Impaired thromboxane synthesis in preactivated human blood platelets: Agonist-specific, irreversible desensitization to thrombin

Hj, Reimers; Rk, Baker; Jh, Joist

doi:10.1016/0049-3848(87)90385-9

Cited by 3 publications

(3 citation statements)

References 32 publications

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“…In platelet-rich plasma (PRP) samples, DMSO final concentration did not exceed 0.25% (35 mM). Based on our experiments and reports from others, it is known that DMSO final concentrations <0.5% are absolutely unable to influence human platelet function (22) or to modify platelet cGMP concentrations; in particular, our experiments performed in healthy subjects (n = 4) demonstrated that platelet cGMP with DMSO at the final concentrations of 0, 30, and 60 mM was 16.5 ± 1.4, 15.9 ± 1.3, and 15.9 ± 1.2 pmol/10 9 platelets, respectively (NS). Protocol.…”

Section: Methodsmentioning

confidence: 91%

Insulin Increases Guanosine-3′ ,5′-Cyclic Monophosphate in Human Platelets: A Mechanism Involved in the Insulin Anti-Aggregating Effect

et al. 1994

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To investigate whether insulin reduces platelet aggregability through a modulation of the guanosine-3',5'-cyclic monophosphate (cGMP) concentrations, we determined by a radioimmunoassay the cGMP values in the platelet-rich plasma (PRP) obtained from 17 healthy volunteers and incubated for 3 min with different concentrations of human recombinant insulin (0, 240, 480, 720, 960, and 1,920 pM). Insulin induced a dose-dependent cGMP increase, from 18.5 +/- 3.3 to 42.0 +/- 6.4 pmol/10(9) platelets (P = 0.0001). This increase was completely blunted when PRP was preincubated for 20 min with the tyrosine kinase inhibitor genistein (10 microM) or with the guanylate cyclase inhibitor methylene blue (10 microM), but the increase remained highly significant (P = 0.003 and 0.009) when PRP was preincubated for 20 min with the phosphodiesterase inhibitor 3-isobutyl-1-methyl-xanthine (IBMX, 500 microM) or with the nitric oxide synthase inhibitor NG-mono-methyl-L-arginine (L-NMMA, 30 microM). Finally, the insulin-induced decrease of platelet aggregability to collagen and ADP was completely blunted when PRP was preincubated with 10 microM of the guanylate cyclase inhibitor methylene blue. This study demonstrates that the platelet anti-aggregatory effect exerted by insulin is attributable to the insulin-induced increase of cGMP that is due to a direct receptor-mediated platelet guanylate cyclase activation.

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Section: Methodsmentioning

confidence: 91%

Insulin Increases Guanosine-3′ ,5′-Cyclic Monophosphate in Human Platelets: A Mechanism Involved in the Insulin Anti-Aggregating Effect

et al. 1994

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“…The data of the present study are consistent with the hypothesis that, in patients with acute or chronic renal allograft rejection, impaired platelet thromboxane synthesis in vitro is a consequence of platelet activation in vivo, leading to secretion of platelet granular constituents and the circulation of partially and/or completely depleted platelets. Indeed, activation of platelets in vitro can result in refractoriness, including a decreased capacity for thromboxane synthesis, upon further stimulation with platelet agonists (4,(29)(30)(31)(32).…”

Section: Discussionmentioning

confidence: 99%

In Vitro Thromboxane Synthesis of Depleted Blood Platelets Following Renal Transplantation

Scharf

1990

Thromb Haemost

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SummaryRenal transplant rejection is associated with platelet activation in vivo which may lead to partially α- and γ-granule-depleted platelets that continue to circulate. These “exhausted” platelets are hemostatically defective. Tb quantitate the extent of platelet granule depletion following kidney transplantation, we determined intraplatelet levels of β-thromboglobulin (βTG), platelet factor 4 (PF4), and serotonin (5-hydroxytryptamine, 5-HT) ex vivo in Tiiton X-1O0-treated platelet lysates. To explore biochemical alterations of partially depleted platelets, we studied platelet thromboxane A2 (TXA2) synthesis in citrated plateletrich plasma (PRP) upon stimulation with thrombin or collagen in 45 recipients of renal allografts and 10 healthy volunteers. The patients were divided into subjects with acute and chronic allograft rejection (N = 15), those with compensated renal failure after kidney transplantation but without evidence of allograft rejection (N = 15), and those with functioning renal transplant (N = 15). The mean intraplatelet content of βTG (38.6 ± 4.2 μE/109 platelets), PF4 (11.8 ± 1.8 μg/109 platelets), and 5-HT (274 ± 31 μg/109 platelets) in patients with acute or chronic renal allograft rejection was significantly lower than in other recipients off < idney transplants or healthy volunteers (βTG: 59.9±4.7 μgl 109 platelets; PF4: 20.4±2.3 ¼g/n platelets; s-rrr: 46lraB ngl 10e platelets; p < 0.ffi5 in all casls). Platelet TxB2 formation upon stimulation with thrombin (10 U/ml) or collagen (6.25 ¼g/ml) for 5 min was significantly reduced in patients with acute or chronic renal allograft rejection (2.25±0.29 and 0.641 0.08 nmoUl0e platelets for thrombin- and collagen-stimulated platelets, respectively) compared to that of healthy volunteers (4.72± 0.60 and 1.35 ± 0.12 nmol/109 platelets, respectively; p <0.05 in all cases). In contrast, platelet TXB2 formation of patients with functioning kidney transplant or those with compensated renal failure but without evidence of transplant rejection did not differ significantly from that of normals. These results confirm that platelets with reduced levels of α- and β-granular constituents are detectable in the circulation following kidney transplantation when acute or chronic renal allograft rejections occur. These platelets are incapable of forming normal amounts of thromboxane upon stimulation with thrombin and collagen in vitro. This dysfunction of thromboxane synthesis, due to alterations in the platelet arachidonate pathway, may reflect the previous activation of platelets in vivo associated with acute or chronic renal allograft rejection.

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“…5 U/ml), showing that they were still functional (data not shown and Reimers et al, 1987;KinloughRathbone et al, 1991). Moreover, these cells have fully re-established both their adenylate energy charge and the ionic gradients across their membranes, indicating their metabolic recovery (data not shown).…”

Section: Methodsmentioning

confidence: 99%

analysis of CD9, CD32 and p67 signalling: use of degranulated platelets indicates direct involvement of CD9 and p67 in integrin activation

et al. 1997

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Summary. The use of agonist monoclonal antibodies (mAbs) to probe the signalling function of platelet membrane proteins is severely limited by the dependence of the mAb effect on Fc-FcgRII interaction. Furthermore, in addition to its anchoring role, the FcgRII receptor itself generates a stimulation signal resulting in granule secretion. Platelet stimulation by the released granule contents can then further obscure the original activation signal. Here we demonstrate that these problems are largely overcome by the use of platelets which had been degranulated with thrombin prior to stimulation with mAbs. We found that, like intact cells, degranulated platelets could also be activated and induced to aggregate by mAbs against a 67 kD membrane protein (known as PTA1) and CD9, and by crosslinked CD32 (FcgRII). However, the signal generated by crosslinked FcgRII was weak compared with that induced by the other monoclonal antibodies. Thus, by diminishing the FcgRII signal contribution, we have succeeded for the first time to clearly dissect the target antigen signal from that generated by FcgRII. In addition to differences in the degree of aggregation, analysis of the signals generated by each mAb showed differences in Ca 2+ fluxes and protein phosphorylation. Moreover, the signals generated by CD9 and PTA1 antigens differed significantly in their sensitivity to PKC inhibition or ADP-ribosylation of the small GTP-binding protein rhoA. Despite these differences, the signals initiated by all three antigens converged to a common signalling pathway which included activation of tyrosine kinase(s). The pattern of protein phosphorylation strongly resembled that induced by gpIIb/IIIa-mediated platelet interaction with macromolecular ligands and by mutual cell contact. The multiple intercellular links formed by mAb would have a similar effect since the Fc-receptor anchorage required for antigen stimulation is already known to be provided by adjacent cells. The present findings suggest that the function of both CD9 and PTA1 antigens is closely associated with gpIIb/IIIa activation.

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Impaired thromboxane synthesis in preactivated human blood platelets: Agonist-specific, irreversible desensitization to thrombin

Cited by 3 publications

References 32 publications

Insulin Increases Guanosine-3′ ,5′-Cyclic Monophosphate in Human Platelets: A Mechanism Involved in the Insulin Anti-Aggregating Effect

Insulin Increases Guanosine-3′ ,5′-Cyclic Monophosphate in Human Platelets: A Mechanism Involved in the Insulin Anti-Aggregating Effect

In Vitro Thromboxane Synthesis of Depleted Blood Platelets Following Renal Transplantation

analysis of CD9, CD32 and p67 signalling: use of degranulated platelets indicates direct involvement of CD9 and p67 in integrin activation

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