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Харьковская медицинская академия последипломного образования, Украина
Участник первенства: Национальное первенство по научной аналитике - "Украина";
Открытое Европейско-Азиатское первенство по научной аналитике;
A comparative cytology screening of scraping from buccal mucosa in patients with leukoplakia mollisand healthy persons was made. The increase in 1,9 - 2,3 times of the amount of the cells of advanced stages of differentiation (surface non-squamous and horny) in lesions are determined, that points at the hyperkeratosis and gives a common ground of this pathology with true leukoplakia. The authors regard the predominance of the number of leukocytes and mucus in patients as the presence of exudative component in leukoplakia mollis.
Keywords: leukoplakia, leukoplakia mollis, buccal epithelium, hyperkeratosis, parakeratosis, cytology.
Проведено сравнительное цитологическое исследование соскоба со слизистой оболочки щеки у больных мягкой лейкоплакией и здоровых лиц. Установлено увеличение в очагах поражения в 1,9- 2,3 раза клеток поздних стадий дифференцировки (поверхностных неороговевающих и ороговевших), что указывает на гиперкератоз и сближает данную патологию с истинной лейкоплакией. Преобладание количества лейкоцитов и слизи у больных авторы расценивают как наличие экссудативного компонента при мягкой лейкоплакии.
Ключевые слова: лейкоплакия, мягкая лейкоплакия, буккальный эпителий, гиперкератоз, паракератоз, цитологическое исследование.
Foreword. Leukoplakia mollisis a benign lesion of the oral mucosa. In 1935 Cannon described the benign nevoid epithelial dysplasia of the oral mucosa, which differs in the clinical and histological picture from the then-known similar diseases. This disease was called "nаеvus spongiosus Cannon." Later in the literature the description of similar changes of the oral mucosa appeared under the different names: "family folded dysplasia of the oral mucosa," "white pleated gingivostomatitis", "leukoedema", "buccal biting" . In 1964 B.M. Pashkov and E.F. Belyaeva described similar with паеvus spongiosus Cannonbenign change of the oral mucosa characterized by periodically desquamation of upper epithelium, and called it a leukoplakia mollis. Many experts believe these signs in the mucosa are different clinical variants of the same disease having nevoid character.
The etiology of the disease is unknown. There are cases of hereditary leukoplakia mollis , it is pointed to the relation of the frequency of its occurrence with smoking and drinking hot beverages , to a multifactorial disease , to the chewing of coca leaves. 
Morphological studies of leukoplakia mollis concerned only the histological, histochemical and electron microscopic studies [7, 9, 14].
Information about the cytological characteristics of lesions is scarce. It is indicated, that there is observed a large amount of epithelial cells both enucleated and nucleated in the impression smears . However, this overall picture of cytological parameters of the oral mucosa is a characteristic for different pathological conditions [3, 8]. There is no information about the degree of differentiation of exfoliative buccal epithelium, the presence of other cellular elements.
The aim of the study is to investigate the cytological picture of the lesion with oral leukoplakia mollis tacking into account the degree of epithelium differentiation, the presence of other cells.
Materials and methods. 22 patients were selected for the study. 12 of them had leukoplakia mollis, and 10 have intact mucosa. The materials for the study were the scrapings from lesions in the intermediate area of the buccal mucosa. Before smears the hygienic assessment of the oral cavity according to Fedorov-Volodkina index was performed. At this we selected patients with hygienic index equal to 1, which corresponds to the normal range. The material was transferred to glass slides, the smear was made, dried, fixed in methanol, stained with azur-eosin according to Romanovsky-Giemsa for 30 min. We counted the epithelial cells of various stages of differentiation in smears. In each specimenunder the low magnification (eye lens 10 and field lens 8) the calculation of 300 epithelial cells was made, among them the cells of different stages of differentiation by the proposals of D.E. Lange were studied: 1 - basal, 2 - parabasal, 3 - intermediate cells of 1 type, 4 - intermediate cells of 2 type, 5 - surface cells (with pyknotic nucleus), 6 – horny enucleated cells . The maturation of the buccal epithelium was expressed as an index of cells differentiation (ICD) : ICD = 1a + 2б+ 3в+ 4г+ 5д+ 6e where 1, 2, 3, 4, 5, 6 are the numerals designation of the described above stages of cells differentiation; a, б, в, г, д, еare the percentage of cells of corresponding stage of differentiation.
Statistical processing of materials was made on parametric criteria (share - P, the standard error - p). Statistically significant differences between the indexes of two independent groups was made on nonparametric tests (Clopper-Pearson confidence interval) using the statistical software package AtteStat 10.8.4. for MS Excel.
Findings. The obtained data showed that both patients with leukoplakia mollisand healthy individuals have the presence of cells of 3, 4, 5 and 6 stages of differentiation in cytological specimens(Table).
The composition of the buccal epithelium in test persons, % (Р±р)
Morphological type of epithelium
Stage of differentiation
having leukoplakia mollis
Intermediate of I type
(95% CI: 3.61-5.69)
(95% CI: 2.31-3.42)
Intermediate of II type
(95% CI: 54.79-59.63)
(95% CI: 24.20-27.06)
(95% CI: 34.29-39.01)
(95% CI: 66.43-69.49)
(95% CI: 1.03-2.31)
(95% CI: 3.01-4.25)
Cytological picture was different between the groups of test persons in the quantitative representation of the epithelial cells of different stages of differentiation, hematogenous cellular elements and the quality of background in the specimens.
When leukoplakia mollisthe cells of surface layers predominated in specimens: 1.9 times greater (p <0.05) the cells of surface non-horny epithelium were detected and 2.3 times (p <0.05) greater the surface squamous epithelium were detected. Accordingly the ratio of cells of the intermediate layer of the epithelium changed (Fig. 1). Thus, patients have 2.3 times less (p <0.05) in cells of the 4th stage of differentiation, and 1.6 (p <0.05) in cells of the 3rd stage of differentiation. The total proportion of the intermediate layer of the epithelium in patients with leukoplakia molliswas 28.43 ± 0.50%, and in healthy persons - 61,79 ± 0.87% (p <0.05).
Index of epithelial differentiation in the patients having leukoplakia molliswas 472,29 (ICD = 1х0+2х0+3х2.82+4х25.61+5х67.97+6х3.59), and in healthy persons – 433.37 (ICD = 1х0+2х0+3х4.57+4х57.22+5х36.62+6х1.58).
In cytological specimensthe difference in the number of hematogenous cells was also observed. When leukoplakia mollisamong all cellular elements the leukocyte amount was 33.72 ± 0.64% (95% CI: 32.47-35.00), which is 1.4 times greater than healthy individuals have - 24.06 ± 0.92% (95% CI: 22.7-25.92). In 9 of the 12 specimens from patients with leukoplakia mollisthe leukocyte occupied all the visual field (Fig. 2). In healthy individuals the great quantity of leukocyte was observed in 3 cases of 10.
Fig. 1. Buccal cells: 1 - healthy persons, 2 – having leukoplakia mollis. Stained with azure-eosin. Increasing x 80.
Fig. 2. Cytological picture of patients with leukoplakia mollis. On the background of buccal epithelium there are many leukocytes of different degrees of degeneration. Stained with azure-eosin. Increasing x 80.
Background in specimens also differed. For the microbial landscape with leukoplakia it was indicative the presence ofcoccoid and filamentous flora, and for the intact mucosa it was indicative the predominance of coccoid flora. In the most of scraping specimens (10 of 12) of leukoplakia site it was determined the mucus, sometimes covering the layers of the epithelium (Fig. 3). In scrapes of healthy individuals the mucus was observed in 2 of 10 specimens.
Discussion. The data obtained by the ICD in patients with leukoplakia mollisand healthy individuals differs from those proposed by I.A. Bykova and her co-authors, they are a little below these criteria. The authors believe that in norm the oral ICD can range from 450 to 560. In ulcerative inflammation and hyperkeratosis the index decreases and increases, respectively . The difference between our data can be explained by the fact that we took the material from buccal mucosa using the method of scraping but not touch smear. However, in patients having leukoplakia mollis the ICD ishigher than in healthy individuals. Such excess of index is associated with its shift to the right, that is with an increased content of the cell of late stage of differentiation - the 5th and 6th.
Fig. 3. Cytological picture of patients with leukoplakia mollis. On the surface of epithelial cells there is an accumulation of mucus. Stained with azure-eosin. Increasing x 80
A range of histological studies of leukoplakia mollis revealed the significant parakeratosis, deficiency of full keratinization without formation of enucleated cells [7, 9]. However, it is known that buccal mucosa relates to a lining, not chewing type , so its epithelial layer consists of non-horny cells. The exception is the intermediate zone of cheeks where normally horny epithelium can be found. Therefore, the presence in cytological specimens the epithelium with nucleus, to our opinion, cannot be regarded as parakeratosis, that is an evidence of the physiological differentiation of buccal epithelium. And the increase of horny cellsduring leukoplakia mollis, that we have established, can be considered as a hyperkeratotic reaction of mucosa, which makes this picture rather similar to the true leukoplakia.
The increase of the number of inflammatory cells (leucocytes) in patients with leukoplakia molliscan indicate on the inflammatory component of the pathology. Although according to some data received in histological study there was no inflammatory reaction in the tissue stroma [7, 9].
Conclusions. The foregoing leads to the conclusion: