Dataset Information

Influence of TGF-?1 expression in endothelial cells on smooth muscle cell phenotypes and MMP production under shear stress in a co-culture model.

ABSTRACT: Recently, our group has contrasted an endothelial cell-smooth muscle cell (EC-SMC) co-culture model with 3D-cultured SMCs and found that SMCs could respond to high shear stress (SS), which has not been explored before. SMCs were not directly exposed to the flow but were under an EC monolayer; therefore, it is necessary to explore the influence of EC on SMC behaviors under high SS for understanding the mechanism of SMC response to various magnitudes of SS. In the present study, TGF-?1 expression in ECs in an EC-SMC co-culture model was suppressed by an siRNA transfection method. Next, phenotypic changes were observed and MMP-2 and -9 productions were measured in SMCs in the co-culture model after 72-h flow exposure to different SS levels. We confirmed that TGF-?1 expression in ECs could influence SMC phenotypic change under SS conditions and that TGF-?1 expression in ECs could also change MMP-2 production but not MMP-9 production in SMCs under SS conditions in the co-culture model. These results could be useful for understanding the mechanisms of SMC response to SS, particularly for understanding signal transduction emanating from ECs.

SUBMITTER: Han X

PROVIDER: S-EPMC6465372 | biostudies-literature | 2019 Apr

REPOSITORIES: biostudies-literature

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Influence of TGF-β1 expression in endothelial cells on smooth muscle cell phenotypes and MMP production under shear stress in a co-culture model.

Han Xiaobo X Sakamoto Naoya N Tomita Noriko N Meng Hui H Sato Masaaki M Ohta Makoto M

Cytotechnology 20190201 2

Recently, our group has contrasted an endothelial cell-smooth muscle cell (EC-SMC) co-culture model with 3D-cultured SMCs and found that SMCs could respond to high shear stress (SS), which has not been explored before. SMCs were not directly exposed to the flow but were under an EC monolayer; therefore, it is necessary to explore the influence of EC on SMC behaviors under high SS for understanding the mechanism of SMC response to various magnitudes of SS. In the present study, TGF-β1 expression ...[more]

PMID: 30707337

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Project description:Collaterals are unique blood vessels present in the microcirculation of most tissues that, by cross-connecting a small fraction of the outer branches of adjacent arterial trees, provide alternate routes of perfusion. However, collaterals are especially susceptible to rarefaction caused by aging, other vascular risk factors, and mouse models of Alzheimer's disease-a vulnerability attributed to the disturbed hemodynamic environment in the watershed regions where they reside. We examined the hypothesis that endothelial and smooth muscle cells (ECs and SMCs, respectively) of collaterals have specializations, distinct from those of similarly-sized nearby distal-most arterioles (DMAs) that maintain collateral integrity despite their continuous exposure to low and oscillatory/disturbed shear stress, high wall stress, and low blood oxygen. Examination of mouse brain revealed the following: Unlike the pro-inflammatory cobble-stoned morphology of ECs exposed to low/oscillatory shear stress elsewhere in the vasculature, collateral ECs are aligned with the vessel axis. Primary cilia, which sense shear stress, are present, unexpectedly, on ECs of collaterals and DMAs but are less abundant on collaterals. Unlike DMAs, collaterals are continuously invested with SMCs, have increased expression of Pycard, Ki67, Pdgfb, Angpt2, Dll4, Ephrinb2, and eNOS, and maintain expression of Klf2/4. Collaterals lack tortuosity when first formed during development, but tortuosity becomes evident within days after birth, progresses through middle age, and then declines-results consistent with the concept that collateral wall cells have a higher turnover rate than DMAs that favors proliferative senescence and collateral rarefaction. In conclusion, endothelial and SMCs of collaterals have morphologic and functional differences from those of nearby similarly sized arterioles. Future studies are required to determine if they represent specializations that counterbalance the disturbed hemodynamic, pro-inflammatory, and pro-proliferative environment in which collaterals reside and thus mitigate their risk factor-induced rarefaction.

Project description:BACKGROUND:Early neointimal hyperplasia of vein graft may be ameliorated via enhancing intravenous surface shear stress. Cellular processes including proliferation, apoptosis and migration of endothelial cells (ECs) and vascular smooth muscle cells (VSMCs) may play very important roles in the process of neointimal hyperplasia of vein graft; and mitogen-activated protein kinase (MAPK) pathways including extracellular signal-regulated kinase (ERK1/2) and p38 pathways play vital roles in regulating a large variety of cellular processes. This study evaluated the impacts of shear stress and MAPK pathways on cellular processes of ECs in a co-culture system with VSMCs, and aimed to test the hypothesis that high shear stress suppresses proliferation and migration but promotes apoptosis of ECs co-cultured with VSMCs via down-regulating MAPK pathway. METHODS:Primary ECs and VSMCs derived from porcine great saphenous vein were collected, respectively. 4-7 generation of cells were used as work cells. ECs and VSMCs were co-cultured and synchronized under high and low shear stress using Parallel-Plate Flow Chamber system. And then, ECs co-cultured with VSMCs were incubated with U0126 (ERK1/2 inhibitor) or PD98059 (p38 inhibitor) under different shear stress. Proliferation, apoptosis and migration of ECs in a co-culture system with VSMCs were detected by 4,5-dimethyl-2-thiazolyl (MTT) assay and bromodeoxyuridine (BrdU) assay, fluorescent-activated cell sorting (FACS) technique, and Transwell assay separately. Each test repeated 3 times. Additionally, protein expressions of ERK1/2 and p38 MAPK were detected by using Western blot, respectively. RESULTS:Under higher level of shear stress condition, proliferation and migration of ECs co-cultured with VSMCs were suppressed, while cell apoptosis was promoted. And blocking ERK1/2 pathway by U0126 or blocking p38 pathway by PD98059, proliferation and migration of ECs co-cultured with VSMCs were further suppressed, while cell apoptosis was further promoted. Additionally, protein expressions of phosphorylation of ERK1/2 and p38MAPK were decreased under higher level of shear stress condition, and were further reduced by blocking ERK1/2 or p38 pathway under shear stress condition. CONCLUSIONS:High shear stress may suppress proliferation and apoptosis of ECs in a co-culture system with VSMCs but promote cell migration via down-regulating ERK1/2 and p38 MAPK pathways.

Dataset Information

Influence of TGF-?1 expression in endothelial cells on smooth muscle cell phenotypes and MMP production under shear stress in a co-culture model.

Publications

Influence of TGF-β1 expression in endothelial cells on smooth muscle cell phenotypes and MMP production under shear stress in a co-culture model.

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