OBJECTIVE: Jingfang Granules have been recommended for the prevention and treatment of corona virus disease 2019 (COVID-19). Through chemical analysis and bioactivity evaluation, this study aims to elucidate the potential effective components of Jingfang Granules. METHODS: The inhibitory acti-vities of Jingfang Granules extract against 3-chymotrypsin-like protease (3CL^pro), papain like protease (PL^pro), spike protein receptor-binding domain (S-RBD) and human cyclooxygenase-2 (COX-2) were evaluated using enzyme assay. The antitussive effects were evaluated using the classical ammonia-induced cough model. The chemical constituents of Jingfang Granules were qualitatively and quantitatively analyzed by liquid chromatography-mass spectrometry (LC/MS). The 3CL^pro and PL^pro inhibitory activities of the major compounds were determined by enzyme assay, molecular docking, and site-directed mutagenesis. RESULTS: Jingfang Granules exhibited 3CL^pro and PL^pro inhibitory activities, as well as COX-2 inhibitory and antitussive activities. By investigating the MS/MS behaviors of reference standards, a total of fifty-six compounds were characterized in Jingfang Granules. Sixteen of them were unambiguously identified by comparing with reference standards. The contents of the 16 major compounds were also determined, and their total contents were 2 498.8 μg/g. Naringin, nodakenin and neohesperidin were three dominating compounds in Jingfang Granules, and their contents were 688.8, 596.4 and 578.7 μg/g, respectively. In addition, neohesperidin and naringin exhibited PL^pro inhibitory activities, and the inhibition rates at 8 μmol/L were 53.5% and 46.1%, respectively. Prim-O-glucosylcimifugin showed significant inhibitory activities against 3CL^pro and PL^pro, and the inhibitory rates at 8 μmol/L were 76.8% and 78.2%, respectively. Molecular docking indicated that hydrogen bonds could be formed between prim-O-glucosylcimifugin and amino acid residues H163, E166, Q192, T190 of 3CL^pro (binding energy, -7.7 kcal/mol) and K157, D164, R166, E167, T301 of PL^pro(-7.3 kcal/mol), respectively. Site-directed mutagenesis indicated amino acid residue K157 was a key active site for the interaction between prim-O-glucosylcimifugin and PL^pro. CONCLUSION Prim-O-glucosylcimifugin, neohesperidin, and naringin as the major compounds from Jingfang Granules could inhibit severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus proteases 3CL^pro and PL^pro. The results are valuable for rational clinical use of Jingfang Granules.
Amino Acids ; Ammonia ; Antitussive Agents ; COVID-19 ; Chymases ; Coronavirus 3C Proteases ; Cyclooxygenase 2 ; Cyclooxygenase 2 Inhibitors ; Cysteine Endopeptidases/metabolism* ; Humans ; Molecular Docking Simulation ; Papain ; Peptide Hydrolases ; SARS-CoV-2 ; Spike Glycoprotein, Coronavirus ; Tandem Mass Spectrometry

OBJECTIVETo assess the association of tag single nucleotide polymorphisms (tag SNPs) of chymase gene (CMA1) with essential hypertension in Yi population from Yunnan, China.

METHODSA case-control study was carried out. Four tag SNPs(rs1956921, rs1800876, rs5244 and rs1885108) were genotyped in 303 patients with essential hypertension and 312 healthy controls using polymerase chain reaction - restriction fragment length polymorphism(PCR-RFLP) method.

RESULTSNo significant difference in genotypic and allelic distributions of the four polymorphisms was detected between the two groups(P>0.05), and the same results existed in the females. The frequencies of rs1956921 C allele and a C-T haplotype constructed with rs1956921 and rs5244 were greater in male patients compared with male controls(P<0.01).

CONCLUSIONThe rs1956921 C allele of the CMA1 gene and the C-T haplotype constructed with rs1956921 and rs5244 may be risk factors for essential hypertension in ethnic Yi males from Yunnan.

Adult ; Alleles ; Asian Continental Ancestry Group ; ethnology ; genetics ; Blood Pressure ; China ; ethnology ; Chymases ; genetics ; Essential Hypertension ; Female ; Humans ; Hypertension ; ethnology ; genetics ; physiopathology ; Male ; Middle Aged ; Polymorphism, Single Nucleotide

Fatal anaphylactic shock is common in forensic practice. However, it is difficult to diagnose for lacking specific pathological and morphologic changes in forensic autopsy. The application of some biochemical indicators is of great significance. This paper reviews the biological characteristics of some biochemical indicators and detection methods. The forensic application, problems and prospects of these indicators are also introduced in details. The stable biochemical indicators, IgE, tryptase and chymase, show great potential and advantages in the identification of fatal anaphylactic shock in forensic medicine.
Anaphylaxis/metabolism* ; Autopsy ; Biomarkers ; Chymases ; Forensic Medicine ; Humans ; Tryptases

PURPOSE: The mast cell plays a pivotal role in the human immune response. Crosslinking of 2 IgE molecules bound to the high affinity IgE receptor (FcepsilonRI) on the surface of the mast cell results in mast cell degranulation and the release of several proinflammatory mediators. Patients with type-I allergy have increased levels of IgE in the blood compared to healthy individuals. METHODS: In a 6-week culture system of stem cells to human mast cells we investigated the effect of the concentration of IgE. The mast cells were cultured with different concentrations of IgE for the last 10 days of the maturation period. It was observed how the IgE concentration affects the histamine release, FcepsilonRI density on the mast cell surface and the concentration of other mediators. RESULTS: A clear correlation between IgE concentration in culture medium and the release of histamine upon activation was observed. It showed a bell-shaped dose response curve, with maximal response around an IgE-concentration of 250 ng/mL. Furthermore, the sensitivity of the mast cells and surface density of FcepsilonRI on mast cell surface was also influenced by the IgE concentration in the culture medium. CONCLUSIONS: IgE in the culture medium during the last 10 days of mast cell maturation influences the release of the preformed mediator histamine after mast cell activation and the density of FcepsilonRI on the mast cell surface. The release of the de novo synthetized mediator prostaglandin D2 and the expression of chymase and tryptase are not influenced by IgE in culture medium.
Chymases ; Histamine ; Histamine Release ; Humans ; Hygiene Hypothesis ; Hypersensitivity ; Immunoglobulin E ; Mast Cells ; Prostaglandin D2 ; Stem Cells ; Tryptases

PURPOSE: The mast cell plays a pivotal role in the human immune response. Crosslinking of 2 IgE molecules bound to the high affinity IgE receptor (FcepsilonRI) on the surface of the mast cell results in mast cell degranulation and the release of several proinflammatory mediators. Patients with type-I allergy have increased levels of IgE in the blood compared to healthy individuals. METHODS: In a 6-week culture system of stem cells to human mast cells we investigated the effect of the concentration of IgE. The mast cells were cultured with different concentrations of IgE for the last 10 days of the maturation period. It was observed how the IgE concentration affects the histamine release, FcepsilonRI density on the mast cell surface and the concentration of other mediators. RESULTS: A clear correlation between IgE concentration in culture medium and the release of histamine upon activation was observed. It showed a bell-shaped dose response curve, with maximal response around an IgE-concentration of 250 ng/mL. Furthermore, the sensitivity of the mast cells and surface density of FcepsilonRI on mast cell surface was also influenced by the IgE concentration in the culture medium. CONCLUSIONS: IgE in the culture medium during the last 10 days of mast cell maturation influences the release of the preformed mediator histamine after mast cell activation and the density of FcepsilonRI on the mast cell surface. The release of the de novo synthetized mediator prostaglandin D2 and the expression of chymase and tryptase are not influenced by IgE in culture medium.
Chymases ; Histamine ; Histamine Release ; Humans ; Hygiene Hypothesis ; Hypersensitivity ; Immunoglobulin E ; Mast Cells ; Prostaglandin D2 ; Stem Cells ; Tryptases

Oxidative stress has been paid increasing attention to as an important causative factor for diabetic vascular complications. Among possible various sources, accumulating evidence has indicated that NAD(P)H oxidase may be the most important source for reactive oxygen species production in diabetic vascular tissues. The mechanisms underlying activation and up-regulation of NAD(P)H oxidase has been supposed to be mediated by high glucose-induced protein kinase C (PKC) activation. In this review article, activation of local renin-angiotensin II system induced by chymase activation is also shown to amplify such a PKC-dependent activation of NAD(P)H oxidase. Additionally, human evidence showing the beneficial effect of antioxidants on diabetic vascular complications. Bilirubin has been recognized as a strong endogenous antioxidant. Here markedly lower prevalence of vascular complications is shown in diabetic patients with Gilbert syndrome, a congenital hyperbilirubinemia, as well as reduced markers of oxidative stress and inflammation. Lastly, statin, angiotensin II receptor blocker, chymase inhibitor, bilirubin and biliverdin, PKC beta isoform inhibitor, and glucagon-like peptide-1 analog, are shown to serve as antioxidants and have some beneficial effect on diabetic vascular complications, via inhibiting PKC-NAD(P)H oxidase activation, supporting the notion that this mechanism may be an effective therapeutic target for preventing diabetic vascular complications.
Angiotensin II ; Antioxidants ; Bilirubin ; Biliverdine ; Chymases ; Diabetic Angiopathies ; Gilbert Disease ; Glucagon-Like Peptide 1 ; Humans ; Hydroxymethylglutaryl-CoA Reductase Inhibitors ; Hyperbilirubinemia ; Inflammation ; NADPH Oxidase ; Oxidative Stress ; Oxidoreductases ; Prevalence ; Protein Kinase C ; Reactive Oxygen Species ; Receptors, Angiotensin ; Up-Regulation

To screen potential hamster chymase 2 inhibitors, a high-throughput screening (HTS) model was established. Recombinant hamster chymase 2 with active form was cloned and expressed in E. coli. The HTS model with total volume of 50 microL in 384-well microplate was based on fluorescence analysis and was proved sensitive as well as specific (Z' = 0.84). A total of 40 080 samples (including 28 060 compounds and 12 020 natural products) were screened, and 613 samples with inhibition greater than 90% were selected for further rescreening. Finally, compounds J16647 and J16648 were identified with high inhibitory activity on chymase 2, and whose IC50 values were 0.823 and 0.690 micromol x L(-1), respectively.
Animals ; Chymases ; analysis ; antagonists & inhibitors ; Cricetinae ; Enzyme Inhibitors ; analysis ; pharmacology ; Escherichia coli ; metabolism ; High-Throughput Screening Assays ; methods ; Inhibitory Concentration 50 ; Rats ; Structure-Activity Relationship

OBJECTIVE: To explore the application value of serum total IgE, tryptase and chymase in the identification of death caused by drug anaphylactic shock. METHODS: The general information from 235 cases of non-drug anaphylactic shock and 32 cases of drug anaphylactic shock were analyzed. The serum IgE level had been detected in the cases. Ten cases caused by coronary disease and 10 cases caused by sudden manhood death syndrome were selected from non-drug anaphylactic shock cases for the control group. Expressions of tryptase and chymase in the lung and heart were detected using immunohistochemistry method. The number and IOD of positive mast cells were counted. RESULTS: In the drug anaphylactic shock group, the IgE value of 18 samples (56.25%) was significantly higher than the normal upper limit of 120 IU/mL. In the non-drug anaphylactic shock group, the IgE value of 67 samples (28.51%) was higher than 120 IU/mL. The expressions of tryptase and chymase were significantly increased in lung and myocardial tissue in drug anaphylactic shock group (P < 0.05). CONCLUSION Tryptase and chymase are more superior than that of the serum total IgE in the diagnosis of death caused by drug anaphylactic shock, and are more suitable in forensic practice.
Adolescent ; Adult ; Aged ; Anaphylaxis/pathology* ; Autopsy ; Case-Control Studies ; Cause of Death ; Child ; Child, Preschool ; Chymases/metabolism* ; Death, Sudden, Cardiac/pathology* ; Drug Hypersensitivity ; Female ; Forensic Pathology ; Humans ; Immunoglobulin E/blood* ; Immunohistochemistry ; Infant ; Lung/pathology* ; Male ; Middle Aged ; Myocardium/pathology* ; Tryptases/metabolism* ; Young Adult

OBJECTIVETo evaluate the roles of plasma mast cell carboxypeptidase and chymase in the diagnosis of allergic diseases by measuring the contents of both in children.

METHODSA total of 59 children with allergic diseases and 53 healthy children were recruited into the study. Plasma levels of mast cell carboxypeptidase and chymase were measured using ELISA.

RESULTSThe plasma levels of mast cell carboxypeptidase and chymase in children with allergic children were 1.089 ± 0.752 ng/mL and 0.905(0.375-2.318) ng/mL, respectively, which were significantly higher than those in healthy children [0.593 ± 0.380 ng/mL and 0.454 (0.097-1.077) ng/mL respectively; P<0.05]. There was a significantly positive correlation between plasma mast cell carboxypeptidase and chymase levels in children with allergic diseases (r=0.684, P<0.01).

CONCLUSIONSPlasma levels of mast cell carboxypeptidase and chymase increase in children with allergic diseases, suggesting that mast cell carboxypeptidase and chymase may serve as the indexes for the diagnosis of allergic diseases.

Adolescent ; Carboxypeptidases ; blood ; Child ; Child, Preschool ; Chymases ; blood ; Female ; Humans ; Hypersensitivity ; diagnosis ; enzymology ; Infant ; Infant, Newborn ; Male ; Mast Cells ; enzymology

OBJECTIVE: To explore the expression of tryptase and chymase in human lung tissue of anaphylactic shock and its value for forensic medicine. METHODS: With ten carbon monoxide poisoning cases as control group, the levels of tryptase and chymase were observed by immunofluorescence and analyzed using the Image Analyze and the Image-pro plus 5.0.2. The positive mast cells were counted and the levels of the tryptase and chymase were calculated respectively. RESULTS: There was a statistically significant difference (P < 0.05) for the tryptase and chymase concentrations in the lung tissue between the anaphylactic shock group and the control group. CONCLUSION The levels of the tryptase and the chymase expression are greatly increased in human lung tissue of anaphylactic shock, which may provide the morphological evidence and reference for the diagnosis of anaphylactic shock in forensic practice.
Adolescent ; Adult ; Anaphylaxis/pathology* ; Cadaver ; Carbon Monoxide Poisoning/pathology* ; Child ; Child, Preschool ; Chymases/metabolism* ; Female ; Fluoroimmunoassay/methods* ; Forensic Pathology ; Humans ; Infant ; Lung/pathology* ; Male ; Mast Cells/enzymology* ; Middle Aged ; Staining and Labeling ; Tryptases/metabolism* ; Young Adult