Microwell arrays for monitoring phenotypic heterogeneity in vascular cell populations

Research output: Contribution to conferenceAbstract

Abstract

Significant remodeling of the vascular wall underlies cardiovascular disease resulting in the formation of atherosclerotic plaques populated with macrophage and smooth muscle cells (SMCs). These SMCs are thought to arise from the vessel wall, as mature SMCs de-differentiate from a contractile to a migratory, proliferate phenotype. However, the remodeling process is not fully understood and uncertainties remain over plaque cell origins and the plasticity of cells within the vascular wall. Both drug development and regenerative medicine have been restricted by these uncertainties. Recently, through a combination of time-lapse, high-speed fluorescence and 3D reconstruction microscopy, we demonstrated unambiguously [1] that freshly isolated mature, contractile SMCs can rapidly transform into not only a migratory but a phagocytic phenotype, a characteristic behaviour of macrophage. Results also showed strong heterogeneity in the proliferative capacity of SMCs [2] and the presence of other highly proliferative cell types in vascular wall that readily interact with SMCs. To better understand vascular call fate, including characterizing the phenotype of cell subpopulations, we employed SU-8 microfabrication to create a series of addressable microwell arrays that enable screening at the single cell level of large numbers of freshly isolated vascular cells. By incorporating microwells of different areas (from 60x60 to 180x180) and seeding with a cell suspension of appropriate density (either a pure SMC population or a mixed vascular population), cells sedimented stochastically across the microwell arrays such that many wells contained single cells. These cells were characterized by imaging in situ prior to tracking them for >1 week as they were induced to de-differentiate in culture. To validate this approach, variation in the proliferation of individual cells was tracked and the expression of SMC markers (e.g. SMA) following phenotypic modulation quantified. This microwell array approach, which is amenable to drug screening applications, will enable detailed characterization of phenotypic changes in vascular cell sub-populations, providing new insights to inform tissue engineering applications.
LanguageEnglish
Number of pages1
Publication statusPublished - 24 Sep 2018
EventNanoBio2018 Conference - Heraklion, Greece
Duration: 24 Sep 201828 Sep 2018

Conference

ConferenceNanoBio2018 Conference
CountryGreece
CityHeraklion
Period24/09/1828/09/18

Fingerprint

Blood Vessels
Muscle
Cells
Smooth Muscle Myocytes
Monitoring
Population
Macrophages
Screening
Phenotype
Uncertainty
Microfabrication
Microtechnology
Tissue engineering
Preclinical Drug Evaluations
Regenerative Medicine
Plasticity
Atherosclerotic Plaques
Tissue Engineering
Microscopic examination
Fluorescence

Keywords

  • cardiovascular disease
  • vascular cell population
  • smooth muscle cells
  • vascular wall remodelling

Cite this

@conference{197fe818c4204af1bb092a01e90b7160,
title = "Microwell arrays for monitoring phenotypic heterogeneity in vascular cell populations",
abstract = "Significant remodeling of the vascular wall underlies cardiovascular disease resulting in the formation of atherosclerotic plaques populated with macrophage and smooth muscle cells (SMCs). These SMCs are thought to arise from the vessel wall, as mature SMCs de-differentiate from a contractile to a migratory, proliferate phenotype. However, the remodeling process is not fully understood and uncertainties remain over plaque cell origins and the plasticity of cells within the vascular wall. Both drug development and regenerative medicine have been restricted by these uncertainties. Recently, through a combination of time-lapse, high-speed fluorescence and 3D reconstruction microscopy, we demonstrated unambiguously [1] that freshly isolated mature, contractile SMCs can rapidly transform into not only a migratory but a phagocytic phenotype, a characteristic behaviour of macrophage. Results also showed strong heterogeneity in the proliferative capacity of SMCs [2] and the presence of other highly proliferative cell types in vascular wall that readily interact with SMCs. To better understand vascular call fate, including characterizing the phenotype of cell subpopulations, we employed SU-8 microfabrication to create a series of addressable microwell arrays that enable screening at the single cell level of large numbers of freshly isolated vascular cells. By incorporating microwells of different areas (from 60x60 to 180x180) and seeding with a cell suspension of appropriate density (either a pure SMC population or a mixed vascular population), cells sedimented stochastically across the microwell arrays such that many wells contained single cells. These cells were characterized by imaging in situ prior to tracking them for >1 week as they were induced to de-differentiate in culture. To validate this approach, variation in the proliferation of individual cells was tracked and the expression of SMC markers (e.g. SMA) following phenotypic modulation quantified. This microwell array approach, which is amenable to drug screening applications, will enable detailed characterization of phenotypic changes in vascular cell sub-populations, providing new insights to inform tissue engineering applications.",
keywords = "cardiovascular disease, vascular cell population, smooth muscle cells, vascular wall remodelling",
author = "Michele Zagnoni and Sandison, {Mairi E.}",
year = "2018",
month = "9",
day = "24",
language = "English",
note = "NanoBio2018 Conference ; Conference date: 24-09-2018 Through 28-09-2018",

}

Zagnoni, M & Sandison, ME 2018, 'Microwell arrays for monitoring phenotypic heterogeneity in vascular cell populations' NanoBio2018 Conference, Heraklion, Greece, 24/09/18 - 28/09/18, .

Microwell arrays for monitoring phenotypic heterogeneity in vascular cell populations. / Zagnoni, Michele; Sandison, Mairi E.

2018. Abstract from NanoBio2018 Conference, Heraklion, Greece.

Research output: Contribution to conferenceAbstract

TY - CONF

T1 - Microwell arrays for monitoring phenotypic heterogeneity in vascular cell populations

AU - Zagnoni, Michele

AU - Sandison, Mairi E.

PY - 2018/9/24

Y1 - 2018/9/24

N2 - Significant remodeling of the vascular wall underlies cardiovascular disease resulting in the formation of atherosclerotic plaques populated with macrophage and smooth muscle cells (SMCs). These SMCs are thought to arise from the vessel wall, as mature SMCs de-differentiate from a contractile to a migratory, proliferate phenotype. However, the remodeling process is not fully understood and uncertainties remain over plaque cell origins and the plasticity of cells within the vascular wall. Both drug development and regenerative medicine have been restricted by these uncertainties. Recently, through a combination of time-lapse, high-speed fluorescence and 3D reconstruction microscopy, we demonstrated unambiguously [1] that freshly isolated mature, contractile SMCs can rapidly transform into not only a migratory but a phagocytic phenotype, a characteristic behaviour of macrophage. Results also showed strong heterogeneity in the proliferative capacity of SMCs [2] and the presence of other highly proliferative cell types in vascular wall that readily interact with SMCs. To better understand vascular call fate, including characterizing the phenotype of cell subpopulations, we employed SU-8 microfabrication to create a series of addressable microwell arrays that enable screening at the single cell level of large numbers of freshly isolated vascular cells. By incorporating microwells of different areas (from 60x60 to 180x180) and seeding with a cell suspension of appropriate density (either a pure SMC population or a mixed vascular population), cells sedimented stochastically across the microwell arrays such that many wells contained single cells. These cells were characterized by imaging in situ prior to tracking them for >1 week as they were induced to de-differentiate in culture. To validate this approach, variation in the proliferation of individual cells was tracked and the expression of SMC markers (e.g. SMA) following phenotypic modulation quantified. This microwell array approach, which is amenable to drug screening applications, will enable detailed characterization of phenotypic changes in vascular cell sub-populations, providing new insights to inform tissue engineering applications.

AB - Significant remodeling of the vascular wall underlies cardiovascular disease resulting in the formation of atherosclerotic plaques populated with macrophage and smooth muscle cells (SMCs). These SMCs are thought to arise from the vessel wall, as mature SMCs de-differentiate from a contractile to a migratory, proliferate phenotype. However, the remodeling process is not fully understood and uncertainties remain over plaque cell origins and the plasticity of cells within the vascular wall. Both drug development and regenerative medicine have been restricted by these uncertainties. Recently, through a combination of time-lapse, high-speed fluorescence and 3D reconstruction microscopy, we demonstrated unambiguously [1] that freshly isolated mature, contractile SMCs can rapidly transform into not only a migratory but a phagocytic phenotype, a characteristic behaviour of macrophage. Results also showed strong heterogeneity in the proliferative capacity of SMCs [2] and the presence of other highly proliferative cell types in vascular wall that readily interact with SMCs. To better understand vascular call fate, including characterizing the phenotype of cell subpopulations, we employed SU-8 microfabrication to create a series of addressable microwell arrays that enable screening at the single cell level of large numbers of freshly isolated vascular cells. By incorporating microwells of different areas (from 60x60 to 180x180) and seeding with a cell suspension of appropriate density (either a pure SMC population or a mixed vascular population), cells sedimented stochastically across the microwell arrays such that many wells contained single cells. These cells were characterized by imaging in situ prior to tracking them for >1 week as they were induced to de-differentiate in culture. To validate this approach, variation in the proliferation of individual cells was tracked and the expression of SMC markers (e.g. SMA) following phenotypic modulation quantified. This microwell array approach, which is amenable to drug screening applications, will enable detailed characterization of phenotypic changes in vascular cell sub-populations, providing new insights to inform tissue engineering applications.

KW - cardiovascular disease

KW - vascular cell population

KW - smooth muscle cells

KW - vascular wall remodelling

UR - https://etp-nanomedicine.eu/event/nanobio-2018/

M3 - Abstract

ER -