CHAPTER II REVIEW OF RELATED LTERATURE Platelets are critical in haemostasis and the development of arterial thrombi

CHAPTER II
REVIEW OF RELATED LTERATURE

Platelets are critical in haemostasis and the development of arterial thrombi. Damaged endothelium activates platelets which respond by adhering and aggregating. Their release of thromboxane A2 and adenosine diphosphate (ADP) amplifies and propagates the process by stimulating surrounding platelets. The production of thrombin via the coagulation cascade is also accelerated, stabilising the thrombus by the conversion of fibrinogen to fibrin. (Gurvinder, 2014) Platelets play a central role in cardiovascular thrombosis. Platelet aggregation caused due to a ruptured artherosclerotic plaque could eventually lead to vascular occlusion. (Philipose et al., 2010) According to Paolo Gresele, inhibition of platelet aggregation has shown great benefit for the treatment and prevention of ischaemic cardiovascular disease. Platelet inhibition can be achieved either by blockade of membrane receptors or by interaction with intracellular signalling pathways like cyclooxygenase and adenosine -5′-diphosphate pathway. While receptor antagonism may provide high specificity, the inhibition of platelet signal transduction may display broader effects, suppressing platelet activation regardless of the initial stimulus. Platelet-mediated thrombosis is a major pathophysiologic
mechanism underlying coronary thrombosis. Platelets adhere to ruptured or eroded plaques, are activated, aggregate, and release secondary messengers, which produce further thrombosis and vasoconstriction and serve as a surface for activation of the clotting cascade. Therefore,
inhibition of platelet activation or aggregation is a very effective method of preventing coronary thrombosis. The various existing antiplatelet agents, which will be explain later on, can act at different points in the platelet to inhibit the cascade of platelet activation, amplification, and aggregation. (Ducrocq et al., 2017)

Grapefruit is an hybrid between the orange and the pomelo, but no one discovered when it was first grown. It was first documented in 1750 by a Welshman in Barbados who described it as a “forbidden fruit,” according to the Purdue University horticulture department. Until the 19th century, it was also called the shaddock, named after a sea captain who is said to have brought the seeds of a pomelo to Jamaica. It has also been called paradise fruit thus giving its scientific name, Citrus paradisi. (Szalay, 2016) According to Organic facts, Citrus paradisi or in simple term, grapefruit has many health benefits because of its high fiber and low calories, also they contain bioflavonoids and other plant chemicals that protect against serious diseases like cancer, heart disease, and the formation of tumors. On the other hand, according to Top Tropicals, Psidium guajava or guava is well known throughout the tropics, many people are familiar with it because of the large number of products made from this aromatic fruit. The plant is well distributed throughout the tropics and subtropics. This is a low growing tree up to 25 feet high and about the same spread. It has been hailed as one of the super fruits due to the numerous health benefits it offers. It indeed is a powerhouse of nutrients since it can act as anti-inflammatory and even in anti platelet activity. This humble fruit is extraordinarily rich in vitamin C, lycopene and antioxidants that are beneficial for skin. (Ahuja, 2018) According to Total Health, grapefruit and guava has a high salicylate content. It was stated by different articles that guava and grapefruit has both acetylsalicylic acid or salicylates, lycopene, and flavonoids which can help to act in platelet aggregation.

Bioflavonoids or flavonoids are widely dispersed group of chemical compounds with health-related characteristics that are based on their antioxidant activity like inhibition of platelet aggregation, anticancer, anti-inflammatory and antiviral activities. (Mallick et al., 2016) There are three kinds of flavonoids present in citrus fruits particularly in Citrus paradise and these are flavanones, flavones and anthocyanins. The four main flavanones are naringenin, isosakuranetin, eriodictyol and hesperetin. Flavanones are very numerous thus making a major contribution to taste properties of citrus fruits but as in the case of all flavonoids, some are desirable and some are undesirable. (Adeel et al., 2015) The role of flavonoids has been studied to prevent and treat cardiovascular diseases due to several reasons including antioxidant and anti-inflammatory activity, as increase dietary intake of antioxidants may prevent atherosclerosis, since increased cholesterol and occlusion is correlated. (Mallick et al., 2016) Citrus flavonoids may also exert neuroprotective effects since they are involved in the modulation of neuronal activities and mental health including brain plasticity, behaviour, mood, depression, and cognition. In this regard, it has been demonstrated that hesperidin can protect neurons against various types of insults associated with many neurodegenerative diseases. Also, naringin has proven to exert neuroprotective effects through anti-inflammatory activity on the survival of dopaminergic neurons and on the integrity of the nigrostriatal pathway in animal models of Parkinson’s disease. Natural flavonoids would therefore seem to have important potential as medicaments in the field of mental health, although their use in clinical practice is still a long way off. (Vazquez et al., 2016)

Lycopene, a red plant pigment, is found in number of fruits such as tomatoes, watermelon, papaya, and guava. Of these, tomatoes contribute the largest to the dietary intake of lycopene. Several studies have demonstrated the antiplatelet activity of tomato in vitro as well as the ability of tomato extracts to reduce ex vivo platelet aggregation.The antiplatelet activity of lycopene has not been compared to any of the clinically used antiplatelet agents until now. Moreover, the influence of lycopene on the action of routinely used antiplatelet agents has not been studied. (Sawardekar et al., 2016) Lycopene has demonstrated hypotensive activity in humans (Engelhard 2006), and several human trials indicate a cholesterol-lowering effect (Ried 2011). One mechanism by which lycopene may limit platelet aggregation is by activating cyclic-GMP, a signaling molecule involved in vessel dilation and involvement in inhibition of phosphodiesterase pathway. Two studies examined the effects on platelet function in healthy human volunteers: High dose and low dose and the results show that both exhibited significant reductions in platelet aggregation up to 6 hours after ingestion. Standardized bioactives suppress platelet adhesion and aggregation by reversibly inhibiting P-selectin and GPIIb/IIIa, two receptors necessary for clot formation. (Kennedy, 2006)

Aspirin belongs to a class of medications called nonsteroidal anti-inflammatory drugs (NSAIDs). Aspirin and other NSAIDs, for example, ibuprofen and naproxen are widely used to treat fever, pain, and inflammatory conditions such as arthritis, tendonitis, and bursitis. Aspirin is known chemically as acetyl salicylic acid and often abbreviated as ASA. Aspirin also has an important inhibitory effect on platelets in the blood. This antiplatelet effect is used to prevent blood clot that promote the clotting of blood inside arteries, particularly in individuals who have atherosclerosis (narrowing of the arteries) or are otherwise prone to develop blood clots in their arteries. (Ogbru et al., 2016) It is the most widely use analgesic, antipyretic, and anti-inflammatory agent in the world and remains the standard for which all other NSAIDs are compared. Aspirin is comprised of the active compounds acetic acid and salicylic acid, forming acetylsalicylic acid. Aspirin inhibits the biosynthesis of prostaglandins by means of an irreversible acetylation and consequent inactivation of COX; thus, aspirin inactivates COX permanently. This is an important distinction among the NSAIDs because aspirin’s duration action is related to the turnover rate of cyclooxygenases in various target tissues. The duration of action of other NSAIDs, which competitively inhibit the active sites of the COX enzymes, relates more directly to the time course of drug disposition. Because platelets are devoid of the ability to produce additional cyclooxygenase, thromboxane synthesis is arrested. (Williams et al., 2011) Aspirin impairs platelet aggregation usually attributed to inhibition of cyclooxygenase-1(COX-1) and subsequent reduction in thromboxane A2 production, although other mechanisms have been implicated. A dose of 81 mg of aspirin daily can permanently affect platelets in circulation, such that the length of drug effect is related to platelet lifespan, not drug half life. Other NSAIDs impair thromboxane A2 production as well but the effects are related to drug half life. (Lanzkowsky, 2011) Hence, aspirin is the cornerstone of antiplatelet therapy and is the most commonly used antiplatelet agent for prophylaxis of thrombotic vascular disorders. (Sawardekar et al., 2016)

Platelet adhesion, activation and aggregation to the injured vessel wall are crucially involved in the pathogenesis of thrombus formation. (Jing et al., 2013) The antiplatelet agents in current clinical use include the cyclooxygenase inhibitors (e.g., aspirin), phosphodiesterase inhibitors (e.g., dipyridamole), adenosine-5′-diphosphate (ADP) receptor pathway inhibitors (e.g., ticlopidine and clopidogrel), and platelet glycoprotein IIb/IIIa receptor antagonists (e.g., abciximab, tirofiban, and eptifibatide). (Sawardekar et al., 2016) Cyclooxygenase-1 (COX-1) catalyses the transformation of arachidonic acid to the unstable intermediate prostaglandin PGH2. Arachidonic acid is essential for the brain, liver, and organs, according to the National institutes of Health. A study published in 2013 in the American Journal of Cell Physiology reports that arachidonic acid appears to help increase muscle mass, and a study published in 2012 in the Journal of Clinical Psychopharmacology found that arachidonic acid supplementation, when combined with docosahexaenoic acid (DHA), helps improve social interaction in people with autism spectrum disorders. Subsequently, thromboxane synthase acts on PGH2 to form TXA2, a transient biological product that induces platelet aggregation and is a powerful vasoconstrictor. As stated, aspirin acts primarily by interfering with the biosynthesis of cyclic prostanoids: TXA2, prostacyclin, and other prostaglandins. It irreversibly inhibits COX-1 by acetylation of serine-530 and induces a long-lasting functional defect in the platelets. The resultant decrease in production of prostaglandins and TXA2 probably accounts for much of aspirin’s antithrombotic effect.The plasma half-life of aspirin is only 20 minutes in circulating blood. Salicylate does not affect COX-1 or COX-2 activity. (Altman et al., 2004) Phosphodiesterases (PDEs), by catalysing the hydrolysis of cAMP and cGMP to inactive 5?-AMP and 5?-GMP, limit intracellular levels of cyclic nucleotides and thus regulate the amplitude, duration and compartmentation of cyclic nucleotide signalling. Functionally, phosphodiesterases can be classified in terms of their affinity and rates of degradation for cAMP and cGMP. To date, more than 60 different isoforms of PDE have been described in mammalian tissues, grouped into 11 broad families (PDE1–PDE11) based on differences in their structure, kinetics, regulatory properties and sensitivity to chemical inhibitors. Alternative splicing and transcription start sites also contribute to the multiplicity of the different isoforms, many of which possess species-specific tissue and/or cellular distribution. Current data suggest that individual isozymes modulate distinct regulatory pathways in cells. Dipyridamole was synthesized more than 50 years ago and initially used as a coronary vasodilator.The antithrombotic effects of dipyridamole were evaluated in several animal models. Thrombus formation on air-injured carotid arteries of rabbits was reduced by dipyridamole, while salicylate increased it. In rabbits, the accumulation of radioactive fibrinogen at balloon angioplasty-treated carotid arteries was significantly reduced by dipyridamole in comparison to heparin. (Gresele et al., 2011) Two researchers from Journal of Clinical and Medical Genomics, Fang Jing and Wei Zhang stated that platelet aggregation is basically involved in pathogenesis of thrombus formation and the formation of a hemostatic thrombus during blood vessel injury is a highly regulated event which ensures that a blood clot is sufficiently stable but not overly robust to cause thrombus propagation and vessel occlusion. (Owaynat et al., 2013) Adenosine diphosphate (ADP) is one of the main platelet-activating factors. Platelet ADP signaling pathways are mediated by the P2Y1 and P2Y12 receptors, which play a key role in platelet activation and aggregation processes. The P2Y1 and P2Y12 are G-coupled receptors and are both required for platelet aggregation. However, ADP-stimulated effects are mediated mainly by P2Y12receptor activation, which leads to sustained platelet aggregation and stabilization of the platelet aggregate. The P2Y1 receptor is responsible for an initial weak and transient phase of platelet aggregation and change in platelet shape. Also, the cytochrome P450 enzymes are responsible for the conversion of clopidogrel into its active metabolite and the metabolism of proton pump inhibitors (PPIs), which may also inhibit CYP enzymes. (Przespolewski et al., 2018). Clopidogrel is currently the most commonly used P2Y12 receptor inhibitor. It has a more favorable safety profile compared with ticlopidine, as first shown in the CLASSICS (The Clopidogrel Aspirin Stent International Cooperative Study trial). However, its main limitation is the broad interindividual variability in its antiplatelet effects. (Angiolillo et al.) According to NHS inform, clopidogrel is an antiplatelet medicine. This means it reduces the risk of blood clots forming. Normally, when there is a cut or break in a small blood vessel, a blood clot forms to plug the hole until the blood vessel heals. Small cells in the blood called platelets cause the blood to clot. When a platelet detects a damaged area of a blood vessel, it produces a chemical that attracts other platelets and makes them stick together to form a blood clot. It reduces the ability of the platelets to stick together and reduces the risk of clots forming. This protects you from having a stroke or heart attack. Influencing factors for clopidogrel-mediated platelet inhibition have only been evaluated by one or two different test systems in the same population so far. Since previous studies revealed poor correlations between the various platelet function tests, the identification of influencing variables for clopidogrel response may vary from one test system to the next. We therefore investigated whether the influencing factors for clopidogrel-mediated platelet inhibition depend on the used assay. (Gremmel et al., 2011) Glycoprotein (GP) IIb/IIIa antagonists are a new class of potent drugs that profoundly inhibit platelet function by blocking the key receptor involved in platelet aggregation. (Ferguson et al., 2012) The glycoprotein (GP) IIb/IIIa receptor mediates the final common pathway of platelet aggregation. In particular, the GP IIb/IIIa receptor is a heterodimer consisting of the alpha IIb and beta3-subunits. By competing with fibrinogen and von Willebrand factor (vWF) for GP IIb/IIIa binding, GP IIb/IIIa antagonists interfere with platelet cross-linking and platelet-derived thrombus formation. Therefore agents blocking the GP IIb/IIIa receptor are very potent platelet inhibitors.

Numerous clinical trials have been conducted to define the optimal use of GP IIb/IIIa inhibitors. The number of GPIIb/IIIa receptors occupied by the drugs was assayed by flow cytometry using two CD61 surface fluorescein antibodies. (Zaragosa et al., 2016) Currently, GP IIb/IIIa inhibitors are mostly indicated for patients undergoing percutaneous coronary interventions (PCI) in the setting of acute coronary syndrome. (Angiolillo et al.) It is a member of the ?3 subfamily of integrins, which also includes ?V?3. GPIIb/IIIa functions as a receptor for fibrinogen and several adhesion proteins sharing an arginine-glycine-aspartic acid (RGD) sequence. GPIIb/IIIa antagonists, through blockade of the receptor, prevent platelet aggregation. Among the three GPIIb/IIIa antagonists used in therapy, abciximab is an anti-?3 monoclonal antibody, while tirofiban and eptifibatide mimic the binding sequence of the fibrinogen ligand. The use of inhibitors of glycoprotein IIb/IIIa (GPIIb/IIIa) has provided dramatic results in terms of the prevention of acute stent thrombosis and a reduction in major adverse coronary events in patients subjected to percutaneous coronary intervention. (Giordano et al., 2015) Platelets have a central role in thrombus formation. At the cellular level, thrombosis is initiated by platelets tethering to subendothelial von Willebrand factor (vWF) via the glycoproteinIb. GPIb?- vWF interactions mediates the initial adhesion step of platelets to the extracellular matrix (ECM) at high shear rates (;500). GPIb? may also contribute to platelet adhesion to the intact vessel wall by interacting with P-selectin exposed on activated endothelial cells. (Jing et al., 2013).

Now that inhibitors of anti platelet agents were fully understood and the components of Citrus paradisi and Psidium guajava that can affect the anti platelet activity were already explained, the researchers aim to compare the effect of both fruit, guava and grapefruit, as platelet inhibitors since it was stated that they contain the same composition like the presence of flavonoids or lycopene. Since aspirin is the major antiplatelet drug, everyone should know that there’s an adverse effect of taking too much aspirin to human body can cause severe pain in the stomach, fatigue, fever, rapid breathing, nausea, vomiting, dizziness, cloudy mind, seizures, hyperactivity, inability to hear and it worst cases it can lead to coma.. According to John Horn and Philip Hansten which are both Doctor of Pharmacy, aspirin must be taken in low-dose because it can prevent and help for myocardial infarction and other cardiovascular diseases, but there is evidence that concurrent use of NSAIDs may inhibit the antiplatelet effect of aspirin. It stated that antiplatelet drugs like aspirin has both advantages and disadvanatges thus, having an alternative to antiplatelet like using natural products instead of those chemicals can be beneficial for the people who are highly need of it. Today, there is increasing demand for natural bioactive compounds as people express more concern about their health, especially in connection with health-giving diets.(Vazquez et al., 2016) Many intake of drugs in the body may cause disease or deficiency since human body cannot process all those drugs thus using fruits or leaves particularly guava and grapefruit may help or contribute to the inhibition of platelet. Antiplatelet drugs should be of proven efficacy in the secondary prevention of ischemic events without incurring an unacceptable increase in the risk of bleeding. (Gachet, 2015)

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