TY - JOUR

T1 - Differential Internalization of Thrombin-Derived Host Defense Peptides into Monocytes and Macrophages

AU - Hansen, Finja C.

AU - Nadeem, Aftab

AU - Browning, Kathryn L.

AU - Campana, Mario

AU - Schmidtchen, Artur

AU - Van Der Plas, Mariena J.A.

N1 - Funding Information: This work was supported by grants from Alfred Österlunds Foundation, Hudfonden (Edvard Welanders Stiftelse and Finsenstiftelsen), O.E. och Edla Johanssons Foundation, Thelma Zoégas Foundation, the Royal Physiographic Society, and Åke Wibergs foundation awarded to M.J.A.P., from Knut and Alice Wallenberg Foundation, the Swedish Strategic Research Foundation, the Swedish Research Council (2017-02341, 2020-02016) and the Swedish Government Funds for Clinical Research (ALF) to A.S., and from LEO Foundation to K.L.B. Publisher Copyright: © 2021 The Author(s). Published by S. Karger AG, Basel.

PY - 2022

Y1 - 2022

N2 - Proteolytic cleavage of thrombin generates C-terminal host defense peptides exerting multiple immunomodulatory effects in response to bacterial stimuli. Previously, we reported that thrombin-derived C-terminal peptides (TCPs) are internalized in monocytes and macrophages in a time- and temperature-dependent manner. In this study, we investigated which endocytosis pathways are responsible for the internalization of TCPs. Using confocal microscopy and flow cytometry, we show that both clathrin-dependent and clathrin-independent pathways are involved in the internalization of the prototypic TCP GKY25 in RAW264.7 and human monocyte-derived M1 macrophages, whereas the uptake of GKY25 in monocytic THP-1 cells is mainly dynamin-dependent. Internalized GKY25 was transported to endosomes and finally lysosomes, where it remained detectable for up to 10 h. Comparison of GKY25 uptake with that of the natural occurring TCPs HVF18 and FYT21 indicates that the pathway of TCP endocytosis is not only cell type-dependent but also depends on the length and composition of the peptide as well as the presence of LPS and bacteria. Finally, using neutron reflectometry, we show that the observed differences between HVF18 and the other 2 TCPs may be explained partially by differences in membrane insertion. Taken together, we show that TCPs are differentially internalized into monocytes and macrophages.

AB - Proteolytic cleavage of thrombin generates C-terminal host defense peptides exerting multiple immunomodulatory effects in response to bacterial stimuli. Previously, we reported that thrombin-derived C-terminal peptides (TCPs) are internalized in monocytes and macrophages in a time- and temperature-dependent manner. In this study, we investigated which endocytosis pathways are responsible for the internalization of TCPs. Using confocal microscopy and flow cytometry, we show that both clathrin-dependent and clathrin-independent pathways are involved in the internalization of the prototypic TCP GKY25 in RAW264.7 and human monocyte-derived M1 macrophages, whereas the uptake of GKY25 in monocytic THP-1 cells is mainly dynamin-dependent. Internalized GKY25 was transported to endosomes and finally lysosomes, where it remained detectable for up to 10 h. Comparison of GKY25 uptake with that of the natural occurring TCPs HVF18 and FYT21 indicates that the pathway of TCP endocytosis is not only cell type-dependent but also depends on the length and composition of the peptide as well as the presence of LPS and bacteria. Finally, using neutron reflectometry, we show that the observed differences between HVF18 and the other 2 TCPs may be explained partially by differences in membrane insertion. Taken together, we show that TCPs are differentially internalized into monocytes and macrophages.

KW - Bacterial infection

KW - Caveolin-dependent endocytosis

KW - Clathrin-dependent endocytosis

KW - Host defense peptides

KW - Lysosomes

KW - Thrombin

U2 - 10.1159/000520831

DO - 10.1159/000520831

M3 - Journal article

C2 - 34937021

AN - SCOPUS:85121767245

VL - 14

SP - 418

EP - 432

JO - Journal of Innate Immunity

JF - Journal of Innate Immunity

SN - 1662-811X

IS - 5

ER -