Multistability in platelets and their response to gold nanoparticles.

The nanoparticle (NP) response of platelets is shown to be critically dependent on extent of preactivation of platelets by an agonist like ADP. A transition from de-aggregatory to aggregatory state is triggered in the presence of gold NPs (AuNP) only in such critical conditions. Adhered and suspended platelets respond differentially to NPs. Preactivation in the adhered state induced by shear force explains such observation. The NP effect is associated with enhanced release reaction, tyrosine phosphorylation and CD62P expression level. Unlike cancer cells, whose response is maximal when NP size is optimal (within the range 50 - 70 nm), the platelet response monotonically increases with reduction of the AuNP size. The uptake study, using quenching of quinacrine hydrochloride fluorescence by AuNP, indicates that accumulation 18 nm AuNP is several-fold higher than the 68 nm AuNP. It is further shown that AuNP response can provide a simple measure for thrombotic risk associated with nano-drugs. FROM THE CLINICAL EDITOR: Platelet aggregation can be triggered in the presence of gold nanoparticles (AuNP). Platelet response monotonically increases with reduction of the AuNP size. AuNP response can provide a simple measure for thrombotic risk associated with nano-drugs.

Platelet responsiveness to yohimbine hydrochloride and MRS2179 in the context of the interaction between collagen and epinephrine in acute coronary syndrome.

Acute coronary syndrome (ACS) covers a spectrum of clinical conditions ranging from unstable angina, Non-ST segment elevation myocardial infarction (NSTEMI), or ST segment elevation myocardial infarction (STEMI). This study encompasses patients with acute coronary syndrome, who were receiving the dual antiplatelet therapy of aspirin and clopidogrel. The focus of the study was to gain insight into the role of selective P2Y1 antagonism using MRS2179 in such cases as well as its effects, if any, on collagen-epinephrine interaction. All the cases showed greater potency of inhibition of the interaction when yohimbine hydrochloride (YH), a blocker of alpha2A-adrenoreceptor, was used compared to MRS2179, a P2Y1 antagonist, although there was variability in responsiveness to the antiplatelet drugs. These findings indicate that alpha2A-adrenoreceptors of platelets in this group play a major role in precipitating the interactive effect of collagen and epinephrine. The dose-response effect as studied by platelet aggregometry showed that the required molar concentration to block the interactive effect in the case of YH was less than that of MRS2179. Hence, it is postulated that although there may be an impairment of collagen-induced aggregation by MRS2179, the interactive effect of collagen-epinephrine may not be impaired by MRS2179 as efficaciously as YH.

Dual antiplatelet drug resistance in patients with acute coronary syndrome.

AIMS AND OBJECTIVES: Antiplatelet therapy is a cornerstone in the management of the atherosclerotic vascular disease. Aspirin and clopidogrel are the two most commonly used antiplatelet drugs in its management. Recently, there has been a concern about the development of resistance to one or both antiplatelet agents with potentially devastating consequences. In this study we tried to assess the in vitro resistance to antiplatelet agents in patients presenting with acute coronary syndrome (ACS). MATERIALS AND METHODS: 144 patients presenting with ACS, who were not on any antiplatelet therapy prior to hospital admission were evaluated in this study. Baseline clinical data was obtained before giving the oral loading dose of aspirin and clopidogrel. Patients received a loading dose of 325 mg of aspirin and 300 mg of clopidogrel followed by a daily dose of 150 mg. of aspirin and 75 mg.of clopidogrel. After 7 days of dual antiplatelet therapy, platelet aggregation pattern was analyzed using optical aggregometer (chrono-log). Response to aspirin and clopidogrel was assessed by interaction with collagen (2microg/ml) and Adenosine diphosphate (ADP) (10micro/ml) respectively. The results were analyzed. Response to doubling the dose of antiplatelet agents was also observed in 6 aspirin resistant patients, 12 clopidogrel resistant patients and in 6 patients resistant to the effect of dual antiplatelet agents. RESULTS: There were 22 patients (15.27%) who showed poor response to aspirin, 28 patients (19.44%) to clopidogrel (primary non-responder) and 18 patients (12.5%) showed a primary non-responsiveness to both the antiplatelet agents in the usual doses. After dose doubling, all 6 aspirin resistant patients showed adequate response but 4 out of 12 clopidogrel resistant patients showed inadequate response. CONCLUSIONS: This pilot study brings out a disquieting picture of 12.5% patients suffering from ACS showing resistance to the antiplatelet effects of both aspirin and clopidogrel in the conventional dose. A long-term prospective randomized controlled trial is required to give an insight into this problem and its clinical consequences.

Antiplatelet drug resistance in patients with recurrent acute coronary syndrome undergoing conservative management.

AIMS AND OBJECTIVES: Recurrent ischemic events continue to occur despite combination anti-platelet therapy. Currently aspirin, clopidogrel and dual resistance are increasingly recognized entities. The relationship of such resistance to recurrent ischemic events is largely unknown. In this study, we tried to gain an insight into the role of antiplatelet drug resistance with recurrent Acute Coronary Syndrome (ACS). MATERIALS AND METHODS: The antiplatelet effect of aspirin and clopidogrel was studied in 40 recurrent ACS patients and 170 patients with first episode of ACS, after > or = 7 days of dual antiplatelet therapy. Platelet aggregation study was done with optical aggregometer. Resistance to aspirin and clopidogrel was defined as > or = 50% aggregation with collagen and ADP respectively. RESULTS: Aspirin, clopidogrel and dual drug resistance were encountered respectively in 35%, 72.5% and 32.5% patients with recurrent ACS. The corresponding figures for the patients with first episode of ACS were 25.3%, 42.3% and 18.8% respectively. P values for the comparisons were 0.237 for aspirin, 0.0007 for clopidogrel and 0.084 for dual drugs. Patients with recurrent ACS were relatively younger and had a higher prevalence of conventional risk factors like hypertension, diabetes and elevated LDL. CONCLUSION: Antiplatelet drug resistance is likely to play an important role in recurrent ACS alongside other conventional risk factors. Further research is required in this field to have a definitive conclusion.

Cell selective response to gold nanoparticles.

Gold nanoparticles (GNPs) are considered a potential probe to detect cancer. The present article investigates whether GNPs, even in the absence of any specific functionalization, induce any cell-specific response. We report GNP-induced death response in human carcinoma lung cell line A549. In contrast, the two other cell lines tested, BHK21 (baby hamster kidney) and HepG2 (human hepatocellular liver carcinoma), remained unaffected by GNP treatment. The specificity of the induction of the death response in A549 cells implies that GNPs do not universally target all cell types. Flow-cytometric studies indicated that the response was dose dependent and had a threshold effect (in A549). Gradual increase in GNP concentration induces a proportional cleavage of poly(ADP-ribose) polymerase. The programmed nature of the death response is implied, because such cleavage follows activation of caspases. Notably, at higher GNP concentration there was an asymmetric accumulation of GNPs in the periphery outside the cell nucleus of the A549 cells. This was confirmed by confocal microscopy, a green scattering (possibly, surface-enhanced Raman effect) appearing on selective z-slices of the image.

Platelet aggregation profile as a marker of hydroxyurea bioavailability through nitric oxide generation in chronic myelogenous leukemia.

Platelet aggregation profiles were studied in chronic myelogenous leukemia patients who were undergoing hydroxyurea therapy. Nitric oxide (NO) generation induced by hydroxyurea was measured from the altered aggregatory response, in which the platelet suspension exhibits a de-aggregatory behaviour. NO caused platelet de-aggregation by generation of cyclic guanidine monophosphate through the activation of soluble guanylate cyclase (SGC). The fact that the observed response is specific to NO was confirmed by the reversal of the de-aggregatory behaviour in the presence of (1)H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), an inhibitor of SGC. Among the subjects studied, one subset showed an hydroxyurea-induced de-aggregatory effect that was inhibited by ODQ, whereas another subset did not show any such effect. The observed inter-individual variability in platelet aggregometric response after the ingestion of drugs may be an indicator for NO generation from hydroxyurea, and this may help to explain the drug efficacy encountered in such cases.

A study of aspirin resistance in type 2 diabetes.

Aspirin is currently known to give inadequate protection against coronary artery disease in diabetes compared to person without it. We evaluated 97 consecutive patients with type 2 diabetes for assessing laboratory aspirin resistance and attempted to assess the impact of various clinical and biochemical parameters on it. Thirty-eight patients (39.1%) were found to be less sensitive to the action of aspirin, 7 persons (7.2%) were found to be resistant and 31 persons (31.9%) were aspirin semi-responders. Only total cholesterol, LDL-cholesterol and triglyceride had statistically significant impact on aspirin resistance (p<0.05). Three persons out of 9 with some form of macrovascular disease had aspirin resistance.

Insensitivity to the alpha2-adrenergic receptor blocker yohimbine hydrochloride and occurrence of spontaneous platelet macroaggregation (SPMA) in diabetes.

We report here a study of platelet aggregation in diabetes, induced by epinephrine and its inhibition by yohimbine hydrochloride (YH), an alpha(2)-adrenergic receptor-blocking agent. Interestingly, emergence of spontaneous platelet macroaggregation (SPMA) was observed in six out of 75 cases in the absence of any agonist. The SPMA cases were strongly associated with insensitivity to YH (in contrast with non-SPMA cases) when epinephrine was used as an agonist. We suggest that the observed correlation is a result of over expression of platelet alpha(2)-adrenoceptors in such subjects. The quantitative nature of the effect is supported by the observation that addition of YH at higher concentration (more than 5 microM) led to restoration of the adrenergic receptor-blocking activity of the said agent. Eventually for non-SPMA subjects YH exhibited blocking activity even at lower concentration. The aggregation profile and the platelet morphology of the SPMA cases had distinctive features as compared to microaggregates formed in other diabetic subjects (non-SPMA cases).

Structural insights in platelet receptor synergism-antiplatelet therapy in post-ischemic cerebrovascular events.

Synergy between agonists of platelet aggregation, namely, ADP and epinephrine, has been studied in patients having a history of cerebrovascular ischemic event. There is a significant variability of responsiveness among individuals towards clopidogrel, which is a specific inhibitor of the low-affinity human purinergic receptor (P2Y12). For responders of clopidogrel, simultaneous application of ADP and epinephrine at sub-threshold concentrations (i.e., concentration below the threshold concentration at which aggregation occurs) leads to platelet aggregation, which is followed by deaggregation. For non-responders of the drug, the synergism seems to be stronger, showing no deaggregatory pattern. The inhibition of synergism by yohimbine hydrochloride (YH), a blocker of alpha2A-adrenoreceptors is more pronounced in non-responders. A simple structural model based on receptor-receptor interaction is proposed to explain the synergism. The model explains synergy in terms of cooperative interaction between the low-affinity ADP receptor P2Y12 (Swiss Prot:Q9H244) and the alpha2A-adrenoreceptor (Swiss Prot:P08913). It follows that the synergistic effect can be achieved in only one of the two 3D structures for the alpha2A-adrenoreceptor P08913 permitted by homology modeling, as there is a better docking interface with the Q9H244. The synergism itself and the observed dichotomous phenomenon in relation to inhibition of synergism among responders and non-responders can be accounted for, if the interacting receptors on the dynamic membrane interface compete with the clopidogrel binding.