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12. Yelins'ka AM, Kostenko VO. Lipid peroxidation and antioxidant protection in periodontal tissues under the action of local pathogenic factor on gums in rats exposed to modeled systemic inflammatory response. Problemy ekologii ta medytsyny. 2017; 21(5-6):62-4.
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ПОКАЗНИКИ ОКИСНО-Н1ТРОЗАТИВНОГО СТРЕСУ У ТКАНИНАХ ПАРОДОНТА ЩУР1В
ПРИ М1СЦЕВОМУ УШКОДЖЕНН1 ЯСЕН НА ТЛ1 ЛШОПОЛIСАХАРИД-ШДУКОВАНОÏ СИСТЕМНОÏ ЗАПАЛЬНОÏ В1ДПОВ1Д1 Слшська А.М., Костенко В. О.
В експеримент на 40 бших щурах дослiджено показники окисно-штрозативного стресу в тканинах пародонта за умов дп на ясна мiсцевого патогенного фактора (5% розчину пдроксиду натрiю) при моделюваннi системно!' запально'1 вiдповiдi (СЗВ). Останню моделювали шляхом внутрiшньочеревного введення лiпополiсахариду Salmonella typhi (трогенал) у дозi 0,4 мкг/кг маси протягом 1-го тижня 3 рази, протягом наступних 7-ми тижшв - 1 раз у тиждень. Показано, що вiдтворення СЗВ супроводжуеться збiльшенням у тканинах пародонта продукцп супероксидного анiон-радикала ( О 2 ), дизрегуляцiею циклу оксиду азоту з одночасною активацiею його NO-синтазно'1 (NOS) i нiтрат / штритредуктазно'1 складових, збiльшенням концентрацп пероксиштрит-йошв. Нанесення лугу на ясна тдвищуе в тканинах продукцiю О 2 i актившсть NOS без порушення функцiонування циклу оксиду азоту (NO) i збшьшення вмiсту пероксинiтрит-йонiв. Аплжащя на ясна 5% розчину пдроксиду натрто на тлi СЗВ викликае збiльшення в тканинах пародонта генерацп О 2 НАДН- i НАДФН-залежними джерелами, дизрегуляцiю циклу NO з тдвищенням концентрацп пероксинiтрит-йонiв, що перевершуе такi при окремому впливi зазначених системного i мiсцевого чинниюв.
Ключовi слова: системна запальна вщповщь, гострий гiнгiвiт, окислювально-нiтрозативного стрес, пародонт.
Стаття надшшла 5.06.18 р.
INDICATORS OF OXIDATIVE-NITROSATIVE STRESS IN PERIODONTAL TISSUES OF RATS EXPOSED
TO LOCAL IRRITATION OF GUMS DURING LIPOPOLYSACCHARIDE-INDUCED SYSTEMIC INFLAMMATORY RESPONSE Yelinska A.M., Kostenko V.O. The study carried out on 40 white rats was aimed at assessing the indices of oxidative-nitrosatitistic stress in periodontal tissues under the condition of combined effect of both local and systemic factors. Local irritation of the gums was induced by the local pathogenic factor (5% sodium hydroxide solution) during the simulation of systemic inflammatory response (SIR) induced by intraperitoneal administration of lipopolysaccharide Salmonella typhi (pyrogenalum) in a dose of 0.4 ^g/kg of weight 3 times during the first week, and once a week for the following 7 weeks. It has been shown that simulation of SIR is accompanied by an increase in the production of superoxide anion-radical ( О 2 ) in periodontal tissues, by the dysregulation of the nitrogen oxide cycle with the simultaneous activation of its NO-synthase (NOS) and nitrate / nitrite reductase components, as well as by the increase in peroxinitrite ion concen-tration. The application of alkali onto the gum increases the .О 2 production and the activity of NOS without disturbing the functioning of the nitric oxide (NO) cycle and increasing in the peroxynitrite ion content in the tissues. The application of 5% sodium hydroxide solution onto the- gums against the background of SIR results in an increase in О 2 generation by NADH- and NADPH-dependent sources in periodontal tissues, the dysregulation of the NO cycle with the increase in peroxinitrite ion concentration that exceeds relevant indices obtained at separate action of these systemic and local factors.
Key words: systemic inflammatory response, acute gingivitis, oxidative-nitrosative stress, periodontium.
Рецензент Срошенко Г. А.
DOI 10.26724/2079-8334-2019-3-69-187-190
UDC 611.24-06:616.45-001.1/3-026.374-036.11]-018-092.9:616-08
G. А. Yeroshenko, М.М. Kopiev, S.M. Bilash, K.V. Shevchenko, \.l. Yachinin I krainian Medical Stomatological Academy, Pollava
RESTRUCTURING OF RAT LUNGS IN ACUTE IMMOBILIZATION STRESS
AND ITS CORRECTION
e-mail: [email protected]
Currently, the search for advanced effective and safe methods of preventing and treatment of stress disorders is one of the priority tasks of medical science. The paper was aimed at the morphological study of the corrective effect of torasemide on rat lungs in experimental acute immobilization stress compared to the stress-protective effect of mexidol. Taking into account the norms of bioethics, the study was involved 30 mature albino male rats. Group I (controls) (n=10) involved animals, exposed to stress without correction; Group II (n=10) involved rats, exposed to stress after administration of mexidol; Group III (n=10) involved rats, exposed to stress that was corrected with torasemide. Correction with pharmacological drugs, especially torasemide, reduces stress effect on the lungs. Histologically, torasemide contributes to retention of the epithelial lining integrity, local visualization of intra-alveolar erythrocytes, and plethora in the capillaries of the interalveolar septa. In the epithelial lining of the bronchi, desquamation of individual cells was sometimes observed. The findings of the study have shown that torasemide has a pronounced stress-protective effect on the lung tissue, which is more powerful than correction with mexidol.
Keywords: lungs, stress, correction, torasemide, mexidol, morphology.
The work is a fragment of the research project "Regularities of morphogenesis of organs, tissues and neurovascular formations in normal conditions, pathology and under the effect of exogenous factors ", state registration No. 0118U004457.
Notwithstanding the long-time history of stress existence and continuous search for novel solutions, the problem of stress remains extremely relevant for the medical and scientific community to date. The issue on finding and choosing the advanced effective and safe methods of the prevention and treatment of stress disorders is one of the priorities for medical investigators [2, 3, 5, 6, 8, 9, 13, 14]. Studies conducted at the Ukrainian Medical Stomatological Academy show that acute immobilization stress causes structural changes
© G.A. Yeroshenko, M.M. Koptev, 2019 , 87
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in the lungs of rats that can be a precursor of the development of pulmonary edema [10, 12, 15]. Therefore, in our opinion, torasemide, a loop diuretic with long-lasting effect, widely used in cardiogenic pulmonary edema, can be used as a potential stress protector. The study of the corrective effect of torasemide on rat lung in acute stress was performed in comparison with mexidol, known as a stress- protective drug.
The purpose of the work was to study the stress-protective effect of torasemide on lungs in experimental acute immobilization stress.
Materials and methods. The total of 30 Wistar albino male rats of 240-260 g body weight aged 8-10 months old were involved into morphological study. Group I (controls) (n=10) involved animals, exposed to simulated acute immobilization stress without correction; Group II (n=10) involved rats, exposed to simulated acute immobilization stress after intraperitoneal administration of stress protective drug mexidol; Group III (n=10) involved rats, exposed to simulated acute immobilization stress after administration of torasemide. Simulated acute immobilization stress was induced by six-hour immobility of rats, lying on the back. Correction procedure included intraperitoneal administration of 100 mg/kg/ body weight mexidol or 0,1 mg torasemide 20 minutes prior the period of immobilization.
The rats were killed by decapitation under thiopentone anesthesia within 2 hours after immobilization. Macroscopic analysis of the lungs and collection of material for histological study has been carried out after dissection of rats' chest. Subsequently, pieces of pulmonary tissue were fixed in 10% neutral formalin and after dehydration in spirits of the ascending densities they were embedded into paraffin according to conventional technique. The microtome specimens were stained with hematoxylin and eosin [1, 7].
The study was performed in compliance with the requirements of international principals of the "European Convention for the Protection of Vertebrate Animals Used for Experimental and Other Scientific Purposes" (Strasbourg,1985) and corresponding Law of Ukraine "For the Protection of Pet Animals" (No.3446-IV, as of 21.02.2006, Kyiv) [4, 11].
Results of the study and their discussion. Examination of lungs after dissection of the chest wall has shown that in experimental animals, exposed to simulated acute immobilization stress without pharmacocorrection (Group I), numerous pleural hemorrhages were observed under the visceral pleura, the number of which was particularly significant on the diaphragm surfaces. The lungs of the rats from Group I had an elastic dough consistency and smooth surfaces. The color of the lungs was red pink with marked numerous polymorphic hemorrhages on its surfaces and in the lung tissue.
Macroscopically, less pronounced stress effect on the lungs with scarce hemorrhages under the visceral pleura and in the lung tissue was noted in rats, administered with mexidol and torasemide.
The histological study showed the adverse effect of acute stress on the lung tissue. The acute stress reaction without pharmacocorrection caused a significant enlargement of alveoli, thinning of interalveolar septa, destruction and detachment of the alveolar epithelium. Numerous alveolar macrophages were noted, sometimes forming continuous chains, their cytoplasm was densely filled with phagocytic material (fig. 1A).
Destructive phenomena were also noted in the intrapulmonary bronchi, manifested by disintegrated epithelial layer with accumulation of cellular detritus and red blood cells in the bronchial lumens, hyperhydration of the mucous membrane, peribronchial leukocytic infiltration. Blood stasis was noted in all sections of the microvasculature; lumens were filled with red blood cells; numerous foci of erythrocyte diapedesis in the interstitium of the lungs and lumens of alveoli were observed (fig. 1B).
Histologically, correction with pharmacological drugs, torasemide, in particular, showed less apparent stress effect on the lungs.
The correction of the outcomes of the effect of acute stress reaction with mexidol showed no desquamation of the alveolar epithelium. Interalveolar septa were thickened and impregnated with erythrocytes, which were also found in the lumens of the alveoli (fig. 2A).
In the wall of small- and medium-sized bronchi a local desquamation of epithelial cells was noted; leukocytic infiltrates
Fig. 1. Alveoli (A) and bronchiole (B) of rat after exposure to simulated acute immobilization stress. Microimage. H&E stain: Lens: 40: Ocular lens:15. A: 1 - alveolar lumen; 2 - alveolar wall; 3 - intra-alveolar macrophage; 4 - desquamated alveolocytes in the alveolar lumens; 5 - macrophages with phagocytic material; B: 1 - lumen of the bronchiole; 2 - epithelium of the bronchiole mucosa; 3 - hyperhydrated lamina propria of the bronchial mucosa; 4 - adventitious membrane; 5 - enlargement of the lateral intracellular fissures; 6 -cellular detritus in the lumen.
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were visualized peri-bronchially (fig. 2B). The arteries were spasmodic; the nuclei of the endothelial cells protruded into the lumens, where hemocytes were not found. Blood stasis and sludge of the red blood cells was detected in the veins.
Histological study has found that epithelial lining of alveoli retained its integrity in a group of animals, administered with torasemide in simulated immobilization stress. The capillaries of interalveolar septa were plethoric, intra-alveolar erythrocytes were locally visualized (fig. 3A). The epithelial lining of the bronchi was solid, but sporadic desquamation of individual cells in the lumen was
Fig. 2. Alveoli and bronchi (B) of rat after exposure to simulated acute ^ -P 1 1 +
immobilization stress and correction with mexidol. Microimage. H&E stain: Lens: 40: noted. Groups °f leukocytes,
Ocular lens:15: 1 - alveolar lumen; 2 - plethora of the vessels and diapedesis of the red containing mainly macrophages blood cells in the interalveolar septum; 3 - local desquamation of the epithelial cells of . "j i-n
the bronchial mucosa; 4 - peribronchial leukocytic infiltration. were detected peri-bronchially.
Hemocytes were found in the arterial lumens. Plasma had heterogeneous optic density. Blood perfusion was unhampered in the venules (fig. 3B).
Apparently, stress causes alterations in rat organs with geodynamic disorders in the adjacent stroma. Subsequently, multiple perivascular hemorrhages and leukocyte infiltrates occurs [3, 10]. The response is typical for many organs, including lungs, and confirmed by the findings of previous studies [12, 15].
Accordingly, the findings [6, 13] showed that administration of Fig. 3. Alveoli ^ and bronchi,(B) of rat after exposure to simulated acuf mexidol to laboratory animals
immobilization stress and correction with torasemide. Microimage. H&E stain: Lens: 40: J
Ocular lens:15: 1 - alveolar lumen; 2 - red blood cells in the arterial lumen; 3 - prevented the development of Selye's
peribronchial leykocytic infiltration. triad, namely, it increased the relative
weight of the thymus, normalized the relative weight of the adrenal glands and prevented ulceration in the
gastric mucosa. The findings of the study revealed the stress-protective effect of mexidol on the lung tissue,
which prevented desquamation of alveolocytes and reduced the intensity of leukocytic infiltration.
However, disturbances of blood perfusion in the microvasculature and the local desquamation of epithelial
cells in the bronchial tree were noted.
Torasemide contributes to minimization of the development of edema of the stroma of the lungs,
prevents the development of hypoxia in the tissue and alterations of organs, thereby providing support for
homeostasis of the lung tissue.
Thus, the study shows that torasemide has a pronounced stress-protective effect on the lung tissue,
more powerful than mexidol. The use of torasemide as an anti-stress drug may be possible after the follow
up thorough studies on the effect of torasemide on other organs, in particular, thymus, adrenal glands,
stomach, liver and kidneys.
Torasemide, a loop diuretic with long-lasting effect, has a pronounced stress-protective effect on the lungs, more powerful than the corrective effect of mexidol. Further research is needed to resolve the issue of the use of torasemide as a stress-protector.
Prospects for further research will encompass the study of stress protective effect of torasemide on other target organs in experimental stress.
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1. Bahriy MM, Dibrova VA, redaktory. Metodyky morfolohichnykh doslidzhen: monohrafiya. Vinnytsya: Nova knyha; 2016. 328 s. [in Ukrainian]
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5. Karnaukh YeV. Antistressovoye kardioprotektornoye deystviye piratsetama pri emotsionalnom stresse po kriteriyu ogranicheniya stress-indutsirovannoy fermentemii i proteoliza. Yeksperimental'na i klinichna meditsina. 2013; (1):43-46. [in Russian]
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9. Omelchenko OYe, Tarasenko LM, Devyatkina TO, vynakhidnyky; Ukrayinska medychna stomatolohichna akademiya, patentovlasnyk. Sposib korektsiyi ultserohennoho efektu hostroho stresu u shchuriv. Patent Ukrayiny № 58721. 26 Kvit 2011. [in Ukrainian]
10. Shkurupiy OA. Eksperymentalne obhruntuvannya rozvytku nekardiohennoho nabryaku lehen u khirurhichnykh khvorykh pry obmezheniy rukhoviy aktyvnosti. Visnyk problem biolohiyi i medytsyny. 2015; 125(4, 2):68-70. [in Ukrainian]
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13. Lutsenko RV, Vakhnenko AV, Vlasova EV. Research of the protection actions of derived 2-oxoindole in acute stress. Wiadomosci lekarskie. 2017;70(1):57-61.
14. Lutsenko RV, Sydorenko AH, Bobyriov VM. Anhedonia at experimental models of chronic stress and its correction. Wiadomosci lekarskie. 2017;70(4):745-750.
15. Pronina OM, Koptev MM, Bilash SM, Yeroshenko GA. Response of hemomicrocirculatory bed of internal organs on various external factors exposure based on the morphological research data. CBiT Meg^HHH Ta öionorii. 2018; 63(1):153-7.
\СТРУКТУРН1 ЗМ1НИ ЛЕГЕНЬ ЩУР1В ПРИ ГОСТРОМУ ШМОБШ1ЗАЦШНОМУ СТРЕСОВ1 ТА IX КОРЕКЦ1Я Срошенко Г.А., Коптев М.М., Биаш С.М., Шевченко К.В., Ячмшь А.1.
У наш час пошук сучасних ефективних i безпечних методiв профшактики та лкування стресових розладiв продовжуе залишатися одним iз прюритетних завдань для медично! науки. Метою роботи було вивчення на морфолопчному рiвнi корегувального впливу торасемщу на легеш щурiв при експериментальному гострому iммобiлiзацiйному стрес у порiвняннi iз стреспротекторною дieю мексидолу. 1з урахуванням норм бюетики, дослщження було виконане на 30 дорослих бших щурах-самцях. I, контрольну групу, склали 10 тварин, яю зазнали стресового впливу без корекцй, до II групи увшшло 10 щурiв, дiя стресу у яких вщбувалася тсля введення мексидолу, у III групи (10 особин) переб^ стресово! реакци проходив на ™ корекцп торасемщом. Корекщя фармаколопчними засобами, особливо торасемщом, зменшуе стресовий вплив на легеш. На пстолопчному рiвнi при використанш торасемщу епiтелiальна вистилка альвеол зберiгала цшсшсть, локально вiзуалiзувалися iнтраальвеолярнi еритроцити, у катлярах мiжальвеолярних перегородок визначалося
повнокров'я. В епiтелiальнiй вистилцi бронхiв подекуди спостерталася десквамацiя окремих клiтин.Проведене дослщження свiдчить, що на легеневу тканину торасемщ мае виразну стреспротекторну дш, потужнiшу за корекцiю мексидолом.
Ключов1 слова: легенi, стрес, корекцiя, торасемщ, мексидол, морфологiя.
Стаття надiйшла 16.10.2018
СТРУКТУРНЫЕ ИЗМЕНЕНИЯ В ЛЕГКИХ КРЫС ПРИ ОСТРОМ ИММОБИЛИЗАЦИОННОМ СТРЕССЕ И ИХ КОРРЕКЦИЯ Срошенко Г.А., Коптев М.Н., Билаш С.М., Шевченко К. В., Ячминь А.И.
В наше время поиск современных эффективных и безопасных методов профилактики и лечения стрессовых расстройств продолжает оставаться одной из приоритетных задач медицинской науки. Целью работы было изучение на морфологическом уровне корректирующего влияния торасемида на легкие крыс при экспериментальном остром иммобилизационном стрессе по сравнению со стресспротекторным воздействием мексидола. С учетом норм биоэтики, исследование было выполнено на 30 взрослых белых крысах-самцах. I, контрольную группу, составили 10 животных, подвергшихся стрессовому воздействию без коррекции, во II группу вошло 10 крыс, влияние стресса на которых происходило после введения мексидола, в III группе (10 особей) стрессовая реакция протекала на фоне коррекции торасемидом. Коррекция фармакологическими средствами, особенно торасемидом, уменьшает стрессовое воздействие на легкие. На гистологическом уровне при использовании торасемида эпителиальная выстилка альвеол сохраняла целостность, локально визуализировались
интраальвеолярные эритроциты, в капиллярах межальвеолярных перегородок определялось полнокровие. В эпителиальной выстилке бронхов иногда наблюдалась десквамация отдельных клеток. Выполненное исследование показывает, что на легочную ткань торасемид оказывает выраженное стресспротекторное воздействие, более мощное, чем коррекция мексидолом.
Ключевые слова: легкие, стресс, коррекция, торасемид, мексидол, морфология.
Рецензент Старченко I.I.
