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The contribution of MIB 1 in the accurate grading of vulvar intraepithelial neoplasia

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To determine the interobserver variation in scoring presence and grade of vulvar intraepithelial neoplasia (VIN) in haematoxylin/eosin (H/E) slides, MIB 1 slides, and the combined use of H/E and MIB 1 slides. 10 slides were stained with H/E and MIB 1 with each of the following diagnoses: normal vulvar skin, VIN 1, VIN 2, and VIN 3. Six observers first scored the H/E slides separately from the MIB 1 slides and second the combined H/E and MIB 1 slides. Unweighted group kappa for MIB 1 was 0.62 and the weighted group kappa was 0.91. This was significantly better than the unweighted group kappa for H/E slides (0.47, p = 0.023) as well as the weighted group kappa for H/E slides (0.82, p = 0.014). There was no improvement by the combined use of H/E and MIB 1 slides. VIN 2 is far less confused with VIN 3 in the combined use of H/E and MIB 1 slides (9%) than in H/E slides (38%) (p = 0.007). There is a tendency to grade VIN in a two tailed grading system rather than a three tailed grading system, which became more apparent with the combined use of H/E and MIB 1 slides. The interobserver variation with sole use of MIB 1 is better than with the use of H/E stain in VIN. The use of MIB 1 in grading VIN diminishes confusion between VIN 2 and VIN 3 fourfold. A two tailed grading system for VIN seems already to work in daily practice.
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... It is widely accepted to be a subtle and difficult histopathological diagnosis as it can easily be mistaken for a benign dermatitis or epithelial hyperplasia 5 and may be difficult to distinguish from the often present background of lichen sclerosus. 17 To our knowledge, only three prior studies have addressed the interobserver variability of VIN [19][20][21] showed a good agreement of 40 specimens with normal skin and VIN 1-3, of which possibly some of the VIN cases may be of the non-HPV type although this is not further clarified. Apparently, there is no literature that focuses on the histopathological diagnosis of DVIN. ...
... Besides the use of these histopathological criteria in making the right diagnosis, there may also be a role for immunohistochemistry. [21][22][23] The use of MIB1 can be helpful to distinguish between normal vulvar epithelium and DVIN as the basal cell layer in DVIN shows a higher proliferation index (percentage of MIB1-positive cells), than normal vulvar epithelium, where the basal cell layer often is negative for MIB1. 23 Furthermore, in DVIN a strong positive staining of the (supra)basal cell layers with p53 can be seen. ...
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No published data concerning intraobserver and interobserver variability in the histopathological diagnosis of differentiated vulvar intraepithelial neoplasia (DVIN) are available, although it is widely accepted to be a subtle and difficult histopathological diagnosis. In this study, the reproducibility of the histopathological diagnosis of DVIN is evaluated. Furthermore, we investigated the possible improvement of the reproducibility after providing guidelines with histological characteristics and tried to identify histological characteristics that are most important in the recognition of DVIN. A total number of 34 hematoxylin and eosin-stained slides were included in this study and were analyzed by six pathologists each with a different level of education. Slides were reviewed before and after studying a guideline with histological characteristics of DVIN. Kappa statistics were used to compare the interobserver variability. Pathologists with a substantial agreement were asked to rank items by usefulness in the recognition of DVIN. The interobserver agreement during the first session varied between 0.08 and 0.54, which slightly increased during the second session toward an agreement between -0.01 and 0.75. Pathologists specialized in gynecopathology reached a substantial agreement (kappa 0.75). The top five of criteria indicated to be the most useful in the diagnosis of DVIN included: atypical mitosis in the basal layer, basal cellular atypia, dyskeratosis, prominent nucleoli and elongation and anastomosis of rete ridges. In conclusion, the histopathological diagnosis of DVIN is difficult, which is expressed by low interobserver agreement. Only in experienced pathologists with training in gynecopathology, kappa values reached a substantial agreement after providing strict guidelines. Therefore, it should be considered that specimens with an unclear diagnosis and/or clinical suspicion for DVIN should be revised by a pathologist specialized in gynecopathology. When adhering to suggested criteria the diagnosis of DVIN can be made easier.Modern Pathology advance online publication, 1 February 2013; doi:10.1038/modpathol.2012.235.
Thesis
It has been proposed that there are two types of squamous cell carcinoma of the vulva, one being associated with 'high-risk' human papillomavirus (HPV) with a recognisable pre-malignant lesion (vulval intraepithelial neoplasia; VIN). A second, which appears not to be associated with HPV and about which there, has been controversy relating to the pre-malignant stage. Clinical and histological association has linked non-neoplastic epithelial disorders (e.g. lichen sclerosus; LS) with this form of vulval cancer; however, epidemiological evidence has shown that few patients will progress to develop cancer. Little research has been undertaken to assess the pre-malignant potential of these lesions. The aim of this study was to assess the malignant potential of such lesions as well as the molecular pathology of vulval cancer itself and to elucidate if the pathways to vulval cancer from VIN and LS are different. Using archival tissue and frozen tissue from invasive carcinoma, epidermis adjacent to the cancer (LS, squamous hyperplasia (SH) and VIN) and normal vulval skin, different techniques were used to assess molecular changes both in vulval malignancy and non-neoplastic changes. Such markers were also used in a clinicopathological analysis of survival for vulval cancer. Furthermore, since the treatment of non-neoplastic epithelial disorders may affect such molecular processes these were also analysed. Furthermore blood analysis and clinical features were assessed along with molecular changes to distinguish the similarities or different features between the vulval cancers associated with HPV and the adjacent lesions and vulval cancers associated with non-neoplastic epithelial disorders of the vulva. Neoplastic and non-neoplastic cell cycle and apoptotic proteins were found to demonstrate abnormal expression, particularly in relation to corticosteroid treatment in LS and in neoplastic changes or adjacent to neoplasia. HPV-16 appeared to play a role in the development of both VIN and vulval cancer associated with VIN. Pathways leading to vulval cancer all appear to involve the G1/S pathway though through different means.
Chapter
Vulvar squamous neoplasia clinical management depends on the identification of low-grade squamous intraepithelial lesions (exophytic and flat condyloma), high-grade squamous intraepithelial lesions (classic and differentiated vulvar intraepithelial neoplasias [VINs]), early invasive squamous cell carcinoma, and the different squamous carcinoma morphologies and variants. This chapter provides a comprehensive review of these topics, including the key questions faced by the pathologist during the different stages of the diagnosis process of each of these lesions. Among the updates for this edition, the reader will find (1) new information on the complications of human papillomavirus (HPV) infection in children (genital warts and recurrent respiratory papillomatosis), (2) VIN terminology, gross and frozen section exams pitfalls, and (3) invasive squamous cell carcinoma prevention, clinical/epidemiological background, and outcome. Particularly, this new edition introduces the reader to a recently described category of preinvasive squamous neoplasia named differentiated exophytic vulvar/squamous intraepithelial lesions (DE-VILs or DE-SILs). Although not yet entirely clarified in respect to their pathogenesis and link to squamous carcinoma, these lesions are almost exclusively HPV negative and seen in association with HPV-negative squamous cell and verrucous carcinomas.
Chapter
General approach to the patient and the problemGenitocrural dermatologyMale genital dermatologyFemale genital dermatologyPerineal and perianal dermatologyUmbilical dermatology
Article
Flat low-grade squamous intraepithelial lesion (LSIL) of the vulva [vulvar intraepithelial neoplasia (VIN) 1, flat condyloma] is an uncommon entity with poorly understood biological behavior. We aimed to determine the risk of subsequent vulvar high-grade squamous intraepithelial lesion (HSIL) or carcinoma following a diagnosis of vulvar LSIL/VIN 1, as well as the frequency and predictive value of p16 immunohistochemical expression in this setting. Of the 51 included cases, p16 positivity (diffuse block staining) was identified in 2 (4%). Follow-up data were available in 34 cases, of which 2 (5.9%) developed subsequent vulvar HSIL, including 1/2 p16-positive cases and 1/32 p16-negative cases. The difference in HSIL frequency between p16-positive and p16-negative cases was not statistically significant (P=0.116 for VIN 2+, P=0.061 for VIN 3). For the 18 patients with treatment information available, 10 (56%) received medical or surgical treatment after biopsy. Our results indicate that flat vulvar LSIL is infrequently p16 positive, and that few patients with vulvar LSIL develop subsequent vulvar HSIL. Despite the use of destructive treatment in some cases, the data provide support for the nonpreneoplastic nature of the entity. Immunohistochemical expression of p16 may not be a predictor of HSIL risk in vulvar LSIL, although this result may also be related to the very low rates of both p16 positivity and subsequent vulvar HSIL in our sample. It is clear that vulvar LSIL is distinct from LSIL in other lower anogenital sites in terms of its behavior and p16 expression frequency.
Article
Aims Differentiating between human papilloma virus-dependent vulvar low-grade and high-grade squamous intraepithelial lesions (LSILs and HSILs) remains difficult in selected cases. Stathmin, a protein involved in cell cycle progression, might be a useful additional marker for this differentiation. The aim of this study was to investigate the additional diagnostic value of stathmin expression in vulvar intraepithelial neoplastic (VIN) lesions. Methods Immunohistochemical analysis was used to evaluate stathmin, P16 and Ki67 expression in 91 samples, including LSILs (n=16), HSILs (n=50), differentiated VIN (dVIN; n=10), lichen sclerosis (LS; n=10) and normal vulvar tissue (n=5). Results Stathmin was expressed in more than one-third of the epithelium in all HSILs and in 20% of LSILs. P16 and Ki67 were expressed in more than one-third of the epithelium in 94% of HSILs and in 13% and 40% of LSILs, respectively. Stathmin was expressed in more than one-third of the epithelium in 10% of the dVIN and in none of the LS or normal lesions. P16 and Ki67 expression was not present in more than one-third of the epithelium in any of these lesions. The sensitivity of stathmin for differentiating between LSILs and HSILs was 100% compared with a sensitivity of 94% for both p16 and Ki67. The specificity of stathmin, p16 and Ki67 was 80%, 87% and 60%, respectively. Conclusions Stathmin is a highly sensitive and specific biomarker for the diagnosis of vulvar HSIL. In addition to the more commonly used immunohistochemical markers p16 and Ki67, stathmin can be a useful diagnostic tool for identifying HSILs, especially in cases in which differentiating between LSIL and HSIL is difficult.
Article
Objective. The mature eukaryotic translation initiation factor 5A contains the unusual amino acid hypusine, formed post-translationally from a specific lysine residue and essential for proliferation of eukaryotic cells. We hypothesized that the major eIF5A isoform, eIF5A-1, is an in situ biomarker for proliferation. NIH-353, a polyclonal immunoreagent specific for hypusine-containing eIF5A-1, was used to test this proposal in biopsies of vulvar high-grade intraepithelial neoplasia (VIN), characterized by the presence of proliferating cells throughout the thickness of the epithelium.Methods. Formalin-fixed and paraffin-embedded archival samples with an independently established diagnosis of VIN 3 were stained immunohistochemically after antigen retrieval, employing NIH-353 and, for comparison, the standard Ki-67 antibody.Results. NIH-353 labeled neoplastic keratinocytes throughout the thickness of the epithelium in all VIN 3 samples. Malignant cells in a case of focally invasive squamous cell carcinoma also stained strongly for mature, hypusine-containing eIF5A-1. Epithelium adjacent to these lesions, though still of apparently normal morphology, was immunoreactive throughout its full thickness. At inflammatory foci of lesional sites, solitary reactive lymphocytes were positive, as were individual proliferating cells within dermal appendages. The submucosal stroma lacked reactive cells.Conclusion. NIH-353 identifies mature eIF5A-1 as an in situ biomarker for proliferation. Like Ki-67, this immunoreagent promises broad applicability in histopathological diagnosis and may be helpful in outcome prediction. In contrast to Ki-67, NIH-353 visualizes a molecular target for antineoplastic therapy, and thus may guide the development and clinical testing of drugs that, like the fungicide ciclopirox, inhibit hypusine formation and cell proliferation.
Article
Vulvar intraepithelial neoplasia (VIN) is a precancerous skin disorder of the vulva. It is currently classified on the basis of histological findings as VIN1, −2, or −3. Clinically useful tumour markers do not yet exist. Up to 90% of cases of VIN3 are associated with human papillomavirus (HPV). Even with appropriate treatment, approximately 5% of women with VIN3 will develop cancer of the vulva, necessitating appropriate long-term follow-up. The two main aims of management of VIN are to prevent cancer and to resolve symptoms. Research into VIN has been limited by the rarity of the disease. Most data come from published case series focusing on VIN3.Surgical local excision is a common treatment for VIN2/3. Rotational skin flap procedures are increasingly used when a large area of vulval tissue requires excision. The carbon dioxide laser is also employed for either ablation or excision of high-grade VIN, especially in non-hairy vulval skin. Ablative therapies do not allow histological assessment and unrecognised malignancy may be missed. Medical treatments, such as topical 5% imiquimod cream, or photodynamic therapy with topical 5-aminolaevulinic acid, result in comparatively poor clearance rates of VIN2/3 and long-term outcomes have not been reported. The importance of treatment-related morbidity on sexuality should not be overlooked. HPV vaccines offer an alternative approach. Early work on therapeutic HPV vaccination for high-grade VIN shows that vaccines can stimulate an immune response but not significant clinical improvement. Prophylactic vaccination may be an exciting way to prevent HPV-related diseases, including VIN.
Article
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Article
Formalin-fixed and paraffin-embedded tissue specimens of normal and dysplastic cervical epithelia (five CIN1, seven CIN2, five CIN3, and five normal) were assessed by an immunoperoxidase technique, using the monoclonal antibody MIB1, regonizing a formalin-fixation-resistant epitope on the cell proliferation-associated Ki-67 antigen. An image analysis system was used to measure four parameters associated with proliferative activity: the Ki-67 labelling index (LI), the number of Ki-67-positive nuclei per unit length of basement membrane, and the maximum value and 90th percentile of the relative distances of Ki-67-positive nuclei from the basement membrane. All these four proliferation-related parameters were highly correlated with the grade of dysplastic change in the epithelium (0·90<r<0·97,p<0·0001). The best correlation was found for the 90th percentile of the relative distance and with this parameter all CIN lesions could be correctly classified. The means and standard deviations of the Ki-67 LIs in normal epithelium, CIN1, CIN2, and CIN3 lesions were 0·07±+0·03, 0·16±0·03, 0·25±0·06, and 0·39±0·06, respectively. These findings support the theory that CIN involves a progressive dysfunction of the proliferative activity of cervical epithelial cells. Image analysis of MIB1 is a promising alternative method for the objective, reproducible, and reliable classification of dysplastic changes in cervical epithelium.
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A previously described coefficient of agreement for nominal scales, kappa, treats all disagreements equally. A generalization to weighted kappa (Kw) is presented. The Kw provides for the incorpation of ratio-scaled degrees of disagreement (or agreement) to each of the cells of the k * k table of joint nominal scale assignments such that disagreements of varying gravity (or agreements of varying degree) are weighted accordingly. Although providing for partial credit, Kw is fully chance corrected. Its sampling characteristics and procedures for hypothesis testing and setting confidence limits are given. Under certain conditions, Kw equals product-moment r. The use of unequal weights for symmetrical cells makes Kw suitable as a measure of validity.
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In this article, vulvar anatomy and histology are reviewed, and nonneoplastic and neoplastic epithelial disorders are presented. Included are lichen sclerosus, squamous cell hyperplasia, vulvar intraepithelial neoplasia, vulvar Paget's disease, and squamous cell carcinoma. Current terminology, tumor measurements, and staging for vulvar carcinoma are reviewed.