Vascular prothesis material Vascular prothesis material modification to enhance modification to enhance endothelial cell adhesion endothelial cell adhesion T. Markkula, T. Markkula, F. Pu, R.L. Williams, J.A. Hunt F. Pu, R.L. Williams, J.A. Hunt Department of Clinical Engineering Department of Clinical Engineering University of Liverpool University of Liverpool
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Vascular prothesis material modification to enhance endothelial cell adhesion T. Markkula, F. Pu, R.L. Williams, J.A. Hunt Department of Clinical Engineering.
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Vascular prothesis material Vascular prothesis material modification to enhance modification to enhance endothelial cell adhesionendothelial cell adhesion
Vascular prothesis material Vascular prothesis material modification to enhance modification to enhance endothelial cell adhesionendothelial cell adhesion
T. Markkula,T. Markkula, F. Pu, R.L. Williams, J.A. Hunt F. Pu, R.L. Williams, J.A. HuntT. Markkula,T. Markkula, F. Pu, R.L. Williams, J.A. Hunt F. Pu, R.L. Williams, J.A. Hunt
Department of Clinical EngineeringDepartment of Clinical EngineeringUniversity of LiverpoolUniversity of Liverpool
L929 fibroblasts on PET
PET surface cleaned ultrasonically for 30 min with 70 % Ethanol and for 30 min with water prior to cell culturing
Replacement of blood vessels Replacement of blood vessels with artificial implantswith artificial implants
PTFE and PET most commonly PTFE and PET most commonly usedused
>6 mm >6 mm ØØ prosthesis OK prosthesis OKSmaller grafts thrombosisSmaller grafts thrombosis
Vascular graftsVascular grafts
ImprovementsImprovements
Find a new material
Modify existing materials
Engineer new tissue
• Endothelial cell lining of inner surface of prosthesis
Mouse antihuman monoclonal antibodies conjugated with FITC, RPE and CyC were used to target CD31, CD54, CD51/61, CD106, CD62E, CD62P and CD62L.The isotope IgG1-k was used for negative control
Immunohistochemistry
ABC immunostaining protocol was used to visualise the quantified expression.
Flow cytometry (FACS)
Mouse antihuman monoclonal antibodies conjugated with FITC, RPE and CyC were used to target CD31, CD54, CD51/61, CD106, CD62E, CD62P and CD62L.The isotope IgG1-k was used for negative control
Immunohistochemistry
ABC immunostaining protocol was used to visualise the quantified expression.
0
20
40
60
80
100
120
CD 54 CD 106 CD 62EAdhesion Molecules
% P
osi
tives
Control
TNF-α
T-PETN-PET
T-PTFE
N-PTFE
P1D1
G G
G
P
P
P P
P
D
P P
P P P
0
20
40
60
80
100
120
CD 54 CD 106 CD 62EAdhesion Molecules
% P
osi
tives
Control
TNF-α
T-PETN-PET
T-PTFE
N-PTFE
P1D7
G
G
G
P P
P P P
D
P
G
P
G
Expression of adhesion molecules Expression of adhesion molecules of endothelial cells on PET and of endothelial cells on PET and PTFEPTFE
NH3-plasma treated PET Untreated PET
CD54 - ICAM, P1, D1
Immunohistochemical staining
NH3-plasma treated PTFE Untreated PTFE
CD54 - ICAM, P1, D1
Immunohistochemical staining
Cell adhesion and Cell adhesion and proliferationproliferation
0
2
4
6
8
10
0 1 7Time (days)
No.
Cel
ls x
1000
0 / s
q.cm
Control
TNF-a
T-PET
N-PET
T-PTFE
N-PTFE
Passage 1
G1 G1
G1
G1
G2
P
•Plasma treatment of PET and PTFE with ammonia appeares to be a powerful method to enhance cell attachment
•The modification of PET and PTFE slightly alter the profile of adhesion molecules expressed but not significantly
•Plasma treatment of PET and PTFE with ammonia appeares to be a powerful method to enhance cell attachment
•The modification of PET and PTFE slightly alter the profile of adhesion molecules expressed but not significantly
Cell growth conclusionsCell growth conclusions
What will be done...What will be done...
• Surface chemistry of samples will be determined using CHEMICAL DERIVATIZATION with XPS...
• The whole range of treatments will be tested with endothelial cell / macrophage co-cultures
What wasn’t presented here...What wasn’t presented here...
• Plasma treatment alters the attachment of macrophages to endothelial cells...
• Macrophage numbers, attachment site and endothelial cell adhesion molecule expressions are altered