Dataset Information

Static spatial growth restriction micropatterning of endothelial colony forming cells influences their morphology and gene expression.

ABSTRACT:

Background

Endothelialization of small diameter synthetic vascular grafts is a potential solution to the thrombosis and intimal hyperplasia that plague current devices. Endothelial colony forming cells, which are blood-derived and similar to mature endothelial cells, are a potential cell source. Anisotropic spatial growth restriction micropatterning has been previously shown to affect the morphology and function of mature endothelial cells in a manner similar to unidirectional fluid shear stress. To date, endothelial colony forming cells have not been successfully micropatterned. This study addresses the hypothesis that micropatterning of endothelial colony forming cells will induce morphological elongation, cytoskeletal alignment, and changes in immunogenic and thrombogenic-related gene expression.

Methods

Spatially growth restrictive test surfaces with 25 ?m-wide lanes alternating between collagen-I and a blocking polymer were created using microfluidics. Case-matched endothelial colony forming cells and control mature carotid endothelial cells were statically cultured on either micropatterned or non-patterned surfaces. Cell elongation was quantified using shape index. Using confocal microscopy, cytoskeletal alignment was visualized and density and apoptotic rate were determined. Gene expression was measured using quantitative PCR to measure KLF-2, eNOS, VCAM-1, and vWF.

Results

Endothelial colony forming cells were successfully micropatterned for up to 50 hours. Micropatterned cells displayed elongation and actin alignment. Micropatterning increased the packing densities of both cell types, but did not affect apoptotic rate, which was lower in endothelial colony forming cells. KLF-2 gene expression was increased in micropatterned relative to non-patterned endothelial colony forming cells after 50 hours. No significant differences were seen in the other genes tested.

Conclusions

Endothelial colony forming cells can be durably micropatterned using spatial growth restriction. Micropatterning has a significant effect on the gross and subcellular morphologies of both cell types. Further study is required to fully understand the effect of micropatterning on endothelial colony forming cell gene expression.

SUBMITTER: Hagen MW

PROVIDER: S-EPMC6561595 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Publications

Static spatial growth restriction micropatterning of endothelial colony forming cells influences their morphology and gene expression.

Hagen Matthew W MW Hinds Monica T MT

PloS one 20190612 6

<h4>Background</h4>Endothelialization of small diameter synthetic vascular grafts is a potential solution to the thrombosis and intimal hyperplasia that plague current devices. Endothelial colony forming cells, which are blood-derived and similar to mature endothelial cells, are a potential cell source. Anisotropic spatial growth restriction micropatterning has been previously shown to affect the morphology and function of mature endothelial cells in a manner similar to unidirectional fluid shea ...[more]

PMID: 31188903

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Project description:Artificial small-diameter vascular grafts remain an unmet need in modern medicine, due to the thrombosis and neointimal hyperplasia that plague currently available synthetic devices. Tissue engineering techniques, including in vitro endothelialization, could offer a solution to this problem. A potential minimally invasive source of patient autologous endothelium is endothelial colony-forming cells (ECFCs), endothelial-like outgrowth products of circulating progenitors. While ECFCs respond to shear stress similar to mature endothelial cells (ECs), their response to luminal topographic micropatterning (TMP), a biomaterial modification with the potential to flow-independently, enhance the attachment, migration, gene expression, and function of mature ECs, remains unstudied. In this study, case-matched carotid endothelial cells (CaECs) and blood-derived ECFCs are statically cultured on polyurethane substrates with micropatterned pitches (pitch = peak to peak distance) ranging from 3-to 14 μm. On all pattern pitches tested, both CaECs and ECFCs showed significant and robust alignment to the angle of the micropatterns. Using a novel cell-by-cell image analysis technique, it was found that actin fibers similarly and significantly aligned to the angle of micropatterned features on all pitches tested. Microtubules analyzed through the same novel approach showed significant alignment on most pitches examined, with a greater variation in fiber angle overall. Interestingly, only CaECs showed significant cellular elongation, and notably to a lower degree than previously seen either in vivo due to flow or in vitro due to spatial growth restriction micropatterning, but consistent with earlier studies of TMP. Neither cell type displayed any significant micropattern-driven changes in the expression of KLF-2 or the downstream adhesion molecules it regulates. These results demonstrate that TMP flow-independently affects ECFC morphology, but that alignment alone is insufficient to drive protective changes in EC and ECFC function.

Project description:BackgroundSuccessful pregnancies are nowadays possible after kidney transplantation but are associated with a higher incidence of maternal and fetal complications. Immunosuppressive therapy causes cardiovascular side effects but must be maintained during pregnancy. Little is known about the consequences of maternal kidney transplantation on offspring's endothelial health. Endothelial colony forming cells (ECFCs) represent a highly proliferative subtype of endothelial progenitor cells and are crucial for vascular homeostasis, repair and neovascularization. Therefore, we investigated whether maternal kidney transplantation affects fetal ECFCs' characteristics.MethodsECFCs were isolated from umbilical cord blood of uncomplicated and post-kidney-transplant pregnancies and analyzed for their functional abilities with proliferation, cell migration, centrosome orientation and angiogenesis assays. Further, ECFCs from uncomplicated pregnancies were exposed to either umbilical cord serum from uncomplicated or post-kidney-transplant pregnancies.ResultsPost-kidney-transplant ECFCs showed significantly less proliferation, less migration and less angiogenesis compared to control ECFCs. The presence of post-kidney-transplant umbilical cord serum led to similar functional aberrations of ECFCs from uncomplicated pregnancies.ConclusionsThese pilot data demonstrate differences in ECFCs' biological characteristics in offspring of women after kidney transplantation. Further studies are needed to monitor offspring's long-term cardiovascular development and to assess possible causal relationships with immunosuppressants, uremia and maternal cardiovascular alterations.ImpactPregnancy after kidney transplantation has become more common in the past years but is associated with higher complications for mother and offspring. Little is known of the impact of maternal kidney transplantation and the mandatory immunosuppressive therapy on offspring vascular development. In this study we are the first to address and detect an impairment of endothelial progenitor cell function in offspring of kidney-transplanted mothers. Serum from post-transplant pregnancies also causes negative effects on ECFCs' function. Clinical studies should focus on long-term monitoring of offspring's cardiovascular health.

Dataset Information

Static spatial growth restriction micropatterning of endothelial colony forming cells influences their morphology and gene expression.

Background

Methods

Results

Conclusions

Publications

Static spatial growth restriction micropatterning of endothelial colony forming cells influences their morphology and gene expression.

Similar Datasets

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