Female ; Humans ; Infant ; Leukocyte Elastase ; isolation & purification ; Neutropenia ; congenital ; enzymology

Human leukocyte elastase is an important selection target of inflammation and cancer. In this paper, a high throughput screening model was established for screening human leukocyte elastase inhibitors from thousands of strains of actinomycetes. As a result, a strain, N01WA-735 with potent suppression activity was isolated. Firstly, the strain N01WA-735 was identified as Streptomyces according to morphology and biochemical analysis. The Streptomyces N01WA-735 was processed by solvent extraction, silica column chromatography, Sephadex LH-20 column chromatography and crystallization to get a pure active compound named N01WA-735E. Its chemical structure was elucidated as the same as that of the compound named BE-52440A by physicochemical properties and spectral data of UV, MS, 1H-NMR and 13C-NMR respectively. The compound showed a strong inhibitory activity against human leukocyte elastase with IC50 of 3.02 micromol/L. The compound is reported as a human leukocyte elastase inhibitor for the first time.
Humans ; Leukocyte Elastase ; antagonists & inhibitors ; Protease Inhibitors ; isolation & purification ; metabolism ; Streptomyces ; isolation & purification ; metabolism

Nonsteroidal antiinflammatory drugs(NSAIDs) are known as clinically effective agents for treatment of inflammatory diseases. Inhibition of cyclooxygenase has been thought to be a major facet of the pharmacological mechanism of NSAIDs. However, it is difficult to ascribe the antiinflammatory effects of NSAIDs solely to the inhibition of prostaglandin synthesis. Human neutrophil elastase (HNElastase; HNE, EC 3.4.21.37) has been known as a causative factor in inflammatory diseases. To investigate the specific relationship between HNElastase inhibition and specificity of molecular structure of several NSAIDs, HNElastase was purified by Ultrogel AcA54 gel filtration, CM-Sephadex ion exchange, and HPLC (with TSK 250 column) chromatography. HNElastase was inhibited by aspirin and salicylate in a competitive manner and by naproxen, ketoprofen, phenylbutazone, and oxyphenbutazone in a partial competative manner, but not by ibuprofen and tolmetin. HNElastase-phenylbutazone-complex showed strong Raman shifts at 200, 440, 1124, 1194, 1384, 1506, and 1768 cm(-1). The Raman bands 1194, 1384, and 1768 cm(-1) may represent evidences of the conformational change at -N=N-phi radical, pyrazol ring, and -C=O radical of the elastase-drug complex, respectively. Phenylbutazone might be bound to HNElastase by ionic and hydrophobic interaction, and masked the active site. Inhibition of HNElastase could be another mechanism of action of NSAIDs besides cyclooxygenase inhibition in the treatment of inflammatory diseases. Different inhibition characteristics of HNE-lastase by NSAIDs such as aspirin, phenylbutazone-like drugs and ineffective drugs could be important points for drawing the criteria for appropriate drugs in clinical application.
Anti-Inflammatory Agents, Non-Steroidal/pharmacology* ; Chromatography, Affinity ; Computer Simulation ; Enzyme Inhibitors/pharmacology ; Human ; Isoenzymes/isolation & purification ; Isoenzymes/antagonists & inhibitors ; Ketoprofen/pharmacology ; Leukocyte Elastase/isolation & purification ; Leukocyte Elastase/antagonists & inhibitors* ; Models, Molecular ; Naproxen/pharmacology ; Phenylbutazone/analogs & derivatives ; Salicylates/pharmacology ; Spectrum Analysis, Raman

OBJECTIVETo investigate the semen quality and its influencing factors in preconception males in Nanjing area so as to provide some evidence for working out effective intervention measures.

METHODSTotally 687 men receiving preconceptional physical examination were enrolled in this study. A questionnaire survey was conducted among the subjects along with an analysis of their semen quality.

RESULTSThe median of sperm concentration was 63.3 x 10(6)/ml (95% CI [19.88-119] x 10(6)/ml). The median of grade a sperm was 33.03% (95% CI [19.38-55.05]%), that of grade a + b sperm was 52.08% (95% CI [39.53-69.37]%), and that of teratosperm was 91.75% (95% CI [69-100]%). The median concentration of seminal plasma PMN-elastase was 195.55 ng/ml (95% CI [76.16-3330.38] ng/ml) and that of seminal plasma zinc was 7.62 μmol/L (95% CI [1.5-23, 45] μmol/L). The positive rates of Ureaplasma urealyticum (UU), Chlamydia trachomatis (CT), and Gardnerella vaginalis (GV) were 42.4%, 0.3%, and 2.4%, respectively. The median of sperm DNA fragmentation index (DFI) of those whose wives had a history of adverse pregnancy was 20.25% (95% CI [2.15-68.25]%). Multivariate logistic regression analysis suggested that mental stress (OR 1.567, 95% CI [1.081-2.27]) and sedentariness (OR 1.772, 95% CI [1.211-2.592]) were independent risk factors for asthenospermia.

CONCLUSIONThe sperm quality of preconception males in Nanjing area is not encouraging, and it can be improved by changing undesirable lifestyle and reducing mental stress.

Adult ; Asthenozoospermia ; etiology ; China ; Chlamydia trachomatis ; isolation & purification ; DNA Fragmentation ; Gardnerella vaginalis ; isolation & purification ; Humans ; Leukocyte Elastase ; analysis ; Male ; Preconception Care ; Semen ; microbiology ; Semen Analysis ; statistics & numerical data ; Sperm Count ; statistics & numerical data ; Spermatozoa ; Ureaplasma urealyticum ; isolation & purification

To study the coumarins of Anemone raddeana Regel, the compounds were separated by silica gel column chromatography and HPLC. Their structures were identified by their physicochemical property and spectral analysis. Two new compounds were isolated and identified as 4, 7-dimethoxyl-5-methyl-6-hydroxy coumarin (1) and 4, 7-dimethoxyl-5-formyl-6-hydroxycoumarin (2). The bioassays indicated that compounds 1 and 2 could significantly inhibit the proliferation of cancer cell, and showed the agonist effect on the transactivity of retinoic acid receptor-alpha (RARalpha). In addition, the two compounds had inhibitory effect against human leukocyte elastase (HLE).
Anemone ; chemistry ; Antineoplastic Agents, Phytogenic ; chemistry ; isolation & purification ; pharmacology ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Coumarins ; chemistry ; isolation & purification ; pharmacology ; Humans ; Inhibitory Concentration 50 ; Leukocyte Elastase ; metabolism ; Molecular Structure ; Plants, Medicinal ; chemistry ; Receptors, Retinoic Acid ; genetics ; metabolism ; Retinoic Acid Receptor alpha ; Rhizome ; chemistry ; Transcriptional Activation

Severe congenital neutropenia (SCN) is a heterogeneous group of disorders with a defect in granulopoiesis causing marked neutropenia and severe bacterial infections. A 17-month-old girl (patient 1) was admitted due to cervical lymphadenitis caused by methicillin-resistant Staphylococcus aureus, with neutropenia. She had Pseudomonas aeruginosa sepsis and peritonitis with perforated appendicitis at 8-month of age. Her sister, a 37-month-old girl (patient 2), had recurrent stomatitis with profound neutropenia, and her mother, a 32-yr-old woman (patient 3), had had recurrent stomatitis until her early 20s with neutropenia. We found an ELANE gene mutation (c.597+1G > A) from them in direct DNA sequencing analysis. Patients 1 and 2 did not respond to granulocyte colony stimulating factor and patient 1 was treated with prolonged antibiotics and excision. We demonstrated inherited SCN cases showing different severity even with the same mutation of the ELANE gene in a family.
Adult ; Child, Preschool ; DNA Mutational Analysis ; Female ; Granulocyte Colony-Stimulating Factor/therapeutic use ; Humans ; Infant ; Leukocyte Elastase/*genetics ; Methicillin-Resistant Staphylococcus aureus/isolation & purification ; Mutation/genetics ; Neutropenia/*congenital/diagnosis/drug therapy/genetics ; Pedigree ; *Phenotype ; Polymorphism, Single Nucleotide ; Recurrence ; Staphylococcal Infections/diagnosis/microbiology ; Stomatitis/diagnosis ; Tomography, X-Ray Computed