Fig 2: GDF15 is a bona fide heart-derived hormone that regulates liver GH signaling ADesign of AAV9-mGDF15 shRNA construct to specifically knockdown GDF15 in Cre+ cardiomyocytes.B–GCardiac Gdf15 expression quantified by qPCR (B), plasma GDF15 concentrations measured by ELISA (C), cardiac Bnp expression quantified by qPCR (D), liver phosphorylated STAT5 level measured by ELISA (E), and plasma IGF1 (F) and GH concentrations (G) measured by ELISA in 9- to 10-day-old littermate control and aKO?KO mice (n = 8–12 mice per group) that received pericardial injection of AAV9-control or Gdf15 shRNA at 2 days of age. *P < 0.05, **P < 0.01, and ***P < 0.001 by t-test. Values are mean + s.e.m.

Fig 3: GDF15 is a candidate heart-derived hormone that inhibits liver GH signaling and body growth APlasma IGF1 concentrations (ng/ml) in 7-day-old weight- and gender-matched littermate WT mice injected with control or different proteins were measured by ELISA (n = 3–5 mice per group, daily i.p. injection from 5 days of age).B–GLiver phosphorylated and total STAT5 and JAK2 as well as ß-actin (loading control) determined by Western blot (B); liver expression of Igf1, Igfbp3, and Igfals quantified by qPCR (C); plasma IGF1 (D), IGFBP3 (E), and GH concentrations (F) measured by ELISA; and daily body weight (G) in weight- and gender-matched littermate WT mice injected with control or GDF15 (n = 5 per group, daily i.p. injection from 3 days of age).HOvernight-fasted (in DMEM) WT mouse primary hepatocytes were first treated with different concentrations of GDF15 for 30 min and then with 20 ng/ml GH for 15 min. Cellular levels of phosphorylated STAT5, total STAT5, and ß-actin (loading control) were determined by Western blot.Data information: *P < 0.05, **P < 0.01, and ***P < 0.001 between control and GDF15 by t-test. All values are mean + s.e.m.Source data are available online for this figure.

Fig 4: Gdf15 expression in control and knockout hearts. (A) Representative pictures of GDF15 immunostaining in P10 hearts. (B) Quantification of Gdf15+ cells based on immunostaining in A. n = 2 mice per genotype. (C) Feature plot showing Gdf15+ nucleus distribution in P10 hearts. (D) Quantification of Gdf15+ nuclei in control and knockout hearts based on sNucDrop-seq. (E) Distribution of Gdf15+ nuclei in each cell type. (**) P < 2.2 × 10-16 by ?2 test in B and D.

Fig 5: GDF-15 expression in the lung of IPF patients. (A) GDF-15 protein expression in the lung tissues of subjects with HC, S-IPF, and AE-IPF by WB. (B) The quantification of A showed that the protein expression of GDF-15 was significantly upregulated in AE-IPF patients compared with S-IPF subjects and HCs (p = 0.007 and p < 0.001, respectively). (C) GDF-15 mRNA expression in the lung of AE-IPF patients was significantly increased compared with HC, S-IPF cases by RT-PCR (p < 0.001 and p < 0.026, respectively) (n = 4, each group). (D–G) GDF-15 expression in the lung measured by IHC. (D) The moderately positive expressions of GDF-15 in the cytoplasm of type II AECs and macrophages in HC (IHC, ×200). (E) The moderately positive expressions of GDF-15 in inflammatory cells and macrophages in the alveolar septum and mild positive expressions in the bronchial epithelial cells and fibroblasts in S-IPF patients (IHC, ×200). (F) GDF-15 moderately positive expressions in inflammatory cells, macrophages, and fibroblasts in the interstitial spaces in AE-IPF case (IHC, ×200). (G) The moderately positive expressions of GDF-15 in the bronchial epithelial cells and fibroblasts in the same AE-IPF patient as Figure 2F (IHC, ×400). (Notes: macrophages ; type II AEC ; inflammatory cell ; bronchial mucosa epithelial cells ; fibroblast ).