Antiplatelet Resistance in Coronary Artery Bypass Grafting: A Systematic Review

Background This systematic review examines the occurrence and implications of resistance to primary antiplatelet agents, aspirin and clopidogrel, often utilised in patients undergoing coronary artery bypass grafting (CABG), alongside the methodologies for assessment of such resistance. Methods An extensive literature search across various databases such as PubMed, MEDLINE via Ovid, Embase, and Cochrane CENTRAL until May 2024 was conducted to identify studies evaluating antiplatelet resistance in on-pump and off-pump CABG patients. Following quality assessment, only high-quality studies were incorporated into this review. Results This review included 19 studies with 3,915 patients, four of which were randomised controlled trials and 15 were observational studies. Aspirin resistance incidence ranged from 11.0% to 51.5%, while clopidogrel resistance was 22%. Antiplatelet resistance, assessed through a wide variety of methods, was associated with a 13 times increase in the risk of vein graft occlusion and increased rates of mortality, myocardial infarction, and target vessel revascularisation in the case of clopidogrel resistance. The effect of cardiopulmonary bypass on antiplatelet resistance remains ambiguous. Conclusion The academic literature lacks a standardised definition for antiplatelet resistance. Assessment methodologies greatly vary, leading to noninterchangeable outcomes. While aspirin resistance has a conflicting overall significant impact on adverse outcomes, clopidogrel resistance correlates with poorer clinical outcomes.


Introduction
In patients undergoing coronary artery bypass grafting (CABG), the two commonly used antiplatelet agents are aspirin and clopidogrel.Early aspirin within 48 hours after CABG serves to mitigate mortality risk and the incidence of organ ischemia in the brain, kidneys, heart, and gastrointestinal tract [1].Te initiation of aspirin soon after CABG surgery has also been substantiated to signifcantly enhance the patency of vein grafts without increasing the bleeding risk [2].Aspirin functions by irreversibly acetylating the platelet cyclooxygenase (COX) enzyme, hence inhibiting the conversion of arachidonic acid to thromboxane A2 (TxA2).Due to its inherent chemical instability, TxA2 undergoes conversion into the stable, inactive thromboxane B2 (TxB2).Te resultant metabolite is 11-dehydro-TxB2, and both are detectable in urine.
Dual antiplatelet therapy, incorporating aspirin and clopidogrel, mitigates the incidence of thrombotic complications following acute coronary syndrome (ACS) [3].Clopidogrel is an adenosine diphosphate (ADP) receptor antagonist.It inhibits platelet activation by binding to the P2Y12 receptor irreversibly.Tis dual therapy leads to a reduction in all-cause mortality and improves vein graft patency, exerting more signifcant efects on ACS patients undergoing CABG surgery [4,5].
However, antiplatelet resistance noted in a subset of patients has been implicated in early graft failure, attributed to suboptimal responsiveness to the antiplatelet agents administered [6].Tough a range of tests exists for assessing antiplatelet resistance, their precision varies, and correlations among them are not consistent [7].Tis systematic review intends to explore contemporary practices, the application of assessment methodologies, and the ramifcations of antiplatelet resistance in patients undergoing CABG surgery.

Methods
Te present systematic review was conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines [8].Ethical approval or patient consent was not sought as this review solely relied on preexisting published studies.

Search Strategy.
A systematic search was carried out across major databases of PubMed, MEDLINE via Ovid, Embase, and the Cochrane Library Database until May 2024 to identify eligible studies using Boolean operators to achieve maximum sensitivity.Te search terms used are "((CABG) OR (Coronary artery bypass graft * ) OR (Cardiac surgery)) AND ((Antiplatelet) OR (Aspirin) OR (Clopidogrel) OR (Ticagrelor) OR (Antithrombotic)) AND ((Mortality) OR (Morbidity) OR (Graft patency) OR (Survival)) AND ((Resistance) OR (platelet mapping)) NOT (stent)."Bibliographies of relevant studies were also screened manually to identify additional suitable studies.

Study Selection and Data
Extraction.Te inclusion criteria include human studies, where patients underwent CABG surgery and had received a minimum of one antiplatelet agent during the perioperative phase.Te included studies must also provide at least one outcome related to antiplatelet resistance, such as vein graft failure, mortality, or morbidity.Animal studies, case reports, case series, reviews, and non-English articles were excluded.
Two authors independently conducted the database search (MST and SE), reviewed articles for potential relevance, extracted data, and assessed the quality and risk of bias in the included studies.Discrepancies were reconciled through consensus or, if needed, consultation with the third author (AK).

Quality Assessment of Included Studies.
Te quality of the observational cohort studies was evaluated using the Newcastle-Ottawa Scale (NOS), designating scores above six as indicative of high-quality studies [9].For the assessment of randomised controlled trials, the Cochrane risk of bias assessment tool was used [10].

Characteristics of the Studies.
Te systematic search identifed 237 studies in total as described in the PRISMA fowchart (Figure 1).After removing 89 duplicates, titles and abstracts of the remaining 148 studies were screened.A further 129 studies were excluded leaving 19 studies for fulltext review, after which all 19 studies were found to meet inclusion criteria.Terefore, a total of four randomised controlled trials and 15 observational cohort studies with a total of 3,915 patients were included in the systematic review [11][12][13][14][15][16][17][18][19][20][21][22][23][24][25][26][27][28].Te specifc characteristics of these studies are outlined in Table 1.Each of the 15 observational studies achieved a score of 6 on the Newcastle-Ottawa Scale (NOS), thus afrming them as good-quality observational studies (Supplementary Table 1).Similarly, the four randomised controlled trials were deemed low-risk and qualifed as highquality studies (Supplementary Figure 1) [10].Te graphical summary is illustrated in Figure 2.

Assessment of Antiplatelet
Resistance.Te evaluation of antiplatelet resistance is characterised by considerable variation, employing an array of diferent platelet function tests and utilising the downstream metabolites of arachidonic acid breakdown, such as serum thromboxane B2 (TxB2) or its urinary metabolite, 11-dehydro-TxB2.Tese metabolites serve to refect the impact of aspirin on platelet function.

Light Transmission Aggregometry.
Te preparation of platelet-rich plasma (PRP) involves centrifuging a 5 ml anticoagulated blood sample at 150g for 10 minutes at room temperature.Subsequently, the PRP is calibrated to a platelet count ranging from 150,000 to 300,000 µl.Light transmission aggregometry is then employed to assay the samples, necessitating the addition of 0.05 ml of arachidonic acid.However, the process can also be undertaken without the adjustment of the platelet count, and alternatives such as type I collagen and ADP may be used in lieu of arachidonic acid [11].Te degree of aggregation is plotted as a function of time and represented as the total percentage of aggregation at the fve-minute mark [14,24].

Impedance Platelet Aggregometry. Multiple Electrode
Aggregometry (MEA) assesses platelet aggregation through the continual monitoring of alterations in electrical impedance, attributable to the activation and subsequent adherence of platelets to metal sensor electrodes across 3-5 distinct channels [12,15,16,19,[21][22][23]29].Each channel utilises a whole blood sample, with arachidonic acid added to evaluate the impact of aspirin (ASPItest), ADP for assessing the efect of P2Y12 platelet inhibitors (ADPtest), or thrombin receptor agonist peptide for measuring the impact of glycoprotein IIb/IIIa inhibitors (TRAPtest) [12,16,19,23].Additionally, collagen can be employed as an alternative to arachidonic acid for ascertaining the efect of aspirin [22].Te resultant aggregation data are presented as an arbitrary area under the curve (AUC) or expressed as an aggregation unit over time (AU x min).

Platelet Function Assay (PFA).
Te PFA-100 (Dade Behring, Germany) is a commercially accessible pointof-care platelet functionality assay that gauges platelet activation under considerable shear stress by aspirating whole blood through cartridges coated with either collagen/epinephrine (CEPI) or collagen/ADP (CADP) [11,25].Te evaluation is documented as Aperture Closure Time (CT), denoting the duration required for ensuing platelet activation to occlude the apertures within the CEPI and CADP cartridges.

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Surgery Research and Practice 3.2.4.VerifyNow Assay.Te VerifyNow system (Accumetrics, San Diego, CA, USA) represents a cartridge-based rapid assay apparatus assessing aspirin impact on platelet reactivity through the VerifyNow Aspirin Test, utilising arachidonic acid as an agonist.Conversely, the VerifyNow P2Y12 Test gauges the direct inhibition of clopidogrel on P2Y12 receptors [13,26].Aspirin test fndings are represented as Aspirin Reaction Units (ARUs), while P2Y12 test results are documented as P2Y12 Reaction Units (PRUs).

Tromboelastogram (TEG).
Heparinised whole blood is used in the TEG assay (Haemoscope Corp, Niles, IL, USA) to evaluate the platelet function in terms of clot maximum amplitude with added arachidonic acid (MA AA ) or without a platelet agonist (MA 0 ), which is compared with kaolinactivated TEG assay (MA KH ) to derive percent of platelet aggregation using the formula: %MA AA � [(MA AA − MA 0 )/ (MA KH − MA 0 )] × 100% [22].Te result is reported as a percent aggregation of platelets.

3.2.6.
Whole-Blood Flow Cytometry.Antiplatelet resistance can be quantifed via a process involving blood incubation with or devoid of arachidonic acid (1.0 mmol/L) for a duration of two minutes, followed by the addition of radiolabeled antibodies targeting CD41a or CD62P receptors on platelets.Postfxation of the samples with 1% paraformaldehyde, the ensuing analysis is conducted using a fuorescent cell sorter (Becton-Dickinson FACScan; BD Immunocytometry Systems, San Jose, CA, USA) [22].Te outcome is articulated as the percentage augmentation in the expression of the CD62P receptor following activation.
In the context of the VerifyNow system, values of aspirin reaction units (ARUs) >550 and P2Y12 reaction units (PRUs) >230 are interpreted as aspirin and clopidogrel resistance, respectively, Nevertheless, the PRU cutof point for clopidogrel resistance could be as low as 188 in certain cases [13,26,27].Aspirin resistance is defned as a collagen and/or epinephrine (CEPI) closure time of <193 seconds in the PFA-100 system [25].

Antiplatelets Used in the Studies.
All investigations incorporated aspirin as the principal antiplatelet treatment, with clopidogrel supplementing aspirin to form a dual antiplatelet therapy in several instances [15,24,26,30,31].Tough clopidogrel is invariably administered at a dosage of 75 mg, the dosage of aspirin displays variability in the range of 80-325 mg, with 100 mg being the most frequently prescribed dosage.One study employed a postoperative loading dose of intravenous aspirin of 500 mg [18].
As for TxB2 measurements, inhibition exceeding 90% was not obtained until fve days after surgery, and merely 34% of patients had platelet inhibition by this point [14,15].Inadequate inhibition of TxB2 was observed with a dosage of 100 mg aspirin, but this was not the case when the dosage was increased to 325 mg [11].Surgery Research and Practice

Surgery Research and Practice
In patients who had demonstrated aspirin resistance perioperatively, this resistance had dissipated in all instances when retested at 6-month and 12-month follow-ups [17,18,24].

Efect of Cardiopulmonary Bypass (CPB).
Te efect of CPB on aspirin resistance remains ambiguous within the literature.Platelet aggregation and thromboxane exhibit notable suppression subsequent to of-pump CABG, whereas such substantial inhibition is not observed after onpump CABG [28].A separate investigation delineates CPB duration as an independent predictor of aspirin nonresponse [23].Contrarily, several studies have asserted that CPB does not signifcantly infuence aspirin resistance [17,18].

Vein Graft Occlusion.
Antiplatelet resistance serves as a predictive factor for graft occlusion [13].Aspirin resistance, when concomitant with compromised vein graft endothelial integrity, precipitates graft thrombosis and failure within a few days post-CABG [22].Furthermore, late occlusion of vein grafts exhibits a thirteen-fold increase in risk (expressed as an odds ratio) in the presence of aspirin resistance [25].Dual antiplatelet therapy represents a potent predictor of vein graft patency and is associated with a decreased incidence of vein graft occlusion [13].

Mortality, Myocardial Infarction (MI), and Stroke.
Tere was no signifcant diference observed in mortality rates, MI, or stroke incidents between patients demonstrating aspirin resistance and those without it.One study demonstrated this during a 6-month follow-up [30,31].A further two studies demonstrated it during the 12-month follow-up periods [16,23].However, it is noteworthy to mention that all patients who died during the 12-month follow-up duration in the other two studies had previously displayed signs of aspirin resistance [17,18].
Te addition of clopidogrel to the aspirin did not result in a decrease in adverse outcomes or an increase in bleeding incidents, except in a specifc population with younger obese patients (age <65 years, BMI >30), where the incidence of adverse events was lower compared to aspirin monotherapy [30].Tis is because patients with a BMI >30, who are aspirin resistant, have worse adverse outcomes compared to those without it [31].Moreover, in patients resistant to clopidogrel and undergoing of-pump CABG, a high residual platelet reactivity is linked with elevated mortality rates, MI, and target vessel revascularisation [26].

Postoperative Immediate Blood
Loss.Te volume of postoperative blood loss 12 hours after surgery was observed to be higher in patients sensitive to preoperative aspirin in comparison to those displaying preoperative aspirin resistance, with mean volumes amounting to 555 ml and 406 ml, respectively [27].Although the chest drain output was comparable within the frst hour following surgery, a greater blood loss was recorded in the aspirin-sensitive group at both the 6-hour and 12-hour marks.Furthermore, these patients exhibited a higher risk of requiring blood transfusion in the postoperative period [19].

Discussion
True resistance to the inhibition of thromboxane A2-essentially, resistance to the biochemical efects of aspirin-is an infrequent phenomenon.Conversely, the incidence of thrombotic events and suboptimal clinical outcomes in spite of aspirin usage in patients could be attributable to an array of mechanisms extending beyond mere inhibition of the COX-1 enzyme [34].Consequently, the terminology "antiplatelet resistance" lacks a universal defnition in the literature.Nevertheless, antiplatelet resistance, when detected with in vitro platelet assays, is associated with adverse clinical outcomes in patients receiving antiplatelet therapy [35][36][37][38].
Reports of antiplatelet resistance in patients undergoing cardiac surgery vary due to difering cutof values for measurements, even when using identical assessment methodologies.Moreover, these inconsistencies are amplifed when employing disparate measurement methods.For instance, one study identifed aspirin resistance with light aggregometry when platelet aggregation was ≥20%, while others set the threshold at >30% [17,18,24].
Tese variant assessment methods yield difering results in determining antiplatelet resistance, thus compromising the comparability between studies [11].A patient labelled as resistant to antiplatelets in one study might not receive the same categorisation in another study utilising a diferent assessment method.Furthermore, the range of aspirin dosages used across individual studies might infuence the manifestation of aspirin resistance.
Moreover, despite the predominant focus on aspirin resistance in studies assessing antiplatelet resistance, there is a lack of information concerning resistance to other antiplatelet agents, such as clopidogrel.Although many assessment methods provide the capability to test clopidogrel resistance by using ADP as an alternative to arachidonic acid as a substrate, this capability is not widely employed.Although clopidogrel is commonly used in numerous studies, only Mannacio et al. reported the incidence of clopidogrel resistance [13,21,26,29,30].
Prior studies involving noncardiac surgery cohorts have shown a correlation between antiplatelet resistance and increased cardiovascular thrombotic events and mortality rates [37,39,40].While a number of randomised controlled trials and observational studies did not report a signifcant diference in adverse outcomes in cardiac surgery patients, including mortality, stroke, and myocardial infarction, these studies did not examine graft patency or patient symptoms [16,23,30,31].Te absence of diferences in adverse outcomes may be due to the potentially transient nature of aspirin resistance [17,18,24].Notably, aspirin resistance is linked to decreased blood loss in the immediate postoperative period, which could be interpreted as a prothrombotic feature when compared with the aspirinsensitive population [27].

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Surgery Research and Practice Better clinical outcomes were observed in a subset of younger (<65 years) and obese aspirin-resistant patients when dual antiplatelet therapy with clopidogrel was administered [30].Additionally, all patients who died during the follow-up period were initially identifed as having perioperative aspirin resistance [17,18].Youn et al. documented worse outcomes in patients undergoing cardiac surgery with clopidogrel resistance [26].Te assessment of this patient cohort could be further augmented with followup coronary angiograms and/or computed tomography coronary angiography.
Antiplatelet resistance is not limited to CABG surgery alone.Tis resistance contributes to poor clinical outcomes in cardiovascular disease, whether managed medically or through percutaneous coronary interventions, as well as in cerebrovascular diseases, including stroke and neurointerventional procedures [37,[41][42][43].A comprehensive meta-analysis including 2,930 patients demonstrated that antiplatelet resistance is associated with a signifcantly higher incidence of cardiovascular events (odds ratio (OR): 3.85; 95% confdence interval (CI): 3.08-4.80)and mortality (OR: 5.99; 95% CI: 2.28-15.72)[37].Consequently, individuals with antiplatelet resistance are at an elevated risk of long-term morbidity and mortality.

Limitations
Tis systematic review is subject to certain limitations.Predominantly, the studies incorporated in this review are observational cohort studies as opposed to randomised controlled trials.Due to the deployment of diverse assessment methods for antiplatelet resistance and their varied results, conducting a meta-analysis is impracticable.Further compounding this issue is the absence of a uniform defnition for antiplatelet resistance across diferent methods.Moreover, the results could potentially be swayed by various surgical techniques and vein graft handling and management, whose precise efects remain largely obscure, alongside the infuence of antiplatelet resistance.
Future investigations with adequately powered randomised controlled trials are required to explore the outcomes of antiplatelet resistance.Tis should encompass the resistance of other antiplatelet agents, thus moving beyond a narrow focus on aspirin as the main antiplatelet agent.To better understand the clinical signifcance of resistance to antiplatelet medication, more extensive imaging studies need to be undertaken to take into account the quality of the grafted coronary artery.

Conclusion
Te existing literature lacks a consistent defnition of antiplatelet resistance.Te methods used to evaluate antiplatelet resistance vary signifcantly, leading to diverse and noninterchangeable results.Although the focus of these studies predominantly rests on aspirin resistance, information regarding other antiplatelets like clopidogrel and ticagrelor remains scarce.Antiplatelet resistance in patients undergoing CABG surgery is correlated with an elevated rate of vein graft occlusion.While aspirin resistance has a conficting impact on overall adverse outcomes, the presence of clopidogrel resistance is associated with worsened outcomes in CABG patients.

Figure 1 :
Figure 1: PRISMA fow diagram for study search and selection.

Figure 2 :
Figure 2: Graphical summary of the included studies.

Table 1 :
Characteristics of the included studies.
Surgery Research and Practice cultured at 37 °C for a 24-hour duration and subsequently centrifuged at 700 × g for 15 minutes

Table 2 :
Summary of antiplatelet resistance measurement methods.