Rosanna Nano

Human Leukocyte Antigen and Antigen Processing Machinery Component Defects in Astrocytic Tumors

Purpose: To determine the frequency of abnormalities in human leukocyte antigen (HLA) and antigen processing machinery (APM) component expression in malignant brain tumors. This information may contribute to our understanding of the immune escape mechanisms used by malignant brain tumors because HLA antigens mediate interactions of tumor cells with the hosts immune system. Experimental Design: Eighty-eight surgically removed malignant astrocytic tumors, classified according to the WHO criteria, were stained in immunoperoxidase reactions with monoclonal antibody recognizing monomorphic, locus-specific, and allospecific determinants of HLA class I antigens, β2-microglobulin, APM components (LMP2, LMP7, TAP1, TAP2, calnexin, calreticulin, and tapasin), and HLA class II antigens. Results: HLA class I antigens were lost in ∼50% of the 47 glioblastoma multiforme (GBM) lesions and in ∼20% of the 18 grade 2 astrocytoma lesions stained. Selective HLA-A2 antigen loss was observed in ∼80% of the 24 GBM lesions and in ∼50% of the 12 grade 2 astrocytoma lesions stained. HLA class I antigen loss was significantly (P < 0.025) correlated with tumor grade. Among the APM components investigated, tapasin expression was down-regulated in ∼20% of the GBM lesions analyzed; it was associated, although not significantly, with HLA class I antigen down-regulation and tumor grade. HLA class II antigen expression was detected in ∼30% of the 44 lesions analyzed. Conclusion: The presence of HLA antigen defects in malignant brain tumors may provide an explanation for the relatively poor clinical response rates observed in the majority of the T cell–based immunotherapy clinical trials conducted to date in patients with malignant brain tumors.

Diagnostic Potential of Autofluorescence for an Assisted Intraoperative Delineation of Glioblastoma Resection Margins

Abstract The intrinsic autofluorescence properties of biological tissues can be affected by the occurrence of histological and biochemical alterations induced by pathological processes. In this study the potential of autofluorescence to distinguish tumor from normal tissues was investigated with the view of a real-time diagnostic application in neurosurgery to delineate glioblastoma resection margins. The autofluorescence properties of nonneoplastic and neoplastic tissues were analyzed on tissue sections and homogenates by means of a microspectrofluorometer, and directly on patients affected by glioblastoma multiforme, during surgery, with a fiber-optic probe. Scan-microspectrofluorometric analysis on tissue sections evidenced a reduction of emission intensity and a broadening of the main emission band, along with a redshift of the peak position, from peritumoral nonneoplastic to neoplastic tissues. Differences in both spectral shape and signal amplitude were found in patients when the glioblastoma lesion autofluorescence was compared with those of cortex and white matter taken as healthy tissues. Both biochemical composition and histological organization contribute to modify the autofluorescence emission of neoplastic, with respect to nonneoplastic, brain tissues. The differences found in the in vivo analysis confirm the prospects for improving the efficacy of tumor resection margin delineation in neurosurgery.

Clinical lessons from the first applications of BNCT on unresectable liver metastases.

After a long series of studies on the effects of neutron irradiation of 10 B loaded neoplastic cells both in culture and in animal experiments, we started the clinical application of BNCT on humans affected by liver metastases of a radically resected colon adenocarcinoma. The procedure we adopted includes a first surgical phase, with hepatectomy; a radiotherapeutic phase, in which the isolated liver, washed and chilled, is extracorporeally irradiated with thermal neutrons; and then a second surgical phase for the reconnection of the liver to the patient. Until now two patients have been subjected to the BNCT treatment. The first one survived 44 months with a good quality of life, and died because of diffuse recurrences of his intestinal tumour. The second patient had the same early perioperative course, but after 33 days a worsening of a dilatative cardiomyopaty, from which he was suffering, determined a cardiac failure and eventually death. This clinical experience, although limited, has shown that extracorporeal neutron irradiation of the liver is a feasible procedure, able to ensure the complete destruction of liver metastases and a possible long lasting survival. In our patients neutron irradiation caused massive cellular necrosis highly specific to tumour cells, whereas normal cells were mostly spared. Nevertheless, the impact of such a traumatic operation on the patients organism must be taken into account. Finally, we have to be aware that the fight against tumour rarely leads to a complete victory. We now have an innovative weapon which is both powerful and partly unsettled: it must be refined and above all used.

Sympathectomy or Doxazosin, But Not Propranolol, Blunt Myocardial Interstitial Fibrosis in Pressure-Overload Hypertrophy

The adaptive changes that develop in the pressure-overloaded left ventricular (LV) myocardium include cardiomyocyte hypertrophy and interstitial fibrosis. Although the former is known to depend to a sizeable extent on sympathetic (over)activity, little information exists whether the same applies to the latter, ie, whether excess catecholamine exposure contributes to the imbalance between collagen deposition by fibroblasts and degradation by matrix metalloproteases (MMPs), eventually leading to LV collagen accumulation. Sprague-Dawley rats were subjected to abdominal aortic banding (B) or sham operation (S) and treated with &bgr;-blockade (Bb, oral propranolol, 40 mg/kg per day), chemical sympathectomy (Sx, 6-hydroxydopamine, 150 mg/kg intraperitoneal twice per week) or vehicle (Vh). Ten weeks later, systolic blood pressure, LV weight, collagen abundance (computer-aided histology), zymographic matrix metalloproteinase (MMP)-2 activity and its specific tissue inhibitor concentration (TIMP-2) were measured. Both sympathectomy and &bgr;-blockade failed to attenuate the banding-induced blood pressure elevation but significantly attenuated the attendant LV hypertrophy. As expected, pressure-overload hypertrophy was associated with interstitial fibrosis (collagen: 4.37±1.23% BVh versus 1.23±0.44% SVh, P<0.05), which was abolished by sympathectomy (2.55±1.31%, P=not significant versus SSx) but left unchanged by &bgr;-blockade (4.11±1.23%, P<0.05 versus both SBb and BSx). &bgr;-blockade, but not sympathectomy, was also associated with an increased TIMP-2/MMP-2 ratio (P<0.05), indicating reduced interstitial collagenolytic activity. In separate groups of banded and sham-operated rats, treatment with the &agr;-receptor blocker doxazosin (10 mg/kg per day) displayed similar antifibrotic and biochemical effects as sympathectomy. Thus in the course of experimental pressure overload, the sympathetic nervous system plays a major pro-fibrotic role, which is mediated via &agr;-adrenergic but not &bgr;-adrenergic receptors.

Neutron autoradiography imaging of selective boron uptake in human metastatic tumours

The ability to selectively hit the tumour cells is an essential characteristic of an anti-tumour therapy. In boron neutron capture therapy (BNCT) this characteristic is based on the selective uptake of (10)B in the tumour cells with respect to normal tissues. An important step in the BNCT planning is the measurement of the boron concentration in the tissue samples, both tumour and healthy. When the tumour is spread through the healthy tissue, as in the case of metastases, the knowledge of the different kinds of tissues in the sample being analysed is crucial. If the percentage of tumour and normal tissues cannot be evaluated, the obtained concentration is a mean value depending on the composition of the different samples being measured. In this case an imaging method that could give information both on the morphology and on the spatial distribution of boron concentration in the sample would be a fundamental support. In this paper, the results of the boron uptake analysis in the tumour and in the healthy samples taken from human livers after boron phenylalanine (BPA) infusion are shown; boron imaging was performed using neutron autoradiography.

Thyroid-related autoantibodies and celiac disease: A role for a gluten-free diet?

An association between celiac disease and other autoimmune disorders—such as insulin-dependent diabetes, Addisons disease, systemic lupus erythematous, rheumatoid arthritis, alopecia areata, and autoimmune endocrine diseases—has been described. The aim of this study was to evaluate the prevalence of celiac disease in 100 patients with autoimmune thyroid disease. Moreover, the monitoring of patients with concomitant celiac and autoimmune thyroid diseases, after a gluten-free diet or a gluten-containing diet, can give important insights into the effect of dietary habits in thyroid autoantibodies modulation. In our study, the prevalence of celiac disease in patients with autoimmune thyroid disease was 2%. In these two celiac patients, the serologic markers became undetectable 6 months after beginning a gluten-free diet. However, thyroid autoantibodies did not positively correlate with dietary habits.

STYRENE-INDUCED ALTERATIONS IN THE RESPIRATORY TRACT OF RATS TREATED BY INHALATION OR INTRAPERITONEALLY

Although exposure to styrene occurs primarily via inhalation, the action of this agent on the respiratory tract has scarcely been investigated. This article describes morphological and biochemical changes occurring in the respiratory tract of rats after either inhalation of styrene vapors (300 ppm, 6 h/d, 5 d/wk, for 2 wk) or systemic (ip) treatment with 40 or 400 mg/kg styrene for 3 consecutive days. Electron microscopy analysis showed diffuse cell damage involving the tracheal, bronchiolar, and alveolar epithelium. In the tracheal epithelium, several cell types were affected. Ciliated cells presented vacuolation, detachment of cilia, blebbing of the apical cytoplasm, and compound cilia. Most secretory cells showed scant secretory granules and blebbings. Dense bodies and fibrillary inclusions were seen in intermediate and basal cells. Styrene also caused alterations of cytoplasmic components in type II pneumocytes and bronchiolar cells as well as thickness of the alveolar wall. These abnormalities were accompanied by depletion of glutathione (GSH) in the lung tissue. Pneumotoxic effects of systemic administration of styrene were dose dependent and tended to be more severe than those seen in the animals exposed for longer periods to styrene by inhalation. Metabolic activation of styrene and subsequent cell damage induced by the reactive metabolite styrene oxide may be involved in the sequence of events culminating in the toxic insult to the respiratory tract.

How to Study Boron Biodistribution in Liver Metastases from Colorectal Cancer

Summary The purpose of this study was to evaluate boron distribution for a safe and effective BNCT (Boron Neutron Capture Therapy) of liver metastases. Samples both from healthy and tumour liver parenchyma were analysed, after i.v. boron administration, by: a particles counting under neutron irradiation; morphological analysis by standard haematoxylin-eosin staining; neutron autoradiography. Our method was unaffected by the cytological heterogeneity inside tumour nodules; it demonstrated selective boron distribution in tumour tissue and predicted estimated mean therapeutic doses in tumour and safety doses in healthy tissue. The time interval for efficient BNCT was 2 to 4 hours after i.v. boron administration.

Gallic acid exerts a protective or an anti-proliferative effect on glioma T98G cells via dose-dependent epigenetic regulation mediated by miRNAs

Glioblastoma multiforme (GBM) is the most malignant primary brain tumor in adulthood, characterized by very high recurrence. Following the limited results for conventional therapies, novel therapeutic agents are under investigation. Among the putative new molecules, gallic acid (GA) represents a promising new anticancer drug. The anticancer effect of this drug has been based on its antioxidant effects. The aim of the present study was to investigate the toxic effects of GA on the T98G human glioblastoma cell line and its capacity to modulate the expression of microRNAs targeting the genes involved in tumor growth and invasion. Cytotoxicity, clonogenic ability and cell migration after GA treatment were tested. Moreover, the expression of miRNAs that target genes for antioxidant mitochondrial enzymes (miR-17-3p), p-21 protein (miR-21-5p) and ATM (miR-421-5p) was determined by qRT-PCR. The results confirmed in the T98G cells the anti-proliferative effect of GA reported for other glioma cell lines and showed that the miRNA expression changes depending on GA concentrations. Different GA concentrations can determine a protective or a toxic effect on tumor cells. Thus, the key for GA to induce a specific anticancer action is to use an optimal concentration that avoids these twin effects.

Histochemical study of cardiac mast cells degranulation and collagen deposition: interaction with the cathecolaminergic system in the rat

Although their role in the cardiovascular system is still largely unknown, mast cells are present in the myocardium of both experimental animals and humans. Interestingly, cathecolaminergic nerve fibres and mast cells are often described in close morphological and functional interactions in various organs. In the present study we investigated the effects of chronic interference with beta-adrenergic receptors (via either sympathectomy or beta-blockade) on cardiac mast cell morphology/activation and on interstitial collagen deposition. In rats subjected to chemical sympathectomizy with the neurotoxin 6-hydroxydopamine (6-OHDA) we observed a significant increase of mast cell density, and in particular of degranulating mast cells, suggesting a close relationship between the cardiac catecholaminergic system and mast cell activation. In parallel, chronic 6-OHDA treatment was associated with increased collagen deposition. The influence of the beta-adrenergic receptor component was investigated in rats subjected to chronic propranolol administration, that caused a further significant increase in mast cell activation associated with a lower extent of collagen deposition when compared to chemical sympathectomy. These data are the first demonstration of a close relationship between rat cardiac mast cell activation and the catecholaminergic system, with a complex interplay with cardiac collagen deposition. Specifically, abrogation of the cardiac sympathetic efferent drive by chemical sympathectomy causes mast cell activation and interstitial fibrosis, possibly due to the local effects of the neurotoxin 6-hydroxydopamine. In contrast, beta-adrenergic blockade is associated with enhanced mast cell degranulation and a lower extent of collagen deposition in the normal myocardium. In conclusion, cardiac mast cell activation is influenced by beta-adrenergic influences.