The Role of p16/Ki67 Dual Staining in Cervical Cancer Screening.

Cervical cancer screening has enabled a decrease in the incidence and mortality of cervical cancer. Various screening modalities have been studied to date. In many countries, screening is still based on cervical cytology, where cervical cells obtained either on glass or in a liquid medium are examined under a microscope. However, the fact that the vast majority of cervical cancers are a result of persistent infection with high-risk human papillomaviruses (hr-HPV) has led to the implementation of primary HPV screening in many countries. Taking into consideration the fact that the majority of HPV infections are transient and do not cause cervical precancer, effective triage methods are needed to prevent an increase in colposcopy referrals. Among these, the most extensively investigated are HPV genotyping, HPV methylation, and p16/Ki67 dual staining. In this manuscript, we briefly summarize the current knowledge regarding different screening strategies for the prevention of cervical cancer, with a focus on p16/Ki67 dual staining. In addition, we provide an explanation regarding the rationale for the use of various screening modalities based on the molecular biology of cervical cancer and cervical precancerous lesions.

P16/Ki-67 Immunostaining in the Triage of Postmenopausal Women With Low-Grade Cytology Results.

OBJECTIVE: The interpretation of postmenopausal smears and the gynecological treatment of these patients can often be difficult. The objective of this study was to assess the performance of p16/Ki-67 dual-stained cytology as a triage of atypical squamous cells of undetermined significance and low-grade intraepithelial lesion cytology results in postmenopausal women. METHODS: All consecutive atypical squamous cells of undetermined significance and low-grade intraepithelial lesion smears in 1-year period were collected and p16/Ki-67 immunostaining was performed retrospectively. The results were compared with histology results or long-term cytology follow-up in cases with no biopsy. RESULTS: The sensitivity of p16/Ki-67 immunostaining for the detection of cervical intraepithelial neoplasia (CIN) 2 and CIN 3 was 57.1% and 85.0%, respectively. The specificity for the detection of CIN 2 was 94.3% and CIN 3 92.4%. Negative predictive values for the detection of CIN 2 and CIN 3 were 96.3% and 99.6%, respectively. CONCLUSIONS: Dual p16/Ki-67 immunostaining is a useful additional method in postmenopausal patients with low-grade cytology. Considering the high specificity and negative predictive value in our study, we believe that it could be helpful in avoiding unnecessary referrals to colposcopy and thus reduce the cost of the program.

Triaging Atypical Squamous Cells-Cannot Exclude High-Grade Squamous Intraepithelial Lesion With p16/Ki67 Dual Stain.

OBJECTIVES: The management of women with Pap cytology results categorized as "atypical squamous cells, cannot exclude high-grade squamous intraepithelial lesion" (ASC-H) can often be challenging. We assessed the performance of p16/Ki67 dual-stained cytology as a potential triage tool for the detection of biopsy-confirmed high-grade squamous intraepithelial lesions (HSILs) in women with ASC-H. MATERIALS AND METHODS: Women with ASC-H were followed for a period of 36 months by repeat cytology and colposcopy. p16/Ki67 dual staining was performed retrospectively on the initial Pap cytology slide with ASC-H interpretation. Results were compared with the final histological diagnosis and/or cytological follow-up for at least 1 year. All outcomes were defined as clinically significant at the threshold of HSIL (cervical intraepithelial neoplasia grade 2 or worse). RESULTS: One hundred sixty-nine cases fulfilled all conditions to be included in the study group. The rate of histologically confirmed HSIL was 57.4%, low-grade squamous intraepithelial lesion was diagnosed in 17.7% of the patients and normal histology or follow-up in 24.9% of the patients. Overall sensitivity and specificity of p16/Ki67 dual stain were 95% and 72%, respectively. Overall positive likelihood ratio of p16/Ki67 in detection of HSIL was 3.41, considerably increasing pretest probability from 57% to posttest probability of 82%. CONCLUSIONS: A substantial subset of patients with ASC-H interpretations is associated with an appreciable risk of clinically significant cervical disease. p16/Ki67 dual stain can provide additional valuable information that may lead to higher-quality management of women with ASC-H, especially when initial colposcopy or biopsy results do not show HSIL lesions.

Results of total hip arthroplasty using a bionic hip stem.

PURPOSE: The trabecular-orientated bionic hip stem was designed to mimic the natural force transmission through the femur in total hip arthroplasty, resulting in supposedly longer prosthesis survivability. The aim of this study was to compare the second-generation bionic hip stem to a standard uncemented hip stem. METHODS: A group of 18 patients (21 hips) who underwent total hip arthroplasty with a bionic stem (bionic group) was compared with a historic group of 12 patients (12 hips) treated with standard anatomic hip stem (control group). During the first year after the procedure, the densitometric measurements of the bone around the prosthesis were taken. Radiographic and clinical assessments were additionally performed preoperatively and at the three month, six month, one year and three year follow-ups in the bionic group. RESULTS: In the bionic group, one patient was revised for aseptic loosening and 16 patients (19 hips) were available to the final follow-up. A significant decrease of bone mineral density was found in Gruen zones 3, 4 and 5 in the bionic group, and in zone 7 in both groups. The bionic group had a significantly higher bone mineral density in Gruen zone 1 at the one year follow-up. At the final follow-up, all prostheses were radiologically stable in both groups. CONCLUSIONS: Provided that a good implant position is achieved, comparable short-term results can be obtained using a bionic stem. Still, a decrease of bone mineral density in Gruen zone 7 occurred in both groups. Further studies are required to determine survivability of the bionic stem.

P16/Ki67 Dual Staining in Glandular Cell Abnormalities of the Uterine Cervix.

Very limited information exists about the role of p16/Ki67 dual staining on glandular cells in detecting glandular precancerous lesions and cervical adenocarcinoma. In this study, we investigated the diagnostic accuracy of p16/Ki67 dual staining for the detection of glandular and squamous lesions on the uterine cervix and for cancer of the upper reproductive tract. We performed a retrospective analysis of prospectively collected data on 96 patients with glandular cell abnormalities. We analyzed the diagnostic accuracy of p16/Ki67 dual staining for atypical glandular cells, not otherwise specified (AGC-NOS); atypical glandular cells, favor neoplastic (AGC-FN); adenocarcinoma in situ (AIS); and A-CA (cervical adenocarcinoma). A separate analysis for the detection of squamous precancerous lesions and squamous-cell carcinoma (CIN3+) and for cancer of the upper reproductive tract (EC/OC) was performed. Among patients who had normal histology or a low-grade lesion on final analysis, only 8.5% had positive dual staining. On the other hand, 85.7% of patients with AIS+ on final histology had positive dual staining. The respective specificities of p16/Ki67 dual staining on AGC-NOS for the detection of AIS+ (adenocarcinoma in situ or cervical adenocarcinoma), CIN3+ and EC/OC were 91.5%, 88.7% and 86.4%. High specificity values of p16/Ki67 dual staining on cervical smears labelled as AGC-NOS for the detection of CIN3+ and AIS+ suggest that this method might be a useful addition in cervical cancer screening.

Identification of women with high grade histopathology results after conisation by artificial neural networks.

BACKGROUND: The aim of the study was to evaluate if artificial neural networks can predict high-grade histopathology results after conisation from risk factors and their combinations in patients undergoing conisation because of pathological changes on uterine cervix. PATIENTS AND METHODS: We analysed 1475 patients who had conisation surgery at the University Clinic for Gynaecology and Obstetrics of University Clinical Centre Maribor from 1993-2005. The database in different datasets was arranged to deal with unbalance data and enhance classification performance. Weka open-source software was used for analysis with artificial neural networks. Last Papanicolaou smear (PAP) and risk factors for development of cervical dysplasia and carcinoma were used as input and high-grade dysplasia Yes/No as output result. 10-fold cross validation was used for defining training and holdout set for analysis. RESULTS: Bas eline classification and multiple runs of artificial neural network on various risk factors settings were performed. We achieved 84.19% correct classifications, area under the curve 0.87, kappa 0.64, F-measure 0.884 and Matthews correlation coefficient (MCC) 0.640 in model, where baseline prediction was 69.79%. CONCLUSIONS: With artificial neural networks we were able to identify more patients who developed high-grade squamous intraepithelial lesion on final histopathology result of conisation as with baseline prediction. But, characteristics of 1475 patients who had conisation in years 1993-2005 at the University Clinical Centre Maribor did not allow reliable prediction with artificial neural networks for every-day clinical practice.

Comparison of Conservative Treatment of Cervical Intraepithelial Lesions with Imiquimod with Standard Excisional Technique Using LLETZ: A Randomized Controlled Trial.

(1) Background: There are limited data on the success of conservative treatment of high-grade cervical squamous intraepithelial lesions (HSIL) with imiquimod directly compared to standard of treatment with LLETZ. (2) Methods: Patients aged 18-40 with histological HSIL (with high-grade cervical intraepithelial neoplasia, CIN2p16+ and CIN3), were randomly assigned to treatment with imiquimod or LLETZ. The primary outcome was defined as the absence of HSIL after either treatment modality. The secondary outcomes were the occurrence of side effects. (3) Results: 52 patients were allocated in each group and were similar regarding baseline characteristics. In the imiquimod group, 82.7% of patients completed treatment, which was successful in 51.9%. All patients in the LLETZ group completed treatment, which was successful in 92.3% (p < 0.001). In the subgroup of CIN2p16+ patients, treatment with imiquimod was not inferior to LLETZ (73.9% vs. 84.2%, p = 0.477). During and after treatment, no cases of progression to cancer were observed. Side effects and severe side effects (local and systemic) were more prevalent in the imiquimod than in the LLETZ group (88.5% vs. 44.2% (p-value < 0.001) and 51.9% vs. 13.5% (p-value < 0.001), respectively). (4) Conclusion: Generally, in patients with HSIL, LLETZ remains the gold standard of treatment. However, in a subgroup analysis of patients with CIN2p16+, the success rate was comparable between the two treatment modalities. Due to the prevalence of side effects, the treatment compliance with imiquimod use may, however, present a clinically important issue.

Case report: femoral neuropathy secondary to total hip arthroplasty wear debris.

Femoral nerve palsy after THA is well known, but delayed palsy is rare. We describe a 58-year-old man who had progressive thigh pain, weakness, and numbness develop 13 years after cementless arthroplasty of his left hip. Plain radiographs showed substantial liner wear. MRI of the lumbar spine was unrevealing and EMG showed a peripheral neurogenic process involving the left femoral nerve. The large intrapelvic cystic mass was confirmed by an abdominopelvic CT scan. Percutaneous aspiration of the cyst was performed. Cultures of the fluid were negative and cytopathologic examination showed necrotic debris without malignant cells. Biopsy revealed necrosis and abundant foreign body granulation tissue with polarizable debris. During surgical removal of the cyst, a defect of the inner acetabular wall was noted. After subsequent revision arthroplasty with allograft bone, the patient's clinical symptoms improved and his EMG returned to normal.

Rapid prescreening as a quality assurance measure in cervical cytology.

OBJECTIVE: To assess the diagnostic validity of rapid prescreening (RPS) as a quality control method in our laboratory. STUDY DESIGN: All consecutive routine conventional Pap smears (n = 2,521) underwent RPS before full screening. Thirty to 60 seconds were allowed to prescreen each slide. The threshold for pathologic smear was atypical squamous cells of undetermined significance (ASCUS) or more. Smear adequacy, infections and presence of endometrial cells, if observed, were not noted on the worksheet. All smears subsequently underwent routine full screening. Results of RPS and full screening were compared. RESULTS: Of a total of 2,521 smears, 294 were diagnosed as abnormal (ASCUS and more) after final diagnosis. Of those, 158 were also detected on RPS. The average sensitivity of RPS was 62.2%. Twenty-five cases were detected on RPS only, among them 15 ASCUS, 8 low-grade squamous intraepithelial lesions and 2 high-grade squamous intraepithelial lesions. CONCLUSION: RPS is an efficient quality control method. It reduces false negative rate, but in our case, it affects the work flow in our laboratory, and in case of its implementation to the routine praxis the daily workload of 50 smears per day should be lowered.

Interobserver variability and accuracy of p16/Ki-67 dual immunocytochemical staining on conventional cervical smears.

BACKGROUND: p16/Ki-67 dual immunocytochemical staining (DS) has been proven as a sensitive and specific test for triage of HPV positive women with good reproducibility and accuracy. However, implementation of the test into an organized screening program (OSP) is not easy. The aims of this study were to compare the performance and agreement of DS results among three Slovenian cytopathological laboratories involved in the national OSP, and to define cases where staining results can be difficult to interpret. METHODS: Cervical smears were obtained for DS from 129 women referred to colposcopy. Smears were evaluated blindly in three laboratories by a cytotechnologist and a cytopathologist after initial training. Results were positive, suspicious, negative or inadequate. Five characteristics of DS staining were recorded. After primary evaluation, an extensive expert-led additional training was undertaken, including a discussion of difficult cases and a practical exam. Smears were re-evaluated and results compared to primary evaluation. RESULTS: After the additional training, the overall percentage of agreement among the three laboratories increased from 77.5 to 89.9% and kappa increased from 0.70 to 0.86. Sensitivity for CIN2+ increased in two laboratories, to 90.5 and 85.7%, without the loss of specificity (75.8%). In one laboratory, the sensitivity slightly decreased from 90.5 to 88.9%, but the specificity increased from 63.6 to 68.2%. Difficult cases had significantly less DS cells, weak intensity of p16 staining, suboptimal cell morphology and background staining compared to positive cases. CONCLUSION: Additional expert-led training and discussion of difficult cases are necessary for accurate interpretation of DS in laboratories involved in OSP. The most difficult cases were those with single stained cells and weak p16 staining. Training protocol for safe implementation of p16/Ki-67 DS in OSP is proposed.

Postmenopausal vaginal atrophy correlates with decreased estradiol and body mass index and does not depend on the time since menopause.

OBJECTIVE: To evaluate the relationship between morphologic cell characteristics in Papanicolaou (Pap) smears and serum estradiol, body mass index (BMI) and the time elapsed since menopause. Study design. In 92 women Pap smears were grouped into atrophic and mature cell patterns and compared with estradiol, BMI and the time since menopause. RESULTS: Forty-one patients with mature cell pattern were on average 7.1 years from menopause and 51 patients with atrophic pattern 8.2 years, but this difference was not significant. Estradiol in patients with mature cell pattern was significantly higher (52.1 +/- 48.5 pmol/l) than in patients with atrophic pattern (25.6 +/- 40.0 pmol/l). Similarly, BMI was significantly higher (27.9 +/- 4.2 kg/m(2)) in patients with mature cell pattern than in patients with atrophic pattern (25.7 +/- 3.8 kg/m(2)). There was no significant correlation between the time since menopause and estradiol among patients with mature and atrophic cell pattern. The same was true for the correlation between the time from menopause and BMI in patients with mature and atrophic pattern. CONCLUSIONS: Estradiol and BMI are associated with vaginal cell maturation and atrophy in postmenopausal women. Vaginal cell atrophy does not depend on the time since menopause.

Clinicopathological characteristics of cervical cancer between 2003 and 2005, after the introduction of a national cancer screening program in Slovenia.

OBJECTIVE: An organized cervical cancer (CC) screening program was introduced in Slovenia in 2003. With the purpose of clinical audit we analyzed the clinicopathological characteristics of CC patients for the period between 2003 and 2005. STUDY DESIGN: The retrospectively collected data of 450 CC patients were presented at three Advisory Boards of Gynecologic Oncology in Slovenia. They were stratified for stage, tumor characteristics and treatment methods according to gynecologic examination attendance in the last 5 years preceding the diagnosis of CC (attenders vs. nonattenders). RESULTS: In the period of observation, 242 women (53.8%) visited their gynecologists in the 5-year period prior to diagnosis of CC. Squamous cell carcinoma was present in 378 women (84.0%), adenocarcinoma in 45 (10.0%), adenosquamous carcinoma in 24 (5.3%) and other types in 3 women (0.7%). Attenders were significantly more frequently diagnosed with squamous cell carcinoma than nonattenders (chi-square=5.13; P<0.05). Attenders were significantly more frequently diagnosed in stage IA than in stage IB (chi-square=22.35; P<0.01). Similarly, in attenders stage I was significantly more frequent than stage II (chi-square=18.81; P<0.01). Pathologic smears of women with CC in the last 5-year period were most frequently evaluated as Pap II in the years 2003 and 2004 (in 39.1 and 26.4% of women, respectively) and as Pap III (in 27.9% of women) in 2005. Surgery was performed in 282 women (62.7%), radiotherapy in 158 (35.1%), symptomatic therapy in 9 (2.0%) and chemotherapy alone in 1 (0.2%) woman. In attenders, surgery alone was the most frequent treatment method (chi-square=91.18; P<0.01). CONCLUSION: Only in attenders a significant redistribution of CC stages in favor of early stages is observed, and in these women more conservative and less extensive treatment methods could be applied.

Osteopenia and osteoporosis can be predicted from Pap test.

OBJECTIVE: To determine sensitivity and specificity of Pap tests for osteopenia and osteoporosis using bone mineral density (BMD) with dual x-ray absorptiometry (DXA) as the reference standard. STUDY DESIGN: DXA measurement was performed on 136 routine Pap smears. Results of DXA measurement were expressed in T-scores, indicating degree of deviation compared to a young adult population of same age and gender. Smears were grouped into atrophic and mature cell patterns. Using a stereologic analysis, mean areas of squamous cells, their nuclei and their cytoplasm were estimated. RESULTS: There was significant positive correlation between cell area and T-score (p < 0.001), as well as between cytoplasm area and T-score (p < 0.001). There was no significant relationship between nucleus area and T-score (p > 0.05). Mean T-scores of patients with atrophic cells were significantly lower than mean T-scores of patients with mature cell patterns (p < 0.001). The group including patients with atrophic or mature cells had a sensitivity of 61.4% and specificity of 86.4%, with positive predictive value of 95.9% in detecting patients with osteopenia or osteoporosis. CONCLUSION: Women with atrophic smear pattern are susceptible to osteopenia or osteoporosis; many cases could be detected with routine Pap test without additional costs.

Cervical cancer screening: A Slovenian experience.

In Slovenia, opportunistic screening was introduced in regular gynaecological practice in 1960. The proportion of population screened was unknown, as well as there were no standards for quality assurance and control. Despite great number of smears read, there were no major changes in invasive cervical cancer incidence in the period 1979 till 1993, but in 1994 the incidence rate started to increase again to reach its peak in 1997 (23,1/100.000, 241 new cases). Based on the experiences from the countries with effectively organised screening programmes, a decision was made in 1996 by the Minister of Health to nominate a group of experts to prepare a proposal for organised cervical cancer screening programme after testing the methodology in pilot study. In the pilot the central computerised information system (Screening Registry) was gradually established to register all smears from the whole country, to identify women who do not attend for screening to send them invitation for screening and to monitor screening activity and its quality. The aim of pilot was also to develop guidelines for quality assurance and control of all procedures involved in cervical cancer screening and treatment of intraepithelial lesions. In three years since the beginning of the national programme, nearly 70% of women in the target age group were registered with at least one smear. All other results are presented in regular programme reports. There is still place for further development of the programme, but the incidence of cervical cancer already started to decline especially among younger women, who attend for screening more often than those aged over 50.

Effect of etidronate in preventing periprosthetic bone loss following cemented hip arthroplasty: a randomized, double blind, controlled trial.

INTRODUCTION: Periprosthetic bone loss occurs after insertion of a total hip prosthesis and is often a result of stress shielding or altered loading of the proximal femur. Preventing the bone loss, which may threaten the prosthesis survival, with an antiresorptive drug would be highly advantageous. MATERIALS AND METHODS: Our study investigated the effect of cyclic etidronate therapy on periprosthetic, contralateral hip, and spine bone mineral density (BMD) in a one-year, prospective, randomized, double-blind study in 31 patients after cemented hip arthroplasty. Etidronate was taken orally in a regimen repeated every 14 weeks, and periprosthetic BMD was measured with dual energy X-ray absorptiometry (DXA) in the total periprosthetic area and in the seven Gruen zones at 1 week (baseline), 6 weeks, 3 months, 6 months, and 12 months postoperatively. RESULTS: In the etidronate group there were significant temporal BMD decreases measured in Gruen zones 2, 3, 6, and 7 and in the entire proximal femur; the greatest decrease was 12.9% and was measured in zone 7 at six months. Also in the etidronate group, there was a significant 2.8% temporal BMD increase in the spine at 12 months. In the placebo group there were significant temporal BMD decreases measured in Gruen zones 1, 2, 3, 5, 6, and 7 and in the entire proximal femur; the greatest decrease was 25.5% and was measured in zone 7 at 12 months. There were no significant differences between the mean BMD measurements of the etidronate and placebo groups. CONCLUSION: These findings suggest that cyclic etidronate therapy has no significant effect in suppressing periprosthetic bone loss after cemented hip arthroplasty.

Morphological analysis of squamous cells in routine Pap smears as a predictor of bone mineral density in asymptomatic women.

OBJECTIVE: The present study attempted to find out possible coherence between morphologic characteristics in Pap smears and bone mineral density (BMD) as measured by DEXA. STUDY DESIGN: DEXA measurement (with the result expressed as T-score) was performed in 79 women in whom Pap smears for routine cervical cancer screening were obtained. The smears were grouped into atrophic and mature cell patterns. Using astereological analysis, the mean areas of squamous cells, their nuclei and cytoplasm were estimated. RESULTS: The mean areas of cells and cytoplasm were significantly lower at lower T-scores (P < 0.01), while the mean areas of nuclei were not (P > 0.5). T-scores were significantly lower in the atrophic cell pattern group (P < 0.001). The study group indicated concurrently high sensitivity of 80.9% and specificity of 78.1%, with positive predictive value of 84.4%. CONCLUSION: These results suggest that a significant number of women with low BMD could be identified parallel with the routine Pap test for cervical cancer screening without additional costs.

Cervical cancer screening practices in central and eastern Europe in 2012.

The burden of cervical cancer in central and eastern Europe is generally higher compared to western or northern Europe due to a history of mostly opportunistic cervical cancer screening practices and due to the strong influence of political and economic changes in post-communist transition. This article describes the current cervical cancer screening practices, organizational plans for the future, and main obstacles that need to be overcome in 16 countries in central and eastern Europe: Albania, Bosnia and Herzegovina, Bulgaria, Croatia, the Czech Republic, Estonia, Hungary, Latvia, Lithuania, Montenegro, Poland, Romania, Serbia, Slovakia, Slovenia and The former Yugoslav Republic of Macedonia. Unfortunately, only a few countries have managed to establish an organized and well-functioning cervical cancer screening program in recent years, whereas most countries in the region are still struggling with implementation-related issues of organized cervical cancer screening. Encouragingly, even in the countries where only opportunistic screening is performed, well-prepared plans and strategies have been established for switching to organized screening in the near future.

Pilomatrix carcinoma of the clitoris.

Pilomatrix carcinoma, a malignant counterpart of pilomatrixoma, is a rare skin neoplasm composed of basaloid and shadow cells that characterize differentiation toward the hair matrix. The authors present a case of pilomatrix carcinoma of the clitoris, a very unusual location not previously reported. Diagnostic criteria and differential diagnoses are discussed. Pilomatrix carcinoma should be differentiated from benign pilomatrixoma and other carcinomas with shadow cells, including basal cell carcinoma with matrical differentiation and metastases of visceral carcinomas with shadow cells.