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 Table of Contents  
ORIGINAL ARTICLE
Year : 2017  |  Volume : 6  |  Issue : 1  |  Page : 1-4

Cytomorphometric analysis of exfoliated buccal mucosal cells in smokers and tobacco chewers


1 Department of Oral Medicine and Radiology, Saraswati-Dhanwantari Dental College and Hospital and Post-Graduate Research Institute, Parbhani, India
2 Department of Oral Medicine and Radiology, Pandit Deendayal Upadhyay Dental College and Hospital, Solapur, Maharashtra, India

Date of Web Publication18-Aug-2017

Correspondence Address:
Abhishek Singh Nayyar
Department of Oral Medicine and Radiology, Saraswati Dhanwantari Dental College and Hospital, Post-Graduate Research Institute, Parbhani - 431 401, Maharashtra
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/ijhi.ijhi_4_17

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  Abstract 


Aims and Objectives: Exfoliative cytology is a minimally invasive procedure that has been used for the detection of early changes in clinically normal oral mucosa of tobacco users. This study highlights the cause-effect relationship between tobacco usage in varied forms and the structural cellular and nuclear alterations in different age groups. Materials and Methods: This study was conducted with different age groups who were tobacco users as a planned, cross-sectional study. Results: The results of the study revealed that the mean nuclear diameter (ND) and cytoplasmic diameter (CD) showed a statistically significant difference in the control group and in smokers and tobacco chewers. Conclusion: Increase in ND and decreased CD, as well as an altered nucleo-cytoplasmic ratio, appear to be due to chronic tobacco usage and cytomorphometric analysis can be used to detect these cellular and subcellular alterations at an early enough stage before actually a frank, morphological change can be appreciated in relation to the oral mucosa.

Keywords: Cytomorphometry, oral mucosa, tobacco


How to cite this article:
Nayyar AS, Kartheeki B, Sinha P, Zakaria KA, Chandrakant CR, Kulkarni A. Cytomorphometric analysis of exfoliated buccal mucosal cells in smokers and tobacco chewers. Int J Histopathol Interpret 2017;6:1-4

How to cite this URL:
Nayyar AS, Kartheeki B, Sinha P, Zakaria KA, Chandrakant CR, Kulkarni A. Cytomorphometric analysis of exfoliated buccal mucosal cells in smokers and tobacco chewers. Int J Histopathol Interpret [serial online] 2017 [cited 2017 Sep 22];6:1-4. Available from: http://www.ijhi.org/text.asp?2017/6/1/1/213271




  Introduction Top


Oral cancers constitute the fourth most common cancers amongst males and the sixth most common cancers in females.[1] The incidence of oral cancers varies amongst different countries of the world and different regions within the same country specifying the role of environmental factors in the pathogenesis of these malignancies. Chronic tobacco usage has been attributed as a major risk factor in the causation of oral cancers. A strong causal association between cancers of the oral cavity and pharynx with the use of tobacco in any form and alcohol are well established now.[2] Majority of oral cancers are diagnosed at advanced stages resulting in a poorer prognosis and survival rates amongst patients at the time of first diagnosis and intervention.[3] In addition, despite advances in the therapeutic options, the morbidity and mortality rates of oral cancers have risen in the past decade leading to increased treatment costs and complications secondary to such advanced treatments.[1] An early diagnosis of oral cancers is, therefore, critical in the successful treatment of the affected patients.[2] Oral exfoliative cytology is a simple, non-invasive and painless procedure that involves microscopic analysis of the cells collected from the surface of the oral mucosa due to their exfoliation owing to a lack of cohesion, a characteristic of malignant cells.[4] However, this technique also has some added disadvantages in the form of the unpredictiveness of the sample obtained, certain unavoidable technical errors and an incorrect interpretation of the findings. With advanced imaging techniques, computerised systems and the use of quantitative techniques to verify the reliability of the results, this procedure is gaining popularity once again.[5] Exfoliative cytology can be used as a diagnostic adjunct in the early detection of such changes even in the absence of obvious clinical manifestations. Easy and fast implementation, adequate diagnostic value, non-invasiveness, cost-effectiveness and reproducibility are some of the definite advantages of this technique.[2] There have been conflicting reports in the literature with regard to the effects of tobacco on oral cavity.[6] The present study was, therefore, undertaken to evaluate the cytomorphometric analysis in such patients so as to predict their potential for future frank malignant conditions so that appropriate and timely intervention could avoid such dreadful sequel in such patients.


  Materials and Methods Top


Forty five individuals with either a habit of smoking or chewing tobacco and fifteen healthy controls were selected from the outpatient department for the study. The study group was in the age range of 20–60 years which was divided into three groups: Group 1 comprising individuals <25 years, Group 2 comprising individuals between 25 and 50 years and a Group 3 comprising individuals >50 years of age.

Inclusion criteria

All individuals having tobacco habit either in the form smoking or chewing tobacco were included in this study group.

Exclusion criteria

Individuals with tobacco-associated oral lesions with clinically evident co-existent haematological disorders were excluded from this study.

Scrapings were made from the buccal mucosa with moistened wooden spatula smeared on to a clear glass slide and immediately fixed with 95% ethanol for a minimum of 15 min. The smears were visualised in ×100 magnification and focused on the stage micrometre scale. In all the cases, the nuclear diameter (ND) and the cytoplasmic diameter (CD) were measured in both horizontal as well as the vertical axes. Only clearly defined cells were measured excluding the clumped or folded cells. 100 cells were measured from each smear and the mean ND and CD were obtained for each case [Figure 1]. The nucleo-cytoplasmic (NC) ratio was, also, calculated for each case.
Figure 1: Nuclear diameter and cytoplasmic diameter measured in both horizontal and vertical axes in micrometre

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  Results Top


This study group was in the age range of 20–60 years which was divided into three groups: Group 1 comprising individuals <25 years, Group 2 comprising individuals between 25 and 50 years and Group 3 comprising individuals >50 years of age. The study individuals had a habit of smoking and/or chewing tobacco. The mean values of ND, CD and NC ratio were calculated amongst these groups [Table 1],[Table 2],[Table 3] and [Graph 1 [Additional file 1] ],[Graph 2 [Additional file 2] ],[Graph 3 [Additional file 3] ]. Statistically significant difference was observed in all the parameters between the control group, smokers and tobacco chewers.
Table 1: Width of nucleus: Descriptive statistics

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Table 2: Width of cells: Descriptive statistics

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Table 3: Nucleo-cytoplasmic ratio: Descriptive statistics

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  Discussion Top


Oral exfoliative cytology is a simple and non-invasive diagnostic adjunct that can be used for an early detection of potentially malignant lesions. Variables such as ND, CD, NC ratio, nuclear shape, continuity of the nuclear membrane, optical density and nuclear texture are assessed by cytomorphometric analysis.[7] All major forms of tobacco are known to have a cause-effect relationship with oral cancers. During malignant transformation, changes occur at the cellular and subcellular levels well before clinical changes become obvious. High-risk potentially malignant lesions can be identified well in early enough stages to reduce the mortality, morbidity and cost of treatment associated with oral cancers.[8] In a study conducted on South Indian population, cytomorphological alterations in the form of a reduction in CD and increase in ND have been reported in buccal squames of tobacco users. In comparison to the non-smokers, there was an average increase of 5–16.5% in the ND as was reported in the smokers.[9] In the present study, significantly altered values were observed for all the three measured parameters (ND, CD and NC ratio) in the group using various forms of tobacco. These changes were directly related to the duration and frequency of habit. Increase in nuclear size is considered to be a cellular adaptation in response to an oral epithelial lesion. As there is decreased turnover rate of the oral epithelial cells which causes the cells to remain in the cell cycle for longer periods, a subsequent delay in the cell division leads to an increase in the nuclear size. The size of the nucleus and cytoplasm decline with aging.[10] The cellular changes seen in the present study can be partly attributed to the fact that tobacco on consumption releases various byproducts such as nitrosamine and nitrosonornicotine which can influence cellular morphometry. In tobacco chewers, there is a close contact of these byproducts, thus, facilitating their infiltration into the mucosa, which might be held responsible for the pronounced cellular changes seen in such patients. Another study reported that the ND of the cells in individuals, who had smoking habits, chewed betel quid and/or practised both these habits, was significantly greater than that of the control group. They also reported that the CD of individuals who chewed betel quid and practised both these habits was significantly smaller than that of the control group.[11] The present study emphasises the importance of an early recognition of cellular alterations caused by tobacco usage. The results of the present study revealed a decrease in CD and an increase in ND and NC ratio in the cells isolated from the buccal mucosa and gingiva of tobacco users which could be used as early indicators of the dysplastic changes in the oral mucosa calling for an early intervention to prevent the development of frank malignant changes.


  Conclusion Top


Increase in ND and decreased CD as well as altered NC ratio appear to be due to tobacco and cytomorphometric analysis can be used to detect these cellular and subcellular alterations at an early enough stage before actually a frank, morphological change can be appreciated in relation to the affected oral mucosa.

Acknowledgement

The authors would like to thank all the patients who contributed in the study without whom this study would not have been feasible.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
  References Top

1.
Ide R, Mizoue T, Fujino Y, Hoshiyama Y, Sakata K, Tamakoshi A, et al. Cigarette smoking, alcohol drinking, and oral and pharyngeal cancer mortality in Japan. Oral Dis 2008;14:314-9.  Back to cited text no. 1
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2.
Orellana-Bustos AI, Espinoza-Santander IL, Franco-Martínez ME, Lobos-James-Freyre N, Ortega-Pinto AV. Evaluation of keratinization and AgNORs count in exfoliative cytology of normal oral mucosa from smokers and non-smokers. Med Oral 2004;9:197-203.  Back to cited text no. 2
    
3.
Sampaio Hde C, Loyola AM, Gomez RS, Mesquita RA. AgNOR count in exfoliative cytology of normal buccal mucosa. Effect of smoking. Acta Cytol 1999;43:117-20.  Back to cited text no. 3
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4.
Diniz-Freitas M, García-García A, Crespo-Abelleira A, Martins-Carneiro JL, Gándara-Rey JM. Applications of exfoliative cytology in the diagnosis of oral cancer. Med Oral 2004;9:355-61.  Back to cited text no. 4
    
5.
Pektas ZO, Keskin A, Günhan O, Karslioglu Y. Evaluation of nuclear morphometry and DNA ploidy status for detection of malignant and premalignant oral lesions: Quantitative cytologic assessment and review of methods for cytomorphometric measurements. J Oral Maxillofac Surg 2006;64:628-35.  Back to cited text no. 5
    
6.
Hashemipour MA, Aghababaie M, Mirshekari TR, Asadi-Shekaari M, Tahmasbi-Arashlow M, Tahmasbi-Arashlow F, et al. Exfoliative cytology of oral mucosa among smokers, opium addicts and non-smokers: A cytomorphometric study. Arch Iran Med 2013;16:725-30.  Back to cited text no. 6
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7.
Einstein TB, Sivapathasundharam B. Cytomorphometric analysis of the buccal mucosa of tobacco users. Indian J Dent Res 2005;16:42-6.  Back to cited text no. 7
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8.
Buch AC, Patel SS, Chandanwale SS, Kumar H, Patel KM, Sunita A. Study of oral exfoliative cytology in tobacco chewers of Western India. Int J Pharma Bio Sci 2014;4:138-43.  Back to cited text no. 8
    
9.
Khot K, Deshmane S, Bagri-Manjarekar K, Warke D, Kotak K. A cytomorphometric analysis of oral mucosal changes in tobacco users. J Nat Sci Biol Med 2015;6 Suppl 1:S22-4.  Back to cited text no. 9
    
10.
Bhisey RA. Chemistry and toxicology of smokeless tobacco. Indian J Cancer 2012;49:364-72.  Back to cited text no. 10
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11.
Ramaesh T, Mendis BR, Ratnatunga N, Thattil RO. The effect of tobacco smoking and of betel chewing with tobacco on the buccal mucosa: A cytomorphometric analysis. J Oral Pathol Med 1999;28:385-8.  Back to cited text no. 11
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    Figures

  [Figure 1]
 
 
    Tables

  [Table 1], [Table 2], [Table 3]



 

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