The effect of oxLDL on microvesicle release from platelets, measured by a sensitive flow cytometry method.
Microvesicles (MVs) are submicron vesicles with sizes of 0.1-1.0-µm in diameter, released from various cell types upon activation or apoptosis. Their involvement in a variety of diseases has been intensively investigated. In blood, platelets are potent MV secretors, and oxLDL, a platelet ligand, ind...
| Main Authors: | , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Frontiers Media S.A.
2015-11-01
|
| Series: | Frontiers in Cardiovascular Medicine |
| Subjects: | |
| Online Access: | http://journal.frontiersin.org/Journal/10.3389/fcvm.2015.00037/full |
| _version_ | 1818907293546184704 |
|---|---|
| author | Tine Bo Nielsen Morten Hjuler Nielsen Aase eHandberg Aase eHandberg |
| author_facet | Tine Bo Nielsen Morten Hjuler Nielsen Aase eHandberg Aase eHandberg |
| author_sort | Tine Bo Nielsen |
| collection | DOAJ |
| description | Microvesicles (MVs) are submicron vesicles with sizes of 0.1-1.0-µm in diameter, released from various cell types upon activation or apoptosis. Their involvement in a variety of diseases has been intensively investigated. In blood, platelets are potent MV secretors, and oxLDL, a platelet ligand, induce platelet activation and thus potentially MV secretion. This interaction occurs through binding of oxLDL with CD36, located on the platelet membrane. In this study we investigated the effect of in vitro incubation of platelets with oxLDL on MV release. Furthermore, we compared the results obtained when separating MVs larger than 0.5-µm as a measure of results obtained from less sensitive conventional flow cytometers with MVs below the 0.5-µm limit. MV size-distribution was analysed in plasma from 11 healthy volunteers (4 females, 7 males). MVs were identified as < 1-μm and positive for lactadherin binding and cell specific markers. Platelet rich plasma (PRP) was incubated without and with oxLDL or LDL (as control) to investigate the impact on platelet activation, evident by release of MVs. Size-calibrated fluorescent beads were used to establish the MV gate, and separate small- and large-size vesicles. CD41+ and CD41+CD36+ MVs increased by 6-8 fold in PRP, when left at room temperature, and the presence of cell specific markers increased. Total MV count was unaffected. Incubations with oxLDL did not increase the MV release or affect the distribution of small- and large-size MVs. We found a large inter-individual variation in the fraction of small- and large-size MVs of 73%. In conclusion, we propose that pro-coagulant activity and activation of platelets induced by interaction of platelet CD36 with oxLDL may not involve release of MVs. Furthermore, our results demonstrate great inter-individual variability in size-distribution of platelet derived MVs and thereby stresses the importance for generation of standardized protocols for MV quantification by flow cytometry. |
| first_indexed | 2024-12-19T21:52:49Z |
| format | Article |
| id | doaj.art-00b39653616f4c5cb1b222735402def8 |
| institution | Directory Open Access Journal |
| issn | 2297-055X |
| language | English |
| last_indexed | 2024-12-19T21:52:49Z |
| publishDate | 2015-11-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Cardiovascular Medicine |
| spelling | doaj.art-00b39653616f4c5cb1b222735402def82022-12-21T20:04:20ZengFrontiers Media S.A.Frontiers in Cardiovascular Medicine2297-055X2015-11-01210.3389/fcvm.2015.00037168174The effect of oxLDL on microvesicle release from platelets, measured by a sensitive flow cytometry method.Tine Bo Nielsen0Morten Hjuler Nielsen1Aase eHandberg2Aase eHandberg3Aalborg University HospitalAalborg University HospitalAalborg University HospitalAalborg UniversityMicrovesicles (MVs) are submicron vesicles with sizes of 0.1-1.0-µm in diameter, released from various cell types upon activation or apoptosis. Their involvement in a variety of diseases has been intensively investigated. In blood, platelets are potent MV secretors, and oxLDL, a platelet ligand, induce platelet activation and thus potentially MV secretion. This interaction occurs through binding of oxLDL with CD36, located on the platelet membrane. In this study we investigated the effect of in vitro incubation of platelets with oxLDL on MV release. Furthermore, we compared the results obtained when separating MVs larger than 0.5-µm as a measure of results obtained from less sensitive conventional flow cytometers with MVs below the 0.5-µm limit. MV size-distribution was analysed in plasma from 11 healthy volunteers (4 females, 7 males). MVs were identified as < 1-μm and positive for lactadherin binding and cell specific markers. Platelet rich plasma (PRP) was incubated without and with oxLDL or LDL (as control) to investigate the impact on platelet activation, evident by release of MVs. Size-calibrated fluorescent beads were used to establish the MV gate, and separate small- and large-size vesicles. CD41+ and CD41+CD36+ MVs increased by 6-8 fold in PRP, when left at room temperature, and the presence of cell specific markers increased. Total MV count was unaffected. Incubations with oxLDL did not increase the MV release or affect the distribution of small- and large-size MVs. We found a large inter-individual variation in the fraction of small- and large-size MVs of 73%. In conclusion, we propose that pro-coagulant activity and activation of platelets induced by interaction of platelet CD36 with oxLDL may not involve release of MVs. Furthermore, our results demonstrate great inter-individual variability in size-distribution of platelet derived MVs and thereby stresses the importance for generation of standardized protocols for MV quantification by flow cytometry.http://journal.frontiersin.org/Journal/10.3389/fcvm.2015.00037/fullFlow CytometryMicrovesiclesmicroparticlesplateletsCD36extracellular vesicles |
| spellingShingle | Tine Bo Nielsen Morten Hjuler Nielsen Aase eHandberg Aase eHandberg The effect of oxLDL on microvesicle release from platelets, measured by a sensitive flow cytometry method. Frontiers in Cardiovascular Medicine Flow Cytometry Microvesicles microparticles platelets CD36 extracellular vesicles |
| title | The effect of oxLDL on microvesicle release from platelets, measured by a sensitive flow cytometry method. |
| title_full | The effect of oxLDL on microvesicle release from platelets, measured by a sensitive flow cytometry method. |
| title_fullStr | The effect of oxLDL on microvesicle release from platelets, measured by a sensitive flow cytometry method. |
| title_full_unstemmed | The effect of oxLDL on microvesicle release from platelets, measured by a sensitive flow cytometry method. |
| title_short | The effect of oxLDL on microvesicle release from platelets, measured by a sensitive flow cytometry method. |
| title_sort | effect of oxldl on microvesicle release from platelets measured by a sensitive flow cytometry method |
| topic | Flow Cytometry Microvesicles microparticles platelets CD36 extracellular vesicles |
| url | http://journal.frontiersin.org/Journal/10.3389/fcvm.2015.00037/full |
| work_keys_str_mv | AT tinebonielsen theeffectofoxldlonmicrovesiclereleasefromplateletsmeasuredbyasensitiveflowcytometrymethod AT mortenhjulernielsen theeffectofoxldlonmicrovesiclereleasefromplateletsmeasuredbyasensitiveflowcytometrymethod AT aaseehandberg theeffectofoxldlonmicrovesiclereleasefromplateletsmeasuredbyasensitiveflowcytometrymethod AT aaseehandberg theeffectofoxldlonmicrovesiclereleasefromplateletsmeasuredbyasensitiveflowcytometrymethod AT tinebonielsen effectofoxldlonmicrovesiclereleasefromplateletsmeasuredbyasensitiveflowcytometrymethod AT mortenhjulernielsen effectofoxldlonmicrovesiclereleasefromplateletsmeasuredbyasensitiveflowcytometrymethod AT aaseehandberg effectofoxldlonmicrovesiclereleasefromplateletsmeasuredbyasensitiveflowcytometrymethod AT aaseehandberg effectofoxldlonmicrovesiclereleasefromplateletsmeasuredbyasensitiveflowcytometrymethod |