ARACHIDONIC ACID (AA) INDUCED AGGREGATION OF RAT PRO- MEGAKARYOBLASTS (RPM)

Platelet aggregation can be induced in vitro by a variety of platelet agonists acting upon membrane receptors. Since platelets have only a limited ability to synthesise proteins, these receptors must be present in megakaryocytes. This was investigated using an eternal line line of RPMs. Cells were suspended in rat platelet free plasma (PFP) at a concentration of 10⁵ cells ml^-1 . 200μl aliquots of this were placed in a light aggregometer. For this suspension, and for an aliquot of PFP, light transmission was adjusted to zero and 100% respectively. Addition of ADP (plasma concentrations 1-100μM), thrombin (0.5-5 I.U. ml^-1 ), and adrenaline (0.1-1 I.U. ml^-1 ) to the suspension caused no change in transmission. However, addition of AA (1.5-6mM) increased light transmission, indicating RPM aggregation. Radioimmunoassay (RIA) on the resultant supernatant showed no thromboxane B2 was produced. Scanning and transmission electron microscopy showed the aggregate to be composed of non-lysed cells. Aggregation of RPMs was not inhibited by preincubation with PGL (1500ng ml^-1 ) indomethacin (100μg ml^-1 ) or fenoprofen (100²μg ml^-1 ). However, preincubation with aspirin (30μM) blocked aggregation.

These results indicate that RPMs can aggregate in response to AA. Mechanism of this is unlike that observed for platelets, since PGI₂, indomethacin and fenoprofen did not block aggregation. The response was only inhibited by aspirin. Indomethacin, fenoprofen and aspirin are all known inhibitors of cyclo-oxygenase. In addition, aspirin also blocks 12-lipoxygenase. Therefore, this may suggest that the effect of AA on RPMs is mediated via 12-lipoxygenase pathway. Further investigations are in progress