Expression of Pluripotency Factors OCT4 and LIN28 Correlates with Survival Outcome in Lung Adenocarcinoma.

Background and Objectives: Lung adenocarcinoma is a leading cause of cancer-related mortality despite recent therapeutic advances. Cancer stem cells have gained increasing attention due to their ability to induce cancer cell proliferation through self-renewal and differentiation into multiple cell lineages. OCT4 and LIN28 (and their homologs A and B) have been identified as key regulators of pluripotency in mammalian embryonic (ES) and induced stem (IS) cells, and they are the crucial regulators of cancer progression. However, their exact role in lung adenocarcinoma has not yet been clarified. Materials and Methods: The aim of this study was to explore the role of the pluripotency factors OCT4 and LIN28 in a cohort of surgically resected human lung adenocarcinomas to reveal possible biomarkers for lung adenocarcinoma prognosis and potential therapeutic targets. The expressions of OCT4, LIN28A and LIN28B were analyzed in formalin-fixed, paraffin-embedded tissue samples from 96 patients with lung adenocarcinoma by immunohistochemistry. The results were analyzed with clinicopathologic parameters and were related to the prognosis of patients. Results: Higher OCT4 expression was related to an improved 5-year overall survival (OS) rate (p < 0.001). Nuclear LIN28B expression was lower in stage I and II tumors (p < 0.05) compared to advanced stage tumors. LIN28B cytoplasmic expression was associated with 5-year OS rates not only in univariate (p < 0.005), but also in multivariate analysis (where age, gender, histopathological subtype and stage were used as cofactors, p < 0.01 HR = 2.592). Patients with lower LIN28B expression showed improved 5-year OS rates compared to patients with increased LIN28B expression. Conclusions: Our findings indicate that OCT4 and LIN28B are implicated in lung adenocarcinoma progression and prognosis outcome; thus, they serve as promising prognostic biomarkers and putative therapeutic targets in lung adenocarcinomas.

Antibodies to aquaporins are frequent in patients with primary Sjögren's syndrome.

Objectives: Several aquaporins (AQPs) are present in the salivary glands, likely contributing to their secretions. AQP dysfunction may contribute to the salivary gland dysfunction in SS. Antibodies to AQP4 and AQP1 are detected in neuromyelitis optica and are believed to play a pathogenic role. We aimed to search for antibodies to several AQPs in the sera from SS patients in an effort to shed light on the pathogenic mechanisms of SS. Methods: We searched for antibodies to six AQPs in the sera of 34 SS patients without neurological findings using ELISAs with synthetic peptides corresponding to the three extracellular domains of each AQP, radioimmunoassays with AQPs, Western blots and competition experiments with cell-embedded AQPs. Results: Thirteen (38.2%) SS patients had antibodies to extracellular domains of AQP1 (two), AQP3 (one), AQP8 (six) or AQP9 (four); none had AQP4 or AQP5 antibodies. Each patient had antibodies to only one extracellular domain. AQP binding was further verified by radioimmunoassay with intact AQPs, western blots and AQP-transfected cells. In contrast, none of the 106 healthy controls or 68 patients with other autoimmune diseases had antibodies to intact AQPs. Expression of AQP8 (the major antibody target) on human salivary glands was shown by immunohistochemistry. Patients with anti-AQP antibodies had more severe xeropthalmia compared with anti-AQP-negative patients, suggesting a potential pathogenic role of these antibodies. Conclusion: Antibodies to AQPs (especially to AQP8 and AQP9) are frequent in SS patients. The likely important role of AQPs in salivary gland secretions justifies further research.

Occipital lobe gumma: a case report and review of the literature.

Syphilis has plagued humanity for thousands of years. Despite the measures of precaution against its transmission and the advancement of modern pharmacology, late-stage phenomena like intracerebral gumma are not uncommon even today. We present a complex case, which has misled the physicians twice. Additionally, we performed a review of the contemporary literature about the common location, clinical findings and up-to-date treatment of intracerebral gummas.

Molecular classification of nonsmall cell lung cancer using a 4-protein quantitative assay.

BACKGROUND: The importance of definitive histological subclassification has increased as drug trials have shown benefit associated with histology in nonsmall-cell lung cancer (NSCLC). The acuity of this problem is further exacerbated by the use of minimally invasive cytology samples. Here we describe the development and validation of a 4-protein classifier that differentiates primary lung adenocarcinomas (AC) from squamous cell carcinomas (SCC). METHODS: Quantitative immunofluorescence (AQUA) was employed to measure proteins differentially expressed between AC and SCC followed by logistic regression analysis. An objective 4-protein classifier was generated to define likelihood of AC in a training set of 343 patients followed by validation in 2 independent cohorts (n = 197 and n = 235). The assay was then tested on 11 cytology specimens. RESULTS: Statistical modeling selected thyroid transcription factor 1 (TTF1), CK5, CK13, and epidermal growth factor receptor (EGFR) to generate a weighted classifier and to identify the optimal cutpoint for differentiating AC from SCC. Using the pathologist's final diagnosis as the criterion standard, the molecular test showed a sensitivity of 96% and specificity of 93%. Blinded analysis of the validation sets yielded sensitivity and specificity of 96% and 97%, respectively. Our assay classified the cytology specimens with a specificity of 100% and sensitivity of 87.5%. CONCLUSIONS: Molecular classification of NSCLC using an objective quantitative test can be highly accurate and could be translated into a diagnostic platform for broad clinical application.

Standardization of epidermal growth factor receptor (EGFR) measurement by quantitative immunofluorescence and impact on antibody-based mutation detection in non-small cell lung cancer.

Challenges in measurement of epidermal growth factor receptor (EGFR) protein expression have led to conflicting data on its prognostic value and discontinuation of its use for prediction of response. Herein is described a quantitative standardized assay for EGFR and its use in a series of retrospective cohorts of patients with non-small cell lung cancer (NSCLC). The AQUA technology of quantitative immunofluorescence was used in conjunction with Western blot analysis to calculate the absolute concentration of EGFR in two independent NSCLC cohorts (170 from Yale New Haven Hospital and 335 from Sotiria and Patras Hospitals in Greece). EGFR and mutated EGFR were measured using D38B1 antibody and two mutation-specific antibodies. All patients positive or borderline for mutation-specific antibody were genotyped. A threshold for reproducible detection of EGFR was defined as 0.85 ng/µg total protein. EGFR expression demonstrated no prognostic value in either cohort. The mutation rate was 1.79% in the Yale cohort, and 1.52% in the Sotiria/Patras cohort, with no antibody detection-based false-positive cases. No mutations were detected for EGFR concentrations <1.46 ng/µg total protein. In summary, accurate measurement of EGFR still shows no prognostic value in NSCLC. In these two population-based cohorts, the antibody-based EGFR mutation rate was lower than has been frequently reported.

Prognostic value of novel biomarkers in astrocytic brain tumors: nuclear receptor co-regulators AIB1, TIF2, and PELP1 are associated with high tumor grade and worse patient prognosis.

Estrogen receptors alpha (ERα) and beta (ERß) and their co-regulatory proteins are key components of complex signaling networks that specifically regulate the growth and development of various tissues and tumors. Still, their protein expression profiles and possible role in the pathogenesis of astrocytic tumors remain largely unknown. The purpose of the present study is to evaluate the differential protein expression of ΕRα, ERß, and their co-activators, AIB1, TIF2, and PELP1 in astrocytic tumors of World Health Organization (WHO) grade II-IV, using immunohistochemistry. Potential correlations with clinicopathological parameters and patient prognosis were also explored. ERα protein expression was undetectable while ERß levels were significantly decreased with progression of tumor grade (P < 0.001). High expression of ERß was an independent favorable prognostic factor on multivariate analysis (P = 0.003). Expression of AIB1, TIF2, and PELP1 was not correlated with ERß expression and followed an opposite trend, with increasing levels in high-grade relative to low-grade tumors (P < 0.001). Univariate survival analysis revealed that high AIB1, TIF2, and PELP1 expression was associated with worse prognosis (P = 0.049, P = 0.033, and P = 0.020, respectively). ERß and ER co-activators AIB1, TIF2, and PELP1 appear to play an important role in the pathogenesis and progression of astrocytic tumors and might have prognostic significance. The mechanisms underlying their involvement in astrocytic tumorigenesis, as well as their utility for prognostic and therapeutic purposes merit further investigation.

Heat shock factor 1 in brain tumors: a link with transient receptor potential channels TRPV1 and TRPA1.

Novel data report a "cross-talk" between Heat-Shock Factor 1 (HSF1) and the transient receptor potential vanilloid 1 cation channel (TRPV1) located in the cell membrane, introducing these channels as possible drug targets for the regulation of HSF1 activation. This study aims to investigate the co-expression of TRPV1 and HSF1 in human brain tumors. Additionally, the expression of the transient receptor potential ankyrin 1 channel (TRPA1), which is co-operated with TRPV1 in a plethora of cells, was studied. Immunohistochemical staining for HSF1, TRPV1 and TRPA1 expression was quantitatively analyzed in paraffin-embedded semi-serial tissue sections from 74 gliomas and 71 meningiomas. mRNA levels of HSF1, TRPV1 and TRPA1 were evaluated using real-time PCR. Although HSF1 was significantly increased compared with TRPV1/TRPA1 (p ≤ 0.001) in both gliomas and meningiomas, high co-expression levels for HSF1, TRPV1 and TRPA1 were found in 62.50% of diffuse fibrillary astrocytomas (WHO, grade II), 37.50% of anaplastic astrocytomas (WHO, grade III), 16.32% of glioblastomas multiforme (WHO, grade IV), and 42.25% of meningiomas (WHO, grade I and II). Correlation analysis revealed a relationship of HSF1 with TRPV1/TRPA1 in diffuse fibrillary astrocytomas (WHO, grade II) and benign meningiomas (WHO, grade I) contrary to glioblastomas multiforme (WHO, grade IV) and high grade meningiomas (WHO, grade II). Importantly, TRPA1 and TRPV1 expression levels were significantly increased in meningiomas compared with astrocytic tumors (p < 0.05). In conclusion, HSF1 and TRPV1/TRPA1 co-expression may be implicated in the pathogenesis of human brain tumors and should be considered for the therapeutic approaches for these tumors.

H3K4 Methylation Status and Lysine Specific Methyltransferase KMT2C Expression Correlate with Prognosis in Lung Adenocarcinoma.

BACKGROUND: Genetic events cannot account for the complexity of human carcinogenesis alone. Mutations of epigenetic regulators and aberrations of their expression patterns have been detected in various human malignancies. Methylation of histone H3 at lysine 4 (H3K4me), is an evolutionarily conserved histone modification that marks regions of active transcription and regulates cell growth, migration, and invasion. The MLL/KMT2 family of histone methyltransferases specifically methylate histone H3 at lysine 4. OBJECTIVE: The aim of this study was to explore the role of KMT2C/MLL3 as well as key histone modification activating markers, such as H3K4me2 and H3K4me3 in a cohort of surgically resected human lung adenocarcinomas in an effort to reveal possible biomarkers for lung adenocarcinoma diagnosis and prognosis and potential therapeutic targets. METHOD: The immunohistochemical expression of KMT2C/MLL3, H3K4me2 and H3K4me3 was analyzed in formalin fixed paraffin embedded tissue from 96 patients with lung adenocarcinoma. Results were associated with clinicopathologic parameters and patient's prognosis. RESULTS: Nuclear expression of KMT2C/MLL3 in epithelial cells was independently associated with shorter overall survival. Cytoplasmic H3K4me2 expression was associated withT stage and nuclear H3K4me2 expression was associated with female gender and patients' prognosis. The latter association persisted after multivariate analysis. No association was found between H3K4me3 expression and clinicopathological data or disease outcome in our cohort of patients. CONCLUSION: These results suggest that the pattern of histone modifications and KMT2C/MLL3 expression can be used as an independent prognostic factor in lung adenocarcinoma, revealing that chromatin remodeling is criticallyinvolved in cancer progression.

High expression of BCL-2 predicts favorable outcome in non-small cell lung cancer patients with non squamous histology.

BACKGROUND: Bcl-2 promotes cell survival by inhibiting adapters needed for the activation and cleavage of caspases thus blocking the proteolytic cascade that ultimately dismantles the cell. Bcl-2 has been investigated as a prognostic factor in non small cell lung cancer (NSCLC) patients with conflicting results. METHODS: Here, we quantitatively assessed Bcl-2 expression in two large and independent cohorts to investigate the impact of Bcl-2 on survival. AQUA(R), a fluorescent-based method for analysis of in situ protein expression, was used to measure Bcl-2 protein levels and classify tumors by Bcl-2 expression in a cohort of 180 NSCLC patients. An independent cohort of 354 NSCLC patients was used to validate Bcl-2 classification and evaluate outcome. RESULTS: Fifty % and 52% of the cases were classified as high expressers in training and validation cohorts respectively. Squamous cell carcinomas were more likely to be high expressers compared to adenocarcinomas (63% vs. 45%, p = 0.002); Bcl-2 was not associated with other clinical or pathological characteristics. Survival analysis showed that patients with high BCL-2 expression had a longer median survival compared to low expressers (22 vs. 17.5 months, log rank p = 0.014) especially in the subset of non-squamous tumors (25 vs. 13.8 months, log rank p = 0.04). Multivariate analysis revealed an independent lower risk for all patients with Bcl-2 expressing tumors (HR = 0.53, 95% CI 0.37-0.75, p = 0.0003) and for patients with non-squamous tumors (HR = 0.5, 95% CI 0.31-0.81, p = 0.005). CONCLUSIONS: Bcl-2 expression defines a subgroup of patients with a favorable outcome and may be useful for prognostic stratification of NSCLC patients.

Histologic-type specific role of cell cycle regulators in non-small cell lung carcinoma.

BACKGROUND: Lung cancer is the most lethal type of cancer in humans. Cell cycle alterations have commonly been encountered in lung cancer and may have prognostic value. MATERIALS AND METHODS: This study investigates the immunohistochemical expression of the important cell cycle regulators phosphatase and tensin homolog deleted on chromosome 10 (PTEN), p27, Cks1, and Skp2 in 128 non-small cell lung carcinomas (64 adenocarcinomas, 46 squamous cell carcinomas, and 18 large cell undifferentiated carcinomas) and adjacent non-neoplastic lung tissue. RESULTS: PTEN and p27 were always highly expressed in non-neoplastic lung whereas Cks1 and Skp2 were not expressed in normal tissue. Decreased PTEN expression was noted in 19/64 adenocarcinomas, 15/46 squamous cell carcinomas, and 7/18 undifferentiated large cell carcinomas. Reduced expression of p27 was noted in 28/64, 19/46, and 6/18 of the tumors, respectively. Increased expression of Cks1 was seen in 38/64, 26/46, and 11/18 and increased expression of Skp2 in 29/64, 30/46, and 14/18 of the tumors, respectively. An inverse relationship between p27 and Skp2 levels was found in adenocarcinomas and between p27 and Cks1 levels in squamous cell carcinomas. Decreased PTEN and p27 expression were associated with advanced tumor stage in squamous cell carcinomas. Univariate analysis showed that high p27 and PTEN and low Cks1 expression correlated with increased survival in patients with squamous cell carcinoma independently of tumor stage. CONCLUSIONS: Aberrant expression of PTEN, p27, Cks1, and Skp2 is a common feature of all three major types of non-small cell lung cancer NSCLC, but seems to be involved in the progression of squamous cell carcinoma alone.

Expression patterns of leptin receptor (OB-R) isoforms and direct in vitro effects of recombinant leptin on OB-R, leptin expression and cytokine secretion by human hematopoietic malignant cells.

Several studies have implicated leptin in the pathophysiology of neoplasias. We investigated the direct effect of leptin on malignant hematopoietic tissue that included: primary acute myeloid leukemia (AML) cells, leukemic cell lines and bone marrow biopsies from multiple myeloma (MM) patients. PBMC, T-cells, B-cells and monocytes from healthy subjects served as controls. We defined the patterns of OB-R isoform expression in AML cells and leukemic cell lines in comparison to control cells by RT-PCR. rLeptin upregulated the expression of OB-R and endogenous leptin in AML blasts and certain cell lines but not in control cells. Cytometric Bead Array analysis of pro- and anti-inflammatory cytokines showed that rleptin upregulates IL-6 secretion by AML cells, various cytokines by the leukemic cell lines tested and IL-10 secretion by control PBMC, contributed by monocytes. Western immunoblotting revealed that the effect of rleptin was independent of JAK-2/phospho-JAK-2 protein levels. Finally, MM biopsies stained positive for leptin and, to a lesser extend, OB-R. Immunoreactivity was confined mostly to the nucleus of the myeloma cells. Normal myelocytes, promyelocytes and megakaryocytes stained weakly positive, and erythroid cells were constantly negative. We propose that the leptin/OB-R system is strongly and directly involved in supporting the growth of hematopoietic malignancies.

The potential role of Bcl-2 expression, apoptosis and cell proliferation (Ki-67 expression) in cases of gastric carcinoma and correlation with classic prognostic factors and patient outcome.

BACKGROUND: This study investigated the presence of apoptosis and proliferation in gastric cancer and assesses their possible correlation with classic prognostic markers and patients' survival. PATIENTS AND METHODS: The study comprised 110 patients with gastric carcinoma who underwent gastrectomy for therapeutic reasons, and did not receive any pre- or postoperative treatment. Patients were followed up for 3.5-140 months. Thick paraffin sections (4 microm) were subjected to immunohistochemistry using anti-Bcl-2 and anti-Ki-67 antibodies and to in situ hybridization [TUNEL method-apoptotic body index (ABI)]. Morphological and immunohistochemical results were correlated with clinicopathologic parameters. RESULTS: Bcl-2 protein was detected in 67% of adenocarcinomas with increased expression in low-grade and early-stage tumors. Bcl-2 expression did not correlate with Ki-67 index, ABI or patients' survival. Ki-67 expression was correlated with a poorer survival rate. Apoptosis was more frequently observed in advanced stage and high-grade tumors. Cox analysis revealed that tumor stage and grade, as well as Ki-67 index, constituted independent prognostic factors. CONCLUSION: This study included patients with gastric cancer none of whom received any additional pre- or post-operative treatment. Thus the prognostic value of each marker studied was not affected by additional treatments. Bcl-2 expression in advanced-stage and high-grade gastric carcinomas, indicate that Bcl-2 is involved in early stage of tumor development. Ki-67 expression constitutes an independent prognostic factor regarding the outcome of patients with gastric cancer. The positive association between apoptosis and proliferation suggests that apoptosis might reflect not only cell loss but also the proliferative activity. However, further research is required in order to determine if these markers may provide useful information for the prediction of prognosis in patients with colorectal carcinoma.

Apoptotic mechanisms within the retina in Staphylococcus epidermidis experimental endophthalmitis.

AIM: To investigate the potential involvement of apoptosis and its regulators Bcl-2, Bax, and Fas within the retina in Staphylococcus epidermidis experimental endophthalmitis. METHODS: Endophthalmitis was induced in 48 male Lewis rats by unilateral 25-mircrol intravitreal injection of 7,000 viable organisms of slime-producing S. epidermidis strain ATCC 35983 (experimental group). Forty-eight other Lewis rats received a similar sterile normal saline injection (control group). The injected eyes were graded for clinical inflammation and were removed in groups at 6, 12, 24, 48, 72, and 168 hours post-injection. After surgical separation, retinal tissue specimens were fixed, and paraffin sections underwent hematoxylin-eosin staining, immunohistochemistry against Bcl-2, Bax, and Fas, and TUNEL assay for detection of apoptotic cells. Following morphometric analysis, the apoptotic body index (ABI) was calculated. RESULTS: While Bcl-2 expression was absent, Bax and Fas expression and apoptosis in ganglion cells, bipolar cells, and photoreceptors, were significantly higher in the experimental group compared to the control group (P < 0.05). In the experimental group, inflammation peaked at 24 hours, Bax and Fas expression at 48 hours and the ABI at 72 hours post-injection. CONCLUSION: Apoptosis is increased within the retina in S. epidermidis experimental endophthalmitis through upregulation of Bax and Fas, peaking soon after peak inflammation.

The role of postoperative radiotherapy in the management of patients with thymic tumors -- a retrospective study.

BACKGROUND: Thymomas are the most common tumor arising in the anterior mediastinum. Surgery is the cornerstone for the management of these tumors. The role of postoperative radiotherapy in Masaoka stage II thymomas remains controversial, but it is well established in the advanced stages. The aim of this study was to investigate the role of postoperative radiotherapy in the overall management of thymomas, and the evaluation of potential prognostic factors. PATIENTS AND METHODS: Between 1989 and 2007, 41 thymoma patients underwent surgery and 27 of them received radiotherapy with a curative intent. The Masaoka staging system was used. The histopathological records and specimens of patients were thoroughly reviewed. Clinical and radiological evaluations took place every 6 months. The mean patient follow-up was 69 months (range: 2-212). RESULTS: DFS (disease free survival), TS (total survival) and DSS (disease specific survival) differed significantly between stages and histological types (p<0.04). Stage I patients were managed only surgically, with none recurring or dying. Concerning stage II patients, TS was significantly longer in non-irradiated cases (10/21) (p=0.025). Stage III (n=8) and IV (n=8) patients underwent postoperative radiotherapy, with 4/8 of stage IV disease also receiving induction chemotherapy. Six out of 8 stage III-IV patients recurred (1 distant and 5 intrathoracic failure), out of whom 4 died due to disease progression despite further treatment (all type C histology). The mean DFS and TS for stage III patients were 49.2 and 50.3 months respectively, with the corresponding values for stage IV being 14.5 and 29.1 months. Patients with myasthenia had a favorable outcome and the ones with complete resection a significantly longer DFS (p=0.0003) and DSS (p=0.039). The Cox regression analysis showed that myasthenia and tumor size are important prognostic factors for DFS (p<0.05). CONCLUSION: Myasthenic patients have a more favorable prognosis. Radiotherapy can be omitted in totally resected stage I-II patients, whereas it is beneficial in more advanced stages.

Transient receptor potential vanilloid (TRPV) channel expression in meningiomas: prognostic and predictive significance.

The TRPV1-4 members of TRPV cation channel subfamily are mainly regarded as polymodal receptors that may be activated by diverse changes in cellular microenvironment and endogenous and exogenous agents. Abnormal expression of these channels has been reported in various tumors but not in meningiomas. Meningioma cells are thought to originate from arachnoid cap cells due to cytological and functional similarities between the two types of cells. To investigate the expression profile of TRPV1-4 channels in meningiomas and compare with TRPV1-4 channel expression in leptomeninges, we used immunohistochemistry in formalin-fixed, paraffin-embedded semi-serial tissue sections from 175 meningiomas with different grades and histological subtypes, and normal brain or meningioma specimens that contained leptomeninges. The labeling index (LI), defined as the percentage of positive (labeled) cells out of the total number of tumor cells counted, was determined. Leptomeninges were TRPV1-4 immunonegative. A significant percentage of tumors exhibited TRPV1-4 channel expression which was independent of the proliferation index of the tumors but was significantly associated with histopathological subtypes. The TRPV1 and TRPV3 immunoexpression was decreased whereas TRPV4 immunoexpression was significantly greater in high-grade (WHO, grade II and III) as compared with low-grade (WHO, grade I) meningiomas. Additionally, TRPV4 emerged as an independent predictor for the degree of malignancy using the binary logistic regression model [dependent variable: grade I versus higher grades (II and III)]. Kaplan-Meier analysis for 102 patients showed no significant association of TRPV1-4 expression with overall survival. The above data support that TRPV1-4 channels are implicated in meningioma pathogenesis, and TRPV4 has predictive significance in the disease.

p75 and TrkC neurotrophin receptors demonstrate a different immunoreactivity profile in comparison to TrkA and TrkB receptors in human normal pituitary gland and adenomas.

BACKGROUND/AIMS: Recent knowledge indicates that neurotrophins play a significant role in neuroendocrine systems through their specific receptors TrkA, TrkB, TrkC and low-affinity p75(NTR) receptor. TrkA and TrkB receptors have been previously detected in numerous endocrine cells in human anterior pituitary and adenomas. In the present study, the localization of p75(NTR) and TrkC along with TrkA and TrkB receptors was investigated. METHODS: Semi-serial paraffin-embedded sections of 5 human normal pituitaries and 30 adenomas were immunostained using specific antibodies. RESULTS: Expression of p75(NTR) receptor was demonstrated in the intricate capillary and reticulin network in the anterior pituitary and in the pericapillary tissue and pituicytes in the posterior lobe. p75(NTR) immunoreactivity was absent from all adenomas. In normal anterior pituitary, a few scattered cells showed weak TrkC immunoreactivity in contrast to a high percentage of endocrine cells distributed throughout the pars distalis and pars intermedia which exhibited strong TrkA and/or intermediate TrkB immunoreactivity. Double immunohistochemistry demonstrated TrkA immunoreactivity in more than 80% of lactotropes and 70% of corticotropes and to a lesser extent in other cell types. Furthermore, in the majority of adenomas, independently of type, sex and age, a high percentage of TrkA- and/or TrkB-positive cells was detected. Interestingly, TrkC expression appeared to be increased in some adenomas compared to normal pituitary. Endothelial cells and perivascular connective tissue were always TrkB-immunostained. CONCLUSION: The above findings support a potential role of all neurotrophins, through their different receptors, in pituitary functions.

Expression of EMT inducers integrin-linked kinase (ILK) and ZEB1 in phyllodes breast tumors is associated with aggressive phenotype.

Phyllodes tumors (PTs) of the breast constitute an uncommon group of mammary fibroepithelial lesions with ambiguous biologic behavior. Recent evidence suggests that epithelial mesenchymal transition (EMT), a driving force of cancer progression is implicated in PTs pathogenesis. Integrin-linked kinase (ILK), a focal adhesion kinase, has been implicated in cancer and EMT and represents a novel cancer therapeutic target. In this study, we aimed to investigate ILK and EMT markers expression in phyllodes breast tumors in relation to tumor grade. Expression of ILK and EMT markers E-cadherin, ß-catenin, Ν-cadherin, vimentin, Snail, ZEB1 and Twist was evaluated by immunohistochemistry in paraffin-embedded tissue sections from 96 human phyllodes breast tumors (48 benign, 27 borderline, 21 malignant). Cytoplasmic and nuclear immunopositivity of ILK were observed in both the epithelial and the stromal component of phyllodes breast tumors and were significantly higher with increasing tumor grade. An EMT-related expression profile consisting of decreased membranous and increased nuclear/cytoplasmic immunoreactivity of E-cadherin and ß-catenin and increased expression of N-cadherin, vimentin, Snail, ZEB1 and Twist was observed in tumor epithelial and stromal component and was significantly associated with malignant phyllodes breast tumor histopathology. Interestingly, there was a significant correlation of ILK expression with all of the EMT markers examined. Our results suggest that EMT significantly contributes to phyllodes tumor pathogenesis and originally implicate ILK and ZEB1 in phyllodes tumors malignant phenotype.

ILK Expression in Colorectal Cancer Is Associated with EMT, Cancer Stem Cell Markers and Chemoresistance.

BACKGROUND/AIM: Epithelial-mesenchymal transition (EMT) and cancer stem cells (CSC) are critically implicated in cancer metastasis and chemoresistance. Herein, we investigated integrin-linked kinase (ILK)'s role in human colon cancer (CRC) progression and chemoresistance in relation to EMT and CSC markers. PATIENTS AND METHODS: Expression of ILK, EMT and CSC markers were evaluated by immunohistochemistry in 149 CRC samples. We also generated colon cancer cells resistant to 5-FU and oxaliplatin and studied the effect of ILK inhibition on drug response by MTT assay and on EMT and CSC markers' expression. RESULTS: ILK expression in human CRC correlates with EMT and CSC markers and is associated with metastasis and chemoresistance. ILK inhibition increases sensitivity of resistant cells to 5-FU and oxaliplatin and reduces the levels of EMT and CSC markers in 5-FU resistant cells. CONCLUSION: ILK overexpression in human CRC associates with EMT and CSC traits, contributing to tumor progression and chemoresistance.

Expression of the regulatory cell cycle proteins p21, p27, p14, p16, p53, mdm2, and cyclin E in bone marrow biopsies with acute myeloid leukemia. Correlation with patients' survival.

Cell cycle control is a crucial event in normal hematopoiesis, and abnormalities of regulatory cell cycle genes have been found to contribute to the development of many hematologic malignancies. The present study investigates the immunohistochemical expression of seven essential cell cycle proteins (p21, p27, p14, p16, p53, mdm2, and cyclin E) in paraffin-embedded sections from 42 bone marrow biopsies obtained from an equal number of patients with newly diagnosed acute myeloid leukemia (AML). This study revealed (i) a high frequency of p53+/mdm2-/p21-phenotype, which is probably a result of p53 gene mutation and/or inhibition of mdm2 action by p14(ARF); (ii) expression of p27+/cyclinE-phenotype in most cases, suggesting that p27 may act as a potent cyclin-dependent kinase inhibitor; (iii) expression of p16 only in very few cases; and (iv) no relationship between the expression of any of the above proteins and survival as well as histologic subtype.

Increased Bax/Bcl-2 ratio up-regulates caspase-3 and increases apoptosis in the thymus of patients with myasthenia gravis.

BACKGROUND: In this study the possible relation of Bax (an apoptosis promoter) to Bcl-2 (an apoptosis inhibitor) ratio with the apoptosis co-ordination enzyme, caspase-3, in the thymus of patients with myasthenia gravis (MG) was investigated in correlation with long-term clinical prognosis. PATIENTS AND METHODS: The study included 46 patients (17M/29F, mean age 36.60 +/- 16.09 yr) with MG, who underwent thymectomy for treatment. The clinical staging (Osserman classification) included: stage 1-5, IIA-21, IIB-17, III-3. The pathology of the thymus showed: hyperplasia-26, atrophy-8, thymoma B1 and B2 type-9, thymoma B3 type (well differentiated thymic carcinoma)-3. The patients were evaluated 39-166 (mean 91.87 +/- 38.38) months after thymectomy. At the end of the follow-up period, the patients were classified as follows: group A: complete stable remission, group B: pharmacological remission + minimal manifestations + improvement + deterioration. Paraffin sections of thymic tissue were subjected to: a) immunohistochemistry (bax, bcl-2, caspase-3 protein); b) in situ hybridization (bax, bcl-2 mRNA); and c) TUNEL-stain (apoptotic cells). Bax to bcl-2 mRNA and protein ratio was determined for each sample by dividing the % bax (+) cells by the % bcl-2 (+) cells. RESULTS: Follow-up data were available for 39/46 patients: 13/39 patients belonged to group A and 26/39 to group B. The Bax/Bcl-2 mRNA and protein ratios were increased towards advanced disease stages (+370% for mRNA and +391% for protein, from MG stage I to stage III). These ratios were correlated with caspase-3 expression (r = 0.782 and 0.583, p < 0.01) and apoptosis (r = 0.591 and 0.358 p < 0.01 and p < 0.05). All the 13 cases in group A had a Bax/Bcl-2 ratio < 1 (mean +/- SD: 0.58 +/- 0.04 for mRNA and 0.62 +/- 0.03 for protein), whereas all the 26 cases of group B had a ratio > 1 (1.47 +/- 0.07 for mRNA and 1.52 +/- 0.18 for protein). The Kaplan-Meier survival curve showed higher, free of disease, survival in group A (p = 0.0082). Cox regression analysis revealed that the Bax/Bcl-2 ratio was an independent prognostic factor, however the p-value was marginally significant (95% CI: 1.078-44.073, p = 0.041). CONCLUSION: This study has demonstrated that in patients with MG who underwent thymectomy: a) the Bax/Bcl-2 ratio may up-regulate caspase-3 expression and modulate apoptosis associated with progress of the disease; b) the Bax/Bcl-2 ratio < 1 was associated with complete stable remission after thymectomy; and c) Bax/Bcl-2 ratio was an independent predictive marker for therapeutic response after thymectomy.