G protein‐coupled chemokine receptors and their peptidergic ligands are interesting therapeutic targets due to their involvement in various immune‐related diseases, including rheumatoid arthritis, ...multiple sclerosis, inflammatory bowel disease, chronic obstructive pulmonary disease, HIV‐1 infection and cancer. To tackle these diseases, a lot of effort has been focused on discovery and development of small‐molecule chemokine receptor antagonists. This has been rewarded by the market approval of two novel chemokine receptor inhibitors, AMD3100 (CXCR4) and Maraviroc (CCR5) for stem cell mobilization and treatment of HIV‐1 infection respectively. The recent GPCR crystal structures together with mutagenesis and pharmacological studies have aided in understanding how small‐molecule ligands interact with chemokine receptors. Many of these ligands display behaviour deviating from simple competition and do not interact with the chemokine binding site, providing evidence for an allosteric mode of action. This review aims to give an overview of the evidence supporting modulation of this intriguing receptor family by a range of ligands, including small molecules, peptides and antibodies. Moreover, the computer‐assisted modelling of chemokine receptor–ligand interactions is discussed in view of GPCR crystal structures. Finally, the implications of concepts such as functional selectivity and chemokine receptor dimerization are considered.
LINKED ARTICLES This article is part of a themed section on the Molecular Pharmacology of G Protein‐Coupled Receptors (GPCRs). To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2012.165.issue‐6. To view the 2010 themed section on the same topic visit http://onlinelibrary.wiley.com/doi/10.1111/bph.2010.159.issue‐5/issuetoc
Induced pluripotent stem cells (iPSC) offer an unprecedented opportunity to model human disease in relevant cell types, but it is unclear whether they could successfully model age‐related diseases ...such as Parkinson's disease (PD). Here, we generated iPSC lines from seven patients with idiopathic PD (ID‐PD), four patients with familial PD associated to the G2019S mutation in the Leucine‐Rich Repeat Kinase 2 (LRRK2) gene (LRRK2‐PD) and four age‐ and sex‐matched healthy individuals (Ctrl). Over long‐time culture, dopaminergic neurons (DAn) differentiated from either ID‐PD‐ or LRRK2‐PD‐iPSC showed morphological alterations, including reduced numbers of neurites and neurite arborization, as well as accumulation of autophagic vacuoles, which were not evident in DAn differentiated from Ctrl‐iPSC. Further induction of autophagy and/or inhibition of lysosomal proteolysis greatly exacerbated the DAn morphological alterations, indicating autophagic compromise in DAn from ID‐PD‐ and LRRK2‐PD‐iPSC, which we demonstrate occurs at the level of autophagosome clearance. Our study provides an iPSC‐based in vitro model that captures the patients' genetic complexity and allows investigation of the pathogenesis of both sporadic and familial PD cases in a disease‐relevant cell type.
The aim of this paper is to study how grape ripeness and ethanol concentration affect the extraction of color and phenolic compounds from skins and seeds during the maceration/fermentation process. ...Simulated maceration assays were carried out with the grapes at three stages of berry development (vitis vinifera cv. Tempranillo) and different percentages of ethanol in the maceration media. Both ripeness and ethanol content have a considerable effect on the extraction of color and phenolic compounds. Of these two factors, ripeness increases the extractability most. The presence of ethanol in the medium facilitates anthocyanin and especially proanthocyanidin extraction, but it also decreases copigmentation phenomena, which can decrease the color intensity. The higher the ethanol concentration is in the maceration media, the higher the astringency of proanthocyanidins. Keywords: Anthocyanin; proanthocyanidins; ethanol; ripeness; extraction; HPLC; grapes; skins; seeds
Huntington's disease (HD) is a neurodegenerative disorder caused by an abnormal expansion of a CAG repeat encoding a polyglutamine tract in the huntingtin (Htt) protein. The mutation leads to ...neuronal death through mechanisms which are still unknown. One hypothesis is that mitochondrial defects may play a key role. In support of this, the activity of mitochondrial complex II (C-II) is preferentially reduced in the striatum of HD patients. Here, we studied C-II expression in different genetic models of HD expressing N-terminal fragments of mutant Htt (mHtt). Western blot analysis showed that the expression of the 30 kDa Iron-Sulfur (Ip) subunit of C-II was significantly reduced in the striatum of the R6/1 transgenic mice, while the levels of the FAD containing catalytic 70 kDa subunit (Fp) were not significantly changed. Blue native gel analysis showed that the assembly of C-II in mitochondria was altered early in N171-82Q transgenic mice. Early loco-regional reduction in C-II activity and Ip protein expression was also demonstrated in a rat model of HD using intrastriatal injection of lentiviral vectors encoding mHtt. Infection of the rat striatum with a lentiviral vector coding the C-II Ip or Fp subunits induced a significant overexpression of these proteins that led to significant neuroprotection of striatal neurons against mHtt neurotoxicity. These results obtained in vivo support the hypothesis that structural and functional alterations of C-II induced by mHtt may play a critical role in the degeneration of striatal neurons in HD and that mitochondrial-targeted therapies may be useful in its treatment.
Opioids, such as morphine, are the most clinically useful class of analgesic drugs for the treatment of acute and chronic pain. However, the use of opioids is greatly limited by the development of ...severe adverse side effects. Consequently, drug discovery efforts have been directed towards improving the therapeutic profile of opioid‐based treatments. Opioid receptors are members of the family of GPCRs. As such, the recent GPCR paradigms of biased agonism and allosterism may provide novel avenues for more effective analgesics. Biased agonism (or functional selectivity) has been described for all the opioid receptor family members. Furthermore, the first allosteric modulators of opioid receptors have very recently been described. However, identification and quantification of biased agonism in a manner that is informative to medicinal chemists and drug discovery programmes still remains a challenge. In this review, we examine the progress, to date, towards identification and quantification of biased agonism and allosterism at the μ‐opioid receptor in the context of its implications for the discovery of better and safer analgesics.
Linked Articles
This article is part of a themed section on Opioids: New Pathways to Functional Selectivity. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-2
Ocean acidification is receiving increasing attention because of its potential to affect marine ecosystems. Rare CO2 vents offer a unique opportunity to investigate the response of benthic ecosystems ...to acidification. However, the benthic habitats investigated so far are mainly found at very shallow water (less than or equal to 5 m depth) and therefore are not representative of the broad range of continental shelf habitats. Here, we show that a decrease from pH 8.1 to 7.9 observed in a CO2 vent system at 40 m depth leads to a dramatic shift in highly diverse and structurally complex habitats. Forests of the kelp Laminaria rodriguezii usually found at larger depths (greater than 65 m) replace the otherwise dominant habitats (i.e. coralligenous outcrops and rhodolith beds), which are mainly characterized by calcifying organisms. Only the aragonite-calcifying algae are able to survive in acidified waters, while high-magnesium-calcite organisms are almost completely absent. Although a long-term survey of the venting area would be necessary to fully understand the effects of the variability of pH and other carbonate parameters over the structure and functioning of the investigated mesophotic habitats, our results suggest that in addition of significant changes at species level, moderate ocean acidification may entail major shifts in the distribution and dominance of key benthic ecosystems at regional scale, which could have broad ecological and socio-economic implications.
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•High-resolution XRF-CS data were calibrated to obtain quantitative concentration.•Calibration method increases the correlation coefficient and reduces the data deviation.•WLS ...regression method reduced significantly the errors of calibrated element concentration.•Low errors allow assessing metal pollution levels according to sediment quality guidelines.•Errors of the calibrated element concentration must be evaluated in future calibration studies.
X-ray fluorescence core scanners (XRF-CS) allow rapid, non-destructive, continuous and high-resolution analyses of the elemental composition of sediment cores, providing large sets of semi-quantitative data. These data can be converted to quantitative data through the linear regression approach using a relatively small number of discrete samples analyzed by techniques providing absolute concentrations. However, a precise characterization of the errors associated with the linear function is required to evaluate the quality of the calibrated element concentrations. Here we present a calibration of high-resolution XRF-CS for six metals (Ti, Mn, Fe, Zn, Pb and As) measured in heavily contaminated marine deposits so that absolute concentrations are obtained. In order to determine the best linear function for conversion of XRF data, we have tested three regression methods: the ordinary least-squares (OLS), which does not consider the standard error in any variable (x and y), the weighted ordinary least-squares (WOLS), which considers the weighted standard error of the vertical variable (y), and the weighted least-squares (WLS), which incorporates the standard error in both x and y variables. We demonstrate that the calibration method presented in this study significantly increases the correlation coefficient, higher than r2 = 0.94, and reduces both the data deviation and the errors of the linear function for the three regression methods. Nonetheless, the WLS appears as the best regression method to minimize errors in the calibrated element concentrations. Our results open the door to use calibrated XRF-CS data to evaluate marine sediment pollution according to the levels of the strictest sediment quality guidelines (SQG) with errors lower than 0.4%–2% for Fe, 1%–7% for Zn, 3–14% for Pb and 5%–16% for Mn. They highlight the robustness of the calibration procedure here presented for accurate and precise quantification of element concentrations from XRF-CS semi-quantitative data.
Pastoral fire effects on soil function in mountain shrublands. After experimental shrub-to-shrub burnings a transient pulse of available N forms and a decrease of microbial biomass N (MBN) and total ...N were found in burned soils.
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•Soil function effects of burning and extensive grazing were studied in shrublands.•Burning increased soil N availability but the effect disappeared after one year.•Burning decreased total N, MBC and N and P-enzyme activities in the mid-term.•Seasonal variation of microbial parameters was more important than burning effects.•Extensive grazing had no negative effects on burned soils.
In Europe, rural depopulation and the abandonment of pastoral practices in mountain areas trigger deep changes in the landscape, which result in the accumulation of lignified fuels and the increased risk of fires, a sensitive issue in southern areas of the continent. From prehistory, a pyric herbivory has been practiced in European mountain regions. Pastoral fires created open communities, amenable to wild and domestic grazing, and herbivores perpetuated them by controlling fuel accumulation. In the last decades, the declining of extensive herbivory has given a prominent role to prescribed fires in order to preserve open communities. As a consequence, a new scenario of increased burning frequency and reduced herbivory emerges, which may affect the soil function in different ways. Our aim was to evaluate the effects of experimental burnings on soil function and nutrients cycling of a mountain gorse (Ulex gallii Planch.) shrubland, with the absence/presence of extensive grazing. We performed traditional “bush-to-bush” burnings in three experimental mountain plots and analysed seasonally along a 2-year period the soil function in relation to C-cycle (dissolved organic C, microbial biomass C and glucosidase activity), N-cycle (inorganicN forms, dissolved organic N, microbial biomass N and urease activity), P-cycle (phosphatase activity) and overall bacterial catabolic activity. Fire effects were time dependent and extensive grazing had a low influence on them. Fire originated a transient pulse of inorganic-N forms in the short term, which disappeared after 1year, and increased dissolved organic N forms, which attenuated with time. In burned areas, a decrease of total N and microbial biomass N, and a slow-down of N- and P-cycle enzymatic activities were observed in the mid-term coinciding with a decrease of soil moisture. Since a higher burning frequency is a feasible situation that may affect mountain, nutrient-poor soils, the enduring effects of prescribed fires need to be taken into account to establish the optimal date of burning and the adequate recurrence regime that avoid negative impacts on the soil function and minimize the loss of nutrients from the soil reservoir.
Genetically modifying autologous T cells to express an anti-CD19 chimeric antigen receptor (CAR) has shown impressive response rates for the treatment of CD19+ B cell malignancies in several clinical ...trials (CTs). Making this treatment available to our patients prompted us to develop a novel CART19 based on our own anti-CD19 antibody (A3B1), followed by CD8 hinge and transmembrane region, 4-1BB- and CD3z-signaling domains. We show that A3B1 CAR T cells are highly cytotoxic and specific against CD19+ cells
, inducing secretion of pro-inflammatory cytokines and CAR T cell proliferation.
, A3B1 CAR T cells are able to fully control disease progression in an NOD.Cg-
/SzJ (NSG) xenograph B-ALL mouse model. Based on the pre-clinical data, we conclude that our CART19 is clearly functional against CD19+ cells, to a level similar to other CAR19s currently being used in the clinic. Concurrently, we describe the implementation of our CAR T cell production system, using lentiviral vector and CliniMACS Prodigy, within a medium-sized academic institution. The results of the validation phase show our system is robust and reproducible, while maintaining a low cost that is affordable for academic institutions. Our model can serve as a paradigm for similar institutions, and it may help to make CAR T cell treatment available to all patients.
In Huntington's disease (HD), striatal medium spiny neurons (MSNs) are particularly sensitive to the presence of a CAG repeat in the huntingtin (HTT) gene. However, there are many evidences that ...cells from the peripheral immune system and central nervous system (CNS) immune cells, namely microglia, play an important role in the etiology and the progression of HD. However, it remains unclear whether MSNs neurodegeneration is mediated by a non-cell autonomous mechanism. The homeostasis in the healthy CNS is maintained by several mechanisms of interaction between all brain cells. Neurons can control microglia activation through several inhibitory mechanisms, such as the CD200-CD200R1 interaction. Due to the complete lack of knowledge about the CD200-CD200R1 system in HD, we determined the temporal patterns of CD200 and CD200R1 expression in the neocortex, hippocampus and striatum in the HD mouse models R6/1 and HdhQ111/7 from pre-symptomatic to manifest stages. In order to explore any alteration in the peripheral immune system, we also studied the levels of expression of CD200 and CD200R1 in whole blood. Although CD200R1 expression was not altered, we observed and increase in CD200 gene expression and protein levels in the brain parenchyma of all the regions we examined, along with HD pathogenesis in R6/1 mice. Interestingly, the expression of CD200 mRNA was also up-regulated in blood following a similar temporal pattern. These results suggest that canonical neuronal-microglial communication through CD200-CD200R1 interaction is not compromised, and CD200 up-regulation in R6/1 brain parenchyma could represent a neurotrophic signal to sustain or extend neuronal function in the latest stages of HD as pro-survival mechanism.
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