Chronic Diseases of Respiratory Organs Diagnostics
Diagnostics and Analyser of Respiratory Organs Inflammation Activity at Chronic Obstructive Pulmonary Disease, Bronchial Asthma and Tuberculosis Using Determination of Hydrogen Peroxide Content in Biosubstrates
Tech Area / Field
- MED-DID/Diagnostics & Devices/Medicine
- CHE-ANL/Analytical Chemistry/Chemistry
- INS-DET/Detection Devices/Instrumentation
8 Project completed
Senior Project Manager
Malakhov Yu I
VNIIEF, Russia, N. Novgorod reg., Sarov
- Moscow State University / Department of Chemistry, Russia, Moscow\nCentral Tuberculosis Research Institute, Russia, Moscow\nJoint Stock Company PRACTIC-NC, Russia, Moscow, Zelenograd
- Alfred Wegener Institute for Polar and Marine Research, Germany, Bremerhaven\nUniversitat de Barcelona / Department de Medecina, Spain, Barcelona\nQueen Mary, University of London / Interdisciplinary Centre in Biomedical Materials, UK, London\nUniversity of Lund / Department of Analytical Chemistry, Sweden, Lund
Project summaryChronic diseases of the respiratory organs (first of all, chronic obstructive pulmonary disease (COPD) and bronchial asthma (BA)) take one of the leading places among diseases of humans leading to high disablement and mortality. Only in the USA the number of people suffering from COPD is estimated to achieve 14 million (about 14% of smokers and 3% of nonsmokers) according to the 3rd U.S. National Health and Nutrition Examination Survey. Though for other countries these data are less exact, the World Organization of Health (WOH) places the COPD at the twelfth place according to the level of its occurrence in the world. According to the level of mortality the COPD takes the sixth place in the world (4th place in the USA) and is the only disease in industrial countries, mortality from which increases. Besides, the significance of COPD as the reason for mortality is very likely underestimated as this disease contributes to cases of death from other more pronounced diagnoses. Particularly, it is to be noted that in case of COPD its occurrence and mortality from it increases both in the developing countries and in industrial countries. All this leads to the fact that according to the predictions of the WOH by 2020 the COPD will take the fifth place in the world in its occurrence and the third place in mortality. Thus, in the coming 20 years the chronic obstructive diseases of respiratory organs, probably, will become one of the main problems of modern medicine.
The pathogenetic basis of both the COPD and BA is chronic inflammation in the respiratory tract, which in each case has its peculiar features and forms a clinical picture of these diseases. One of the components of the inflammatory process is an oxidative stress – synthesis of the excess quantity of active forms of oxygen (AFO) and active forms of nitrogen (AFN) in the locus of inflammation. The oxidative processes are evolutionally produced means for anti-infection protection of the organism. They damage structural components of the antigenes when they are acted upon by AFO (superoxide anion-radical O2-, singlet oxygen O2, radicals HOO and HO, hydrogen peroxide H2O2). However, in case of COPD and BA the AFO for various reasons are not capable to act upon the antigene efficiently, but the organism continues to produce them in excess quantity, which causes exhaustion of anti-oxidant protection and damage of its tissues. The modern approaches to therapy of this process, as a rule, have a delayed clinical effect and for their use to be optimized fair monitoring of oxidative reactions in the diseased organ is needed. Thus, diagnostics of the oxidative stress peculiarities and expression and thereby of inflammation is the basis for formation of the therapy strategy and subsequent monitoring of the pathologic process. Besides, monitoring of peculiarities of the oxidative stress may become an important criterion for differential diagnostics of various obstructive processes (e.g., COPD and BA), which remains to be an actual task of pulmonology.
Unfortunately, the modern methods for diagnostics of inflammation peculiarities and expression either are not sufficiently informative (investigation of peripheral blood) or invasive and traumatic (bronchoscopy, biopsy, lavage) or expensive and inaccessible for every day and everywhere clinical practice (investigation of the exhaled air composition). In particular, nowadays, the widely used analyses of systemic substrates (general analysis of blood, analysis of proteins of acute phase) may not reflect peculiarities of inflammation of organs, first of all due to the fact that these inflammatory changes may be caused by organs and systems of another localization. Besides, inflammation processes in respiratory organs are not always accompanied by inflammatory changes of systemic substrates or due to peculiarities of this inflammation (e.g., allergic inflammation in the respiratory tract at BA) or due to formation of fibrous changes in the locus of disease (e.g., at tuberculosis). The use of substrates of organs (lavage liquid, bioptates of a lung tissue, exhaled breath condensate, exhaled air, sputum, and induced sputum) in diagnostics of inflammation has an advantage over the system (indirect) methods due to considerably higher specificity with respect to the studied localization of inflammation, higher sensitivity and more complete reflection of inflammation process peculiarities. Unfortunately, many substrates of tissues as applied to the respiratory organs may be obtained only by invasive and highly traumatic methods (bronchoscopy, bronchoalveolar lavage, biopsy), which does not make possible continuous monitoring of inflammation and these methods may be used only at high quality clinical centers. Therefore, presently it becomes actual to develop and improve new means of diagnostics based on the analysis of substrates of organs obtained by noninvasive methods (exhaled air composition, exhaled breath condensate, sputum, induced sputum).
The noninvasive analysis based on the control of the exhaled air composition relies on the detection of the oxidative stress products such as nitrogen oxide NO (a component of the AFN-system of protection), pentane, hexane (witnesses of the AFO effect on the lipides of membranes, 8-isoprostane (a product of AFO metabolism), and hydrogen peroxide H2O2. The latter takes a special place among the markers of the oxidative stress. The system of anti-oxidant organism protection (superoxide-dismutasa and other anti-oxidasas) converts highly active radical AFO formed during the oxidative stress to less active molecular forms, first of all to H2O2. The hydrogen peroxide can move from the locus of inflammation into the surrounding media (including the exhaled air) and thus its concentration in the exhaled air and biologic substrates unambiguously reflects the extent of the oxidative stress expression and is considered today as a direct marker of this process. As activation of inflammation as a rule is accompanied by activation of the oxidation processes, the level of H2O2 in exhaled air may serve as a direct quantity criterion of the extent of the inflammation process expression.
The hydrogen peroxide content is most often determined in the exhaled breath condensate (EBC), but its determination is also possible in other biologic substrates: nasal separated, saliva, tear liquid, urine, blood, lavage liquid, and if special equipment is available, directly in the tissue of the studied organ. As H2O2 concentration in exhaled air is directly connected with oxidative processes in the locus of inflammation, and collection of EBC is noninvasive and can be easily made in conditions of any medical institution, determination of the H2O2 content in the EBC is a promising diagnostic criterion at chronic diseases of respiratory organs having inflammatory genesis (COPD, BA, tuberculosis, sarcoidosis, fibrosing alveolitis etc.).
High diagnostic significance of H2O2 content determination in the EBC at COPD and BA has been demonstrated by numerous investigations. However, the presently existing methods of H2O2 determination are difficult to use and expensive. In most investigations the use is made of the fluorometric method of analysis based on the analytical reaction of H2O2 with peroxidasa of horse radish. Unfortunately, presently this method of investigation cannot be used in wide clinical practice. The reason for this is high cost of equipment and consumable materials and difficulties in performing this analysis. That is why this method is used only at research centers and well-equipped clinics. Besides, this method does not allow rapid analyses to be performed, which complicates monitoring of inflammation.
All this shows the necessity of developing a new procedure for H2O2 determination in the EBC and other biological substrates that would be deprived of the above disadvantages. In particular, this is possible due to direct detection of H2O2 in substrates with the help of new generation sensors are developing by us.
The goal of project is to develop a new method for diagnostics of the inflammation process activity in patients suffering from chronic obstructive pulmonary disease using new highly sensitive sensors of hydrogen peroxide, which will make possible rapid, highly accurate, cheap analysis of H2O2 content in biologic substrates. From technical point of view, this project is aimed at creation of sensors and instruments intended for the use in clinical practice in treatment of patients with various obstructive and nonobstructive diseases of respiratory organs.
The proposed method of analysis is based on the use of chemical sensors allowing direct determination of H2O2 in biosubstrates. The presence of the increased hydrogen peroxide content must become a reliable marker of the inflammation activity determining the scope of applied anti-inflammatory therapy and control of its efficiency. The developed sensors are to be used for hydrogen peroxide determination in vitro (EBC) and to be adapted for the use in vivo (directly in diseased organs). In the last case additional sensors are to be developed in the form of miniature biocompatible sensors capable to work in the organism of human as well as special measuring equipment for receiving and processing of signals from them.
The Project applies to the following fields:
- Medical laboratory and clinical investigations.
- Instrument making for medicine.
- Manufacturing technology for special-purpose sensors.
The relevant experience has been gained by the supposed Project contractors:
- Russian Federal Nuclear Centre – All-Russia Research Institute of Experimental Physics, Sarov, Nizhny Novgorod Region;
- Office of State, Central Research Institute of Tuberculosis of the Russian Academy of Medical Sciences, Moscow
- Chemical Department of the M.V. Lomonosov Moscow State University, Moscow.
- Joint Stock Company “Practic NC”, Zelenograd, Moscow.
Specific goals of the Project:
- Development of a new instrument and method for clinical diagnostics of the inflammation process in patients with chronic pulmonary diseases (COPD, BA, tuberculosis, sarcoidosis etc.) based on determination of the hydrogen peroxide content in biosubstrates (exhaled breath condensate, saliva, tear liquid, urine, blood, lavage liquid, directly in lung parenchyma).
- Testing of the developed instrument and method in vivo on experimental animals (mice).
- Clinical testing of the developed method on volunteers: healthy and suffering from chronic inflammatory pulmonary diseases of various natures (COPD, BA, tuberculosis, sarcoidosis and fibrosing alveolitis) with various clinical symptoms of the inflammation process activity.
- Development of the sensor construction suitable for determination of the hydrogen peroxide concentration in biosubstrates.
- Investigation of the metrological and technical characteristics of sensors for hydrogen peroxide in model solutions (in vitro), development of algorithms and methods for processing signals from sensors.
- Development, manufacture and determination of main characteristics of the model analyzer of hydrogen peroxide in biosubstrates. Testing in model solutions (in vitro).
- Manufacturing of model sensors and a model analyzer for determination of the hydrogen peroxide concentration directly in the opening of bronchi, parenchyma of lungs, blood (in vivo), exhaled air (in vitro) with the detection threshold not exceeding 10-7 mole/l.
- Proposals on introduction of the developed diagnostics methods into the clinical practice of pulmonology.
- Proposals on commercialization of the developed sensors, device and diagnostics methods.
The scientific-and-technical approach and methods for performing investigations are based on the data reported in scientific publications of the executors of the project and other investigators [1-38].
The Project satisfies all objectives of the ISTC since it helps a group of Russian scientists, who have knowledge and skills in the field of weapons of mass destruction, re-orient themselves to peaceful activities and assists in solving an important national and international scientific-and-technical problem, namely, development of a new method and means for clinical diagnostics of the inflammation process in patients with chronic lung diseases including patients suffering from COPD, bronchial asthma and tuberculosis.