A new governance model for the conservation and enhancement of Italian ancient transhumance routes.

Transhumance routes mark the natural path followed by herds during their seasonal movements. These routes are disappearing in many European countries. In Italy, a few fragments survive in Molise, in southern Italy, called Tratturi and run along grassy paths. The Tratturi are considered to be of archaeological, economic and landscape interest, sources of cultural and environmental attraction, protected by a national law. However, the landscapes connected to the Tratturi are disappearing both as a consequence of non-use and as a consequence of the advancement of wooded areas, and this poses a problem related to the governance system that involves many stakeholders, both at international, national and local level. The study, through the use of the combined approach of Stakeholder Basic Analysis and Social Network Analysis, identifies the role of the stakeholders involved, their power, their interest and their ability to support the decision-making system. Furthermore, the paper uses multivariate statistical analysis to detect the key stakeholders. A new model of governance, allowing to preserve the Tratturi and to enhance the services they can provide, is introduced, and it is named Authority for the Conservation and Enhancement of Tratturi (ACET). This study can help to identify appropriate management models for ancient routes based on all stakeholders involved, and more generally to implement shared and co-responsible planning choices in socio-ecological systems.

Modelling food policies in Italian urban agendas in the time of Covid-19: Experiences, challenges and opportunities.

Global food systems are broken and in need of profound change. These imbalances and vulnerabilities are particularly strong in cities, where most of the global population lives and that are at the core of the major challenges linked with food production and consumption. The food system transition needs cities as key game-changers towards more sustainable, equitable, healthier and fairer food systems. Against this backdrop, the present article analyses the role of food policies within urban policies, with a focus on Italian cities. In particular, the article discusses data collected from representatives of 100 municipalities across Northern, Central and Southern Italy. Moreover, it addresses the types of policies and initiatives adopted at the local level, the main obstacles encountered, the role of national and international city networks and the impact of Covid-19 on urban food security, with the aim to identify potential models of urban food policies as a structural component of a broader urban agenda. By doing this, the article aims at filling a research gap in current literature, as it is the first large-scale survey on urban food policies in Italy, identifying models of urban food policies that are already being developed within broader urban development agendas.

How did Italian diversified farms tackle Covid-19 pandemic first wave challenges?

The spread of the Covid-19 pandemic in Italy, in the period March-May 2020, quickly triggered a deep crisis, causing an immediate economic slowdown and consequently a strong contraction in domestic demand and trade. The food supply chain faced severe difficulties, although its anti-cyclical nature allowed for greater resilience compared to other economic sectors. In this framework of ongoing uncertainty, it is important to understand the response of farms to the crisis, and their role in the sustainability of the post-pandemic food supply chain, even for future policy interventions in the short and medium term. The purpose of the paper is to investigate how diversification affected the response of farms to the Covid-19 crisis, and explore whether the changes required by the post-crisis scenario can produce opportunities for their activities. The study investigates a sample of fifteen farms in central Italy through semi-structured interviews, performing a lexicon-based text and sentiment analysis. The results highlight the importance of farm diversification in dealing with the Covid-19 crisis, and emphasise the role of diversified farms for the sustainability of the agri-food system. These results have interesting policy implications, particularly regarding support for the competitiveness of farms by improving sustainable logistics, electronic commerce and exchanges of knowledge and innovations among farmers; these measures should be taken into account to target the future agricultural, rural and food policies, at both national and local level.

Development of advanced therapies in Italy: Management models and sustainability in six Italian cell factories.

On November 10, 2014, the representatives of all six certified Good Manufacturing Practices (GMP) cell factories operating in the Lombardy Region of Italy convened a 1-day workshop in Milan titled "Management Models for the Development And Sustainability of Cell Factories: Public-Private Partnership?" The speakers and panelists addressed not only the many scientific, technological and cultural challenges faced by Lombardy Cell Factories, but also the potential impact of advanced therapy medicinal products (ATMPs) on public health and the role played by translational research in this process. Future perspectives for research and development (R&D) and manufacturing processes in the field of regenerative medicine were discussed as well. This report summarizes the most important issues raised by the workshop participants with particular emphasis on strengths and limitations of the R&D and manufacturing processes for innovative therapeutics in Lombardy and what can be improved in this context while maintaining GMP standards. The participants highlighted several strategies to translate patient-specific advanced therapeutics into scaled manufacturing products for clinical application. These included (i) the development of a synergistic interaction between public and private institutions, (ii) better integration with Italian regulatory agencies and (iii) the creation of a network among Lombardy cell factories and other Italian and European institutions.

IGKV3 proteins as candidate "off-the-shelf" vaccines for kappa-light chain-restricted B-cell non-Hodgkin lymphomas.

PURPOSE: An increasing set of B-cell non-Hodgkin lymphomas (B-NHL) show a biased usage of IGKV3-20 and IGKV3-15 immunoglobulin genes, a feature that could be exploited for the development of ready-to-use, broadly applicable cancer vaccines. EXPERIMENTAL DESIGN: The immunogenic properties of clonal IGKV3-20 and IGKV3-15 proteins were analyzed with particular focus on their ability to elicit cross-reactive responses against molecularly related IGKV proteins expressed by different B-cell lymphoproliferative disorders. RESULTS: IGK+ lymphoma patients show humoral and T-cell responses to IGKV3-20 and IGKV3-15 proteins and IGKV3-specific cytotoxic T lymphocytes (CTL) can be easily induced ex vivo. IGKV3-20-specific CTLs cross-react against different IGKV3 proteins, an effect mediated by the presence of 21 shared, sometimes promiscuous, T-cell epitopes, presented by common HLA class I allele products, thus assuring a broad HLA coverage of IGKV3-based vaccines. Many natural epitope variants are carried by IGK light chains expressed by a broad spectrum of B-NHLs and we show that IGKV3-20-specific CTLs cross-react also against several of these variant epitopes. Both humoral and CTL-specific responses were induced by KLH-conjugated IGKV3-20 protein in HLA-A2-transgenic mice and coinjection of IGKV3-20-specific CTLs with IGKV3-20+ or IGKV3-15+ lymphoma cells into SCID mice totally prevented tumor growth, thus confirming the ability of these effectors to mediate efficient and cross-reactive cytotoxic responses also in vivo. CONCLUSIONS: These results provide the rationale to exploit IGKV3 proteins as "off-the-shelf" vaccines for a large fraction of lymphoma patients.

Cryopreservation of turkey semen by the pellet method: effects of variables such as the extender, cryoprotectant concentration, cooling time and warming temperature on sperm quality determined through principal components analysis.

This study was designed to identify the best pellet cryopreservation procedure for the cryosurvival of turkey semen among 192 different treatments established by variations and permutations of seven conditions used in the freezing/thawing process. These conditions were: diluent (IGGKPh, SPh or Tselutin); dilution rate (1:3 vs. 1:4); cooling time (45 vs. 60 min); dimethylacetamide (DMA) concentration as cryoprotectant (6 vs. 8%); equilibration time in DMA (1 vs. 5 min); semen drop volume (50 vs. 80 µL) and thawing temperature (60 vs. 75 °C). Through principal components analysis (PCA), post-thaw sperm quality data (mobility, viability and membrane functional integrity) were reduced to a single output variable (Sperm Quality) indicating overall post-thaw semen quality. All treatments induced a significant reduction in semen quality after warming (P < 0.01), though one set of seven conditions, or treatment, was identified by PCA to generate the highest Sperm Quality score and a further five treatments yielded a score not significantly different (P > 0.05) from this best score. Although still not fulfilling the requirements for commercial application, our findings serve to identify the critical steps in turkey sperm cryopreservation that need to be assessed in future studies.

Immune signatures in human PBMCs of idiotypic vaccine for HCV-related lymphoproliferative disorders.

Hepatitis C virus (HCV) is one of the major risk factors for chronic hepatitis, which may progress to cirrhosis and hepatocellular carcinoma, as well as for type II mixed cryoglobulinemia (MC), which may further evolve into an overt B-cell non-Hodgkin's lymphoma (NHL). It has been previously shown that B-cell receptor (BCR) repertoire, expressed by clonal B-cells involved in type II MC as well as in HCV-associated NHL, is constrained to a limited number of variable heavy (VH)- and light (VL)-chain genes. Among these, the VK3-20 light chain idiotype has been selected as a possible target for passive as well as active immunization strategy. In the present study, we describe the results of a multiparametric analysis of the innate and early adaptive immune response after ex vivo stimulation of human immune cells with the VK3-20 protein. This objective has been pursued by implementing high-throughput technologies such as multiparameter flow cytometry and multiplex analysis of cytokines and chemokines.

Toward cell therapy using placenta-derived cells: disease mechanisms, cell biology, preclinical studies, and regulatory aspects at the round table.

Among the many cell types that may prove useful to regenerative medicine, mounting evidence suggests that human term placenta-derived cells will join the list of significant contributors. In making new cell therapy-based strategies a clinical reality, it is fundamental that no a priori claims are made regarding which cell source is preferable for a particular therapeutic application. Rather, ongoing comparisons of the potentiality and characteristics of cells from different sources should be made to promote constant improvement in cell therapies, and such comparisons will likely show that individually tailored cells can address disease-specific clinical needs. The principle underlying such an approach is resistance to the notion that comprehensive characterization of any cell type has been achieved, neither in terms of phenotype nor risks-to-benefits ratio. Tailoring cell therapy approaches to specific conditions also requires an understanding of basic disease mechanisms and close collaboration between translational researchers and clinicians, to identify current needs and shortcomings in existing treatments. To this end, the international workshop entitled "Placenta-derived stem cells for treatment of inflammatory diseases: moving toward clinical application" was held in Brescia, Italy, in March 2009, and aimed to harness an understanding of basic inflammatory mechanisms inherent in human diseases with updated findings regarding biological and therapeutic properties of human placenta-derived cells, with particular emphasis on their potential for treating inflammatory diseases. Finally, steps required to allow their future clinical application according to regulatory aspects including good manufacturing practice (GMP) were also considered. In September 2009, the International Placenta Stem Cell Society (IPLASS) was founded to help strengthen the research network in this field.

Modulation of JC virus transcription by C/EBPbeta.

The polyomavirus JC (JCV) causes the demyelinating disease progressive multifocal leukoencephalopathy (PML). Infection by JCV is very common in childhood after which the virus enters a latent state, which is poorly understood. Under conditions of severe immunosuppression, especially AIDS, JCV may reactivate to cause PML. Expression of JC viral proteins is regulated by the JCV non-coding control region (NCCR), which contains an NF-kappaB binding site previously shown to activate transcription. We now report that C/EBPbeta inhibits basal and NF-kappaB-stimulated JCV transcription via the same site. Gel shift analysis showed C/EBPbeta bound to this region in vitro and ChIP assays confirmed this binding in vivo. Further, a ternary complex of NF-kappaB/p65, C/EBPbeta-LIP and JCV DNA could be detected in co-immunoprecipitation experiments. Mutagenesis analysis of the JCV NCCR indicated p65 and C/EBPbeta-LIP bound to adjacent but distinct sites and that both sites regulate basal and p65-stimulated transcription. Thus C/EBPbeta negatively regulates JCV, which together with NF-kappaB activation, may control the balance between JCV latency and activation leading to PML. This balance may be regulated by proinflammatory cytokines in the brain.

Early growth response-1 protein is induced by JC virus infection and binds and regulates the JC virus promoter.

JC virus (JCV) is a human polyomavirus that can emerge from a latent state to cause the cytolytic destruction of oligodendrocytes in the brain resulting in the fatal demyelinating disease, progressive multifocal leukoencephalopathy (PML). Previous studies described a cis-acting transcriptional regulatory element in the JCV non-coding control region (NCCR) that is involved in the response of JCV to cytokines. This consists of a 23 base pair GGA/C rich sequence (GRS) near the replication origin (5112 to +4) that contains potential binding sites for Sp1 and Egr-1. Gel shift analysis showed that Egr-1, but not Sp1, bound to GRS. Evidence is presented that the GRS gel shift seen on cellular stimulation is due to Egr-1. Thus, TPA-induced GRS gel shift could be blocked by antibody to Egr-1. Further, the TPA-induced GRS DNA/protein complex was isolated and found to contain Egr-1 by Western blot. No other Egr-1 sites were found in the JCV NCCR. Functionally, Egr-1 was found to stimulate transcription of JCV late promoter but not early promoter reporter constructs. Mutation of the Egr-1 site abrogated Egr-1 binding and virus with the mutated Egr-1 site showed markedly reduced VP1 expression and DNA replication. Infection of primary astrocytes by wild-type JCV induced Egr-1 nuclear expression that was maximal at 5-10 days post-infection. Finally, upregulation of Egr-1 was detected in PML by immunohistochemistry. These data suggest that Egr-1 induction may be important in the life cycle of JCV and PML pathogenesis.

Skewed T-cell differentiation in patients with indolent non-Hodgkin lymphoma reversed by ex vivo T-cell culture with gammac cytokines.

The unfavorable clinical evolution in indolent non-Hodgkin lymphomas suggests defective control of neoplastic growth by the immune system. To address this issue, we evaluated phenotype, function, and maturation profile of CD4(+) and CD8(+) T cells from peripheral-blood, lymph nodes, or bone marrow of patients with B-cell non-Hodgkin lymphoma (NHL) at diagnosis. T cells from these patients frequently showed an activated but apoptosis-prone phenotype with low frequency of tumor-reactive T cells showing a TH2/Tc2 functional profile in the response to autologous tumor. In peripheral blood or in lymph nodes and bone marrow, and, in comparison to healthy donors, patients' T cells showed a skewed differentiation toward Tnaive and Tcentral memory stages, with low expression of granzyme B and perforin. T-cell culture with autologous tumor in the presence of IL-2, IL-15, and autologous bone marrow-derived cells led to massive T-cell expansion and to differentiation of cytotoxic factor(+) CD8(+) T cells releasing IFN-gamma and killing autologous B-cell tumor in an HLA-class I-restricted fashion. These results suggest impaired T-cell differentiation to effector stage in patients with B-cell NHL, but indicate that T-cell responsiveness to gammac cytokines is retained, thus allowing to promote generation of antitumor T cells for immune intervention.

Dendritic cell viability is decreased after phagocytosis of apoptotic tumor cells induced by staurosporine or vaccinia virus infection.

BACKGROUND AND OBJECTIVES: Dendritic cells (DC) primed with tumor antigens can effectively mediate the regression of a variety of established solid malignancies in both murine and human models. Several experimental studies indicate that apoptotic bodies are an optimal source of tumor antigens for ex vivo priming of DC. However, the clinical use of killed tumor cells as a source of antigens will require an optimal methodology to induce effective tumor cell apoptosis. DESIGN AND METHODS: The goal of this study was to compare the efficiency of three agents for inducing neoplastic B lymphocyte apoptosis; staurosporine, infection by modified vaccinia (MVA) viral particles and ultraviolet C (UVC) radiation. RESULTS: The three methods were finely tuned to induce apoptosis in more than 90% of tumor cells after 24 hours of exposure. However, the viability of monocyte-derived DC, loaded with B-cell tumor apoptotic bodies induced by staurosporine or MVA viral particles, decreased dramatically within 48 hours after phagocytosis of the killed neoplastic cells. The persistence of the apoptosis-inducing agents in the apoptotic bodies and not in the tumor supernatant, was responsible for the observed damage to DC viability. In contrast, DC viability was not affected after uptake of tumor cells killed through UVC-irradiation. Furthermore, B-lymphoblastic cell line (LCL)-specific T cells were reactivated by DC loaded with apoptotic bodies induced by UVC-rays. INTERPRETATION AND CONCLUSIONS: Since the method used to induce tumor cell apoptosis might be detrimental to DC viability, these findings should be considered when designing anticancer vaccination programs.