This is the first study to compare the effects on platelet function of different immunosuppressive regimens that are based on monotherapy. All renal transplant patients showed preactivated platelets compared with those of patients with hypertension. However, the "newer" immunosuppressive agents tacrolimus and mycophenolate mofetil seemed to have fewer unfavorable effects on platelet CD62 expression and PAC1 expression and aggregation. Whether this finding is accompanied by fewer cardiovascular events remains to be elucidated.

A controversial discussion as to whether human platelets are capable of regulated protein synthesis has been ongoing for over half a century. A previous study has suggested that human platelets synthesize large amounts of interleukin 1beta (IL-1beta) in response to external cues and in a physiologically significant manner. However, cytokines such as IL-1beta are generally considered to be products of leukocytes and it could not be completely excluded that contaminating leukocytes may have contributed to the IL-1beta results in platelet preparations. It was therefore our intention to investigate whether residual leukocytes had an impact on thrombin-induced IL-1beta synthesis. Using various methods to reduce the level of contaminating leukocytes, we found that IL-1beta production in platelet-rich suspensions is dependent on the presence of leukocytes, as it was decreased by reducing the number of leukocytes. In addition, we found that thrombin-induced IL-1beta synthesis was completely eliminated in leukocyte-free platelet preparations and could be restored by adding leukocytes. IL-1beta synthesis could be detected in platelet suspensions contaminated with at least 1 leukocyte per 10(5) platelets. This study demonstrated that platelets are incapable of synthesizing detectable amounts of IL-1beta on their own. We suggest that any IL-1beta synthesis detected is a by-product of leukocytes contaminating the platelet preparations. Thus, the hypothesis that platelets producing IL-1beta, provide a new link between thrombosis and inflammation needs to be reconsidered.

Thromboelastography methods have been used to predict or monitor treatment of haemophilia patients with recombinant activated factor VII (rFVIIa). However, neither of the two thromboelastographic methods (ROTEM and TEG) has as yet been validated. This multi-centre, randomised trial compared both methods in terms of intra- and inter- patient variability following in vivo and ex vivo rFVIIa administration to haemophilia A and B patients with and without inhibitors. Patients ((3)16 years old) received the same intravenous rFVIIa dose (45, 90 or 180 microg/kg) twice, 1-12 weeks apart. Blood samples were collected pre-dose and 15, 60, 120 and 240 minutes post-dose for ROTEM and TEG analysis. Pre-dose samples were also spiked ex vivo with rFVIIa (0.6, 1.2 or 2.4 microg/ml), to correspond to the three in vivo doses. Twenty-six haemophilia A and four haemophilia B patients were enrolled. A significant treatment effect was observed with in vivo rFVIIa (p<0.05) with more pronounced effects in inhibitor (n=14) versus non-inhibitor (n=16) patients. There was a strong positive correlation between ROTEM and TEG parameters. Intra- and inter-patient variation was large for all thromboelastography parameters at all time points and rFVIIa doses. Intra-patient variation was generally lower for non-inhibitor than inhibitor patients, and lower following ex vivo spiking versus in vivo rFVIIa administration. In conclusion, there was a clear effect of rFVIIa on all thromboelastography parameters, but the large intra- and inter-patient variability following in vivo rFVIIa administration renders the use of our method unsuitable for dose-response prediction for haemophilia patients in the clinical setting.

No abstract

Although the administration of recombinant coagulation factor VIIa (rFVIIa) is a well established treatment in haemophilia with inhibitory antibodies, monitoring the therapeutic efficacy is still a problem. This is because the complete haemostatic effect in vivo depends on negatively charged surfaces provided by exposed lipids from activated platelets which are not present in standard clinical assays using plasma. The thrombin generation assay, however, measures the endogenous thrombin potential (ETP) in platelet-rich plasma (PRP) and this assay might be useful for monitoring the haemostatic response to rFVIIa. We characterized the in vitro concentration-response relationship of rFVIIa and the thrombin generation parameters ETP, PEAK and TIME TO PEAK using platelet-rich plasma (PRP) and rFVIIa at concentrations between one and five times the therapeutic dose. We also studied the effect of inhibiting tissue-factor and the intrinsic coagulation pathway using excess TF-neutralizing antibodies and corn trypsin inhibitor (CTI), respectively. There was a sigmoid relationship between ETP and PEAK and the dose of rFVIIa. Increasing rFVIIa concentrations between 100 and 500 U/ml resulted in a progressive increase in ETP, whereas supratherapeutical concentrations led to a plateau phase. The plateau phase differed between patients, suggesting a biological variation in the maximum ETP. Neutralization of plasma TF with TF-Ab partially decreased FVIIa efficacy. The inhibitory effect of CTI on rFVIIa-induced thrombin generation via the intrinsic pathway was negligible. The thrombin generation assay using PRP is a useful test for determining the sufficient efficacy of rFVIIa in blood. Once a plateau level is reached, higher doses of rFVIIa have no additional effect on haemostatic efficacy. The variation in plateau levels between subjects indicates that there are inter-individual differences in the level of thrombin activity that can be generated. High doses of rFVIIa are effective even in the absence of TF.