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This is an open access article distributed under the terms of the Creative Commons Attribution License. ; The activation of G-protein coupled receptors by agonist compounds results in diverse biological responses in cells, such as the endocytosis process consisting in the translocation of receptors from the plasma membrane to the cytoplasm within internalizing vesicles or endosomes. In order to functionally evaluate endocytosis events resulted from pharmacological responses, we have developed an image analysis method -the Q-Endosomes algorithm- that specifically discriminates the fluorescent signal originated at endosomes from that one observed at the plasma membrane in images obtained from living cells by fluorescence microscopy. Mu opioid (MOP) receptor tagged at the carboxy-terminus with yellow fluorescent protein (YFP) and permanently expressed in HEK293 cells was used as experimental model to validate this methodology. Time-course experiments performed with several agonists resulted in different sigmoid curves depending on the drug used to initiate MOP receptor endocytosis. Thus, endocytosis resulting from the simultaneous activation of co-expressed MOP and serotonin 5-HT 2C receptors by morphine plus serotonin was significantly different, in kinetics as well as in maximal response parameters, from the one caused by DAMGO, sufentanyl or methadone. Therefore, this analytical tool permits the pharmacological characterization of receptor endocytosis in living cells with functional and temporal resolution. ; This work was supported by the SAF2010-15663 grant from the Spanish Government (MINECO). ; Peer Reviewed

The activation of G-protein coupled receptors by agonist compounds results in diverse biological responses in cells, such as the endocytosis process consisting in the translocation of receptors from the plasma membrane to the cytoplasm within internalizing vesicles or endosomes. In order to functionally evaluate endocytosis events resulted from pharmacological responses, we have developed an image analysis method –the Q-Endosomes algorithm– that specifically discriminates the fluorescent signal originated at endosomes from that one observed at the plasma membrane in images obtained from living cells by fluorescence microscopy. Mu opioid (MOP) receptor tagged at the carboxy-terminus with yellow fluorescent protein (YFP) and permanently expressed in HEK293 cells was used as experimental model to validate this methodology. Time-course experiments performed with several agonists resulted in different sigmoid curves depending on the drug used to initiate MOP receptor endocytosis. Thus, endocytosis resulting from the simultaneous activation of coexpressed MOP and serotonin 5-HT2C receptors by morphine plus serotonin was significantly different, in kinetics as well as in maximal response parameters, from the one caused by DAMGO, sufentanyl or methadone. Therefore, this analytical tool permits the pharmacological characterization of receptor endocytosis in living cells with functional and temporal resolution. ; This work was supported by the SAF2010-15663 grant from the Spanish Government (MINECO).

Impregnation treatments are one of the alternatives to protect concrete-based building and monuments from weathering degradation. However, it is important to consider the chemical compatibility of the reaction products with the building material. The impregnation product studied here consists of a silica oligomer able to polymerize, by a simple sol-gel process, inside the pore structure of concrete. In this work, we investigate the ability of this impregnation treatment to produce C-S-H gel in contact with cement paste. A complete characterization of the reaction products demonstrated that the silanol groups from silica oligomers react with the portlandite present in the cement paste generating a material with the chemical, structural and morphological features of C-S-H gel. Simultaneously, the 29Si NMR results indicate that the Sisingle bondO units are incorporated into the existing C-S-H, increasing its chain length. These results open the way for a simple concrete structures repairing procedure. ; This project has received funding from the European Union's Horizon 2020 - Research and Innovation Framework Programme under grant agreement No 760858; and from Spanish Government/FEDER-EU under research project MAT2017-84228-R. ; Peer reviewed

Abstract
Phenotypic heterogeneity is an important mechanism for regulating bacterial virulence, where a single regulatory switch is typically activated to generate virulent and avirulent subpopulations. The opportunistic pathogen Acinetobacter baumannii can transition at high frequency between virulent opaque (VIR-O) and avirulent translucent subpopulations, distinguished by cells that form opaque or translucent colonies. We demonstrate that expression of 11 TetR-type transcriptional regulators (TTTRs) can drive cells from the VIR-O opaque subpopulation to cells that form translucent colonies. Remarkably, in a subpopulation of VIR-O cells, four of these TTTRs were stochastically activated in different combinations to drive cells to the translucent state. The resulting translucent subvariants exhibited unique phenotypic differences and the majority were avirulent. Due to their functional redundancy, a quadruple mutant with all four of these TTTRs inactivated was required to observe a loss of switching from the VIR-O state. Further, we demonstrate a small RNA, SrvS, acts as a "rheostat," where the levels of SrvS expression influences both the VIR-O to translucent switching frequency, and which TTTR is activated when VIR-O cells switch. In summary, this work has revealed a new paradigm for phenotypic switching in bacteria, where an unprecedented number of related transcriptional regulators are activated in different combinations to control virulence and generate unique translucent subvariants with distinct phenotypic properties.

That the preservation of twentieth concrete heritage is an area scantly explored can be attributed to a lack of appreciation for such a young material. In most cases conservation is broached from a technical perspective with little regard for heritage value. Ongoing assessment of the condition of structures is the primary strategy to minimise such misguided action. This study involved characterising the condition of the concrete in a number of singular elements forming part of the Eduardo Torroja Institute for Construction Science headquarters at Madrid, Spain, a modernist compound listed by the city of Madrid as a protected asset. The in situ findings using non-destructive and laboratory techniques revealed the core concrete to be in good condition. The surface material, however, exhibits signs of durability issues calling for conservation treatments and techniques compatible with the preservation of the integrity and authenticity of this young heritage material. ; This study was funded by the European Union's Horizon H2020 Research and Innovation Pro-gramme under Grant Agreement No. 760858 (In-novaConcrete) and by the Regional Government of Madrid (TopHeritage-CM; S2018/NMT_4372). The professional support provided by the CSIC's 'Open Heritage: Research and Society (PTI-PAIS)' Interdisciplinary Thematic Platform is gratefully ac-knowledged.

That the preservation of twentieth concrete heritage is an area scantly explored can be attributed to a lack of appreciation for such a young material. In most cases conservation is broached from a technical perspective with little regard for heritage value. Ongoing assessment of the condition of structures is the primary strategy to minimise such misguided action. This study involved characterising the condition of the concrete in a number of singular elements forming part of the Eduardo Torroja Institute for Construction Science headquarters at Madrid, Spain, a modernist compound listed by the city of Madrid as a protected asset. The in situ findings using non-destructive and laboratory techniques revealed the core concrete to be in good condition. The surface material, however, exhibits signs of durability issues calling for conservation treatments and techniques compatible with the preservation of the integrity and authenticity of this young heritage material. ; This study was funded by the European Union's Horizon H2020 Research and Innovation Pro-gramme under Grant Agreement No. 760858 (In-novaConcrete) and by the Regional Government of Madrid (TopHeritage-CM; S2018/NMT_4372). The professional support provided by the CSIC's 'Open Heritage: Research and Society (PTI-PAIS)' Interdisciplinary Thematic Platform is gratefully ac-knowledged.

Nanometer-thick epitaxial Co films intercalated between graphene (Gr) and a heavy metal (HM) substrate are promising systems for the development of spin–orbitronic devices due to their large perpendicular magnetic anisotropy (PMA). A combination of theoretical modeling and experiments reveals the origin of the PMA and explains its behavior as a function of the Co thickness. High quality epitaxial Gr/Con/HM(111) (HM = Pt,Ir) heterostructures are grown by intercalation below graphene, which acts as a surfactant that kinetically stabilizes the pseudomorphic growth of highly perfect Co face-centered tetragonal (fct) films, with a reduced number of stacking faults as the only structural defect observable by high-resolution scanning transmission electron microscopy (STEM). Magneto-optic Kerr effect (MOKE) measurements show that such heterostructures present PMA up to large Co critical thicknesses of about 4 nm (20 ML) and 2 nm (10 ML) for Pt and Ir substrates, respectively. X-ray magnetic circular dichroism (XMCD) measurements show an inverse power law of the anisotropy of the orbital moment with Co thickness, reflecting its interfacial nature, that changes sign at about the same critical values. First principles calculations show that, regardless of the presence of graphene, ideal Co fct films on HM buffers do not sustain PMAs beyond around 6 mLs due to the in-plane contribution of the inner bulk-like Co layers. The large experimental critical thicknesses sustaining PMA can only be retrieved by the inclusion of structural defects that promote a local hcp stacking such as twin boundaries or stacking faults. Remarkably, a layer resolved analysis of the orbital momentum anisotropy reproduces its interfacial nature, and reveals that the Gr/Co interface contribution is comparable to that of the Co/Pt(Ir). ; Financial support from MINECO (Grant Nos. RTI2018-097895-B-C41, RTI2018-097895-B-C42 and RTI2018-097895-B-C43 (FUN-SOC), PID2019-103910GB-I00, FIS2016-78591-C3-1-R and FIS2016-78591-C3-2-R (SKYTRON), PGC2018-098613-B-C21 (SpOrQuMat), PCI2019-111908-2 and PCI2019-111867-2 (FLAGERA 3 grant SOgraphMEM)], from Regional Government of Madrid (grant number P2018/NMT-4321 (NANOMAGCOST-CM)) and from Gobierno Vasco-UPV/EHU (grant numbers GIU18/138 and IT-1246-19). We acknowledge experiments at ALBA BL29 via proposal no. 2019023333. IMDEA-Nanociencia acknowledges support from the "Severo Ochoa" Program for Centres of Excellence in R&D (MINECO, Grant SEV-2016-0686). ; Peer reviewed

Hepatocellular carcinoma (HCC) is the third most common cause of cancer-related deaths worldwide. There is increasing interest in developing specific markers to serve as predictors of response to sorafenib and to guide targeted therapy. Using a sequencing platform designed to study somatic mutations in a selection of 112 genes (HepatoExome), we aimed to characterize lesions from HCC patients and cell lines, and to use the data to study the biological and mechanistic effects of case-specific targeted therapies used alone or in combination with sorafenib. We characterized 331 HCC cases in silico and 32 paired samples obtained prospectively from primary tumors of HCC patients. Each case was analyzed in a time compatible with the requirements of the clinic (within 15 days). In 53% of the discovery cohort cases, we detected unique mutational signatures, with up to 34% of them carrying mutated genes with the potential to guide therapy. In a panel of HCC cell lines, each characterized by a specific mutational signature, sorafenib elicited heterogeneous mechanistic and biological responses, whereas targeted therapy provoked the robust inhibition of cell proliferation and DNA synthesis along with the blockage of AKT/mTOR signaling. The combination of sorafenib with targeted therapies exhibited synergistic anti-HCC biological activity concomitantly with highly effective inhibition of MAPK and AKT/mTOR signaling. Thus, somatic mutations may lead to identify case-specific mechanisms of disease in HCC lesions arising from multiple etiologies. Moreover, targeted therapies guided by molecular characterization, used alone or in combination with sorafenib, can effectively block important HCC disease mechanisms. ; FUNDING: Grants from ISCIII, co-financed by the European Union (FEDER) (PI16/00156), Ramón and Cajal research program from MINECO (RYC-2013-14097) and FUNDACIÓN LUCHAMOS POR LA VIDA to JPV. Grants from ISCIII (RD06/0020/0107-RD012/0036/0060) to MAP. Grant from ISCIII (Ref. PIE15/00079) to JC & JPV. NGD is a recipient of a UC-IDIVAL pre-doctoral fellow. I.V. was also supported by the Ramón and Cajal research program.