Limits...
The Marine-Derived Kinase Inhibitor Fascaplysin Exerts Anti-Thrombotic Activity.

Ampofo E, Später T, Müller I, Eichler H, Menger MD, Laschke MW - Mar Drugs (2015)

Bottom Line: This was associated with a decreased platelet aggregation.Fascaplysin also decreased PLA formation after PMA but not PAR-1-AP and ADP stimulation.Fascaplysin exerts anti-thrombotic activity, which represents a novel mode of action in the pleiotropic activity spectrum of this compound.

View Article: PubMed Central - PubMed

Affiliation: Institute for Clinical and Experimental Surgery, Saarland University, 66421 Homburg/Saar, Germany. emmanuel.ampofo@uks.eu.

ABSTRACT

Background: The marine-derived kinase inhibitor fascaplysin down-regulates the PI3K pathway in cancer cells. Since this pathway also plays an essential role in platelet signaling, we herein investigated the effect of fascaplysin on thrombosis.

Methods: Fascaplysin effects on platelet activation, platelet aggregation and platelet-leukocyte aggregates (PLA) formation were analyzed by flow cytometry. Mouse dorsal skinfold chambers were used to determine in vivo the effect of fascaplysin on photochemically induced thrombus formation and tail-vein bleeding time.

Results: Pre-treatment of platelets with fascaplysin reduced the activation of glycoprotein (GP)IIb/IIIa after protease-activated receptor-1-activating peptide (PAR-1-AP), adenosine diphosphate (ADP) and phorbol-12-myristate-13-acetate (PMA) stimulation, but did not markedly affect the expression of P-selectin. This was associated with a decreased platelet aggregation. Fascaplysin also decreased PLA formation after PMA but not PAR-1-AP and ADP stimulation. This may be explained by an increased expression of CD11b on leukocytes in PAR-1-AP- and ADP-treated whole blood. In the dorsal skinfold chamber model of photochemically induced thrombus formation, fascaplysin-treated mice revealed a significantly extended complete vessel occlusion time when compared to controls. Furthermore, fascaplysin increased the tail-vein bleeding time.

Conclusion: Fascaplysin exerts anti-thrombotic activity, which represents a novel mode of action in the pleiotropic activity spectrum of this compound.

Show MeSH

Related in: MedlinePlus

Effect of fascaplysin on leukocytic CD11b expression. (A–C) Whole blood was incubated with 10 μM fascaplysin (black bars, n = 6) or vehicle (DMSO, white bars, n = 6) for 0.5 h followed by stimulation with PAR-1-AP, ADP or PMA. Data are given in % of vehicle. Unstimulated whole blood served as negative control (grey bars, n = 6). Expression of CD11b on all leukocytes (A), monocytes (B) and granulocytes (C) were assessed by flow cytometry using double fluorescence staining (CD45/CD11b). Mean ± SEM. * p < 0.05 vs. vehicle.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4663553&req=5

marinedrugs-13-06774-f005: Effect of fascaplysin on leukocytic CD11b expression. (A–C) Whole blood was incubated with 10 μM fascaplysin (black bars, n = 6) or vehicle (DMSO, white bars, n = 6) for 0.5 h followed by stimulation with PAR-1-AP, ADP or PMA. Data are given in % of vehicle. Unstimulated whole blood served as negative control (grey bars, n = 6). Expression of CD11b on all leukocytes (A), monocytes (B) and granulocytes (C) were assessed by flow cytometry using double fluorescence staining (CD45/CD11b). Mean ± SEM. * p < 0.05 vs. vehicle.

Mentions: The formation of PLA is not only dependent on the activation of platelets, but also on the expression of leukocytic adhesion molecules. In this context, several studies reported that the leukocytic integrin Mac-1 (CD11b/CD18) mediates the binding to GPIIb/IIIa on platelets via fibrinogen bridging [28,29,30]. Accordingly, we further assessed the expression of CD11b on leukocytes by means of flow cytometry. We found that fascaplysin treatment markedly increases the levels of CD11b following PAR-1-AP or ADP stimulation. However, PMA-induced CD11b levels were not affected by exposure of the cells to fascaplysin (Figure 5A). The additional analysis of different leukocyte subpopulations revealed that only granulocytes showed a similar expression pattern, but not monocytes (Figure 5B,C). This indicates that fascaplysin-induced up-regulation of CD11b expression primarily occurs in granulocytes.


The Marine-Derived Kinase Inhibitor Fascaplysin Exerts Anti-Thrombotic Activity.

Ampofo E, Später T, Müller I, Eichler H, Menger MD, Laschke MW - Mar Drugs (2015)

Effect of fascaplysin on leukocytic CD11b expression. (A–C) Whole blood was incubated with 10 μM fascaplysin (black bars, n = 6) or vehicle (DMSO, white bars, n = 6) for 0.5 h followed by stimulation with PAR-1-AP, ADP or PMA. Data are given in % of vehicle. Unstimulated whole blood served as negative control (grey bars, n = 6). Expression of CD11b on all leukocytes (A), monocytes (B) and granulocytes (C) were assessed by flow cytometry using double fluorescence staining (CD45/CD11b). Mean ± SEM. * p < 0.05 vs. vehicle.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4663553&req=5

marinedrugs-13-06774-f005: Effect of fascaplysin on leukocytic CD11b expression. (A–C) Whole blood was incubated with 10 μM fascaplysin (black bars, n = 6) or vehicle (DMSO, white bars, n = 6) for 0.5 h followed by stimulation with PAR-1-AP, ADP or PMA. Data are given in % of vehicle. Unstimulated whole blood served as negative control (grey bars, n = 6). Expression of CD11b on all leukocytes (A), monocytes (B) and granulocytes (C) were assessed by flow cytometry using double fluorescence staining (CD45/CD11b). Mean ± SEM. * p < 0.05 vs. vehicle.
Mentions: The formation of PLA is not only dependent on the activation of platelets, but also on the expression of leukocytic adhesion molecules. In this context, several studies reported that the leukocytic integrin Mac-1 (CD11b/CD18) mediates the binding to GPIIb/IIIa on platelets via fibrinogen bridging [28,29,30]. Accordingly, we further assessed the expression of CD11b on leukocytes by means of flow cytometry. We found that fascaplysin treatment markedly increases the levels of CD11b following PAR-1-AP or ADP stimulation. However, PMA-induced CD11b levels were not affected by exposure of the cells to fascaplysin (Figure 5A). The additional analysis of different leukocyte subpopulations revealed that only granulocytes showed a similar expression pattern, but not monocytes (Figure 5B,C). This indicates that fascaplysin-induced up-regulation of CD11b expression primarily occurs in granulocytes.

Bottom Line: This was associated with a decreased platelet aggregation.Fascaplysin also decreased PLA formation after PMA but not PAR-1-AP and ADP stimulation.Fascaplysin exerts anti-thrombotic activity, which represents a novel mode of action in the pleiotropic activity spectrum of this compound.

View Article: PubMed Central - PubMed

Affiliation: Institute for Clinical and Experimental Surgery, Saarland University, 66421 Homburg/Saar, Germany. emmanuel.ampofo@uks.eu.

ABSTRACT

Background: The marine-derived kinase inhibitor fascaplysin down-regulates the PI3K pathway in cancer cells. Since this pathway also plays an essential role in platelet signaling, we herein investigated the effect of fascaplysin on thrombosis.

Methods: Fascaplysin effects on platelet activation, platelet aggregation and platelet-leukocyte aggregates (PLA) formation were analyzed by flow cytometry. Mouse dorsal skinfold chambers were used to determine in vivo the effect of fascaplysin on photochemically induced thrombus formation and tail-vein bleeding time.

Results: Pre-treatment of platelets with fascaplysin reduced the activation of glycoprotein (GP)IIb/IIIa after protease-activated receptor-1-activating peptide (PAR-1-AP), adenosine diphosphate (ADP) and phorbol-12-myristate-13-acetate (PMA) stimulation, but did not markedly affect the expression of P-selectin. This was associated with a decreased platelet aggregation. Fascaplysin also decreased PLA formation after PMA but not PAR-1-AP and ADP stimulation. This may be explained by an increased expression of CD11b on leukocytes in PAR-1-AP- and ADP-treated whole blood. In the dorsal skinfold chamber model of photochemically induced thrombus formation, fascaplysin-treated mice revealed a significantly extended complete vessel occlusion time when compared to controls. Furthermore, fascaplysin increased the tail-vein bleeding time.

Conclusion: Fascaplysin exerts anti-thrombotic activity, which represents a novel mode of action in the pleiotropic activity spectrum of this compound.

Show MeSH
Related in: MedlinePlus