Intracellular Ascorbate Prevents Endothelial Barrier Permeabilization by Thrombin

Parker, W. H.; Qu, Zhi-chao; May, James M.

doi:10.1074/jbc.m115.662098

Cited by 19 publications

(24 citation statements)

References 57 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The cause of thrombin-induced endothelial hyperpermeability involved activation of the small GTPase RhoA and Rho kinase, which increased the phosphorylation of MLC. In addition, Parker et al [43] demonstrated that thrombin induced endothelial barrier destabilization, at least partly, by inhibiting the activation of Rap1 and Rac1 and increasing the phosphorylation and distribution of VE-cadherin, the phosphorylation of MLC, and actin polymerization. Consistent with the results of previous studies, we observed that after endothelial cells were exposed to 0.5 mmol/L OA, the expression of pMLC was increased in the AT III siRNA group compared with that in the control siRNA group.…”

Section: Discussionmentioning

confidence: 99%

ITRAQ-Based Proteomics Analysis of Acute Lung Injury Induced by Oleic Acid in Mice

Zhu

Zhang

et al. 2017

Cell Physiol Biochem

View full text Add to dashboard Cite

Background/Aims: This study was conducted to investigate the relationship between differentially expressed proteins (DEPs) and the pathogenesis of oleic acid (OA)-induced acute lung injury (ALI) in mice. Methods: Eight-week-old male C57BL/6 mice were injected with OA through the tail vein and sacrificed 6 hours after OA administration to identify protein expression levels in lung tissue using isobaric tags for relative and absolute quantification (iTRAQ) technology. Then, DEPs such as antithrombin III (AT III), 12-lipoxygenase (12-LO), dedicator of cytokinesis 2 (DOCK2), polycystin-2 and plasminogen were identified by western blotting. Subsequently, we focused on investigating the effect of AT III on endothelial integrity using siRNA interference technology. The levels of IL-6, IL-1β, TNF-α and TGF-β expression were detected using an enzyme-linked immunosorbent assay (ELISA). Alterations in the tight junction component ZO-1 and the phosphorylation of myosin light chain (pMLC) were determined by western blotting. The stress fiber F-actin were also detected by immunofluorescence staining. In addition, endothelial permeability was determined via a transwell permeability assay. Results: A total of 5152 proteins were found to be expressed in lung tissues from the OA-treated and saline-treated mice. Among these proteins, 849 were differentially expressed between the two groups, including 545 upregulated and 304 downregulated proteins. After AT III knockdown, the levels of inflammatory factors and endothelial permeability were elevated, the expression of ZO-1 was decreased, and the expression of F-actin and pMLC was increased. All these results illustrated that AT III knockdown exaggerated the disruption of endothelial integrity mediated by OA. Conclusion: These findings using iTRAQ technology demonstrate, for the first time, differences in the lung tissue expression levels of proteins between OA-treated mice and saline-treated mice. This study reveals that 12-LO, DOCK2 and especially AT III may be candidate biomarkers for OA-induced acute lung injury.

show abstract

Section: Discussionmentioning

confidence: 99%

ITRAQ-Based Proteomics Analysis of Acute Lung Injury Induced by Oleic Acid in Mice

Zhu

Zhang

et al. 2017

Cell Physiol Biochem

View full text Add to dashboard Cite

show abstract

“…Whatever the source of oxidative stress, it is clear that increasing cellular antioxidant capacity might be beneficial in decreasing VEGF-induced hyperpermeability. In this regard, we have shown that vitamin C, or ascorbic acid, prevents endothelial barrier leakage due to oxidized low density lipoprotein [14] and more recently due to thrombin [15]. Although ascorbate is considered an important cellular antioxidant, in both basal [16] and thrombin-activated cells [15], the vitamin enhanced the function of eNOS to generate nitric oxide (NO).…”

Section: Introductionmentioning

confidence: 99%

“…In this regard, we have shown that vitamin C, or ascorbic acid, prevents endothelial barrier leakage due to oxidized low density lipoprotein [14] and more recently due to thrombin [15]. Although ascorbate is considered an important cellular antioxidant, in both basal [16] and thrombin-activated cells [15], the vitamin enhanced the function of eNOS to generate nitric oxide (NO). It is known from a previous study that ascorbate promotes NO generation by recycling the crucial eNOS co-factor tetrahydrobiopterin [17].…”

Section: Introductionmentioning

confidence: 99%

Ascorbic acid prevents VEGF-induced increases in endothelial barrier permeability

et al. 2015

Self Cite

View full text Add to dashboard Cite

Vascular endothelial growth factor (VEGF) increases endothelial barrier permeability, an effect that may contribute to macular edema in diabetic retinopathy. Since vitamin C, or ascorbic acid, can tighten the endothelial permeability barrier, we examined whether it could prevent the increase in permeability due to VEGF in human umbilical vein endothelial cells (HUVECs). As previously observed, VEGF increased HUVEC permeability to radiolabeled inulin within 60 min in a concentration-dependent manner. Loading the cells with increasing concentrations of ascorbate progressively prevented the leakage caused by 100 ng/ml VEGF, with a significant inhibition at 13 μM and complete inhibition at 50 μM. Loading cells with 100 μM ascorbate also decreased basal generation of reactive oxygen species and prevented the increase caused by both 100 ng/ml VEGF. VEGF treatment decreased intracellular ascorbate by 25%, thus linking ascorbate oxidation to its prevention of VEGF-induced barrier leakage. The latter was blocked by treating the cells with 60 μM L-NAME (but not D-NAME) as well as by 30 μM sepiapterin, a precursor of tetrahydrobiopterin that is required for proper function of endothelial nitric oxide synthase (eNOS). These findings suggest that VEGF-induced barrier leakage uncouples eNOS. Ascorbate inhibition of the VEGF effect could thus be due either to scavenging superoxide or to peroxynitrite generated by the uncoupled eNOS, or more likely to its ability to recycle tetrahydrobiopterin, thus avoiding enzyme uncoupling in the first place. Ascorbate prevention of VEGF-induced increases in endothelial permeability opens the possibility that its repletion could benefit diabetic macular edema.

show abstract

“…Ascorbate loading of HUVECs also completely prevented the endothelial barrier leak caused by short-term treatment with the inflammatory agent thrombin [111]. In that study, ascorbate acting through NO increased cyclic GMP and prevented the thrombin-induced decrease in cyclic AMP.…”

Section: Ascorbate Functions That May Decrease Pericyte Loss and Endomentioning

confidence: 95%

Ascorbic acid repletion: A possible therapy for diabetic macular edema?

May

2016

Free Radical Biology and Medicine

Self Cite

View full text Add to dashboard Cite

Macular edema poses a significant risk for visual loss in persons with diabetic retinopathy. It occurs when plasma constituents and fluid leak out of damaged retinal microvasculature in the area of the macula, causing loss of central vision. Apoptotic loss of pericytes surrounding capillaries is perhaps the earliest feature of diabetic vascular damage in the macula, which is also associated with dysfunction of the endothelium and loss of the otherwise very tight endothelial permeability barrier. Increased oxidative stress is a key feature of damage to both cell types, mediated by excess superoxide from glucose-induced increases in mitochondrial metabolism, as well as by activation of the receptor for advanced glycation end products (RAGE). The latter in turn activates multiple pathways, some of which lead to increased oxidative stress, such as those involving NF-κB, NADPH oxidase, and endothelial nitric oxide synthase. Such cellular oxidative stress is associated with low cellular and plasma ascorbic acid levels in many subjects with diabetes in poor glycemic control. Whether repletion of low ascorbate in retinal endothelium and pericytes might help to prevent diabetic macular edema is unknown. However, cell culture studies show that the vitamin prevents high-glucose and RAGE-induced apoptosis in both cell types, that it preserves nitric oxide generated by endothelial cells, and that it tightens the leaky endothelial permeability barrier. Although these findings need to be confirmed in pre-clinical animal studies, it is worth considering clinical trials to determine whether adequate ascorbate repletion is possible and whether it might help to delay or even reverse early diabetic macular edema.

show abstract

Intracellular Ascorbate Prevents Endothelial Barrier Permeabilization by Thrombin

Cited by 19 publications

References 57 publications

ITRAQ-Based Proteomics Analysis of Acute Lung Injury Induced by Oleic Acid in Mice

ITRAQ-Based Proteomics Analysis of Acute Lung Injury Induced by Oleic Acid in Mice

Ascorbic acid prevents VEGF-induced increases in endothelial barrier permeability

Ascorbic acid repletion: A possible therapy for diabetic macular edema?

Contact Info

Product

Resources

About