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Year : 2012  |  Volume : 55  |  Issue : 2  |  Page : 267-268
Microphotometric nuclear DNA analysis of atypical squamous cells of the uterine cervix


Division of Cytopathology, Institute of Cytology and Preventive Oncology (ICMR), I-7, Sector-39, Noida, Uttar Pradesh, India

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Date of Web Publication3-Jul-2012
 

How to cite this article:
Kashyap V. Microphotometric nuclear DNA analysis of atypical squamous cells of the uterine cervix. Indian J Pathol Microbiol 2012;55:267-8

How to cite this URL:
Kashyap V. Microphotometric nuclear DNA analysis of atypical squamous cells of the uterine cervix. Indian J Pathol Microbiol [serial online] 2012 [cited 2020 Feb 20];55:267-8. Available from: http://www.ijpmonline.org/text.asp?2012/55/2/267/97910


Sir,

 Pap smear More Details is the standard screening tool, in low resource settings, used to test for the presence of atypical cells of the uterine cervix that could become intraepithelial lesions or cancerous. The Bethesda system (TBS) 2001 has largely replaced previous classifications of Pap smears and is dividing the smears in two groups- low grade squamous intraepithelial lesions (LSIL) or high grade squamous intraepithelial lesions (HSIL). [1] A third category, atypical squamous cells (ASC) is used to classify minimal cellular changes that do not satisfy the criteria for the LSIL or HSIL. Atypical squamous cells are further divided into atypical squamous cells of undetermined significance (ASC-US) and atypical squamous cells-cannot exclude high grade lesions (ASC-H). The management of ASC-US is a triage with HR-HPV testing and colposcopy is recommended for ASC-H. Deoxy ribonucleic acid (DNA) aneuploidy is another important indicator to predict the biologic potential of squamous intraepithelial lesions. By nuclear DNA quantitation, the intraepithelial lesions could be euploid, polyploid or aneuploid. Changes having euploid or polyploid DNA distribution are most likely related to stimulated hyperplastic, non-neoplastic process secondary to infection, inflammation, repair or other unidentified agents. Those changes having an aneuploid DNA pattern are true neoplastic processes associated with significant alterations in the chromosomes and mitoses [2],[3],[4] . This study was aimed to reveal aneuploid DNA pattern of atypical squamous cells with high risk, HR-HPV DNA positivity.

For the present report 3000 Pap smears were screened and diagnosed as per TBS 2001. A total of 60 (2%) cases were diagnosed as atypical squamous cells (ASC) consisting 50 (1.7%) cases of undetermined significance (ASC-US) and 10 (0.3%) cases of atypical squamous cells- cannot exclude hyperplasia (ASC-H) on the basis of cytomorphological features. The smears were reviewed and atypical squamous cells were marked by glass pencil on the back of glass slide and after destaining, processed for Feulgen staining and DNA ploidy analysis [2] . For control, 50 normal intermediate cells were measured and their mean DNA value was considered as 'Diploid'. About 10-15 atypical squamous cells (ASC) of each category i.e. ASC-US and ASC-H were analyzed per smear for microphotometric DNA analysis and the cases were categorized as diploid, polyploid and aneuploid. Cervical scrapes of all ASC cases were also tested for HR- HPV 16 DNA by PCR amplification [5] .

The 50 cases, cytologically diagnosed as ASC-US showed intermediate squamous cells with mildly enlarged nuclei (2-3 times the size of normal intermediate cell), slightly hyperchromatic, round to oval in shape with minimal irregularities. Amongst them 30 (60%) cases had diploid, 15 (30%) had polyploid and 5 (10%) had aneuploid DNA pattern. 2/5 aneuploid ASC-US cases were positive for HR-HPV 16. The 10 (0.3 %) cases which were cytologically diagnosed as ASC-H showed cells resemble parabasal cells in size, nuclei were hyperchromatic with uneven chromatin and irregular nuclear membrane, amongst them 3 had diploid, 4 had polyploid and 3 had aneuploid DNA pattern. 2/4 polyploid and 2/3 aneuploid ASC-H cases were positive for HR-HPV 16 [Table 1].
Table 1: DNA ploidy analysis of Atypical Squamous Cells (ASC) of
Uterine Cervix


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The cytology classification (TBS) of Pap smears is accepted and used widely. The combination of HPV screening and Pap smear cytology is a powerful test for the detection of potential carcinoma precursor. The DNA aneuploidy has been regarded as an objective marker for progression and so far DNA aneuploidy has not been found in normal or in reactive squamous cells of the cervix. The development of DNA ploidy abnormalities in cervical cells occurs frequently in the presence of HR-HPV infection. The increase in DNA content from diploid to aneuploid in individual cells may signal gross cell cycle disturbances as a result of viral regulation. The virus infection induces regulatory changes, such as endoreplication and delayed cytoplasm differentiation that progress to high morphologic cellular atypia. [3] In this study, the DNA ploidy pattern of ASC-US and ASC-H was demonstrated and it was observed that 5 (10%) of the ASC-US showed aneuploid DNA pattern and 2 (4%) ASC-US were positive for HR-HPV 16, however 4/10 (40%) ASC-H cases with polyploid and aneuploid DNA pattern were positive for HPV 16. The measurement of DNA content conversely provides objective information and increase the positive predictive value of atypical cells for HSIL/cervical carcinoma. This study also reveals that the combination of DNA cytometry with HPV typing of atypical cells could be a strong tool to separate cases of HSIL from those of LSIL.

 
   References Top

1.Solomon D, Davey D, Kurman R, Moriarty A. O'Connor D, Prey M, et al. The 2001 Bethesda system: Terminology for reporting results of cervical cytology. JAMA 2002;287:2114-9.  Back to cited text no. 1
    
2.Kashyap V, Das BC. DNA aneuploidy and infection of human papillomavirus type 16 preneoplastic lesions of the uterine cervix: Correlation with progression to malignancy. Cancer Letters 1998;123:47-52.  Back to cited text no. 2
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3.Bollmann R, Mehes G, Torka R, Speich N, Schmitt C, Bollmann M. Determination of features indicating progression in atypical squamous cells with undetermined significance: Human papillomavirus typing and DNA ploidy analysis from liquid-based cytologic samples. Cancer 2003;99:113-7.  Back to cited text no. 3
    
4.Melsheimer P, Vinokurova S, Wentzensen N, Bastert G, von knebel Doeberitz M. DNA aneuploidy and integration of human papillomavirus type E6/E7 oncogenes in intraepithelial neoplasia and invasive squamous cell carcinoma of the cervix uteri. Clin Cancer Res 2004;10:3059-63  Back to cited text no. 4
    
5.Das BC, Sharma JK, Gopalkrishna V, Luthra UK. Analysis by polymerase chain reaction of the physical state human papillomavirus type 16 DNA in cervical precancerous lesions. J Gen Virol 1992;73:2327-36.  Back to cited text no. 5
    

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Correspondence Address:
Veena Kashyap
Division of Cytopathology, Institute of Cytology and Preventive Oncology (ICMR), I-7, Sector-39, Noida, Uttar Pradesh
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0377-4929.97910

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