liu.seSearch for publications in DiVA
Change search
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • harvard1
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • oxford
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
The ATP-gated P2X1 receptor plays a pivotal role in activation of aspirin-treated platelets by thrombin and epinephrine
Linköping University, Department of Medicine and Health Sciences, Pharmacology . Linköping University, Faculty of Health Sciences.
Linköping University, Department of Clinical and Experimental Medicine, Clinical Chemistry. Linköping University, Faculty of Health Sciences.
Linköping University, Department of Medicine and Health Sciences, Pharmacology . Linköping University, Faculty of Health Sciences.
Linköping University, Department of Medicine and Health Sciences, Pharmacology . Linköping University, Faculty of Health Sciences.
Show others and affiliations
2008 (English)In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 283, no 27, 18493-18504 p.Article in journal (Refereed) Published
Abstract [en]

Human platelets express protease-activated receptor 1 (PAR1) and PAR4 but limited data indicate for differences in signal transduction. We studied the involvement of PAR1 and PAR4 in the cross-talk between thrombin and epinephrine. The results show that epinephrine acted via alpha(2A)-adrenergic receptors to provoke aggregation, secretion, and Ca(2+) mobilization in aspirin-treated platelets pre-stimulated with subthreshold concentrations of thrombin. Incubating platelets with antibodies against PAR4 or the PAR4-specific inhibitor pepducin P4pal-i1 abolished the aggregation. Furthermore, platelets pre-exposed to the PAR4-activating peptide AYPGKF, but not to the PAR1-activating peptide SFLLRN, were aggregated by epinephrine, whereas both AYPGKF and SFLLRN synergized with epinephrine in the absence of aspirin. The roles of released ATP and ADP were elucidated by using antagonists of the purinergic receptors P2X(1), P2Y(1), and P2Y(12) (i.e. NF449, MRS2159, MRS2179, and cangrelor). Intriguingly, ATP, but not ADP, was required for the epinephrine/thrombin-induced aggregation. In Western blot analysis, a low concentration of AYPGKF, but not SFLLRN, stimulated phosphorylation of Akt on serine 473. Moreover, the phosphatidyl inositide 3-kinase inhibitor LY294002 antagonized the effect of epinephrine combined with thrombin or AYPGKF. Thus, in aspirin-treated platelets, PAR4, but not PAR1, interacts synergistically with alpha(2A)-adrenergic receptors, and the PI3-kinase/Akt pathway is involved in this cross-talk. Furthermore, in PAR4-pretreated platelets, epinephrine caused dense granule secretion, and subsequent signaling from the ATP-gated P2X(1)-receptor and the alpha(2A)-adrenergic receptor induced aggregation. These results suggest a new mechanism that has ATP as a key element and circumvents the action of aspirin on epinephrine-facilitated PAR4-mediated platelet activation.

Place, publisher, year, edition, pages
2008. Vol. 283, no 27, 18493-18504 p.
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:liu:diva-20726DOI: 10.1074/jbc.M800358200PubMedID: 18480058OAI: oai:DiVA.org:liu-20726DiVA: diva2:235795
Available from: 2009-09-18 Created: 2009-09-18 Last updated: 2017-12-13Bibliographically approved
In thesis
1. The thrombin receptors PAR1 and PAR4 and their relative role in platelet activation
Open this publication in new window or tab >>The thrombin receptors PAR1 and PAR4 and their relative role in platelet activation
2009 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

Many blood cell mechanisms in the human body are working all the time to maintain haemostasis in the blood vessels. Once a wound arises platelets are alerted via different substances to cover the wound and prevent loss of blood. Most of the times these mechanisms do stop the blood, and further heal the wound. During other circumstances the platelet-covering continues to form a thrombus, preventing the blood to flow and instead causes myocardial infarction or stroke. There are several risk factors triggering development of circulatory diseases such as obesity, lack of exercise, smoking, infection and stress.

This thesis describes the interaction between the two platelet thrombin receptors PAR1 and PAR4, together with the interaction of the oral pathogen Porphyromonas gingivalis (with thrombin-like gingipains), and the cross talk with the stress hormone epinephrine and its α2A adrenergic receptor. Until now PAR1 is thought to be the most important thrombin receptor due to its high affinity for thrombin. From a phylogenetical and patophysiological point of view there must be a reason why platelets express two different thrombin receptors. Today PAR4 is considered less important, but this thesis implies that PAR4 plays an important role in platelet signaling and haemostasis.

The results show that bacteria pre-stimulated platelets, followed by epinephrine gives a strong and full aggregation and calcium mobilization, in both aspirinated and non-aspirinated human platelets. The amount of bacteria does not itself, or epinephrine alone give aggregation or calcium mobilization. This mechanism is dependent on both Rgp type gingipain released from P. gingivalis, and PARs in an interaction with the α2A adrenergic receptor.

Further, results reveal that PAR4 interacts and cross talks with the platelet α2A-adrenergic receptor in aspirinated platelets. Neither of the two platelet purinergic P2Y-receptors (P2Y12 and P2Y1) contribute to this action, but the purinergic P2X1 does. In aggregation studies a low dose of PAR4 activating peptide (AP), but not PAR1-AP, followed by epinephrine results in a strong aggregation and in a calcium mobilization. ATP secretion measurements did reveal that ATP was released during epinephrine stimulation, which indicate that ATP and P2X1 have a key role in this event. By blocking P2X1 both aggregation and calcium mobilization were abolished, but not by blocking P2Y12 and P2Y1. Inhibition of PI3-kinase, both epinephrine-induced calcium mobilization and aggregation were significant reduced. In non-aspirinated platelets PAR1 synergizes with the α2A adrenergic receptor and P2X1.

In conclusion, this thesis suggests that PAR4 plays an intriguing and important role in platelets with inactived cyclooxygenase 1.  The results described in this thesis contribute to an increased knowledge of the platelet thrombin receptors.

Place, publisher, year, edition, pages
Linköping: Linköping University Electronic Press, 2009. 43 p.
Series
Linköping Studies in Health Sciences. Thesis, ISSN 1100-6013 ; 104
National Category
Pharmacology and Toxicology
Identifiers
urn:nbn:se:liu:diva-19958 (URN)978-91-7393-562-3 (ISBN)
Presentation
2009-09-04, Eken, Hälsouniversitetet, Campus US, Linköpings Universitet, Linköping, 13:00 (Swedish)
Opponent
Supervisors
Available from: 2009-09-18 Created: 2009-08-20 Last updated: 2009-09-18Bibliographically approved
2. The role of platelet thrombin receptors PAR1 and PAR4 in platelet activation
Open this publication in new window or tab >>The role of platelet thrombin receptors PAR1 and PAR4 in platelet activation
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Platelets play a pivotal role in coagulation and haemostasis. Their most prominent task is to seal damaged blood vessels by the formation of a platelet plug at the damaged area. Once the injury is covered, platelets retract the coagulum to close the wound and allow the blood to flow freely in the vessel. Platelets are strongly activated by the essential enzyme thrombin, formed in the coagulation cascade. Activation of the platelet thrombin receptors PAR1 and PAR4 leads to shape change, secretion of granule content, and aggregation, all of which can be accomplished by each receptor individually. However more and more findings indicate that there are differences between the receptors and that they have different physiological functions.

This thesis presents studies performed to elucidate the relative role of PAR1 and PAR4 in platelet activation and coagulation.

We have studied the effects on platelet activation and coagulation, and revealed a possible physiological role for PAR4 in the stabilisation of the coagulum. We also investigated the relative role of PAR1 and PAR4 in the cross-talk between thrombin and epinephrine with and without inhibition of COX-1. We demonstrated that PAR4 interacts with adrenergic receptors and causes an aggregation of platelets dependent on released ATP and its receptor P2X1, thereby circumventing the inhibition by aspirin. Not only is this an interesting specific role for PAR4, but it may also be of clinical importance considering that COX-1 inhibition is the most common treatment for patients with cardiovascular disease to prevent thrombosis.

We show that the number of PAR1 receptors varied between donors and that this variation was correlated to the response on receptor activation. The number of PAR1 receptors on the platelet surface was decreased after PAR1 stimulation but increased after stimulation of other receptors.

In a final attempt to elucidate the nature of PAR1 and PAR4 we used mathematics to evaluate the effect of co-stimulation of the receptors. We found a strong synergistic effect for both platelet activation and aggregation. This indicates that PAR1 and PAR4 interact in a yet unknown way to regulate or amplify the effect of each other rather than merely transmitting the incoming signal the same way.

Place, publisher, year, edition, pages
Linköping: Linköping University Electronic Press, 2009. 70 p.
Series
Linköping University Medical Dissertations, ISSN 0345-0082 ; 1156
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-51935 (URN)978-91-7393-505-0 (ISBN)
Public defence
2009-12-04, Linden, Hälsouniversitetet, Campus US, Linköpings Universitet, Linköping, 13:00 (English)
Opponent
Supervisors
Available from: 2009-11-24 Created: 2009-11-24 Last updated: 2015-03-13Bibliographically approved
3. The role of platelet thrombin receptors PAR1 and PAR4 in health and disease
Open this publication in new window or tab >>The role of platelet thrombin receptors PAR1 and PAR4 in health and disease
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Blood cells are continuously flowing in our systems maintaining haemostasis in the arteries and veins. If a vessel is damaged, the smallest cell fragments in the blood (platelets) are directed to cover the wound and plug the leakage to prevent blood loss. Most of the time platelets stop the blood leak without any difficulties. During other, pathological, circumstances, platelets continue to form a thrombus, preventing the blood flow and may cause myocardial infarction or stroke.

Thrombin is the most potent platelet agonist and is a product created in the coagulation cascade. This thesis is focused on the interactions between the two platelet thrombin receptors; protease activated receptors 1 (PAR1) and PAR4 in vitro. We have investigated potential differences between these receptors in several situations associated with cardiovascular disease.

First we studied interactions between PAR1 and PAR4 and the oral pathogen Porphyromonas gingivalis (which secretes enzymes, gingipains, with properties similar to thrombin). Here we showed that P. gingivalis is signaling mainly, but not exclusively, via PAR4. Our second study showed that the cross-talk between the stress hormone epinephrine and thrombin occur exclusively through PAR4 if the key-substance ATP is present and cyclooxygenase-1 inhibited by aspirin. The third study investigated platelet secretion, with focus on the protein plasminogen activator inhibitor 1(PAI-1), an inhibitor of the fibrinolytic process responsible for dissolving a formed clot. Here we showed that PAI-1 secretion and synthesis was more sensitive to stimulation through PAR1 than PAR4. Finally this thesis describes differences between PAR1 and PAR4 in cell-signaling pathways regulating the stability of a platelet aggregate, where PAR4 seems to be of importance to create stable platelet aggregates and that this stability is dependent on ADP activation via P2Y12 and cell signaling via PI3-kinase.

Until now, PAR1 has been considered to be the most important thrombin receptor, due to its high affinity for thrombin. However, there must be a reason why platelets express two different thrombin receptors. This thesis highlights several situations where PAR4 plays a complementary and important role in platelet signaling and haemostasis.

In conclusion, this thesis suggests that PAR4 plays a major role in calcium signaling and the induction of sustained aggregation, while PAR1 shows a more prominent role in platelet secretion and synthesis. This thesis also reveals new interactions between platelet thrombin receptors and the ADP-, ATP- and epinephrine receptors. The results described in this thesis contribute to an increased knowledge of the platelet thrombin receptors and their interplay in situations such as infection, stress, fibrinolysis, and platelet aggregation.

Place, publisher, year, edition, pages
Linköping: Linköping University Electronic Press, 2011. 63 p.
Series
Linköping University Medical Dissertations, ISSN 0345-0082 ; 1261
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-71395 (URN)978-91-7393-067-3 (ISBN)
Public defence
2011-11-04, Berzeliussalen, Hälsouniversitetet, Campus US, Linköpings universitet, Linköping, 13:00 (English)
Opponent
Supervisors
Available from: 2011-10-14 Created: 2011-10-14 Last updated: 2011-11-28Bibliographically approved

Open Access in DiVA

No full text

Other links

Publisher's full textPubMedLink to Licentiate Thesis (Martina NylanderLink to Ph.D. Thesis (Karin Vretenbrant Öberg)

Authority records BETA

Grenegård, MagnusVretenbrant-Öberg, KarinNylander, MartinaDésilets, StéphanieLindström, Eva GRamström, SofiaLindahl, Tomas L

Search in DiVA

By author/editor
Grenegård, MagnusVretenbrant-Öberg, KarinNylander, MartinaDésilets, StéphanieLindström, Eva GRamström, SofiaLindahl, Tomas L
By organisation
Pharmacology Faculty of Health SciencesClinical ChemistryDepartment of Clinical Chemistry
In the same journal
Journal of Biological Chemistry
Medical and Health Sciences

Search outside of DiVA

GoogleGoogle Scholar

doi
pubmed
urn-nbn

Altmetric score

doi
pubmed
urn-nbn
Total: 243 hits
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • harvard1
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • oxford
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf