Significance of a first-time atypical Papanicolaou smear in a young, high-risk African-American and Latino-American population

The first atypical Papanicolaou smear in young, sexually active Latino and African-American women of low socioeconomic status may be predictive of underlying cervical neoplasia and human papillomavirus infection of significant quantity. The optimal management of first-time atypia on routine Pap smear has not been established. In many clinics, colposcopically directed sampling of the cervix is recommended only if atypia persists following specific or nonspecific treatment of cervicitis or after an arbitrarily determined time interval. Others recommend immediate colposcopic evaluation. To determine the best approach to the first-time atypical Pap smear in young minority women at high risk for the development of cervical cancer, 250 such patients were evaluated with colposcopically directed biopsy of the cervix prior to any form of therapy. Pap smears were repeated at the time of colposcopy. Histologically, there was evidence of cervical intraepithelial neoplasia in 41% of patients and human papillomavirus infection in 86%. Repeat Pap smears predicted the presence of cervical intraepithelial neoplasia in only 24% of patients. Immediate colposcopic evaluation represents the most prudent approach to the first-time atypical Pap smear in young, high-risk minority women.     

Natural history of false-negative papanicolaou smears: a prospective study using screening colposcopy in addition to cytology

The regular Papanicolaou (Pap) smear is cornerstone of women's preventive healthcare. The introduction of the regular Pap smear as a screening tool for cervical cancer has markedly decreased the number of deaths from cervical cancer. During the past decade, however, the rate of death from cervical cancer has remained relatively static. This screening method is known to have a high rate of false-negative results; therefore, serial examinations are necessary for optimal sensitivity. The sensitivity of the routine pelvic examination is further increased with the addition of colposcopy to cytologic screening. Patients identified as having disease by colposcopy and biopsy but not by Pap smear (false-negative), are followed up, and the progression of their disease is documented until the Pap smear reflects the known change. A total of 276 patients whose Pap tests showed "no abnormal cells" and mild dysplasia (CIN-1 [cervical intraepithelial neoplasia]) by colposcopy and biopsy, who opted not to be treated, were followed up as a function of time. The results of the Pap test on the 276 patients progressed to a low-grade squamous intraepithelial lesion (LGSIL) or higher within 42 months. In this study, false-negative Pap tests that were identified by simultaneously performed colposcopy became positive within 42 months; therefore, careful repetitive screening is necessary. The author argues that a definitive, prospective study on the addition of colposcopy or similar adjunct procedure to the routine pelvic examination, in selected cases, is needed.     

Significant reduction in the rate of false-negative cervical smears with neural network-based technology (PAPNET Testing System)

False-negative cervical Pap smears may lead to disability or death from carcinoma of the uterine cervix. New computer technology has led to the development of an interactive, neural network-based vision instrument to increase the accuracy of cervical smear screening. The instrument belongs to a new class of medical devices designed to provide computer-aided diagnosis (CADx). To test the instrument's performance, 487 archival negative smears (index smears) from 228 women with biopsy-documented high-grade precancerous lesions or invasive cervical carcinoma (index women) were retrieved from the files of 10 participating laboratories that were using federally mandated quality assurance procedures. Samples of sequential negative smears (total 9,666) were retrieved as controls. The instrument was used to identify evidence of missed cytological abnormalities, including atypical squamous or glandular cells of undetermined significance (ASCUS, AGUS), low-grade or high-grade squamous intraepithelial lesions (LSIL, HSIL) and carcinoma. Using the instrument, 98 false-negative index smears were identified in 72 of the 228 index women (31.6%, 95% confidence interval [CI]: 25% to 38%). Disregarding the debatable categories of ASCUS or AGUS, there were 44 women whose false-negative smears disclosed squamous intraepithelial lesions (SIL) or carcinoma (19.3%; 95% CI: 14.2% to 24.4%). Unexpectedly, SILs were also identified in 127 of 9,666 control negative smears (1.3%; 95% CI: 1.1% to 1.5%). Compared with historical performance data from several participating laboratories, the instrument increased the detection rate of SILs in control smears by 25% and increased the yield of quality control rescreening 5.1 times (P < 0.0001). These data provide evidence that conventional screening and quality control rescreening of cervical smears fail to identify a substantial number of abnormalities. A significant improvement in performance of screening of cervical smears could be achieved with the use of the instrument described in this report.     

Hantavirus pulmonary syndrome treated with inhaled nitric oxide

BACKGROUND: A retrospective study was done to assess the correlation between endometrial cells on routine cervical cytology and carcinoma of the endometrium. METHODS: In a 4-year period, endometrial cells of some type were identified on the Papanicolaou (Pap) smears of 61 women, of whom 52 had further diagnostic evaluation of the endometrium. Data were analyzed with a multivariate stepwise logistic regression. RESULTS: The results indicated an association of endometrial cells in Pap smears with carcinoma of the endometrium in seven patients (13.5%). In 45 patients (86.5%), the final diagnosis was benign. Factors that impacted the diagnosis of carcinoma were the findings of atypical or cancerous endometrial cells on Pap smear and abnormal vaginal bleeding. CONCLUSIONS: These data indicate the importance of further diagnostic evaluation with endometrial sampling in postmenopausal patients with endometrial cells seen in Pap smears, especially those with abnormal bleeding.     

Evaluation of the ThinPrep Pap test as an adjunct to the conventional Pap smear

OBJECTIVE: To evaluate the ThinPrep Pap test as an adjunct to the conventional Pap smear. DESIGN AND SETTING: Prospectively collected cervical samples were split for independent screening at a large specialised private gynaecological pathology practice in Sydney. MAIN OUTCOME MEASURES: Detection of additional significant abnormalities (cervical intraepithelial neoplasia 1, or more severe); changed management recommendations from "repeat smear in 12 months" or "...six months" to "colposcopy", a reduction in unsatisfactory reports. RESULTS: 35,560 paired (split-sample) conventional and ThinPrep slides were prepared. Significant abnormalities were detected in 724 conventional smears (2%). Additional significant abnormalities were found in 85 ThinPrep slides whose corresponding conventional smear was negative or unsatisfactory even after review, representing a 12% increase in the detection of significant abnormalities. As a result of the addition of ThinPrep, management recommendations were changed from "repeat smear in 12 months" or "...six months" to "colposcopy" for 89 of 1669 women whose conventional Pap smears showed minor non-specific changes or papillomavirus. There were 1258 conventional smears (3.5%) that were unsatisfactory compared with 235 ThinPrep slides (0.7%); for only 74 samples (0.2%) were both slides unsatisfactory. CONCLUSIONS: The addition of the ThinPrep Pap test improves detection and clinical management of cervical abnormalities, and reduces the number of unsatisfactory samples which would otherwise require repeat tests.     

Mcm2 is a target of regulation by Cdc7-Dbf4 during the initiation of DNA synthesis

The initiation of DNA synthesis is an important cell cycle event that defines the beginning of S phase. This critical event involves the participation of proteins whose functions are regulated by cyclin dependent protein kinases (Cdks). The Mcm2-7 proteins are a family of six conserved proteins that are essential for the initiation of DNA synthesis in all eukaryotes. In Saccharomyces cerevisiae, members of the Mcm2-7 family undergo cell cycle-specific phosphorylation. Phosphorylation of Mcm proteins at the beginning of S phase coincides with the removal of these proteins from chromatin and the onset of DNA synthesis. In this study, we identified DBF4, which encodes the regulatory subunit of a Cdk-like protein kinase Cdc7-Dbf4, in a screen for second site suppressors of mcm2-1. The dbf4 suppressor mutation restores competence to initiate DNA synthesis to the mcm2-1 mutant. Cdc7-Dbf4 interacts physically with Mcm2 and phosphorylates Mcm2 and three other members of the Mcm2-7 family in vitro. Blocking the kinase activity of Cdc7-Dbf4 at the G1-to-S phase transition also blocks the phosphorylation of Mcm2 at this defined point of the cell cycle. Taken together, our data suggest that phosphorylation of Mcm2 and probably other members of the Mcm2-7 proteins by Cdc7-Dbf4 at the G1-to-S phase transition is a critical step in the initiation of DNA synthesis at replication origins.     

Pap smear screening in an urban STD clinic. Yield of screening and predictors of abnormalities

BACKGROUND: Pap smear screening studies in STD clinics have reported high rates of squamous intraepithelial lesions (SIL); however, there are limited data on levels of unsatisfactory smears of characteristics associated with cytologic abnormalities. GOAL: To assess the yield to Pap smear screening in an STD clinic and to evaluate the rates of and risk factors for atypia, low-grade SIL (LSIL), and high-grade SIL (HSIL). STUDY DESIGN: A chart review of the clinic records of women undergoing Pap smear screening between 1991 and 1994 was conducted. Results were assessed from two different screening protocols, the first using a Dacron swab to obtain the endocervical sample and the second using a cytobrush. RESULTS: Of 2034 Pap smears, 1313 (64.6%) were negative, 202 (9.9%) were unsatisfactory, 257 (12.6%) were atypical, 211 (10.4%) had LSIL, and 51 (2.5%) had HSIL. With the change to the cytobrush protocol, the rate of unsatisfactory smears decreased from 14.4% to 3% (p < 0.001), atypia increased from 10% to 16.7% (p < 0.001), and HSIL rose from 1.7% to 3.7% (p < 0.001). By multivariate analysis, atypia was associated with genital warts (odds ratio (OR) 1.53, 95% confidence interval (CI): 1.16-2.02); LSIL with younger age (p < 0.001, for trend), black race (OR 1.51, 95% CI: 1.08-2.10), genital warts (OR 1.81, 95% CI: 1.33-2.47), and an abnormal appearance of the cervix on examination (OR 2.49, 95% CI: 1.85-3.35); and HSIL with a previous abnormal Pap smear (OR 2.48, 95% CI: 1.08-2.10). Overall, abnormality rates were significantly higher in adolescents (35.5%) than older women (21.7%) (p < 0.01). CONCLUSIONS: Obtaining satisfactory Pap smears among women undergoing screening in an STD clinic is feasible and cytologic abnormalities are common. These results continue to support the need for Pap smear screening in STD clinics, but the high rates in adolescents, a group in whom the natural history of cytologic abnormalities has not been well-defined, raise questions about the need to develop age-appropriate screening and management strategies.     

Quality of life of women with stress urinary incontinence with or without pollakiuria]

BACKGROUND: The optimal management of low grade Papanicolaou (Pap) smear abnormalities remains controversial. This center's experience with recommending cytologic follow-up for women with atypical cells of undetermined significance (ASCUS) or low grade squamous intraepithelial lesions (LSIL) was reviewed to determine outcome and patient/physician compliance. METHODS: The records were reviewed on women with Pap smears reported as either ASCUS (320) or LSIL (112) who did not have a history of dysplasia. The cytologic and colposcopic follow-up for a 2-year period was obtained from the laboratory data base that includes the colposcopy and cancer referrals for this region. Repeat Pap smear in 6 months was recommended. If patients subsequently demonstrated high grade SIL (HSIL) or persistent ASCUS or LSIL over three time intervals, colposcopic evaluation was recommended. RESULTS: The outcome was determined by the most significant diagnosis among the follow-up Pap smears or colposcopic biopsies. 29% of patients were lost to follow-up. Of the remaining patients, 70.5% reverted to normal or benign cellular changes, 25.3% persisted as ASCUS or LSIL, and 5.2% progressed to HSIL. The majority of patients (68%) were referred for colposcopy for persistent mildly abnormal Pap smears. The timing of referral ranged from 3-30 months. CONCLUSIONS: These results suggest that cytologic follow-up of women with low grade Pap smear abnormalities will identify a large number whose smears will regress to normal. A small but significant proportion of women showed subsequent HSIL. Most HSIL was detected within 1 year of the initial abnormal Pap smear and the majority of intervening Pap smears also were abnormal. Approximately one third of patients did not have follow-up within the study system and their outcome was uncertain. Although the recommendations are standard, patterns of follow-up and referral to colposcopy varied widely, suggesting that the guidelines need to be reinforced to both patients and physicians. [See editorial on pages 1-4, this issue.]     

Update on the Papanicolaou smear: new issues for the 1990s

The Papanicolaou (Pap) smear is a complex examination that has undergone refinement in recent years. It is now widely accepted that a false-negative rate of perhaps 10 to 20% is inevitable for a variety of reasons. The diagnosis of "atypical cells of undetermined significance" is not a benign diagnosis and is often followed by a more severe lesion on follow-up. The Pap smear appears to be an inefficient method for detecting glandular neoplasms, and clinical features remain the most important diagnostic clue. A Pap smear and cervical biopsy should correlate in 75% or more of cases. In cases of noncorrelation, a positive Pap smear diagnosis is supported in the vast majority of cases.     

The significance of endometrial cells in the cervical smear

Presence of atypical endometrial cells in a cervical smear is quite unusual; but if they are found the woman needs extra attention because of the risk of adenocarcinoma of the uterus. Postmenopausal women sometimes also have endometrial cells without atypia in the cervical smear, as a sign of endometrial pathology. We conducted a retrospective follow-up study of these women. From 1978 to 1989, 52 smears from a total of about 50,000 postmenopausal women were classified as Pap IIIA, due to the presence of normal endometrial cells. Histological follow-up was performed in 25 cases and 3 times an adenocarcinoma of the endometrium was diagnosed (6%). The smears from 142 women (pre- and postmenopausal) contained atypical endometrial cells and therefore were classified as Pap IIIA or higher. Histological follow-up was performed in 104 cases. In 48% (n = 68) an adenocarcinoma was diagnosed. In 75% of these cases (n = 51) the atypical cells were graded as severe or worse (> or = Pap IIIB). Cervical smears are not taken to detect pathology of the endometrium, but whenever atypical cells are found in the smear there is a strong indication for further (gynaecological) examination.     

Mcm2 and Mcm3 are constitutive nuclear proteins that exhibit distinct isoforms and bind chromatin during specific cell cycle stages of Saccharomyces cerevisiae

The Mcm2-7 proteins are a family of conserved proteins whose functions are essential for the initiation of DNA synthesis in all eukaryotes. These patients are constitutively present in high abundance in actively proliferating cells. In Saccharomyces cerevisiae, the intracellular concentrations of Mcms are between 100 and 500 times the number of replication origins. However, these proteins are limiting for the initiation of DNA synthesis at replication origins. Our studies indicate that only a small fraction of Mcm2 and Mcm3 tightly associates with chromatin, from late M phase to the beginning of the S phase. The rest of the Mcm2 and Mcm3 proteins are disturbed to both the cytoplasm and nucleoplasm in relatively constant levels throughout the cell cycle. We also show that S. cerevisiae Mcm3 is a phosphoprotein that exists in multiple isoforms and that distinct isoforms of Mcm2 and Mcm3 can be detected at specific stages of the cell cycle. These results suggest that the localization and function of the Mcm proteins are regulated by posttranslational phosphorylation in a manner that is consistent with a role for the Mcm proteins in restricting DNA replication to once per cell cycle.     

Association of RPA with chromosomal replication origins requires an Mcm protein, and is regulated by Rad53, and cyclin- and Dbf4-dependent kinases

Eukaryotic cells use multiple replication origins to replicate their large genomes. Some origins fire early during S phase whereas others fire late. In Saccharomyces cerevisiae, initiator sequences (ARSs) are bound by the origin recognition complex (ORC). Cdc6p synthesized at the end of mitosis joins ORC and facilitates recruitment of Mcm proteins, which renders origins competent to fire. However, origins fire only upon the subsequent activation of S phase cyclin-dependent kinases (S-CDKs) and Dbf4/Cdc7 at the G1/S boundary. We have used a chromatin immunoprecipitation assay to measure the association with ARS sequences of DNA primase and the single-stranded DNA binding replication protein A (RPA) when fork movement is inhibited by hydroxyurea (HU). RPA's association with origins requires S-CDKs, Dbf4/Cdc7 kinase and an Mcm protein. The recruitment of DNA primase depends on RPA. Furthermore, early- and late-firing origins differ not in the timing of their recruitment of an Mcm protein, but in the timing of RPA's recruitment. RPA is recruited to early but not to late origins in HU. We also show that Rad53 kinase is required to prevent RPA association with a late origin in HU. Our data suggest that the origin unwinding accompanied by RPA association is a key step, regulated by S-CDKs, Dbf4/Cdc7 and Rad53p. Thus, in the presence of active S-CDKs and Dbf4/Cdc7, Mcms may open origins and thereby facilitate the loading of RPA.     

Formation of a preinitiation complex by S-phase cyclin CDK-dependent loading of Cdc45p onto chromatin

Cdc45p, a protein essential for initiation of DNA replication, associates with chromatin after "start" in late G1 and during the S phase of the cell cycle. Binding of Cdc45p to chromatin depends on Clb-Cdc28 kinase activity as well as functional Cdc6p and Mcm2p, which suggests that Cdc45p associates with the prereplication complex after activation of S-phase cyclin-dependent kinases (CDKs). As indicated by the timing and the CDK dependence, binding of Cdc45p to chromatin is crucial for commitment to initiation of DNA replication. During S phase, Cdc45p physically interacts with minichromosome maintenance (MCM) proteins on chromatin; however, dissociation of Cdc45p from chromatin is slower than that of MCMs, which indicates that the proteins are released by different mechanisms.     

Screening for cervical cancer

As part of a continuing series on how the work of pathology laboratories contributes to patient care, this article looks at the Papanicolaou (Pap) or cervical smear test, which involves the microscopical examination of cells recovered by scraping the surface of the cervix. The incidence, causes and aetiology are described and the organisation of cervical screening is shown. Finally, the range of findings from a positive smear test is explained.     

Efficacy of ThinPrep preparation of cervical smears: a 1,000-case, investigator-sponsored study

The diagnoses of 1,000 pairs of conventional Papanicolaou (Pap) smears and ThinPrep preparations were compared. Cervical cells were collected using an Ayre spatula and endocervical brush. The conventional smear was made first, the collection devices were rinsed into PreservCyt solution, and the slides were prepared using the ThinPrep Processor. The diagnoses of the paired smears agreed in 988 of the 1,000 cases (98.8%), including 949 negatives, 28 atypicals, 9 low grade squamous intraepithelial lesions (LGSIL), and 2 high grade squamous intraepithelial lesions (HGSIL). Five cases where LGSIL or HGSIL was found on the ThinPrep slide were negative or atypical on the conventional smear. No conventional smear abnormalities were missed on the ThinPrep slide. Although not statistically significant, this difference indicates that the ThinPrep method gives a better diagnosis of abnormalities than the conventional method. The ThinPrep method was acceptable to participating physicians and ThinPreps were easier and faster to screen than conventional smears.     

Screening histories of incidence cases of cervical cancer and high grade SIL. A comparison

OBJECTIVE: To compare the cytologic history of patients with cervical cancer with that of patients with high grade cervical intraepithelial neoplasia (CIN) in order to analyze the causes of screening failure. STUDY DESIGN: In 337 patients treated for high grade CIN and 86 women treated for cancer of the cervix, all cytologic reports from the last five years before diagnosis were reviewed, and slides of normal smears taken within three years of diagnosis were reexamined. RESULTS: Among patients with cancer, 32/86 (37.2%) never had a smear, for 28/86 (32.6%) the time interval between screenings was greater than three years, 12/66 (18.2%) were incorrectly managed after the first abnormal smear, and 7/69 (10.1%) had a normal smear within three years of diagnosis. Conversely, among patients with high grade CIN, 45/337 (13.4%) never had a smear, 60/337 (17.8%) had a time interval between screenings greater than three years, 120/337 (35.6%) were incorrectly managed after the first abnormal smear, and 100/333 (30.0%) had a normal smear within three years of diagnosis. When compared with high grade CIN, cancer of the cervix was associated with absent or insufficient screening (adjusted odds ratio [aOR] = 3.28, 95% confidence interval (95% CI) = 1.86-5.80) but neither with incorrect management of the first abnormal smear (aOR = 1.32, 95% CI = 0.46-3.75) nor with a normal smear within three years of diagnosis (aOR = 0.22, 95% CI = 0.10-0.50). CONCLUSION: Better participation of patients is necessary to improve the efficacy of screening. Conversely, reducing the interval between smears to less than three years would cause a major increase in cost and earlier diagnosis of high grade CIN, but not a significant decrease in the number of cancers.     

Down-regulation of cyclin F levels during nerve growth factor-induced differentiation of PC12 cells

Recent experiments in budding yeast and Xenopus have provided new insights into the regulation of eukaroytic DNA replication. The multi-subunit origin recognition complex plays a key role in initiation, remaining bound at origins of replication during most of the cell cycle. Early in the cell cycle, Cdc6 and the Mcm proteins 'reset' chromatin for another round of DNA replication. Cyclin-dependent kinases appear to play a dual role, both in activating replication origins and blocking the formation of new pre-replicative complexes; thus limiting replication to once per cell cycle.     

Usefulness of hybrid capture HPV DNA assay as a diagnostic tool for human papillomavirus infection

The purpose of this study was to determine the usefulness of the hybrid capture HPV DNA assay, a new nonradioactive solution hybridization assay, as a diagnostic tool for human papillomavirus infection. In a total of 234 women, samples for the hybrid capture assay and polymerase chain reaction (PCR) assay were obtained by wiping a swab across the cervix and external os (either a Dacron swab or a cotton swab was used). The papanicolaou smear test (Pap smear) was carried out on all 234 women. Tissue samples for biopsy were obtained by colposcopy from 118 of the women. Fisher exact test was used for statistical analyses. Using the hybrid capture assay, HPV DNA of high- and intermediate-oncogenic-risk type was detected in 23 (13.9%) of 166 samples from women with Pap smear Class I or II, and 48 (70.6%) of 68 with Pap smear Class III, IV or V (p < 0.0001). The HPV DNA type was detected in 18 (29.0%) of 62 samples from those with no evidence of cervical intraepithelial neoplasia and 44 (78.6%) of 56 with cervical intraepithelial neoplasia or squamous cell carcinoma (p < 0.0001). Correlation of the test results between the hybrid capture test and PCR was determined by using the 217 samples in which both test results were available (PCR test results were not obtainable in 17 samples. When PCR is set as a gold standard, the hybrid capture test has high sensitivity (74.6%) and specificity (92.7%). These findings suggest that the hybrid capture HPV DNA assay is a useful method for diagnosing HPV infection in the clinic.     

Nuclear accumulation of Saccharomyces cerevisiae Mcm3 is dependent on its nuclear localization sequence

BACKGROUND: The proteins of the Mcm2-7 family are required for the initiation of DNA replication. In Saccharomyces cerevisiae the nuclear envelope does not break down during the mitotic phase of the cell cycle. Large nuclear proteins, such as the Mcm proteins, which accumulate in the nucleus during specific portions of the cell cycle, must have regulated mechanisms to direct their entry into the nucleus. RESULTS: We have identified a nuclear localization sequence (NLS) in Mcm3, and demonstrated that it is necessary for the translocation of Mcm3 into the nucleus and sufficient for directing Escherichia coli beta-galactosidase to the nucleus. Immediately adjacent to the nuclear localization sequence are four potential sites for phosphorylation by Cdc28. Mutagenesis of all four sites has no immediate phenotypic effect on cell growth or viability, nor does it affect nuclear accumulation of Mcm3, although two-dimensional protein gel analysis has shown that at least some of these sites are normally phosphorylated in vivo. Substitution of the Mcm3 NLS by the SV40 large T-antigen NLS also directs the nuclear accumulation of the Mcm3-T-antigen protein, although cell growth is compromised. Replication activity in cells bearing either the Mcm3-Cdc28 phosphorylation site mutations or the Mcm3 T-antigen NLS substitution, as measured by plasmid stability assays, is comparable to activity in wild-type cells. CONCLUSIONS: The Mcm3 protein is imported into the nucleus by a specific NLS. The cell cycle specific nuclear accumulation of Mcm3 appears to be a result of nuclear retention or nuclear targeting, rather than nuclear import regulated through the NLS.