Altered expression of the costimulatory molecules CD80, CD86, CD152, PD-1 and ICOS on T-cells from paracoccidioidomycosis patients: Lack of correlation with T-cell hyporesponsiveness

T-cell proliferative hypo responsiveness, a hallmark of paracoccidioidomycosis immune responses, underlies host`s failure in controlling fungus spread, being reversible with antifungal treatment. The mechanisms leading to this hypoproliferation are not well known. Since costimulatory molecules have been shown to profoundly regulate T-cell immune responses, we investigated the hypothesis that the determinants of the responder versus tolerant state may be the regulated expression of, or signaling by, costimulatory molecules. Expression of CD80, CD86, CD28, CD152, ICOS and PD-1 costimulatory molecules were examined on T-cells and monocytes harvested from stimulated and unstimulated PBMC cultures of active paracoccidioidomycosis patients and healthy individuals cured of past paracoccidioidomycosis. Stimuli were gp43, the immunodominant component of Paracoccidioides brasiliensis, and a Candida antigen. While CD28 expression, critical for optimal T-cell activation, was comparable between patients and controls, CD152, PD-1 and ICOS, which preferentially deliver negative signaling, were overexpressed on patients` stimulated and unstimutated T-cells. PBMC cultures were carried out in presence of the respective blocking antibodies which, however, failed to restore T-cell proliferation. CD80 and CD86 were equally expressed on patients` and controls` monocytes, but overexpressed on patients` T-cells. Blockade with the respective blocking antibodies on day 4 of the culture also did not restore T-cell proliferation, while, on day 0, differentially inhibited Candida and gp43 responses, suggesting that different antigens require different costimulatory pathways for antigen presentation. Our data favors the hypothesis, raised from other foreign antigen models, that prolonged in vivo antigen exposure leads to an adaptive tolerance T-cell state which is hardly reverted in vitro. (C) 2008 Elsevier Inc. All rights reserved.

Better CD4+T Cell Recovery in Brazilian HIV-Infected Individuals Under HAART Due to Cumulative Carriage of SDF-1-3`A, CCR2-V64I, CCR5-D32 and CCR5-Promoter 59029A/G Polymorphisms

Polymorphisms of chemokines and chemokine-receptors genes have been shown to influence the rate of progression to AIDS; however, their influence on response to HAART remains unclear. We investigated the frequency of the SDF-1-3`A, CCR2-64I, CCR5-D32 and CCR5-Promoter-59029-A/G polymorphisms in Brazilian HIV-1-infected and uninfected individuals and their influence on CD4+ T-cell evolution HIV-1 infected individuals before and during HAART. Polymorphism detection was done in a transversal study of 200 HIV-1-infected and 82 uninfected individuals. The rate of CD4+ T cell increase or decrease was studied in a cohort of 155 HIV-1 infected individuals on pre and post-HAART. Polymorphisms were determined by PCR associated with RFLP. The rate of CD4+ T-cell decline or increase was also determined. HIV-1 infected and uninfected subjects showed, respectively, frequencies of 0.193 and 0.220 for SDF-1-3`A, of 0.140 and 0.110 for CCR2-V64I, of 0.038 and 0.055 for CCR5-D32, and of 0.442 and 0.390 for CCR5-P-59029-A/G. HIV-1-infected subjects carrying one, two or three of these four polymorphisms showed better CD4+ T-cell recovery than HIV-1-infected subjects carrying the four wild-type alleles (+2.7, +1.6, +3.5, and -0.9 lymphocytes/mu l/month, respectively). Regression logistic analysis showed that the CCR5-D32/CCR2-V64I association was predictor of positive CD4+ T cell slope after HAART. The distribution of polymorphisms did not differ between HIV-1-infected and uninfected individuals, but differed from more homogenous ethnic groups probably reflecting the miscegenation of the Brazilian population. We add further evidence of the role of these polymorphisms by showing that the CD4 gain was influenced by carriage of one or more of the polymorphisms studied here. These results highlight the possibility that these genetic traits can be useful to identify patients at risk for faster progression to AIDS or therapeutic failure.

PTCH1 gene mutations in exon 17 and loss of heterozygosity on D9S180 microsatellite in sporadic and inherited human basal cell carcinomas

Background Basal cell carcinomas (BCCs) are the most frequent human cancer that results from malignant transformation of basal cells in the epidermis. Gorlin syndrome is a rare inherited autosomal dominant disease that predisposes with multiple BCCs and other birth defects. Both sporadic and inherited BCCs are associated with mutations in the tumor suppressor gene PTCH1, but there is still uncertainty on the role of its homolog PTCH2. Objectives To search for mutations and genomic instability in sporadic and inherited BCCs. Methods DNA obtained from leukocytes and tumor cells was amplified by polymerase chain reaction regarding five exons of PTCH1 and PTCH2 and neighboring microsatellites. Exons were sequenced and compared with the GenBank database. Results Only D9S180, of six microsatellites, showed loss of heterozygosity in three BCCs (two sporadic and one inherited). One sporadic BCC presented the mutation g. 2885G>C in exon 17 of PTCH1, which predicts the substitution p.R962T in an external domain of the protein. In addition, the leukocytes and tumor cells of one patient with Gorlin syndrome showed the mutation g. 2839T>G in the same exon and gene, which predicts a p.E947stop and truncated protein. All control and tumor samples presented IVS9 + 217T in intron 9 of PTCH1. Conclusion Mutations found in the PTCH1 gene and neighboring repetitive sequences may have contributed to the development of the studied BCCs.

Primary Cutaneous Blastoid Mantle Cell Lymphoma-Case Report

Mantle cell lymphoma (MCL) commonly involves extranodal sites, usually as a manifestation of disseminated disease. In rare cases, MCLs may arise as a primary tumor in the skin. Blastoid mantle cell lymphoma (BV-MCL) is a rare variant and has a more aggressive clinical course. The phenotype of BV-MCL is characterized as CD20(+), CD5(+), cyclin D1(+), CD23(-), and CD10(-). Interphase fluorescence in situ hybridization shows a characteristic t(11; 14) fusion pattern. We report a case of a BV-MCL arising in skin as primary cutaneous MCL with the characteristic immunophenotype and translocation.

Predictors of higher number of stages in Mohs micrographic surgery for the treatment of squamous cell carcinoma of the head

BACKGROUND - Squamous cell carcinomas of the skin of the bead are better treated with Mobs micrographic surgery which has the lowest recurrence rates and allows spare normal tissue. There are some characteristics of squamous cell carcinoma that can be related to a higher number of surgical stages. OBJECTIVE - To study characteristic of head squamous cell carcinoma that predicts a higher number of Mohs surgical stages. METHODS - A retrospective analysis of 51 squamous cell carcinomas of the bead treated with Mobs surgery was performed to determine risk factors for a higher number of surgical stages. The characteristics analyzed were clinical limits, morphology, recurrence, histological differentiation and size and compared to the number of surgical stages. The analysis was performed by Fisher`s exact test and multivariate logistic regression. RESULTS - The recurrent squamous cell carcinomas showed a tendency for a higher number of stages (p=0,081). The Odds Ratio for a higher number of Mobs stages was three for inaccurate limits; although not statistically significant, it corroborates clinical and previous publication. CONCLUSION - Clinical characteristics of squamous cell carcinoma as recurrence and inaccurate limits would not predict, but could indicate tendency of a higher number of Mobs micrographic surgery stages.

SOX2 is an amplified lineage-survival oncogene in lung and esophageal squamous cell carcinomas

Lineage-survival oncogenes are activated by somatic DNA alterations in cancers arising from the cell lineages in which these genes play a role in normal development(1,2). Here we show that a peak of genomic amplification on chromosome 3q26.33 found in squamous cell carcinomas (SCCs) of the lung and esophagus contains the transcription factor gene SOX2, which is mutated in hereditary human esophageal malformations(3), is necessary for normal esophageal squamous development(4), promotes differentiation and proliferation of basal tracheal cells(5) and cooperates in induction of pluripotent stem cells(6-8). SOX2 expression is required for proliferation and anchorage-independent growth of lung and esophageal cell lines, as shown by RNA interference experiments. Furthermore, ectopic expression of SOX2 here cooperated with FOXE1 or FGFR2 to transform immortalized tracheobronchial epithelial cells. SOX2-driven tumors show expression of markers of both squamous differentiation and pluripotency. These characteristics identify SOX2 as a lineage-survival oncogene in lung and esophageal SCC.

Acinar cell carcinoma of the pancreas: is the absence of neuroendocrine component related to a more malignant behavior?

Background/Aims: Acinar cell carcinomas are uncommon malignant tumors of the pancreas, accounting for 1-2% of all the cases of exocrine pancreatic tumor. Some authors have estimated acinar cell tumors to be as aggressive as ductal adenocarcinoma of the pancreas whereas other series showed acinar cell tumors to have a favorable clinical outcome. This discrepancy in prognosis may be related to the cellular components of the tumor. Methodology: With the aim to evaluate the possible relationship between the presence of neuroendocrine differentiation and behavior of these tumors, the authors reviewed all patients presenting acinar cell carcinoma of the pancreas in the last 5 years with emphasis in the immunohistochemical evaluation. Results: Four patients presented neuroendocrine differentiation on immunohistochemical evaluation and had a more benign outcome. Two patients without neuroendocrine component had a disseminated disease at presentation. This data suggests that this tumor is less aggressive than ductal adenocarcinoma and even with nodal involvement, long term survival after complete resection can be achieved. Conclusions: It is possible that the absence of neuroendocrine component may be related to a less favorable outcome and adjuvant therapy may be necessary. Due to the rarity of this pancreatic tumor, this relationship remains to be confirmed with a multicentric study including a larger number of patients.

Molecular evidence of human T-cell lymphotropic virus types 1 and 2 (HTLV-1 and HTLV-2) infections in HTLV seroindeterminate individuals from Sao Paulo, Brazil

Background: Using enzyme immunoassays and Western blot (Wb) tests, HTLV serodiagnosis yields indeterminate results in a significant number of cases. Objective: To determine the prevalence of HTLV infection among HTLV-seroindeterminate individuals. Study design: We studied peripheral blood mononuclear cells from 65 anti-HTLV Wb-seroindeterminate individuals by attempting to amplify proviral DNA sequences (tax and pot) to identify HTLV-I and HTLV-2 infections. Results: These 65 specimens exhibited predominantly (43%) anti-HTLV antibodies to gag-coded antigens in the absence of anti-p24 on Wb analysis. Tax proviral sequences were detected in 6 (9.2%) samples. According to restricted fragment polymorphism analysis (RFLP), we identified HTLV-1 proviral DNA in 4 samples. HTLV-2 in one and sequences from both in another. Nested PCR for the pot region was positive in 3 (4.6%) specimens, which were also positive for tax sequences. After hybridization HTLV-1 infection was confirmed in 2 samples (3.1%) and HTLV-2 in another (1.5%). Detection of a single HTLV DNA sequence may be due to infection by defective provirus, but its significance remains undefined. In this cohort, no Wb reactivity pattern was predictive of proviral detection. HTLV-I infection was demonstrated in an individual who had Wb reactivity to gag-coded antigens only. Conclusions: This emphasizes the importance of clinical and laboratory follow-up of HTLV-seroindeterminate individuals from endemic areas. (c) 2009 Elsevier B.V. All rights reserved.

Establishment and characterization of androgen-independent human prostate cancer cell lines, PcBra1, PcBra2, and PcBra3

One of the main obstacles for understanding biological events involved in cancer is the lack of experimental models for in vitro studies especially for prostate cancer (PC).There are a limited number of PC cell lines being the majority originated from metastatic tumors mostly acquired from American Tissue Cell Culture which demands importation an expensive and bureaucratic process. Also it is well known that there are ethnic differences between populations concerning the behavior of tumors and the research based on cell lines derived from Brazilians should be interesting. Our aim was to develop tumor cell lines from primary PC.

CYP1A2*1C, CYP2E1*5B, and GSTM1 polymorphisms are predictors of risk and poor outcome in head and neck squamous cell carcinoma patients

Head and neck squamous cell carcinoma (HNSCC) is associated with environmental factors, especially tobacco and alcohol consumption. Most of the carcinogens present in tobacco smoke are converted into DNA-reactive metabolites by cytochrome P450 (CYPs) enzymes and detoxification of these substances is performed by glutathione S-transferases (GSTs). It has been suggested that genetic alterations, such as polymorphisms, play an important role in tumorigenesis and HNSCC progression. The aim of this study was to investigate CYP1A1, CYP1A2, CYP2E1, GSTM1, and GSTT1 polymorphisms as risk factors in HNSCC and their association with clinicopathologic data. The patients comprised 153 individuals with HNSCC (cases) and 145 with no current or previous diagnosis of cancer (controls). Genotyping of the single nucleotide polymorphisms (SNPs) of the CYP1A1, CYP1A2, and CYP2E1 genes was performed by PCR-RFLP and the GSTM1 and GSTT1 copy number polymorphisms (CNPs) were analyzed by PCR-multiplex. As expected, a significant difference was detected for tobacco and alcohol consumption between cases and controls (P < 0.001). It was observed that the CYP1A2*1D (OR = 16.24) variant and GSTM1 null alleles (OR = 0.02) confer increased risk of HNSCC development (P < 0.001). In addition, head and neck cancer alcohol consumers were more frequently associated with the CYP2E1*5B variant allele than control alcohol users (P < 0.0001, OR = 190.6). The CYP1A2*1C polymorphism was associated with tumor recurrence (log-rank test, P = 0.0161). The CYP2E1*5B and GSTM1 null alleles were significantly associated with advanced clinical stages (T3 + T4; P = 0.022 and P = 0.028, respectively). Overall, the findings suggested that the genetic polymorphisms studied are predictors of risk and are also associated with tumor recurrence, since they are important for determining the parameters associated with tumor progression and poor outcomes in HNSCC. (C) 2009 Elsevier Ltd. All rights reserved.

Bioinformatics construction of the human cell surfaceome

Cell surface proteins are excellent targets for diagnostic and therapeutic interventions. By using bioinformatics tools, we generated a catalog of 3,702 transmembrane proteins located at the surface of human cells (human cell surfaceome). We explored the genetic diversity of the human cell surfaceome at different levels, including the distribution of polymorphisms, conservation among eukaryotic species, and patterns of gene expression. By integrating expression information from a variety of sources, we were able to identify surfaceome genes with a restricted expression in normal tissues and/or differential expression in tumors, important characteristics for putative tumor targets. A high-throughput and efficient quantitative real-time PCR approach was used to validate 593 surfaceome genes selected on the basis of their expression pattern in normal and tumor samples. A number of candidates were identified as potential diagnostic and therapeutic targets for colorectal tumors and glioblastoma. Several candidate genes were also identified as coding for cell surface cancer/testis antigens. The human cell surfaceome will serve as a reference for further studies aimed at characterizing tumor targets at the surface of human cells.

Glial cell line-derived neurotrophic factor added to a sciatic nerve fragment grafted in a spinal cord gap ameliorates motor impairments in rats and increases local axonal growth

Purpose: The aversive nature of regenerative milieu is the main problem related to the failure of neuronal restoration in the injured spinal cord which however might be addressed with an adequate repair intervention. We evaluated whether glial cell line-derived neurotrophic factor (GDNF) may increase the ability of sciatic nerve graft, placed in a gap promoted by complete transections of the spinal cord, to enhance motor recovery and local fiber growth. Methods: Rats received a 4 mm-long gap at low thoracic level and were repaired with a fragment of the sciatic nerve. GDNF was added (NERVE+GDNF) or not to the grafts (NERVE-GDNF). Motor behavior score (BBB) and sensorimotor tests-linked to the combined behavior score (CBS), which indicate the degree of the motor improvement and the percentage of functional deficit, respectively, and also the spontaneous motor behavior in an open field by means of an infrared motion sensor activity monitor were analyzed. At the end of the third month post surgery, the tissue composed by the graft and the adjacent regions of the spinal cord was removed and submitted to the immunohistochemistry of the neurofilament-200 (NF-200), growth associated protein-43 (GAP-43), microtubule associated protein-2 (MAP-2), 5-hidroxytryptamine (serotonin, 5-HT) and calcitonin gene related peptide (CGRP). The immunoreactive fibers were quantified at the epicenter of the graft by means of stereological procedures. Results: Higher BBB and lower CBS levels (p < 0.001) were found in NERVE+GDNF rats. GDNF added to the graft increased the levels of individual sensorimotor tests mainly at the third month. Analysis of the spontaneous motor behavior showed decreases in the time and number of small movement events by the third month without changes in time and number of large movement events in the NERVE+GDNF rats. Immunoreactive fibers were encountered inside the grafts and higher amounts of NF-200, GAP-43 and MAP-2 fibers were found in the epicenter of the graft when GDNF was added. A small amount of descending 5-HT fibers was seen reentering in the adjacent caudal levels of the spinal cords which were grafted in the presence of GDNF, event that has not occurred without the neurotrophic factor. GDNF in the graft also led to a large amount of MAP-2 perikarya and fibers in the caudal levels of the cord gray matter, as determined by the microdensitometric image analysis. Conclusions: GDNF added to the nerve graft favored the motor recovery, local neuronal fiber growth and neuroplasticity in the adjacent spinal cord.

Evaluation of inverted papilloma and squamous cell carcinoma by nasal contact endoscopy

Background: Contact endoscopy (CE) was initially described as a method used in the analysis of uterine and vocal folds histology. The first nasal cavity CE studies achieved promising results regarding its use for the differentiation between benign and malignant lesions, considering that biopsy might cause some complications, especially bleeding. This study described and compared the findings of CE on inverted papilloma and nasosinusal squamous cell carcinoma (SCC) and tested the effectiveness of this exam as a noninvasive method for in vivo differentiation between these tumors. Methods: The patients included in this study were divided into group A, patients diagnosed with inverted papilloma, and group B, patients diagnosed with SCC. CE results were compared among themselves. CE images were presented to examiners not experienced with the method. Results: Twenty-two patients were examined, 13 in group A and 9 in group B. The main relevant differences in CE findings between those two groups were corkscrew vessels, presence of mitoses, keratinization and nuclear pleomorphism in carcinoma, and vacuolated cells in papilloma. The examiners were capable of defining the diagnosis of these nasal tumors only based on CE images. Conclusion: CE may be a useful noninvasive exam to be used in the in vivo diagnosis of inverted papilloma and nasosinusal SCC, which may enable better preoperative planning. (Am J Rhinol Allergy 24, 210-214, 2010; doi: 10.2500/ajra.2010.24.3467)

Imiquimod 5% Cream for the Treatment of Periocular Basal Cell Carcinoma

Purpose: The objective of this pilot study was to evaluate the efficacy and safety of 5% imiquimod cream in the treatment of periocular basal cell carcinoma (BCC) through the analysis of a case series. Methods: Eight subjects with primary nodular BCC of the eyelid were recruited. Treatment lasted 10 to 16 weeks. The average follow-up time was 11.7 months. Results: Of a total of 10 lesions, 80% resolved clinically and histologically and have remained asymptomatic since. Conclusion: Imiquimod cream 5% was shown to be an attractive alternative to surgical treatment of periocular BCC. Future studies with larger samples and longer follow-up periods are expected to provide more accurate information on the efficacy and safety of the drug.

Claudin and p53 expression in vulvar lichen sclerosus and squamous-cell carcinoma

Aims Vulvar squamous-cell carcinoma (SCC) is a rare gynaecological cancer. Vulvar SCC has been shown to develop from vulvar intraepithelial neoplasias, which are related to lichen sclerosus (LS). Most studies to date have compared vulvar SCC with LS only morphologically, but no detailed molecular analysis has been performed. The objective was to compare claudin and p53 expression in these diseases and determine if there was any association with expression and vulvar SCC progression. Methods Immunohistochemical analysis was performed in order to determine expression of p53 and claudin 1, 2, 3, 4, 5, 7 and 11 in human vulvar tissue samples from LS, SCC and control patients. Results Claudin 1, 2, 3, 4 and 5 were expressed comparably in the three groups. Claudin 7 and 11 expression was significantly decreased in LS and SCC samples compared with the control group. Expression of p53 was significantly increased in SCC and LS patient samples compared with the control group. Conclusions Claudin 7 and 11 were not expressed in LS and SCC. However, there was no significant difference in expression of any of the claudins between the LS and SCC samples. Furthermore, p53 expression is the highest in SCC patients and lowest in the control group. However, expression of p53 did not vary between samples from isolated LS and LS associated SCC patients, suggesting that increased p53 expression is not the determining factor in the progression of LS lesions to SCC.