Assessment of quantitative methods for enhancement measurement on vessel wall magnetic resonance imaging evaluation of intracranial atherosclerosis.

PURPOSE: Quantitative measures of vessel wall magnetic resonance imaging (vwMRI) for the evaluation of intracranial atherosclerotic disease (ICAD) offers standardization not available with previously used qualitative approaches that may be difficult to replicate. METHODS: vwMRI studies performed to evaluate ICAD that had caused a stroke were analyzed. Two blinded reviewers qualitatively rated culprit lesions for the presence of enhancement on T1 delay alternating with nutation for tailored excitation (DANTE) SPACE images. At least 3 months later, quantitative analysis was performed of the same images, comparing lesion enhancement to reference structures. Cohen's kappa and intraclass correlation coefficients were calculated to assess agreement. Ratios of enhancement of lesions to references were compared to qualitative ratings. RESULTS: Studies from 54 patients met inclusion criteria. A mean of 49 (90.7%) lesions were qualitatively rated as enhancing, with good inter-rater agreement (κ = 0.783). Among reference structure candidates, low infundibulum demonstrated the highest inter-rater agreement on pre- and post-contrast imaging. The ratio of percentage increase in plaque signal following contrast to the same measure in low infundibulum demonstrated the highest agreement with qualitative assessment, with highest agreement seen with a ratio of 0.8 set as a threshold (κ = 0.675). CONCLUSION: Quantitative metrics can yield objective data to better standardize techniques and acceptance of vwMRI evaluation of ICAD. The low infundibulum had the highest inter-rater agreement on both pre- and post-contrast images and is best suited as a normally enhancing reference structure. Such quantitative techniques should be implemented in future research of vwMRI for the evaluation of ICAD.

The role of the multidrug resistance protein-1 in modulation of endothelial cell oxidative stress.

Glutathione (GSH) is the major source of intracellular sulfhydryl groups. Oxidized GSH (GSSG) can be recycled to GSH by the GSH reductase or exported from the cell. The mechanism by which GSSG is exported and the consequence of its export from endothelial cells has not been defined previously. We found that human endothelial cells express the multidrug resistance protein-1 (MRP1) and use this as their major exporter of GSSG. Oscillatory shear stress, which is known to stimulate endothelial cell production of reactive oxygen species, decreased intracellular GSH. In contrast, laminar shear significantly increased intracellular GSH. Oscillatory shear also caused a robust export of GSSG that was prevented by the MRP1 inhibitor MK571 and by MRP1 small interfering RNA. MRP1 inhibition prevented the decline in intracellular GSH, preserved the intracellular GSH Nernst potential, and reduced apoptosis caused by oscillatory shear. In aortas of hypertensive mice, endothelial disulfide export was doubled, and this was prevented by MK571 and was not observed in aortas of hypertensive MRP1-/- mice. Further, the altered endothelium-dependent vasodilatation caused by hypertension was ameliorated in MRP1-/- mice. GSSG export by MRP1 leads to a perturbation of endothelial redox state and ultimately endothelial cell apoptosis. Endothelial MRP1 may provide a novel therapeutic target for prevention of vascular disease.

Oscillatory shear stress upregulation of endothelial nitric oxide synthase requires intracellular hydrogen peroxide and CaMKII.

We have previously shown that hydrogen peroxide (H(2)O(2)) upregulates endothelial nitric oxide synthase (eNOS) expression via a calcium/calmodulin-dependent protein kinase II (CaMKII)-mediated mechanism whereas it also acutely activates eNOS enzyme. We hypothesized that oscillatory shear stress (OSS), which stimulates endogenous H(2)O(2), would have effects on eNOS expression and function similar to that of exogenous H(2)O(2). Exposure of bovine aortic endothelial cells to OSS (+/-15 dynes/cm(2)) increased eNOS mRNA expression by 3-fold. Pretreatment with either polyethylene glycol-catalase (PEG-CAT, a scavenger of H(2)O(2)) or KN93, an inhibitor of CaMKII, abolished this response. OSS activated CaMKII in an H(2)O(2)-dependent fashion whereas unidirectional laminar shear stress (LSS) inhibited CaMKII phosphorylation. Inhibition of c-Src (essential for LSS upregulation of eNOS) had no effect on OSS upregulation of eNOS. Additionally, OSS stimulated NO* production acutely. Scavenging of H(2)O(2) by PEG-CAT attenuated OSS stimulation of NO* by 50% whereas it had no effect on LSS regulation of NO* production. These data suggest that intracellular H(2)O(2) and CaMKII mediate OSS upregulation of eNOS. The acute activation of eNOS by OSS also partially requires H(2)O(2). As OSS has been shown previously to stimulate sustained production of superoxide (O(2)*-) which would inactivate NO*, these responses may represent attempted compensation to restore NO* bioavailability in areas exposed to OSS. Simultaneous stimulation of O(2)*- and NO* by this mechanism, however, could facilitate peroxynitrite formation and protein nitration, which may enhance atherosclerotic lesion formation. Both OSS and LSS upregulate eNOS expression but via different signaling mechanisms.