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Table of Contents   
ORIGINAL RESEARCH  
Year : 2012  |  Volume : 23  |  Issue : 5  |  Page : 561-564
Rose Bengal staining - diagnostic aid for potentially malignant and malignant disorders: A pilot study


1 Department of Oral Pathology & Microbiology, Desh Bhagat Dental College & Hospital, Sri Muktsar Sahib, Punjab, India
2 Department of Oral and Maxillofacial Pathology and Microbiology, Sinhgad Dental College and Hospital, Pune, India
3 Department of Oral and Maxillofacial Pathology and Microbiology, College of dental sciences, Davangere, Karnataka, India

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Date of Submission28-May-2011
Date of Decision25-Nov-2011
Date of Acceptance04-Mar-2012
Date of Web Publication19-Feb-2013
 

   Abstract 

Background: The word "Cancer" itself is sufficient to cause a fear in the minds of people. Early detection of oral potentially malignant and malignant disorders is still a diagnostic challenge for most of the clinicians. In the present study, we have evaluated the efficacy of Rose Bengal staining in the detection of oral premalignant and malignant lesions.
Aim: The aim of the study was to evaluate the efficacy of the Rose Bengal staining for the identification of dysplastic areas clinically, and then correlating the findings with the histological grading of dysplasia.
Methods and Materials: A total of 20 patients were selected for the study. Patients had been evaluated clinically and histopathologically along with the grading of dysplasia, and Rose Bengal stain was painted over the lesional mucosa with the patient's consent. Incisional biopsies were taken from the stained area of the Rose Bengal dye, and studied histopathologically. Grading of dysplasia and intensity of the Rose Bengal staining were found to be correlated. Chi square test was performed and a statistical significance of P<0.001 was observed. Statistical significance was defined as P<0.001.
Results and Conclusion: Grading of dysplasia and the intensity of Rose Bengal staining were directly proportional to each other according to this study. P value was found to be significant. Thus Rose Bengal stain can be used as a diagnostic aid in the detection of oral potentially malignant and malignant disorders.

Keywords: Malignant disorders, potentially malignant disorders, Rose Bengal stain

How to cite this article:
Mittal N, Palaskar S, Shankari M. Rose Bengal staining - diagnostic aid for potentially malignant and malignant disorders: A pilot study. Indian J Dent Res 2012;23:561-4

How to cite this URL:
Mittal N, Palaskar S, Shankari M. Rose Bengal staining - diagnostic aid for potentially malignant and malignant disorders: A pilot study. Indian J Dent Res [serial online] 2012 [cited 2020 Jan 24];23:561-4. Available from: http://www.ijdr.in/text.asp?2012/23/5/561/107326
The word "Cancer" itself is sufficient to cause a fear in the minds of people. Emphasis has been placed on early detection and diagnosis of potentially malignant and malignant disorders. Adjuncts for detection of lesions include toluidine blue staining, exfoliative cytology, use of Vizilite and Velscope etc. As there are some drawbacks associated with each technique, the research for the ideal material that would help in the early detection of oral cancer is still going on. Rose Bengal stain (RB), the 4, 5, 6, 7-tetrachloro-2′,4′,5′,7′-tetraiododerivative of fluorescein, can stain the desquamated ocular epithelial cells. [1] As observed by Norn, with an exposure as brief as 1 second, Rose Bengal predominantly stains the cell membranes, an increasing the concentration or time of exposure produces predominant nuclear staining. [2] RB staining has been even used to delineate the extent of the corneal and conjunctival neoplasms. [3] Therefore, such characteristics of RB stain have prompted the researches to test its potential for the detection of oral precancerous and malignant lesions. [1]

According to the previous studies, lesions highly stained by RB have a higher likelihood to be oral squamous cell carcinoma (OSCC) or epithelial dysplasia (DP), than those showing a mild staining. Thus, RB staining might have the potency to be used as a diagnostic aid to detect oral precancerous or malignant lesions for clinicians. [1]

Diagnosis only by clinical examination may lead to misdiagnosis and therapeutic errors, therefore clinical and histopathological features should always be correlated to obtain the confirmatory diagnosis. Thus, the aim of the present study was to correlate the clinical and histopathological features of potentially malignant and malignant disorders to obtain a confirmatory diagnosis, along with evaluation of the efficacy of Rose Bengal Stain in determining the grade of dysplasia in these lesions. [1]


   Materials and Methods Top


This study was conducted in the Department of Oral and Maxillofacial Pathology, M. M. College of Dental Sciences and Research, Mullana. The study group encompassed a total of 20 cases. The criteria to include the patient in the present study were: (i) white lesions with oral tobacco or betelnut related habits, and (ii) lesions causing a suspicion of malignancy. All the patients were evaluated clinically and provisional diagnosis was obtained.

Staining procedure

  • The patients were asked to rinse their mouth with distilled water to clean the lesions for 1 minute;
  • 1% Rose Bengal solution was applied with a cotton tip for 2 minutes;
  • Patients were asked to rinse their mouth for one minute with distilled water to remove excess RB solution;
  • Oral examination of the location, size, morphology and surface characteristics of the sites stained was carried out, and the intensity of the staining was evaluated. After this, incisional biopsies were taken from the stained area of Rose Bengal dye under infiltration with adrenaline with the patient's consent. After the fixation, processing and hematoxylin and eosin stain, slides were examined histopathologically and grading of dysplasia and intensity of Rose Bengal staining were correlated.
Statistical analysis

Chi square test was performed and a statistical significance of P<0.001 was obtained. Statistical significance was defined as P<0.001.


   Results Top


Out of the 20 cases, 10 cases were diagnosed as homogeneous leukoplakia, 7 cases as speckled leukoplakia, 1 case as verrucous carcinoma and 2 cases as carcinoma of buccal mucosa and floor of the mouth. Out of the 10 cases of diagnosed homogeneous leukoplakia, 8 cases were Rose Bengal staining positive [Figure 1] and [Figure 2] and 2 cases were negative [Figure 3] and [Figure 4]. In [Figure 3] mild coloring of the lesion could have been because of remnants of stain due to inadequate rinsing of the lesion with distilled water after staining. Histopathologically, 9 cases showed mild dysplasia and 1 showed moderate dysplasia. 2 cases of homogeneous Leukoplakia, showing mild dysplasia, were false negative for Rose Bengal staining. Out of the 7 cases diagnosed as speckled leukoplakia, all cases were Rose Bengal staining positive [Figure 5] and [Figure 6]. Histopathologically, 1 case showed mild dysplasia, 5 showed moderate dysplasia and 1 showed severe dysplasia. The case diagnosed as verrucous carcinoma histopathologically was also positive for Rose Bengal staining. Clinically 2 cases diagnosed as carcinoma of buccal mucosa and floor of the mouth were positive for Rose Bengal staining clinically [Figure 7] and [Figure 8]. Further, these cases were histopathologically diagnosed as well differentiated squamous cell carcinoma [Table 1]. When the intensity of staining and grading of dysplasia were compared, it was observed that intensity of staining was more in high grade dysplasia.
Figure 1: Mild staining with Rose Bengal of homogeneous leukoplakia in one of the patients in the study

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Figure 2: Mild dysplasia

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Figure 3: Negative staining with Rose Bengal of homogeneous leukoplakia in one of the patients in the study

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Figure 4: Mild dysplasia

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Figure 5: Positive intense staining with Rose Bengal of non homogeneous leukoplakia in one of the patients in the study

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Figure 6: Moderate dysplasia

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Figure 7: Positive very intense staining with Rose Bengal of Squamous cell carcinoma in one of the patients in the study

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Figure 8: Well differentiated SCC

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Table 1: Histopathological diagnosis of the lesions in the patients in the study

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Efficacy of Rose Bengal staining method

Sensitivity and diagnostic accuracy of RB staining method to detect dysplasia and carcinoma was found to be 90%. p value was found less than 0.001, which was significant.


   Discussion Top


Considering the increased incidence of oral cancer globally, it is crucial to find measures which will help in the early detection and diagnosis of this condition, thus helping in reducing the patient morbidity and mortality. This pilot study shed light on an effective diagnostic aid that can be used for oral potentially malignant and malignant disorders. Norn established the concept that Rose Bengal stains the cells, wherever there is poor protection of the surface epithelium by the preocular tear film. This concept has also been extended to the interpretation of other lesions, such as epithelial dendrites of herpes simplex and zoster, dysplasia or squamous metaplasia of conjunctival squamous neoplasms. [2] Mucus or mucous layer may block the Rose Bengal uptake. A primary epithelial abnormality i.e. dysplasia, metaplasia, virus infected cells or other forms of epithelial keratitis, can render the inability of epithelium to interact with the mucous layer, thus allowing the Rose Bengal staining. False negative results could be due to late clinical expression of genetically induced changes in the cells or inability of the stain to penetrate the deeper layers of epithelium showing the dysplastic changes.

RB (vital dye) [4] staining results also seemed to be more promising in detection of dysplasia in precancerous or clinically benign lesions, when compared with toluidine blue. [1] Ge-fei-Du et al. (2007) concluded in his study that Rose Bengal staining may be better than toluidine blue staining. [1] 1% Toluidine blue is an effective method of picking up malignant changes in premalignant lesions, [5] and Rose Bengal stain can also be used effectively for the same purpose. In case of low grade dysplasia, there would be more possibilities for these lesions to loose out on follow-up as they are often asymptomatic and neglected by patients. In our study, even the cases of mild dysplasia were detected by Rose Bengal staining. Rose Bengal stain had also been used for the staining of cystic disorders of corneal epithelium [6] as well as the diagnostic test for Sjogren syndrome in eyes. [7] RB staining was even used to delineate the extent of corneal and conjunctival neoplasms. [3] Rose Bengal (RB) disodium is a well-known intravenous diagnostic agent that undergoes rapid hepatic excretion in an unmetabolized form; [8],[9] however, in a new formulation it may also be used for chemoablation of localized tumours. It could also benefit patients with metastatic melanoma. [10]

Further studies are required for better understanding the potential of Rose Bengal stain in detecting the oral potentially malignant and malignant disorders. A control group was not used in the study, as the research was done as an attempt to know about the feasibility of the rose Bengal stain in relation to oral potentially malignant and malignant disorders, and with the intention that even low grade dysplasia should not be ignored. In future, further research work can establish the Rose Bengal stain as a valuable diagnostic test in the detection of oral potentially malignant and malignant disorders.

In our study, sensitivity and diagnostic accuracy of RB staining method to detect dysplasia and carcinoma was found to be 90%. These results were comparable with the toluidine blue staining in the previous studies.


   Conclusion Top


Rose Bengal staining can be used as a valuable diagnostic test in the detection of oral potentially malignant and malignant disorders.

 
   References Top

1.Du GF, Li CZ, Chen HZ, Chen XM, Xiao Q, Cao ZG, et al. Rose bengal staining in detection of oral precancerous and malignant lesions with colorimetric evaluation: A pilot study. Int J Cancer 2007;120:1958-63.  Back to cited text no. 1
[PUBMED]    
2.Feenstra RP, Tseng SC. What Is Actually Stained by Rose Bengal. Arch Ophthalmol 1992;110:984-93.  Back to cited text no. 2
[PUBMED]    
3.Wilson FM II. Rose Bengal staining of epibular squamous neoplasms. Ophthalmic Surg 1976;7:21-3.  Back to cited text no. 3
    
4.Kim J. The use of vital dyes in corneal diseases. Curr opin ophthalmol 2000;11:241-7.  Back to cited text no. 4
[PUBMED]    
5.Hegde MC, Kamath PM, Shreedharan S, Dannana NK, Raju RM. Supravital staining: Its role in detecting the early malignancies. Indian J Otolaryngol Head Neck Surg 2006;58:31-4.  Back to cited text no. 5
    
6.Bron AJ, Tripathi RC. Cystic disorders of the corneal epithelium.1 Clinical aspects. Br J Ophthalmol 1973;57:361-75.  Back to cited text no. 6
[PUBMED]    
7.Coll J, Porta M, Prat JR, Cebollada JG, Tomas S. Sjogren's syndrome: A stepwise approach to the use of diagnostic tests. Ann Rheum Dis 1992;51:607-10.  Back to cited text no. 7
    
8.Kubin RH, Grodsky GM, Carbone JV. Investigation of rose Bengal conjugation. Proc Soc Exp Biol Med 1960; 104:650-3.  Back to cited text no. 8
[PUBMED]    
9.Pirotte J. Suitability criteria for compartmental analysis of the plasma clearance curve of exogenous cholephils. 131I rose bengal fulfils these criteria. Biomedicine 1979;30:211-15.  Back to cited text no. 9
[PUBMED]    
10.Thompsona JF, Herseya P, Wachterd E. Chemoablation of metastatic melanoma using intralesional Rose Bengal. Melanoma Res 2008;18:405-11.  Back to cited text no. 10
    

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Correspondence Address:
Nitasha Mittal
Department of Oral Pathology & Microbiology, Desh Bhagat Dental College & Hospital, Sri Muktsar Sahib, Punjab
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0970-9290.107326

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    Figures

  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7], [Figure 8]
 
 
    Tables

  [Table 1]

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