Immature Squamous Metaplasia Research Articles

Inhibition of cytosine 5-hydroxymethylation during progression of cancer precursor lesions in the uterine cervix.

Methylation and hydroxymethylation of cytosine moieties in CpG islands of specific genes are epigenetic processes shown to be involved in the development of cervical (pre)neoplastic lesions. We studied global (hydroxy)methylation during the subsequent steps in the carcinogenic process of the uterine cervix by using immunohistochemical protocols for the detection of 5-methylcytosine (5-mC) and 5-hydroxymethylcytosine (5-hmC) in paraffin-embedded tissues of the normal epithelia and (pre)malignant lesions. This approach allowed obtaining spatially resolved information of (epi)genetic alterations for individual cell populations in morphologically heterogeneous tissue samples. The normal ectocervical squamous epithelium showed a high degree of heterogeneity for both modifications, with a major positivity for 5-mC in the basal and parabasal layers in the ectocervical region, while 5-hmC immunostaining was even more restricted to the cells in the basal layer. Immature squamous metaplasia, characterized by expression of SOX17, surprisingly showed a decrease of 5-hmC in the basal compartments and an increase in the more superficial layers of the epithelium. The normal endocervical glandular epithelium showed a strong immunostaining reactivity for both modifications. At the squamocolumnar junctions, a specific 5-hmC pattern was observed in the squamous epithelium, resembling that of metaplasia, with the typical weak to negative reaction for 5-hmC in the basal cell compartment. The reserve cells underlying the glandular epithelium were also largely negative for 5-hmC but showed immunostaining for 5-mC. While the overall methylation status remained relatively constant, about 20% of the high-grade squamous lesions showed a very low immunostaining reactivity for 5-hmC. The (pre)malignant glandular lesions, including adenocarcinoma in situ (AIS) and adenocarcinoma showed a progressive decrease of hydroxymethylation with advancement of the lesion, resulting in cases with regions that were negative for 5-hmC immunostaining. These data indicate that inhibition of demethylation, which normally follows cytosine hydroxymethylation, is an important epigenetic switch in the development of cervical cancer.

Expression of the O-Linked N-Acetylglucosamine-containing Epitope H (O-GlcNAcH) in Human Uterine Cervical Mucosa.

Epitope H contains an O-linked N-acetylglucosamine (O-GlcNAcH) residue in a specific conformation or environment, recognized by a site-specific monoclonal mouse IgM antibody H. O-GlcNAcH occurs in several normal and pathological cells and in several polypeptides, including keratin-8 and vimentin, on the latter in cells under stress. In this work, we studied the distribution of O-GlcNAcH on cells of endocervical mucosa in 60 specimens of endocervical curettings, 10 of which contained 15 inflamed polyps. In our results, expression of O-GlcNAcH was weak in the mucosa with <5% mucin-secreting cells and up to 30% of the polyps staining positively. All non-ciliated, non-mucin-secreting cells, normal and hyperplastic 'reserve' cells, as well as the cells of immature squamous metaplasia, showed strong diffuse cytoplasmic staining for O-GlcNAcH. In mature squamous epithelium, fewer than 5% of basal cells and all the intermediate and superficial cells showed cytoplasmic staining for O-GlcNAcH, whereas parabasal cells were negative. All ciliated cells showed patchy or diffuse cytoplasmic staining. Nuclear staining for O-GlcNAcH was weak with fewer than 5% of hyperplastic 'reserve' and ciliated cells staining positively. Moreover, mucosal fibroblasts were negative, whereas all stromal cells of the polyps showed strong cytoplasmic staining for O-GlcNAcH. O-GlcNAcH is: a) differentially expressed among the cellular elements of mucosa and polyps, b) upregulated in mucin-secreting cells of polyps, c) induced in stromal cells of inflamed polyps, and d) can be used as a marker to differentiate between 'reserve' (positive) and parabasal (negative) cells, which have similar morphology using conventional cytological stains.

CIN grades possessing different HPV RNA location patterns and RNAscope is helpful tool for distinguishing squamous intraepithelial lesions in difficult cervical cases

Background and objectivesThe precise grading and characterization of cervical intraepithelial neoplasia (CIN) has been the focus of pathologists for a long time. This study aimed to explore known strategies for the grading of CINs.MethodsAfter routine H&E review, 85 lesions graded CIN 1, 2, or 3 were investigated primarily by HPV RNAscope to detect HR-HPV and LR-HPV, in combination with an HPV-DNA test and P16/Ki67 immunohistochemistry (IHC). Then, the 85 cases were divided into a control group (49 cases) and a test group (36 cases). The former consisted of cases with consistency between morphology, HPV DNA detection and P16/Ki67 IHC. We used them to evaluate HPV RNA distribution patterns in CINs of different grades. The latter were ambiguous cases in which pathologists could not confirm the diagnosis because of inconsistencies between morphology, HPV DNA detection and P16/Ki67 IHC. We reassessed them by comparison to the pattern in the control group.ResultsThe expression patterns of HPV mRNA signals were different in different CIN lesions. LSIL/CIN1 lesions were mostly expressed in superficial epithelium with diffuse clustered nuclear or cytoplasmic staining; HSIL/CIN2 were characterised by nuclear/cytoplasmic punctate or diffuse cluster nuclear staining in the mid-surface layer, and scattered nuclear/cytoplasmic punctate staining in basal and parabasal cells; whereas HSIL/CIN3 showed full-thickness nucleus/cytoplasmic scattered staining with a punctate pattern. According to the staining pattern, we corrected the diagnosis of 22 cases (22/36, 61.1%).ConclusionBecause of its distinct location pattern, HPV RNAscope has obvious advantages over the HPV-DNA test, and combined with P16/Ki67 IHC, it can help pathologists correctly grade CIN. In addition, it can effectively discriminate true CIN from normal or CIN mimic lesions, such as immature squamous metaplasia, atrophy, and inflammatory/reactive changes. Therefore, HPV RNAscope is a valuable auxiliary diagnostic test to avoid the overtreatment and undertreatment of CIN lesions.

Immature Squamous Metaplasia of Endocervix – A Great Mimicker of Invasive Squamous Cell Carcinoma of Cervix

Immature Squamous Metaplasia of Endocervix – A Great Mimicker of Invasive Squamous Cell Carcinoma of Cervix

Stratified Mucin Producing Intraepithelial Lesion (SMILE): Report of Two Cases

Abstract Introduction/Objective Stratified mucin producing intraepithelial lesion (SMILE) is an uncommon variant of adenocarcinoma in-situ (AIS), often confused with immature squamous metaplasia or high-grade intraepithelial lesion (HG-SIL). This premalignant lesion is often found in proximity to HG-SIL, AIS and invasive carcinomas. SMILE is also a precursor for Invasive Stratified Mucinous Carcinoma. As was first described in the original article, SMILE lesions are considered derived from the reserve cells at the transformation zone where HPV infections lead to varied phenotypes. Methods Two cases of SMILE were collected from our files. A 31-year-old female with a previous abnormal Pap smear result. She underwent cervical biopsy and subsequent LEEP.The second patient, a 26-year-old female, underwent a cervical biopsy after an abnormal Pap smear. Results Histologically, stratified cells with presence of conspicuous mucin production with abundant cytoplasm separating cells in mid and lower layer of epithelium accompanied with conspicuous spacing of nuclei in the lower to middle epithelial layer are seen. Additionally, there is nuclear pleomorphism with absence of gland formation. Mucicarmine stain was performed and showed positive staining in mucin producing dysplastic cells throughout the epithelium. Immunoperoxidase stains demonstrated positivity for p16 (nuclear and cytoplasmic) and p63 in the basal cells. The Ki67 proliferation index was high (approximately 90%). These results supported the diagnostic interpretation of SMILE. Conclusion SMILE is a rare variant of glandular precursor lesion in the cervix. It is frequently associated with HG-SIL and/or AIS, but may rarely appear as an isolated finding. The differential diagnosis include adenocarcinoma in-situ, atypical immature squamous metaplasia and squamous intraepithelial lesion. Since these lesions are cervical glandular intraepithelial neoplasms, the morphologic recognition of a SMILE in a biopsy is important for patient follow up, particularly in cases were only squamous intraepithelial lesion is present in the sample.

Clinical Utility of Biomarkers in Diagnosis and Management of High Grade Cervical Intraepithelial Neoplasia (HSIL)

Objective of the study: The increasing inter- and intra-observer variability in the diagnosis of CIN by histopathology has led to the advent of different biomarkers. The aim of this study is to evaluate Ki-67 and p16 INK4a marker in differentiating CIN lesions and the clinical utility of it in the management of high grade squamous intraepithelial lesions (HSIL).&#x0D; Materials and methods: This cross-sectional study has been carried out among the biopsy proved cases of CIN2 and CIN3 in which P16 and Ki-67 immunostain was performed in the Colposcopy clinic, Bangabandhu Sheikh Mujib Medical University (BSMMU) and Department of Pathology, BSMMU respectively from 14-3-17 to 30-05-18. Thirty-two cases were incorporated in the study by purposive sampling. Data was analyzed using SPSS version 17.&#x0D; Results: Among 32 cases of HSIL, 25 were diagnosed as CIN 2 (78.13 %) and 7 cases as CIN 3 (21.87 %) by histopathology. All the cases were subjected to immunohistochemistry with P16 and Ki 67. Regarding the status of P16, it was found positive only in 32% cases of CIN 2 but in 100% cases of CIN 3. Similarly, Ki67 was found positive in 36 % of CIN2 cases but in all cases of CIN 3 lesions. Thus immunostain confirmed one third histopathologically diagnosed CIN 2 cases. On the other hand, all cases of CIN 3 were confirmed as HSIL with the immunostain. All the cases of CIN 2 were reviewed for consensus opinion. Only 8 (32%) cases of CIN 2 were both immunostain positive , the review diagnosis by consensus opinion was also CIN 2 in those cases. One case (4%) of CIN 2 was P16 negative but Ki 67 positive, which ultimately revealed as immature squamous metaplasia. Sixteen cases of CIN 2 were negative for both P16 and Ki 67. Among these, majority cases (52%) , consensus opinion revealed as CIN 1 and 12% cases as immature squamous metaplasia. Therefore, an HSIL diagnosed by histopathology could be false positive in a significant number of cases and overtreatment of a false positive CIN 2 lesion can affect future reproductive status, also have some psychological consequences and economic burden in the health sector.&#x0D; Conclusion: By using the biomarkers, overtreatment and undertreatment of CIN can be avoided. All cases of CIN 2 should be carefully dealt with either review of the slides by consensus opinion or by doing immunohistochemistry if facility is available.&#x0D; Bangladesh J Obstet Gynaecol, 2019; Vol. 34(2): 79-86

The Challenge of Pathological Diagnosis for Precancerous Cervical Lesions

Cervical cancer is the third most common cancer after breast and colorectal cancers worldwide, and the second most common gynecological malignancy after endometrial cancer. Cervical cancer screening was first described by Papanicolaou in 1941 using the PAP smear test. The incidence of invasive cervical cancer clearly reduced with the common use of PAP smear in developed countries. The basic methods used for diagnosis of premalignant lesions of the cervix are determination with colposcopy, biopsy and HPV DNA typing. Cervical premalignant lesions cannot be observed with the naked eye, other than exophytic or papillary lesions of condyloma acuminatum. Condyloma acuminatum are simultaneously LSIL. LSIL and HSIL differentiation cannot be made with colposcopy. Due to better repeatability and interobserver compliance, the World Health Organization (WHO) recommends a 2-layer HSIL/LSIL system. In the 3-layer CIN system, CIN1 is equivalent to LSIL, while CIN3 equivalent is HSIL. The lesions are frequently encountered in routine biopsies, and in some cases, differential diagnosis may be difficult. Basal cell hyperplasia, atrophy, reactive and repair-induced atypia and immature squamous metaplasia may mimic precancerous lesions. Although histomorphology is gold standard, P16 and Ki-67 are beneficial immunohistochemical ancillary testes. However, it should be kept in mind that p16 can be positive in LSIL, and negative in HSIL.

Ectopic prostatic tissue in vagina - case report

During embryonic development, both glandular parenchyma of prostate in a male fetus and paraurethral Skene’s glands in a female fetus develop from pelvic urethra endoderm. This common embryological origin results in finding prostatic tissue even in the lower genital tract in women. Herein, the authors present a case of polypoid lesion in vagina consisting solely of glandular elements without squamous component, which stained negative for prostatic markers, but morphologically identical to normal prostatic tissue. A patient, 57-years-old came to the Clinic, with a cystic change in the petiole, which size being 30×17×15 mm, on the left vaginal wall adjacent to introitus. The cystic change was excised and a preparation was sent to a histopathological examination. The histopathological examination included serial Hematoxylin and Eosin stained sections, along with immunohistochemical panel. It was concluded that it was ectopic prostatic tissue or ectopic paraurethral gland tissue within vaginal mucosa. Partially, epithelium at the surface of our polypoid lesion showed immature squamous metaplasia, vaginal intraepithelial neoplasia - VaIN II/III, and transitional cell metaplasia. Gland formations having benign features were within base of the change, at the resection lines, while dysplastic epithelium was fully removed.

Role of p16/INK4a and Ki-67 as specific biomarkers for cervical intraepithelial neoplasia: An institutional study.

BACKGROUND:P16/INK4a and Ki-67 have emerged as important biomarkers for the detection of high-risk human papilloma virus (HR-HPV) associated dysplastic changes in the cervical biopsy samples. The increasing inter- and intra-observer variability in the diagnosis of dysplastic lesions and immature squamous metaplasia on histopathology has led to the advent of these biomarkers. This study was taken up with an aim to study their role in increasing the diagnostic accuracy in equivocal cases on histopathology.MATERIALS AND METHODS:Fifty cervical biopsy specimens were stained with p16/INK4a and Ki-67 consisting of 10 cases each of cervical intraepithelial neoplasia (CIN I/II/III) along with five cases of squamous metaplasia. Histopathological diagnosis was considered as the gold standard. Statistical analysis was done by kappa statistics, and P value was calculated.RESULTS:The sensitivity and specificity of p16/INK4a and Ki-67 were 76.2%, 87.5%, 90.5%, and 87.5%, respectively. The overall agreement of both the immunostains with histopathological diagnosis was statistically significant (P < 0.05) and the diagnostic accuracy improved when both the stains were used in conjunction.CONCLUSION:Ki-67 and p16/INK4a can be used as complimentary tests in differentiating dysplastic and nondysplastic lesions and help in confirming the histopathological diagnosis. They aid in recognition of dysplasias caused by HR-HPV, which have higher tendency to progress to neoplasia. However, further research is advocated before the widespread use of these markers for screening of dysplasias.

Role of p16, Ki67 and CK17 in differentiating benign lesions, cervical intraepithelial neoplasia(CIN) and atypical immature squamous metaplasia(AIM) of uterine cervix

Cervical intraepithelial neoplasia (CIN) is a premalignant lesion capable of progressing to cervical cancer. Despite the existing well-defined criteria, the histopathological diagnosis is subject to high rates of discordance among pathologists. Aim: To study the role of p16, Ki67 and CK17 in differentiating benign lesions, cervical intraepithelial lesions(CIN) and atypical immature squamous metaplasia (AIM)and to improve intra and interobserver reproducibility of diagnosis of cervical neoplasia. Material and Methods: In a cross sectional study, a total of 75 cervical biopsies including benign lesions (n=24), AIM (n=28), CIN (n=23) were studied and analyzed immunohistochemically using p16, Ki67 and CK17 immunomarkers. Data was evaluated using chi-square test. Results: p16 and Ki 67positivity were observed in 91.3% and 78.26% of CIN and 28.57% of AIM respectively. None of the benign lesions expressed p16 and Ki67while CK17 positivity was observed in 46.42% of CIN and 100% of AIM with 12.5% of benign lesions. Conclusion: The three biomarkers (p16, CK17 and Ki67) had a high degree of sensitivity and specificity and appear to be a useful and reliable diagnostic adjunct to improve the routine diagnosis and reduce interobserver variability in cervical biopsy specimens. Immunohistochemical markers such as p16 alone or with Ki67 represents important tool for the pathologists in distinguishing high grade cervical dysplasia from its benign mimics such as AIM and reactive inflammatory lesion thus avoiding overtreatment. Cervical intraepithelial neoplasia (CIN) is a premalignant lesion capable of progressing to cervical cancer. Despite the existing well-defined criteria, the histopathological diagnosis is subject to high rates of discordance among pathologists. Aim: To study the role of p16, Ki67 and CK17 in differentiating benign lesions, cervical intraepithelial lesions(CIN) and atypical immature squamous metaplasia (AIM)and to improve intra and interobserver reproducibility of diagnosis of cervical neoplasia. Material and Methods: In a cross sectional study, a total of 75 cervical biopsies including benign lesions (n=24), AIM (n=28), CIN (n=23) were studied and analyzed immunohistochemically using p16, Ki67 and CK17 immunomarkers. Data was evaluated using chi-square test. Results: p16 and Ki 67positivity were observed in 91.3% and 78.26% of CIN and 28.57% of AIM respectively. None of the benign lesions expressed p16 and Ki67while CK17 positivity was observed in 46.42% of CIN and 100% of AIM with 12.5% of benign lesions. Conclusion: The three biomarkers (p16, CK17 and Ki67) had a high degree of sensitivity and specificity and appear to be a useful and reliable diagnostic adjunct to improve the routine diagnosis and reduce interobserver variability in cervical biopsy specimens. Immunohistochemical markers such as p16 alone or with Ki67 represents important tool for the pathologists in distinguishing high grade cervical dysplasia from its benign mimics such as AIM and reactive inflammatory lesion thus avoiding overtreatment. Normal 0 false false false EN-IN X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:Table Normal; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-qformat:yes; mso-style-parent:; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin-top:0cm; mso-para-margin-right:0cm; mso-para-margin-bottom:10.0pt; mso-para-margin-left:0cm; line-height:115%; mso-pagination:widow-orphan; font-size:11.0pt; font-family:Calibri,sans-serif; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:Times New Roman; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin;}

P16INK4a Expression in Cervical Lesions Correlates with Histologic Grading - a Tertiary Level Medical Facility Based Retrospective Study

p16INK4a is a tumor-suppressor protein and cyclin-dependent kinase (cdk) inhibitor that blocks cdk4- and cdk6-mediated pRb phosphorylation to inhibit E2F-dependent transcription and cell-cycle progression. Because the E7 protein of high-risk HPVs inactivates pRB, the resulting overexpression of p16INK4a may be a good marker for infection with high risk HPV types. Immunostaining of p16INK4a allows precise identification of even small CIN or cervical cancer lesions in biopsy sections and can help reduce inter-observer variation in the histopathological interpretation of cervical biopsy specimens. The aims of the present study were to evaluate the expression of p16 INK4a in cervical biopsies and to compare the grade of cervical neoplasia with intensity of staining. The study covered 110 cervical biopsy tissue blocks over a period of 2 years, (85 cases of CIN of varying grade and invasive cervical cancers and 25 of non-neoplastic lesions). Immunostaining with p16INK4a antibodies followed standard operating procedures. The results showed an increasing trend for p16INK4a immunoreactivity from benign to higher grade lesions. Out of 25 cases of non dysplasia (15 cervicitis &10 immature squamous metaplasia), 8%(2/25) showed P16INK4a expression (grade 1). Among low grade lesions like CIN1, 32% (8/25) cases demonstrated P16INK4a expression (grade 1). Some 52.3% (11/21) of CIN2 cases proved positive. The intensity of p16INK4a expression in CIN 2 was grade 1 in 33%, grade 2 in 14% and grade 3 in 4.7% of cases. All the CIN3 lesions and cervical squamous cell carcinomas exhibited grade 3 anti p16INK4a antibody staining. The association of p16INK4a expression with histologic grade of cervical pathology was highly significant (χ2-value:51.81, p<0.0001). The staining intensity increase with higher grade disease was also statistically significant (χ2-value :133.95, p<0.0001).

Expression of HPV16/18, p16, CK17, Ki-67 in cervical squamous epithelial lesions

Objective To investigate the human papillomavirus 16/18 (HPV16/18) infective status and its relationship with the expression of p16, CK17, Ki-67 in immature squamous metaplasia(IM), cervical intraepithelial neoplasia (CIN) and cervical carcinomas (CC). Methods In situ hybridization (ISH) and immunohistochemistry were applied to detect the expression of HPV16/18, p16, CK17, Ki-67 on tissues from 30 IM, 60 CIN and 30 CC. Results The expression rates of HPV16/18 in LSIL, HSIL and CC were 30 % (9/30), 60 % (18/30) and 77 % (23/30), higher than that in IM [6.7 % (2/30)]. HPV16/18 was positively expressed with the progression of cervical squamous epithelial lesions. The expression rates of CK17 in IM was 90 % (27/30), higher than that in LSIL, HSIL and CC [17 % (5/30), 10 % (3/30), 6.7 % (2/30)]. The expression rates of p16, Ki-67 were 83 % (25/30) and 87 % (26/30) in LSIL, 90 % (27/30) and 93 % (29/30) in HSIL, 97 % (29/30) and 97 % (29/30) in CC, higher than that in IM [13 % (4/30) and 10 % (3/30)]. The expression of p16, Ki-67 were particularly seen in HPV16/18-positive HSIL and CC, and the correlation were observed. Conclusion HPV16/18 infection is highly associated with the degree of squamous intraepithelial neoplasia and carcinomas of cervix and upregulated p16 and Ki-67, which suggested that HPV16/18 maybe cause mutation of p16 gene. With the progression of cervical squamous epithelial lesions, positive expression rates of CK17 were decreased compared with increased of HPV16/18, p16, Ki-67. Combined detection of HPV16/18, p16, Ki-67 is helpful for diagnosis and classification of cervical squamous epithelial lesions. Key words: Human papillomavirus 16/18; CK17; p16; Ki-67; Immature squamous metaplasia; Cervical intraepithelial neoplasia; Cervical squamous cell carcinomas

Immunohistochemistry and in situ hybridization for the diagnosis and classification of squamous lesions of the anogenital region.

Distinguishing anogenital squamous intraepithelial lesions from benign conditions and mimics may be problematic. Immunohistochemistry for surrogate markers of HPV infection, such as Ki-67, p16, and ProEx™ C, may aid the diagnosis in equivocal cases. The main diagnostic pitfall in the diagnosis of LSIL is the occurrence of "pseudokoilocytes" in benign squamous mucosa, which may lead to overdiagnosis. When interpreted correctly, Ki-67 is a sensitive and specific marker for dysplasia in mature squamous epithelium and is therefore useful for confirmation of LSIL and condyloma. A Ki-67 positive result is defined as the presence of a cluster of at least two strongly stained epithelial nuclei in the upper two-thirds of the epithelial thickness. With such a definition, there is almost complete concordance between consensus diagnosis of LSIL/condyloma confirmed by detection of HPV DNA and positive Ki-67. A related proliferation marker, ProEx™ C, has similar staining patterns and utility for the diagnosis of low grade dysplasia. The differential diagnosis of HSIL includes atypical immature squamous metaplasia and atrophy. A marker with high sensitivity and specificity for the detection of HSIL in cervical, vulvar, and anal mucosa is p16. A 2-tier scoring system is used to evaluate p16 staining. No staining or a discontinuous, patchy nuclear and cytoplasmic staining pattern is considered as a negative result. A positive result is defined as diffuse and strong staining of cells of the basal and parabasal layers of the squamous epithelium, with or without staining of superficial cell layers. New markers that are undergoing evaluation for their clinical utility include stathmin-1, phosphorylated S6, and SOX2. Confirmation of the diagnosis of dysplasia by HPV detection in tissue sections using HPV capsid protein immunohistochemistry, HPV DNA or HPV RNA in situ hybridization offers lower sensitivity as compared to immunohistochemistry for surrogate markers and therefore has more limited utility in this context.

Gynaecological Screening for Cervical and Vulvar Malignancies in a Cohort of Systemic Sclerosis Patients: Our Experience and Review of the Literature

Background. Increased incidence of cancer was frequently reported in scleroderma (SSc), but no association with gynaecological malignancies was described in literature. Objectives. To investigate gynaecological neoplasms in SSc patients. Methods. In this cross-sectional analysis, we evaluated 80 SSc patients, living in the same geographical area. We considered all patients undergoing gynaecological evaluation, including pap test as screening for cervical cancer, between January 2008 and December 2014. Results. 55 (68.7%) patients were negative and 20 (25%) presented inflammatory alterations, while cancer or precancerous lesions were found in 5 (6.2%) cases (2 showed cervical cancer (one of them in situ), 1 vulvar melanoma, 1 vulvar intraepithelial neoplasia, and 1 endocervical polyp with immature squamous metaplasia). The frequency of cervical cancer in our series seems higher in comparison to the incidence registered in the same geographical area. The presence of atypical cytological findings correlated with anti-Scl70 autoantibodies (p = 0.022); moreover, the patients with these alterations tended to be older (median 65, range 46–67), if compared to the whole series (p = 0.052). Conclusions. A relatively high frequency of gynaecological malignancies was found in our SSc series. In general, gynaecological evaluation for SSc women needs to be included in the routine patients' surveillance.

Fascin expression in cervical normal squamous epithelium, cervical intraepithelial neoplasia, and superficially invasive (stage IA1) squamous carcinoma of the cervix

The aims of this study were to: investigate fascin expression in normal cervix, cervical intraepithelial neoplasia (CIN), and stage IA1 squamous cell carcinoma (SCC).Fascin immunostaining was performed in cervical biopsies showing normal squamous epithelium (n=10), CIN 1 (n=10), CIN 2-3 without invasion (n=11), and CIN 2-3 adjacent to SCC (n=40); SCC was also present in 27 of the latter cases.Fascin expression in normal squamous epithelium was restricted to basal and parabasal cells, whereas there was increased staining in immature squamous metaplasia and in most CIN lesions. Full thickness staining was more frequent in high grade CIN adjacent to invasion than in CIN 2-3 alone. Eighteen SCCs (67%) were fascin positive and seven cases showed accentuated staining at the tumour-stromal interface (invasive front). There was no consistent relationship between fascin expression in CIN lesions and in corresponding carcinomas. Fascin staining in reactive stromal cells sometimes made identification of the invasive neoplastic cells difficult.Fascin is overexpressed in most CIN lesions and this may be a marker of increased invasive potential in high grade CIN. However, fascin staining does not distinguish low and high grade CIN or in situ and invasive squamous neoplasia. Therefore fascin has limited diagnostic utility in demonstrating superficial stromal invasion.

Keratin 17 in premalignant and malignant squamous lesions of the cervix: proteomic discovery and immunohistochemical validation as a diagnostic and prognostic biomarker

Most previously described immunohistochemical markers of cervical high-grade squamous intraepithelial lesion (HSIL) and squamous cell carcinoma may help to improve diagnostic accuracy but have a minimal prognostic value. The goals of the current study were to identify and validate novel candidate biomarkers that could potentially improve diagnostic and prognostic accuracy for cervical HSIL and squamous cell carcinoma. Microdissected tissue sections from formalin-fixed paraffin-embedded normal ectocervical squamous mucosa, low-grade squamous intraepithelial lesion (LSIL), HSIL and squamous cell carcinoma sections were analyzed by mass spectrometry-based shotgun proteomics for biomarker discovery. The diagnostic specificity of candidate biomarkers was subsequently evaluated by immunohistochemical analysis of tissue microarrays. Among 1750 proteins identified by proteomic analyses, keratin 4 (KRT4) and keratin 17 (KRT17) showed reciprocal patterns of expression in the spectrum of cases ranging from normal ectocervical squamous mucosa to squamous cell carcinoma. Immunohistochemical studies confirmed that KRT4 expression was significantly decreased in squamous cell carcinoma compared with the other diagnostic categories. By contrast, KRT17 expression was significantly increased in HSIL and squamous cell carcinoma compared with normal ectocervical squamous mucosa and LSIL. KRT17 was also highly expressed in immature squamous metaplasia and in endocervical reserve cells but was generally not detected in mature squamous metaplasia. Furthermore, high levels of KRT17 expression were significantly associated with poor survival of squamous cell carcinoma patients (Hazard ratio=14.76, P=0.01). In summary, both KRT4 and KRT17 expressions are related to the histopathology of the cervical squamous mucosa; KRT17 is highly overexpressed in immature squamous metaplasia, in HSIL, and in squamous cell carcinoma and the level of KRT17 in squamous cell carcinoma may help to identify patients who are at greatest risk for cervical cancer mortality.

Role of p16, CK17, p63, and Human Papillomavirus in Diagnosis of Cervical Intraepithelial Neoplasia and Distinction From Its Mimics

Diagnosis of cervical intraepithelial neoplasia (CIN), the precursor forms of cervical cancer, can be tricky and it has led to discrepancy between pathologists in distinguishing them from its mimics such as atypical immature metaplasia (AIM), immature squamous metaplasia (ISM), reactive atypia (RA), atrophy, and basal cell hyperplasia (BCH). To overcome this problem this study aims at using immunohistochemical (IHC) markers p16, p63, CK17, and human papillomavirus (HPV) to differentiate CIN from its mimics. This study analyzed 350 cervical samples with histomorphological diagnosis of CIN and its mimics and the utility of IHC markers p16, p63, CK17, and HPV in distinction was analyzed. p16 showed 67.76% sensitivity and 99.4% specificity whereas HPV showed 57.9% sensitivity and 91.6% specificity in detecting CIN. CK17 and p63 did not show any significance in distinguishing CIN from its mimics. After IHC of AIM cases, 66.7% were reclassified as CIN III, 27.8% as ISM with reactive atypia (ISMRA), and 5.5% case as immature condyloma. In total, 3.7% of diagnosis was upgraded to CIN and 0.6% of pre-IHC diagnosis was downgraded from CIN to reactive lesions. IHC panel comprising p16, p63, CK17, and HPV are useful adjuncts in distinguishing CIN from its mimics particularly when histomorphology has overlapping morphological features.

Immature squamous metaplasia (focal atypical epithelial hyperplasia) of the pancreatic duct—immunohistochemical distinction from intraductal carcinoma

Immature squamous metaplasia of the pancreatic duct (ISMPD) can be difficult to differentiate from an intraductal carcinoma of the pancreas (ICP), and little is known about the pathological nature of ISMPD. The aim of this study was to analyse 20 ISMPD and 10 ICP tissue samples. ISMPD shares some characteristics with ICP. Seven of 20 ISMPD samples were covered by a layer of pancreatic duct epithelium, whereas this was not seen in the ICP samples. Immunohistochemistry of ISMPD revealed positivity for p63 (100%), cytokeratin 5/6 (95%), cytokeratin 7 (95%), cytokeratin 20 (10%), and MUC-1 (95%), and the samples were negative for p53, carcinoembryonic antigen (CEA), and bcl-2. In contrast, ICP was positive for p63 (40%), p53 (10%), cytokeratin 7 (90%), cytokeratin 20 (20%), CEA (30%), and MUC-1 (80%), and negative for cytokeratin 5/6. However, in 84% (16) of the ISMPD samples, cytokeratin 7 was expressed only by an epithelial layer at the apical surface; this expression pattern was not found in any of the 10 ICP samples. The mean Ki67 labelling index was 1.0% in ISMPD and 18.5% in ICP. Our study suggests that immunohistochemical staining for cytokeratin 5/6 and Ki67 constitutes the best combination for differentiating ISMPD from ICP.

P16INK4a Immunoprofiles of Squamous Lesions of the Uterine Cervix–Implications for the Reclassification of Atypical Immature Squamous Metaplasia

p16(INK4a) immunoprofiles of non-precancerous and dysplastic squamous cervical lesions were defined and applied to the reclassification of atypical immature squamous metaplasia (AIM). The immunoexpression of cytokeratin 17 (CK 17) in AIM was also evaluated. Totally, 295 cervical cone biopsies representing squamous metaplasia, reactive changes, koilocytosis, flat condyloma, CIN I, CIN II, CIN III and AIM were subjected to p16(INK4a) immunohistochemistry. AIM cases were analyzed using CK 17 antibody. Typical p16(INK4a) immunoprofiles for the metaplastic, LSIL/HPV and HSIL phenotypes were recorded and used for the categorization of AIM into particular phenotype groups. Results were correlated with CK 17 immunoexpression. All CIN II and CIN III lesions, all but one case of CIN I and all flat condylomas overexpressed p16(INK4a). Other non-precancerous lesions, including koilocytosis, were predominantly negative. Contrary to the sporadic and focal immunostaining, diffuse positivity was associated with the dysplastic features of the lesion. CIN II and CIN III were characterized by a diffuse, strong/weak, full-thickness staining, whereas CIN I showed a heterogeneous diffuse/focal, weak/strong, lower half positivity. One third of AIM lesions may be reclassified as HSIL, one third as LSIL/HPV and one third shows metaplastic phenotype. All AIM cases with metaplastic and LSIL/HPV phenotypes expressed CK 17 diffusely, whereas focal positivity slightly prevailed in AIM with HSIL phenotype. We conclude that p16(INK4a) immunohistochemistry is a supporting method for the differential diagnosis of cervical lesions, which may be especially useful for the reclassification of AIM. The efficacy of CK 17 immunohistochemistry seems to be controversial for these purposes.

Expression of defensins in the lungs in acute respiratory distress syndrome associated with influenza pneumonia

Summary. We investigated paraffin slices of the lungs from 35 people died during the 2009 epidemic of influenza A / H1N1. The exudative stage of acute respiratory distress syndrome (ARDS) was diagnosed in 10 cases, the proliferative stage in 16 and the fibrotic stage in 9 cases. Expression of α–defensins (HNP 1–3) in lung tissue was determined by the immunohistochemistry. The highest HNP 1–3 expression level was found during the exudative stage in neutrophils; it was similar in macrophages and alveolocytes type 2. α–defensins were also expressed by the epithelium of terminal bronchioles. During the proliferative stage, the HNP 1–3 expression in neutrophils remained at the high level, it was increased in alveolocytes type 2 and was unchanged in macrophages. During this stage, HNP 1–3 additionally expressed by immature alveolocytes and squamous metaplasia epithelial cells in the terminal bronchioles. During the fibrotic stage, the HNP 1–3 expression in neutrophils was reduced but remained the highest in comparison with that in the other cells; it did not changed in the alveolocytes type 2 and the macrophages. In this stage, HNP 1–3 also expressed in proliferating fibroblasts.