Fig. 2

PDAC tumor cells with different organ-specific metastatic potentials showed different capacities in modulating the metabolism gene methylation in CAFs and also the heterogeneity of CAFs in metastatic sites. a The schema shows that wild-type C56Bl/6 mice orthotopically implanted with primary pancreatic KPC tumors spontaneously developed liver or lung metastases, which were dissected for CAF isolation by using magnetic beads coupled with anti-FAP antibodies. Two KPC cell lines from mice that developed both liver and lung metastases were orthotopically implanted in the pancreas of 40 mice (20 mice per cell line). As previously shown [6, 7], the majority of mice died from primary tumor growth without metastases. Of 40 mice, 7 of them developed liver metastasis only and 5 developed lung metastasis only. CAFs were isolated from dissected liver and lung metastases. b Percentage of methylation in the NQO-1 gene in CAFs from multiple liver metastases was measured by MSP. Methylation percentage of NQO-1 in normal liver fibroblasts was used as a baseline methylation level for comparison. A1-7: 7 mice developed liver metastasis spontaneously. Note that methylation levels of the NQO-1 gene were elevated in CAFs isolated from liver metastases compared to normal liver fibroblasts. c Percentage of methylation in the NQO1 gene in CAFs from multiple lung metastases was measured by MSP. Methylation percentage of NQO-1 in normal lung fibroblasts was used as a baseline methylation level for comparison. B1-5: 5 mice developed lung metastasis spontaneously. Note that the gene methylation levels of NQO-1 remained at baseline levels in CAFs isolated from lung metastases compared to normal lung fibroblasts. d Heatmap was generated using TPM scores based on the RNA sequencing analysis of mouse moMSC (mMSC) after co-cultured with Liver Met tumor cells (mMSC + liver) and Lung Met tumor cells (mMSC + lung) to compare the expression of iCAF and myCAF signature genes. Red color indicates upregulation in co-culture compared to mono-culture, blue color indicates downregulation in co-culture compared to mono-culture. e Heatmap was generated using TPM based on the RNA sequencing analysis of mouse CAFs isolated from liver metastasis (4545 liver CAF) and from lung metastasis only (3403 lung CAF) to compare the expression of iCAF and myCAF signature genes. f Diagram illustration shows that CAFs in liver metastasis are reprogrammed by PDAC cells with liver metastasis potential and subsequently lose part of their heterogeneity. Note that CAFs in lung metastasis are not reprogrammed by KPC cells with lung metastasis potential and thus maintain their heterogeneity. Image created using Biorender