PROJECT # 1The expertise of the Institute of Biophysics (IBF) in light scattering techniques and spectroscopy will be

focalised to investigate both A beta-peptides conformational properties and aggregation kinetics as a function of the experimental conditions (ex. concentration, pH, temperature, metal ions concentration and type).

This Research Unit concentrates on the characterization of the A beta-peptide mutated systems, synthesized by the Research Unit IBB-CNR, to the purpose of obtaining model systems able to reproduce, by controlling

the experimental conditions, oligomeric states of the A beta-peptide. For this reason it is important to ascertain that mutations do not alter drastically A beta-peptides properties, although a higher stability for the

oligomeric states is achieved. The goals of the IBF can be summarized in the following issues:A beta-peptides stability and tendency to aggregate. Combined time resolved static and dynamic light

scattering measurements, at low and large angles, allow to follow the evolution in the scattering intensity as well as in the hydrodynamic radii distribution function relative to the species in solution. Therefore both

results give information on the growth of mass and hydrodynamic size of the aggregates.Conformational studies. Different size aggregates can be separated and fractionated by Size Exclusion

Chromatography. By using CD, fluorescence and microscopy on the separated fractions as well as in the native solutions, structural properties can be studied. These results can be faced to toxicity assays relative to

the different species in order to link structural and toxicity effects.Molecular modelling. The different attitude of the peptides synthesized to form fibrillar aggregates could be

studied by molecular modelling. Recent works have reported computer simulations on interaction studies in the presence of two or more A beta-peptides. Due to the high conformational freedom of the A beta-peptide,

it is required either the implementation of challenging simulation methods (as an ex. replica-exchange simulations) to guarantee a sufficiently large conformational space sampling, or the use of a more simple

coarse-grained model for the Abeta-peptide.Interaction A beta-peptides membranes. By using fluorescence correlation spectroscopy (FCS), the

interaction of the A beta-peptides with model membranes can be detected. Labelled peptides are used to measure differences of their hydrodynamic properties when free in solution or when is present a lipid bi-

layer. The occurrence of the interaction, together with the onset of any aggregation process due to the peptides can be also monitored by confocal microscopy technique.

References

Picone, P., R. Carrotta, G. Montana, R. Nobile, P. L. San Biagio, and M. Di Carlo. 2008. Beta-amyloid oligomers and fibrillar aggregates induce different apoptotic pathways in LAN5 neuroblastoma cell

cultures. Biophysical Journal in press.Di Carlo, M., R. Carrotta, G. Montana, and P. Picone. 2008. Amyloid b-protein: Physiological and

toxicalogical role In Biophysical inquiry into protein aggregation and amyloid diseases. D. Bulone, and P. L. San Biagio, editors. Transworld Research Network, Kerala, India. 233 - 252.Carrotta, R., J.

Barthès, A. Longo, V. Martorana, M. Manno, G. Portale, and P. L. San Biagio. 2007. Large size fibrillar bundles of the alzheimer amyloid beta-protein. Eur. Biophys. J. 36:701-709.

Carrotta, R., M. Di Carlo, M. Manno, G. Montana, P. Picone, D. Romancino, and P. L. San Biagio. 2006. Toxicity of recombinant beta-amyloid prefibrillar oligomers on the morphogenesis of the sea urchin

Paracentrotus lividus. The FASEB journal : official publication of the Federation of American Societies for Experimental Biology. 20:1916-1917.

CNR - ISTITUTO DI BIOFISICA

PROJECT # 2Insulin has long been known to form amyloid fibrils under given conditions (1). The formation of insulin fibrils is important not only for the development of reliable formulations or delivery systems, but also for modelling

the basic properties of protein self-organization. Insulin aggregation displays a wide variety of morphologies, from small oligomeric filaments to huge floccules or elongated mature fibrils (2-4). These structures

eventually assemble to form a gel. In this study, we plan to focus on the formation of such a gel, made of insulin amyloid fibrils, upon incubation

at high temperature and low pH. By light scattering and rheological techniques, we are able to monitor the development of the structural and mechanical properties of fibrillar aggregates, up to the dynamical arrest of

the sample, which marks the onset of gelation. Preliminary results have shown that the formation of large structures is correlated with the growth of the elastic modulus of the solution. The structural and dynamical

properties of these non ergodic-fibrillar gels will be characterized by large and small angle dynamic light scattering (5,6).

References

1. J. Brange, L. Andersen, E. D. Laursen, G. Meyn, and E. Rasmussen (1997) Towards under standing insulin fibrillation. J. Pharm. Sci. 86:517-525.

2. A. Podestà , G. Tiana, P. Milani, and M. Manno (2006) Early Events in Insulin Fibrillization Studied by Time-Lapse Atomic Force Microscopy. Biophys. J. 90:589-597.

3. M. Manno, E. F. Craparo, V. Martorana, D. Bulone, and P. L. San Biagio (2006) Kinetics of insulin aggregation: Disentanglement of amyloid brillation from large-size cluster formation. Biophys. J.

90:4585-4591.4. M. Manno, E. F. Craparo, A. Podestà , D. Bulone, R. Carrotta, V. Martorana, G. Tiana, and P. L. San

Biagio (2007) Kinetics of Different Processes in Human Insulin Amyloid Formation. J. Mol. Biol. 366:258 274.

5. M. Manno, D. Bulone, V. Martorana, and P. L. San Biagio (2004) Ergodic to non-ergodic transition monitored by scattered light intensity statistics. Physica A 341:40-54.

6. D. Bulone, D. Giacomazza, V. Martorana, J. Newman and P. L. San Biagio (2004) Ordering of agarose near the macroscopic gelation point. Phys. Rev. E 69:041401.

PROJECT # 3 Gel matrices based on hydrogels are widely used as encapsulation and release systems in food, cosmetic, agricultural or pharmaceutical applications. The design of matrices with structural properties suitable for a

specific use requires the knowledgement of new control parameters for gel properties, such as texture, strength, or porosity. Our activity in this field is aimed to understand, on a physical chemical basis, how the

release is modulated by the matrix properties. Drug-release systems have typically a multicomponent nature and the release mechanism is strongly dependent on the particular device used. Molecular diffusion of the

drug, swelling or shrinking or just dissolving of the gel matrix, direct intermolecular interaction between polymeric matrix and drug or surfactant are some of the main factors to be considered in projecting

controlled release formulations. Also, we investigate the changes in the structural properties of the matrix induced by the introduction of the particles to be released.

The experimental activity in this field takes advantage by the simultaneous use of different techniques such as static and dynamic light scattering at both large and small angle, fluorescence correlation spectroscopy,

and rheology.

CNR - ISTITUTO DI BIOFISICA

References

Monge, M. E., M. R. Negri, D. Giacomazza, and D. Bulone. 2008. Correlation between rheological properties and limonene release in pectin gels using an electronic nose. Food Hydrocolloids 22:916 - 922.

Montana, G., M. L. Bondi, R. Carrotta, P. Picone, E. F. Craparo, P. L. San Biagio, G. Giammona, and M. Di Carlo. 2007. Employment of cationic solid-lipid nanoparticles as RNA carriers. Bioconjugate

Chemistry 18:302-308.Mangione, M. R., D. Giacomazza, G. Cavallaro, D. Bulone, V. Martorana, and P. L. San Biagio. 2007.

Relation between structural and release properties in a polysaccharide gel system. Biophys. Chem. 129:18 - 24.

Giacomazza, D., D. Bulone, V. Martorana, G. Cavallaro, and P. L. San Biagio. 2005. K+ and Na+ effects on the gelation properties of k-Carrageenan. Biophysical Chemistry 113:129 -135.

Monge, M. E., D. Bulone, D. Giacomazza, M. R. Negri, and D. L. Bernik. 2004. Electronic Nose screening of limonene release from multicomponent essential oils encapsulated in pectin gels. Combinatorial

Chemistry & High Throughput Screening 7:337 - 326.Monge, M. E., D. Bulone, D. Giacomazza, D. L. Bernik, and M. R. Negri. 2004. Detection of flavor release

from pectin gels using electronic noses. Sensors & Actuators B - Chemical 101:28-38.

Facilities at BIOPHYSICS INSTITUTELIGHT SCATTERING LAB

• N.RO 2 BROOKHAVEN SM200 SPECTROMETERS• N.RO 2 BROOKHAVEN BI9000 PHOTON CORRELATORS• N.RO 1 ARGON LASER• N.RO 1 HE‐NE LASER

RHEOLOGY LAB• TA AR1000 RHEOMETER • TA AR‐G2 RHEOMETER• LAMINAR VERTICAL FLOW CABINET SERIL VBH

SPECTROSCOPY 1 LAB• UV‐VIS SHIMADZU UV2401PC SPECTROPHOTOMETER• N.RO 2 SHIMADZU GC 2010 FID E GC2010/GCMS QP2010S GAS CHROMATOGRAPHY• UV‐VIS JASCO SPECTROPHOTOMETER• JASCO P‐1020 POLARIMETER

SPECTROSCOPY 2 LAB• JASCO J‐815 CIRCULAR DICHROISM• JASCO FP6500 SPETTROFLUORIMETER• NIKON MICROSCOPE

LOW ANGLE LIGHT SCATTERING LAB

• HIGH RESOLUTION CCD CAMERA• N.RO 2 HE‐NE LASER

CNR - ISTITUTO DI BIOFISICA

CHEMISTRY LAB

• ELECTRONIC NOSE EOJ 38 ‐ SACMI• HPLC SHIMADZU LC20AT‐SPD• HAMAMATSU C9413‐01 FLUORESCENCE CORRELATOR SPECTROMETER

MICROCAL LAB

• HART SCIENTIFIC 707 DIFFERENTIAL MICROCALORIMETER• DMA58 DENSIMETER

More facilities accessible in Palermo for IBF Researcher- AFM

- Time resolved fluorescence- Two photon apparatus

CNR - ISTITUTO DI BIOFISICA

Dr. Giovanna BARBIERI (1 student)“Effects of the Major Histocompatibility Complex class II (MHC II) engagement and of the meso-tetra(4-sulfonatophenyl) porphynate derivatives on melanoma cells”

Melanoma, a solid tumour that arise from pigment producing melanocytes, is the most aggressive and lethal type form of cutaneous cancer in Caucasian light skin population. The extreme resistance of melanoma to the available tumour treatment has stimulated the investigation of immunotherapy approaches and the identification of new chemotherapeutic agents. Based on this, we will define the intracellular signalling that melanomas use to frustrate an effective anti-tumour response exerted by the cross-talk between the melanoma tumour cells and their immune counterparts. Moreover we will identify the molecular targets and the cytotoxic properties of new chemotherapeutic agents to improve the currently available chemotherapy.

The “Istituto di Biomedicina e Immunologia Molecolare”, is equipped with hoods at laminar flow and incubators for cells cultures; electrophoresis materials for proteins and DNAs; ultracentrifuges and refrigerated centrifuges for eppendorf and falcon tubes; fluorescence microscopy; spectrophotometers; -20 and –80 freezers; FACScan apparatus; autoclaves.

References of Barbieri-Costa group

1) V. Deffrennes, J. Vedrenne, M.-C. Stolzengerg, J. Piskurish, G. Barbieri, J. P. Ting, D. Charron and C. Alcaide-Loridan. Constitutive expression of MHC calss II genes in melanoma cell lines results from a transcription abnormally initiated from the promoter III of CIITA. J. Immunol. 167, (2001), 98-106.2) W. Wiszniewski, M.-C. Fondanèche, F. Le Deist, M. Kanariou, F. Selz, G. Barbieri, N. Brousse, V. Steimle, C. Alcaide-Loridan, D. Charron, A. Fischer and B. Lisowska-Grospierre. Mutation in the class II transactivator leading to mild immunodeficiency. J. Immunol. 167, (2001) 1787-1794.3) R. Naves, A.M. Lennon, G. Barbieri, L. Reyes, G. Puga, L. Salas, V. Deffrennes, M. Rosemblatt, M. Fellous, D. Charron, C. Alcaide-Loridan and M.R. Bono. MHC calls II-deficient tumor cell lines with a defective expression of the class II transactivator. Int. Immunol. 14 (5), (2002) 481-491.4) G. Barbieri, V. Deffrennes, T. Prod’homme, J. Vedrenne, F. Baton, C. Cortes, A. Fischer, M.R. Bono, B. Lisowka-Grospierre, D. Charron and C. Alcaide-Loridan. Isoforms of the class II transactivator protein. Int. Immunol. 14 (8), (2002) 839-848.5) T. Prod’homme, B. Drenou, C. De Ruyffelaere, G. Barbieri, W. Wiszniewski, C. Bastard, D. Charron and C. Alcaide-Loridan. Defective class II transactivator expression in a B lymphoma cell line. Leukemia 18(4), (2004) 832-40.6) M.A. Costa, L. Pellerito, V. Izzo, T. Fiore, C. Pellerito, M. Melis, M.T. Musmeci and G. Barbieri.Diorganotin(IV) and triorganotin(IV) complexes of meso-tetra(4-sulfonatophenyl)porphine induce apoptosis in A375 human melanoma cells. Cancer Letters, 238, (2006) 284–294.7) M.A. Costa, L. Gulino, L. Pellerito, T. Fiore, C. Pellerito and G. Barbieri. Effects of two organotin(IV)(sulfonatophenyl)porphynate on the MAPKs and on the growth of A375 human melanoma cells. Oncology Reports (accepted for publication).

Molecular Allergology labScientific responsible: Dr. Paolo COLOMBO (1 student)

Identification of allergens from Parietaria pollen by means of DNA recombinant technology. Expression of recombinant allergens in E.coli. Immunological reactivity to allergens in healthy and allergic subjects

Technologies:cDNA cloning, Protein purification, Western blots, Cell culture

References (last three years):

1: Bonura A, Gulino L, Trapani A, Di Felice G, Tinghino R, Amoroso S, Geraci D, Valenta R, Westritschnig K, Scala E, Mari A, Colombo P. Isolation, expression and immunological characterization of a calcium-binding protein from Parietaria pollen. Mol Immunol. 2008 May;45(9):2465-73. Epub 2008 Mar 4.2: Nasta F, Corinti S, Bonura A, Colombo P, Di Felice G, Pioli C. CTLA-4 regulates allergen response by modulating GATA-3 protein level per cell. Immunology. 2007 May;121(1):62-70. Epub 2007 Feb 20.3: Colombo P, Bonura A. Parietaria pollen allergens. Eur Ann Allergy Clin Immunol. 2006 Sep;38(7):224-5. Review. 4: Bonura A, Corinti S, Artale A, Di Felice G, Amoroso S, Melis M, Geraci D,Colombo P. A hybrid expressing genetically engineered major allergens of the Parietariapollen as a tool for specific allergy vaccination. Int Arch Allergy Immunol. 2007;142(4):274-84. Epub 2006 Nov 22.5: Bonura A, Artale A, Marino M, Amoroso S, Marcucci F, Geraci D, Colombo P. Cross-reactivity between Parietaria species using the major rParj1 and rParj2 allergens. Allergy Asthma Proc. 2006 Sep-Oct;27(5):378-82.

Dr. Marta DI CARLO (1 student)

The topic of our research concerns the study of the beta amyloid a peptide involved in Alzheimer’s disease. This pepide, under some conditions, from its native conformational state is converted into higly ordered fibrillar aggregates. Such transitions can give rise to pathological conditions ranging from neurodegenerative disorders to systemic amyloidoses. Aim of our research project is to use beta amyloid to study:

!ibrillogenesis oxidativestress Celldeath modulationofgeneexpression

Used techniques:

cell colturebacteria colturenucleic acids extractionprotein extractionelectrophoreses for proteins and nucleic acidPCRWestern blotmicroscopy

Recent references1: Pellicanò M, Picone P, Cavalieri V, Carrotta R, Spinelli G, Di Carlo M. The sea urchin embryo: A model to study Alzheimer's beta amyloid induced toxicity. Arch Biochem Biophys. 2008 Dec 25. [Epub ahead of print]2: Romancino DP, Montana G, Cavalieri V, Spinelli G, Di Carlo M. EGFR signalling is required for Paracentrotus lividus endomesoderm specification. Arch Biochem Biophys. 2008 Jun 1;474(1):167-74. Epub 2008 Mar 26.3: Sabbatini ME, Rodríguez M, di Carlo MB, Davio CA, Vatta MS, Bianciotti LG. C-type natriuretic peptide enhances amylase release through NPR-C receptors in the exocrine pancreas. Am J Physiol Gastrointest Liver Physiol. 2007 Nov;293(5):G987-94. Epub 2007 Aug 16.4: Montana G, Bondì ML, Carrotta R, Picone P, Craparo EF, San Biagio PL, Giammona G, Di Carlo M. Employment of cationic solid-lipid nanoparticles as RNA carriers. Bioconjug Chem. 2007 Mar-Apr;18(2):302-8. Epub 2007 Jan 25.5: Santangelo C, Matarrese P, Masella R, Di Carlo MC, Di Lillo A, Scazzocchio B, Vecci E, Malorni W, Perfetti R, Anastasi E. Hepatocyte growth factor protects rat RINm5F cell line against free fatty acid-induced apoptosis by counteracting oxidative stress. J Mol Endocrinol. 2007 Feb;38(1-2):147-58.6: Carrotta R, Di Carlo M, Manno M, Montana G, Picone P, Romancino D, San Biagio PL.Toxicity of recombinant beta-amyloid prefibrillar oligomers on the morphogenesis of the sea urchin Paracentrotus lividus. FASEB J. 2006 Sep;20(11):1916-7. Epub 2006 Jul 3.

Research Group: Oncobiology Scientific responsible: Dr. Melchiorre CERVELLO (1 student)

Our main field of interest is the study of the molecular mechanisms which contribute to the acquisition of peculiar biological characteristics of neoplastic cells, such as loss of cell growth control and resistance to apoptosis. In particular, in recent years we have focused our attention on the study on the role of cyclooxygenase-2 (COX-2) during hepatocarcinogenesis. In addition, we have studied the role of the beta-catenin system in the control of hepatoma cell proliferation. Another aspect of great interest studied by my research group is related to the characterization and definition of the molecular targets for particular drugs, such as proteasome, NF-κB, Raf/MEK/ERK, PI3K/AKT/mTOR and COX-2 inhibitors. In collaboration with clinicians we have also started a new project for the molecular classification of hepatocellular carcinoma (HCC) by molecular profiling technologies of RNA and microRNA extracted from tumoral and nontumoral tissues (mainly liver cirrhosis) from patients with HCC. In addition, we have also studying the presence of free circulating DNA and RNA in plasma and serum from HCC and LC patients in order to identify biomarkers for early detection of cancer.

Techniques used in the lab:Cell culture RNA and DNA purificationcDNA synthesis/RT-PCRCell transfectionSDS-PAGEWestern blottingImmunofluorescenceCytofluorimetryCell growth assays

References (last three years):

1. Notarbartolo M, Lo Cicero S, Meli M, Poma P, Labbozzetta M, Cervello M, D'Alessandro N. Induction of apoptosis by the adenosine derivative IB-MECA in parental or multidrug-resistant HL-60 leukemia cells: possible relationship to the effects on inhibitor of apoptosis protein levels. Chemotherapy. (2005), 5, 272-279

2. Lampiasi N, Foderà D, D'Alessandro N, Cusimano A, Azzolina A, Tripodo C, Florena AM, Minervini MI, Notarbartolo M, Montalto G, Cervello M. The selective cyclooxygenase-1 inhibitor SC-560 suppresses cell proliferation and induces apoptosis in human hepatocellular carcinoma cells. Int J Mol Med. (2006), 17, 245-252 Licciardi M, Campisi M, Cavallaro G, Cervello M, Azzolina A, Giammona G. Synthesis and characterization of polyaminoacidic polycations for gene delivery. Biomaterials (2006), 27, 2066-2075

3. Cervello M, Montalto G. Cyclooxygenases in hepatocellular carcinoma. World J Gastroenterol. (2006), 12: 5113-5121 Poma P, Notarbartolo M, Labbozzetta M, Sanguedolce R, Alaimo A, Carina V, Maurici A, Cusimano A, Cervello M, D'Alessandro N. Antitumor effects of the novel NF-kappaB inhibitor dehydroxymethyl-epoxyquinomicin on human hepatic cancer cells:

analysis of synergy with cisplatin and of possible correlation with inhibition of pro-survival genes and IL-6 production. Int J Oncol. (2006), 28, 923-930 M Soresi, L, Giannitrapani L, D’Antona F, Florena AM, La Spada E, Terranova A, Cervello M, D’Alessandro N, Montalto G. Interleukin-6 and its soluble receptor in patients with liver cirrhosis and hepatocellular carcinoma. World J Gastroenterol. (2006), 12, 2563-2568

4. Labbozzetta M, Notarbartolo M, Poma P, Giannitrapani L, Cervello M, Montalto G, D'Alessandro N. Significance of autologous interleukin-6 production in the HA22T/VGH cell model of hepatocellular carcinoma. Ann N Y Acad Sci. (2006), 1089, 268-275

5. Giannitrapani L, Soresi M, La Spada E, Cervello M, D'Alessandro N, Montalto G. Sex hormones and risk of liver tumor. Ann N Y Acad Sci. (2006), 1089, 228-236

6. Bondì ML, Craparo EF, Giammona G, Cervello M, Azzolina A, Diana P, Martorana A, Cirrincione G. Nanostructured Lipid Carriers-Containing Anticancer Compounds: Preparation, Characterization, and Cytotoxicity Studies. Drug Delivery (2007), 14: 61-67

7. Lampiasi N, Azzolina A, Montalto G, Cervello M. Histamine and spontaneously released mast cell granules affect the cell growth of human hepatocellular carcinoma cells. Exp Mol Med (2007), 39: 248-94

8. Giannitrapani L, Soresi M, Ingrao S, La Spada E, Vuturo O, Florena AM, Cervello M, Montalto G. Response to antiviral therapy and hepatic expression of cyclooxygenases in chronic hepatitis C. Eur J Gastroenterol Hepatol. (2007), 19:927-933

9. Cusimano A, Foderà D, D’Alessandro N, Lampiasi N, Azzolina A, Montalto G, Cervello M Potentiation of the antitumor effects of both selective cyclooxygenase-1 and cyclooxygenase-2 inhibitors in human hepatic cancer cells by inhibition of the MEK/ERK pathway. Cancer Biol Ther (2007), 6: 1461-1468

10. McCubrey JA, Milella M, Tafuri A, Martelli AM, Lunghi P, Bonati A, Cervello M, Lee JT, Steelman LS. Targeting the Raf/MEK/ERK pathway with small molecule inhibitors . Curr Opin Investig Drugs (2008), 9: 614-630

11. Steelman SL, Stadelman KM, Chappell WH, Horn S, Bäsecke J, Cervello M, Nicoletti F, Libra M, Martelli AM, McCubrey JA. Akt as a Therapeutic Target in Cancer. Expert Opin Ther Targets (2008), 12:1139-1365

12. McCubrey JA, Bäsecke J, Cervello M, Martelli AM, Franklin RA. GSK-3beta is a critical mediator of tetrandrine induced cell cycle arrest and cytotoxicity. Cancer Biol Ther. (2008), 7: 1079

13. Misaghian N, Ligresti G, Steelman LS, Bertrand FE, Bäsecke J, Libra M, Nicoletti F, Stivala F, Milella M, Tafuri A, Cervello M, Martelli AM, McCubrey JA.Targeting the leukemic stem cell: the Holy Grail of leukemia therapy. Leukemia (2008), in press

Dr. Patrizia GUARNERI (1 Student)

The laboratory of Neuroscience is composed by senior researchers and young investigators, some of them with temporary positions and others included in training programs. The activities of research are devoted to the study of neurodegenerative mechanisms related to retinopathies and neurodegenerative diseases with the precise intent to identify molecular targets for diagnosis and cure of diseases. Special interests are: 1) The role of steroids in the retinal physiopathology. Studies are currently looking at the role of estrogen

and androgen receptors as possible regulators of gene expression and function in the retina and as key players in the neurodegenerative processes occurring in retinal ischemia. Biochemical, molecular and pharmacological approaches together with an analysis of retinal proteoma are carried out using retinal explants, an in vitro model of retinal ischemia and intraocular treatments with drugs.

2) The role of metal ion dishomeostasis in relation to the amyloidogenesis, angiogenesis and neurodegneration in models of proliferative retinopathies, such as diabetic retinopathy and exudative age macular disease. This study has the precise goal to develop novel approaches for diagnosis and therapy throughout the collaboration with drug designer’s people and to provide evidence for improving the knowledge and treatment of Alzheimer’s disease in view of its many similarities with these retinopathies.

3) The characterization of the pathogenic mechanisms of the CLN8 variant form of the late-infantile neuronal ceroid lipofuscinosis, which is an autosomic recessive neurodegenerative disease. The CLN8 gene encodes a transmembrane protein mainly located at the endoplasmic reticulum, whose function is still unknown. Studies are in progress to characterize the CLN8 function through protein-protein interaction systems and to define the death pathways involved in the neurodegeneration of selective CNS structures in a mouse CLN8 model. At present, studies related to the involvement of apoptosis, inflammation, autophagy and ER/UPR stress are conducted.

4) The identification of VAP family proteins as probable modifiers of various ALS phenotypes linked to the different rates of progression throughout studies on muscle biopsy and primary fibroblast cultures of SALS patients.

Techniques used: cell culture, animal handling and treatment, western-blotting, 2DE, protein-protein interaction, split-ubiquitin two hybrid system, co-immunoprecipitation studies, antibody purification, energetic metabolism, oxidative stress, lipid peroxidation, southern and northern blotting, DNA laddering, RT-PCR, transcription and transduction in vitro, EMSA/DNaseI footprinting, transient transfection in cells, construction of cDNA libraries in E. coli and Streptomycetes, GST-Pull Down, over-expression of genes in E. coli and purification of proteins, optic and electron microscopy of cells and tissues, immuno-histo and cytochemistry.

References (last five years):

1: Cascio C, Russo D, Drago G, Galizzi G, Passantino R, Guarneri R, Guarneri P. 17beta-estradiol synthesis in the adult male rat retina. Exp Eye Res. 2007 Jul;85(1):166-72. Epub 2007 Feb 16.2: Guarneri R, Russo D, Cascio C, D'Agostino S, Galizzi G, Bigini P, Mennini T,Guarneri P. Retinal oxidation, apoptosis and age- and sex-differences in the mnd mutantmouse, a model of neuronal ceroid lipofuscinosis. Brain Res. 2004 Jul 16;1014(1-2):209-20.3: Guarneri P, Cascio C, Russo D, D'Agostino S, Drago G, Galizzi G, De Leo G,Piccoli F, Guarneri M, Guarneri R. Neurosteroids in the retina: neurodegenerative and neuroprotective agents in retinal degeneration. Ann N Y Acad Sci. 2003 Dec;1007:117-28. Review.

Mark GJOMARKAJ’s group activity (1 student)

We have been involved over the past years in investigating the mechanisms of inflammation and remodeling in bronchial asthma. We participated as active partner to European research projects (ENFUMOSA, BIOAIR, GA2LEN) supported by the European Union in the context of the IV, the V, and the VI framework programmes, respectively. These studies allowed us to identify biomarkers detectable by non invasive methods in the airways (exhaled breath condensate, exhaled NO) and in peripheral blood and induced sputum, as well as by invasive methods in bronchial biopsies. These biomarkers allow to monitor the disease activity and to identify specific patient’s phenotypes at putative risk to develop a severe disease and to develop airway remodeling. We also demonstrated that the improvement of the effects of the combined therapy (glucocorticoids and beta agonists) is also due to the increase of some anti-inflammatory effects of the two combined drugs with respect to their single use. With regard to drug resistance, non responder patients to GC therapy have an altered expression of the GC receptors. These studies are also supported by pharmaceutical industries. Our group also investigates the molecular mechanisms involved in chronic obstructive pulmonary disease (COPD). Also these studies were performed on peripheral blood, induced sputum, and surgical lung samples from COPD patients. Laser microdisection, real-time PCR, GeArray, bidimensional electrophoresis, western blot analysis, and zimography are the techniques used for these studies. These studies are supported by different pharmaceutical industries. Finally, our group has published a number of manuscripts showing the mechanisms involved in the inflammatory processes during pleural inflammation. These mechanisms were investigated in both neoplastic and non neoplastic pleuritis. These studies were supported by two two years grants of the Alberta Lung Association (Canada).

Publications year 20081:Pleural mesothelial cells express both BLT2 and PPARalpha and mount an integrated response to pleural leukotriene B4.Pace E, Ferraro M, Mody CH, Melis MR, Scafidi V, Bonanno A, Profita M, Giarratano A, Gjomarkaj M.J Immunol. 2008 Nov 15;181(10):7292-9.

2: A case-control study of the relation between plasma selenium and asthma in European populations: a GAL2EN project.Burney P, Potts J, Makowska J, Kowalski M, Phillips J, Gnatiuc L, Shaheen S, Joos G, Van Cauwenberge P, van Zele T, Verbruggen K, van Durme Y, Derudder I, Wohrl S, Godnic-Cvar J, Salameh B, Skadhauge L, Thomsen G, Zuberbier T, Bergmann KC, Heinzerling L, Renz H, Al-Fakhri N, Kosche B, Hildenberg A, Papadopoulos NG, Xepapadaki P, Zannikos K, Gjomarkaj M, Bruno A, Pace E, Bonini S, Bresciani M, Gramiccioni C, Fokkens W, Weersink EJ, Carlsen KH, Bakkeheim E, Loureiro C, Villanueva CM, Sanjuas C, Zock JP, Lundback B, Janson C.Allergy. 2008 Jul;63(7):865-71. Erratum in: Allergy. 2008 Dec;63(12):1647.

3:Important research questions in allergy and related diseases: nonallergic rhinitis: a GA2LEN paper.Bousquet J, Fokkens W, Burney P, Durham SR, Bachert C, Akdis CA, Canonica GW, Dahlen SE, Zuberbier T, Bieber T, Bonini S, Bousquet PJ, Brozek JL, Cardell LO, Crameri R, Custovic A, Demoly P, van Wijk RG, Gjomarkaj M, Holland C, Howarth P, Humbert M, Johnston SL, Kauffmann F, Kowalski ML, Lambrecht B, Lehmann S, Leynaert B, Lodrup-Carlsen K, Mullol J, Niggemann B, Nizankowska-Mogilnicka E, Papadopoulos N, Passalacqua G, Schünemann HJ, Simon HU, Todo-Bom A, Toskala E, Valenta R, Wickman M, Zock JP.Allergy. 2008 Jul;63(7):842-53. Review.

4: The relation between paracetamol use and asthma: a GA2LEN European case-control study.

Shaheen S, Potts J, Gnatiuc L, Makowska J, Kowalski ML, Joos G, van Zele T, van Durme Y, De Rudder I, Wöhrl S, Godnic-Cvar J, Skadhauge L, Thomsen G, Zuberbier T, Bergmann KC, Heinzerling L, Gjomarkaj M, Bruno A, Pace E, Bonini S, Fokkens W, Weersink EJ, Loureiro C, Todo-Bom A, Villanueva CM, Sanjuas C, Zock JP, Janson C, Burney P; Selenium and Asthma Research Integration project; GA2LEN.Eur Respir J. 2008 Nov;32(5):1231-6. Epub 2008 Jun 25.

5: Cysteinyl leukotriene-1 receptor activation in a human bronchial epithelial cell line leads to signal transducer and activator of transcription 1-mediated eosinophil adhesion.Profita M, Sala A, Bonanno A, Siena L, Ferraro M, Di Giorgi R, Montalbano AM, Albano GD, Gagliardo R, Gjomarkaj M.J Pharmacol Exp Ther. 2008 Jun;325(3):1024-30. Epub 2008 Feb 27.

6: Acetylcholine mediates the release of IL-8 in human bronchial epithelial cells by a NFkB/ERK-dependent mechanism.Profita M, Bonanno A, Siena L, Ferraro M, Montalbano AM, Pompeo F, Riccobono L, Pieper MP, Gjomarkaj M.Eur J Pharmacol. 2008 Mar 17;582(1-3):145-53. Epub 2008 Jan 9.

7: Cigarette smoke increases Toll-like receptor 4 and modifies lipopolysaccharide-mediated responses in airway epithelial cells.Pace E, Ferraro M, Siena L, Melis M, Montalbano AM, Johnson M, Bonsignore MR, Bonsignore G, Gjomarkaj M.Immunology. 2008 Jul;124(3):401-11. Epub 2008 Jan 22.

Cell Stress and Environment Research UnitGroup Leader: Dr. Valeria Matranga

tel +39-091-6809551; fax +39-091-6809557email: valeria.matranga@ibim.cnr.it

Research activitiesDevelopment of the sea urchin embryo, including embryotoxicity tests,

regeneration studies, aquaculture and biomineralization projects. Read more at: http://www.ibim.cnr.it/matranga/matranga.htm

Techniques used: biochemistry, cell and molecular biologyInstruments: optical microscopes equipped with epifluorescence and

micromanipulators; aquaria and cell culture rooms; instruments for the extraction, separation and analysis of proteins and nucleic acids

Litterature:Matranga V. and Y. Yokota (2008) Responses of marine organisms to physical and

chemical impacts. Cell Biology and Toxicology 24:471-474.Filosto, S. Roccheri, M. C., Bonaventura, R., and V. Matranga (2008) Environmentally

relevant cadmium concentrations affect development and induce apoptosis of Paracentrotus lividus larvae cultured in vitro. Cell Biology and Toxicology 24: 603-610.

Kiyomoto M., F. Zito, C. Costa, V. Poma, S. Sciarrino and V. Matranga (2007) Skeletogenesis by transfated secondary mesenchyme cells is dependent on extracellular matrix–ectoderm interactions in Paracentrotus lividus sea urchin embryos. Dev Growth Diff 49: 731-741

Bonaventura R., V. Poma, R. Russo, F. Zito and V. Matranga (2006) Effects of UV-B radiation on the development and hsp 70 expression in sea urchin cleavage embryos. Marine Biology 149: 79-86

Matranga V. (2005). Marine Molecular Biotechnology: Echinodermata – Volume N° 39 of the series Progress in Molecular and Subcellular Biology, Springer Press, Berlin, Germany, pp 1-264