Heitor Ricardo Cosiski Marana

Enrichment of regulatory T cells in invasive breast tumor correlates with the upregulation of IL‐17A expression and invasiveness of the tumor

Breast cancer is a leading cause of neoplasia‐associated death in women worldwide. Regulatory T (Treg) and Th17 cells are enriched within some tumors, but the role these cells play in invasive ductal carcinoma (IDC) of the breast is unknown. We show that CD25+CD4+ T cells from PBMCs and tumor express high levels of Foxp3, GITR, CTLA‐4, and CD103, indicating that tumor‐infiltrating Treg cells are functional and possibly recruited by CCL22. Additionally, we observed upregulation of Th17‐related molecules (IL‐17A, RORC, and CCR6) and IL‐17A produced by tumor‐infiltrating CD4+ and CD8+ T lymphocytes. The angiogenic factors CXCL8, MMP‐2, MMP‐9, and vascular endothelial growth factor detected within the tumor are possibly induced by IL‐17 and indicative of poor disease prognosis. Treg and Th17 cells were synchronically increased in IDC patients, with positive correlation between Foxp3, IL‐17A, and RORC expression, and associated with tumor aggressiveness. Therefore, Treg and Th17 cells can affect disease progression by Treg‐cell‐mediated suppression of the effector T‐cell response, as indicated by a decrease in the proliferation of T cells isolated from PBMCs of IDC patients and induction of angiogenic factors by IL‐17‐producing Th17. The understanding of regulation of the Treg/Th17 axis may result in novel perspectives for the control of invasive tumors.

A morphometric study of maternal smoking on apoptosis in the syncytiotrophoblast

Objectives: To study syncytiotrophoblast apoptosis in the placenta of smoking and non‐smoking pregnant women. Methods: Twelve neonates, pregnancies and placentas were available for study. Eight mothers smoked during pregnancy and the remaining four were non‐smokers used as control subjects. The main outcome measure was the apoptotic syncytiotrophoblast index for each group. Apoptosis was detected by immunohistochemistry using the TUNEL method and quantitatively measured using a Merz grid. The apoptotic syncytiotrophoblast index was calculated as the ratio of mean apoptotic labeling to percent terminal villus area using high‐power field microscopy. Results: Significant differences in apoptotic syncytiotrophoblast index were observed between the control group (15.06 ± 3.72) and the smoker group (1.66 ± 1.74) (P < 0.0001, Mann‐Whitney test), but no differences were detected in clinical or morphometric data between groups. Conclusions: The human placental syncytiotrophoblast undergoes apoptosis and this process is associated with inhibition of apoptosis by the smoking habit. The same way as the presence of trophoblast apoptosis is associated with modifications of the maternal‐fetal exchange, the inhibitory effect of the smoking habit on syncytiotrophoblast could be responsible for the poor prognosis of pregnancy in the presence of maternal smoking.

Tumor-infiltrating CD4+ T lymphocytes in early breast cancer reflect lymph node involvement

BACKGROUND The role of immune system in the pathogenesis and progression of breast cancer is a subject of controversy, and this stimulated us to investigate the association of the immunophenotype of tumor-infiltrating lymphocytes in early breast cancer with the spread of tumor cells to axillary lymph nodes. METHODS Tumor samples from 23 patients with early breast cancer from the Department of Gynecology and Obstetrics of Ribeirão Preto Medical School (USP) were obtained at the time of biopsy and submitted to an enzyme-digestion procedure for the extraction of tumor-infiltrating lymphocytes. The lymphocytes extracted were analyzed by dual-color flow cytometry with monoclonal antibodies in these combinations: CD3 FITC/CD19 PE, CD3 FITC/CD4 PE, CD3 FITC/CD8 PE, and CD16/56 PerCP, which are specific for immunophenotyping of T and B lymphocytes, helper and cytotoxic T lymphocytes, and natural killer (NK) cells. The mean percentage of these cells was used for comparing groups of patients with or without lymph node metastasis. RESULTS The mean value for T-lymphocyte infiltration was 24.72 +/- 17.37%; for B-lymphocyte infiltration, 4.22 +/- 6.27%; for NK-cell infiltration, 4.41 +/- 5.22%, and for CD4(+) and CD8(+) T-lymphocyte infiltration, 12.43 +/- 10.12% and 11.30 +/- 15.09%, respectively. Only mean values of T- and CD4(+) T-lymphocyte infiltration were higher in the group of patients with lymph node metastasis, while no differences were noted in the other lymphocyte subpopulations. CONCLUSION The association of tumor-infiltrating CD4(+) T lymphocytes with lymph node metastasis suggests a role for these cells in the spread of neoplasia to lymph nodes in patients with early breast cancer.

Apoptosis induced by neoadjuvant chemotherapy in breast cancer

Aim: To evaluate the relationship between apoptosis induced by chemotherapy and clinical response in breast cancer. Methods: Apoptosis index (AI), mutant p53 and Bcl‐2 protein expression were evaluated in 44 breast tumour samples from patients submitted to neoadjuvant chemotherapy. Objective response (OR) to primary chemotherapy was observed in 37 patients (84%) and no response (NR) in seven. AI was measured by the rate of apoptotic cells identified using morphological criteria. p53 and Bcl‐2 protein expression were evaluated using an immunoperoxidase staining technique. Results: The median AI change observed between pre‐chemotherapy AI and post‐chemotherapy AI was 0.84 in the OR group and 0.01 in the NR group, (rho = 0.4; p = 0.006). There was no change in Bcl‐2 protein expression following chemotherapy. In the OR group, p53 protein expression was positive in 41.6% of patients before and in 22.2% after chemotherapy (difference = 16.6%; p = 0.03). No change was detected in the NR group. Conclusion: A positive correlation was found between the increase in AI and clinical response to neoadjuvant chemotherapy in locally advanced breast cancer.

Expression of Hypoxia-inducible factor 1-α and Vascular endothelial growth factor-C in locally advanced breast cancer patients

BACKGROUND: Locally advanced breast cancers are more prevalent in underdeveloped countries. Targeted therapy has been improved to identify hallmarks that are specific to these subtypes of tumors. OBJECTIVES: We aimed to prospectively assess the expression of Hypoxia inducible factor-1 α and vascular endothelial growth factor-C in locally advanced breast cancer patients. METHODS: Thirty women underwent incisional biopsies for the histopathological diagnosis of breast carcinoma and participated in neoadjuvant chemotherapy. The association of Hypoxia inducible factor-1 α and vascular endothelial growth factor-C with age, tumor size, histological grade, clinical staging, hormonal and axillary status, clinical and pathological response after neoadjuvant chemotherapy, expression of estrogen and progesterone receptors, and the presence of c-erbB-2 antigen was studied. RESULTS: Hypoxia inducible factor-1 α expression and Vascular endothelial growth factor-C expression were observed in 66.7% and 63.3% of all patients, respectively, and were marginally associated with each other (p = 0.06). Among the studied variables, only positive axillary status was associated with the presence of HIF-1α (p = 0.02). Complete pathological response was significantly associated (p = 0.04) with the expression of vascular endothelial growth factor-C prior to neoadjuvant chemotherapy. CONCLUSION: We concluded that Hypoxia inducible factor-1 α was associated with a poor prognosis and that vascular endothelial growth factor-C could be used as a predictive factor in locally advanced breast cancer patients with complete pathological response after neoadjuvant chemotherapy.

Genital Manifestation of Graft-vs.-Host Disease: A Series of Case Reports

INTRODUCTION After hematopoietic stem cell transplantation (HSCT), many patients present genital graft-vs.-host disease (GVHD) that can culminate with sexual problems, which are poorly dimensioned. AIM We hope to draw attention to the need to perform genital biopsy to diagnose genital GVHD, and thus to call attention to the need to incorporate careful attention to sexual health in the treatment of these patients. METHODS Five allogeneic stem cell transplant recipients complaining of coital pain after HSCT were clinically diagnosed for genital GVHD. Genital biopsies were given for histological analysis, and microphotographs of the corresponding marked field in the slide were taken. Specimens were evaluated by the site pathologist and then sent to a reference pathologist, each blinded to the histological findings. A literature search was performed in PubMed/MEDLINE (1966-2009) for cross-sectional and cohort studies or trials related to genital GVHD. Expert opinions peer reviews and case reports were also considered. MAIN OUTCOME MEASURES HSCT, genital GVHD, genital biopsy. RESULTS The biopsy showed evidence of dilated apoptotic cells in the basal layer and detachment of the epithelial lining of the mucosa, hyalinization and thickening of collagen fibers, capillary ectasia, and mononuclear inflammatory infiltrate of the submucosa. Three patients presented vulval lesion such as leucoplasia and ulcer on the large lip. Histological analyses showed evidence of epithelial hyperplasia and influx of inflammatory cells to the epithelial surface, intercellular edema and spongiosis, apoptotic bodies on the basal layer of the epithelium, spongiosis, and nuclear vacuolization. A common treatment based on corticotherapy resulted in complete remission of coetaneous or mucous genital lesions in all five patients. CONCLUSION Genital biopsy is important to differentially diagnose GVHD and secondary symptoms due to hypoestrogenism. Prevention is the most important step in controlling the evolution GVHD in the vagina to prevent vaginal obstruction and sexual dysfunction.

Fibromatosis of the male breast: a case report with immunohistochemistry study and review of the literature

mitted to the Outpatient Breast Oncology Service, Department of Health of Sao Carlos, Brazil, with a 3-month history of a painless subcutaneous mass with rapid growth. Clinical examination revealed an ill-defined mass located in the upper inner quadrant of his left breast tissue measuring of 3.5 cm in diameter. The mass was firm and was fixed to the pectoral fascia and to skin. There was no axillary or cervical lymphadenopathy or nipple discharge. At ultrasound, the lesion manifested as an irregular, hypoechoic mass with posterior acoustic shadowing, suggestive of malignancy. Fine-needle aspiration cytology was performed, but the result was inconclusive because the aspirate did not yield sufficient epithelial cells for diagnosis. The tumor was removed by en bloc resection with wide excision, the greater pectoral muscle being spared. During surgery the lesion was found to be firmly adherent to the underlying pectoralis major. The appearance was that of an infiltrative tumor, and the diagnosis of epithelial neoplasm was considered in the differential diagnosis. There was no history of trauma to the chest wall, so that a fibrous reaction to trauma was not considered in the differential diagnosis. The surgical specimen was fixed in 4% formalin. Se

Expression of aldehyde dehydrogenase after neoadjuvant chemotherapy is associated with expression of hypoxia-inducible factors 1 and 2 alpha and predicts prognosis in locally advanced breast cancer

OBJECTIVE: To analyze the expression of hypoxia-inducible factors (hypoxia-inducible factor 1A and hypoxia-inducible factor 2A) and aldehyde dehydrogenase proteins in patients with locally advanced breast carcinoma who were subjected to neoadjuvant chemotherapy. METHODS: We included 90 patients with histologically confirmed stage II and III breast carcinoma who were treated with neoadjuvant chemotherapy between 2000 and 2005. Immunohistochemistry for aldehyde dehydrogenase, hypoxia-inducible factor 1A, and hypoxia-inducible factor 2A was performed before and after neoadjuvant chemotherapy. We analyzed the influence of clinical and pathological features on clinical and pathological response, disease-free survival, and overall survival. RESULTS: An objective clinical response to neoadjuvant chemotherapy was observed in 80% of patients, with 12% showing a complete pathological response. Among all clinical and pathological parameters, only the expression of hypoxia-inducible factor 1A was associated with a pathological response. A positive association was found between expression of aldehyde dehydrogenase and that of hypoxia-inducible factor 1A before and after chemotherapy. Aldehyde dehydrogenase expression was associated with expression of hypoxia inducible-factor 2A in tumors after neoadjuvant treatment. In a univariate analysis, prognosis was influenced by age, pathological response, metastasis to axillary lymph nodes after neoadjuvant chemotherapy, overexpression of hypoxia-inducible factor 2, and the presence of aldehyde dehydrogenase-positive cells within the primary tumor after neoadjuvant chemotherapy. In a multivariate analysis, only age and the presence of aldehyde dehydrogenase-positive cells after chemotherapy were associated with reduced overall survival. CONCLUSION: The presence of aldehyde dehydrogenase-positive cells within the residual tumor after neoadjuvant chemotherapy is associated with an increase in the expression of hypoxia-inducible factor 2A and with poor prognosis in patients with locally advanced breast cancer.

Impact of surgical staging in locally advanced cervical cancer and subsequent chemotherapy.

Surgical staging (SS) is the gold standard for determination of the true extent of a patients disease and is an important prognostic factor in cervical cancer. We investigated whether lymph node dissection (LND) prior to chemotherapy (CT) followed by radical surgery (RS) could modified overall (OS) and disease‐free survival (DFS).

Fatores de risco para recidiva após tratamento de lesöes provocadas pelo HPV no trato genital feminino

Purpose: evaluation of the risk factors [lesion grade, seropositivity for type 1 acquired immunodeficiency virus (HIV-1) and association with pregnancy ] for relapse of human papillomavirus (HPV) induced lesions of the female genital tract. Patients and Methods: seventy patients with a clinical, colposcopic and cytologic diagnosis of HPV infection were studied. Clinical follow-up lasted at least 6 months after the initial treatment, thus permitting the evaluation of the therapeutic results. Twenty-seven of these patients were pregnant and 12 were seropositive for HIV-1. The remaining 44 patients were not in the pregnancy-puerperium cycle and 14 of them were HIV-1 positive. According to cytologic criteria, the cervical lesions were classified as changes associated with HPV or grade I cervical intraepithelial neoplasia (CIN I) (low grade lesions) or CIN II/III (high grade lesions). Data were analyzed statistically by the exact Fisher test, with the level of significance set at p<0.05. The therapeutic scheme for lesions limited to the uterine cervix was cryo- or electrocautery (EC), whereas topical 5-fluorouracil was used for the diffused lesions through the vaginal wall. For the lesions in the vulvoperineal region, 80% trichloroacetic acid was used, and when they were voluminous, EC was applied. Among the pregnant women, a cryocautery was used for lesions limited to the cervix and EC for diffuse lesions. Results: among the HIV-1-negative pregnant women there was an 87.5% rate of recurrence when the lesions were in the cervix-vagina, and no recurrence when the lesions were vulvoperineal. In contrast, seropositive pregnant women presented 100% recurrence regardless of the site of the lesion. Among nonpregnant HIV negative women, 20 and 24% recurrence was observed in the cervix-vagina and in the vulvoperineal region, respectively, as opposed to 87.5 and 100% recurrence, respectively, for the same regions among HIV positive women. The lesions associated with CIN showed a higher frequency of recurrence with increasing CIN grade and a synergistic effect with the association of HIV-1 and pregnancy. Conclusions: the recurrence rate for women treated for HPV-induced lesions is high and the association with pregnancy, HIV and increased grade of the intraepithelial lesions are synergistic factors in the determination of therapeutic failure. The site of implantation of HPV-induced lesions is of prognostic significance only when the infection is not associated with HIV.