Fig 2: Pathological and immunohistochemical findings in primary tumors (P), patient-derived xenografts (PDXs, F0), and PDX from pleural effusion (PE0) and peritoneal effusion (A0) samples. (a,b) Similar pathological findings among P, F0, and A0 for cases 1 and 3 (i.e., serous carcinoma). The tumor cells mainly had a solid architecture. Papillary growths and a fibrous branch were often observed. The nuclei had strong atypia. The tumor cells were positive for AE1/AE3 and negative for p53. The percent Ki-67 labeling of each tumor ranged from 30% to 33%. (c) In case 5 (i.e., carcinosarcoma), the primary tumor had two malignant components: epithelial and sarcomatous components. The high-grade epithelial carcinoma component was positive for AE1/AE3, while the sarcomatous component was positive for CD10. Interestingly, similar pathological and immunohistochemical features were found between P and A0. The percent Ki-67 labeling was 26% in P and 27% in A0. (d) In case 6 (i.e., low-grade serous carcinoma), similar pathological and immunohistochemical features were found between P and A0; the tumor cells were small papillae-containing cells with uniform nuclei and inconspicuous mitotic activity. The tumor cells were positive for AE1/AE3 and negative for p53 and had 6% Ki labeling. (e) In case 7 (i.e., clear cell carcinoma), similar pathological and immunohistochemical features were found among P, F0, and PE0. The tumors exhibited a solid pattern. The solid architecture consisted of sheets of clear cells separated by delicate septa. The tumor cells were positive for AE1/AE3 and negative for p53. Ki-67 labeling was 7% in P, 7% in F0, and 6% in PE0. The scale bar indicates 100 µm.

Fig 3: Relationships between serum NEP levels and tissue NEP expression levels (adipose and LA tissues). A: Relationship between serum NEP levels and NEP protein (a) and mRNA expression (b) in the SAT (n = 44). B: Relationship between serum NEP levels and NEP protein (a) and mRNA expression (b) in EAT (n = 58). C: Relationship between serum NEP levels and NEP protein (a) and mRNA expression (b) in the LA (n = 61). The r- and p-values are shown.

Fig 4: Comparison of echocardiographic findings and gene expression levels between PAF patients with atrial low NEP and high NEP expression. A: Age, BMI, and echocardiographic findings in PAF patients (n = 21) with low or high atrial NEP mRNA expression. B-F: Comparison of NEP mRNA (B), NEP protein level (C), serum concentration of NEP (D), ANP mRNA (E), and fibrosis-related gene expression (F) between PAF patients (n = 25) with low and high NEP mRNA expression. For some genes, actual data (mean ± standard deviation) are shown in the figure. Red bar, high NEP (n = 12–13); blue bar, low NEP (n = 11–12). *p < 0.05, **p < 0.01.

Fig 5: Multiple linear regression analysis and partial correlation analysis between atrial NEP mRNA expression level and fibrosis-related gene expression level. A: Multiple regression analysis. Multivariate regression analysis showed that 1) log (TGFB1) and log (NOX4) were independent variables for predicting log (COL1A1) expression (brown arrow). 2) Log (NOX4) and log (NLRP3) were independent variables for predicting log (NEP) expression levels (green arrow). 3) Log (NEP) was an independent variable for predicting log (NLRP3), log (TGFB1), and log (NOX4) (red arrow). β-values are shown. B: Partial correlation analysis of NOX4, TGFB1, NLRP3, and NEP. Log (NEP) expression level was significantly positively correlated with log (NOX4) (r = 0.41) and log (NLRP4) (r = 0.39) expression level, but not with log (TGFB1). The partial correlation coefficients (r) are also shown. The lines connecting each index have a partial correlation coefficient of 0.25 or more.