Fig. 1
Elevated ac4C modification and increased NAT10 expression are correlated with enhanced PNI and poor prognosis in PDAC. (A-B) Heatmap of UPLC/MS-MS showing upregulation of RNA ac4C levels in PDAC tissues with PNI compared to those without PNI from GDPH cohort (A, p = 0.007) and SYSMH-cohort (B, p = 0.023). (C) Quantification of ac4C level in PDAC tissue with or without PNI from UPLC/MS-MS analysis. (D) RNA dot blot assays indicating a consistent increase in ac4C levels in three paired PDAC tissues (T) when compared to levels in paired normal (N) pancreas (upper panel). The lower panel compares between PDAC tissues with PNI and those without PNI. (E) Kaplan-Meier curves for OS and PFS in patients with PAAD from TCGA database with higher (n = 101) or lower (n = 76) expression of NAT10 mRNA. (F) Kaplan-Meier curves for OS and DFS in patients with pancreatic cancer from the E-MTAB-6134 database with higher (n = 163) or lower (n = 125) expression of NAT10 mRNA. (G) Representative immunofluorescence staining of NAT10 (Green) and CK19 (Red) in PDAC. The nuclei are stained with DAPI (blue). Scale bar, 50 μm. (H) Representative immunohistochemistry images of NAT10 protein in clinical PDAC tissues. Scale bar: 50 μm. (I) Immunoblotting of NAT10 in three paired samples of PDAC and normal tissue (upper), and another six PDAC tissues with (n = 3) or without (n = 3) PNI (lower). (J) Representative immunohistochemistry images of NAT10 in PDAC cases from our cohort with low and high expression. Red circle, perineural nerve in the pancreas. Scale bar: 50 μm. (K) Representative immunohistochemistry images of NAT10 staining in clinical samples with or without PNI. Red circle indicates the nerve. Scale bar: 200 μm (left); 50 μm (right). (L) Percentages of PNI and NAT10 expression in PDAC from different clinical cohorts. n = 62 in the SYSMH cohort; n = 232 in the GDPH cohort. (M-N) Kaplan-Meier analysis for OS in patients with PDAC from the SYSMH (M) and GDPH (N) cohort with higher or lower NAT10 expression. *p < 0.05, **p < 0.01, ***p < 0.001. (C), Student’s t-test. (E-F) and (M-N), log-rank test. (L), χ2 test