Improved endocervical sampling with the Cytobrush.

OBJECTIVE: To evaluate the effectiveness of the Ayre wooden spatula, the cotton-tipped swab and the Zelsmyr Cytobrush in obtaining endocervical cells. DESIGN: Cross-sectional comparison study. SETTING: Family practice unit. PATIENTS: All postpubertal, nonpregnant women who underwent a routine Papanicolaou smear during a 7-month period. INTERVENTIONS: The three devices were used in each patient in a randomized sequence. An experienced cytotechnologist blinded to the device used evaluated the slides for overall epithelial cellularity (graded from 0 [acellular specimen] to 12 [overloaded sample]), density (the number of groups of five or more endocervical cells) and size of cell clusters (5 to 10 cells per cluster [score of 1], 11 to 100 [2] or more than 100 [3]). MAIN RESULTS: Samples from 2 of the 136 women were rejected because of improper labelling of the slides or failure to use all three devices. Seventy-six (57%) of the smears obtained with the spatula and 71 (53%) with the swab had no endocervical cells, as compared with only 14 (10%) obtained with the Cytobrush (p = 0.001). The overall cellularity (and standard deviation [SD]) of the smears obtained with the Cytobrush (5.69 [SD 1.17], p = 0.001) and the spatula (5.70 [SD 1.46], p = 0.001) was significantly greater than the cellularity of those obtained with the swab (4.31 [SD 1.17]). The Cytobrush yielded significantly more groups of endocervical cells (109.84 per slide) than either the spatula (4.17) or the swab (6.25) (p = 0.001). The Cytobrush also produced larger cell clusters (1.56 [SD 0.67], p = 0.001) than either the swab (0.83 [SD 1.70]) or the spatula (0.64 [SD 0.67]). CONCLUSIONS: The Cytobrush and the spatula should be used instead of the spatula alone or the spatula and the swab for collecting endocervical cells.     

Comparison of spatula and nonspatula methods for cervical sampling

A comparison between nonspatula (cotton swab and Cytobrush) cervical sampling methods and spatula (wooden Ayre spatula and plastic extended-tip Szalay Cyto-Spatula) sampling methods was made in 109 cases. Based on the presence of endocervical cells, there were statistically significant qualitative differences between the non-spatula methods as well as between the spatula methods, but not between the Cytobrush and Cyto-Spatula smears or the cotton swab and Ayre spatula smears. In all kinds of inflammatory lesions, the spatula samples were more accurate and diagnostic than the nonspatula ones. In all cases of cervical intraepithelial neoplasia and in most cases of squamous metaplasia, the Cyto-Spatula sample was the most accurate. It is concluded that the Szalay Cyto-Spatula method is superior to the other cervical sampling methods because it provides well-preserved cells from both the endocervix and the ectocervix in one smear. The Cytobrush should be used in conjunction with spatula sampling (combination method) for effective sampling of the cervix. The Cytobrush alone is effective mainly for endocervical sampling while the Ayre spatula alone is effective mainly for ectocervical sampling; the cotton swab is ineffective for both endocervical and ectocervical sampling.     

Comparative evaluation of seven cell collection devices for cervical smears

OBJECTIVE: To compare the most commonly used cervical sampling devices. STUDY DESIGN: We examined seven cytology sampling devices (Cytobrush, Cervex brush, Szalay spatula, Papex spatula, WrGKK spatula [main social security agency in Vienna], cotton swab and loop). Eight hundred smears were assessed for even distribution of cells, percentage of slide surface covered with cells, and presence and number of endocervical cells. RESULTS: Even distribution of cells was best with the WrGKK spatula. Percentage of slide surface covered with evaluable cells was best with the Cytobrush. Highest ranking for the presence of endocervical cells was found for the Cytobrush. Cotton swabs and loop showed inferior results in all categories. CONCLUSION: The use of cervical cell sampling devices showing the best cytologic results improves the interpretation and validity of cervical smears. Our results suggest that cotton swabs and loops should not be used for cervical cell sampling.     

Do gelatin-coated slides increase cellular retention in oral exfoliative cytology?

A minimum number of cells is required for an accurate cytologic assessment of oral lesions; however, the cell yield in some oral smears is reduced, relating to the site from which the cells were harvested and to the lesion examined. The use of gelatin coating to increase the retention of cells in smears from such sites was assessed. Cells were obtained from four sites within the oral cavities of 20 patients (10 male and 10 female) with healthy mouths. Using a wooden spatula, two samples were obtained from each site; one was transferred to a gelatin-coated slide, the other to an uncoated slide. For each slide, the cell yield was rated on a scale of 0 (no cells), 1 (few cells) and 2 (many cells). The results were then analyzed using the Wilcoxon sign-rank test. The coated and uncoated slides yielded similar results for most sites, except that the coated slides seemed to be advantageous for samples from the ventral surface of the tongue.     

Cervical smears following laser treatment. Comparison of Cervex brush versus Cytobrush-Ayre spatula sampling

For 802 women at initial follow-up after laser treatment of cervical lesions, 421 smears prepared using the Cervex brush were compared with 381 smears prepared using the combination of a Cytobrush plus an Ayre spatula. The smears were graded for adequacy, the presence of endocervical or metaplastic cells and the presence and degree of epithelial abnormalities. Endocervical or metaplastic cells were seen more often in Cytobrush-Ayre spatula smears (94.5%) as compared with Cervex brush smears (88.8%; P = .004). Also, the number of samples classed as inadequate was significantly greater with Cervex brush smears (4.0%) than with Cytobrush-Ayre spatula smears (0.3%; P = .0003). The number of smears showing dysplasia was too small to detect realistic differences between the two sampling methods. These findings suggest that, in women who have had laser treatment of the cervix, Cytobrush plus Ayre spatula sampling produces better-quality smears than does Cervex brush sampling, with regard to both adequacy and the presence of endocervical cells.     

Application of the micronucleus test to exfoliated epithelial cells from the oral cavity of beedi smokers, a high-risk group for oral cancer

The primary sites for occurrence of oral cancer include the buccal mucosa, tongue, alveolus, palate, lip and the floor of the mouth. In this study, an attempt was made to estimate the cytogenetic damage in different regions of the oral mucosa in people habituated to smoking beedi,which is one of the major forms of tobacco consumption in India and believed to be a major risk factor for oral cancer. By using the micronucleus assay on exfoliated cells from the buccal mucosa, palate and tongue of beedi smokers, we examined an early cellular response to the effect of beedi smoking. A total number of 50 randomly selected male subjects were included in the study. Case and control groups (smokers and non-smokers, respectively) comprised 25 subjects each. The difference in mean micronucleated cell count between cases and controls was significant (P <0.01) for buccal mucosa and palate, but not for tongue. The correlation between age and micronucleus cell count was weak for both cases (r=0.27) and controls (r=0.36).     

Quantitative analysis of AgNOR proteins in exfoliative cytology specimens of oral mucosa from smokers and nonsmokers

OBJECTIVE: To determine the cell proliferation rate and possible effects of cigarette smoking on the oral mucosa lining through analysis of silver-stained nucleolar organizer regions (AgNORs) in exfoliative cytology specimens. STUDY DESIGN: Exfoliative cytology was performed on the left side of the border of the tongue and of the floor of the mouth in 25 smoking patients and 25 nonsmoking patients. The inclusion criterion for smokers was the consumption of more than 20 cigarettes per day for a minimum of 30 years. RESULTS: The slides were stained by histochemical AgNOR method. In the nonsmoking group the mean number of AgNORs per nucleus was 2.732 +/- 0.236 in the tongue border and 2.918 +/- 0.195 in the floor of the mouth. In smoking patients the mean number of AgNORs per nucleus was 3.372 +/- 0.375 in the tongue border and 3.245 +/- 0.237 in the floor of the mouth. CONCLUSION: The results suggest higher cell proliferation quantified by the histochemical AgNOR technique in exfoliative cytology specimens obtained from the oral mucosa lining of smokers presenting no clinical alterations.     

The impact of liquid-based oral cytology on the diagnosis of oral squamous dysplasia and carcinoma

OBJECTIVE: Even though diagnostic oral exfoliative cytology is a useful, economical and practical tool in the diagnosis of oral dysplasia and carcinoma, it is not yet extensively used. The results of conventional exfoliative and liquid-based diagnostic cytology in oral potentially malignant lesions (PML) are herein reported and compared with the histological diagnosis. METHODS: Either conventional (89) or liquid-based (384) exfoliative cytology was used for the diagnosis of oral dysplasia/carcinoma in 473 subjects and the results were compared with scalpel biopsy histology. Cells were collected using a Cytobrush device for conventional smears and with a dermatological curette for the liquid-based cytology. The 'curette technique' also allowed for the collection of 'accidental' tissue fragments, utilized as microbiopsies. RESULTS: Histological diagnosis was squamous carcinoma in 96 of 473 cases, high-grade dysplasia (oral intraepithelial neoplasia two to three) in 24 and other lesions in 353 cases. The smears in the conventional cytology group were inadequate in 12.4%, with an 85.7% sensitivity and a 95.9% specificity. There were 8.8% of inadequate specimens in the liquid-based cytology group; sensitivity was 95.1% and specificity was 99.0%. CONCLUSIONS: Although conventional cytology is useful when diagnosing oral PML (better sensitivity and predictive positive value if compared with the cervical smear test with similar specificity) and can improve the accuracy of histological diagnosis, liquid-based cytology gives better results, as it not only enhances both sensitivity and specificity, but also provides material for further investigation (AgNORs, DNA, microbiopsies, etc.).     

Relation between sampling device and detection of abnormality in cervical smears: a meta-analysis of randomised and quasi-randomised studies.

OBJECTIVE: To assess the diagnostic yield of different sampling devices used in cervical screening. DESIGN: Meta-analysis of randomised and quasi-randomised studies. SETTING: All randomised and quasi-randomised studies comparing the yield of cytological or histological abnormalities when two or more different sampling devices were used. SUBJECTS: 85,000 patients included in 29 studies reported in 28 papers. MAIN OUTCOME MEASURES: Pooled relative risk and 95% confidence interval of the yield of mild dysplasia or worse in smears recovered by each sampling method versus each other method with which it was compared; sensitivity or positive predictive value, or both, of cytological versus histological results in six studies from which sufficient data were available. RESULTS: There were no substantial differences in the yield of cytological abnormalities between the Ayre spatula, the Cytobrush, and the cotton swab used alone. There were also no substantial differences in the yield of cytological abnormalities between the extended tip spatula, the Ayre spatula combined with the Cytobrush or cotton swab, or the Cervex brush. The Ayre spatula, Cytobruah, or cotton swab used alone generally performed significantly worse than the combinations, the extended tip spatula, or the Cervex brush. There were no substantial differences in sensitivity or positive predictive value between the sampling methods. CONCLUSIONS: These results support the use of either the extended tip spatula, a combination of any spatula plus the Cytobrush or cotton swab, or the Cervex brush for cervical screening.     

Cytologic and cytometric analysis of oral mucosa in Alzheimer's disease

OBJECTIVE: To analyze cytomorphologically the buccal mucosa of patients with Alzheimer's disease (AD). STUDY DESIGN: Brush biopsies were obtained from 10 patients with AD and 9 age-matched controls without neurologic symptoms from 3 distinct oral sites. RESULTS: A significant reduction in partially keratinized intermediate (red) cells was observed in the buccal mucosa of the AD group. In the AD group, parabasal cells from the floor of the mouth (p = 0.017) and buccal mucosa (p = 0.058) and red cells,from the tongue dorsum (p = 0.013) and buccal mucosa (p = 0.002), exhibited significantly greater nuclear areas. With regard to the nuclear to cytoplasmic (N:C) ratio, intermediate (red) cells from the buccal mucosa and tongue dorsum of AD individuals showed a decrease in this parameter (p <0.0001), while superficial (yellow) cells (from buccal mucosa) (p= 0.042) and parabasal (blue) cells (from the tongue dorsum) (p = 0.003) exhibited an increased N:C ratio. No significant differences were detected in the cells from the floor of the mouth. CONCLUSIONS: Our findings indicate that cytologic and cytometric changes were detectable in the exfoliative cytology of the buccal mucosa and tongue in the AD group.     

Comparison of Cytobrush sampling, spatula sampling and combined Cytobrush-spatula sampling of the uterine cervix

Since the introduction of the Cytobrush for sampling the uterine cervix, some practitioners have ceased taking a concomitant cervical scraping using a spatula. To examine whether Cytobrush sampling alone is adequate for the diagnosis of cervical lesions, the Cytobrush and spatula samples in 444 smears (most with original diagnoses of at least mild dysplasia) were analyzed separately for the presence of diagnostic cells, endocervical cells and squamous cells. Of the 412 smears showing pathologic findings (mild to severe dysplasia or worse), diagnostic cells were present in 400 Cytobrush samples and in 369 spatula samples; the combination of both samples thus gave a 3% gain in correct diagnoses as compared to use of the Cytobrush samples alone. Another 18 smears would have been underdiagnosed based only on the Cytobrush samples. Endocervical cells were present in 95.3% of the Cytobrush samples and 83.8% of the spatula samples; squamous cells were present in 93.9% of the Cytobrush samples and 96.8% of the spatula samples. Analysis confirmed that it is important that the smear should contain both endocervical and squamous cells. A positive relationship between the absence of squamous cells in the Cytobrush sample and the probability of a false-negative assessment was suggested. It thus seems inadvisable to replace the combination sampling method by Cytobrush sampling alone, which may lead to a false-negative diagnosis.     

Cytologic analysis of alterations induced by Smoking and by alcohol consumption

OBJECTIVE: To analyze cytologically the buccal mucosa of smoking and nonsmoking volunteers to determine what cellular changes are induced by cigarettes and alcohol consumption. STUDY DESIGN: In order to evaluate cellular changes induced by smoking and alcohol consumption, exfoliative cytology was used for the analysis of mucosal smears obtained from the buccal mucosa of 25 smokers and 25 nonsmokers. The number of cigarettes consumed, duration of smoking, presence or absence of alcohol ingestion, ingested alcohol dose and frequency of consumption, and most frequently used type of alcoholic beverage were determined using a questionnaire. Three smears from each individual stained by the Papanicolaou method were analyzed quantitatively and qualitatively under a light microscope by 2 experienced examiners in terms of inflammatory and dysplastic alterations and of the degree of epithelial maturation. RESULTS: Although numerous alterations were observed in smokers they corresponded up to only Papanicolaou class II and were not significantly different from nonsmokers (Mann-Whitney and chi2 tests, p < 0.05). A higher proportion of inflammatory cells (polymorphonuclear and mononuclear cells) were obtained from smokers as compared to nonsmokers, while the proportion of bacteria was similar in the 2 groups. CONCLUSION: The findings indicate that even after a short period of cigarette use and alcohol consumption, inflammatory alterations were detectable on exfoliative cytology of the buccal mucosa in a young group, demonstrating the usefulness of cytology for early detection in smokers.     

Diagnostic accuracy of squamous cervical lesions studied in spatula-cytobrush smears

The cytologically positive cases found in 25,300 cervical smears of spatula samples and 6,168 smears prepared by combined spatula-Cytobrush sampling were analyzed. The diagnostic accuracy (the correlation between the cytologic and histologic diagnoses) was the same for both types of sampling. As to the histologic diagnosis, the rates of severe dysplasia, carcinoma in situ and squamous carcinoma in the spatula-Cytobrush group were more than twice as high as in the spatula group. In the spatula group, the majority of abnormal cells was of the mature type. In the spatula-Cytobrush group, the majority of smears contained a mixture of immature and mature abnormal cells. The more immature lesions, which are often located higher in the endocervical canal, seem to be better sampled by the Cytobrush. The results indicate that the Cytobrush reaches areas that a spatula cannot reach, resulting in a higher diagnostic efficiency.     

A comparison between the Accu-Pap device and the extended-tip wooden Ayre spatula for cervical cytology sampling

The wooden Ayre spatula with an extended endocervical tip was compared with both designs of the plastic Accu-Pap sampler in two series of 100 consecutive patients to compare their adequacy in obtaining samples for cervical cytology. There was no significant advantage noted in the smears taken by either spatula. In terms of the sequence of smears, the second smear generally contained more endocervical cells and less often showed an absence of diagnostic cells.     

Cytologic and DNA-cytometric early diagnosis of oral cancer.

OBJECTIVE: The aim of this prospective study was to report on the diagnostic accuracy of conventional oral exfoliative cytology taken from white-spotted, ulcerated or other suspicious oral lesions in our clinic. In addition we checked DNA-image cytometry as an adjuvant diagnostic tool. Our hypothesis is that DNA-aneuploidy is a sensitive and specific marker for the early identification of tumor cells in oral brushings. STUDY DESIGN: 251 cytological diagnoses obtained from exfoliative smears of 181 patients from macroscopically suspicious lesions of the oral mucosa and from clinically seemingly benign oral lesions which were excised for establishing histological diagnoses were compared with histological and/or clinical follow-ups of the respective patients. Additionally nuclear DNA-contents were measured after Feulgen restaining using a TV image analysis system. RESULTS: Sensitivity of our cytological diagnosis on oral smears for the detection of cancer cells was 94.6%, specificity 99.5%, positive predictive value 98.1% and negative predictive value 98.5%. DNA-aneuploidy was assumed if abnormal DNA-stemlines or cells with DNA-content greater 9c were observed. On this basis the prevalence of DNA-aneuploidy in smears of oral squamous cell carcinomas in situ or invasive carcinomas was 96.4%. Sensitivity of DNA-aneuploidy in oral smears for the detection of cancer cells was 96.4%, specificity 100%, positive predictive value 100% and negative 99.0%. The combination of both techniques increased the sensitivity to 98.2%, specificity to 100%, positive predictive value to 100% and negative to 99.5%. CONCLUSIONS: Brush cytology of all visible oral lesions, if they are clinically considered as suspicious for cancer, are an easily practicable, cheap, non-invasive, painless, safe and accurate screening method for detection of oral precancerous lesions, carcinoma in situ or invasive squamous cell carcinoma in all stages. We conclude that DNA-image cytometry is a very sensitive, highly specific and objective adjuvant tool for the early identification of neoplastic epithelial cells in oral smears.     

Comparison of the Cytobrush®, dermatological curette and oral CDx® brush test as methods for obtaining samples of RNA for molecular analysis of oral cytology

M. D. Reboiras‐López, M. Pérez‐Sayáns, J. M. Somoza‐Martín, P. Gayoso‐Diz, F. Barros‐Angueira, J. M. Gándara‐Rey and A. García‐García Comparison of the Cytobrush^®, dermatological curette and oral CDx^® brush test as methods for obtaining samples of RNA for molecular analysis of oral cytology Objective: Interest in oral exfoliative cytology has increased with the availability of molecular markers that may lead to the earlier diagnosis of oral squamous cell carcinoma. This research aims to compare the efficacy of three different instruments (Cytobrush, curette and Oral CDx brush) in providing adequate material for molecular analysis. Methods: One hundred and four cytological samples obtained from volunteer healthy subjects were analysed using all three instruments. The clinical and demographical variables under study were age, sex and smoking habits. The three instruments were compared for their ability to obtain adequate samples and for the amount of RNA obtained using quantitative real‐time polymerase chain reaction (PCR‐qRT) analysis of the Abelson (ABL) housekeeping gene. Results: RNA of the ABL gene has been quantified by number of copies. Adequate samples were more likely to be obtained with a curette (90.6%) or Oral CDx (80.0%) than a Cytobrush (48.6%); P < 0.001. Similarly, the RNA quantification was 17.64 ± 21.10 with a curette, 16.04 ± 15.81 with Oral CDx and 6.82 ± 6.71 with a Cytobrush. There were statistically significant differences between the Cytobrush and curette (P = 0.008) and between the Cytobrush and OralCDx (P = 0.034). There was no difference according to the demographical variables. Conclusions: Oral exfoliative cytology is a simple, non‐invasive technique that provides sufficient RNA to perform studies on gene expression. Although material was obtained with all three instruments, adequate samples were more likely to be obtained with the curette or Oral CDx than with a Cytobrush. The Oral CDx is a less aggressive instrument than the curette, so could be a useful tool in a clinical setting.     

Controlled trial of a new cervical spatula.

A wooden spatula was designed to scrape the varied distribution of epithelial abnormalities of the cervix as seen at colposcopy. The efficiency of the spatula in obtaining dyskaryotic cells and improving the cellular quality of smears was compared with that of the Ayre spatula in a controlled trial. More than 17,000 smears were taken from women aged 14-86 years by more than 200 smear takers from 74 centres. Twenty two per cent more dyskaryotic smears were obtained with the trial spatula, and the cellular quality of the smears was improved in all age groups. Although it was associated with a slightly increased risk of bleeding, 83% of users preferred the trial spatula.     

Quantitative cytomorphologic analyses of Papanicolaou-stained smears from oral lichen planus

OBJECTIVE: To apply quantitative techniques to cytologic smears in order to improve the diagnostic sensitivity of cytology in the detection of malignant change in the oral cavity. STUDY DESIGN: Eighty-two patients with lesions of oral lichen planus were investigated. A total of 247 Papanicolaou-stained buccal smears underwent nuclear and cytoplasmic area measurements using a TV image analyzer. RESULTS: The cytomorphologic results of this study suggest that lesions of oral lichen planus contain smaller cells and nuclei than those observed in clinically normal oral mucosal smears. There was a statistically significant reduction in nuclear area (P < .001) and cytoplasmic area (P < .05) for the oral lichen planus smears. CONCLUSION: The TV image analysis system could provide a valuable method of quantifying cell changes in oral smears collected from oral mucosa as a method of monitoring lesions of oral lichen planus.     

Local variations in the staining patterns of keratin antigens and blood group antigens in normal rodent oral epithelia

Summary All rodent oral epithelia are orthokeratinized. However, morphological, immunohistochemical and biochemical studies have shown that regional differences exist. In the present study, intraregional variations in differentiation patterns of rat oral epithelia are demonstrated using monoclonal anti-keratin antibodies AE1 and AE2 and antibodies to blood group antigens B and H. Well-defined areas of rat buccal and hard palate epithelium differed from the general staining patterns of these epithelia. These areas were associated with a papillary surface contour. These local variations were not found in the strain of mice examined. The results suggest that physiologically different vertical compartments of keratinocytes exist within one and the same region of rat oral mucosa, a phenomenon previously recognized in detail only in the epithelium of dorsal tongue. The papillary structures may have some functional significance related to the processing of food similar to that suggested for lingual filiform papillae.     

The surface structure of the completely and incompletely orthokeratinized oral epithelium in the rat: a light, scanning and transmission electron microscope study.

Mucosa from the hard and soft palates, molar gingiva, cheek and dorsal surface of the tongue of the rat was examined in the light microscope, following Mallory's triple connective tissue stain, and in the scanning and transmission electron microscopes. The epithelium covering the hard palate, gingiva, the smooth band of mucosa at the junction of the hard and soft palates, intermediate zones of the soft palate, fungiform papilla-like structures in the central zone of the soft palate, the fungiform papillae, and the more superficial part and posterior surfaces of the filiform papillae of the tongue all exhibited complete orthokeratinization. The oral surfaces of the epithelial cells in all these areas had a honeycomb pattern of interconnecting ridges surrounding depressions. Imprints of the overlying cells that had been desquamated were apparent, and the lateral boundaries between the cells were formed by two raised ridges separated by a gap. The epithelium covering the cheek, central zone of the soft palate apart from the fungiform papilla-like structures, lateral zones of the soft palate, gingival crevice, and the mucosa between the fungiform and filiform papillae of the tongue all exhibited incomplete orthokeratinization. The oral surfaces of the epithelial cells in all these areas were relatively smooth and did not exhibit a honeycomb pattern of interconnecting ridges. Imprints of the overlying cells that had been desquamated and the lateral boundaries between the cells were only very occasionally found. In the transmission electron microscope the outlines of the cells were compatible with the surface patterns seen in the scanning electron microscope. The possible relationships between the degree of orthokeratinization and ultrastructure of the various epithelia are discussed.