PMC

a Experimental sample for measuring the hinge states (HSs). The numbers—label the four y-directional hinges. b Eigen frequencies for the sample shown in a at k_y = π/a, obtained from full-wave simulation. The blue dots represent the bulk and surface states, and the red dots indicate the HSs. c, Eigen profiles for the two states highlighted in red in b (∣p∣ denotes the absolute value of acoustic pressure). The color indicates the amplitude of the acoustic pressure. d–g Experimentally measured dispersions for hinge 1 d, hinge 2 e, hinge 3 f, and hinge 4 g. The gray dots represent simulated eigenfrequencies of the bulk and SSs, and the red dots indicate simulated hinge bands. The solid blue lines denote the projections of the nodal rings. h–k Experimentally measured acoustic intensity distributions on two surfaces adjacent to hinge 1 h, hinge 2 i, hinge 3 j, and hinge 4 k. The red star indicates the position of the speaker and the red dashed line highlights the position of the hinge. The operating frequencies of the speaker are chosen as: 3078 Hz (hinge 1 and hinge 3), 3076 Hz (hinge 2) and 3080 Hz (hinge 4), which are around the eigenfrequencies of the HSs.

Top and side views of the investigated hinge designs. The hinges of the two most commonly implanted bileaflet mechanical heart valves are studied: the St Jude Medical (SJM) valve and the CarboMedics (CM) valve. In order to assess the effect of the hinge gap width on the flow, two configurations of the SJM hinge are investigated: a SJM hinge with a regular hinge gap width (SJM regular hinge) and a SJM hinge with a larger than regular hinge gap width (SJM large hinge, not shown). The terminology used to describe the hinge recess is included. Note that the images are to scale and that the leaflet is in its fully closed position

(a–c) The hinge analog dose required for optimal p-Met inhibition was determined by immunoblotting for p-Met following treatment with HGF (10 ng/ml) and each hinge analog at several concentrations (100-1 nM). β-actin served as the loading control. (a) Representative p-Met blots demonstrating inhibition of HGF activity by HGF Hinge at each concentration and optimal inhibition observed at 1 nM. (b) MSP Hinge inhibited Met activation at 10 and 1 nM, but no inhibition was observed at 100 nM. (c) Norleual suppressed HGF-dependent Met activation at each concentration with maximal inhibition at 1 nM. (d–f) To quantitatively assess hinge analog inhibition of Met activation, LM-P cells were treated with PBS (control), HGF (20 ng/ml), or HGF + HGF Hinge, HGF + MSP Hinge, or HGF + Norleual (all at 1 nM) for 10 min prior to lysate collection. (d) Images of representative immunoblots demonstrating the increase in p-Met and p-Erk expression by HGF and the inhibition of the HGF-dependent phosphorylation of Met and Erk by the hinge analogs. Total Met and Erk levels remained relatively unchanged regardless of treatment. β-actin was included as a loading control. (e) Quantitative analysis of p-Met bands by densitometry. p-Met bands were normalized to total Met and expressed as p-Met/Met. HGF alone group was significantly different from all other groups, indicating inhibition of HGF activity by the hinge analogs. (*** P < 0.001, n=3). (f) Densitometric analysis of p-Erk bands demonstrates inhibition of p-Erk expression by MSP Hinge and Norleual. (** P < 0.01, n=3). The difference between HGF and the HGF Hinge group did not reach significance. Data is expressed as p-Erk/β-actin. (g) LM-P cells were treated with HGF Hinge, MSP Hinge, Norleual (all at 1 nM), or PBS (control) without addition of HGF and immunoblotted for p-Met. No significant change in the basal level of Met phosphorylation was observed.

Schematic representation of Ab configurations. Schematic representation of different Abs’ configurations. (A) HcAb has its llama hinge linker: bivalent; (B) Camelid VHH: monovalent; (C) VHH-(G4S)3-VHH: bivalent; (D) VHH-(G4S)3-VHH-(G4S)3-VHH: trivalent; (E) Human classic Ab with its hinge linker: bivalent; (F) Llama VHH-human IgG4 hinge linker-IgG4 Fc chimeric HcAb: bivalent; (G) Llama VHH-human CH1-IgG4 hinge linker-IgG4 Fc chimeric heavy chain: bivalent; (H) VHH-llama long linker-VHH-human IgG4 hinge linker-IgG4 Fc: tetravalent; (I) VHH-(G4S)3 linker-VHH-human CH1-IgG4 hinge linker-IgG4 Fc chimeric Ab: tetravalent; (J) VHH-human IgG4 hinge linker-IgG4 Fc-llama long linker-VHH chimeric Ab: tetravalent. Thus, “(G4S)3” is a 15-peptide linker (amino acid sequence: GGGGSGGGGSGGGGS). “IgG4-hinge linker” means the hinge region between the human Fab and human IgG4 Fc (amino acid sequence: PPCPSCPAPEFLGGPS). “Llama long linker” means the hinge region between the llama Fab and llama IgG2 Fc (amino acid sequence: EPKIPQPQPKPQPQPQPQPKPQPKPEPECTCPKCP).