key: cord-1006067-2sck9dph
authors: Maiti, Smarajit; Banerjee, Amrita; Kanwar, Mehak
title: In silico Nigellidine (N. sativa) bind to viral spike/active-sites of ACE1/2, AT1/2 to prevent COVID-19 induced vaso-tumult/vascular-damage/comorbidity
date: 2021-03-18
journal: Vascul Pharmacol
DOI: 10.1016/j.vph.2021.106856
sha: 761232bbcd50ebeecdf0d84f8c07a143ae8a7c39
doc_id: 1006067
cord_uid: 2sck9dph

COVID-19, a global-pandemic binds human-lung-ACE2. ACE2 causes vasodilatation. ACE2 works in balance with ACE1. The vaso-status maintains blood-pressure/vascular-health which is demolished in Covid-19 manifesting aldosterone/salt-deregulations/inflammations/endothelial-dysfunctions/hyper-hypo- tension, sepsis/hypovolemic-shock and vessel-thrombosis/coagulations. Here, nigellidine, an indazole-alkaloid was analyzed by molecular-docking for binding to different Angiotensin-binding-proteins (enzymes, ACE1(6en5)/ACE2(4aph)/receptors, AT1(6os1)/AT2(5xjm)) and COVID-19 spike-glycoprotein(6vsb). Nigellidine strongly binds to the spike-protein at the hinge-region/active-site-opening which may hamper proper-binding of nCoV2-ACE2 surface. Nigellidine effectively binds in the Angiotensin- II binding-site/entry-pocket (−7.54 kcal/mol, −211.76, Atomic-Contact-Energy; ACE-value) of ACE2 (Ki 8.68 and 8.3 μmol) in comparison to known-binder EGCG (−4.53) and Theaflavin-di-gallate (−2.85). Nigellidine showed strong-binding (Ki, 50.93 μmol/binding-energy −5.48 kcal/mol) to mono/multi-meric ACE1. Moreover, it binds Angiotensin-receptors, AT1/AT2 (Ki, 42.79/14.22 μmol, binding-energy, −5.96/−6.61 kcal/mol) at active-sites, respectively. This article reports the novel binding of nigellidine and subsequent blockage of angiotensin-binding proteins. The ACEs-blocking could restore Angiotensin-level, restrict vaso-turbulence in Covid patients and receptor-blocking might stop inflammatory/vascular impairment. Nigellidine may slowdown the vaso-fluctuations due to Angiotensin-deregulations in Covid patients. Angiotensin II-ACE2 binding (ACE-value −294.81) is more favorable than nigellidine-ACE2. Conversely, nigellidine-ACE1 binding-energy/Ki is lower than nigellidine-ACE2 values indicating a balanced-state between constriction-dilatation. Moreover, nigellidine binds to the viral-spike, closer-proximity to its ACE2 binding-domain. Taken together, Covid patients/elderly-patients, comorbid-patients (with hypertensive/diabetic/cardiac/renal-impairment, counting >80% of non-survivors) could be greatly benefited.

At the initial phase of infection of SARS CoV2, ACE2 is occupied by the peripheral spike glycoprotein of the virus which is then internalized. In this situation angiotensin II level become high so the blood pressure increases and in the hypertensive individuals this situation becomes even more detrimental for the drastic pressure increase. In elderly persons it may results in cerebral, cardiac and renal vascular failure and death [1, 2] .But, in the post infection stage and after few cycles of viral generations (considering its extreme high rate of propagation) SARS CoV2 stimulatesACE2 J o u r n a l P r e -p r o o f expression on different tissues including the vast beds of arterial endothelial cells and even in several tissue surfaces other than lung [3, 4] . It makes the system more vulnerable to viral infection.

At the same time extra ACE2 utilizes more angiotensin II to form angiotensin 1-7 peptides which may results in sudden vasodilatation (creating vasogenic or vasoplegic shock,) and hypovolemic shock in association with cardiogenic shock (3) . In addition, in advanced viral infection state sepsis shock in response to cytokine storm makes the situation worsen. So, the persons already having an infection/inflammation history with hypertensive/vascular disorder both or alone become extremely vulnerable and morbid in nature [5] . The present status of death versus infection of countrywise/global ratios and elderly (having more possibility several disorders/aging processes) persons decisively suggests the comorbidity situation is the strongest determinant of the disease severity and death.

As explained in H. Li, L. Liu, D. , low infection, moderate infection and severe infection can be represented by no co-morbidity, less number of co-morbidity and more number of co-morbidity situations respectively. Renin-angiotensin system, aldosterone and other steroid regulations are the major and direct determinant of blood pressure and vascular health in an individual. Disorder in the peripheral vasculature may impair three organ-associated vascular systems; namely cardio-vascular, reno-vascular and cerebro-vascular systems. These are also the main target of the age and possible gender associated anomalies [7, 8] . Age related impairment of the immune system has been reported [9, 1] . Moreover, it has been recently demonstrated that SARS-CoV-2 can infect podocytes and tubular epithelial cells of the kidney [10] . Taken together, comorbidity is the main factor and possibly can explain the gap between numbers of deaths versus infection in different geographical locations.

Angiotensin II being the main factor of pressure regulation and ACE2 being the specific target of spike glycoprotein of SARS CoV-2, the upstream and downstream regulations of angiotensin II metabolism and functions could best explain about the co-morbidity situations and eventually the therapeutic targeting of this regulatory pathway may protect from the severity of the co-morbidity situation. In addition to the binding affinity between viral spikes to ACE2, only ACE2 expression can be a disease severity factor. So targeting the spike protein is also great therapeutic approach. Earlier finding suggests that tea flavonoids like Epigallocatechin and theaflavin gallate may block the central channel of the spike protein [11] . In the current study we showed that nigellidine In classical physiology, it is well-known that angiotensinogen is converted to angiotensin I which is further converted to angiotensin II with the help of ACE. Angiotensin II is a vasoconstriction agent. Now the ACE2 can convert the angiotensin II to angiotensin (1-7) peptides. These peptides are altogether vasodilatation agents. ACE2 performs pivotally counter-balancing to ACE by catalyzing angiotensin II which is the key regulator in the renin-angiotensin-aldosterone system. In case of lower function of ACE2, angiotensin II remain high and performs as the ligand function to AT1 and AT2 surface receptors of several tissues. Binding to AT1 has a strong vasoconstriction function and that is opposite when the binding is on AT2. Other than vaso-status determination, this bunch of peptides and their target enzymes/receptors have other function like inflammatory, gene expressions, signal transductions function. Those states can perform in multi-phasic manner ending up with good or bad outcome. Very often in case of impairment of this tight regulation angiotensin II may act as a double sided sword. So, the up and down stream regulations of Angiotensin II functions which is targeted by SARS CoV-2 has been screened. Here, the focus has been pointed on possible drug targets to this system to forbid the co-morbid situation during this viral infection. Our study suggests that nigellidine an indazole alkaloid strongly resists the SARS Cov-2 associated ACE2 deregulations at its different regulatory steps with most balanced outcome. It is proposed that medications which inhibit the RAS ACE-dependent pathway may be beneficial in treating COVID-19 [12, 13] . The basal level of Ang II (basal constriction state) is depleted. It is called post sepsis viral shock, hypotonic/hypovolemic shock. These extra ACE2 may bind with the nigellidine and reduce extra Angiotensin II breakdown. It will maintain normal physiological vaso-state (constriction/dilatation state).

In this background, our objectivity was to screen a suitable molecule which can effectively bind to the angiotensin binding proteins (enzymes ACEs and receptors ATs) and as a whole downregulates Angiotensin II mediated vaso-functions to restore the normal vaso-state and vascularinflammatory activities. From a bunch of molecules tested, we found that nigellidine can effectively perform this function which could be helpful in the comorbid and other Covid patients. Moreover, nigellidine was found to bind in the viral spike glycoprotein that is close proximity of RBD and ACE2.

For the first time, we showed that nigellidine can bind to the active binding locations of several angiotensin binding proteins. This finding may have great clinical implications in the present pandemic situation of Covid infection. Further studies are necessary in this regard.

Protein tertiary structure retrieval J o u r n a l P r e -p r o o f Journal Pre-proof Present study has represented the interaction pattern of nigellidine with different Angiotensin receptors like Angiotensin Converting Enzyme 1 (ACE1), Angiotensin Converting Enzyme 2 (ACE2), AT1 and AT2 and COVID 19 spike glycoprotein. The 3D structures of these receptors were retrieved from Protein Data Bank (PDB) in .pdb file written in flat file format (https://www.rcsb.org/). That PDB ID, used in this study were 6en5, 4aph, 6os1, 5xjm and 6vsb representing ACE1, ACE2, AT1, AT2 and COVID 19 spike glycoprotein respectively.

The selected PDB structures were found with different molecules like H 2 O, NAG, BAM etc. including Angiotensin molecules in Angiotensin receptors. Whereas, COVID 19 spike glycoprotein showed with multiple NAG unit at different position. So, for molecular docking study different attached molecules were removed from the receptor molecules using Pymol molecular visualization software [14] . After removal, receptor molecules were saved in .pdb file for further analysis.

The ligand molecule, nigellidine is a rare indazole-type alkaloid also known as 6,7,8,9- Where T[s,t] is the pre-calculated value of the atom pair s and t and |s−t| is representing the Euclidean distance between s and t. The more favourable ACE value is the lower value which indicated the rigid complex formation between receptor and ligand. Here in this study nigellidine was docked with all the selected Angiotensin receptors and COVID 19 spike glycoproteins and analysed using ACE value.

The molecular docking study between each Angiotensin receptors, spike glycoprotein with ligand nigellidine was again performed using Autodock software. Autodock is an automated docking tool for small molecule docking upon known receptors 3D structure and explores the ligands atomic affinity grids. Autodock have application in structure-based drug designing, lead optimization, X-ray crystallography or NMR structure analysis, virtual screening (HTS), combinational library design, protein-protein docking and chemical mechanism study. Autodock analyze the receptor-ligand docking from different angle through Binding energy, Ligand efficiency, Intermolecular energy, Desolvation energy, Electrostatic energy, Total energy, Torsion energy and Inhibition constant. The 

Each sets of molecular docking were analyzed using Pymol molecular visualization software [14] . The presentation of docking structures and interactive bonds were represented through technical and transparent surface analysis mainly. On the other hand the 2D interactions were analyzed through LIGPLOT software [17] . LIGPLOT basically reads the ligand binding sites of 3D structure in PDB format and then it unrolls the positions according to their rotatable bonds and becomes flattening then in 2D form.

To understand the binding between different docked molecules, each pairs were individually subjected to PRODIGY tool on HADDOCK server. Binding affinity analysis were analyzed between AT1-Nigellidine, AT2-Nigellidine, ACE1-Nigellidine, ACE1-Nigellidine, AT1-Angiotensin, AT2-Angiotensin, ACE1-Angiotensin, ACE1-Angiotensin.

The nCoV is denoted as the spike glycoprotein of novel coronavirus or SARS CoV2. This protein is 

Angiotensin Converting Enzyme 2 are usually found as the outer surface protein of different cell types like lungs, arteries, heart, kidney and intestines. ACE2 acts on angiotensin II, an 8 amino acid long vasoconstrictor peptide, to convert it into angiotensin (1 -7). These converted angiotensin function as vasodilator. Thus ACE2 controls high blood pressure. In present study ACE2 was analysed for angiotensin II and nigellidine binding. As per the 3D structure of ACE2 it possesses an internal channel of angiotensin binding (Fig. 2) . ACE2 functions as a carboxypeptidase and it cleaves a single residue from angiotensin I (AngI). That finally generates Ang1-9, and a single residue from angiotensin II (AngII) to generate Ang1-7 [18] .

Angiotensin II is the main ligand of ACE2. As a result of which we also found the -9. with a value of -11.6 ΔG (kcal mol-1) and it also showed some dissociation constant value of 6.20E-09. Whereas, molecular docking of ACE2 with nigellidine showed the lower binding energy of -7.54 and -6.73 kcal/mole (S Table 1 ), lower binding efficiency of -5.6 ΔG (kcal mol-1) at the angiotensin binding site (S Table 3 ). But no dissociation constant value was observed for ACE2-nigellidine interaction. And the complete internal channel was found to block by the various posture of nigellidine (S Figure 2) . Even if the accumulation of nigellidine at the exit site of angiotensin was maximum. Nigellidine was also found to block the entry pocket of ACE2 with highest affinity and low binding energy value of -7.54 and -7.52 kcal/mol ( fig. 2 ). Though the binding energy of nigellidine (-7.54 and -6.73) and the binding affinity (-5.6 ΔG (kcal mol-1)) were lower than angiotensin binding, it was found as a potent inhibitor of ACE2. Ligand efficiency represents the binding energy of per atom of ligand to its receptor [20] .

Here Table 1 ). In relation to that intermolecular energy, representing the lower barriers between two docked structures [21] , also supported the rigid binding with negative value of -6.56, (-7. very less or nun error value [24] . From all aspects, nigellidine binding to ACE1, ACE2, AT1 and AT2

were non-erroneous and facilitate from all significant aspects. 

ACE1 acts on Angiotensin I and converts it into Angiotensin II. This is being converted to Angiotensin 1-17 by ACE2. And thus the blood pressure of the body is being regulated. Though becoming functionally different the sequence of two proteins shares common amino acid combination and positions at maximum sites (S- fig. 1 ). Some major amino acid changes were also observed within two sequences. Whereas, the sequential diversity imprint vary less in structural diversity (data not shown).

AT1 and AT2 both were classified as the Angiotensin receptor. Though they are different subtypes they shares very close relation in respect to protein sequences and tertiary structural conformation (S- fig. 1 ). Both were mainly composed of alpha helix. In present study, AT1 (PDB ID:6OS1) and AT2 (PDB ID:5XJM) were analyzed for nigellidine binding affinity.

In AT1, Angiotensin binds to site 1 according to PDB structure 6OS1 (Fig.4a) . But another location (site 2) of Angiotensin binding was also observed during molecular docking study of AT1 and

Angiotensin. At site1, amino acids of angiotensin TYR 4, LYS 5, HIS6, ARG2 and ALA8 formed bonds with PHE182, LYS199, TYR87, ASP1281, ASP1281 and LYS199 amino acids of AT1. During nigellidine docking, remarkable atomic contact energy value was observed at those two sites of Angiotensin binding (Fig. 4b) . Highest ACE value of -227.09 was observed at site 2. According to autodock analysis also the site 2 was found with highest affinity and binding energy value of -7.12. Whereas, most of the nigellidine interactions were observed at site one. At site 1 maximum ACE value of -129.61 was found to block ASP1281, with which HIS6 molecule of Angiotensin usually binds (Fig. 4b) . Other stable bond was formed with ARG167 and nearby amino acids were CYS180, TRP84, ALA21, ASP16, VAL179, TYR92 and TYR87. Among them LYS5 of Angiotensin usually interact with TYR87. According to autodock, out of 10 best positions of nigellidine only one was found at site 1 with binding energy value -5.96 kcal/mol and Ki of 42.79 µmol. Where, nigellidine formed bond with TYR184 (Fig.4c) which indirectly hampered the binding of Angiotensin TYR4 with PHE182 (Fig. 4d) .

Likewise in AT2, site 1 and site 2 were observed with Angiotensin binding (Fig. 5a) . According to PDB structure 5XJM, Angiotensin was found at site 1. Highest ACE value (-235.57) of nigellidine binding was at site 2. Maximum affinity of nigellidine binding was at site 1 (Fig. 5b) . Nigellidine with J o u r n a l P r e -p r o o f ACE value -116.91 was found to interact with ARG182, the usual amino acid in Angiotensin-AT2

interaction. The other stable bond was found with MET128 and nearby amino acids were THR125, LEU124, ILE211, MET197, LYS215, PHE272 and TRP269. Among which LYS215 was another Angiotensin binding site. According to autodock, highest affinity of nigellidine was observed at site 1 with binding energy value of -6.61 kcal/mol (Fig. 5c) . Other two postures of nigellidine with binding energy value of -6.51 and -6.46 kcal/mol were also found there (S-table1). Nigellidine was found to compete with Angiotensin upon binding with ARG182. On the other hand it also hampered the Angiotensin -LYS215 binding. At this location, the Ki values were also lower (14.22, 16.79 & 18.36 µmol) than AT1-nigellidine binding (42.79 µmol). So, from the above analysis it could be concluded that AT2 is more prone to nigellidine then AT1.

In the present study, different Angiotensin receptors were analyzed for nigellidine binding or inhibition criterion. Where nigellidine was found to block different amino acids at which Angiotensin usually binds. Here the Angiotensin binding amino acids at different receptors like AT1, AT2 and ACE2 were analyzed ( Figure 6 ). The angiotensin attached in the NMR structure of ACE2 was of 8 amino acids (DRVYIHPF). Whereas, Angiotensin attached with AT1 and AT2 were of 7 amino acids i.e. RVYKHPA and RVYIHPI respectively. The sequential differences between AT1 and AT2 were 

The patients of sepsis manifested the characteristics for septic shock and finally sepsis according to the Sepsis-3 International Consensus. Greatly enhanced immunological responses and J o u r n a l P r e -p r o o f severe inflammatory responses are responsible for these conditions [26] . Observations that were with negative for bacterial and fungal infections in 76% sepsis patients in a COVID-19 cohort suggests that post infection immunological and other physiological manifestations are mostly due to this virus [5] . In this condition, understanding the mechanism of viral sepsis is required in SARS CoV-2 patients. Alterations of the vaso-state function due to severe influence on ACE-angiotensinaldosterone regulations generate strong pathology especially in the elderly comorbid patients.

Moreover, drastic increase in pro-inflammatory factors like TNFα, IL-1β, IL-6, granulocyte-colony stimulating factor, IFN-ϒ worsens the situations significantly in those patients. [27, 28] . Report suggests that SARS-CoV-2 could exploit species-specific interferon-driven upregulation of ACE2, a tissue-protective mediator during lung injury, to enhance infection [29] .

Viral RNA isolated from the T lymphocytes isolated from peripheral blood and some tissues like spleen, lymph-nodes other various organs suggests ACE might be expressed in these tissues and can cause the direct viral infection. This adds extra burden of ACE-angiotensin deregulations. SJ [30, 31] . Septic hypotension after COVID effects due to higher expression of ACE2 in the large endothelial bed of the vascular/circulatory system raise the questions of using ACE inhibitor therapy with angiotensin II receptor blockers in these patients. Several researchers argued in favour of or against this hypothesis. Some explained that this therapy decrease the inflammation but other analyzed that ACE-expression would increase in this situation and would favour more viral entry SH [32] [33] [34] . Further research is needed to explore whether these drugs inhibit or aid viral entry.

Report suggests that combination dosage with drugs like ACE inhibitor enalapril and the angiotensin II receptor blocker candesartan might have some beneficial effects in the Covid infected patients in the patients with Chronic Pulmonary Heart Disease [35] . Ang II binding receptors AT 1 and 2 has a great implication in the patho-physiology of Covid outcome. AT1 is present in endothelial cells, leukocytes and platelets. In endothelial cells, Angiotensin binding to this receptor induces NADPH dependant oxidative stress that in one hand induce both ATs dependant proinflammatory cytokines induction. On the other hand the oxidative stress induces AP1 regulated Eand P selectin [36] . This process is supported by CD11/18 operated induction in the endothelial cell also via AT1.

Endothelial cellular activation of oxidative stress is also occurring via AT1 dependant platelet activation of CD40/CD40L signalling. This may results in wide spread thrombotic functions in the blood vessels which is noticed in the large number of Covid patients and also in the autopsy studies [37, 38] . AT2 receptor acts like a double sided sword. In a very narrow window it antagonizes angiotensin-induced AT1 functions. Though AT1 induced vasoconstriction may be outperformed by the AT2 regulated vasodilatation but in several conditions over-activation of AT2 may results in severe hypotonic shock [40] . Moreover, AT2 bound with angiotensin induces smooth muscle cell apoptosis via GATA-6 activation and FasL-Fas involvement [41] . GATA activation by MEK/ERK1/2 may result in growth and differentiation [41] and the Fas signalling may activate Caspase 8 and result in extrinsic apoptosis [42] . All these could have been occurring in the current situation in patients of severe Covid condition. Our present study suggests that nigellidine has an affinity for binding to the AT2-angiotensin binding site and may terminate this series of adverse events. AT2 activation may also stimulate bradikynin and NO dependant cGMP signalling [43] . And phospholipase A 2 dependant arachidonic acid production and K+ release from the cells that result in hyperpolarization and decreased excitability [44] .

Initially it was surprising to us that how a single molecule can bind effectively in so many . 1 ). But during high viral load or a higher rate of viral replication, over dose of nigellidine will control the sudden fluctuation of blood pressure and spike binding to ACE2. Nigellidine mainly act as the inhibitor of angiotensine conversion by ACE2 or inhibits the carboxypeptidase activity of ACE2 which ultimately controls the vasodilatation and may control blood pressure fluctuation.

In our current study, we were trying to screen a drug which has wide range of affinity towards the angiotensin binding proteins (enzymes and receptors). So that it can block the proteins and arrest the angiotensin II enzymatic and receptor signalling at the sub threshold levels (figure 7).

In Covid patients, the angiotensin II induced vaso-dysfunction and impaired vascular events will be strictly restored. Not only that, Angiotensin II and its receptor AT1/2-mediated downstream inflammatory signalling will also be blocked. Here, for the first time we demonstrated that nigellidine an indazole alkaloid strongly bind to the ACE2 and then ACE1 enzymes. Moreover, it can block the AT1/2 receptors. During attachment with ACE2 receptor it becomes exposed. Nigellidine bound at the hinge region which controls the flexibility of spike glycoprotein attachment site.

Nigellidine and ACE2 binding was presented. Angiotensine usually bound to ACE2 at an internal channel. Nigellidine was also found to block that internal channel. Nigellidine was also found to block the entry pocket of Angiotensine within ACE2. Figure 3 .Nigellidine ACE mono-meric binding. Nigellidine was usually bound to ACE1 at its internal channel and outer surface of the ACE1 also. The active site location of ACE1 was represented which interacted with D7 and S8. The involved common amino acids were also blocked by nigellidine like ALA332. In silico Nigellidine (N. sativa) bind to viral sp ACE1/2, AT1/2 to prevent COVID-19 induced tumult/vascular-damage/comorbidity

Are patients with hypertension and diabetes mellitus at increased risk for COVID-19 infection?

Hypertension, the renin-angiotensin system, and the risk of lower respiratory tract infections and lung injury: implications for COVID-19

Is the ACE2 Overexpression a Risk Factor for COVID-19 Infection?

Clinical characteristics of coronavirus disease 2019 in China

Clinical course and risk factors for mortality of adult in patients with COVID-19 in Wuhan, China: a retrospective cohort study

SARS-CoV-2 and viral sepsis: observations and hypotheses

Angiotensin-converting enzyme-2 (ACE2), SARS-CoV-2 and pathophysiology of coronavirus disease 2019 (COVID-19)

Role of age, comorbidity and renin-angiotensin-aldosterone system in COVID-19. Effects of ACE inhibitors and angiotensin receptor blockersKardiologiia

Changesin primary lymphoid organs with aging

Is the kidney a target of SARS-CoV-2?

Epigallocatechin-Gallate and Theaflavin-Gallate Interaction in SARS CoV-2 Spike-Protein Central-Channel with Reference to the Hydroxychloroquine Interaction: Bioinformatics and Molecular Docking Study

The renin-angiotensin system -a therapeutic target in COVID-19?

Urgent need for evaluating agonists of Angiotensin-(1-7)/ Mas receptor axis for treatment of patients with COVID-19

Pymol: An open-source molecular graphics tool

Taking geometry to its edge: fast unbound rigid (and hinge-bent) docking

Protein-protein binding site identification by enumerating the configurations

LIGPLOT: a program to generate schematic diagrams of protein-ligand interactions

A crucial role of angiotensin converting enzyme 2 (ACE2) in SARS coronavirus-induced lung injury

Epigallocatechin-3-gallate inhibits human angiotensin-converting enzyme activity through an autoxidationdependent mechanism

The maximal affinity of ligands

Tipping the Scales: Spectroscopic Tools for Intermolecular Energy Balances

Electric Potential

Electricity and Magnetism

Another look at measures of forecast accuracy

Liver injury in COVID-19: management and challenges

The third international consensus definitions for sepsis and septic shock (Sepsis-3)

Clinical features of patients infected with 2019 novel coronavirus in Wuhan

Longitudinal characteristics of lymphocyte responses and cytokine profiles in the peripheral blood of SARS-CoV-2 infected patients. medRxiv (2020)

SARS-CoV-2 Receptor ACE2 Is an Interferon-Stimulated Gene in Human Airway Epithelial Cells and Is Detected in Specific Cell Subsets across Tissues

The pathology of influenza virus infections

Multiple organ infection and the pathogenesis of SARS

Increasing host cellular receptor-angiotensin-converting enzyme 2 (ace2) expression by coronavirus may facilitate 2019-nCoV infection. bioRxiv (2020)

Inhibitors of RAS might be a good choice for the therapy of COVID-19 pneumonia

Effect of angiotensin-converting enzyme inhibition and angiotensin II receptor blockers on cardiac angiotensin-converting enzyme 2

Angiotensin converting enzyme inhibitors and pulmonary hypertension

Angiotensin II type 1 receptor signaling contributes to platelet-leukocyte-endothelial cell interactions in the cerebral microvasculature

Characteristics and outcomes of patients hospitalized for COVID-19 and cardiac disease in Northern Italy

COVID-19 associated coagulopathy: The crowning glory of thrombo-inflammation concept

Abnormal coagulation parameters are associated with poor prognosis in patients with novel coronavirus pneumonia

Is the use of RAS inhibitors safe in the current era of COVID-19 pandemic

Angiotensin II-inducible smooth muscle cell apoptosis involves the angiotensin II type 2 receptor, GATA-6 activation, and FasL-Fas engagement

ANG II induces apoptosis of human vascular smooth muscle via extrinsic pathway involving inhibition of Akt phosphorylation and increased FasL expression

Angiotensin II type 2 receptor overexpression activates the vascular kinin system and causes vasodilation

Cardiovascular effects of the angiotensin type 2 receptor

El-Mottaleb, Renal function and arterial bloodpressure alterations after exposure to acetaminophen with apotential role of Nigella sativaoil in adult male rats

Possible mechanism(s) for relaxant effect of aqueous andmacerated extracts fromNigella sativaon tracheal chains ofguinea pig

Data Availability: All data are available upon request

The authors declare they read the article and agreed to published its current revised form in the Journal Vascular Pharmacology In silico Nigellidine (N. sativa) bind to viral spike/active-sites of ACE1/2, AT1/2 to prevent COVID-19 induced vaso-tumult/vascular-damage/comorbidity