Dear Hiwa, your question is not at all easy to answer.

In our hands, thrombin at 0.3 U/ml (which is quite high) provokes marked VASP Ser157 phosphorylation in isolated human platelets, whereas thrombin has only minor effects on Ser239 phosphorylation. It has been reported that VASP Ser157 phosphorylation is predominately achieved by PKA activation via an increase in cAMP, whereas that of Ser239 is mainly due to cGMP/PKG signalling. Further, we found that, despite 'extensive' VASP phosphorylation at both serine-residues by increasing cAMP with adenosine and cGMP with SNAP, full platelet aggregation occured when we combined thrombin (Gq) with epinephrine (Gi/z) (so far unpublished data).

Together, although I'm not sure if you could 'claim' this, thrombin in fact induces VASP Ser157 phosphorylation. Thrombin, the major effector serine-protease of the coagulation cascade, is certainly one of the most potent platelet agonist. As nature is smart, there may be a -whatsoever- negative feed-back mechanism to prevent an overall thrombotic event.

Finally, 'some researchers believe...this or that'...means nothing!

The point is: go to lab, perform serious research, obtain data, and publish them.

If they are right or wrong ...will show the coming decade.

Best regards,

Knut

Dear Dr. Knut Fälker

thanks a lot for your good information. I study exercise physiology. there are so many investigations that report exercise training induce thrombin generation (specifically with high intensity) after on session, may be because of increased shear stress, that result in increased platelet activation. but after a period of training (for example 4 weeks) by conditioning to exercise, despite the presence of shear stress and of course thrombin, the level of platelet activation decrease significantly. I think in this situation thrombin play different role with different signaling pathway by VASP phosphorylation.

I would be very appreciated if let know your idea about that

best wishes

hiwa

Dear Hiwa,

as we perform in vitro sudies using isolated platelets and pharmacological compounds, an exaggeration to in vivo studies are mostly merely speculative. You may also consider that frequent exercise, and therfore blood circulation and especially shear, will increase the generation and release of nitric oxid (NO) from endothelial cells, which via an increase of cGMP results in an increase of platelet VASP Ser157 phosphorylation in an extent comparable to that induced by thrombin. As initially mentioned, VASP Ser157 may be difficult to explain when considering all parameters under physiological/in vivo conditions.

Best regards,

Knut

Dear Dr. Knut Fälker

Thanks a lot again for your kind reply. As you mentioned diffuse of NO (generated by endothelial cells) into platelet and also PCG-I2 that bind to its receptor is main inhibitory pathway specially after exercise training. we are going to investigate it in different project. But we like to consider the role of thrombin in reduction of platelet activation after conditioning to exercise. as you mentioned " is not at all easy to answer" and " VASP Ser157 may be difficult to explain when considering all parameters under physiological/in vivo conditions". Gambryan et al. (2010) suggest this role for collagen and thrombin by activating catalytic subunit of PKA after dissociating it from NFkB-IkB-PKA complex (http://www.ncbi.nlm.nih.gov/pubmed/20356841 )

please see Figure 8 in page 18362 of this article. may be there is a factor that link thrombin to IKK. My question (or my problem) is that: after measuring thrombin and VASP and finding possible relation (statistically) between them, which factor strengthen the possible relation? PKAc, IKK or IkB-P? or may be another factor?

thank you so much

Best regards

HIwa

Dear Hiwa, I of course know the work from Stepan Gambaryan and Ulrich Walter (as I'm from Germany as well). However, don't add more confusion to your work. I suggest to consider to assess phosphorylation of VASP at both Ser157 as well as Ser239 (For an overall view take a look at Albert Smolesnki's review in JTH from 2012). In our hands, the cAMP-elevator adenosine (or PGI2) induces prominent Ser157 phosphorylation, whereas the NO-donor SNAP induces half maximal Ser157 phosphorylation, which to a similar extent is provoked by thrombin. In contrast, Ser239 is prominently phosphorylated in Response to SNAP, only minor by adenosine, and merely by thrombin. Hence, when you compare platelet VASP phosphorylation (from untrained vs trained individuals) and see an effect on Ser157 phosphorylation that may be due to either an increase/decrease of adenosin/PGI2 or thrombin-generation (and NFkB (to be discussed )), respectively. However, when you observe an increase in Ser239 phosphorylation under those conditions the inhibitory effect may be attributed to an increase of NO by the endothelium due to frequent exercise/increase in flow.

For your complete confusion, please find one of our figures (confidential) attached.

Best regards,

Knut

Dear Dr. Knut Fälker

I read your answers and suggestions again and again. based on your explain and also the information in the figure (that you send me) I convinced that it is better to see both Ser157 and Ser239. but because of some research problems in Iran, probably we investigate ser157/239 in two pathway (NO/PGI2 and Thrombin) by two research program.

I already read Albert Smolesnki's review. that is so grate article.

first I should plan a good study design and exercise program to be approved.

thanks for the article and Figure that you sent to me

best wishes

Hiwa