Expression of hnRNPK & Claudin-4 in HCV-Induced Early HCC and Adjacent Liver Tissue

Abstract

Background: HCC in Egypt usually occurs in HCV cirrhotic livers with poor prognosis due to late diagnosis. High hnRNPK & low Claudin-4 profiles indicate Epithelial Mesenchymal Transition (EMT), malignant transformation and high-grade tumours.

Aim: We studied the immunohistochemical expression of hnRNPK and Claudin-4 in HCV induced early HCC (eHCC) and adjacent liver tissue in Egyptian patients to improve eHCC detection in cirrhotic livers with better curative therapy options.

Method: We studied the immunohistochemical expression of hnRNPK and Claudin-4 in 100 Egyptian patients resection specimens of HCV induced early HCC (eHCC) and adjacent liver tissue, in order to improve eHCC detection in cirrhotic livers, thus improving their therapeutic options.

Results: Early HCC grade significantly directly correlated with nuclear hnRNPK/5HPFs count and inversely correlated with Claudin-4 expression %, with a converse correlation between hnRNPK and Claudin-4. Moreover in eHCC, combined hnRNPK ≥ 30/5HPFs & Claudin-4 ≥ 40% significantly distinguished low grade eHCC (G1) from high grade eHCC (G2&G3), with sensitivity 97% & specificity 69.7% for hnRNPK ≥ 30/5HPFs, and with sensitivity 70% & specificity 94.3% for Claudin-4 ≥ 40%. Moreover in the adjacent liver, both markers expressions significantly directly correlated with each other and with METAVIR fibrosis score but not with activity. Furthermore, 58% of eHCCs showed hnRNPK ≥ 30 Claudin-4 < 40% profile, indicating EMT type 3, compared to 26% with hnRNPK ≥ 30 Claudin-4 ≤ 10% profile in adjacent cirrhotic/ precirrhotic liver, with significant use of combined hnRNPK ≥ 30/5HPFs & Claudin 4 ≤ 10% as eHCC prediction cut offs in cirrhosis (p < 0.05).

Conclusion: Combination of hnRNPK and Claudin-4 can indicate early HCC development in HCV cirrhotic livers using hnRNPK ≥ 30/5HPFs & Claudin-4 ≤ 10% cut offs. Also, combination of hnRNPK ≥ 30/5HPFs & Claudin-4 ≥ 40% can distinguish low grade eHCC (G1) from high grade eHCC (G2&G3).

Keywords: Claudin-4; EMT; Metavir; cirrhosis; eHCC; hnRNPK.

Figure 1

H&E Low (G1) & High (G2 & G3) early HCC (eHCC) and the adjacent liver. (A) High (G2-3) early HCC (eHCC) shows moderate nuclear anaplasia and pseudoglandular forms. The adjacent liver shows steatosis (H&E ×100). (B) High grade (G2-3) early HCC (eHCC), shows moderate to high nuclear anaplasia, pseudoglandular forms and bile (H&E × 200). (C) Low grade (G1) early HCC (eHCC) shows low grade nuclei and compact architecture, among a background cirrhotic liver (H&E × 100)

Figure 2

Immunohistochemistry expression hnRNPK & Claudin-4 in eHCC and adjacent liver. (A) Cirrhotic liver nodules are showing nuclear hnRNPK (in focus), (IHC, DAB, ×100). (B) High grade (G2-3) early HCC (eHCC), showing nuclear (in focus) and cytoplasmic hnRNPK expression, (IHC, DAB, ×200).(C) Low grade (G1) early HCC (eHCC) and cirrhotic liver nodules with foci of weak cytoplasmic Claudin-4 (in focus), compared to normal moderate Claudin-4 in proliferated bile ductules (as an internal control) (IHC, DAB, ×200). (D) High grade (G2) early HCC (eHCC) shows weak cytoplasmic Claudin-4 (in focus), (IHC, DAB, ×100)

Figure 3

Statistical analysis charts for evaluation of hnRNPK & Claudin-4 expression in early HCC (eHCC) & adjacent liver. (A) Values of mean nuclear hnRNPK count /5HPFs & mean Claudin-4% of expression in early HCC (eHCC) & adjacent liver. (B) Cut offs of hnRNPK/5HPFs & Claudin-4 expression% among the studied cases regarding early HCC (eHCC) grade. (C) hnRNPK & Claudin-4 expression profiles in early HCC (eHCC) regarding Epithelial Mesenchymal Transition (EMT). (D) hnRNPK & Claudin-4 expression profiles in cirrhotic/precirrhotic adjacent liver regarding Epithelial Mesenchymal Transition (EMT)