Fangfang Liu ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Barrier Function ◽  
Epithelial Barrier ◽  
Intestinal Epithelial ◽  
Intestinal Epithelial Barrier ◽  
Epithelial Barrier Function

In vitro and in vivo studies show that c9, t11-CLA, but not t10, c12-CLA isomer, impairs intestinal epithelial barrier function in IPEC-J2 cells and mice via activation of GPR120-[Ca2+]i and the MLCK pathway.

14 COLONIC EPITHELIAL CELL-SPECIFIC AFTIPHILIN KNOCKDOWN REGULATES EPITHELIAL BARRIER FUNCTION THROUGH MYOSIN LIGHT CHAIN-ASSOCIATED ACTIN ORGANIZATION IN VITRO AND INTESTINAL LENGTH IN VIVO

2020 ◽  
Vol 26 (Supplement_1) ◽  
pp. S28-S28
Author(s):  
Ivy Ka Man Law ◽  
Carl Rankin ◽  
Charalabos Pothoulakis
Keyword(s):  
Epithelial Cell ◽  
Barrier Function ◽  
Intestinal Epithelial Cell ◽  
Epithelial Barrier ◽  
Intestinal Epithelial ◽  
Knock Out ◽  
Epithelial Barrier Function ◽  
Actin Organization

Abstract Background and Aims Colonic epithelial integrity is often compromised during colonic inflammation and Inflammatory Bowel Disease. Aftiphilin (AFTPH) is a downstream target of microRNA-133a and its expression is reduced in colonic tissues of wild type mice from experimental colitis models and colonic biopsies from patients with ulcerative colitis. We have previously shown that AFTPH is involved in regulating intestinal epithelial barrier function and actin organization in human colonic epithelial cells in vitro (DDW 2016). On the other hand, our results suggested that global aftiphilin knock-out is embryonic lethal in mouse models (DDW 2019). Here, we further examined the role of AFTPH in regulating actin organization in vitro and characterize the colonic epithelial cell-specific aftiphilin knock-out mice. Methods Human colonic epithelial NCM460 cells were transfected with si-RNA against AFTPH to achieve transient AFTPH gene-silencing. Stable AFTPH knock-down clones were generated by transducing Caco2-BBE cells with recombinant lentivirus carrying sh-AFTPH or control sh-RNA. To create intestinal epithelial cell-specific aftiphilin knock-out mice, Aftph flox/flox mice were cross-bred with B6.Cg-Tg(Vil1-cre)997Gum/J mice, which express Villin-driven Cre recombinase (Vil-Cre), to generate intestinal epithelial cell-specific aftiphilin knock-out mice (Aftph Vil-/Vil-). Protein expression of F- and G-actin and p70S6K were detected using Western blot. Tissues from various organs were collected with Aftph Vil-/Vil- and its wildtype counterparts at 12 weeks. Results Results from western blot analysis showed that F-/G-actin ratio in AFTPH gene-silenced NCM460 cells were 0.6±0.17 fold, when compared to the treatment control. In addition, AFTPH gene-silencing in human colonic epithelial cells activated p70S6K, a kinase that is involved in actin organization, when compared to treatment control (1.2±0.15 vs. 2.0±0.15, p=0.0354). Furthermore, transepithelial electric resistance (TER) of Caco2-BBE cells deficient in AFTPH is significantly lower than that of control cells (0.5±0.07 fold). Lastly, in vivo intestinal epithelial cell-specific Aftph knock-out increased the length of small intestine, when compared to that of wild type mice (30.7±0.33 vs. 34.8±0.97, p=0.02), while the tissue weight of spleen to body weight was reduced (0.30±0.011 vs. 0.26±0.006, p=0.0169). Summary and Conclusions Our results indicate that AFTPH directly regulates epithelial barrier function and actin organization through mediating F-/G-actin ratio in human colonic epithelial cells, possibly through p70S6K. Importantly, intestinal epithelial cell-specific knock-out in vivo increased intestinal length and reduced size of the spleen. Our results suggested that AFTPH is crucial in regulating colonic epithelial barrier function in vitro and intestinal homeostasis.

Breast Milk and Formula Feeding Affect Intestinal Epithelial Barrier Function in Vivo and in Vitro

2012 ◽  
Vol 172 (2) ◽  
pp. 302
Author(s):  
V. Poroyko ◽  
T. Mirzapoiazova ◽  
E.M. Carlisle ◽  
M.S. Caplan ◽  
J. Alverdy ◽  
...  
Keyword(s):  
Breast Milk ◽  
Barrier Function ◽  
Formula Feeding ◽  
Epithelial Barrier ◽  
Intestinal Epithelial ◽  
Intestinal Epithelial Barrier ◽  
Epithelial Barrier Function

The Role of Polyphenols in the Modulation of Plasma Non-Enzymatic Antioxidant Capacity (NEAC)

2012 ◽  
Vol 82 (3) ◽  
pp. 228-232 ◽  
Author(s):  
Mauro Serafini ◽  
Giuseppa Morabito
Keyword(s):  
Antioxidant Capacity ◽  
Dietary Intervention ◽  
Ex Vivo ◽  
The Body ◽  
Dietary Polyphenols ◽  
Enzymatic Antioxidant ◽  
Endogenous Antioxidant

Dietary polyphenols have been shown to scavenge free radicals, modulating cellular redox transcription factors in different in vitro and ex vivo models. Dietary intervention studies have shown that consumption of plant foods modulates plasma Non-Enzymatic Antioxidant Capacity (NEAC), a biomarker of the endogenous antioxidant network, in human subjects. However, the identification of the molecules responsible for this effect are yet to be obtained and evidences of an antioxidant in vivo action of polyphenols are conflicting. There is a clear discrepancy between polyphenols (PP) concentration in body fluids and the extent of increase of plasma NEAC. The low degree of absorption and the extensive metabolism of PP within the body have raised questions about their contribution to the endogenous antioxidant network. This work will discuss the role of polyphenols from galenic preparation, food extracts, and selected dietary sources as modulators of plasma NEAC in humans.

scholarly journals Heparan sulfate and heparin interactions with proteins

2015 ◽  
Vol 12 (110) ◽  
pp. 20150589 ◽  
Author(s):  
Maria C. Z. Meneghetti ◽  
Ashley J. Hughes ◽  
Timothy R. Rudd ◽  
Helena B. Nader ◽  
Andrew K. Powell ◽  
...  
Keyword(s):  
Heparan Sulfate ◽  
Ex Vivo ◽  
Sequence Specificity ◽  
Structure Activity ◽  
Multiple Binding Sites ◽  
Multiple Binding ◽  
Heparin Activity

Heparan sulfate (HS) polysaccharides are ubiquitous components of the cell surface and extracellular matrix of all multicellular animals, whereas heparin is present within mast cells and can be viewed as a more sulfated, tissue-specific, HS variant. HS and heparin regulate biological processes through interactions with a large repertoire of proteins. Owing to these interactions and diverse effects observed during in vitro , ex vivo and in vivo experiments, manifold biological/pharmacological activities have been attributed to them. The properties that have been thought to bestow protein binding and biological activity upon HS and heparin vary from high levels of sequence specificity to a dependence on charge. In contrast to these opposing opinions, we will argue that the evidence supports both a level of redundancy and a degree of selectivity in the structure–activity relationship. The relationship between this apparent redundancy, the multi-dentate nature of heparin and HS polysaccharide chains, their involvement in protein networks and the multiple binding sites on proteins, each possessing different properties, will also be considered. Finally, the role of cations in modulating HS/heparin activity will be reviewed and some of the implications for structure–activity relationships and regulation will be discussed.

scholarly journals The pathophysiologic role of the protein kinase Cδ pathway in the intervertebral discs of rabbits and mice: In vitro, ex vivo, and in vivo studies

2012 ◽  
Vol 64 (6) ◽  
pp. 1950-1959 ◽  
Author(s):  
Michael B. Ellman ◽  
Jae-Sung Kim ◽  
Howard S. An ◽  
Jeffrey S. Kroin ◽  
Xin Li ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Ex Vivo ◽  
Intervertebral Discs ◽  
In Vivo Studies ◽  
Protein Kinase Cδ
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