Id |
Subject |
Object |
Predicate |
Lexical cue |
T83 |
0-2 |
Sentence |
denotes |
3. |
T84 |
3-24 |
Sentence |
denotes |
Materials and Methods |
T85 |
26-30 |
Sentence |
denotes |
3.1. |
T86 |
31-52 |
Sentence |
denotes |
Computational Methods |
T87 |
53-284 |
Sentence |
denotes |
Vitual screening on the DrugBank [23] was carried out using the Molecular Operating Environment (MOE) program [28], using four common points of the BK B1 and B2 pharmacophores described previously [25,26] that were used as a query. |
T88 |
285-448 |
Sentence |
denotes |
Previously, a 3D DrugBank database containing the 3D structure, together with a set of conformations of each molecule, was constructed as explained elsewhere [24]. |
T89 |
449-580 |
Sentence |
denotes |
Virtual screening was carried out on the subset of molecules with molecular weight between 200 and 600 (a total of 1703 molecules). |
T90 |
581-856 |
Sentence |
denotes |
Hits identified in the screening process were docked in the 3D models of the B1 and B2 bradykinin receptors, using a set of unique conformations resulting from thorough conformational searches for the diverse ligands studied and rank ordered by means of the MOE program [28]. |
T91 |
858-862 |
Sentence |
denotes |
3.2. |
T92 |
863-878 |
Sentence |
denotes |
In Vitro Assays |
T93 |
880-886 |
Sentence |
denotes |
3.2.1. |
T94 |
887-913 |
Sentence |
denotes |
Ligand Displacement Assays |
T95 |
914-1080 |
Sentence |
denotes |
Raloxifene was tested for its ability to displace reference ligands on human recombinant bradykinin B1 or B2 receptors, respectively, expressed in CHO cells at 20 µM. |
T96 |
1081-1289 |
Sentence |
denotes |
For this purpose, first saturation isotherms were obtained with reference ligands ([3H] des-Arg10-BK (0.35 nM) in the case of B1, and [3H]-bradykinin (0.2 nM) for B2) incubated for 60 min at room temperature. |
T97 |
1290-1423 |
Sentence |
denotes |
Nonspecific binding was evaluated by adding a reference compound (desArg9[Leu8]-BK at 10 µM in the case of B1 and BK at 1 µM for B2). |
T98 |
1424-1656 |
Sentence |
denotes |
Antagonism of raloxifene was measured as a percent inhibition of specific binding of [3H]-bradykinin as control, obtained in the presence of raloxifene at 20 µM and using as reference compounds desArg10-KD for B1 and NPC-567 for B2. |
T99 |
1658-1664 |
Sentence |
denotes |
3.2.2. |
T100 |
1665-1691 |
Sentence |
denotes |
Functional Efficacy Assays |
T101 |
1692-1822 |
Sentence |
denotes |
Measurementent of the efficacy of the compound on the B2 receptor was carried out following the protocol described elsewhere [31]. |
T102 |
1823-2005 |
Sentence |
denotes |
The method is based on measuring differences in intracellular Ca2+ concentrations produced in diverse conditions on CHO cells expressing recombinant human B2 receptor by fluorometry. |
T103 |
2006-2126 |
Sentence |
denotes |
Agonism was measured through the capacity of the compound to increase Ca2+ concentration compared to BK (EC50 ~ 2.4 pM). |
T104 |
2127-2234 |
Sentence |
denotes |
Antagonism was measured through the capacity to antagonize BK based on the reduction of Ca2+ concentration. |