Fig 2: iPSC directed intestinal differentiation in transwell culture results in cells with expected morphology and marker expression through Stage 2. (A) Directed intestinal differentiation occurs through several distinct stages. (B) Transwell cells derived from a panel of iPSC lines express Definitive Endoderm markers SOX17 and CXCR4 at Stage 1. (C) Top row: transwell iPSC panel morphology at Stage 2 differs in amount and size of budding spheroids (arrow heads). Scale bar, 250 µm. Bottom row: CDX2 expression of Stage 2 transwell iPSC-derived cells differs among cell lines in intensity and uniformity. Scale bar, 250 µm.

Fig 3: GCs downregulated CXCR4 via the impaired PGE2/EP4/AKT signaling in EPCs. (A) The relative mRNA expression of EP4 in untreated, GC-treated EPCs, and GC-treated EPCs in the presence of PGE2. (B) The relative protein expression of pAKT/AKT in untreated, GC-treated EPCs, and GC-treated EPCs in the presence of PGE2. (C) The inhibition of PI3K/AKT pathway in GC-treated EPCs in the presence of PGE2. LY294002 was used as a PI3K inhibitor. (D) The relative mRNA expression of CXCR4 in GC-treated EPCs in the presence of PGE2 and PI3K inhibitor. Dexamethasone (100 nM, 24-h treatment) was used as a GC. Untreated: untreated EPCs, 100 nM: GC-treated EPCs, 100 nM + PGE2: GC-treated EPCs in the presence of PGE2, 100 nM + PGE2 + LY: GC-treated EPCs in the presence of PGE2 and PI3K inhibitor. LY: LY294002. The data represent the mean ± SD. n = 3, **P < 0.01, *P < 0.05. The experiment was repeated triplicate. The charts showed the overall mean data from all experiments.

Fig 4: Analyses of CXCR4 expression and migration potential in fresh or cryopreserved MSC and MSCTRAIL cells (passage 5). (a) CXCR4 expression was measured by FACS. Cells were stained by an isotype immunoglobulin G-PE or by an anti-human-CXCR4-PE antibodies, respectively. (b) Three-dimensional collagen gel migration assay of fresh or cryopreserved MSCs and MSCTRAILs compared with fresh MSCTRAILs, respectively, analyzed by Bonferroni multiple comparison statistic test (***P < 0.001).

Fig 5: Acetylated KLF5 is upregulated and positively correlated with CXCR4 in bone metastases of PCa.a Representative images of IHC staining of Ac-KLF5. Red arrows indicate positive Ac-KLF5 staining. Scale bar, 50 µm. b Quantitative analyses of Ac-KLF5 expression in benign tissues, hyperplasia, and localized tumors of the prostate and metastases of PCa from both the visceral and bone tissues. Numbers of samples (n) are indicated in the figure. Data are shown in mean ± S.E.M. NS not significant; *p < 0.05; ***p < 0.001 (two-tailed Student’s t test). c Ac-KLF5 expression is positively correlated with CXCR4 expression in bone metastases of PCa patients, as indicated by representative images of IHC staining in two bone metastasis samples. Black arrow heads indicate stronger Ac-KLF5 and CXCR4 staining in the same cells from consecutive sections. Scale bar, 50 µm. d Pearson analyses of the expression of Ac-KLF5 and CXCR4 in 51 bone metastasis samples of PCa patients. ***p < 0.0001. e A schematic depicting how acetylation of KLF5 leads to bone metastasis by promoting osteoclast differentiation via transcriptionally activating CXCR4, which in turn increases IL-11 secretion. Representative images shown in a and c are statistically analyzed in b and d, respectively. Source data are provided as a Source Data file.