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Novel high-throughput in vitro model for identifying hemodynamic-induced inflammatory mediators of cerebral aneurysm formation.

ABSTRACT: Cerebral aneurysms are thought to develop at locations of hemodynamic shear stress, via an inflammatory process. The molecular mechanism that links shear stress to inflammation, however, is not completely understood. Progress in studying this disease is limited by a lack of a suitable in vitro model. To address this, we designed novel in vitro parallel-plate flow chamber models of a straight artery, a bifurcation, and a bifurcation aneurysm. We compared endothelial cell phenotypes across the 3 different models and among microenvironments within each flow model by cytokine array, ELISA, and relative immunofluorescence. Human aneurysms express interleukin-8 and chemokine (C-X-C motif) ligand 1 (CXCL1), whereas normal arteries do not. The bifurcation aneurysm model showed significantly higher interleukin-8 and CXCL1 levels than both the straight artery and bifurcation models. Within the bifurcation and bifurcation aneurysm models, endothelial cells near the bifurcation or within the aneurysm sac microenvironments have significantly higher expression of CXCL1, and interleukin-8 and CXCL1, respectively, than at the straight proximal segment or the limbs of the bifurcation. Murine aneurysms express CXCL1, and it is the primary ELR+ CXC chemokine expressed, whereas normal arteries do not. CXCL1 antibody blockade results in significantly fewer murine aneurysms (13.3 versus 66.7%; P=0.0078), decreased neutrophil infiltration, and vascular cell adhesion molecule 1 expression than an immunoglobulin G control. We successfully designed and validated a novel hemodynamic model of cerebral aneurysms in vitro. We also show that shear stress-induced CXCL1 plays a critical role in cerebral aneurysm formation.

SUBMITTER: Nowicki KW

PROVIDER: S-EPMC4231007 | biostudies-literature | 2014 Dec

REPOSITORIES: biostudies-literature

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Novel high-throughput in vitro model for identifying hemodynamic-induced inflammatory mediators of cerebral aneurysm formation.

Nowicki Kamil W KW Hosaka Koji K He Yong Y McFetridge Peter S PS Scott Edward W EW Hoh Brian L BL

Hypertension (Dallas, Tex. : 1979) 20140915 6

Cerebral aneurysms are thought to develop at locations of hemodynamic shear stress, via an inflammatory process. The molecular mechanism that links shear stress to inflammation, however, is not completely understood. Progress in studying this disease is limited by a lack of a suitable in vitro model. To address this, we designed novel in vitro parallel-plate flow chamber models of a straight artery, a bifurcation, and a bifurcation aneurysm. We compared endothelial cell phenotypes across the 3 d ...[more]

PMID: 25225207

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Project description:The growth of cerebral aneurysms is linked to local hemodynamic conditions, but the driving mechanisms of the growth are poorly understood. The goal of this study was to examine the association between intraaneurysmal hemodynamic features and areas of aneurysm growth, to present the key hemodynamic parameters essential for an accurate prediction of the growth, and to gain a deeper understanding of the underlying mechanisms. Patient-specific images of a growing cerebral aneurysm in 3 different growth stages acquired over a period of 40 months were segmented and reconstructed. A unique aspect of this patient-specific case study was that while one side of the aneurysm stayed stable, the other side continued to grow. This unique case enabled the authors to examine their aims in the same patient with parent and daughter arteries under the same inlet flow conditions. Pulsatile flow in the aneurysm models was simulated using computational fluid dynamics and was validated with in vitro experiments using particle image velocimetry measurements. The authors' detailed analysis of intrasaccular hemodynamics linked the growing regions of aneurysms to flow instabilities and complex vortex structures. Extremely low velocities were observed at or around the center of the unstable vortex structure, which matched well with the growing regions of the studied cerebral aneurysm. Furthermore, the authors observed that the aneurysm wall regions with a growth greater than 0.5 mm coincided with wall regions of lower (< 0.5 Pa) time-averaged wall shear stress (TAWSS), lower instantaneous (< 0.5 Pa) wall shear stress (WSS), and high (> 0.1) oscillatory shear index (OSI). To determine which set of parameters can best identify growing and nongrowing aneurysms, the authors performed statistical analysis for consecutive stages of the growing CA. The results demonstrated that the combination of TAWSS and the distance from the center of the vortical structure has the highest sensitivity and positive predictive value, and relatively high specificity and negative predictive value. These findings suggest that an unstable, recirculating flow structure within the aneurysm sac created in the region adjacent to the aneurysm wall with low TAWSS may be introduced as an accurate criterion to explain the hemodynamic conditions predisposing the aneurysm to growth. The authors' findings are based on one patient's data set, but the study lays out the justification for future large-scale verification. The authors' findings can assist clinicians in differentiating stable and growing aneurysms during preinterventional planning.

Project description:Paracrine interactions between adipocytes and macrophages contribute to chronic inflammation in obese adipose tissue. Dietary strategies to mitigate such inflammation include long-chain polyunsaturated fatty acids, docosahexaenoic (DHA) and eicosapentaenoic (EPA) acids, which act through PPARγ-dependent and independent pathways. We utilized an in vitro co-culture model designed to mimic the ratio of macrophages:adipocytes in obese adipose tissue, whereby murine 3T3-L1 adipocytes were cultured with RAW 264.7 macrophages in direct contact, or separated by a trans-well membrane (contact-independent mechanism), with 125 µM of albumin-complexed DHA, EPA, palmitic acid (PA), or albumin alone (control). Thus, we studied the effect of physical cell contact versus the presence of soluble factors, with or without a PPARγ antagonist (T0070907) in order to elucidate putative mechanisms. After 12 hr, DHA was the most anti-inflammatory, decreasing MCP1 and IL-6 secretion in the contact system (-57%, -63%, respectively, p ≤ 0.05) with similar effects in the trans-well system. The trans-well system allowed for isolation of cell types for inflammatory mediator analysis. DHA decreased mRNA expression (p<0.05) of Mcp1 (-7.1 fold) and increased expression of the negative regulator, Mcp1-IP (+1.5 fold). In macrophages, DHA decreased mRNA expression of pro-inflammatory M1 polarization markers (p ≤ 0.05), Nos2 (iNOS; -7 fold), Tnfα (-4.2 fold) and Nfκb (-2.3 fold), while increasing anti-inflammatory Tgfβ1 (+1.7 fold). Interestingly, the PPARγ antagonist co-administered with DHA or EPA in co-culture reduced (p ≤ 0.05) adiponectin cellular protein, without modulating other cytokines (protein or mRNA). Overall, our findings suggest that DHA may lessen the degree of MCP1 and IL-6 secreted from adipocytes, and may reduce the degree of M1 polarization of macrophages recruited to adipose tissue, thereby decreasing the intensity of pro-inflammatory cross-talk between adipocytes and macrophages in obese adipose tissue.

Dataset Information

Novel high-throughput in vitro model for identifying hemodynamic-induced inflammatory mediators of cerebral aneurysm formation.

Publications

Novel high-throughput in vitro model for identifying hemodynamic-induced inflammatory mediators of cerebral aneurysm formation.

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