Clinicopathological and follow-up data
Forty-seven patients (30 men and 17 women) were eventually included in the present study, with a mean age of 58.4 ± 11.0 years. Nine patients had lymph node invasion, and 5 patients had distant metastasis at the time of diagnosis. After a follow-up period of 25.7 ± 19.1 months, 32 patients (68.1%) developed intrahepatic recurrence after surgery (Table 1).
Table 1
The comparison of clinicopathological parameters between mild and significant peritumoral DR patients
| Mild DR (n = 25) | Significant DR(n = 22) | P value |
Age | 59.3 ± 9.4 | 57.4 ± 12.8 | 0.489 |
Gender | | | 0.560 |
Male | 15(60.0%) | 15(68.1%) | |
Female | 10(40.0%%) | 7(31.9%) | |
TNM stages | | | |
T | | | 0.491 |
1–2 | 10(40.0%) | 11(50.0%) | |
3–4 | 15(60.0%) | 11(50.0%) | |
N | | | 0.449 |
0 | 21(84.0%) | 17(77.2%) | |
1 | 4(16.0%) | 5(22.8%) | |
M | | | 0.532 |
0 | 23(92.0%) | 19(86.4%) | |
1 | 2(8.0%) | 3(15.7%) | |
Recurrence | | | 0.057 |
0 | 11(44.0%) | 4(18.2%) | |
1 | 14(56.0%) | 18(81.8%) | |
Peritumoral DR Was Related To Local Inflammation And Fibrosis
The correlation analysis showed that peritumoral DR was significantly correlated with local inflammation and fibrosis (r = 0.357, P = 0.008 and r = 0.742, P < 0.001, respectively) (Fig. 2a). Due to the different definition of background DR, the DR grade in peritumor (< 2 cm away from the tumor) and nontumor (> 2 cm away from the tumor) areas was also compared in 30 patients whose liver tissues from both sites were available. The results demonstrated a small difference in local inflammation and fibrosis between them, but with no statistical significance. The DR grade in nontumor area paralleled with that in peritumoral area (r = 0.713, P < 0.001) (Fig. 2b). These results indicated a similar extent of DR and local environment between peritumor and nontumor areas.
Peritumoral DR Was Related To The Prognosis Of ICC
According to the grade of peritumoral DR, patients with ICC were divided into two groups: mild peritumoral DR (grades 1 and 2) (n = 25) and significant DR (grades 3 and 4) (n = 22). Age, gender composition, TNM stages, and tumor differentiation were not significantly different between these two groups. However, the recurrence in the significant DR group was much higher than that in the mild DR group (81.8% vs 56.0%, P = 0.058) (Table 1). The survival analysis showed that patients with significant peritumoral DR had poor overall and disease-free survival (P = 0.01and P = 0.03, respectively) (Fig. 3).
Different Proliferation Of Peritumoral Ductular Cells
According to the previous description, PI was used to mark the extent of proliferation (Fig. 4a). The significant peritumoral DR group showed a higher PI trend of ductular cells compared with the mild peritumoral DR group but failed to achieve statistical significance due to high variation (0.43 ± 0.29 vs 0.28 ± 0.31, P = 0.172). The percentage of high PI (> 50%) ductular cells was also higher in the significant peritumoral DR group (44.44% vs 30.77%, P < 0.01). Undoubtedly, the tumor cells showed much higher PI compared with the other two groups (Fig. 4b).
Different grade of peritumor DR was related to different microenvironments
ICC is a kind of tumor with abundant extracellular matrix (ECM), which plays an indispensable role in tumor progression. Double-fluorescence immunostaining showed the α-smooth muscle actin (α-SMA)-positive fibrosis background and CK19-positive ductular and tumor cells. The results demonstrated more abundant ECM andα-SMA-positive vessels in peritumoral areas in the significant DR group than in the mild DR group, which was similar to that in the tumor (Fig. 5a). The correlation analysis showed that the DR grade was positively related to the portal/septalα-SMA level (Fig. 5b).