Isolation of Mouse Primary Aortic Endothelial Cells by Selection with Specific Antibodies

Endothelial cells (ECs) are key blood-vessel-wall components that play critical roles in the regulation of many physiological processes, including angiogenesis, coagulation, and vascular tone control, and in pathological events such as vessel inflammation and leukocyte infiltration. EC dysfunction is one of the first events associated with the development of atherosclerosis and is sustained throughout progression of the disease. The study of ECs in vitro has become an invaluable tool for investigating these vascular processes at the molecular level, and is widely used in the search for therapeutic targets and strategies. This chapter describes a protocol for the isolation and culture of primary mouse aortic ECs based on antibody-mediated EC selection.Work in V.A.’s laboratory is supported by grants from the Spanish Ministry for Economy and Competitiveness (MINECO) (SAF2013-46663-R), the Fondo Europeo de Desarrollo Regional (FEDER), the Instituto de Salud Carlos III (RD12/0042/0028), the Progeria Research Foundation (Innovator Award 2012, Established Investigator Award 2014), and the European Union (Liphos, Grant Agreement 317916). P.M-S. is supported by an FPU predoctoral fellowship from MINECO. The Centro Nacional de Investigaciones Cardiovasculares (CNIC) is supported by the MINECO and the Pro-CNIC Foundation.S

Isolation of Mouse Primary Aortic Endothelial Cells by Selection with Specific Antibodies

Endothelial cells (ECs) are key blood-vessel-wall components that play critical roles in the regulation of many physiological processes, including angiogenesis, coagulation, and vascular tone control, and in pathological events such as vessel inflammation and leukocyte infiltration. EC dysfunction is one of the first events associated with the development of atherosclerosis and is sustained throughout progression of the disease. The study of ECs in vitro has become an invaluable tool for investigating these vascular processes at the molecular level, and is widely used in the search for therapeutic targets and strategies. This chapter describes a protocol for the isolation and culture of primary mouse aortic ECs based on antibody-mediated EC selection.Work in V.A.’s laboratory is supported by grants from the Spanish Ministry for Economy and Competitiveness (MINECO) (SAF2013-46663-R), the Fondo Europeo de Desarrollo Regional (FEDER), the Instituto de Salud Carlos III (RD12/0042/0028), the Progeria Research Foundation (Innovator Award 2012, Established Investigator Award 2014), and the European Union (Liphos, Grant Agreement 317916). P.M-S. is supported by an FPU predoctoral fellowship from MINECO. The Centro Nacional de Investigaciones Cardiovasculares (CNIC) is supported by the MINECO and the Pro-CNIC Foundation.S

Loss of p27 phosphorylation at Ser10 accelerates early atherogenesis by promoting leukocyte recruitment via RhoA/ROCK

Reduced phosphorylation of the tumor suppressor p27(Kip1) (p27) at serine 10 (Ser10) is a hallmark of advanced human and mouse atherosclerosis. Apolipoprotein E-null mice defective for this posttranslational modification (apoE(-/-)p27Ser10Ala) exhibited increased atherosclerosis burden at late disease states. Here, we investigated the regulation of p27 phosphorylation in Ser10 at the very initial stages of atherosclerosis and its impact on endothelial-leukocyte interaction and early plaque formation. Hypercholesterolemia in fat-fed apoE(-/-) mice is associated with a rapid downregulation of p27-phospho-Ser10 in primary endothelial cells (ECs) and in aorta prior to the development of macroscopically-visible lesions. We find that lack of p27 phosphorylation at Ser10 enhances the expression of adhesion molecules in aorta of apoE(-/-) mice and ECs, and augments endothelial-leukocyte interactions and leukocyte recruitment in vivo. These effects correlated with increased RhoA/Rho-associated coiled-coil containing protein kinase (ROCK) signaling in ECs, and inhibition of this pathway with fasudil reduced leukocyte-EC interactions to control levels in the microvasculature of p27Ser10Ala mice. Moreover, apoE(-/-)p27Ser10Ala mice displayed increased leukocyte recruitment and homing to atherosusceptible arteries and augmented early plaque development, which could be blunted with fasudil. In conclusion, our studies demonstrate a very rapid reduction in p27-phospho-Ser10 levels at the onset of atherogenesis, which contributes to early plaque build-up through RhoA/ROCK-induced integrin expression in ECs and enhanced leukocyte recruitment.Spanish Ministry of Economy and Competitivity (MINECO) [SAF2010-16044, SAF2013-46663-R]; Fondo Europeo de Desarrollo Regional (FEDER); Instituto de Salud Carlos III [RD12/0042/0028]; FPU predoctoral fellowship from MINECO; MINECO; Pro-CNIC FoundationS

A Cryptographic Layer for the Interoperability of CBDC and Cryptocurrency Ledgers

Cryptocurrencies are used in several, distinct use cases, thereby sustaining the existence of many ledgers that are heterogeneous in terms of design and purpose. In addition, the interest of central banks in deploying Central Bank Digital Currency (CBDC) has spurred a blooming number of conceptually different proposals from central banks and academia. As a result of the diversity of cryptocurrency and CBDC ledgers, interoperability, i.e., the seamless transfer of value between users that operate on different ledgers, has become an interesting research problem. In fact, interoperability has been explored both in CBDC and cryptocurrencies, and numerous proposals exist. However, these proposals are tailored to the characteristics of the ledgers for which they are designed. For instance, some rely on trusted hardware, others rely on the scripting capabilities of the underlying ledger or on specific cryptographic assumptions of the transaction authorization mechanism (e.g., adaptor signatures), while others rely on a trusted entity. This fragmentation results in the repetitive development of interoperablity protocols that address the same applications across the various ledgers. In this work, we propose an alternative approach: decouple the transaction authorization details of each ledger from the definition of cross-ledger applications. To do so, we define a middle layer that abstracts the core functionality for authorizing transactions in any ledger and the security notions of interest. This middle layer serves two purposes: (i) it becomes the main cryptographic building block to (re)define cross-ledger applications in a ledger agnostic manner; (ii) for any ledger that exists (and the new to come), it suffices to prove that it is a secure instance of the middle layer to be compatible with cross-ledger protocols. We define two new primitives for our middle layer, the basic payment ledger (BL) and the conditional payment ledger (CL). We prove that the two most common transaction authorization mechanism (digital signatures and zero knowledge proofs) are secure constructions for BL. We also prove that common smart contracts (e.g., HTLC and PTLC) and the combination of adaptor signatures with verifiable timelock puzzles are also secure constructions for CL. Finally, we discuss how to design some popular applications (e.g. atomic swaps between ledgers) using our middle layer

Human-Robot Cooperation in Surface Inspection Aerial Missions

The goal of the work presented in this paper is to facilitate the cooperation between human opera- tors and aerial robots to perform surface inspec- tion missions. Our approach is based on a model of human collaborative control with a mixed ini- tiative interaction. In the paper, we present our human-robot cooperation model based on the combination of a supervisory mode and an as- sistance mode with a set of interaction patterns. We developed a software system implementing this interaction model and carried out several real flight experiments that proved that this ap- proach can be used in aerial robotics for sur- face inspection missions (e.g., in vision based indoor missions). Compared to a conventional tele-operated inspection system, the solution pre- sented in this paper gives more autonomy to the aerial systems, reducing the cognitive load of the operator during the mission development

Age-dependent defective TGF-beta1 signaling in patients undergoing coronary artery bypass grafting

Background: Transforming growth factor beta (TGF-beta 1) is a pleiotropic cytokine, which is deregulated in atherosclerosis; however the role of age in this process is unknown. We aimed to assess whether TGF-beta 1 signaling is affected by age. Methods: Vascular smooth muscle cells (VSMC) were obtained from patients undergoing abdominal surgery. Levels of TGF-beta 1 were measured by ELISA in sera from 169 patients undergoing coronary artery bypass grafting (CABG). The p27 expression was determined by Western blot from internal mammary arteries (IMA) obtained from CABG patients (n = 13). In VSMC from these patients undergoing abdominal surgery, secretion of TGF-beta 1 was determined by ELISA of cell-conditioned media. Results: In VSMC from aged patients we observed a lower TGF-beta 1 secretion, measured as TGF-beta 1 concentration in cell conditioned medium (p < 0.001). This effect was correlated to an age-dependent decrease of p27 expression in IMA from aged CABG patients. In a similar manner, there was an age-dependent decrease of serum TGF-beta 1 levels in CABG patients (p = 0.0195). Conclusions: VSMC from aged patients showed a higher degree of cellular senescence and it was associated to a lower TGF-beta 1 secretion and signaling.S

Grasa de sobrepaso en ovejas con diferente espesor de grasa dorsal, respuesta hormonal y principales variables reproductivas

El objetivo de este estudio fue evaluar el efecto de la grasa de sobrepaso en ovejas con bajo (1 a 2 mm) y alto (3 a 4 mm) espesor de grasa dorsal y sincronización de estro (inicio y duración), niveles séricos de hormona luteinizante (LH), estradiol (E2), progesterona (P4 e insulina (INS), porcentaje de gestación y prolificidad. Cincuenta y nueve ovejas, con dos niveles de grasa dorsal determinado mediante ultrasonografía, bajo y alto (GDb, GDa), sin y con la adición de 150 g de grasa de sobrepaso (0 y 150 g), respectivamente, se asignaron a los siguientes grupos: GDb+0 g (n = 16), GDb+150 g (n = 14), GDa+0 g (n = 14) y GDa+150 g (n = 15). Las ovejas se sincronizaron con esponjas de acetato de fluorogesterona (FGA, 20 mg) por 12 d y, dos días antes de su remoción se aplicó 15 mg de PGF2. El diseño fue completamente al azar con un arreglo factorial 2X2, los resultados fueron analizados mediante el paquete estadístico SAS. No se encontraron diferencias (P>0,05) para las variables inicio y duración del estro, inicio y duración del pico pre-ovulatorio de LH y prolificidad, por efecto de la adición de la grasa; sin embargo, la amplitud del pico de LH y el porcentaje de gestación fueron diferentes entre tratamientos (P<0,05). La concentración de P4en suero fue mayor (P<0,05) en ovejas sin la adición de grasa (0 g). Las concentraciones de E2 e INS se incrementaron (P<0,05) en ovejas con GDa. Se concluye que la adición de grasa de sobrepaso no modificó la respuesta en las variables reproductivas, pero si disminuyó la concentración de P4. Por otro lado, las concentraciones de E2 e INS se incrementaron en ovejas con GDa, lo cual se atribuye a un mejor estado metabólico, nutricional y corporal del animal

Die Rattenfänger von Balingen : wie die Bundeswehr mit Musik neuen Nachwuchs sucht

<div><p>Nuclear lamins are important structural and functional proteins in mammalian cells, but little is known about the mechanisms and cofactors that regulate their traffic into the nucleus. Here, we demonstrate that trafficking of lamin A, but not lamin B1, and its assembly into the nuclear envelope are regulated by sorting nexin 6 (SNX6), a major component of the retromer that targets proteins and other molecules to specific subcellular locations. SNX6 interacts with lamin A <i>in vitro</i> and <i>in vivo</i> and links it to the outer surface of the endoplasmic reticulum in human and mouse cells. SNX6 transports its lamin A cargo to the nuclear envelope in a process that takes several hours. Lamin A protein levels in the nucleus augment or decrease, respectively, upon gain or loss of SNX6 function. We further show that SNX6-dependent lamin A nuclear import occurs across the nuclear pore complex via a RAN-GTP-dependent mechanism. These results identify SNX6 as a key regulator of lamin A synthesis and incorporation into the nuclear envelope.</p></div

Impacts of agricultural irrigation on groundwater salinity

Agricultural irrigation represents the main use of global water resources. Irrigation has an impact on the environment, and scientific evidence suggests that it inevitably leads to salinization of both soil and aquifers. The effects are most pronounced under arid and semi-arid conditions. In considering the varied impacts of irrigation practices on groundwater quality, these can be classed as either direct—the direct result of applying water and accompanying agrochemicals to cropland—or indirect—the effects of irrigation abstractions on groundwater hydrogeochemistry. This paper summarizes and illustrates through paradigmatic case studies the main impacts of irrigation practices on groundwater salinity. Typically, a diverse range of groundwater salinization processes operating concomitantly at different time scales (from days to hundreds of years) is involved in agricultural irrigation. Case studies suggest that the existing paradigm for irrigated agriculture of focusing mainly on crop production increases has contributed to widespread salinization of groundwater resources.This study was supported by CEIMAR “Campus de Excelencia Internacional del Mar” and the projects CADESEM (Grant Number P11-RNM-8115), Junta de Andalucia Regional Government, Spain, and KARSTCLIMA (Grant Number CGL2015-71510-R) and ACUIDESAL (Grant Number CGL2015-67273-R), Spanish Ministry of Science and Technology—FEDER