Transiently overexpressed IL-34 in epidermis led to macrophage enrichment in epidermis. A total of 1.8 nL (30 ng/μL) of the pBLK-krt19-il34-sv40 construct was microinjected into one-cell stage Tg (mpeg1: GFP) (A) and Tg (lyz: EGFP) (B) zebrafish embryos. (A, B) WISH of il34 expression and whole-mount antibody staining of GFP expression in 4 dpf embryo (6 × ). The whole body picture of the fish is made up of two separate images taken by confocal and stitched together in the Photoshop. Insets are high magnification (20 × ) of the corresponding boxed regions (gray dotted rectangles). (C, D) Quantitative analysis of macrophage or neutrophil cell numbers in uninjected and construct-injected embryos' trunk (approximately between the 3rd and 6th somite, shown in gray dotted rectangle), skin, and CHT tail region (approximately between the 13th and 17th somite, shown between two white dotted lines). Data were analyzed by Mann–Whitney U test, ***p < 0.001 compared to control. n = 5, 5 for the 4 dpf injected and control fish. Bars: 200 μm (the line on 6 times objective pictures); 50 μm (the line on 20 times objective pictures). CHT, caudal hematopoietic tissue; dpf, days postfertilization; IL-34, interleukin-34; WISH, whole mount in situhybridization.

 Transiently overexpressed IL-34 in liver led to macrophage enrichment in liver. A total of 1.8 nL (30 ng/μL) of the pBLK-fabp10a-il34-sv40 construct was microinjected into one-cell stage Tg (mpeg1: GFP) (A)and Tg (lyz: EGFP) (B) zebrafish embryos. (A, B) WISH of il34 expression and whole-mount antibody staining of GFP expression in 4 dpf embryo (6 × ). The whole body picture of the fish is made up of two separate images taken by confocal and stitched together in the Photoshop. Insets are high magnification (20 × ) of the corresponding boxed regions (gray dotted areas). (C, D) Quantitative analysis of macrophage or neutrophil cell numbers in uninjected and construct-injected embryos' liver (shown in white dotted area) and CHT tail region (approximately between the 13th and 17th somite, shown between two white dotted lines). Data were analyzed by Mann–Whitney U test, **p < 0.01 compared to control. n = 5, 5 for the 4 dpf injected and control fish. Bars: 200 μm (the line on 6 times objective pictures); 50 μm (the line on 20 times objective pictures). fabp10a, fatty acid binding protein 10a.

Stably overexpressed IL-34 in liver led to macrophage enrichment in liver. (A) WISH of il34 expression and whole-mount antibody staining of GFP expression in 4 dpf embryo (6 × ). The whole body picture of the fish is made up of two separate images taken by confocal and stitched together in the Photoshop. Insets are high magnification (20 × ) of the corresponding boxed regions (gray dotted areas). (B) Quantitative analysis of macrophage cell numbers in Tg (mpeg1: GFP) and Tg (fabp10a: il34; mpeg1: GFP) embryos' liver (shown in white dotted area) and CHT tail region (approximately between the 13th and 17th somite, shown between two white dotted lines). (C) Liver confocal imaging of Tg (mpeg1: GFP) and Tg (fabp10a: il34; mpeg1: GFP) in 8-month-old fish (20 × ). (D) Quantitative analysis of macrophage cell numbers in the livers of Tg (mpeg1: GFP) and Tg (fabp10a: il34; mpeg1: GFP) adult fish. Data were analyzed by Mann–Whitney U test, **p < 0.01, and ***p < 0.001 compared to control. n = 5, 5 for the 4 dpf or 8 mpf IL-34 overexpression and control fish. Bars: 200 μm (the line on 6 times objective pictures); 50 μm (the line on 20 times objective pictures).

 Overexpression of IL-34 led to macrophage migration to the liver. Micrographs of live imaging show the process of a macrophage (labelled by white arrows) passing by the liver (white dotted area) within 28 minutes in control fish (A) and the process of a macrophage (labelled by white arrows) migrating into the liver (white dotted area) within 28 minutes in IL-34 overexpressing fish (B). Bars: 40 μm (white line).