Atherosclerosis (AS) is a prevalent clinical vascular condition and serves as a pivotal pathological foundation for cardiovascular diseases. Understanding the pathogenesis of AS has significant clinical and societal implications, aiding in the development of targeted drugs. Neutrophils, the most abundant leukocytes in circulation, assume a central role during inflammatory responses and closely interact with AS, which is a chronic inflammatory vascular disease. Neutrophil extracellular traps (NETs) are substantial reticular formations discharged by neutrophils that serve as an immune defense mechanism. These structures play a crucial role in inducing dysfunction of the vascular barrier following endothelial cell injury. Components released by NETs pose a threat to the integrity of vascular endothelium, which is essential as it acts as the primary barrier to maintain vascular wall integrity. Endothelial damage constitutes the initial stage in the onset of AS. Recent investigations have explored the intricate involvement of NETs in AS progression. The underlying structures of NETs and their active ingredients, including histone, myeloperoxidase (MPO), cathepsin G, neutrophil elastase (NE), matrix metalloproteinases (MMPs), antimicrobial peptide LL-37, alpha-defensin 1-3, and high mobility group protein B1 have diverse and complex effects on AS through various mechanisms. This review aims to comprehensively examine the interplay between NETs and AS while providing insights into their mechanistic underpinnings of NETs in this condition. By shedding light on this intricate relationship, this exploration paves the way for future investigations into NETs while guiding clinical translation efforts and charting new paths for therapeutic interventions.
Extracellular Traps/physiology* ; Humans ; Atherosclerosis/immunology* ; Neutrophils/physiology* ; Leukocyte Elastase/metabolism* ; Peroxidase/physiology* ; Matrix Metalloproteinases/physiology* ; Cathepsin G/metabolism* ; Cathelicidins ; HMGB1 Protein/physiology* ; Histones ; Animals ; Endothelium, Vascular

OBJECTIVE: To investigate the distribution characteristics of polymorphonuclear neutrophil (PMN) in the lungs during the early stage of severe burns and the mechanism of neutrophil elastase (NE) promoting lung injury. METHODS: 6-8-week-old male C57BL/6J mice were selected for the experiments. A 30% total body surface area (TBSA) III degree burn mouse model was established (severe burn group); the Sham-injury group was treated with 37 centigrade water. In the sodium sivelestat intervention group (SV intervention group), NE competitive inhibitor, sivelestat, 100 mg/kg, was injected via tail vein immediately after injury, while other groups received an equal volume of saline. Ten mice were harvested from each group to observe survival for 72 hours. Respiratory function tests were tested at 0 (immediate), 3, 6, 12, and 24 hours after molding. hematoxylin-eosin (HE) and immunohistochemical staining were used to observe lung tissue structure, inflammatory changes and PMN infiltration. The PMN absolute count in mice lung tissue was detected buy flow cytometry. At 6, 12, and 24 hours after molding, PMN counts and the concentration of NE [enzyme linked immunosorbent assay (ELISA)] in peripheral blood plasma, lung tissue, and bronchoalveolar lavage fluid (BALF) were detected. RESULTS: (1) HE staining results showed that compared with the Sham-injury group, the lungs of mice in the severe burn group showed inflammatory changes and PMN infiltration, with more significant changes at 6 hours. Immunohistochemistry results also confirmed that the expression of NE protein released from PMN significantly increased after 6 hours of severe burn injury [(3.79±0.62)% vs. (0.18±0.05)%, t = 11.56, P < 0.01]. (2) Compared with the Sham-injury group, the number of PMN and the concentration of NE in the peripheral blood and lung tissues in the severe burn group were significantly increased (F values were 13.709, 55.350 and 29.890, 13.286, respectively, all P < 0.01), peaking at 6 hours [plasma PMN count (×10⁹/L): 2.92±1.01 vs. 0.92±0.29, lung tissue PMN absolute count (cells): 48 788.03±11 833.91 vs. 1 516.72±415.35, plasma NE (ng/L): 24 522.71±3 842.92 vs. 7 009.34±4 067.86, lung tissue NE (ng/L): 262 189.04±9 695.13 vs. 65 026.03± 16 016.31, all P < 0.01]. The number of PMN in the lung of severely burned mice was highly correlated with NE concentration (r = 0.892, P < 0.001). There was no significantly difference in the PMN absolute count in the BALF of mice between the Sham-injury group and severe burn group (F = 1.403, P > 0.05). The Sham-injury group and severe burn group contained a small amount of NE in the BALF, and the concentration of NE in the BALF of the severely burned 6 hours and 12 hours groups were significantly higher than those of the Sham-injury group (ng/L: 328.58±158.10, 415.30±240.89 vs. 61.95±15.80, both P < 0.05). (3) Kaplan-Meier survival curve showed that the 72-hour survival rate of mice in the SV intervention group was significantly higher than that in the severe burn group (100% vs. 10%, Log-Rank test: χ² = 19.12, P < 0.001). (4) Compared with the Sham-injury group, all lung function indices of the severe burn group decreased significantly. All lung function indices of SV intervention group improved gradually over time, which were significantly better than those of the severe burn group. (5) Compared with the Sham-injury group, the PMN absolute count in lung tissue and the concentration of NE in plasma and lung tissue were significantly higher in the SV intervention group (F values were 46.709, 3.535, 32.701, respectively, all P < 0.05), with a peak at 6 hours. Compared with the severe burn group, the SV intervention group had a higher PMN absolute count in lung tissue (cells: 8 870.80±7 013.89 vs. 25 974.92±22 240.8, P < 0.05), and higher plasma and lung tissue NE concentrations (ng/L: 14 955.94±3 944.41 vs. 21 972.75±4 573.05, 81 956.87±38 658.35 vs. 168 182.30±83 513.91, both P < 0.01) were significantly decreased. CONCLUSIONS In the early stage of severe burns, there is a significant infiltration of PMN into the lungs. The NE promotes lung injury in the early stage of severe burn, and improve lung injury by inhibiting the action of NE.
Animals ; Burns/metabolism* ; Leukocyte Elastase/metabolism* ; Male ; Mice, Inbred C57BL ; Mice ; Neutrophils/metabolism* ; Lung/metabolism* ; Disease Models, Animal ; Neutrophil Infiltration ; Lung Injury/metabolism* ; Glycine/analogs & derivatives* ; Sulfonamides

OBJECTIVE: To investigate the clinical efficacy of Huang'e Capsules in the treatment of type ⅢA chronic prostatitis, and its effects on the levels of the inflammatory cytokines neutrophil elastase (NE), IL-8 and TGF-β1 in the expressed prostatic secretion (EPS) of the patients. METHODS: We selected 120 patients with type ⅢA chronic prostatitis and randomly assigned them to medication with Huang'e Capsules (the trial group, n = 60) or Levofloxacin and Tamsulosin (the control group, n = 60), both for a course of 4 weeks. We obtained the NIH-CPSI scores and the levels of NE, IL-8 and TGF-β1 in the EPS, and compared them between the two groups before and after treatment. RESULTS: Totally, 116 of the patients completed the study, 59 in the trial and 57 in the control group. The overall clinical effectiveness was significantly higher in the trial group than in the control (89.8% vs 77.2%, P<0.05). Compared with the baseline, the patients of the trial group showed significant decreases after treatment in the total NIH-CPSI scores (32.5±7.4 vs 13.2±5.1), pain symptom scores (13.7±3.9 vs 4.2±2.3), urination symptom scores (6.9±2.4 vs 5.1±3.2) and quality of life (QOL) scores (8.3±2.7 vs 3.7±1.5) (all P< 0.05), and so did the controls in the total NIH-CPSI scores (30.8±7.8 vs 13.7±3.9), pain symptom scores (14.2±4.1 vs 7.8±2.9), urination symptom scores (7.1±2.7 vs 4.9±3.4) and quality of life (QOL) scores (8.1±2.4 vs 5.6±1.9) (all P< 0.05), and the decreases were even more significant in the trial than in the control group in the pain symptom and QOL scores ( P< 0.05). The patients of the trial group also exhibited a marked reduction after treatment in the contents of NE (［1135.4±321.5］ vs ［347.6±207.3］ ng/L, P< 0.05) and IL-8 (［974.9±231.6］ vs ［ 431.3±207.2］ ng/L, P< 0.05) but an elevation in that of TGF-β1 (［591.0±172.1］ vs ［1 402.1 ± 221.5］ ng/L, P< 0.05) in the EPS, and so did the controls in the levels of NE (［1052.8±280.3］ vs ［761.1±225.1］ ng/L, P<0.05), (［1007.5±287.7］ vs ［775.7±182.5］ ng/L, P< 0.05), (［607.8±201.3］ vs ［871.3±192.5］ ng/L, P< 0.05), with even more significant improvement in the trial than in the control group (P< 0.05). CONCLUSION Huang'e Capsules has a significant clinical efficacy and safety in the treatment of type IIIA prostatitis, which can effectively relieve the pain and urination symptoms and improve the levels of the inflammatory cytokines NE, IL-8 and TGF-β1 in the patients.
Humans ; Male ; Prostatitis/metabolism* ; Interleukin-8/metabolism* ; Drugs, Chinese Herbal/therapeutic use* ; Transforming Growth Factor beta1/metabolism* ; Prostate/metabolism* ; Leukocyte Elastase/metabolism* ; Tamsulosin ; Treatment Outcome ; Capsules ; Middle Aged ; Phytotherapy ; Cytokines/metabolism* ; Adult ; Levofloxacin

Objective: To investigate the clinical value of observing perioperative changes of myeloperoxidase (MPO) and neutrophil elastase (NE) in coronary artery circulation in patients underwent valve replacement surgery. Methods: This perspective cohort study was performed in patients who underwent valvular surgery in Nanjing Drum Tower Hospital and Fuwai Hospital from June 2021 to June 2022. Patients were divided into perioperative myocardial injury group and age-, sex- and type of cardiac procedure-matched non-perioperative myocardial injury control group in the ratio of 1∶1. Perioperative myocardial injury was defined as cardiac troponin T (cTnT)>0.8 μg/L on the first postoperative day (POD), and the cTnT level on the second POD increased by more than 10% compared with the cTnT level on the first POD. During the operation, blood samples were collected from the coronary sinus before clamping ascending aorta, and within 5 minutes after de-clamping ascending aorta. Then, the levels of MPO and NE on coronary sinus were continuously measured. The death, severe ventricular arrhythmia, pneumonia, re-intubation, repeat cardiac surgery, extracorporeal membrane oxygenation (ECMO), intra-aortic balloon pump (IABP), continuous renal replacement therapy (CRRT), mechanical ventilation time and the duration of intensive care unit (ICU) were recorded. The levels of MPO and NE and the incidence of clinical outcomes were compared between the myocardial injury group and the control group. The independent risk factors of myocardial injury were analyzed by multivariate logistic regression. Results: A total of 130 patients were enrolled, aged (60.6±7.6) years old, with 59 males (45.4%). There were 65 patients in the myocardial injury group and 65 patients in the control group. During hospitalization, there was no death, ECMO, IABP and CRRT cases in both groups. Compared with the control group, the incidence of severe ventricular arrhythmia (13.8%(9/65) vs. 3.1%(2/65), P=0.03), pneumonia (20.0%(13/65) vs. 3.1%(2/65), P=0.03), re-intubation (6.2%(4/65) vs. 0, P=0.04) was significantly higher in myocardial injury group. The mechanical ventilation time (16.8(10.7, 101.7) h vs. 7.5(4.7, 15.1) h, P<0.01), and the duration of ICU (3.7(2.7, 18.9) vs. 2.7(1.8, 6.9)d, P<0.01) were significantly longer in myocardial injury group compared with the control group. There was no significant difference in the levels of MPO and NE in coronary sinus blood between the two groups before aortic clamping (all P>0.05). However, MPO ((551.3±124.2) μg/L vs. (447.2±135.9) μg/L, P<0.01) and NE ((417.0±83.1)μg/L vs. (341.0±68.3)μg/L, P<0.01) after 5 min aortic de-clamping were significantly higher in myocardial injury group than in the control group. Multivariate logistic regression analysis showed that the levels of NE (OR=1.02, 95%CI: 1.01-1.02, P<0.01), MPO (OR=1.00, 95%CI: 1.00-1.01, P=0.02) and mechanical ventilation time (OR=1.03, 95%CI: 1.01-1.06, P=0.02) were independent risk factors of myocardial injury in patients after surgical valvular replacement. Conclusion: Perioperative myocardial injury is related poor clinical outcomes, perioperative NE and MPO in coronary artery circulation are independent risk factors of perioperative myocardial injury in patients undergoing valve replacement surgery.
Aged ; Humans ; Male ; Middle Aged ; Cohort Studies ; Coronary Circulation ; Leukocyte Elastase ; Peroxidase ; Prognosis ; Retrospective Studies ; Female

Extracellular elastase-like protease is one of the key virulence proteases of Scedosporium aurantiacum. To date, little is known about this enzyme in terms of genetic information, structure, properties and virulence mechanism due to the difficulties in purification caused by its low secretion amount, high specific activity, uncompleted genome sequencing and annotation. This work investigated the gene, structure and enzymatic properties of this enzyme. The S. aurantiacum elastase-like protease from the fungal culture supernatant was analyzed through tandem mass spectrometry (MS/MS) approach, illustrating its primary structure. Bioinformatics tools were employed to predict the conserved domain and tertiary structure, the enzymatic properties were also studied. It turned out that S. aurantiacum extracellular elastase-like protease demonstrated well hydrolysis towards elastin and bovine achilles tendon collagen, with V_max of 18.14 μg/s and 17.57 μg/s respectively, better than fish scale gelatin, with the lowest hydrolysis effect on casein. Its activity towards elastin was lower than that of the elastase from porcine pancreas, with values of K_cat/K_m of 3.541 (μg/s) and 4.091 (μg/s), respectively. It was an alkaline protease, with optimal pH 8.2 and temperature 37 ^oC. Zn²⁺ promoted the enzymatic activity while Ca²⁺, Mg²⁺, Na⁺, elastatinal and PMSF inhibited its activity. Its sequence was similar to Paecilomyces lilacinus secreted serine protease (PDB Entry: c3f7oB_) with multiple conserved fractions each containing more than 7 amino acids, thus suitable for design of PCR primer. This study increased our knowledge on S. aurantiacum extracellular elastase-like protease in terms of structure and enzymatic properties, and may facilitate later studies on protein expression and virulence mechanism.
Animals ; Cattle ; Pancreatic Elastase/genetics* ; Elastin/genetics* ; Tandem Mass Spectrometry ; Serine Proteases/genetics*

Humans ; Mutation ; Congenital Bone Marrow Failure Syndromes/genetics* ; Neutropenia/genetics* ; Leukocyte Elastase/genetics*

A large number of protease inhibitors have been found from leeches, which are essential in various physiological and biological processes. In the curret study, a novel elastase inhibitor was purified and characterized from the leech of Hirudinaria manillensis, which was named HMEI-A. Primary structure analysis showed that HMEI-A belonged to a new family of proteins. HMEI-A exerted inhibitory effects on elastase and showed potent abilities to inhibit elastase with an inhibition constant (K
Amino Acid Sequence ; Animals ; Leeches/chemistry* ; Pancreatic Elastase/antagonists & inhibitors* ; Protease Inhibitors/pharmacology* ; Proteins

OBJECTIVE: To establish and evaluate the model of chronic obstructive pulmonary disease (COPD) with osteoporosis induced by elastase in mice. METHODS: Twenty four healthy female 8-week-old C57BL / 6 mice (weighing about 18 g) were randomly divided into three groups. The control group was given intratracheal drip of normal saline, the experimental group 1 and the experimental group 2 were given intratracheal drip of elastase, the control group and the experimental group 1 were kept for 8 weeks and then killed, the experimental group 2 was kept for 12 weeks and then killed. HE staining was used to evaluate the histopathological changes of lung and tibia in the control and experimental groups. The levels of serum inflammatory factors and broncho alveolar lavage factors (BALF) were detected by ELISA. Micro CT was used to detect the bone mass related parameters of mouse femur. The expression of osteoclastic and osteogenic genes was detected by real-time fluorescence quantitative PCR. RESULTS: Lung histopathology showed that the structure of alveoli in the experimental group was disordered, the walls of alveoli became thin or broken, and the alveoli cavity expanded. IL-6 and TNF-α in BALF were significantly higher than those in control group (<0.001), while IL-1β and TNF-α in serum inflammatory factors were significantly higher than those in control group (<0.001). BV / TV(bone volume fraction), TB.Th(average bone trabecular thickness) and TB.N(average bone trabecular number) in the experimental group were significantly lower than those in the control group (<0.05), TB.Sp (average bone trabecular separation) and BS / BV (bone surface area fraction) in the experimental group were significantly higher than those in the control group (<0.01). Compared with the control group, the expression of osteoclast related marker genes increased in the experimental group (<0.05), but decreased in the experimental group(<0.05). The results of experiment 1 and experiment 2 were time-dependent. CONCLUSION In this study, elastase was used to construct a COPD model with osteoporosis successfully, which provides a suitable animal model for the future study of the pathogenesis of COPD with osteoporosis.
Animals ; Bone Density ; Female ; Mice ; Mice, Inbred C57BL ; Osteoporosis ; etiology ; Pancreatic Elastase ; Pulmonary Disease, Chronic Obstructive ; complications

OBJECTIVE: To delineate the clinical feature and genetic basis of four patients with congenital neutropenia. METHODS: All patients were subjected to whole exome sequencing (WES). Suspected variants were verified by Sanger sequencing. RESULTS: The patients (two boys and two girls), aged 7 to 15 months, suffered from neutropenia and recurrent infections. Bone marrow smears showed a significant decrease in the proportion of rod-shaped and lobulated granulocytes, which suggested impaired development and maturation of bone marrow neutrophils. WES has discovered heterozygous variants (c.496G>A, c.58C>G, c.391G>A and IVS1+5T>A) of the ELANE gene in the patients. Among these, c.58C>G and IVS1+5T>A were unreported previously. Follow up revealed patients 1 and 3 had periodic neutropenia, while patients 2 and 4 had severe congenital neutropenia. After attaining the definite diagnosis, the patients were treated symptomatically. CONCLUSION The main clinical feature of congenital neutropenia is refractory recurrent bacterial infections, for which mutations of the ELANE gene are a common cause. Two novel pathogenic ELANE variants have been discovered in this study.
Congenital Bone Marrow Failure Syndromes/genetics* ; Female ; Genetic Testing ; Humans ; Infant ; Leukocyte Elastase/genetics* ; Male ; Mutation ; Neutropenia/genetics*

Kawasaki disease is a form of vasculitis, mainly in small and medium arteries of unknown origin, occurring frequently in childhood. It is the leading form of childhood-onset acquired heart disease in developed countries and leads to complications of coronary artery aneurysms in approximately 25% of cases if left untreated. Although more than half a century has passed since Professor Tomisaku Kawasaki's first report in 1957, the cause is not yet clear. Currently, intravenous immunoglobulin therapy has been established as the standard treatment for Kawasaki disease. Various treatment strategies are still being studied under the slogan, “Ending powerful inflammation in the acute phase as early as possible and minimizing the incidence of coronary artery lesions,” as the goal of acute phase treatments for Kawasaki disease. Currently, in addition to immunoglobulin therapy, steroid therapy, therapy using infliximab, biological products, suppression of elastase secretion inside and outside the neutrophils, inactivated ulinastatin therapy and cyclosporine therapy, plasma exchange, etc. are performed. This chapter outlines the history and transition of the acute phase treatment for Kawasaki disease.
Aneurysm ; Arteries ; Biological Products ; Coronary Vessels ; Cyclosporine ; Developed Countries ; Heart Diseases ; Immunization, Passive ; Incidence ; Inflammation ; Infliximab ; Mucocutaneous Lymph Node Syndrome ; Neutrophils ; Pancreatic Elastase ; Plasma Exchange ; Prednisolone ; Vasculitis