The isolation and culture of human ovarian microvascular endothelial cells
The isolation and culture of human ovarian microvascular endothelial cells
We have developed a model system for the isolation and culture of human ovarian microvascular endothelial cells (HOMEC).
Follicular aspirates, obtained at oocyte recovery for in vitro fertilisation were filtered to obtain fragments of follicle wall. These were set in Matrigel and cultured to allow the growth of cells through the matrix. Cells formed capillary-like structures in association with the Matrigel and upon emergence onto the uncoated surface of the culture flask, formed the characteristic endothelial cell cobblestone monolayer. Immunocytochemistry demonstrated that the cells possessed all of the endothelial cell specific markers, and also constitutively expressed VCAM-1 which is normally associated with stimulated endothelial cells.
The development of an endothelial/granulosa cell co-culture model showed specific cellular architecture. Granulosa cell clusters were linked together by endothelial capillary-like structures. These studies indicate that there is intercellular communication between the cells of the corpus luteum and that these interactions can be modelled in vitro.
The presence of both VEGF receptors, flt-1 and KDR, and the endothelial nitric oxide synthase was demonstrated by RT-PCR for HOMEC, therefore, all the constituents for important endothelial mechanisms in relation to VEGF action are in place in these cells. Studies showed a dose dependent increase in HOMEC proliferation in response to VEGF. This proliferative effect in HUVEC could be blocked with an anti-flt-1 antibody, indicating that the two VEGF receptors have different functions, and that binding of VEGF to KDR elicits the mitogenic effect of VEGF. Furthermore it was shown that the proliferative effect of VEGF on both HOMEC and HUVEC could be blocked with a nitric oxide synthase inhibitor, demonstrating the importance of nitric oxide production on the mitogenic effect of VEGF.
University of Southampton
Ratcliffe, Kirsty Elizabeth
335352da-60e1-4bb9-80ef-c0a172ee1707
2000
Ratcliffe, Kirsty Elizabeth
335352da-60e1-4bb9-80ef-c0a172ee1707
Ratcliffe, Kirsty Elizabeth
(2000)
The isolation and culture of human ovarian microvascular endothelial cells.
University of Southampton, Doctoral Thesis.
Record type:
Thesis
(Doctoral)
Abstract
We have developed a model system for the isolation and culture of human ovarian microvascular endothelial cells (HOMEC).
Follicular aspirates, obtained at oocyte recovery for in vitro fertilisation were filtered to obtain fragments of follicle wall. These were set in Matrigel and cultured to allow the growth of cells through the matrix. Cells formed capillary-like structures in association with the Matrigel and upon emergence onto the uncoated surface of the culture flask, formed the characteristic endothelial cell cobblestone monolayer. Immunocytochemistry demonstrated that the cells possessed all of the endothelial cell specific markers, and also constitutively expressed VCAM-1 which is normally associated with stimulated endothelial cells.
The development of an endothelial/granulosa cell co-culture model showed specific cellular architecture. Granulosa cell clusters were linked together by endothelial capillary-like structures. These studies indicate that there is intercellular communication between the cells of the corpus luteum and that these interactions can be modelled in vitro.
The presence of both VEGF receptors, flt-1 and KDR, and the endothelial nitric oxide synthase was demonstrated by RT-PCR for HOMEC, therefore, all the constituents for important endothelial mechanisms in relation to VEGF action are in place in these cells. Studies showed a dose dependent increase in HOMEC proliferation in response to VEGF. This proliferative effect in HUVEC could be blocked with an anti-flt-1 antibody, indicating that the two VEGF receptors have different functions, and that binding of VEGF to KDR elicits the mitogenic effect of VEGF. Furthermore it was shown that the proliferative effect of VEGF on both HOMEC and HUVEC could be blocked with a nitric oxide synthase inhibitor, demonstrating the importance of nitric oxide production on the mitogenic effect of VEGF.
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Published date: 2000
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Local EPrints ID: 463991
URI: http://eprints.soton.ac.uk/id/eprint/463991
PURE UUID: 17132470-fb98-4819-9b45-65356fa72ca3
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Date deposited: 04 Jul 2022 21:00
Last modified: 16 Mar 2024 19:06
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Author:
Kirsty Elizabeth Ratcliffe
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