Gyn Atlas Section 3
Negative for Intraepithelial Lesion or Malignancy
Jacalyn L. Papillo, BS, CT(ASCP) & Pamela Smith Piraino, CT(ASCP)
Whether utilizing the ThinPrep Pap Test or the conventional Pap smear, it is essential to have a firm understanding of what constitutes "normal". The ThinPrep process does not significantly alter the cytologic presentation of normal squamous and endocervical cells, as seen from the photomicrographs in the previous chapter on specimen adequacy. However, further discussion is warranted in the following areas: atrophy, pregnancy, post-partum changes, cytolysis, tubal metaplasia, lower uterine segment and benign appearing endometrial cells.
With the progression of atrophy during perimenopause and menopause there is a thinning of the epithelium, which often leads to a corresponding decrease in the number of cells shed during sample collection. Cell maturation is retarded, so that the prominent cell type becomes parabasal. With this shift in hormonal status the cell sample may contain increased glycogen, cellular degeneration and stripped nuclei. A granular background of degenerative cellular debris is generally seen. An inflammatory response is often present and evidence of bleeding is sometimes appreciated. Slight changes in the nuclear contours of the parabasal cells due to degeneration could lead to an over-diagnosis of atypia. It is important to evaluate nuclear size, the nuclear to cytoplasmic ratio and chromatin pattern carefully before rendering a cytologic diagnosis of ASC-US or more severe lesion in atrophic smears. In the case of atrophy with co-existing SIL, a sub-population of atypical cells should exhibit increased N/C ratios, hyperchromasia and irregular nuclear membranes.
Reminder: You may click on any slide image for an enlarged view.
Inflammatory background, decreased maturation and slight degenerative change are present. 40X
Parabasal cells occur in sheets and singly. 40X
Normal endocervical cells are often readily identified in well preserved atrophic smears. 40X
Note low N/C ratio and degenerative vacuoles (not to be mistaken for HPV related changes). 60X
Note increased N/C ratio, hyperchromasia and irregular nuclear membranes in the HSIL cells. 60X
PregnancyThe cytologic patterns associated with excessive hormonal influences, such as pregnancy, are similar with conventional Pap smears and ThinPrep slides. During pregnancy, caution should be used to properly differentiate glycogenated cells from true koilocytes present with HPV-associated changes. This can be done by noting a lack of nuclear irregularity, along with a smooth round vacuolization from glycogen, compared to abnormal nuclei surrounded by a sharp irregular cavitation caused by HPV.
Glycogenated intermediate cells are seen during pregnancy. 60X
Postpartum ChangesDuring the postpartum period, or if breast feeding, a hormonal pattern of low squamous maturation predominates. Parabasal cells are abundant, usually presenting singly. Glycogen may also be present. A background of inflammatory cells and reactive changes is often present and may require additional scrutiny. Adherence to strict cytologic criteria (the lack of an increased N/C ratio, abnormal chromatin and irregular nuclear membranes) should preclude an over-diagnosis of cytologic atypia.
Postpartum changes including single parabasal cells and a background of inflammatory cells. 20X
High power view of parabasal cells during postpartum. Note presence of glycogen and low N/C ratio. 60X
HSIL in a postpartum sample. Note the increased N/C ratio and abnormal chromatin in the HSIL cells as compared to the parabasal cells, apparent even at low magnification. 20X
CytolysisThe familiar pattern of bare nuclei, superficial and intermediate cells and Döderlein bacilli associated with lactobacilli-induced cytolysis can be readily identified at low magnification on ThinPrep slides. The bacilli may be trapped in mucus or seen overlying squamous cells.
Bare nuclei present with Doederlein bacilli overlying intermediate squamous cells. 60X
Tubal MetaplasiaTubal metaplasia is characterized by columnar cells with pleomorphic, hyperchromatic nuclei with increased N/C ratios. As with conventional Pap smears, cells from tubal metaplasia may occasionally be mistaken for atypical endocervical cells and even adenocarcinoma in situ. In order to avoid an over-diagnosis, look for the presence of discrete cytoplasmic vacuoles, terminal bars and cilia.
A strip of columnar cells with hyperchromatic, crowded nuclei. Close observation reveals distinct cytoplasmic vacuoles and cilia possibly representing tubal metaplasia. 60X
Lower Uterine SegmentWith increased vigilance by providers to collect adequate specimens, aggressive sampling may result in the presence of cells from the lower uterine segment in cervicovaginal samples. Cells from the lower uterine segment generally present as a tissue fragment rather than individual cells or small groups of cells. The fragment has well ordered nuclei and nucleoli may be present. A folding over of the fragment on itself may cause a three dimensional appearance.
A fragment of cells from the lower uterine segment, which has folded over itself causing a three dimensional appearance. 20X
Lower uterine segment may have prominent nucleoli. 40X
Benign Appearing Endometrial CellsEndometrial cells may display a wide range of nuclear and cytoplasmic size, as well as architectural configuration. Endometrial cell groups tend to round up in ThinPrep solution. The nuclei are well preserved and appear crisp with distinct, active chromatin; chromocenters are frequently noted.
Bethesda 2001 recommends the reporting of benign appearing exfoliated endometrial cells in women age 40 and above, regardless of clinical history. The general categorization "Other" was included in the Bethesda 2001 lexicon for this purpose. Histiocytes, stromal cells and abraded endometrial cells do not have the same significance as exfoliated endometrial cells and should not be considered in the same context. While most often benign, identification of endometrial cells, particularly if not associated with menses or after menopause, may indicate risk for an endometrial abnormality.
Glandular clusters of endometrial cells with 3-dimensional architecture and distinct chromocenters may be noted. Occasional bean-shaped nuclei aid in identification, as does the comparison of nuclear size with reference intermediate cell nuclei. 60X
3-dimensional group of endometrial cells with round and elongated nuclei; pale, fine and evenly dispersed chromatinand occasional chromocenters are appreciated. Nuclear size equal to reference intermediate cell nuclei. 60X
Pictured is a rounded cluster of endometrial cells with ghost (lysed) red blood cells in the background. Nuclear chromatin is evenly distributed, with occasional chromocenters seen. 60X
Endometrial cells may present as loose clusters of cells with abundant cytoplasm, some vacuolated. Round to oval nuclei with several bean-shaped nuclei are readily identifiable. 60X
A loose cluster of cells with bean-shaped, elongated or spindled nuclei and longitudinal grooves in the nuclear membrane may be appreciated. Nuclear size is similar or smaller than the reference intermediate cell nuclei. 40X
A small cluster of superficial stromal cells with foamy, vacuolated cytoplasm is indistinguishable from histiocytes. 40X
Endometrial cells may coexist with HSIL (low power). Note the squamoid cytoplasm and increased N/C ratios in HSIL cells vs. glandular cluster of endometrial cells with 3-dimensional architecture. 20X
A syncytium of HSIL cells at high power is pictured. Note the abnormal chromatin, enlarged nuclear size and loss of polarity to differentiate from endometrial cells. 60X
Same Patient, Split Sample
Trichomonas VaginalisOn ThinPrep slides, trichomonads may appear slightly smaller than the air dried organisms found on the conventional Pap smear. Because of the rapid fixation, the ThinPrep Pap test slides bear a similarity to a "wet prep" used in the physician's office.
Although proportionately smaller, the organisms maintain their characteristic internal structure and often retain their flagellae lending confidence to the diagnosis. Two findings characteristically found on ThinPrep slides are useful in the detection of T. Vaginalis: trichomonads tend to present either as isolated organisms or in prominent groupings, or microcolonies, often overlying epithelial cells or clustering together.
Candida speciesThe spearing of the epithelial cells with the cells folding along the long axis of the organism is the first clue that Candida spp. is present on the ThinPrep slide. Pseudo-septate hyphae and budding yeasts are preserved and easily detected. Staining may vary from colorless to eosinophilic to gray-brown depending upon patient biology. Laboratories often observe an increased detection of Candida spp. when implementing the ThinPrep Pap Test.
Spearing of squamous cells occurs along the long axis of Candida spp. 40x
Pseudo-hyphae, hyphae and budding identifies the organism. 60x
Herpes Simplex VirusThe classic characteristics of herpes virus infection are retained and readily identified on ThinPrep slides. Cellular changes such as multi-nucleation, nuclear molding, ground-glass chromatin, chromatin margination, and intranuclear viral inclusions aid in the diagnosis.
Intranuclear inclusions, multi-nucleation, and ground-glass chromatin characterize herpes simplex virus. 60x
ActinomycesDelicate and dense branches of filamentous bacteria with accompanying sulphur granules allow for easy identification of the organism. Infection by Actinomyces often accompanies the use of an intrauterine device (IUD) or a pessary.
IUD ChangesPatients with an IUD may occasionally show single or clustered glandular cells with an increase in nuclear to cytoplasmic ratio. Nucleoli may be present and prominent vacuoles may cause a signet ring appearance to the cells. To avoid a diagnosis of AGUS, cellular changes must be correlated with patient history. These changes often accompany an infection with Actinomyces.
Single cells with increased N/C ratio, slight hyperchromasia and some irregularity to the nuclear membrane represent cellular changes caused by an IUD. 40x
Shift in the Flora Suggestive of Bacterial VaginosisSquamous cells may be covered with coccobacilli when the normal lactobacilli are absent. A diagnosis of bacterial vaginosis may be confirmed by bacterial culture.
Chronic Follicular CervicitisThe presence of lymphocytes accompanied by tingible body macrophages may indicate a chronic inflammatory condition or the rupturing of a lymph follicle during sample collection. On the ThinPrep slide the lymphocytes from chronic follicular cervicitis will generally appear in small clusters grouped together. The presence of tingible body macrophages aid in the diagnosis.
Lymphocytes and tingible body macrophages characterize a chronic cervicitis. 60x
RadiationThe same changes found on conventional Pap smears following radiation therapy are seen on the ThinPrep Pap Test slides. Cells may be enlarged, show multinucleation, vacuolization, bi-chromasia, and altered chromatin. Ingested inflammatory cells may also be present.
Multinucleation, vacuolization, altered chromatin and inflammatory cell engulfment are often present post radiation therapy. 40x
Atrophy with inflammationConventional Pap smears from atrophic patients generally contain a uniform background of granular debris, and degenerated inflammatory cells. With ThinPrep slides this background debris is maintained, but is often pooled into small clumps alongside atrophic epithelium.
An atrophic patient sample contains inflammatory cells and debris on ThinPrep Pap Test. 40x
Decidual changesThe cells of patients who are late in their pregnancy (or postpartum) may show changes which can mimic high grade dysplasia. The morphologic characteristics of decidual change on ThinPrep Pap test slides are similar to those found on the conventional Pap smear.
RepairCriteria to classify a reparative process on Thin Prep slides are similar to the criteria as enumerated in The Bethesda System. (1994)
- Minimal nuclear enlargement (11/2 to 2 times normal intermediate cell nucleus)
- Bi and multi-nucleation may occur
- Mild hyperchromasia may be present
- Chromatin finely granular, evenly distributed
- Karyopyknosis and karyorrhexis may occur
- Nuclear membranes smooth
- Prominent single or multiple nucleoli
- Flat sheets with polarity maintained; groups may round up in fluid
- Mitosis and single cells may be noted
- Often accompanied by neutrophils, inflammatory changes and/or organisms
Repair can present with prominent nucleoli, some multiple. 40x
Repair shows smooth nuclear contours and polarity is maintained within group. 60x
Repair may contain mitosis and macronucleoli. 60x
Repair often shows "school of fish" architecture. Polarity is maintained and the N/C ratio of the cells is in the normal range. 40X
- Biscotti CV, et al: Thin-layer Pap test vs conventional Pap smear: Analysis of 400 split samples. J Reprod Med 2002;47(1):9-13.
- Lee KR, et al: Comparison of conventional Papanicolaou smears and a fluid-based, thin-layer system for cervical cancer screening. Obstet Gynecol 1997;90:278-284.
- Solomon, Diane, et al: The 2001 Bethesda System, terminology for reporting results of cervical cytology, JAMA 2002;287(16): 2114-2119.