Search Results (5)

Background/objectives: The porcine intestinal epithelium plays a fundamental role as a defence interface against pathogens. Its alteration can cause severe inflammatory conditions and diseases. Hyperosmotic stress under physiological conditions and upon pathogen challenge can cause malabsorption. Different cell types counteract the osmolarity increase by accumulating organic osmolytes such as betaine, taurine, and myo-inositol through specific transporters. Betaine is known for protecting cells from hyperosmotic stress and has positive effects when fed to pigs. The aim of this study is to demonstrate the modulation of osmolyte transporters gene expression in IPEC-J2 during osmolarity changes and assess the effects of betaine. Methods: IPEC-J2 were seeded in transwells, where differentiate as a polarized monolayer. Epithelial cell integrity (TEER), oxidative stress (NO) and gene expression of osmolyte transporters, tight junction proteins (TJp) and pro-inflammatory cytokines were evaluated. Results: Cells treated with NaCl hyperosmolar medium (500 mOsm/L) showed a TEER decrease at 3 h and detachment within 24 h, associated with an osmolyte transporters reduction. IPEC-J2 treated with mannitol hyperosmolar medium (500 mOsm/L) upregulated taurine (TauT), myo-inositol (SMIT) and betaine (BGT1) transporters expression. A decrease in TJp expression was associated with a TEER decrease and an increase in TNFα, IL6, and IL8. Betaine could attenuate the hyperosmolarity-induced reduction in TEER and TJp expression, the NO increase and cytokines upregulation. Conclusions: This study demonstrates the expression of osmolyte transporters in IPEC-J2, which was upregulated upon hyperosmotic treatment. Betaine counteracts changes in intracellular osmolarity by contributing to maintaining the epithelial barrier function and reducing the inflammatory condition. Compatible osmolytes may provide beneficial effects in therapies for diseases characterized by inflammation and TJp-related dysfunctions. Full article

Dry eye disease (DED) is a common ocular disorder characterized by an inadequate lubrication of the eye by tears leading to inflammation and the alteration of the ocular surface. Current treatments are often limited due to their side effects and ineffectiveness. Thymoquinone (TQ) is a natural compound present in the essential oil of Nigella sativa L., with anti-inflammatory and antioxidant activities. In this study, conventional and hyaluronic acid-coated liposomes were developed to improve TQ activity at ocular level. In the present study, the cytoprotective effects of TQ or TQ liposomes were assessed against oxidative and inflammatory processes in human corneal epithelial cells (HCE-2). Hyperosmolarity conditions (450 mOsm) were used as a model of DED. Interleukin-1β (IL-1β), Interleukin-6 (IL-6) and tumor necrosis factor (TNFα) were quantified by quantitative real-time polymerase chain reaction (RT-qPCR); COX-2 and Phospho-NF-κB p65 (p-p65) by Western blotting (WB). Moreover, the mitochondrial reactive oxygen species (mtROS) levels were measured by MitoSOX assay. The hyperosmotic treatment induced a significant increase of the proinflammatory genes and proteins expression that were significantly decreased in the liposomes-treated cells. The coincubation with hyaluronic acid-coated liposomes significantly reverted the increase of mtROS production, evidently stimulated by the hyperosmotic stress. Our data suggest that TQ-loaded liposomes have potential as a therapeutic agent in dry eye disease, improving the TQ efficacy. Full article

Tear hyperosmolarity plays an essential role in the initiation and progression of dry-eye disease. Under a hyperosmotic environment, corneal epithelial cells experience perturbations in endoplasmic reticulum function that can lead to proinflammatory signaling and apoptosis. In this study, we investigated the effect of tauroursodeoxycholic acid (TUDCA), a chemical chaperone known to protect against endoplasmic reticulum stress, on corneal epithelial cells exposed to hyperosmotic conditions. We found that the expression of the genes involved in the activation of the unfolded protein response and the pro-apoptotic transcription factor DDIT3 were markedly upregulated in patients with Sjögren’s dry-eye disease and in a human model of corneal epithelial differentiation following treatment with hyperosmotic saline. Experiments in vitro demonstrated that TUDCA prevented hyperosmotically induced cell death by reducing nuclear DNA fragmentation and caspase-3 activation. TUDCA supplementation also led to the transcriptional repression of CXCL8 and IL5, two inflammatory mediators associated with dry-eye pathogenesis. These studies highlight the role of hyperosmotic conditions in promoting endoplasmic reticulum stress in the cornea and identify TUDCA as a potential therapeutic agent for the treatment of dry-eye disease. Full article

Duchenne muscular dystrophy (DMD) is characterized by chronic inflammation and fibrotic tissue production by fibroblasts. The promyogenic factor nuclear factor of activated T-cells 5 (NFAT5) is virtually present in all cells, responding to hyperosmolar or pro-inflammatory stress. In embryogenic fibroblasts, absence of NFAT5 results in cell cycle arrest. Here, unaffected skeletal muscle fibroblasts from one healthy donor showed NFAT5 nuclear translocation upon hyperosmolar stress and normal cell viability. Absence of NFAT5 translocation under pro-inflammatory conditions resulted in decreased cell growth (Incucyte ZOOM). In DMD skeletal muscle fibroblasts from one DMD patient, NFAT5 was merely located in the nucleus. Exposure to hyperosmolar conditions or pro-inflammatory cytokines IFN-γ, IL-1β and TNF-α had no influence on NFAT5 physiology (immunofluorescence, western blotting, RT-qPCR). Hyperosmolarity resulted in decreased cell viability and pro-inflammatory stress in unaltered cell growth. These findings suggest that NFAT5 is vital to DMD fibroblast survival. Exposure to pro-inflammatory or hyperosmolar stress in DMD fibroblasts results in an unexpected NFAT5 response, where fibroblasts are not triggered by inflammatory cytokines and do not withstand hyperosmolarity. Chronic inflammation could be viewed as a non-restrictive factor in the formation of fibrosis in DMD. Abnormal NFAT5 physiology could provide a molecular explanation for permanent fibrotic matrix production by DMD fibroblasts. Full article

The aim of this study was to assess the anti-inflammatory and anti-apoptotic effects of KIOM-2015EW, the hot-water extract of maple leaves in hyperosmolar stress (HOS)-induced human corneal epithelial cells (HCECs). HCECs were exposed to hyperosmolar medium and exposed to KIOM-2015EW with or without the hyperosmolar media. Tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6 production and apoptosis were observed, and the activation of mitogen-activated protein kinases (MAPKs) including extracellular signal regulated kinase (ERK), p38 and c-JUN N-terminal kinase (JNK) signaling and nuclear factor (NF)-κB was confirmed. Compared to isomolar medium, the induction of cell cytotoxicity significantly increased in HCECs exposed to hyperosmolar medium in a time-dependent manner. KIOM-2015EW-treatment significantly reduced the mRNA and protein expression of pro-inflammatory mediators and apoptosis. KIOM-2015EW-treatment inhibited HOS-induced MAPK signaling activation. Additionally, the HOS-induced increase in NF-κB phosphorylation was attenuated by KIOM-2015EW. The results demonstrated that KIOM-2015EW protects the ocular surface by suppressing inflammation in dry eye disease, and suggest that KIOM-2015EW may be used to treat several ocular surface diseases where inflammation plays a key role. Full article