{"id":{"repo_id":"de-montfort","oai_identifier":"oai:dora.dmu.ac.uk:2086/25725"},"canonical_url":"https://search.dev.ndltd.org/etd/de-montfort/oai:dora.dmu.ac.uk:2086/25725","repository":{"repo_id":"de-montfort","name":"De Montfort University","base_url":"https://dora.dmu.ac.uk/server/oai/request"},"display":{"title":"THE ROLE OF P2Y RECEPTORS IN HUMAN VASCULAR SMOOTH MUSCLE","abstract":"Excessive or inappropriate vascular smooth muscle (VSM) cell proliferation is a major contributor to proliferative disorders such as atherosclerosis and restenosis following angioplasty. Treatment is by bypass grafting using the internal mammary artery (IMA) or saphenous vein (SV). IMA conduits show greater resistance to restenosis and hence increased patient survival rates than SV, although the mechanism is unknown. At the site of vascular injury, nucleotides in addition to PDGF are released from degranulating platelets, VSM cells and endothelial cells. Proliferative studies in animals show that extracellular nucleotides act as growth factors for VSM cells. PDGF is also implicated in proliferation of VSM cells in vivo and in vitro. However, the role of nucleotides in restenosis and atherosclerosis is still poorly understood, but is clearly of great clinical importance and may lead to improved pharmacological intervention. PDGF-BB increased pHj-thymidine incorporation in SV cells when compared to IMA cells. The response was augmented by ATP in SV cells only. Surprisingly, UTP and UDP reduced the PDGF mediated response in both IMA and SV cells using pH]-thymidine incorporation, a colorimetric proliferation assay and cell counting as indexes of mitogenesis. 2MeSADP and AR-C67085MX were ineffective, ruling out the involvement of P2Yi, P2Yi2, P2Yi3 and P2Yii receptors respectively. This suggests that the proproliferative action of ATP is at P2Y2 receptors, whilst the anti-proliferative action is at P2Y4 (UTP) and P2Y6 (UDP) receptors. In SV cells, the marked downregulation of the cell cycle inhibitor p27'<'pi by PDGF-BB was enhanced by ATP whilst the constitutively expressed p21°'’T cyclin DI and cyclin E protein levels were unchanged. UTP was without effect. Thus, ATP may exert its proliferative effect at the level of the cell cycle by relieving the inhibitory influences of p27kp1. ATP and UTP equipotently elevated intracellular calcium in SV cells in the absence of a detectable InsP3 response in a concentration-dependent manner due to smoothly graded increases in amplitude and recruitment of more cells. The rank order of potency was very similar to that derived for 1321N1 cells expressing the hP2Y2 receptor and is consistent with activation of P2Y2 receptors, although a contribution from P2Y4 receptors cannot be be ruled out. The response to UDP varied considerably between cultures and may reflect differences in P2Y6-receptor expression between patients. 2MeSADP and AR-C67085MX were inactive, confirming the absence of P2Yi and P2Yii receptors. Unexpectedly, the ATP-responsive hP2Yii receptor transfected into 1321N1 cells responded to UTP to release Ca\"'' in the absence of a PLC response. Pertussis toxin pre-treatment and 2-APB differentially affected the responses of ATP and UTP. ATP responses were more resistant to desensitisation than UTP, whilst application of the P2Yii—specific agonist, AR-C69931MX virtually abolished the subsequent response to UTP. These results suggest that ATP and UTP act at the same receptor to recruit distinct signalling pathways. In conclusion, the decreased patency of SV grafts compared to IMA may be explained by both the increased proliferative response to PDGF-BB and the pro-proliferative action of ATP in synthetic SV VSM cells. Both are stored in platelets and released simultaneously at the site of vascular damage or disease. ATP acting at P2Y2 receptors may function as a progression factor in the proliferative response to facilitate the progression of competent cells to synthesise DNA. Importantly UTP and UDP, acting at P2Y4 and P2Y6 receptors respectively, may function as negative controllers of VSM cell proliferation in human artery and vein.","abstract_html":"Excessive or inappropriate vascular smooth muscle (VSM) cell proliferation is a major contributor to proliferative disorders such as atherosclerosis and restenosis following angioplasty. Treatment is by bypass grafting using the internal mammary artery (IMA) or saphenous vein (SV). IMA conduits show greater resistance to restenosis and hence increased patient survival rates than SV, although the mechanism is unknown. At the site of vascular injury, nucleotides in addition to PDGF are released from degranulating platelets, VSM cells and endothelial cells. Proliferative studies in animals show that extracellular nucleotides act as growth factors for VSM cells. PDGF is also implicated in proliferation of VSM cells in vivo and in vitro. However, the role of nucleotides in restenosis and atherosclerosis is still poorly understood, but is clearly of great clinical importance and may lead to improved pharmacological intervention. PDGF-BB increased pHj-thymidine incorporation in SV cells when compared to IMA cells. The response was augmented by ATP in SV cells only. Surprisingly, UTP and UDP reduced the PDGF mediated response in both IMA and SV cells using pH]-thymidine incorporation, a colorimetric proliferation assay and cell counting as indexes of mitogenesis. 2MeSADP and AR-C67085MX were ineffective, ruling out the involvement of P2Yi, P2Yi2, P2Yi3 and P2Yii receptors respectively. This suggests that the proproliferative action of ATP is at P2Y2 receptors, whilst the anti-proliferative action is at P2Y4 (UTP) and P2Y6 (UDP) receptors. In SV cells, the marked downregulation of the cell cycle inhibitor p27&#x27;&lt;&#x27;pi by PDGF-BB was enhanced by ATP whilst the constitutively expressed p21°&#x27;’T cyclin DI and cyclin E protein levels were unchanged. UTP was without effect. Thus, ATP may exert its proliferative effect at the level of the cell cycle by relieving the inhibitory influences of p27kp1. ATP and UTP equipotently elevated intracellular calcium in SV cells in the absence of a detectable InsP3 response in a concentration-dependent manner due to smoothly graded increases in amplitude and recruitment of more cells. The rank order of potency was very similar to that derived for 1321N1 cells expressing the hP2Y2 receptor and is consistent with activation of P2Y2 receptors, although a contribution from P2Y4 receptors cannot be be ruled out. The response to UDP varied considerably between cultures and may reflect differences in P2Y6-receptor expression between patients. 2MeSADP and AR-C67085MX were inactive, confirming the absence of P2Yi and P2Yii receptors. Unexpectedly, the ATP-responsive hP2Yii receptor transfected into 1321N1 cells responded to UTP to release Ca&quot;&#x27;&#x27; in the absence of a PLC response. Pertussis toxin pre-treatment and 2-APB differentially affected the responses of ATP and UTP. ATP responses were more resistant to desensitisation than UTP, whilst application of the P2Yii—specific agonist, AR-C69931MX virtually abolished the subsequent response to UTP. These results suggest that ATP and UTP act at the same receptor to recruit distinct signalling pathways. In conclusion, the decreased patency of SV grafts compared to IMA may be explained by both the increased proliferative response to PDGF-BB and the pro-proliferative action of ATP in synthetic SV VSM cells. Both are stored in platelets and released simultaneously at the site of vascular damage or disease. ATP acting at P2Y2 receptors may function as a progression factor in the proliferative response to facilitate the progression of competent cells to synthesise DNA. Importantly UTP and UDP, acting at P2Y4 and P2Y6 receptors respectively, may function as negative controllers of VSM cell proliferation in human artery and vein.","abstract_has_math":false,"creators":["White, Pamela J."],"institution":"De Montfort University","degree_name":"PhD","degree_level":"Doctoral","degree_discipline":null,"degree_department":null,"school":null,"contributors":[],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2005,"date_issued":"2005-01","date_published":"2005-01","updated_at":"2026-07-24T06:18:44Z","subjects":[],"languages":[],"rights":[],"rights_urls":["https://dora.dmu.ac.uk/bitstreams/74196f6c-67ab-41a2-8f47-7a16aab5fff5/download"],"identifier_entries":[]},"links":{"outbound_url":null,"outbound_label":null,"outbound_source":null},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:creator","label":"Author","values":["White, Pamela J."]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.issued","label":"Date","values":["2005-01"]},{"key":"dc:publisher.department","label":"Dc Publisher Department","values":["Faculty of Health and Life Sciences"]},{"key":"dc:publisher.institution","label":"Dc Publisher Institution","values":["De Montfort University"]},{"key":"dc:relation.isreferencedby","label":"Dc Relation Isreferencedby","values":["https://hdl.handle.net/2086/25725"]},{"key":"dc:type","label":"Dc Type","values":["Thesis or dissertation"]},{"key":"dc:type.qualificationlevel","label":"Dc Type Qualificationlevel","values":["Doctoral"]},{"key":"dc:type.qualificationname","label":"Dc Type Qualificationname","values":["PhD"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:rights","label":"Dc Rights","values":["https://dora.dmu.ac.uk/bitstreams/74196f6c-67ab-41a2-8f47-7a16aab5fff5/download"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.uri","label":"Identifier URI","values":["https://dora.dmu.ac.uk/bitstreams/21241aa4-4fed-4b80-ba43-03c042167384/download"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["Excessive or inappropriate vascular smooth muscle (VSM) cell proliferation is a major contributor to proliferative disorders such as atherosclerosis and restenosis following angioplasty. Treatment is by bypass grafting using the internal mammary artery (IMA) or saphenous vein (SV). IMA conduits show greater resistance to restenosis and hence increased patient survival rates than SV, although the mechanism is unknown. At the site of vascular injury, nucleotides in addition to PDGF are released from degranulating platelets, VSM cells and endothelial cells. Proliferative studies in animals show that extracellular nucleotides act as growth factors for VSM cells. PDGF is also implicated in proliferation of VSM cells in vivo and in vitro. However, the role of nucleotides in restenosis and atherosclerosis is still poorly understood, but is clearly of great clinical importance and may lead to improved pharmacological intervention. PDGF-BB increased pHj-thymidine incorporation in SV cells when compared to IMA cells. The response was augmented by ATP in SV cells only. Surprisingly, UTP and UDP reduced the PDGF mediated response in both IMA and SV cells using pH]-thymidine incorporation, a colorimetric proliferation assay and cell counting as indexes of mitogenesis. 2MeSADP and AR-C67085MX were ineffective, ruling out the involvement of P2Yi, P2Yi2, P2Yi3 and P2Yii receptors respectively. This suggests that the proproliferative action of ATP is at P2Y2 receptors, whilst the anti-proliferative action is at P2Y4 (UTP) and P2Y6 (UDP) receptors. In SV cells, the marked downregulation of the cell cycle inhibitor p27'<'pi by PDGF-BB was enhanced by ATP whilst the constitutively expressed p21°'’T cyclin DI and cyclin E protein levels were unchanged. UTP was without effect. Thus, ATP may exert its proliferative effect at the level of the cell cycle by relieving the inhibitory influences of p27kp1. ATP and UTP equipotently elevated intracellular calcium in SV cells in the absence of a detectable InsP3 response in a concentration-dependent manner due to smoothly graded increases in amplitude and recruitment of more cells. The rank order of potency was very similar to that derived for 1321N1 cells expressing the hP2Y2 receptor and is consistent with activation of P2Y2 receptors, although a contribution from P2Y4 receptors cannot be be ruled out. The response to UDP varied considerably between cultures and may reflect differences in P2Y6-receptor expression between patients. 2MeSADP and AR-C67085MX were inactive, confirming the absence of P2Yi and P2Yii receptors. Unexpectedly, the ATP-responsive hP2Yii receptor transfected into 1321N1 cells responded to UTP to release Ca\"'' in the absence of a PLC response. Pertussis toxin pre-treatment and 2-APB differentially affected the responses of ATP and UTP. ATP responses were more resistant to desensitisation than UTP, whilst application of the P2Yii—specific agonist, AR-C69931MX virtually abolished the subsequent response to UTP. These results suggest that ATP and UTP act at the same receptor to recruit distinct signalling pathways. In conclusion, the decreased patency of SV grafts compared to IMA may be explained by both the increased proliferative response to PDGF-BB and the pro-proliferative action of ATP in synthetic SV VSM cells. Both are stored in platelets and released simultaneously at the site of vascular damage or disease. ATP acting at P2Y2 receptors may function as a progression factor in the proliferative response to facilitate the progression of competent cells to synthesise DNA. 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At the site of vascular injury, nucleotides in addition to PDGF are released from degranulating platelets, VSM cells and endothelial cells. Proliferative studies in animals show that extracellular nucleotides act as growth factors for VSM cells. PDGF is also implicated in proliferation of VSM cells in vivo and in vitro. However, the role of nucleotides in restenosis and atherosclerosis is still poorly understood, but is clearly of great clinical importance and may lead to improved pharmacological intervention. PDGF-BB increased pHj-thymidine incorporation in SV cells when compared to IMA cells. The response was augmented by ATP in SV cells only. Surprisingly, UTP and UDP reduced the PDGF mediated response in both IMA and SV cells using pH]-thymidine incorporation, a colorimetric proliferation assay and cell counting as indexes of mitogenesis. 2MeSADP and AR-C67085MX were ineffective, ruling out the involvement of P2Yi, P2Yi2, P2Yi3 and P2Yii receptors respectively. This suggests that the proproliferative action of ATP is at P2Y2 receptors, whilst the anti-proliferative action is at P2Y4 (UTP) and P2Y6 (UDP) receptors. In SV cells, the marked downregulation of the cell cycle inhibitor p27'<'pi by PDGF-BB was enhanced by ATP whilst the constitutively expressed p21°'’T cyclin DI and cyclin E protein levels were unchanged. UTP was without effect. Thus, ATP may exert its proliferative effect at the level of the cell cycle by relieving the inhibitory influences of p27kp1. ATP and UTP equipotently elevated intracellular calcium in SV cells in the absence of a detectable InsP3 response in a concentration-dependent manner due to smoothly graded increases in amplitude and recruitment of more cells. The rank order of potency was very similar to that derived for 1321N1 cells expressing the hP2Y2 receptor and is consistent with activation of P2Y2 receptors, although a contribution from P2Y4 receptors cannot be be ruled out. The response to UDP varied considerably between cultures and may reflect differences in P2Y6-receptor expression between patients. 2MeSADP and AR-C67085MX were inactive, confirming the absence of P2Yi and P2Yii receptors. Unexpectedly, the ATP-responsive hP2Yii receptor transfected into 1321N1 cells responded to UTP to release Ca\"'' in the absence of a PLC response. Pertussis toxin pre-treatment and 2-APB differentially affected the responses of ATP and UTP. ATP responses were more resistant to desensitisation than UTP, whilst application of the P2Yii—specific agonist, AR-C69931MX virtually abolished the subsequent response to UTP. These results suggest that ATP and UTP act at the same receptor to recruit distinct signalling pathways. In conclusion, the decreased patency of SV grafts compared to IMA may be explained by both the increased proliferative response to PDGF-BB and the pro-proliferative action of ATP in synthetic SV VSM cells. Both are stored in platelets and released simultaneously at the site of vascular damage or disease. ATP acting at P2Y2 receptors may function as a progression factor in the proliferative response to facilitate the progression of competent cells to synthesise DNA. 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