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Background/aim: 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., Materials and Methods: 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)., Results: 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%., Conclusion: 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., (Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.)

Background: 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., Materials and Methods: The state of ALK was analyzed by immunohistochemistry and fluorescent in situ hybridization in 63 MM tissue specimens., Results: None of the 63 MM samples showed overexpression or translocation of ALK., Conclusion: 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.

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 miniinvasive 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.

Background/aim: 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)., Materials and Methods: 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)., Results: 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)., Conclusion: PE-SMRP detection has a superior diagnostic accuracy than S-SMRP detection in the diagnosis of MPM.

Novel contrast agent based systems, which selectively visualize specific cells, e.g., neurons in the brain, would be of substantial importance for the fast developing field of molecular magnetic resonance imaging (MRI). We report here the synthesis and in vitro validation of a Gd(III)-based contrast agent designed to act as an MRI responsive probe for imaging the activity of the enzyme glutamic acid decarboxylase (GAD) present in neurons. Upon the action of the enzyme, the Gd(III) complex increases its hydration sphere and takes on a residual positive charge that promotes its binding to endogenous macromolecules. Both effects contribute in a synergic way to generate a marked relaxation enhancement, which directly reports enzyme activity and will allow activity detection of GAD positive cells in vitro and in vivo selectively.

Soluble mesothelin-related peptide (SMRP) is regarded as an FDA approved biomarker for the diagnosis and monitoring of pleural malignant mesothelioma (MPM). We detected the SMRP levels in pleural effusions (PE) by means of an ELISA and analyzed their diagnostic relevance to differentiate MPM from benign pathology and from non-MPM pleural metastasis. Comparison with cytology in MPM-PE was also performed. We found that SMRP detection in MPM-PE can help the diagnosis of MPM and provide additional diagnostic value to cytology. We concluded that SMRP test may be incorporated into clinical practice of PE from patients suspicious for MPM.

The aim of this study based on the third phase of the architecture of diagnostic research was to assess the sensitivity and specificity of soluble mesothelin-related peptide (SMRP) in pleural exudative effusions (PE) compared to the histology obtained by medical thoracoscopy as the diagnostic gold standard examination. We assessed 104 consecutive thoracoscopies. SMRP concentrations were obtained using an ELISA test. We had 34 mesotheliomas (25 epithelioid and 9 sarcomatoid), 35 pleural metastases, and 35 benign diseases. PE-SMRP were significantly higher in patients with epitheliomorphic mesothelioma (mean ± SD 46.55 ± 44.29 nM) than in patients with sarcomatoid mesothelioma (16.11 ± 25.02 nM; p = 0.061), pleural metastasis (7.52 ± 10.77 nM; p < 0.0001), or benign diseases (5.82 ± 8.86 nM; p < 0.0001). Using ROC curve analysis, PE-SMRP offered an AUC of 0.767 in its ability to differentiate between patients with mesothelioma and all other diagnoses. The diagnostic sensitivity and specificity of PE-SMRP for distinguishing mesothelioma from all other causes of pleural effusion, at a cut-off value of 19.6 nM, were 58.8 and 97.1 %, respectively. PE-SMRP higher than the assumed cut-off of 19.6 nM were observed in 18/25 (72.0%) epitheliomorphic mesotheliomas, 2/9 (22.2%) sarcomatoid mesotheliomas, 5/35 (14.3%) pleural metastases, and 1/35 (2.9%) benign diseases. We conclude that PE-SMRP adds some clinical information in the work-up of patients with a PE of unknown origin: (1) thoracoscopy should always be done in patients with a positive mesothelin; (2) a negative mesothelin does not exclude a malignant disease.

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 naïve 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., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Background: human mammaglobin (hMAM) expression has been reported in pleural effusions (PE). The aim of this study was to assess the clinical relevance of hMAM mRNA in PE from patients who underwent thoracoscopy.
, Material and Methods: A total of 288 patients with PE were studied, 155 of which were diagnosed with malignant and 133 with non-malignant diseases by thoracoscopy. Cells from PE were analyzed by nested hMAM RT-PCR. Statistical analyses were performed to evaluate the diagnostic performance parameters (DPP), the association between hMAM expression and benign or malignant status and the relative risk of cancer for patients with negative thoracoscopy showing hMAM positivity.
, Results: hMAM mRNA was found in 68/288 (23.6%) PE samples of which 51 were from the 155 patients diagnosed with malignant diseases and 17 were from the 133 patients diagnosed with non-malignant diseases. A significant correlation between hMAM expression and malignancy was found (OR=3.04) and the DPP were as follows: sensitivity=32.9%, specificity=87.2%, accuracy=58.0%, positive predictive value=75.0% and negative predictive value=52.7%. Among the patients with negative thoracoscopy (n=133), 5/17 (29.4%) hMAM-positive patients had or developed a tumor during the 18-month follow up period, as compared to 10/116 (8.6%) hMAM-negative patients (relative risk of 4.6 for developing a malignancy).
, Conclusion: These findings suggest a possible application of hMAM RT-PCR detection in PE as to identify a false-negative thoracoscopy in non-specific pleuritis.

This work aims at assessing the in vitro potential of paramagnetic pH sensitive liposomes as imaging tools for visualizing drug-delivery and release processes by Magnetic Resonance Imaging (MRI). pH sensitive liposomes (pSLs) were formulated using the fusogenic phospholipid 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE), the membrane stabilizer D-α-tocopherol-hemisuccinate (THS), and were loaded with several paramagnetic complexes including the clinically approved Gadoteridol (marketed as ProHance™). The proposed formulation allows the fast and full release of Gadoteridol at pH 5.5. The leakage of the imaging reporter from the vesicles was associated with a relaxivity enhancement that allowed its visualization by MRI. It was observed that the release mechanism implies the protonation of the THS basic sites that leads to vesicle aggregation, thus enabling the expression of the fusogenic property of POPE. Attempts for improving the MRI properties of pSLs were pursued through the encapsulation of imaging agents with higher relaxivity than Gadoteridol, but it was observed that the release kinetic can be significantly affected by the probe size. Aiming at preparing stealth pSLs, PEG chains were conjugated to the external surface of the vesicles via cleavable disulphide bridges. Such nanomedicines do not release their content at acidic pH as long as the coating polymer is not removed from the surface. The results obtained suggest that the liposomal formulation investigated in this work has the potential for visualizing drug-delivery and release processes by in vivo MRI preclinical studies., (Copyright © 2011 Elsevier B.V. All rights reserved.)

A novel class of paramagnetic liposome-based systems acting as dual T(1) and CEST (1)H-MRI contrast agents is described. The vesicles contain a shift reagent in the aqueous core and a Gd-complex on the external surface conjugated through a biodegradable linker. As such, the probe can generate T(1) contrast only, but after the cleavage and removal of the Gd-coating, the CEST contrast is switched on., (© The Royal Society of Chemistry 2011)

Background: 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., Materials and Methods: Amplification of hMAM mRNA was performed from CSF cells by RT-PCR., Results: No amplification of hMAM was obtained from the CSF cells., Conclusion: 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.

Four novel MRI Gd(III)-based probes have been synthesized and evaluated for their labeling properties on cultured cell lines K562, C6, and B16. The labeling strategy relies upon the fact that cells display a large number of reactive exofacial protein thiols (EPTs) that can be exploited as anchorage points for suitably activated MRI probes. The probes are composed of a Gd(III) chelate (based on either DO3A or DTPA) connected through a flexible linker to the 2-pyridyldithio chemical function for binding to EPTs. GdDO3A-based chelates could efficiently label cells (up to a level of 1.2 x 10(10) Gd(III) atoms/cell), whereas GdDTPA-based chelates showed poor or no cell labeling ability at all. Among the GdDO3A based compounds, that having the longest spacer (compound GdL1A) showed the best labeling efficacy. The mechanism of EPT mediated cell labeling by GdL1A involves probe internalization without sequestration of the Gd(III) chelate within subcellular structures such as endosomes.

Background: 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., Patients and Methods: 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 OR >2.0 or <0.5 as being clinically relevant. ORs and their corresponding 95% confidence limits (95% CL) were obtained by logistic regression analysis., Results: 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)., Conclusion: 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.

As was reported that human mammaglobin (hMAM) may be expressed in malignant pleural effusions (PEs), we investigated the relevance of hMAM reverse-transcriptase polymerase chain reaction (RT-PCR) for their diagnosis and determination of primary origin. Two hundred and twenty-eight malignant (132 male, 96 female) and 185 benign (132 male, 53 female) PEs were investigated. Statistical analyses evaluated the diagnostic performance parameters in all PEs and in cytologically negative malignant PEs, the association between hMAM and benign or malignant status by the direct index of correlation [diagnostic odds ratio (DOR)], chi test, and P value (P). In addition, the discriminative diagnostic power of hMAM expression, independently in breast cancer, lung cancer (LC), malignant mesothelioma (MM), and other cancers was evaluated. In the entire patient population, hMAM was detected in 45.6% and 5.4% of malignant and benign PEs, respectively, in the male group in 41.7% and 4.5% and in the female group in 51.0% and 7.5% of malignant and benign PEs, respectively. A statistically significant correlation between hMAM and malignancy was found in the entire population (DOR=14.68, P<0.001) and in the male (DOR=15.00, P<0.001) or female (DOR=12.77, P<0.001) groups. hMAM RT-PCR increased the diagnostic rate of malignant PEs as it allowed us to detect as malignant 32.1% of cytologically negative PEs. In female patients the positivity of hMAM indicated with higher probability (50.8%) the origin of PEs from breast cancer but lower probability from LC (17%), MM (9.4%), or other cancers (15.1%), whereas in male patients it indicated with similar probability (about 40%) the origin from LC or MM. Our results suggest that hMAM RT-PCR may provide information both in the diagnosis of PE and in the search for the primary site of neoplasia, either in male or female patients.

The in vitro validation of a new pH mapping MRI method based on a supramolecular poly-beta-cyclodextrin-19F-Gd adduct is reported.

The WT1 gene plays a crucial role in urogenital and gonadal development. Germline WT1 alterations have been described in a wide spectrum of pathological conditions, including kidney diseases, genital abnormalities and Wilms tumor (WT), frequently occurring in combination. We report on a novel WT1 nonsense mutation (c.1105C>T), introducing a premature stop codon in exon 8 (p.Q369X), in a young XY male patient who presented with bilateral cryptorchidism, nystagmus, mild proteinuria and WT, but no sign of severe nephropathy. Although the majority of congenital urogenital abnormalities are not due to constitutional defects of the WT1 gene, our findings provide a rational for considering WT1 mutational analysis as one of the screening options in newborns with congenital defects of the urogenital tract due to the associated high risk of WT.

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.

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.

Leptomeningeal (LM) carcinomatosis is an increasing clinical complication in patients with advanced breast cancer (BC). The LM carcinomatosis diagnostic procedures rely mainly on cerebrospinal fluid (CSF) cytology, although both the amount of CSF and the number of malignant cells remain limiting factors. Therefore, efforts should be made to design new highly sensitive diagnostic tools to detect malignant cells in CSF of BC patients with LM carcinomatosis. In this study, the human Mammaglobin (hMAM) mRNA amplification by RT-PCR was employed to detect metastatic cells in CSF and thus, to diagnose LM carcinomatosis in a BC patient. Our data demonstrate that hMAM transcripts are expressed in the CSF of a BC patient with LM carcinomatosis, hence making RT-PCR for hMAM a potentially suitable test to identify occult BC cells in the brain.

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.

Efficient routes to DOTA-monoamide ligands bearing amino, hydroxyl, aldehyde and maleimido groups are described. These functional groups, which can be spaced at will from the coordination cage, will readily react with suitable groups of targeting moieties. Bioconjugates obtained in this way can be used for diagnostic imaging and therapeutic applications.

The present study investigates the diagnostic significance of human mammaglobin (hMAM) mRNA expression in pleural effusions (PE) from breast cancer (BC) patients. Two hundred and fifty PE samples, including 32 from patients who had diagnosis of BC, 116 from patients with other cancers, and 102 from patients with benign diseases, were subjected to nested reverse-transcription polymerase chain reaction (RT-PCR) for hMAM, and the results were compared with conventional cytology. hMAM was found expressed in 76/250 (30.4%) total PE and in 23/28 (sensitivity of 82.1%) of the PE subgroup owing to metastasis from BC. The specificity for hMAM detection method was 75.7%, whereas accuracy, positive predictive value, and negative predictive value were 76.4%, 30.3%, and 97.1%, respectively. hMAM was also detected in 46/116 (39.6%) PE specimens from other types of cancer and in 7/102 (6.8%) from benign diseases. Comparative analysis of RT-PCR and cytology showed that 14 PE samples from metastatic BC (50%) were positive by both PCR and cytology, 9 (32.1%) were positive only by PCR and 5 (17.9%) were negative by both tests, whereas no cases were found of positive cytology with negative PCR. RT-PCR increased sensitivity of BC effusion detection to 32.1% (McNemar test, P=0.004). We demonstrated that RT-PCR for hMAM test was more sensitive than cytomorphology suggesting that, although hMAM is not BC specific, it may be useful in adjunct to cytology for the routine screening of malignant BC effusions.

Improved LIPOCEST MRI contrast agents with highly shifted intraliposomal water protons were prepared by entrapping neutral polynuclear Tm(III)-based paramagnetic shift reagents in phospholipidic vesicles.

A novel Gd(III) complex with a modified DO3A-like chelating cage has been synthesized and characterized as a candidate contrast agent responsive to the concentration of free thiols in tissues (essentially represented by reduced glutathione, GSH). The novel compound (called Gd-DO3AS-Act) bears a flexible linker ending with a 2-pyridyl-dithio group, that can promptly react with free thiols (XSH) to form mixed disulfides of the form Gd-DO3AS-SX. Compound Gd-DO3AS-Act is characterized by a millimolar relaxivity as high as 8.1 mM(-1) s(-1) (at 20 MHz, 25 degrees C and pH 7.4). Upon reaction with GSH, the Gd-DO3AS-SG covalent adduct is formed and the millimolar relaxivity drops to 4.1 mM(-1) s(-1). Such a decrease in relaxivity is explained on the basis of the formation of an intramolecular coordinative bond between one of the glutathionyl carboxyl groups and the Gd(III) centre, lowering the hydration state of the paramagnetic centre. (1)H-NMR dispersion profiles together with (17)O-NMR transverse relaxation time versus temperature profiles confirm that the hydration of the Gd(III) centre is strongly reduced ongoing from Gd-DO3AS-Act to the Gd-DO3AS-SG adduct. The relaxivity difference brought about by the reaction of Gd-DO3AS-Act with GSH can be enhanced up to 60% in the presence of poly-beta-cyclodextrin.

A novel procedure for making the pH responsiveness of a Gd-based probe independent of its concentration has been developed. The method is based on a ratiometric approach and requires the measurement, at magnetic fields higher than 1 T, of the paramagnetic contribution to the longitudinal and the transverse NMR relaxation rates of water protons in the presence of a Gd-based probe with a rotational mobility in the nanosecond time scale. The method has been tested and validated in vitro by using a macromolecular adduct based on the polyornithine backbone, whose rotational mobility is pH dependent. The results reported in this communication improve substantially the diagnostic potential of MRI responsive agents.

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., (Copyright 2006 John Wiley & Sons Ltd.)

Human mammaglobin (hMAM) has recently been recognized as a breast associated glycoprotein. Although the biological role of hMAM is unknown, it has been previously reported that hMAM gene expression is a marker of low biological and clinical aggressiveness of breast cancer (BC). In this study, 148 cases of BC tissues were investigated for hMAM mRNA expression by reverse transcriptase-polymerase chain reaction (RT-PCR). In order to evaluate its prognostic value, hMAM was correlated with age of patients, type and size of tumor, nodal stage, histologic grade, c-erbB-2 over expression, Ki67 labelling index, estrogen receptor (ER) status and progesterone receptor (PGR) status. Fisher's exact test was used to examine the association between different parameters and hMAM. hMAM was expressed in 138/148 (93%) of BC tissues examined. Among the 10 hMAM negative cases, 8 were invasive ductal carcinomas (microscopically higher G3 grade) and 2 infiltrating lobular carcinomas. We found a significant association (p = 0.020) between absence of hMAM mRNA and G3 histologic grade but not with any other prognostic parameters studied. The present study indicates that lack of hMAM expression is restricted to the BC with G3 grading. Further studies are needed to clarify the biological basis and the clinical significance of our results.

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.

Occult carcinoma cells in peripheral blood of breast cancer (BC) patients is generally associated with poor disease prognosis. Reverse transcriptase-polymerase chain reaction (RT-PCR) is a sensitive method for revealing rare circulating cancer cells. The target mRNA must be carefully chosen, as it must be expressed only by malignant cells. In this study, we developed a nested RT-PCR assay for mammaglobin (hMAM) and assessed both its specificity and its sensitivity in the detection of cancer cells in peripheral blood of BC patients. hMAM mRNA was detected by RT-PCR in 156/165 (95%) of fresh BC tissues analyzed. All samples from 66 healthy blood donors and 151 patients with benign breast disease were hMAM negative as assessed by nested RT-PCR. In contrast, hMAM was detected in 16/137 (12%) of peripheral blood samples deriving from BC patients: 0/9 in stage 0, 1/50 (2%) in stage I, 3/33 (9%) in stage II, 1/18 (5%) in stage III and 11/27 (41%) in stage IV. Using nested RT-PCR, we were able to amplify hMAM transcript of one tumour cell/10(6) normal cells. Our data demonstrate that hMAM mRNA detection by RT-PCR is a specific assay potentially suitable for identification of occult cancer cells in peripheral blood of BC patients.

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.

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-18Ralpha and beta 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-18Ralpha and beta transcripts were expressed in the three B-cell subsets. IL-18Ralpha was detected on the surface of naive, GC, and memory B lymphocytes, and IL-18Rbeta 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.

Three novel heptacoordinated Ln(III) complexes (Ln = Gd and Yb) have been synthesized and investigated by (1)H NMR spectroscopy. These complexes contain two stereogenic centers, one associated with a deltadeltadeltadelta or lambdalambdalambdalambda conformation of the ethylenediamine moieties in the tetraazamacrocycle and the latter arises from the orientation (Delta or Lambda) of the coordinating arms. Evidence has been gained for the occurrence of a fast exchange between all the possible conformers. Upon addition of several (S)-alpha-hydroxy-carboxylate substrates, the formation of stable ternary adducts has been obtained. Their (1)H NMR spectra are consistent with the presence of two diastereoisomers differing in the conformation adopted by the macrocyclic ligand wrapping the lanthanide(III) ion. The interaction leading to the formation of the ternary complexes is enantioselective depending on the hydrophilicity of the alpha-hydroxy-carboxylate.

In this report, we describe a case of poorly differentiate myoepithelial cell rich carcinoma in with morphological findings of large poligonal nests with festoon-like pattern sometimes showing central necrosis, reminiscent of a comedo-like pattern and numerous mitoses. Immunohistochemical staining shows positive reaction for cytokeratin AE/1, CAM 5.2, 34 beta E12, vimentin, smooth muscle actin, EMA, S100 protein and oncogene cERB.b2 and negative for estrogen, progesterone, GFAP and chromogranin. Moreover, this carcinoma show the expression of the mammaglobin mRNA, a highly specific marker of breast epithelial cells that it is not expressed in all breast carcinoma.

A paramagnetic Yb(III) complex bearing six exchangeable amide protons, [Yb(MBDO3AM)](3+), has been investigated with the aim of developing a MRI-CEST (chemical exchange saturation transfer) contrast agent responsive to the concentration of L-lactate. The complex binds the substrate quantitatively to yield [Yb(MBDO3AM)L-lactate](2+). The exchange between the free and the L-lactate-bound complex is slow on the NMR time scale, and the resonances of their corresponding amide protons are sufficiently separated (more than 10 ppm) to allow their selective irradiation. Therefore, the CEST properties of the two forms can be independently assessed. In turn, the resulting saturation transfer to the bulk water signal is dependent on the L-lactate concentration.

The rate of axial water exchange in well-defined series of lanthanide complexes depends on the extent of second sphere hydration which is determined by complex hydrophobicity and the nature of the lanthanide ion and its counter-ion.

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., (Copyright 2002 Wiley-Liss, Inc.)

Three novel Mn(II) complexes bearing benzyloxymethyl functionalities are reported and their ability to enhance water (1H and 17O) relaxation times is investigated in detail. Two of them contain one coordinated water molecule and display relaxivity values only slightly smaller than those shown by the most clinically used contrast agents (e.g. [Gd(DTPA)(H2O)]2-). Moreover, in these Mn(II) chelates the exchange rate of the coordinated water is ca. one order of magnitude higher if compared to the exchange rates previously reported for Gd(III) complexes with octadentate ligands. The occurrence of such fast exchange rates of the coordinated water is exploited in the formation of macromolecular adducts with human serum albumin to attain systems displaying relaxivity values in the upper range of those so far reported for analogous Gd(III) systems. These results strongly support the view that Mn(II) complexes, in spite of the lower effective magnetic moment, can be considered as viable alternatives to the currently used Gd(III) complexes as contrast agents for MRI applications.

The results reported in this work show that tightly assembled adducts formed by trisubstituted GdIII complexes and a beta-CD multimer (Poly-beta-CD, d.p. ca. 12) may represent very interesting candidates for novel MRI applications wherein a high number of paramagnetic ions endowed with high relaxivity (per GdIII ion) are necessary. The relaxivities found for the paramagnetic adducts represent a remarkable step forward on the relaxivity scale. However, a detailed investigation of the determinants of the relaxation enhancement in these systems shows that their relaxivities are still limited by a nonoptimal tauR and a relatively long exchange lifetime of the coordinated water(s). Moreover, the exchange rate of the water molecule(s) coordinated to the GdIII ion further decreases upon binding to the Poly-beta-CD. It is suggested that this finding is related to the structural properties of the supramolecule, which brings a high density of hydroxyl groups into the proximity of the "guest" complexes, and this yields an overall reinforcement of the hydrogen-bonding network involving the coordinated water(s). On the other hand, such a tight arrangement appears responsible for an enhanced contribution to the observed relaxivity arising from water molecules in the second coordination sphere of the metal center.

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 (V(H)) gene sequence displayed a substantial accumulation of point mutations (but no intra-clonal diversification), whereas the productive Ig V lambda (V(lambda)) gene sequence was virtually unmutated. Studies on the Ig V kappa (V(kappa)) locus suggested a receptor revision event (with a switch from kappa to lambda 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., (Copyright 2000 Wiley-Liss, Inc.)

Cyclin D1 is a G1 cyclin participating in the control of cell cycle progression through interaction with the retinoblastoma gene product (pRB). The overexpression of positive regulators (such as cyclin D1) has been reported in a variety of neoplasms, but their role in thyroid tumorigenesis is yet to be established. In our series of 54 thyroid carcinomas, cyclin D1 overexpression (detected by both immunohistochemistry and by Northern blotting) was correlated with prognostic variables, proliferative activity and pRB. Cyclin D1 overexpression was observed in 35% of thyroid carcinomas with a significantly higher expression of this cyclin in neoplastic tissues than in matched normal parenchyma. In well-differentiated carcinomas, the cyclin D1 mRNA overexpression was inversely correlated with nodal status (p = 0.03), while the protein product was higher in tumors from patients less than 40 than patients over 40 years of age. Inversely, there was no significant correlation with gender and tumor status, pRB and with proliferative activity.

Bisalkylation of suitably protected L-glutamic acid and L-lysine derivatives with tert-butyl N-(2-bromoethyl)iminodiacetate 2, followed by deprotection of the omega functional group affords N, N-bis[2-[bis[2-(1, 1-dimethylethoxy)-2-oxoethyl]amino]ethyl]-L-glutamic acid 1-(1, 1-dimethylethyl) ester 4 and N2,N2-bis[2-[bis[2-(1, 1-dimethylethoxy)-2-oxoethyl]amino]ethyl]-L-lysine 1,1-dimethylethyl ester 7. Such compounds feature a carboxylic or an amino group, respectively, which are available for conjugation with a suitable partner via formation of an amide bond. The conjugates, which can be prepared in this way, contain a chelating subunit in which all five acetic residues of DTPA are available for the complexation of metal ions. Direct bisalkylation of glycine with 2 promptly gives N, N-bis[2-[bis[2-(1,1-dimethylethoxy)-2-oxoethyl]amino]ethyl]glycine 11. The latter allows to achieve conjugates in which the central acetic group of DTPA is selectively converted into an acetamide.

The efficiency and selectivity of tumour targeting by several tetra-n-propylporphycene (TPPn) and tetrakis(methoxyethyl)porphycene (TMPn) derivatives have been studied by administering 3.76 micromol/kg of aqueous or liposomal porphycene formulations to BALB/c mice bearing an i.m. implanted MS-2 fibrosarcoma. These 2 parameters have been studied as a function of the type of substituents linked to the 9-position of the macrocycle by amide, ester or ether functional groups. The pharmacokinetic properties appear to be controlled mainly by the degree of porphycene hydrophobicity, as evaluated by measuring their retention times in a C 18 column for HPLC. Thus, the post-injection time (T50) at which the porphycene concentration in the plasma decreases to 50% of the initial value ranged from a few minutes for the less hydrophobic to several hours for the more hydrophobic porphycenes. An increase in hydrophobicity also was accompanied by an enhanced efficiency and selectivity of tumour targeting. The less hydrophobic porphycenes showed a maximum tumour uptake of 0.5-2 nmol/g of tissue at 10-20 min after administration with a tumour/peri-tumoural concentration ratio around 2-3, while those with higher hydrophobicity reached tumour concentrations of 7-8 nmol/g at 24-48 hr after administration with concentration ratios higher than 20.

A case of an important intracranial hemorrhage after a low dose (approx. 500,000 UI) of streptokinase in a 60 year-old woman suffering from myocardial infarction is presented. Clinical, electrocardiographic, echocardiographic, lab and tomographic findings are described. The authors suggest a pharmacokinetic mechanism which could be responsible of a "paradox effect" (a powerful and dangerous effect of the drug when given in low dose) and they wonder whether in case of allergic reactions should it be better not to stop the infusion of the thrombolytic drug and be more liberal with the "symptomatic" drugs. Tha patient is still alive and the clinical conditions slowly progressing.

In the literature, the mitral valve prolapse and bicuspid aorta have been widely discussed as isolated cases or in association with other congenital heart pathologies or systematic illnesses. Nevertheless, they have not been documented contemporarily in the same clinical case. The following case describes a healthy, young, asymptomatic athlete, who has a double valvular heart failure. The defect is occasionally evident during transthoracic echocardiographic examination. The role of echocardiography is stressed taking into consideration the natural lineage and unfavourable reciprocal effect on cardiac hemodynamics, omitting relative implications of familial pathologies. This method is suggested as the means of suitable evaluation for athletes. In fact, this is the best technique to reveral the most precocious modification of cardiac hemodynamic. Consequently, echocardiography allows us to guide and monitor the most appropriate therapy.

Four cases of gastric carcinoma are described that are associated with an osteoclast-like giant cell (OGC) stromal component. The patients were all middle-aged men (range 53-63 years). Microscopically, the tumors were characterized by a bland cytologic appearance, and an either solid or cribriform pattern. Osteoclast-like giant cells were found adjacent to, or intimately intermixed with, the neoplastic cells in the primary gastric masses and in the lymph nodal metastases and were often associated with lymphocytes, histiocytes, and desmoplastic stroma. By immunohistochemistry, mononuclear cells and OGCs showed diffuse positivity for alpha-1-antichymotrypsin, alpha-1-antitrypsin, and CD68. Neoplastic cells that were positive for keratin and CEA, also showed reactivity for vimentin and the latent membrane protein of Epstein-Barr virus in one case. At follow-up, three patients had died at 13, 15, and 24 months after diagnosis, and one is still alive, without evidence of disease, after 120 months. This report describes a novel variant of gastric carcinoma with distinctive and histologic features.

Raynaud's phenomenon is in most cases idiopathic, but it may appear as secondary to other diseases. Since clinical evaluation is often inadequate to distinguish between these two forms, simple noninvasive methods are needed for an early diagnosis of the secondary form. The authors used nailfold capillaroscopy to study 58 patients with Raynaud's phenomenon--44 apparently idiopathic and 14 secondary to connective tissue diseases. In all patients affected by secondary Raynaud's phenomenon nailfold capillaroscopy confirmed the presence of an abnormal pattern with typical systemic sclerosis alterations in 5 cases. Twenty-one (48%) of the other 44 patients presented a normal capillaroscopic pattern, 15 (34%) had alterations compatible with long-term idiopathic Raynaud's phenomenon, and 8 (18%) showed abnormal capillaroscopic patterns, 2 of which were identified as the sclerodermic pattern. Nailfold capillaroscopy distinguishes between primary and secondary Raynaud's phenomenon and may be used to identify patients presenting no evidence of underlying disease who could be at risk of developing connective tissue disease.

Background: Risk of basal cell carcinoma (BCC) recurrence appears to be related to tumor location, tumor size, treatment modality, radicalness of excision, and histologic type. Recurrences of BCC on the trunk and extremities are rare, with 97% of all recurrent lesions being located in the head and neck region., Objective: This report concerns a man who had a BCC of the back removed with electrodesiccation and curettage in 1980 at the age of 36. In the following 10 years the patient experienced five recurrences and finally developed infiltration of the thoracic spine that precluded further attempts at radical excision of the neoplasm., Methods and Results: The pathology of the BCC in our patient revealed a number of histologic features that have been associated with aggressive behaviour. These include an infiltrative growth pattern characterized by an irregular and acute tapered profile of the tumor cell groups and fibroblast-rich stroma, infiltrating invading tumour edges, poorly peripheral palisading, nuclear pleomorphism, and perineural invasion., Conclusion: Basal cell carcinomas that occur in sunlight-protected areas of the trunk of middle-aged individuals can occasionally exhibit aggressive and highly invasive features as well as a marked tendency for local uncontrollable recurrences. The prognosis for patients with this variety of BCC is poor, especially when progressive infiltration of deep tissues does not permit, as in our case, any possible further radical surgical procedure.

The histopathologic features of tissue specimens from 16 patients with acute Epstein-Barr virus-induced infectious mononucleosis, which was confirmed by clinical and serologic methods, are described. The clinical course was usually self-limited (14 patients), but it resulted in the death of two patients, one of whom (patient with renal transplantation) was immunosuppressed. Each lymphoid tissue specimen, including those obtained from the lymph nodes (n = 9), tonsils (n = 5), spleen (n = 1), and appendix (n = 1), showed a nonuniform expansion of nonfollicular areas by a polymorphous population of lymphocytes, including transformed lymphocytes and immunoblasts. In situ hybridization demonstrated Epstein-Barr virus-infected lymphocytes in four of eight tissue specimens that were studied. Other histologic features included Reed-Sternberg-like cells, plasma cells, histiocytes, frequent mitoses, abundance of postcapillary venules, and necrosis. These histologic features should suggest a diagnosis of infectious mononucleosis rather than other processes, either benign or malignant, that can mimic it.