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This article aims at highlighting the most recent and promising research trends, the open challenges and the possible routes to follow in the field of targeted therapy. A highly interdisciplinary viewpoint has been used, trying to evidence and discuss the different opportunities deriving from recent evolutions of nanotechnology, polymer science, robotics and biotechnology. The most used vectors for nanomedicine applications are described, together with the different action strategies reported in the literature, such as passive targeting, site-directed targeting and remotely triggerable drug delivery. Special emphasis is given to magnetically triggered systems and ultrasound-responsive materials, identified as the most promising paradigms. Key competences and system integration strategies derived from robotics are also introduced, focusing the attention on the crucial issue of achieving high controllability of the vector at the micro- and nano-scale. Finally, biocomponents are described, highlighting their potential as functional sensing elements or smart mechanisms to be integrated on board of advanced micro-nano therapeutic devices. The conclusion aims at depicting the importance of novel and improved targeted therapy strategies, to be coupled with the emerging world of predicting and personalized medicine. To this aim, a real merging of skills and approaches, derived from the aforementioned research fields, is recognized as highly desirable and rich of opportunities.

Virus cell‐to‐cell spread has been reported for many different viruses and may contribute to pathogenesis of viral disease. The role played by cell‐to‐cell contact in hepatitis C virus (HCV) transmission was studied in vitro by cell co‐cultivation experiments. A human lymphoblastoid B‐cell line, infected persistently with HCV in vitro (TO.FEHCV), was used as HCV donor [Serafino et al., 2003]; recipient cells were the human hepatoma HepG2 cell line. Both cell types were co‐cultured for 48 hr to allow the cell‐to‐cell contacts. The hepatoma HepG2 cells are not permissive to free‐virus infection, but they were infected successfully using TO.FEHCVcells as source of virus. The kinetics of viral RNA synthesis and the percentage of infected cells were compared in cell‐mediated‐and cell‐free‐viral infection. After co‐cultivation, a consistent proportion of hepatoma cells replicated HCV and stably expressed viral antigens. Virus produced was infectious as demonstrated by the ability to reinfect fresh B‐cells. This cell model shows that permissiveness to HCV infection can be achieved in vitro in non‐permissive hepatoma cells by direct cell‐to‐cell contacts with infected human B‐cells. This mechanism of virus spread may also play a pathogenic role in vivo. J. Med. Virol.78:192–201, 2006. © 2005 Wiley‐Liss, Inc.

Background:Calcium-binding plant allergens can be grouped in different families according to the number of calciumbinding domains (EF hands). Objective:We sought to identify pollens containing crossreactive calcium-binding allergens and to investigate structural and immunologic similarities of members belonging to different families of calcium-binding allergens. Methods:By means of multiple sequence alignment and molecular modeling, we searched for structural similarities among pollen allergens with 2 (Phl p 7, timothy grass; Aln g 4, alder), 3 (Bet v 3, birch) and 4 EF hands (Jun o 4, prickly juniper). Purified recombinant Aln g 4 and Jun o 4 were used to determine the prevalence of IgE recognition in 210 patients sensitized to different pollens and to search, by means of ELISA competition, for the presence of cross-reactive epitopes in pollens from 16 unrelated plant species. IgE cross-reactivity among the allergen families was studied with purified rPhl p 7, rAln g 4, rBet v 3, and rJun o 4 and 2 synthetic peptides comprising the N-terminal and C-terminal EF hands of Phl p 7 by means of ELISA competition. Results:Structural similarities were found by using molecular modeling among the allergens with 2, 3, and 4 EF hands. Pollens from 16 unrelated plants contained Aln g 4- and Jun o 4-related epitopes. Twenty-two percent of the patients with multiple pollen sensitization reacted to at least one of the calcium-binding allergens. A hierarchy of IgE cross-reactivity (rPhl p7 rAln g 4 rJun o 4 rBet v 3) could be established that identified rPhl p 7 as the EF-hand allergen containing most IgE epitopes in the population studied. Conclusion:The demonstration that members of different families of calcium-binding plant allergens share similarities suggests that it may be possible to use representative molecules for the diagnosis and therapy of allergies to EF-hand allergens. (J Allergy Clin Immunol 2002;109:314-20.)

It has been suggested that an atypical course of primary infection by EBV and the reactivation of EBV infection in transplanted patients may induce hepatitis. We explored the possibility to dissect the infectious activity from the ability to promote B lymphocyte proliferation in vivo by injecting in nu/nu mice a low number (2 × 106–0.05 × 106) of cells from CE a normal human bone marrow–derived B cell line. This line carries an endogenous EBV in episomal and linear forms. Twenty nu/nu mice were inoculated subcutaneously with the B cell line CE and a matched group with the cell line RAG obtained by EBV in vitro infection of normal human peripheral blood. The mice injected with the CE line did not develop a lymphoproliferative disease, but 5 of them displayed typical histopathological lesions of chronic hepatitis without involvement of other organs. Similar results were obtained in 2 out of 20 animals in the RAG group. A close association between liver lesions and a previous EBV infection, by putative circulating B lymphoblastoid cells releasing their EBV, was established by PCR and by in situ hybridization with BamHI “W” DNA probe. This latter probe detected the presence of about 15% of positive cells only in affected livers. In addition, the rare detection in some hepatocytes of “A” type Cowdry bodies would suggest the occurrence of continuous EBV replication although at a very low level. These data show that we succeeded in dissecting the infectious from the proliferative activity of the endogenous EBV carrier CE cell line. This provides in addition a promising model for chronic EBV‐associated hepatitis. J. Med. Virol. 66:70–81, 2002. © 2002 Wiley‐Liss, Inc.

Background:Cow’s milk allergy is a common disease of infancy and early childhood. If the baby is not breast-fed, a substitute for cow’s milk formula is necessary. Objective:The aim of this study was to investigate, in vitro and in vivo, the allergenicity of mare’s milk in a population of selected children with severe IgE-mediated cow’s milk allergy. Methods:Twenty-five children (17 male and 8 female) aged 19 to 72 months (median age 34 months) with IgE-mediated cow’s milk allergy were selected for this study. All the children underwent skin prick tests with cow’s milk and mare’s milk and double-blind placebo-controlled oral food challenge (DBPCOFC) with fresh cow’s milk, fresh mare’s milk, and, as placebo, a soy formula (Isomil, Abbott, Campoverde, Italy). We performed immunoblotting of cow’s and mare’s milk developed with IgE from allergic children. Results:All the children showed strong positive skin test responses to cow’s milk (4+); 2 children had positive skin test responses to mare’s milk (2+). All children had positive DBPCOFCs to cow’s milk; one child had a positive DBPCOFC to mare’s milk. No children reacted to the placebo (Isomil). In the cow’s milk, some proteins are able to strongly react with human IgE; when the sera are tested with mare’s milk, the bands corresponding to the same proteins are recognized by a lower percentage of sera. Conclusion:These data suggest that mare’s milk can be regarded as a good substitute of cow’s milk in most children with severe IgE-mediated cow’s milk allergy. It would be prudent, however, to confirm its tolerability by a supervised titrated oral challenge test. (J Allergy Clin Immunol 2000;105:1031-4.)

Background:Cross-reacting carbohydrate determinants (CCDs) are antigenic structures shared by allergenic components from taxonomically distant sources. The case history of a patient with a great discrepancy between skin test and specific IgE results led us to investigate the role of these determinants in his specific case and in an allergic population. Objective:We sought to determine the role of CCDs in causing false-positive and clinically irrelevant results in in vitro tests. Methods:The involvement of CCDs was studied by specific IgE inhibition by using glycoproteins with a known carbohydrate structure. Direct and inhibition assays were performed by commercially available systems, in-house ELISA, and the immunoblotting technique. The binding to the periodate-oxidated carbohydrate structure of glycoproteins and allergenic extracts was also evaluated. A comparative study between skin test and specific IgE responses to the antigens studied was carried out in 428 consecutive allergic subjects. Results:All the tests performed suggested that cross-reacting carbohydrate epitopes were the cause of false-positive specific IgE results in one of the commercial systems and the high reactivity in all the solid-phase in vitro tests. None of the cross-reacting carbohydrate allergens yielded a positive skin test response. Periodate treatment caused variable degrees of reduction of IgE binding to the different antigens studied, indicating that CCDs played a different role in each of them. About 41% of patients allergic to pollen had specific IgE for a glycoprotein, without a positive skin test response to the same molecule. Conclusions:CCDs must be taken into account when evaluating the clinical relevance of positive results in in vitro specific IgE assays, at least in the diagnosis of patients with pollen allergy. Commercial systems should be carefully assessed for the ability to detect specific IgE for carbohydrate determinants to avoid false-positive or clinically irrelevant results. (J Allergy Clin Immunol 1999;103:1005-11.)

Using the CELLO detector, we have measured cross sections for the processγγ→π+π− in the mass range 0.75–1.9 GeV/c2. A partial wave fit to the data indicates the presence of a sizeableS-wave amplitude with signs of resonant behaviour. Values for the γγ width of thef2(1270) are given, showing the model-dependence of this quantity. At higher dipion masses, the model of Brodsky and Lepage is found to give an order of magnitude description of the data.

We have measured the inclusive production of γ, π0 and η ine+e− annihilation at the center of mass energy of 35 GeV. The differential cross sections, extended to the kinematical limit and measured with high accuracy, are found to be in good agreement with previously reported results. Using the measured spectra we determine the average multiplicity for each of these particle species.

We present an analysis of strange particle production frome+e- annihilation into multihadronic final states. The experiment was performed with the CELLO detector at the PETRA storage ring at DESY, the data was taken at a centre of mass energy of 35 GeV with an integrated luminosity of 86 pb-1. The particlesKS0,K*± and ? have been identified by their characteristic decays, and differential cross sections for their production have been obtained. From a comparison ofKS0 andK*± rates the Lund vector meson suppression parameterV/(V+P)S has been determined.

The reactione+e−→e+e−π+π−π0 in the untagged mode has been measured with the CELLO detector at PETRA. The cross section is dominated by exclusivea2(1320) production, whose radiative width is determined to beΓγγ=1.00±0.07(stat.)±0.15(syst.)keV. An angular correlation analysis indicates pure helicity 2 with an upper limit on the helicity 0 contribution of 8.2% (at 95% c.l.). Excitation of the pseudotensor meson π20(1670) is observed for the first time in this reaction. The π2 radiative width and its dependence on interference effects are studied.

We have searched for resonance production in the reaction ???Ks0Kp. No signal was found for the?c and an upper limit for the radiative with keV (95% c.l.) is obtained. For the glueball candidate ?(1440) (previouslyi) the upper limit is derived. In the tagged data sample resonance formation of a spin 1 state at 1420 MeV is observed, which is absent in the untagged data. The mass and width of this state are consistent with those of thef1(1420); an analysis of decay angular distributions favours positive parity.

The expression of the cellular gene coding for the epidermal growth factor (EGF) receptor (EGF-R) was assayed in the presence of hepatitis B virus (HBV) gene expression under different experimental conditions in human hepatoma-derived cells. First, transfection experiments of the well-differentiated HepG2 human hepatoma cell line using different expression vectors of the HBV X-region demonstrated that the X-gene product is capable of inducing EGF-R gene overexpression; in addition, by using a stable in vitro expression system for HBV, it was shown that EGF-R gene expression in these cells is greater than in the uninfected parent cells, and that this results in a three-fold increase in ¹²⁵I-EGF binding. Finally, a CAT-expression assay was performed, indicating that regulatory regions of the EGF-R-gene are target sequences for X-protein trans-activation. Copyright 1993, 1999 Academic Press

We present measurement of the p0?*?, ??*? and ?'?*? form factors. The p0-form factor is for the first time observed in the space-like region. The transition form factor of the ?-meson is determined from its decay modes p+p-p0, p+p-? and the neutral decay mode ??. The decay of the ?' is observed in the decay channels ??, ?p+p- with ???? and in the four charged prong final state stemming from ?p+p- with the ? decaying into p+p-(p0/?). All form factors agree well with a simple ?-pole predicted by the vector meson dominance model and also with the QCD inspired Brodsky-Lepage model.

Multihadronic events measured with the CELLO detector in the energy range 14 to 46 GeV have been analyzed in terms of thrust, jet masses and the asymmetry of the energy-energy correlation. The data have been compared with 2nd order QCD calculations. From a study of the general properties of fragmentation effects, model-independent limits on ΛQCD and αs have been found to be 79 MeV < ΛQCD < 628 MeV and 0.117 < αs < 0.169 (at GeV). The dependence of these results on the renormalization scheme is discussed.

We present the results of a search for Gamma Ray Bursts at energies Eγ ≥ 10 GeV and Eγ ≥ 100 TeV made by the EAS-TOP Air Shower Array in correlation with ∼ 50 events detected by BATSE in the last 3 years.

We have studied the ρ0 production rate in the reaction ξξ→3π+3π− in the energy range 1.6≦Wγγ≦7.5 GeV with the CELLO detector at PETRA. Our analysis points to a substantial yield of ρ0ρ0π+π− events in particular atWγγ>4.0 GeV. We give cross sections for the ρ02π+2π− and ρ0ρ0π+π− final states and calculate upper limits for the reaction γγ→ρ0ρ0 (1700) →ρ0ρ0π+π−.

An asymmetric distribution of deoxy‐5‐methylcytidylic acid‐inhibiting restriction sites (dcm‐sites) takes place in ten human genes regulated by 5‐methylcytosine. These genes are dcm‐site enriched upstream and dcm‐site poor downstream. Along them, there is a scattering of hypermethylatable introns and hypomethylatable exons with a common code: the 5mCpG dinucleotides characterize promoters; Gp5mCs characterize introns; Tp5mCs and Cp5mCs are in small concentrations in exons. Housekeeping genes contain more dcm‐sites when compared with tissue‐specinc genes. This depends on the higher number of dcm‐sites in their promoters and introns. In exons, the relatively lower number of dcm‐sites is almost the same in both housekeeping and tissue‐specific genes. Going from 5' to 3', the average frequency of occurrence of these sites per nucleotide units decreases in introns and increases in exons. This difference is highly discriminated for tissue‐specinc and less discriminated for housekeeping genes.

Abstract: The three reactionse +e−→e +e−e+e−,e + e -→e + e -μ+μ- ande + e -→μ+μ-μ+ μ- have been studied using the CELLO detector at PETRA. The data correspond to 130 pb−1 collected at energies ranging from 35 GeV to 46.8 GeV. A detailed analysis of the distribution of the observed events in phase space shows good agreement with QED to order α4.

Background: Species belonging to the Cupressaceae family are a relevant source of allergens that are present in a wide number of countries. Objective: We sought to identify, purify, and characterize recombinant allergens from Juniperus oxycedrus, a species belonging to the Cupressaceae family. Methods: Double-stranded cDNA was synthesized from mRNA and cloned into the lambda-ZAP expression vector. IgE screening of the library was performed with a pool of sera from subjects allergic to Cupressaceae. A recombinant 6xHis-tagged Juniperus oxycedrus allergen, Jun o 2, was expressed in Escherichia coli and purified by Ni^2^+ affinity chromatography. It was studied further by immunoblotting inhibition with pollen extracts from other Cupressaceae, Oleaceae, Urticaceae, and Graminaceae. The role of protein-bound calcium on the allergen's IgE-binding capacity was tested in a plaque assay in the presence or absence of EGTA. Results: A cDNA coding for a newly identified Juniperus oxycedrus pollen allergen, rJun o 2, was isolated. The deduced amino acid sequence contained four typical Ca^2^+ binding sites and showed a significant sequence similarity to calmodulins. Depletion of Ca^2^+ in the plaque assay led to a loss of IgE-binding capacity of rJun o 2. Immunoblotting inhibition revealed that J. oxycedrus, J. ashei, Cupressus arizonica, C. sempervirens , Parietaria judaica, Olea europaea, and Lolium perenne pollen extracts were able to inhibit IgE binding to blotted rJun o 2 at different concentrations. Conclusion: rJun o 2 contains IgE-binding epitopes shared by taxonomically unrelated species, and therefore it can be regarded as a new panallergen. These findings could contribute to an explanation for the phenomenon of multiple positive test results in polysensitized patients and the potential symptom-eliciting role of allergenic sources previously not encountered. (J Allergy Clin Immunol 1998;101:772-7)

Background: Cupressaceae allergy is a worldwide pollinosis caused by several species. Some species in limited geographic areas pollinate in fall and winter. Juniperus oxycedrus matches these features. Objective: We sought to define the immunochemical, allergologic, and environmental aspects of J. oxycedrus pollen. Methods: Pollen extract from J. oxycedrus was prepared and characterized by biochemical analysis and human specific IgE binding by means of ELISA and immunoblotting. A 3-year phenological study was conducted to define the pollinating period of J. oxycedrus. Forty consecutive patients allergic to cypress were recruited in two areas and divided into two groups according to their exposure to J. oxycedrus pollen. Clinical evaluation, skin prick tests, and specific IgE determination with J. oxycedrus, J. ashei, and Cupressus arizonica extracts were carried out on both groups. Results:J. oxycedrus pollen extract was obtained, and it showed specific IgE binding and wide cross-reactivity with other Cupressaceae species. The extract caused a positive skin test response in all the patients tested, with about 80% of them having detectable specific IgE. Symptoms related to J. oxycedrus pollen exposure were recorded in 72% of the directly exposed patients and occasionally in 9% of the nonexposed patients. In the Mediterranean coastal area considered, J. oxycedrus was the first Cupressaceae species that started to pollinate at the beginning of November and ended in the first part of December. Conclusions:J. oxycedrus represents a newly characterized pollen species of the Cupressaceae family that cross-reacts with other members of the same family. Subjects with cypress allergy have in vivo and in vitro positive test responses for J. oxycedrus and can show symptoms when exposed to its pollen. Finally, the most important feature of J. oxycedrus is its early pollinating period in southern Europe (Italy), causing a further extension of the cypress pollen season in areas where other Cupressaceae species are present. (J Allergy Clin Immunol 1998;101:755-61.)

In the reaction γγ→KS0KS0resonance production of thef2−is observed. For the radiative withis found. The small number of events in thef2,a2mass region is consistent with the assumption of destructivef2−a2interference. From the mass distribution we determine the relative phases between the tensor mesons. Upper limits on the radiative widths of the glueball candidatesf2(1720) andX(2220) are derived.

A fully implantable artificial pancreas (AP) still represents the holy grail for diabetes treatment. The quest for efficient miniaturized implantable insulin pumps, able to accurately regulate the blood glucose profile and to keep insulin stability, is still persistent. This work describes the design and testing of a microinjection system connected to a variable volume insulin reservoir devised to favor insulin stability during storage. The design, the constitutive materials, and the related fabrication techniques were selected to favor insulin stability by avoiding—or at least limiting—hormone aggregation. We compared substrates made of nylon 6 and Teflon, provided with different surface roughness values due to the employed fabrication procedures (i.e., standard machining and spray deposition). Insulin stability was tested in a worst case condition for 14 days, and pumping system reliability and repeatability in dosing were tested over an entire reservoir emptying cycle. We found that nylon 6 guarantees a higher insulin stability than Teflon and that independent of the material used, larger roughness determines a higher amount of insulin aggregates. A dedicated rotary pump featured by a 1-μL delivery resolution was developed and connected through a proper gear mechanism to a variable volume air-tight insulin reservoir. The microinjection system was also able to operate in a reverse mode to enable the refilling of the implanted reservoir. The developed system represents a fundamental building block toward the development of a fully implantable AP and could be advantageously integrated even in different implantable drug delivery apparatus (e.g., for pain management).

A search for GRBs of energy $E>10$GeV has been performed by studying the single particle counting rate of the EAS array EASTOP. No candidates have been observed in coincidence with 292 BATSE GRBs and the typical upper limits obtained on the energy fluence are $F(10~{\rm GeV}\div 1~{\rm TeV})\sim 10 ^{-4}\div 10^{-3}~{\rm erg}~{\rm cm}^{-2}$for the events with zenith angle $\theta<30^{\circ}$. A single candidate has been observed in the "all sky" survey mode during 1.5 103days.

ARGO-YBJ (Astrophysical Radiation with Ground-based Observatory at YangBaJing) is a detector optimized to study small size air showers. It consists of a layer of Resistive Plate Counters (RPCs) covering an area of ~ 6500 m2and will be located in the Yangbajing Laboratory (Tibet, China) at 4300 m a.s.l. ARGO-YBJ will be devoted to a wide range of fundamental issues in cosmic rays and astroparticle physics, including in particular gamma-ray astronomy and gamma-ray bursts physics in the range $10~{\rm GeV} \div 500~{\rm TeV}$. The sensitivity of ARGO-YBJ to detect high energy GRBs is presented.