Blood Vessel Invasion Is an Independent Prognostic Factor in Endometrial Endometrioid Carcinoma Compared to Lymph Vessel Invasion and Myometrial Invasion Pattern.

We studied 115 cases of EEC diagnosed on hysterectomy specimens. Double immunohistochemical staining (D2-40/CD31) was performed in all 115 cases to show LVI and BVI on the same slide. MELF pattern invasion was present in 24/115 (21%) cases. MELF-positive tumors had a higher frequency of LVI than MELF-negative tumors (58% and 23%, respectively); the frequency of BVI was twice as high in MELF-positive tumors in comparison to MELF-negative tumors (25% and 12%, respectively). These differences were significant (p ˂ 0.0001). All tumors with positive BVI also had a concomitant LVI. The presence of MELF invasion had no impact on overall survival, confirming previous studies. 5-year survival rates were almost equal in cases with negative LVSI and cases with positive isolated LVI (98% vs. 97%). However, in cases where BVI was also present, the 5-year survival rate was significantly lower, 63% (p ˂ 0.0001). Furthermore, BVI proved to be an independent prognostic factor for overall survival, disease-free survival, and recurrence in the multivariate analysis. In conclusion, MELF pattern invasion is a good predictor of lymphatic and blood vessel invasion but has no prognostic value. Our results suggest that BVI in EEC has greater clinical value than isolated LVI or myometrial invasion patterns, and the therapeutic approach should be guided by BVI presence. Therefore, we hope this study will promote the routine evaluation of BVI in the context of EEC diagnostic procedures.

Pitfalls in Gynecological Cytology: Review of the Common and Less Frequent Entities in Pap Test.

BACKGROUND: Pitfalls in Pap test could be defined as false positive, false negative, or underdiagnosed results which can lead to unnecessary diagnostic procedures or delayed and inadequate treatment. It can be a consequence of misinterpretation of certain morphological entities which are described in this paper. SUMMARY: The paper presents an overview of the morphological features and look-alikes of the common sources of pitfalls such as atrophy, repair, intrauterine device change, tubal metaplasia, hyperchromatic crowded groups, and radiation changes. Rare causes of pitfalls such as Arias-Stella changes, pemphigus, tumor diathesis per se, rare types of cervical cancer, including verrucous and papillary squamous cell cancer, gastric type, and endometrioid adenocarcinoma are also described. KEY MESSAGES: The awareness of pitfalls in cervical cytology is important for cytopathologists and clinicians to avoid future errors. Review of Pap tests with erroneous diagnosis is important for quality control in cytology laboratory, and it must be considered an educational- and experience-building procedure. Cytopathologist should not pull back in significant diagnoses, especially in human papillomavirus-negative cases.

Complete restoration of fertility in a patient treated for androgen-secreting granulosa cell tumor - Case report.

CASE REPORT: A 35-yr-old patient suffering from secondary amenorrhea for two years before she was diagnosed. Secondary amenorrhea occurred after the first normal vaginal delivery, and it was initially associated with breastfeeding and a formerly diagnosed thyroid disease. Transvaginal ultrasound confirmed a tumorous mass of the right ovary. Blood hormone tests detected high serum inhibin B and Anti-Müllerian hormone levels and high androgen level with no signs of virilization. Surgical treatment was indicated for a definitive diagnosis of suspected sex cord-stromal tumor. Right-sided laparoscopic salpingo-oophorectomy was performed, and the histopathological analysis confirmed the diagnosis of granulosa cell tumor adult type. The oncological team recommended adjuvant chemotherapy after the operation, but the patient did not give an informed consent. One month after surgical treatment, spontaneous menstrual bleeding occurred with normalization of sex hormone levels and the menstrual cycle. Nine months after surgical treatment, the patient was examined again due to secondary amenorrhea. Ultrasound confirmed a vital intrauterine pregnancy. The pregnancy course was normal, and the patient had a full-term spontaneous vaginal delivery of her second child. CONCLUSION: Restoration of fertility after a temporary loss due to hormone-secreting granulosa cell tumor is possible after sparing surgical treatment. The role of adjuvant chemotherapy is controversial, particularly in patients with stage I-II disease because of the rarity of this tumor and the absence of prospective randomized studies.

The Role of CD44 and RHAMM in Endometrial (Endometrioid Type) Cancer: An Immunohistochemical Study.

Hyaluronan controls cell migration, differentiation, and proliferation, and it is involved in tumor invasion. The extracellular matrix containing hyaluronan regulates cell behavior via cell surface receptors such as CD44 and receptor for hyaluronan-mediated motility (RHAMM, CD168). We investigated the expression of CD44 and RHAMM in tissue samples of endometrial cancer and the relation of their expression with clinicopathologic parameters of patients. In order to evaluate the value of CD44 and RHAMM as prognostic factors, we investigated the relation of their expression with patients' survival. Our results demonstrated a statistically significant correlation with the depth of myometrial invasion, lymphovascular invasion (LVSI), The International Federation of Gynecology and Obstetrics stage of disease, and, in the case of RHAMM expression, a significant correlation with histologic tumor grade as well. CD44 expression was present in the cell membrane in all cases, but in a proportion of tumors in the cytoplasm as well. In this group of patients, we noticed a significantly greater number of cases with deeper myometrial invasion and LVSI. Finally, we sorted out the group of tumors with simultaneous strong CD44 and strong RHAMM expression, and found a statistically significant correlation with the depth of myometrial invasion and LVSI. Using an univariate analysis, we demonstrated that, in our sample of patients, CD44 expression showed a statistically significant influence on patients' 5-year survival. However, using a multivariate Cox regression analysis, neither CD44 nor RHAMM confirmed themselves as independent prognostic factors.

Adult Granulosa Cell Tumors of the Ovary: A Retrospective Study of 36 FIGO Stage I Cases with Emphasis on Prognostic Pathohistological Features.

OBJECTIVE: Adult granulosa cell tumors (AGCTs) represent 2%-5% of all ovarian malignancies. The aim of this study was to analyze clinical and pathohistological parameters and their impact on recurrence, overall, and disease-free survival in FIGO stage I AGCT patients. METHODS: The tumor specimens analyzed in this retrospective study were obtained from a total of 36 patients with diagnosis of ovarian AGCT surgically treated at the Department of Gynecology, Rijeka University Hospital Centre, between 1994 and 2012. Clinical, pathological, and follow-up data were collected. RESULTS: The mean age at diagnosis was 54.5 years with a range of 24-84. The majority of the patients, 30 (83%), were in FIGO stage IA, 3 (8%) in stage IC1, 1 (3%) in stage IC2, and 2 (6%) in stage IC3. During follow-up period (median 117.5 months, range 26-276), recurrence occurred in 4 patients (12%) with 2 deaths of the disease recorded. In univariate analysis, the 5-year survival rates were significantly shorter in patients with FIGO substage IC (p = 0.019), with positive LVSI (p = 0.022), with presence of necrosis (p = 0.040), and with hemorrhage (p = 0.017). In univariate analysis, the 5-year disease-free survival rates were significantly shorter in patients treated with fertility surgery (p = 0.004), with diffuse growth pattern (p = 0.012), with moderate and severe nuclear atypia (p = 0.032), and with presence of hemorrhage (p = 0.022). FIGO substage IC proved to be independent predictor for recurrence (OR = 16.87, p = 0.015, and OR = 23.49, p = 0.023, resp.) and disease-free survival (p = 0.0002; HR 20.84, p = 0.02) at the uni- and multivariate analyses. CONCLUSIONS: FIGO substage IC is predictive of recurrence and disease-free survival in patients with early-stage AGCTs. LVSI, presence of necrosis and hemorrhage, diffuse growth pattern, and nuclear atypia in AGCTs seem to be associated with overall and disease-free survival, so these pathological features should be taken into consideration when managing patients with AGCT.

Primary Carcinosarcoma of the Vagina Associated With Differentiated Squamous Intraepithelial Neoplasia in a Patient With Complete Uterine Prolapse: Case Report and Review of the Literature.

Vaginal carcinosarcomas (VCSs) are rare and clinically aggressive neoplasms. Primary vaginal malignancies are among the rarest malignant tumors, so clear management guidelines and optimal therapy, especially in the presence of significant pelvic organ prolapse, has not been determined. Here, we present a case of primary VCS closely associated with differentiated squamous intraepithelial neoplasia (DSIN), from which it appeared to have arisen in a postmenopausal patient with complete uterine prolapse. The unusual presentation of our case with DSIN in the adjacent vaginal epithelium with possible diagnostic pitfalls emphasizes the need for systemic presentation of these cases to help pathologists and clinicians know that such lesions can initially present in a patient with complete uterine prolapse. To our knowledge, this is the first case of vaginal DSIN described in the literature to date.

Role of tumor-associated glycoprotein-72 in the progression of endometrial adenocarcinoma: a proposed study.

Endometrial adenocarcinoma is on the basis of the molecular, immunohistological and clinicopathologic features broadly divided into two groups, referred as type I and type II. Type I appears more frequently and in principle patients have a good prognosis; however a significant number of patients develop local recurrences. We hypothesize that TAG-72, expressed on endometrial carcinoma binds and internalizes endocytic pattern recognition receptors on surrounding tissue antigen presenting cells (dendritic cells and macrophages), powers their anti-inflammatory maturation program and make them capable to elicit or modulated tolerogenic immune response mediated by local T and NK effectors. This could support uncontrolled local tumor growth, deeper tumor invasion into surrounding tissues, frequent local recurrences and/or lymph node metastasis. To test this hypothesis, we propose a semi-quantitative immunohistochemical analysis of TAG-72 expression in endometrial adenocarcinoma samples and to correlate the results with clinical and pathological parameters (age, type and histological grade of the tumor, estrogen and progesterone receptor expression, invasion into the myometrium and capillaries, presence of lymph node metastases, FIGO stage, and TNM classification). It would be worthwhile to investigate the local tissue immune response in the tumor environment using tissue samples removed during surgery. These studies could elucidate the underlying immunopathological mechanisms that govern the early recurrence and possibly distant metastases of TAG-72-expressing adenocarcinomas and might help in deciding the type of treatment to be applied in a selected group of cancer patients including application of biological therapy with anti-TAG-72 antibodies, according the principle of personalized oncology treatments.

Heat-Shock Proteins 70 Induce Pro-Inflammatory Maturation Program in Decidual CD1a(+) Dendritic Cells.

PROBLEM: The aim of the study was to assess possible binding of a mixture of constitutive Hsc70 and inducible Hsp70 forms (HSP70) to Toll-like receptor (TLR) 4 and CD91 receptors on decidual CD1a(+) dendritic cells (DCs) and their influence on DCs maturation status. METHOD OF STUDY: Immunohistology and immunofluorescence of paraffin-embedded first trimetester and term pregnancy decidua were performed together with flow cytometry detection of antigens in DCs after stimulation of decidual mononuclear cells with HSP70. RESULTS: Hsc70 and Hsp70 labeling revealed intracellular and nuclear staining in trophoblast cells. The numbers of Hsc70(+) and Hsp70(+) cells of decidual tissue were higher in early pregnancy decidua than in decidua at term. HSP70 binds CD91 and TLR4 receptors on CD1a(+) DCs and increased the expression of CD83, HLA-DR, CD80, and CD86, but decreased CC receptor (CCR) 5. HSP70 increased CC ligand (CCL) 3 and CCL22. HSP70 in the concentration of 1 µg/mL increased the percentage of interferon-γ and interleukin (IL)-15-expressing cells over the cells expressing IL-4. CONCLUSION: HSP70 binds CD91 and TLR4 on decidual CD1a(+) DCs, causes their maturation, and increases IL-15 in the context of Th1 cytokine/chemokine domination, which could support immune response harmful for ongoing pregnancy.

Expression of CD133 and CD117 in 64 Serous Ovarian Cancer Cases.

The cancer stem cells (CSCs) represent a minority of tumor cells that are able to proliferate and self-renew and might be responsible for tumor initiation and maintenance. The CD133 and CD117 are the most commonly used markers for the putative CSCs, especially for the ovarian CSCs, but its clinical significance remains uncertain. The aim of this study was to compare the immunohistochemical expression of CD133 and CD117 in 64 primary ovarian high grade serous carcinoma and peritoneal metastasis, and to examine their potential clinical role. CD133 expression was mainly seen in the apical/endoluminal cell surface of tumor cells and was found in 61% of the carcinoma samples and 41% of the metastasis. The median of CD133 positive cells in tumors was 1 (0.1-7)%, and in metastases was 0.6 (0.1-6)%. CD117 expression appeared as a cytoplasmic and/or membranous stain and was found in 81% of the carcinoma samples and 77% of the metastasis. The median of CD117 positive cells in tumors was 1 (0.1-8)%, and in metastases was 0.1 (0.1-7)%. Multivariate analysis has shown that patients with high CD133 expression in tumor cells have significantly shorter disease free survival and overall survival (p=0.025 and p=0.014, respectively). Patients with high CD117 expression in tumor cells have significantly shorter disease free survival (p=0.031). Cox's proportional hazards model identified expression of CD133 protein in tumor as an independent prognostic factor. Our study indicates that the immunohistochemical assessment of CD133 and CD117 expression may have potential clinical value in predicting disease progression and prognosis in the high grade serous ovarian cancer. CD133 proved to be an independent prognostic factor in the high grade serous ovarian cancer patients.

The significance of heat-shock protein gp96 and its receptors' CD91 and Toll-like receptor 4 expression at the maternal foetal interface.

PROBLEM: Differences in the expression of gp96 and its receptors were analysed in normal and pathological human pregnancy. MATERIAL AND METHODS: Immunohistology and immunofluorescence of sections from decidual part of term placenta, first trimester normal decidua, missed abortion and blighted ovum decidua were performed together with reverse transcriptase-quantitative polymerase chain reaction and flow cytometry. RESULTS: In missed abortion, gp96 was intensively stained, when compared to normal early pregnancy. The intensity of CD91 and TLR4 was higher in the first trimester pregnancy and blighted ovum, when compared to missed abortion. Decidual part of the term placenta is invaded with gp96⁺ , CD91⁺ and TLR4+ trophoblast. Progesterone-induced blocking factor (PIBF) decreased the frequency of TLR4⁺ T lymphocytes, CD91⁺ T, natural killer (NK) and mature dendritic cells after an 18-h culture. Decidual mononuclear cells (DMCs) treated with PIBF down-regulated CD91, TLR4 and gp96 gene expression. CONCLUSION: The presence of gp96, CD91 and TLR4 at the maternal-foetal interface provides a molecular basis for their interaction, particularly in the absence of PIBF.

Frequency and distribution of lymph node metastases in epithelial ovarian cancer.

BACKGROUND: The aim of this retrospective study was to evaluate the incidence and distribution of nodal metastases in relation to the serous versus nonserous histological subtypes of epithelial ovarian cancer. METHODS: Patients were treated primarily with upfront surgery, including pelvic and paraaortic systematic lymphadenectomy, up to the level of the left renal vein, before any kind of chemotherapy administration. Patients were classified according the tumor histology into 2 groups: serous (including the cases of mixed histology with a serous component) and nonserous group. RESULTS: A total of 173 patients fulfilled the inclusion criteria; 76 and 97 patients had serous and nonserous ovarian carcinoma, respectively. Positive lymph nodes were found in 59.3% (45/76) and 14.4% (14/97) of patients in the serous and nonserous histology groups,respectively. There was no difference in positive node distribution in 3 regions (pelvic and para-aortic regions, below and above the inferior mesenteric artery) between these 2 groups. Early spread including 1 or 2 positive lymph nodes was predominantly found in the para-aortic region in both groups, serous and nonserous, whereas distribution of positive nodes in patients with 3 or more lymph nodes shows equal presence in pelvic and para-aortic regions. CONCLUSIONS: Serous ovarian carcinomas are much more prone to metastasize to lymph nodes than nonserous histological types. However, the pattern of lymph node distribution did not differ between these 2 groups and was similar in the pelvic and para-aortic regions.

Treatment and outcome of stage Ia1 squamous cell carcinoma of the uterine cervix.

OBJECTIVE: To evaluate the treatment and clinical outcome of patients with FIGO stage IA1 cervical squamous cell carcinoma (SCC). METHODS: Medical records, including 5-year follow-up, were reviewed for 276 patients with stage IA1 SCC. RESULTS: As definitive surgery, 152 (55.1%), 72 (26.1%), 40 (14.5%), and 12 (4.3%) patients underwent conization, hysterectomy, hysterectomy with pelvic lymphadenectomy (PL), and radical hysterectomy with PL, respectively. Among these groups, the 5-year recurrence-free rate was 94.1%, 98.6%, 95%, and 100%, respectively, and the survival rate was 98.7%, 98.6%, 100%, and 100%, respectively. Recurrent disease was identified among 12 (4.3%) patients, and was related to the depth of invasion (P<0.001). Eleven (4.0%) of 276 patients were found to have lymph vascular space invasion (LVSI). There were no positive lymph nodes among 52 patients who underwent PL, including those with LVSI. Conization was followed by hysterectomy in 49 patients. In these patients, residual intraepithelial neoplasia was found in 18 women, 3 of whom had no involved margins on previous conization. In the group of patients treated by conization, recurrence correlated with the status of the endocervical and lateral cone margin (P<0.001). CONCLUSION: As a conservative approach, conization is an effective and reasonable treatment option for stage IA1 SCC, especially in actively reproductive women.

Cervical cytology and HPV test in follow-up after conisation or LLETZ.

The patients treated with conservative surgical therapy for cervical intraepithelial neoplasia (CIN) have an increased risk to develop invasive cervical carcinoma compared to the general population. Cervical cytology and HPV test are included in the protocols for the detection of treatment failure. The purpose of the study was to analyse cytology-histology correlation after conisation or Large Loop Excision of the Transformation Zone (LLETZ), resection margin status, compliance to the follow-up protocol and evaluation of cervical cytology and HPV testing in two year period after surgical treatment. We retrospectively reviewed 251 cases of conisation or LLETZ performed between January and December, 2006. Conventional cervical smears were analysed and abnormal cytology was defined as atypical squamous cells of undetermined significance or worse (ASCUS+). The digene Hybrid capture 2 test was used for detection of high-risk HPV types. Histology analysis demonstrated CIN1 + lesion in 234 cases (93.2%) with cytology-histology correlation in 97.9% of cases. A preoperative HPV test was made in 142 histologically confirmed CIN1 + lesions and 137 (96.5%) tested positive. The resection margins were involved in 48 (20.8%) cases. In 24 (10.3%) cases the margins were difficult to determine. Abnormal cytology was found in 33 (15.2%) cases of the 217 (86.5%) patients that attended the post-treatment visits. The post-treatment HPV test was performed on 159 women and it was positive in 25 (15.7%) cases. The complete follow-up control cytology, with at least three Pap smears in the subsequent two years or with second treatment, was registered in only 146 (58.2%) patients. 14/217 (6.5%) patients underwent second treatment with histologically confirmed treatment failure. In all patients with control smear, repeated cytology found HSIL. On six women, the control HPV test was performed. In five cases, it was positive and in one case with histological diagnoses of VAIN2, it was negative. Our study confirms the important role of cervical cytology in the diagnosis of cervical intraepithelial lesions and monitoring after treatment. In the future we will have to improve compliance to the follow-up protocols and use of the HPV test in the selection of women at risk of treatment failure.