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BACKGROUND In the literature, there exist conflicting data on the value of fibulin-3 (FBLN3) for the diagnosis of pleural effusion (PE) in malignant pleural mesothelioma (MPM). Therefore we compared the diagnostic performance of FBLN3 against that of soluble mesothelin-related peptide (SMRP) in a cohort of Italian patients. MATERIALS AND METHODS FBLN3 and SMRP were detected in PE from 33 patients with MPM, 64 with pleural benign lesions and 23 with non-MPM pleural metastases using a commercial enzyme-linked-immunosorbent(ELISA)-assay kit according to manufacturers' instructions. RESULTS Levels of FBLN3 were similar in PE from MPM and PE from other pathologies (geometric mean=68.1 vs. 66.2 ng/ml; p=0.872) in contrast to SMRP levels, which were significantly higher in PE from MPM (geometric mean=14.6 vs. 3.2 nM; p

CTLA-4 function as a negative regulator of T cell-mediated immune response is well established, whereas much less is known about the immunoregulatory role of its soluble isoform (sCTLA-4). No data are available on CTLA-4 expression and prognostic impact in malignant pleural mesothelioma (MPM). We investigated, by immunohistochemistry, CTLA-4 expression in tumor tissues and, by ELISA, sCTLA-4 levels in sera and matched pleural effusions from 45 MPM patients. Prognostic effect of CTLA-4 expression on overall survival (OS) was assessed through Cox regression and prognostic significance expressed as death rate ratio (HR). We found that 56.0 % of MPM tissues expressed CTLA-4 with variable intensity and percentage of positive cells estimated by the immunoreactive score. sCTLA-4 levels were significantly higher in sera (S-sCTLA-4) than in pleural effusions (PE-sCTLA-4) (geometric mean ratio = 2.70, P value = 0.020). CTLA-4 expression at the tissue level was higher in the epithelioid histological subtype than in the sarcomatoid, whereas at the serum level, it was higher in the sarcomatoid subtype. A homogeneous favorable prognostic effect was found for CTLA-4 overexpression in tissue, serum and pleural effusion. Interestingly, only the PE-sCTLA-4 was found to be a statistically significant positive prognostic factor (HR = 0.37, 95 % CI = 0.18-0.77, P value = 0.007). Indeed, PE-sCTLA-4 correlated with CTLA-4 expression in tissues, whereas this latter expression showed a weak association with OS. To confirm our findings, further experimental evidences obtained from a larger cohort of MPM patients are required. However, our results would indicate a positive correlation of PE-sCTLA-4 levels and OS in MPM patients.

We describe a 70 year old woman affected by diffuse pulmonary lymphoid hyperplasia (DPLH). DPLH is a rare clinical-pathological entity generally associated with connective tissue diseases, but we diagnosed our case as an idiopathic DPLH. To the best of our knowledge, this is the first case of idiopathic DPLH reported in medical literature.

We report a case of acute fibrinous and organising pneumonia in Whipple’s disease with lung improvement after antibiotic therapy. In our knowledge this is the first report of Whipple’s disease with acute fibrinous and organising pneumonia.

Hypoxia is a typical hallmark of many solid tumors and often leads to therapy resistance and the development of a more aggressive cancer phenotype. Oxygen content in tissues has been evaluated using numerous different methods for several imaging modalities, but none has yet reached the required standard of spatial and temporal resolution. Magnetic Resonance Imaging (MRI) appears to be the technique of choice and several pO2-responsive probes have been designed for it over the years. In vivo translation is often hampered in Gd-relaxation agents as it is not possible to separate effects that arise from changes in local concentration from those associated with responsive properties. A novel procedure for the MRI based assessment of hypoxia is reported herein. The method relies on the combined use of Gd-DOTP- and Gd-HPDO3A-labeled red blood cells (RBCs) where the first probe acts as a vascular oxygenation-responsive agent, while the second reports the local labeled RBC concentration in a transplanted breast tumor mouse model. The MRI assessment of oxygenation state has been validated by photoacoustic imaging and ex vivo immunofluorescence. The method refines tumor staging in preclinical models and makes possible an accurate monitoring of the relationship between oxygenation and tumor growth.

Purpose: Dysregulated cytokine/cytokine receptor expression may occur in B-cell lymphoproliferative disorders. Little information is available on interleukin-18 receptor (IL-18R) and IL-18 expression in normal and malignant B cells. Our purpose was to investigate this issue in human naive, germinal center (GC) and memory B cells, and in their neoplastic counterparts. Experimental Design: We have evaluated IL-18 expression and production in tonsil naive, GC, and memory B cells and in their presumed neoplastic counterparts by reverse transcription-PCR and ELISA. Moreover, IL-18Rα and β expression was investigated in the same cells by reverse transcription-PCR, flow cytometry, and immunohistochemistry. Results: We found that: (a) IL-18 mRNA was expressed in tonsil naive, GC, and memory B cells. Bioactive IL-18 was secreted by naive and GC, but not by memory B cells; (b) IL-18Rα and β transcripts were expressed in the three B-cell subsets. IL-18Rα was detected on the surface of naive, GC, and memory B lymphocytes, and IL-18Rβ was detected on GC and memory, but not naive, B cells; (c) mantle zone, follicular, marginal zone, Burkitt lymphoma (BL), and B-cell chronic lymphocytic leukemia (B-CLL) cells expressed IL-18 mRNA. B-CLL and BL cells did not produce bioactive IL-18; and (d) lymphoma B cells displayed heterogeneous expression of either or both IL-18R chain mRNA. In contrast, B-CLL cells expressed both IL-18R chains at the mRNA and protein levels. Conclusions: Dysregulated expression of IL-18 and/or IL-18R in chronic B-cell lymphoproliferative disorders may sometimes contribute to tumor escape from the host immune system.

Splenosis can be considered as a benign condition due to the presence of heterotopic splenic tissue in abdomen, thorax and pelvis because of a massive splenic trauma or surgery. Here we report the case of a patient with an unknown hepatic mass, chronic hepatitis C, liver cirrhosis and a previous splenectomy after abdominal trauma. In our case lesion could not be clearly defined by ultrasound, computed tomography and magnetic resonance. Classical features of malignancy were not demonstrated at computed tomography, while at magnetic resonance imaging the differential diagnosis of the lesion appeared quite difficult and debate. Then an ultrasound-guided biopsy demonstrated the liver mass in left hepatic lobe consisted of splenic tissue and some millimetric accessory spleens in the left upper quadrant. So the possibility of an intra-hepatic splenosis should be taken into account in patients with an unknown liver mass and a history of previous abdominal trauma, followed by splenectomy. The conclusive diagnosis of intra-hepatic splenosis was given by ultrasound-guided biopsy.

Mesothelin (SMRP) is regarded as a biomarker of malignant pleural mesothelioma (MPM). Herein, we analyzed the contribution of SMRP detection in pleural effusion and in serum to the diagnosis of MPM with non-positive cytology.The present study included 52 cases of MPM, 43 of pleural benign lesions and 25 of non-MPM pleural metastases. SMRP was measured by MesoMark ELISA (Cis-Bio International Gif/Yvette; France).In non-positive cytology, effusion-SMRP showed higher diagnostic performance than serum-SMRP. We found 38 out of 52 (73.1%) cases of non-positive cytology MPM, out of which 27 (71.0%) were positive for effusion-SMRP (cut-off=12.70 nM) and 18 (47.4%) for serum-SMRP (cut-off=1.08 nM). When cytology, effusion- and serum-SMRP were used in combination, an overall sensitivity in detection of MPM of 78.9% was achieved. The same sensitivity was obtained by combining cytology with effusion-SMRP alone, whereas the combination of serum-SMRP with cytology led to a sensitivity of 61.5%.Detection of both effusion- and serum-SMRP can contribute to improve the diagnosis of MPM with non-positive cytology. However, the analysis of SMRP in effusion makes it unnecessary to test SMRP in the serum.

Malignant mesothelioma (MM) is a particularly aggressive type of primary tumor, associated with exposure to asbestos, and characterized by high mortality. To date, there is no curative therapy for MM. The receptor anaplastic lymphoma kinase (ALK) was found to be mutated in many cases of cancer and used as a target in biological therapies. We investigated whether this pharmacological treatment could also be applicable to MM.The state of ALK was analyzed by immunohistochemistry and fluorescent in situ hybridization in 63 MM tissue specimens.None of the 63 MM samples showed overexpression or translocation of ALK.Our preliminary data exclude the utility of analysis of the ALK gene in MM and suggest that ALK inhibitor therapy is not applicable to MM.

In this study, immunoglobulin variable (Ig V) region genes, c-myc re-arrangement and sequence and p53 status were analyzed in clones derived from a Burkitt's lymphoma cell line (LAM) in which it was previously demonstrated that Epstein-Barr virus (EBV) infection occurred late during lymphomagenesis. Such evidence was based on the finding that 2 groups of cellular clones, characterized by the same c-myc re-arrangement but different EBV-fused termini, were obtained from the LAM cell line. The Ig V gene sequences were identical for the 2 groups of clones with different EBV-fused termini. The Ig variable heavy (VH) gene sequence displayed a substantial accumulation of point mutations (but no intra-clonal diversification), whereas the productive Ig V lambda (Vλ) gene sequence was virtually unmutated. Studies on the Ig V kappa (Vκ) locus suggested a receptor revision event (with a switch from κ to λ chain production) prior to EBV infection. Likewise, it was determined that the mutations observed in both p53 alleles and in the re-arranged c-myc gene occurred before EBV infection. Based on these findings, we present a model for the various steps of lymphomagenesis. It is proposed that stimulation by an antigen or a superantigen initially favored the clonal expansion and accumulation of other cytogenetic changes, including those involved in receptor editing. These events occurred prior to or during the germinal center (GC) phase of B-cell maturation. Thereafter, possibly upon exit of the cells from the GC, EBV infection occurred, further promoting lymphomagenesis. Int. J. Cancer 88:744–750, 2000. © 2000 Wiley-Liss, Inc.

This report discusses the association of ovarian lymphoma and hydroureteronephrosis managed by laparoscopic surgery and endoscopy., Introduction: Ovarian lymphoma is a rare entity, and hydronephrosis from lymphoma is even rarer. Most reports describe a laparoscopic approach to the disease, but we report a case of hydroureteronephrosis associated with ovarian lymphoma managed completely by mini-invasive techniques. Case Report: A 51-year-old woman was referred to us for back pain and renal colic and computed tomography scan findings of right hydroureteronephrosis and a mass in the right mesorectum and uterosacral ligament. After magnetic resonance imaging was performed, the patient underwent laparoscopic adnexectomy and ureterolysis after ureteroscopy and stenting. Histology results showed diffuse B-cell lymphoma of the ovary occluding the ureter without infiltration. The patient has undergone 6 cycles of chemotherapy. Discussion: This is the first report to describe ovarian lymphoma and hydroureteronephrosis managed completely by laparoscopic surgery and endoscopy. Frequency in clinical practice, differential diagnosis, and endoscopic approach are discussed. The advantages of a multidisciplinary endoscopic team are underlined.

Soluble mesothelin-related peptide (SMRP) is regarded as a biomarker of malignant pleural mesothelioma (MPM). Herein, we compared the diagnostic performances of SMRP in matched pleural effusion (PE-SMRP) and serum (S-SMRP).Diagnosis on pleural biopsies was performed for all patients including 43 with MPM, 23 with non-MPM pleural metastases (MTS) and 36 with benign (BNG) pleural diseases. SMRP was measured by a MesoMark ELISA (Cis-Bio International Gif/Yvette; France).Using the receiver operating characteristic (ROC) analysis, 12.70 and 1.08 nM were detected as cut-off values to optimal discrimination for PE-SMRP and S-SMRP, respectively. PE-SMRP showed a better diagnostic accuracy than S-SMRP in MPM vs. MTS+BNG (area under the ROC curve=81.6 vs. 70.5; sensitivity=69.8% vs. 46.5%; specificity=88.1% vs. 84.7%; diagnostic odds ratio (DOR)=17.1 vs. 4.8). In S-SMRP-negative patients, PE-SMRP maintained an acceptable performance (Sensitivity=47.8%; DOR=8.3; p=0.001), whereas in PE-SMRP-negative patients, S-SMRP performed very poorly (Sensitivity=15.4%; DOR=1.2; p=0.858).PE-SMRP detection has a superior diagnostic accuracy than S-SMRP detection in the diagnosis of MPM.

The unambiguous detection of specific neuronal subtypes is up to now only possible with invasive techniques or optical imaging after genetic modification. High field magnetic resonance imaging (MRI) has the ability to visualize the brain structure and anatomy noninvasively, with high resolution--but missing the cell specific and functional information. Here we present a new tool for neuroimaging with MRI, enabling the selective detection of GABAergic neurons under in vivo conditions. The specific imaging contrast is achieved by a novel paramagnetic contrast agent, which responds to the activity of the enzyme glutamic acid decarboxylase--expressed solely by inhibitory neurons. The relaxivity of the complex is increased upon decarboxylation of two glutamic acid moieties, thus allowing increased water access to the inner and outer coordination spheres of the paramagnetic ion. The mechanism and specificity of activation were proven with tissue lysates and further applied to a differentiation protocol for murine embryonic stem cells. The relaxation enhancement was studied quantitatively and revealed decreased longitudinal relaxation times in the inhibitory neuron samples compared to the naive stem cells in vitro and in vivo. Furthermore, this approach offers not only the discrimination of inhibitory, GABAergic neurons in the brain but also may expand the usefulness of MRI for functional imaging on a cellular level.

In breast cancer (BC), metastases to the central nervous system usually arise in women with advanced disease. Diagnosis of leptomeningeal (LM) metastasis is based on neurological symptoms, imaging studies and cytological detection of malignant cells in the cerebrospinal fluid (CSF). However, often these approaches are not sensitive enough to recognize leptomeninges involvement and subsequently to make a diagnosis of LM carcinomatosis. This study investigated the employment of reverse transcriptase-polymerase chain reaction (RT-PCR) for the human mammaglobin (hMAM) gene in a case of BC with cerebral metastases in which the involvement of the leptomeninges was in doubt.Amplification of hMAM mRNA was performed from CSF cells by RT-PCR.No amplification of hMAM was obtained from the CSF cells.RT-PCR for human mammaglobin mRNA of the CSF in BC patients with brain metastases may aid clinical determination of LM involvement and consequently the choice of the most effective therapy regimens for affected patients.

We investigated the association of circulating tumor cells (CTC) with poor prognostic markers in breast cancer (BC) at time of diagnosis. Peripheral blood (PB) samples from 190 patients with invasive BC, 12 patients with in situ BC, before theraphy and/or surgery, and 330 from patients without BC were tested for CTCs by RT-PCR for human mammaglobin (hMAM). hMAM was expressed only in PB of invasive BC (9.5%) and a significant correlation was found between CTCs with lymph node involvement, tumour size and negative ER. We conclude that CTC detention in invasive BC may be an additional poor prognostic indicator.

HER2 expression or gene amplification is crucial for performing targeted theraphy. We assessed p185HER2 (p185) expression, HER2 amplification and copy number of chromosome 17 centromere (CEP17) in pancreatic adenocarcinoma (PAC). We analyzed 26 tumours at diagnosis, 21 metastatic lymph nodes (mLN) and 38 distant metastases (mTS) by immunohistochemistry and by FISH. p185 overexpression was shown in 4% of primary PAC, 6% of mLN and 29% of mTS. Amplification was found restricted to 10% of the mTS and increased CEP17 in 21% of mTS and 16% of mLN. Overexpression of p185 was present in all amplified cases but in only 36% of increased CEP17. In conclusion HER2 amplification and increased CEP17 were mainly related to mTS or mLN and increased CEP17 was not always linked to overexpression of p185.

So far discordant results regarding the significance of tumour cells circulating in peripheral blood (CTCs) of breast cancer (BC) patients have been reported. Our aim was to evaluate the association of indirect CTC detection by amplification of human mammaglobin (hMAM) gene expression with traditional prognostic markers of clinical outcome in BC at the time of diagnosis.Peripheral blood samples from 190 patients with invasive and 12 patients with in situ BC, before therapy and/or surgery, from 184 patients with benign breast disease and from 146 healthy volunteers were tested for hMAM expression by a nested reverse transcriptase-polymerase chain reaction (RT-PCR) assay. Correlations between CTCs and age at diagnosis, tumour type and size, grading, lymph node involvement, oestrogen and progesterone receptor status, HER-2/neu expression and Ki-67/MIB-1 labelling index were assessed through the odds ratio (OR) point estimates, considering OR2.0 or0.5 as being clinically relevant. ORs and their corresponding 95% confidence limits (95% CL) were obtained by logistic regression analysis.Expression of hMAM was found only in peripheral blood of patients with invasive BC (9.5%) and multivariate logistic regression analysis indicated its association with lymph node involvement (pN1-pN3 vs. pN0, OR=5.6, 95% CL=1.4-22.6; p=0.009), tumour size (pT2-pT4 vs. pT1, OR=2.3, 95% CL=0.6-9.0; p=0.207) and negative ER status (OR=2.5, 95% CL=0.6-10.0; p=0.227).Our data show that CTC detection in invasive BC at the time of diagnosis is associated with poor prognosis and may also be used as an additional prognostic indicator.

Cells display on the outer surface of the plasma membrane a large number of protein thiols that can be reversibly labelled with suitably designed Gd(III)-based contrast agents for cell tracking by MRI.

In Burkitt's lymphoma (BL) cells due to a t(8;14) chromosomal translocation c-myc is often placed in proximity to the Emu enhancer of the Ig locus and upregulated. We demonstrated that in BL cells a peptide nucleic acid (PNA), complementary to intronic Emu sequences (PNAEmuwt), specifically blocks the expression of the c-myc oncogene under the Emu enhancer control and inhibits BL cell growth in culture. Here, we investigated whether PNAEmuwt was also able to block tumor growth in SCID mice inoculated with human BL cell lines. After subcutaneous inoculum in mice BL cells reproducibly form tumors. Both pre-treatment of BL cells with PNAEmuwt before inoculum and chronic intravenous administration of PNAEmuwt to mice already inoculated with BL cells selectively caused increased latency of tumor appearance and decreased final tumor size. Tumors from PNAEmuwt-treated animals showed substantial areas of cell necrosis and of c-myc downregulation. Inhibition of tumor growth was specific and was not observed with PNAEmumut carrying sequence mutations and in BL cell lines where the translocated c-myc is not under the control of the Emu enhancer. These data confirm the potential therapeutic value of PNA targeted to regulatory non-coding regions.

Human infection with the sheep nasal botfly Oestrus ovis is sporadic and is often the consequence of an accidental deposit of the larvae by an adult botfly in the eye. This infestation results in external ophthalmomyiasis that, although a very rare condition, is more common among people living close to farming communities. We report three cases of O. ovis infestation which occurred in Italy in a limited area of La Spezia province (Le Cinque Terre), Italy during summer 2004. None of the patients had contact with wild or farm animals.

Gd(III) chelates are often used as reporting probes in magnetic resonance imaging applications. Their use relies on the assumption that their high stability ensures against the release of free ligand and free Gd(3+) ions, which are both highly toxic to living systems. The presence of free Gd(3+) ions may have a profound effect on the contrast in the MR image as they may form highly hydrated, macromolecular complexes endowed with very high relaxivity thus providing erroneous information on the extent of the contrast effect induced by the given reporting probe. Although the stability of a given complex may be high enough, the complexation step could not have been completed. Therefore any Gd complex preparation has to be carefully checked for the content of free ligand and free metal ions. Herein the currently used procedures that allow an assessment of the amounts of free metal ions and free ligand in a solution of a given Gd complex are described in detail.

Purpose: Few data are available in the literature on chemokine receptor expression and migratory capability of mantle cell lymphoma (MCL) B cells. Information on these issues may allow us to identify novel mechanisms of chemokine-driven tumor cell migration.Experimental Design: The research was designed to investigate: (a) expression of CCR1 to CCR7 and CXCR1 to CXCR5 chemokine receptors; and (b) chemotaxis to the respective ligands in MCL B cells and in their normal counterparts, i.e., CD5+ B cells.Results: Malignant B cells from MCL patients and normal counterparts displayed similar chemokine receptor profiles. MCL B cells were induced to migrate by CXCL12 and CCL19, whereas normal CD5+ B cells migrated to the former, but not the latter chemokine. Overnight culture of MCL B cells and their normal counterparts with CXCL12 cross-sensitized other chemokine receptors to their ligands in some tumor samples but not in CD5+ B cells.Conclusions: CCR7 and CXCR4 ligands may play a key role in tumor cell migration and spreading in vivo. CXCL12 may additionally contribute by sensitizing MCL B cells to respond to the ligands of other chemokine receptors.

The recently introduced new class of contrast agents (CAs) based on chemical exchange saturation transfer (CEST) may have a huge potential for the development of novel applications in the field of MRI. In this work we explored the CEST properties of a series of Lanthanide(III) complexes (Ln = Eu, Dy, Ho, Er, Tm, Yb) with the macrocyclic DOTAM-Gly ligand, which is the tetraglycineamide derivative of DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid). These complexes possess two pools of exchangeable protons represented by the coordinated water and the amide protons. Yb-DOTAM-Gly displays the most interesting CEST properties when its amide N-H resonance (16 ppm upfield H2O signal) is irradiated. Up to 70% suppression of the water signal is obtained at pH 8. As the exchange rate of amide protons is base-catalyzed, Yb-DOTAM-Gly results to be an efficient pH-responsive probe in the 5.5-8.1 pH range. Moreover, a ratiometric method has been set up in order to remove the dependence of the observed pH responsiveness from the absolute concentration of the paramagnetic agent. In fact, the use of a mixture of Eu-DOTAM-Gly and Yb-DOTAM-Gly, whose exchangeable proton pools are represented by the coordinated water (ca. 40 ppm downfield H2O signal at 312K) and amide protons, respectively, produces a pH-dependent CEST effect which is the function of the concentration ratio of the two complexes.