Tag Archives: lysyl oxidase
Involvement of Cu-containing amine oxidases in the development of lung pathology in ovalbumin-induced bronchial asthma in guinea pigs
O. Hudkova1*, S. Luhovskyi2, L. Drobot1, N. Latyshko1
1Palladin Institute of Biochemistry, National Academy of Sсiences of Ukraine, Kyiv;
2Dmitry F. Chebotarev Institute of Gerontology, National Academy of Medical Sciences of Ukraine, Kyiv;
*e-mail: ogudkova@biochem.kiev.ua
Received: 03 August 2022; Revised: 15 September 2022;
Accepted: 29 September 2022; Available on-line: 06 October 2022
Bronchial asthma is developed as an immune response to allergen challenges accompanied by inflammation and fibrosis implicated in airway remodeling. To reveal the causative implication of Cu-containing amine oxidases, semicarbazide-sensitive amine oxidase (SSAO), diamine oxidase (DAO), and lysyl oxidase (LOX), in bronchial asthma development we used their irreversible inhibitor, semicarbazide, and ovalbumin-induced pathology in guinea pigs. Semicarbazide was introduced to asthmatic animals via drink or inhalation. At the 16th week after disease induction, the increase in the activity of pro-inflammatory SSAO and DAO in plasma (1.6 and 2 times, respectively) was observed. The introduction of semicarbazide to asthmatic animals via drink or inhalation significantly decreased activities of these enzymes compared to the untreated asthmatic animals. A considerable increase in IL-13 content and LOX activity in the lung tissue of asthmatic animals were observed that evidenced airway inflammation and pulmonary fibrosis development. The uptake of semicarbazide by guinea pigs with bronchial asthma led to normalization of LOX activity. Histological studies confirmed that semicarbazide attenuated morphopathological changes in the lungs of asthmatic animals. Thus, the data obtained indicate the direct participation of the studied enzymes in the progression of pathological processes in atopic bronchial asthma as well as the potential use of semicarbazide as a drug in complex anti-asthmatic therapy.
Purification procedure and assay for the activity of lysyl oxidase
O. O. Gudkova, N. V. Latyshko, O. V. Zaitseva, S. G. Shandrenko
Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: gudkovahelga@gmail.com
The goal of the present study was to extract and purify lysyl oxidase from rodent’s tissues by a fast, simple, effective and inexpensive method and to develop a sensitive, time-saving lysyl oxidase specific activity assay for routine in vitro experiments. Lysyl oxidase was purified by elaborated purification procedure which relies on negative adsorption principle, that is, an effective decrease in the concentration of ballast components by the polar hydrophilic adsorbent and increasing the concentration of the protein of interest. Peroxide-coupled lysyl oxidase activity quantification methods based on luminol chemiluminescence in the presence of horseradish peroxidase as a catalyst and fluorescent detection using folic acid and Cu(II) with 1,5-diaminopentane as the substrate, were designed. Lysyl oxidase was partially purified from urea extracts of rodent’s tissues. Used purification procedure ensures the fast release of 93% of ballast proteins as shown by polyacrylamide gel electrophoresis. Lysyl oxidase specific activity after purification was 10-22-fold higher than that of the original extract. The molecular mass of murine lysyl oxidase from lung and heart was estimated to be ~32 kDa. We elaborated two sensitive methods for lysyl oxidase activity quantification and fast inexpensive procedure for partial enzyme purification useful in bulky in vitro experiments.