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8 Wschodnioeuropejskie Czasopismo Naukowe (East European Scientific Journal) #4(44), 2019 SMI
UDC: 616.311: 579.23-036.632: 599.323.4
Suslova O. V.
candidate of medical sciences Odessa National Medical University Schneider S.A.
doctor of medical sciences, professor, director of the SE "Institute of Dentistry and Maxillofacial Surgery of the National Academy of Medical Sciences of Ukraine"
Tkachenko E.K.
candidate of biological sciences, head of the experimental pathology Sector of the SE "Institute of Dentistry and Maxillofacial Surgery of the National Academy of Medical Sciences of Ukraine"
Nikolaenko K.V.
laboratory assistant of the experimental pathology Sector of the SE "Institute of Dentistry and Maxillofacial Surgery of the National Academy of Medical Sciences of Ukraine"
BIOCHEMICAL AND CYTOMORPHOLOGICAL CHANGES IN THE ORAL MUCOSA OF RATS UNDER THE INFLUENCE OF CHRONIC NUTRITIONAL DEFICIENCY OF POLYPHENOLS
Abstract. The article examines changes in the oral mucosa of rats in conditions of depriving them of plant food. Chronic deprivation of plant food in the oral mucosa caused increased peroxidation processes, insufficient functioning of antioxidant enzymes, increased elastase activity.
Key words: plant polyphenols, deficiency, lipid peroxidation, rats.
The epithelium of the oral mucosa in the course of its vital activity is affected by various pathogenic factors - stresses of different nature, toxicants, xenobiotic drugs, etc.
The composition of polyphenols (PF) of plant origin lead flavonoids, phenolic acids and their derivatives. They are effective adaptogens that normalize systemic and local regulatory processes.
Cellular and humoral regulators of animal tissues have plant analogs, many of which are found in plants and are edible. Thus, the optimal level of resistance of the tissues of the oral cavity largely depends on the level of food intake containing vegetable PF.
The purpose of the study is to study the effect of long-term insufficiency of plant polyphenols in the diet of experimental animals on the state of the oral mucosa.
Materials and methods.
The experiment was conducted on 14 white rats, females, 7 animals per group. Intact animals were kept on a standard vivarium diet. In the 2nd group, rats were fed on a diet devoid of plant components or a polyphenol-free diet (BDP) [1]. Experimental BPR included: whole wheat flour - 30%, whole milk powder - 30%, starch - 20%, sugar - 15%, sodium chloride - 1%. Alfalfa flour was excluded from the diet [1] as containing vegetable PF. The duration of the experiment was 70 days, after which the rats were killed by total bleeding from the heart under thiopental anesthesia (40 mg / kg). All animal experiments were carried out in accordance with the European Convention for the Protection of Vertebrate Animals used in experiments and for other scientific purposes. The object of biochemical studies was the oral mucosa. The level of LPO processes was assessed by the content of malonic dialdehyde (MDA) [2]; determined the activity of antioxidant enzymes:
glutathione peroxidase (GPO) [3] and superoxide dis-mutase (SOD) [4]. The activity of elastase was determined by the method of [5].
Cheek mucosa (SSCH) was dissected, fixed in formalin and embedded in paraffin. Sections 6-8 ^m thick were stained with hematoxylin and eosin, van gieson picrofuchsin and toluidine blue [6]. The counting of mitoses of epithelial cells was performed in the basal and spinous layers of the system with an increase of 15 x 40 [7]. The mitotic index (MI) was calculated based on the total number of cells counted (3000) and expressed as a percentage. Dual-core epithelial cells were determined in the spinous layer. Erosion of the epithelial layer was determined using a micrometer scale with an increase of 8 x 20. The coefficient of erosion of the epithelium (Keroz. Epit.) Was calculated by the ratio of length: the damaged epithelium / epithelium investigated in arbitrary units (the standard units).
On the sketches of the blood vessels of the micro-circulatory bed (ICR), made with an increase of 15x40, the point method determined the specific area of the wall of the blood vessel with the cells forming it and the specific area of its lumen. According to the ratio of the wall area / lumen area, the coefficient of vascular stenosis (CVS) was calculated.
The results of the experiments were processed using the criterion t the reliability of differences in student.
Research results.
The content of rats on a diet devoid of alimentary polyphenols (BPR) for 70 days caused an increase in lipid peroxidation processes - the MDA content in the oral mucosa increased by 25% (p <0.001) (Table 1). At the same time, GPO activity decreased 2.3 times (p <0.001); SOD activity - by 21% (p = 0.05; Table. 1). An increase in the content of MDA may indirectly indicate an increase in inflammatory events in the tissue
SS-5..Ë Wschodnioeuropejskie Czasopismo Naukowe (East European Scientific Journal) #4(44), 2019 9
studied, and a decrease in the activity of antioxidant enzymes indicates a lack of functioning of the antioxidant protection system.
Table 1
Influence of a non-polished diet on MDA content and enzyme activity in the oral mucosa of the rats
(M ± m; p)
Animal groups MDA content (^mol I Активность
g) GPO (mkat / g) SOD (y.e) elastase (nkat / g)
Intact 89,9±7,02 91,2±0,53 0,70±0,06 1,78±0,13
BPR 112±6,58 39,4±6,35 0,55±0,02 2,23±0,18
р<0,001 р<0,001 р=0,05 р=0,07
Note. In tab. 1-3 indicator of reliability p calculated relatively intact group.
SOSH intact rats had the usual structure. The flat keratinizing epithelium was subdivided into basal and spinous cell layers, passing into the stratum corneum. Spiny layer is represented by cells with light cytoplasm and polymorphic light nuclei: the average internuclear distance is 18.0 ± 0.71 microns. Active keratinization
Table 2
The ratio of the number of mitoses and binuclear cells in the epithelium of the mucous membrane of the
was observed in the surface rows of the spinous layer. The quantitative parameters of the proliferative processes of epithelial cells are as follows: mitotic index (MI) - 1.5 ± 0.22%, the number of binuclear cells - 15 ± 1.1% (Table 2).
Animal groups Mitotic index (MI), (%) The number of binuclear cells, (%)
Intact 1,5±0,22 15,0±1,1
BPR 0,39±0,10 р=0,01 23,0±1,4 р=0,006
Own plate SOSCH is represented by fibrous connective tissue. The main types of cells are fibroblasts and histiocytes. Macrophages and leukocytes were found in single copies. Soch is profusely vascularized.
Blood vessels of the ICR had a typical structure. The packing density of the ICR vessels was 12.8 ± 0.6% in the test volume; the coefficient of preservation of the lumen of blood vessels (Xpc) - 0.57 ± 0.03 (Table 3).
Table 3
The state of the blood vessels of the microcirculatory bed of the mucous membrane of the
Own plate SOSCH is represented by fibrous connective tissue. The main types of cells are fibroblasts and histiocytes. Macrophages and leukocytes were found in single copies. Soch is profusely vascularized.
Blood vessels of the ICR had a typical structure. The packing density of the ICR vessels was 12.8 ± 0.6% in the test volume; the coefficient of preservation of the lumen of blood vessels (Xpc) - 0.57 ± 0.03 (Table 3).
Table 3
The state of the blood vessels of the microcirculatory bed of the mucous membrane of the
Animal groups Vessel Packing Density, (%) Vascular lumen conservation factor, Ksps
Intact 12,8±0,60 0,57±0,03
BPR 8,9±0,50 р=0,004 0,48±0,03 p=0,07
In the group of rats kept on a diet devoid of plant food (BPR), the epithelium of SOSH was distinguished by thickness heterogeneity. Foci of hyperkeratosis, parakeratosis and acanthosis of the spinous layer were found in it. Papillomatous structures in the epithelium were found in the form of single specimens. On the border with keratinized cells, small foci of hydropically modified epithelial cells appeared. They were subject to destruction and turned into vesicles, however, no significant damage to the epithelium was found.
The diet, devoid of plant polyphenols, caused slight erosion of the stratum corneum on the surface of the epithelial layer. The average internuclear cell distance decreased by 25% (p = 0.003).
Under the influence of the diet, the indices of pro-liferative processes in social and nutritional careers changed dramatically. Mitotic cell activity decreased by 74% (p = 0.01); at the same time, the number of binuclear epithelial cells increased by 53% (p = 0.006) as compared with the data of intact groups (Table 2). 7% of mitosis-dividing cells had an atypical form, asymmetric mitoses.
In the connective tissue of the lamina propria, the edema of collagen fibers and intercellular substance
was moderately pronounced. In the blood vessels of the ICR revealed swelling of endothelial cells. The state indicators of the ICR changed in the direction of decrease: Xcps decreased by 16% (trend, p = 0.07), and the packing density of the vessels decreased by 30% (p = 0.004; Table 3).
Conclusion
The diet, devoid of plant polyphenols, had a pathogenic effect on the oral mucosa of the rats — peroxide and destructive processes intensified under conditions of insufficient functioning of the antioxidant protection system.
In the epithelial layer of the rat cheek mucosa, violations of cell division were revealed, which manifested themselves by a threefold decrease in the number of mitoses in the basal layer, a significant increase in the number of binuclear cells or incomplete mitoses. The appearance of foci of dyskeratosis and acanthosis, a decrease in the volume of nuclei speak of impaired cell differentiation. Moderate edema of the intercellular structures and a decrease in the vascular packing density indicate disturbances in the microvasculature of the buccal mucosa.
10 Wschodnioeuropejskie Czasopismo Naukowe (East European Scientific Journal) #4(44), 2019 SMI
Reference
1. Prohonchukov A. A. Rukovodstvo po terapevticheskoy stomatologii / A. A. Prohonchukov, N. K. Zhizhina // Pod red. A. I. Evdokimova. - M.: Meditsina, 1967 - 572 s.
2. Vladimirov Yu.A. , Archakov A. I. Perekisnoe okislenie lipidov v biologicheskih membranah. - M.: Nauka. - 1972. - 230 s. A.S.922637 SSSR. MKI 01 33/48.
3. Sposob opredeleniya aktivnosti glutation-peroksidazyi v biologicheskih tkanyah / V. Pahomova, N. Kozlyanina, G. Kryukova. - Opubl. 25.04.82, Byul. #15. - 2 s.
4. Chevari S. Rol superoksiddismutazyi v okislitelnyih protsessah kletki i metod opredeleniya
eYo v biologicheskom materiale / Chevari S., Chaba I., Sekey Y. // Lab. delo. - 1985. - #11. - S. 678-681.
5. Visser L., Blout E. R. The use of p-nitrophenol-N-tret-butyl-oxycarbonyl-a-alaninate as substrate for elastase / L. Visser, E. R. Blout // Biochem. biophys. Acta. - 1972. - Vol. 268.- №1. - P. 275-280.
6. Merkulov G. A. Kurs patologicheskoy tehniki / G.A. Merkulov. - L.: Lengis, 1969. - 423 s.
7 . Avtandilov G. G. Meditsinskaya morfometriya / G. G. Avtandilov. - M.: Meditsina, 1990. - 384 s. -(Rukovodstvo)
UDC (616.311.2 + 616.314.17-008.1-02): 599.323.4
Savelieva N.N.
doctor of medical sciences, associate professor, Department of Dentistry
Kharkiv National Medical University
Schneider S.A.
doctor of medical sciences, professor director of the SE "Institute of Dentistry and Maxillofacial Surgery of the National Academy of Medical Sciences of Ukraine"
Tkachenko E.K.
candidate of biological sciences, head of the experimental pathology Sector of the SE "Institute of Dentistry and Maxillofacial Surgery of the National Academy of Medical Sciences of Ukraine"
Nikolaenko K.V. laboratory assistant of the experimental pathology Sector of the SE "Institute of Dentistry and Maxillofacial Surgery of the National Academy of Medical Sciences of Ukraine"
STUDY OF THE CONDITION OF THE ORAL MUCOSA OF RATS WITH LONG-TERM
ADMINISTRATION OF NIFEDIPINE
Abstract. The article examines the effect of long-term oral nifedipine on biochemical and cytomorphological changes in the oral mucosa of rats. 1 group intact (5 individuals). The 2nd group (7) received per os nifedipine in a dose of 5 mg / kg body weight of rats for 70 days. Nifedipine in the oral mucosa caused an increase in peroxidation processes, inadequate functioning of antioxidant enzymes, an increase in the activity of the pro-inflammatory enzyme - acid phosphatase. According to morphometric studies revealed violations in the epithelial layer and the lamina propria of the oral mucosa.
Key words: lipid peroxidation, antioxidant enzymes, acid phosphatase, nifedipine, epithelial layer, oral mucosa, rats.
The basis for the preservation of the normal physiological state of the oral mucosa is the constant desquamation of epithelial cells damaged and microbes-colonized. Only with the breakdown of the cell cycle and the synthesis of protective proteins [1] do the processes of a "second line of defense" develop - inflammation.
Some xenobiotics of medicinal nature cause side effects in the tissues of the oral cavity. These substances include antianginal agent nifedipine, which with prolonged use can cause inflammation in the oral mucosa.
The aim of the work is to study the effect of long-term use of nifedipine on the state of the oral mucosa of rats.
Materials and methods
In the experiment taken 12 white rats males 1,5-months. age, which were kept on a full diet of vivarium:
1st group (5 individuals) intact, in the 2nd group (7) rats received a suspension of nifedipine (manufactured by Pharmaceutical Company LLC Zdorovye, Ukraine) in a dose of 5 mg / kg body weight of rats for 70 days.
After slaughter, the blood serum and oral mucosa (oral mucosa) were taken from the animals. The level of POL was assessed by the content of malonic dialde-hyde (MDA) [2]. The activity of antioxidant enzymes was determined: glutathione peroxidase (GPO) [3], glutathione reductase (GR) [4], catalase [5]. Acid phosphatase activity was determined by the method of [6].
For cytomorphological studies, fragments of the buccal mucosa were dissected out in rats, fixed in formalin and embedded in paraffin. Sections 10 ^m thick were processed according to Einarson [7] and used for review morphological and morphometric studies. With a small magnification of the microscope, the coefficient of erosion of the epithelium (CEE) was measured, what
