Tumour Infiltrating (TINK) and Tumour Associated (TANK) Natural Killer cells: role in colorectal cancer (CRC) progression and angiogenesis.

Tumor infiltrating immune cells often show a skewed phenotype that reflects attenuation of anti- tumor activity and enhancement of pro-tumor and pro-angiogenic activities. We previously reported that NKs from Non Small Cell Lung Cancer patients are able to acquire the decidual-like CD56+CD16-VEGFhighPlGFhighIL- 8+IFN\uf067low phenotype and promote angiogenesis in vitro. Here, we extended our findings to colorectal cancer (CRC) to verify whether the TINK/TANK polarization may represent a crucial hallmark of solid tumours. We found that CD56+CD16- NK cells predominate in CRC adjacent and tumor tissues, show decreased NKG2D surface expression and impaired cytotoxicity. Further, TINK/TANKs from CRC patients express the decidual NK markers CD9 and CD49a. Secretomic and flow cytometry on CRC peripheral blood NK cells revealed the up-regulation of several pro-angiogenic factors. Molecularly, the STAT-3 and STAT-5 pathway activation was observed in TANKs, suggesting the potential involvement of these signaling pathways in the angiogenic switch. CM by FACS sorted NKs from CRC patients were able to induce HUVEC proliferation, migration, adhesion and the formation of capillary like structures. These functional alterations are related with molecular changes in HUVECs, that include the phosphorylation of AMPK\u3b1, GSK-3\uf062, P70 S6 Kinase and S6 ribosomal protein. Our data demonstrate that TINK/TANKS from CRC patients are switched toward a proangiogenic/ pro-tumor phenotype and function. We propose that TINK/TANKs could represent a relevant biomarker for CRC progression

Role of CD40 and Zeb1 in Shaping Normal and Leukemic Bone Marrow Niche

Acute myeloid leukaemia (AML) is an aggressive disease impairing normal haematopoiesis. Beyond the genomic landscape and the mutational profile of AML cells, increasing evidence demonstrated that bone marrow (BM) microenvironment contributes to leukemia development and, in particular, the immunologic landscape is emerging as a crucial component of leukemic microenvironment correlating with clinical parameters. Leukemic cells express different antigens that are able to activate T-cells triggering an anti-leukemia immune response. Thus, we hypothesized that immunogenic leukemia cells could set up mechanisms for immune escape including the establishment of immunosuppression. To escape from immune recognition leukemic cells could instruct neighbouring mesenchymal (MSCs) cells to trigger inhibitory T cell pathways through the up-regulation of CD40 or autonomously gain immunosuppressive properties activating epithelial to mesenchymal transition-related programs (i.e. ZEB-1). Hence, the aim of this study is to evaluate the impact of ZEB-1 expression on AML cell ability to mold an immunosuppressive microenvironment. Moreover, we also evaluated the role of CD40 on BM-MSCs in shaping the BM tolerogenic niche during leukemia development and in bone marrow transplantation. To this aim we firstly analysed the gene expression profile of 61 AML cases that were divided in “ZEB1low” and “ZEB1high”. The two groups showed difference between the karyotypic features and expressed different levels of immunomodulatory markers. Analysis performed using public datasets, showed that ZEB1high patients displayed a worse overall survival compared with ZEB1low patients. Immunohistochemistry analysis performed on archival BM biopsy showed that ZEB1high cases also showed an expansion of CD3+IL-17A+ cells. To unveil the real contribution of ZEB-1 in shaping a BM microenvironment, we moved from humans to mouse models. Zeb-1 expressing or silenced C1498 murine AML cells were injected intra-bone in immunocompetent mice. Mice injected with Zeb-1+ cells showed a higher engraftment compared with mice injected with silenced cells. This phenotype correlated with an expansion of Th17 and an increase of TIM3+PD1+CD8+ cells within the BM of Zeb-1+ cell injected mice. Molecular studies highlighted that Zeb-1 silencing in C1498 induced a significant reduction of Il-6, Tgfβ and Il-23, cytokines involved in Th17 differentiation and maintenance. In turn, IL-17A promoted AML cell proliferation and aggressiveness in-vitro. Moreover, Zeb-1 silenced cells showed a reduced expression of Arg1 that has been reported to negatively affect the activation of CD8+ T cells leading to the increase of cytotoxic CD8+ cells in mice injected with silenced C1498. In addition, Zeb-1 was able to up-regulate the expression of Cd40 and Ido-1 on BM-MSCs. Given the importance of stromal Cd40 expression in sustaining the progression of other haematological diseases, we hypothesized that its expression can be able to affect AML development. To this end, we injected C1498 cells in Cd40-KO mice and we showed that Cd40 deficiency was associated with graft failure. To evaluate the immune-regulatory role of stromal Cd40, we performed BM transplantation experiments. We observed that Cd40 deficiency leads to the persistent T-cell activation with the expansion of CD4+ INF-γ/TNF-α effector T cells and with the parallel depletion of regulatory T cells due to an increase production of Ox40l. Our results suggest that ZEB1 expression could characterize a group of particularly aggressive and chemotherapy resistant AML whose poor outcome is the result of both intrinsic clone aggressiveness and the immunosuppressive microenvironment initiated by ZEB1

acetyl l carnitine is an anti angiogenic agent targeting the vegfr2 and cxcr4 pathways

Abstract Carnitines play an important role in the energy exchange in cells, and are involved in the transport of fatty acids across the inner mitochondrial membrane. l -Acetylcarnitine (ALCAR) is an acetic acid ester of carnitine that has higher bioavailability and is considered a fat-burning energizer supplement. We previously found that in serum samples from prostate cancer (PCa) patients, 3 carnitine family members were significantly decreased, suggesting a potential protective role of carnitine against PCa. Several studies support beneficial effects of carnitines on cancer, no study has investigated the activities of carnitine on tumor angiogenesis. We examined whether ALCAR acts as an "angiopreventive" compound and studied the molecular mechanisms involved. We found that ALCAR was able to limit inflammatory angiogenesis by reducing stimulated endothelial cell and macrophage infiltration in vitro and in vivo. Molecularly, we show that ALCAR downregulates VEGF, VEGFR2, CXCL12, CXCR4 and FAK pathways. ALCAR blocked the activation of NF-κB and ICAM-1 and reduced the adhesion of a monocyte cell line to endothelial cells. This is the first study showing that ALCAR has anti-angiogenic and anti-inflammatory properties and might be an attractive candidate for cancer angioprevention

Neutrophil and Natural Killer Cell Interactions in Cancers: Dangerous Liaisons Instructing Immunosuppression and Angiogenesis

The tumor immune microenvironment (TIME) has largely been reported to cooperate on tumor onset and progression, as a consequence of the phenotype/functional plasticity and adaptation capabilities of tumor-infiltrating and tumor-associated immune cells. Immune cells within the tumor micro (tissue-local) and macro (peripheral blood) environment closely interact by cell-to-cell contact and/or via soluble factors, also generating a tumor-permissive soil. These dangerous liaisons have been investigated for pillars of tumor immunology, such as tumor associated macrophages and T cell subsets. Here, we reviewed and discussed the contribution of selected innate immunity effector cells, namely neutrophils and natural killer cells, as \u201csoloists\u201d or by their \u201cdangerous liaisons\u201d, in favoring tumor progression by dissecting the cellular and molecular mechanisms involved

Biological Aspects, Advancements and Techno-Economical Evaluation of Biological Methanation for the Recycling and Valorization of CO2

Nowadays, sustainable and renewable energy production is a global priority. Over the past decade, several Power-to-X (PtX) technologies have been proposed to store and convert the surplus of renewable energies into chemical bonds of chemicals produced by different processes. CO2 is a major contributor to climate change, yet it is also an undervalued source of carbon that could be recycled and represents an opportunity to generate renewable energy. In this context, PtX technologies would allow for CO2 valorization into renewable fuels while reducing greenhouse gas (GHG) emissions. With this work we want to provide an up-to-date overview of biomethanation as a PtX technology by considering the biological aspects and the main parameters affecting its application and scalability at an industrial level. Particular attention will be paid to the concept of CO2-streams valorization and to the integration of the process with renewable energies. Aspects related to new promising technologies such as in situ, ex situ, hybrid biomethanation and the concept of underground methanation will be discussed, also in connection with recent application cases. Furthermore, the technical and economic feasibility will be critically analyzed to highlight current options and limitations for implementing a sustainable process

DIAGNÓSTICO AMBIENTAL DE UMA INSTITUIÇÃO DE ENSINO TÉCNICO, INTEGRADO E SUPERIOR

As instituições de ensino, assim como todas organizações, apresentam aspectos ambientais que se não geridos corretamente, acarretam impactos ambientais significativos. Nos dias atuais, as instituições de ensino tem se revelado grandes precursoras no processo de desenvolvimento tecnológico, na preparação de estu­dantes e fornecimento de informações e conhecimento básico para a construção do desenvolvimento sustentável e de uma sociedade justa, porém, ao seguir esta premissa, torna-se indispensável que essas organizações comecem a incorporar os princípios e práticas da sustentabilidade em suas atividades. O presente trabalho aborda a diagnose ambiental de uma instituição pública de ensino técnico, integrado e superior através da identificação dos aspectos ambientais, visando obter dados para embasar ações futuras voltadas a responsabilidade socioambiental na instituição. A instituição estudada está localizada na região sudeste do Estado do Pará. Para execução do trabalho, utilizou-se o método de pesquisa descritiva; levaram-se em conta os aspectos ambientais diretos associados às atividades e serviços da instituição. A Instituição apresenta aspectos ambientais comuns do encontrados em outras instituições de ensino, como: consumo de papel, água, energia elétrica e geração de resíduos sólidos e consumo e queima de combustível. Os aspectos encontrados são passíveis de controle a partir de medidas de pouca complexidade

Human Dental Pulp Mesenchymal Stem Cell-Derived Soluble Factors Combined with a Nanostructured Scaffold Support theGeneratio of a Vascular Network In Vivo

Among all strategies directed at developing new tools to support re-vascularization of damaged tissues, the use of pro-angiogenic soluble factors, derived from mesenchymal stem cells (MSCs), appears a promising approach for regenerative medicine. Here, we compared the feasibility of two devices, generated by coupling soluble factors of human dental pulp mesenchymal stem cells (DPSCs), with a nanostructured scaffold, to support angiogenesis once transplanted in mice. DPSCs were obtained from impacted wisdom tooth removal, usually considered surgical waste material. After 28 days, we verified the presence of active blood vessels inside the scaffold through optical and scansion electron microscopy. The mRNA expression of surface antigens related to macrophage polarization (CD68, CD80, CD86, CD163, CD206), as well as pro-angiogenic markers (CD31, CD34, CD105, Angpt1, Angpt2, CDH5) was evaluated by real-time PCR. Our results demonstrate the capability of DPSC–scaffold and DPSC soluble factors–scaffold to support angiogenesis, similarly to adipose stem cells, whereas the absence of blood vessels was found in the scaffold grafted alone. Our results provide evidence that DPSC-conditioned medium can be proposed as a cell-free preparation able to support angiogenesis, thus, providing a relevant tool to overcome the issues and restrictions associated with the use of cells

Discovery of the 3-Amino-1,2,4-triazine-Based Library as Selective PDK1 Inhibitors with Therapeutic Potential in Highly Aggressive Pancreatic Ductal Adenocarcinoma

Pyruvate dehydrogenase kinases (PDKs) are serine/threonine kinases, that are directly involved in altered cancer cell metabolism, resulting in cancer aggressiveness and resistance. Dichloroacetic acid (DCA) is the first PDK inhibitor that has entered phase II clinical; however, several side effects associated with weak anticancer activity and excessive drug dose (100 mg/kg) have led to its limitation in clinical application. Building upon a molecular hybridization approach, a small library of 3-amino-1,2,4-triazine derivatives has been designed, synthesized, and characterized for their PDK inhibitory activity using in silico, in vitro, and in vivo assays. Biochemical screenings showed that all synthesized compounds are potent and subtype-selective inhibitors of PDK. Accordingly, molecular modeling studies revealed that a lot of ligands can be properly placed inside the ATP-binding site of PDK1. Interestingly, 2D and 3D cell studies revealed their ability to induce cancer cell death at low micromolar doses, being extremely effective against human pancreatic KRAS mutated cancer cells. Cellular mechanistic studies confirm their ability to hamper the PDK/PDH axis, thus leading to metabolic/redox cellular impairment, and to ultimately trigger apoptotic cancer cell death. Remarkably, preliminary in vivo studies performed on a highly aggressive and metastatic Kras-mutant solid tumor model confirm the ability of the most representative compound 5i to target the PDH/PDK axis in vivo and highlighted its equal efficacy and better tolerability profile with respect to those elicited by the reference FDA approved drugs, cisplatin and gemcitabine. Collectively, the data highlights the promising anticancer potential of these novel PDK-targeting derivatives toward obtaining clinical candidates for combatting highly aggressive KRAS-mutant pancreatic ductal adenocarcinomas

“Innovative high pressure/high temperature, multi-sensing bioreactors system for microbial risk assessment in underground hydrogen storage”

This study addresses the microbial risks associated with Underground Hydrogen Storage (UHS), a critical component in the transition towards renewable energy systems, by employing an innovative multi-reactor system (Bio-xplorer) to simulate UHS conditions in two Italian reservoirs. The microbiological risk assessment (MRA) of Reservoir A and B was evaluated by subjecting them to gas mixtures of 10 % H2 and 90 % CH4, and 99 % H2 and 1 % CO2, respectively. In Reservoir A, the stability of pressure and temperature, the negligible optical density, and lack of microbial metabolites suggested a low risk of microbial activation. Molecular analyses confirmed the absence of sulphate- reducing bacteria (SRB) and limited growth of hydrogenotrophic methanogens (HM). Similarly, in Reservoir B, the absence of SRB and limited occurrence of HM indicated a low microbiological risk. Overall, the present work supports the safe and efficient implementation of UHS, a promising mitigation technique for climate change, using an innovative tool for MRA

Structural Manipulations of Marine Natural Products Inspire a New Library of 3-Amino-1,2,4-Triazine PDK Inhibitors Endowed with Antitumor Activity in Pancreatic Ductal Adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) is one of the main aggressive types of cancer, characterized by late prognosis and drug resistance. Among the main factors sustaining PDAC progression, the alteration of cell metabolism has emerged to have a key role in PDAC cell proliferation, invasion, and resistance to standard chemotherapeutic agents. Taking into account all these factors and the urgency in evaluating novel options to treat PDAC, in the present work we reported the synthesis of a new series of indolyl-7-azaindolyl triazine compounds inspired by marine bis-indolyl alkaloids. We first assessed the ability of the new triazine compounds to inhibit the enzymatic activity of pyruvate dehydrogenase kinases (PDKs). The results showed that most of derivatives totally inhibit PDK1 and PDK4. Molecular docking analysis was executed to predict the possible binding mode of these derivatives using ligand-based homology modeling technique. Evaluation of the capability of new triazines to inhibit the cell growth in 2D and 3D KRAS-wild-type (BxPC-3) and KRAS-mutant (PSN-1) PDAC cell line, was carried out. The results showed the capacity of the new derivatives to reduce cell growth with a major selectivity against KRAS-mutant PDAC PSN-1 on both cell models. These data demonstrated that the new triazine derivatives target PDK1 enzymatic activity and exhibit cytotoxic effects on 2D and 3D PDAC cell models, thus encouraging further structure manipulation for analogs development against PDA