Interleukin‐4‐Clicked Surfaces Drive M2 Macrophage Polarization

Abstract

Driving macrophage (Mϕ) polarization into the M2 phenotype provides potential against inflammatory diseases. Interleukin-4 (IL-4) promotes polarization into the M2-Mϕ phenotype, but its systemic use is constrained by dose-limiting toxicity. Consequently, we developed IL-4-decorated surfaces aiming at sustained and localized activity. IL-4 muteins were generated by genetic code expansion; Lys42 was replaced by unnatural amino acids (uAAs). Both muteins showed cell-stimulation ability and binding affinity to IL4Rα similar to those of wt-IL-4. Copper-catalyzed (CuAAC) and copper-free strain-promoted (SPAAC) 1,3-dipolar azide-alkyne cycloadditions were used to site-selectively anchor IL-4 to agarose surfaces. These surfaces had sustained IL-4 activity, as demonstrated by TF-1 cell proliferation and M2, but not M1, polarization of M-CSF-generated human Mϕ. The approach provides a blueprint for the engineering of cytokine-activated surfaces profiled for sustained and spatially controlled activity.

Site-Specific POxylation of Interleukin-4

“…Plk-IL-4 and wt-IL-4 were expressed as described previously. 33 Briefly, E.coli BL21 DE3, encoding for TAG(42)-IL-4 and for the PylRS/tRNA CUA pair, were cultured at 37 °C and the Plk substrate was added at a final concentration of 2 mM at OD 600 = 0.3 in standard TB (Terrific Broth) medium. For wt-IL-4 expression, BL21 (DE3) cells, encoding for wt-IL-4, were used.…”

“…For the enhanced yellow fluorescent protein (eYFP) reporter gene assay, 15.000 HEK 293T cells were cotransfected in 24 well plates with 1 μg of the plasmids pHW0040 (P STAT6 −eYFP) and 1 μg of the constitutive STAT6 expression vector pSTAT6 (Genebank accession N-BC075852.1) as described before. 33 After exchange of the transfection medium against growth medium, supplemented with 1.3 nM of wt-IL-4, Plk-IL-4, and polymer-conjugated IL-4 variants, respectively, HEK 293T cells were stimulated for 48 h at 37 °C and 5% CO 2 . eYFP expression was detected with an Axiovert 200 M inverted microscope (Zeiss, Oberkochen, Germany).…”

TLR7/8 Agonist and SHP2 Inhibitor Loaded Nanoparticle Enhances Macrophage Immunotherapy Efficacy

“…To accomplish that, we performed a macrophage repolarization assay as shown in the schematics in Figure a. Briefly, iBMDMs or RAW264.7 cells were polarized to an M2 phenotype by adding IL‐4 at a concentration of 20 ng mL −1 for 24 h. [ 45 ] The macrophages were then subjected to different treatments (Control, SHP099 Free Drug (SHP FD), LNP, SHP099 Nanoparticle (SHP NP), R848 Nanoparticle (R848 NP), R848 Free Drug (R848 FD)) with an equivalent drug concentration of 10 µM of each drug. After an incubation time of 4 h, the LNPs are then washed away with cold PBS, and fresh media was replaced.…”

Nicotine causes alternative polarization of macrophages via Src‐mediated STAT3 activation: Potential pathobiological implications

“…Wang et al, 2016;Yamaguchi et al, 2017). The M2 or alternative polarization of macrophages is induced by multiple stimuli such as IL-4, IL-10, and IL-13 and exerts anti-inflammatory responses (da Silva et al, 2015;Luhmann et al, 2016;M. Z. Zhang et al, 2017).…”

“…The M1 polarization is stimulated by a bacterial infection, and their cell wall components such as LPS primarily and induces a pro‐inflammatory immune response by releasing the cytokines such as IFN‐γ and TNF‐α, IL‐6, IL‐12, IL‐23, and nitric oxide (NO) production (X. Chen et al, 2019; Sica et al, 2015; Q. Wang et al, 2016; Yamaguchi et al, 2017). The M2 or alternative polarization of macrophages is induced by multiple stimuli such as IL‐4, IL‐10, and IL‐13 and exerts anti‐inflammatory responses (da Silva et al, 2015; Luhmann et al, 2016; M. Z. Zhang et al, 2017). M2 macrophages release a variety of chemokines and cytokines that contribute to tissue repair and remodeling, immunoregulation, and tumor‐promoting processes (Deshmukh et al, 2018; Magatti et al, 2017; Sumitomo et al, 2019).…”