Angiotensin I converting enzyme(ACE)inhibitory activity and antihypertensive effects of rice peptides

In this paper,the antihypertension effect of rice peptide(RP)was studied.With spontaneously hypertensive rats(SHR)and Wistar Kyoto(WKY)as the research objects,RP disposable gastric and long-term gastric irrigation experiments were carried out and systolic blood pressure(SBP)was measured.At the end of the long-term gastric irrigation experiment,the content of nitric oxide(NO),angiotensin-converting enzyme(ACE),angiotensin II(Ang II)and renin in the plasma and the activity of ACE were determined.The results showed that RP could reduce systolic pressure of SHR and had time-dose dependence while high-dose RP signifi cantly reduced systolic pressure by 24.6 and 17.2 mm Hg,respectively after a single and long-term gastric irrigation test.RP also could inhibit the activity of ACE and increase the release of NO.These results suggested that the decompression mechanism of RP is likely to be related to the regulation of the renin-angiotensin system(RAS)and NO.

SARS coronavirus entry into host cells through a novel clathrin- and caveolae-independent endocytic pathway

While severe acute respiratory syndrome coronavirus （SARS-CoV）~as initially thought to enter cells through direct fusion with the plasma membrane, more recent evidence suggests that yirus entry may also involve endocytosis. We have found that SARS-CoV enters cells viapH- and receptor-dependent endocytosis. Treatment of cells with either SARS-COV spike protein or spike-bearing pseudoviruses resulted in the translocation of angiotensin-converting enzyme 2 （ACE2）, the functional receptor of SARS-CoV, from the cell surface to endosomes. In addition, the spike-bearing pseudoviruses and early endosome antigen 1 were found to colocalize in endosomes. Further analyses using specific endocytic path- way inhibitors and dominant-negative Epsl5 as well as caveolin-1 colocalization study suggested that virus entry was mediated by a clathrin- and caveolae-independent mechanism. Moreover, cholesterol- and sphingolipid-rich lipid raft microdomains in the plasma membrane, which have been shown to act as platforms for many physiological signaling pathways, were shown to be involved in virus entry. Endocytic entry of SARS-CoV may expand the cellular range of SARS-CoV infection, and our findings here contribute to the understanding of SARS-CoV pathogenesis, providing new information for anti-viral drug research.

Ferrets: A powerful model of SARS-CoV-2

The rapid spread of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) in recent years not only caused a global pandemic but resulted in enormous social,economic,and health burdens worldwide.Despite considerable efforts to combat coronavirus disease 2019(COVID-19),various SARS-CoV-2 variants have emerged,and their underlying mechanisms of pathogenicity remain largely unknown.Furthermore,effective therapeutic drugs are still under development.Thus,an ideal animal model is crucial for studying the pathogenesis of COVID-19 and for the preclinical evaluation of vaccines and antivirals against SARS-CoV-2 and variant infections.Currently,several animal models,including mice,hamsters,ferrets,and nonhuman primates(NHPs),have been established to study COVID-19.Among them,ferrets are naturally susceptible to SARS-CoV-2 infection and are considered suitable for COVID-19 study.Here,we summarize recent developments and application of SARS-CoV-2 ferret models in studies on pathogenesis,therapeutic agents,and vaccines,and provide a perspective on the role of these models in preventing COVID-19 spread.

Identification of therapeutic drugs against COVID-19 through computational investigation on drug repurposing and structural modification

Global prevalence of coronavirus disease 2019(COVID-19)calls for an urgent development of anti-viral regime.Compared with the development of new drugs,drug repurposing can significantly reduce the cost,time,and safety risks.Given the fact that coronavirus harnesses spike protein to invade host cells through angiotensinconverting enzyme 2(ACE2),hence we see if any previous anti-virtual compounds can block spike-ACE2 interaction and inhibit the virus entry.The results of molecular docking and molecular dynamic simulations revealed that remdesivir exhibits better than expected anti-viral invasion potential against COVID-19 among the three types of compounds including remdesivir,tenofovir and lopinavir.In addition,a positive correlation between the surface area occupied by remdesivir and anti-viral invasion potential was also found.As such,the structure of remdesivir was modified by linking an N-benzyl substituted diamidine derivative to its hydroxyl group through an ester bond.It was found that this compound has a higher anti-viral invasion potential and greater specificity.

The Study of Molecular Mechanisms of Xuanbai Chengqi Decoction(宣白承气汤)in the Treatment of Coronavirus Disease 2019 Based on Network Pharmacology and Molecular Docking Method

Objective:To explore the molecular mechanism of Xuanbai Chengqi Decoction(宣白承气汤)in the treatment of Coronavirus Disease 2019(COVID-19)based on network pharmacology,and to verify by molecular docking technology.Methods:The components and targets of Xuanbai Chengqi Decoction(宣白承气汤)were obtained by TCMSP,targets’information was corrected based on the databases such as UniP rot and DrugBank,and the software Cytoscape3.7.1 was adopted to construct TCM-Component-Target and Component-Target network.The main targets were mapped to the KEGG pathway and the GO biological process with the help of DAVID to further elucidate the potential relationship between the main targets and Xuanbai Chengqi Decoction(宣白承气汤)therapy for COVID-19.In the end,the Swiss Dock platform was adopted for the molecular docking verification of key components and targets.Results:The Component-Target network consists of 35 components and 106 corresponding targets,the main targets include COX-2,NCOA2,PTGS1,HSP90 AB1,PRKACA and PGR,etc.There are 561 GO entries of target mapping(P 0.05),including 155 entries for Biological Processes(BP),147 entries for Cell Composition(CC),and 259 entries for Molecular Function(MF).There are 38 KEGG mapping pathways(P 0.05),including many aspects of infectious disease,immune system and endocrine system,as well as Calcium signaling pathway,VEGF signaling pathway,PI3K-Akt signaling pathway and other processes.Conclusion:The result of molecular docking shows that the affinity of the key components such as beta-sitosterol and stigmasterol are similar to recommended medications for COVID-19.Its effect in the treatment of middle stage of COVID-19 may be related to the blocking of the binding of COVID-19 virus and ACE2,antivirus,and relieving inflammatory storm.

Interferon Regulatory Factor 5 and Renin-Angiotensin-Aldosterone System Polymorphisms in Coronary Artery Disease: An Overview of Experimental and Clinical Studies

Heart diseases are the main cause of mortality in Mexico, being coronary heart disease the most frequent in the country. Its high prevalence makes important the study of the pathophysiology and the search for prognostic factors. Different genes and polymorphisms promote atherogenesis and coronary artery disease, they affect inflammatory and vascular pathological processes. Interferon regulatory factor 5 (IRF5) is associated with coronary heart disease, it promotes chronic inflammation and cytokines release;it could trigger immune reactions and its activating receptors express in the vascular endothelium. Besides, polymorphisms in the renin-angiotensin-aldosterone system (RAAS) are implied with coronary disease, they are found in angiotensinogen (AGT), angiotensin II type 1 receptor (AT1R), angiotensin II type 2 receptor (AT2R), and angiotensin-converting enzyme (ACE) genes. These genetic polymorphisms are associated with a prothrombotic state, endothelial dysfunction, and immune activation. Multiple experimental studies showed that chronic activation of RAAS and chronic expression of IRF5 generates an environment prone to the development of atherosclerosis, and autoimmune and cardiovascular diseases. Studying these specific genes and their relationship with coronary heart disease will allow a better understanding of the pathological process and possibly the quest for new treatments.

Cytoplasmic domain and enzymatic activity of ACE2 are not required for PI4KB dependent endocytosis entry of SARS-CoV-2 into host cells

The recent COVID-19 pandemic poses a global health emergency.Cellular entry of the causative agent SARS-CoV-2 is mediated by its spike protein interacting with cellular receptor-human angiotensin converting enzyme 2(ACE2).Here,by using lentivirus based pseudotypes bearing spike protein,we demonstrated that entry of SARS-CoV-2 into host cells was dependent on clathrin-mediated endocytosis,and phosphoinositides played essential roles during this process.In addition,we showed that the intracellular domain and the catalytic activity of ACE2 were not required for efficient virus entry.Finally,we showed that the current predominant Delta variant,although with high infectivity and high syncytium formation,also entered cells through clathrin-mediated endocytosis.These results provide new insights into SARS-CoV-2 cellular entry and present proof of principle that targeting viral entry could be an effective way to treat different variant infections.

Inhibitory mechanism of angiotensin-converting enzyme inhibitory peptides from black tea

The aim of this work is to discover the inhibitory mechanism of tea peptides and to analyse the affinities between the peptides and the angiotensin-converting enzyme(ACE)as well as the stability of the complexes using in vitro and in silico methods.Four peptide sequences identified from tea,namely peptides I,II,III,and IV,were used to examine ACE inhibition and kinetics.The half maximal inhibitory concentration(IC_(50))values of the four peptides were(210.03±18.29),(178.91±5.18),(196.31±2.87),and(121.11±3.38)μmol/L,respectively.The results of Lineweaver-Burk plots showed that peptides I,II,and IV inhibited ACE activity in an uncompetitive manner,which requires the presence of substrate.Peptide III inhibited ACE in a noncompetitive manner,for which the presence of substrate is not necessary.The docking simulations showed that the four peptides did not bind to the active sites of ACE,indicating that the four peptides are allosteric inhibitors.The binding free energies calculated from molecular dynamic(MD)simulation were-72.47,-42.20,-52.10,and-67.14 kcal/mol(1 kcal=4.186 kJ),r espectively.The lower IC_(50)value of peptide IV may be attributed to its stability when docking with ACE and changes in the flexibility and unfolding of ACE.These four bioactive peptides with ACE inhibitory ability can be incorporated into novel functional ingredients of black tea.

Antibody Cocktail Exhibits Broad Neutralization Activity Against SARS-CoV-2 and SARS-CoV-2 Variants

Severe acute respiratory syndrome coronavirus 2(SARS-Co V-2)has precipitated multiple variants resistant to therapeutic antibodies.In this study,12 high-affinity antibodies were generated from convalescent donors in early outbreaks using immune antibody phage display libraries.Of them,two RBD-binding antibodies(F61 and H121)showed high-affinity neutralization against SARS-Co V-2,whereas three S2-target antibodies failed to neutralize SARS-Co V-2.Following structure analysis,F61 identified a linear epitope located in residues G446–S494,which overlapped with angiotensinconverting enzyme 2(ACE2)binding sites,while H121 recognized a conformational epitope located on the side face of RBD,outside from ACE2 binding domain.Hence the cocktail of the two antibodies achieved better performance of neutralization to SARS-Co V-2.Importantly,these two antibodies also showed efficient neutralizing activities to the variants including B.1.1.7 and B.1.351,and reacted with mutations of N501 Y,E484 K,and L452 R,indicated that it may also neutralize the recent India endemic strain B.1.617.The unchanged binding activity of F61 and H121 to RBD with multiple mutations revealed a broad neutralizing activity against variants,which mitigated the risk of viral escape.Our findings revealed the therapeutic basis of cocktail antibodies against constantly emerging SARS-Co V-2 variants and provided promising candidate antibodies to clinical treatment of COVID-19 patients infected with broad SARS-Co V-2 variants.

Long Term Culture of Human Kidney Proximal Tubule Epithelial Cells Maintains Lineage Functions and Serves as an Ex vivo Model for Coronavirus Associated Kidney Injury

The mechanism of how SARS-CoV-2 causes severe multi-organ failure is largely unknown. Acute kidney injury(AKI) is one of the frequent organ damage in severe COVID-19 patients. Previous studies have shown that human renal tubule cells could be the potential host cells targeted by SARS-CoV-2. Traditional cancer cell lines or immortalized cell lines are genetically and phenotypically different from host cells. Animal models are widely used, but often fail to reflect a physiological and pathogenic status because of species tropisms. There is an unmet need for normal human epithelial cells for disease modeling. In this study, we successfully established long term cultures of normal human kidney proximal tubule epithelial cells(KPTECs) in 2 D and 3 D culture systems using conditional reprogramming(CR) and organoids techniques.These cells had the ability to differentiate and repair DNA damage, and showed no transforming property. Importantly, the CR KPTECs maintained lineage function with expression of specific transporters(SLC34 A3 and cubilin). They also expressed angiotensin-converting enzyme 2(ACE2), a receptor for SARS-CoV and SARS-CoV-2. In contrast, cancer cell line did not express endogenous SLC34 A3, cubilin and ACE2. Very interestingly, ACE2 expression was around twofold higher in 3 D organoids culture compared to that in 2 D CR culture condition. Pseudovirion assays demonstrated that SARS-CoV spike(S) protein was able to enter CR cells with luciferase reporter. This integrated 2 D CR and 3 D organoid cultures provide a physiological ex vivo model to study kidney functions, innate immune response of kidney cells to viruses, and a novel platform for drug discovery and safety evaluation.

Macrophage exosomes transfer angiotensin Ⅱ type 1 receptor to lung fibroblasts mediating bleomycin-induced pulmonary fibrosis

Background:Macrophages are involved in the pathogenesis of idiopathic pulmonary fibrosis,partially by activating lung fibroblasts.However,how macrophages communicate with lung fibroblasts is largely unexplored.Exosomes can mediate intercellular communication,whereas its role in lung fibrogenesis is unclear.Here we aim to investigate whether exosomes can mediate the crosstalk between macrophages and lung fibroblasts and subsequently induce fibrosis.Methods:In vivo,bleomycin(BLM)-induced lung fibrosis model was established and macrophages infiltration was examined.The effects of GW4869,an exosomes inhibitor,on lung fibrosis were assessed.Moreover,macrophage exosomes were injected into mice to observe its pro-fibrotic effects.In vitro,exosomes derived from angiotensin Ⅱ(Ang Ⅱ)-stimulated macrophages were collected.Then,lung fibroblasts were treated with the exosomes.Twenty-four hours later,protein levels ofα-collagen I,angiotensin Ⅱ type 1 receptor(AT1R),transforming growth factor-β(TGF-β),and phospho-Smad2/3(p-Smad2/3)in lung fibroblasts were examined.The Student's t test or analysis of variance were used for statistical analysis.Results:In vivo,BLM-treated mice showed enhanced infiltration of macrophages,increased fibrotic alterations,and higher levels of Ang Ⅱ and AT1R.GW4869 attenuated BLM-induced pulmonary fibrosis.Mice with exosomes injection showed fibrotic features with higher levels of Ang Ⅱ and AT1R,which was reversed by irbesartan.In vitro,we found that macrophages secreted a great number of exosomes.The exosomes were taken by fibroblasts and resulted in higher levels of AT1R(0.22±0.02 vs.0.07±0.02,t=8.66,P=0.001),TGF-β(0.54±0.05 vs.0.09±0.06,t=10.00,P<0.001),p-Smad2/3(0.58±0.06 vs.0.07±0.03,t=12.86,P<0.001)andα-collagen I(0.27±0.02 vs.0.16±0.01,t=7.01,P=0.002),and increased Ang Ⅱ secretion(62.27±7.32 vs.9.56±1.68,t=12.16,P<0.001).Interestingly,Ang Ⅱ increased the number of macrophage exosomes,and the protein levels of Alix(1.45±0.15 vs.1.00±0.10,t=4.32,P=0.012),AT1R(4.05±0.64 vs.1.00±0.09,t=8.17,P=0.001),and glyceraldehyde-3-phosphate dehydrogenase(2.13±0.36 vs.1.00±0.10,t=5.28,P=0.006)were increased in exosomes secreted by the same number of macrophages,indicating a positive loop between Ang Ⅱ and exosomes production.Conclusions:Exosomes mediate intercellular communication between macrophages and fibroblasts plays an important role in BLM-induced pulmonary fibrosis.

Effects of angiotensin Ⅱ receptor blocker usage on viral load,antibody dynamics, and transcriptional characteristics among COVID-19 patients with hypertension

Epidemiological evidence suggests that patients with hypertension infected with severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)are at increased risk of acute lung injury.However,it is still not clear whether this increased risk is related to the usage of renin-angiotensin system(RAS)blockers.We collected medical records of coronavirus disease 2019(COVID-19)patients from the First Affiliated Hospital,Zhejiang University School of Medicine(Hangzhou,China),and evaluated the potential impact of an angiotensin II receptor blocker(ARB)on the clinical outcomes of COVID-19 patients with hypertension.A total of 30 hypertensive COVID-19 patients were enrolled,of which 17 were classified as non-ARB group and the remaining 13 as ARB group based on the antihypertensive therapies they received.Compared with the non-ARB group,patients in the ARB group had a lower proportion of severe cases and intensive care unit(ICU)admission as well as shortened length of hospital stay,and manifested favorable results in most of the laboratory testing.Viral loads in the ARB group were lower than those in the non-ARB group throughout the disease course.No significant difference in the time of seroconversion or antibody levels was observed between the two groups.The median levels of soluble angiotensin-converting enzyme 2(sACE2)in serum and urine samples were similar in both groups,and there were no significant correlations between serum sACE2 and biomarkers of disease severity.Transcriptional analysis showed 125 differentially expressed genes which mainly were enriched in oxygen transport,bicarbonate transport,and blood coagulation.Our results suggest that ARB usage is not associated with aggravation of COVID-19.These findings support the maintenance of ARB treatment in hypertensive patients diagnosed with COVID-19.

The clinical implication of gamma-glutamyl transpeptidase in COVID-19

Background and aim:Coronavirus disease 2019(COVID-19)is a life-threatening disease that predomi-nantly causes respiratory failure.The impact of COVID-19 on other organs remains elusive.Herein,we aimed to investigate the effects of COVID-19 on the hepatobiliary system.Methods:In the current study,we obtained the clinical records and laboratory results from 66 laboratory-confirmed patients with COVID-19 at the Wuhan Tongji Hospital between 10 February 2020 and 28 February 2020.The detailed clinical features and laboratory findings were collected for analysis.Bioinformatics analysis was conducted to evaluate the correlation between gamma-glutamyl transferase(GGT)and severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)entry receptor angiotensin-converting enzyme 2(ACE2).Results:In this cohort,30(51.7%)patients had abnormal liver function on admission,which was asso-ciated with disease severity and enriched in the male and diabetic patients.The elevated levels of direct bilirubin(P¼0.029)and GGT(P¼0.004)were common in patients with severe pneumonia when compared with those with mild pneumonia.In addition,elevated levels of GGT(P¼0.003)and aspartate aminotransferase(AST)(P¼0.007)were positively associated with longer hospital stay.The expression of ACE2 was closely associated with GGT in various human tissues because they shared the common transcriptional regulator hepatic nuclear factor-1 b(HNF1B).Conclusions:Increased GGT levels were common in severe cases and elevated GGT levels were positively associated with prolonged hospital stay and disease severity.Due to the consistent expression with ACE2,GGT is a potent biomarker indicating the susceptibility of SARS-CoV-2 infection.

Angiotensin Converting Enzyme as a Possible Biomarker to Predict Effect of Obstetric Anesthesia on Pregnant Women with COVID-19 Infection

We retrospectively investigated 68 parturients with or without COVID-19 undergone emergency cesarean section with combined spinal-epidural anesthesia(CSEA)from a single tertiary university hospital in Wuhan,China.The cases were divided into 2 groups:patients with COVID-19 pneumonia(Group 1)and cases without COVID-19 pneumonia(Group 2).The patients in Group 1 were later divided into 2 groups:patients with low-angiotensin converting enzyme(ACE)(Group 3)and patients with normal-ACE(Group 4).The ACE levels,blood pressure and anesthesia management between the patients of Group 1 and Group 2,Group 3 and Group 4 were recorded as the primary outcome.The secondary outcome included perioperative symptoms,laboratory parameters and vital signs.Compared with Group 2,the patients in Group 1 had different ACE level and lower blood pressure after CSEA.Compared with Group 4,the patients in Group 3 showed lower SBP after CSEA(127 vs.130 mm Hg,p=0.028),accompanied with more partus matures and younger age(28 vs.32 years,p=0.007).ACE may be a possible biomarker to predict the anesthesia effects on patients with COVID-19 infections undergoing emergency cesarean delivery.