Introduction
Liver fibrosis, characterized by diffuse fibrous deposition and distortion of hepatic vascular architecture in histology, is an intermediate step in the development of cirrhosis resulting from chronic liver disease. Liver sinosoidal endothelial cells (LSECs) are critical gatekeepers of sinusoidal homeostasis. Following injury, LSECs are the first to be affected and initiate a cascade of reactions within the sinusoidal, which contribute to the development and progression of liver fibrosis and portal hypertension. Our preliminary studies have shown that hepatic cyclooxygenase-2 (COX-2) expression was correlated with the severity of inflammation and fibrosis, and its inhibitor could ameliorate intrahepatic fibrosis.
Aims & Methods
The present study aimed to investigate the role of COX-2 on LSECs function and its underlying molecular mechanism.
Results
Ptgs2 deficiency attenuates liver fibrosis
In both inducible and non-inducible mouse models, Ptgs2 knockdown resulted in a reduction in collagen deposition. TAA- or DDC-induced fibrosis-related protein expression (collagen I and α-SMA) was significantly reduced by Ptgs2 deletion.
Ptgs2 deficiency alleviates hepatic inflammatory injury
RNA-seq analysis indicated that down-regulated DEGs by Ptgs2 knockdown were enriched in leukocyte chemotaxis, leukocyte migration, and leukocyte cell-cell adhesion. Consistently, significantly decreased numbers of infiltrating neutrophils and macrophages were observed in Ptgs2-deficient mice. In addition to inflammatory cell infiltration, liver function was also improved suggested by significant decreases in serum levels of ALT and AST.
COX-2 expression is elevated in LSECs in reponse to liver injury
D-IF showed that COX-2 expression was elevated in injury liver tissues. The immunofluorescence for COX-2 in TAA-A and TAA-C groups was also co-stained with LYVE1, a LSEC-specific maker. RNA-seq data from the GEO database further confimed the COX-2 overexpression in murine LSECs isolated from various fibrotic models. In vivo study, COX-2-mPGES-1-EP2 pathway could be up-regulated by pro-inflammatory factor TNFα.
COX-2-PGE2-EP2 promotes leukocyte recruitment through NF-κB pathway
RNA-seq analysis found that inhibition of COX-2 supressed TNF-induced gene transcription which were enriched in leukocyte recruitment, TNFα pathway, and NF-κB pathway. In HHSEC, TNFα activated NF-κB signaling pathway and increased expression levels of chemokines (CXCL1 and CCL2) and adhesion molecules (ICAM1 and VCAM1). This TNFα-induced effect could be potently suppressed by COX-2 inhibitor (Celecoxib), mPGES-1 inhibitor (MF63) or EP2 receptor antagonist (TG4-155). PGE2 also activated NF-κB pathway, which was abolished after EP2 blockage.
COX-2 inhibitors attenuate fibrosis and liver injury
Both Celecoxib and Etoricoxib (both were selective COX-2 inhibitors) were able to decrease collagen deposition, fibrosis-related protein expression, inflammatory cell infiltration and serum transaminase levels in TAA or DDC models.
Conclusion
COX-2 expression in LSECs is up-regulated in liver fibrosis. Upon inflammatory stimulation, COX-2-PGE2-EP2 axis increases chemokines and adhesion molecules through the synergistic activation of NF-κB signaling pathway, leading to leukocyte recruitment and hepatic inflammation. Inhibition of COX-2 by Celecoxib or Etoricoxib attenuate fibrosis and liver injury. The present study sheds light on the regulation of LSECs on leukocyte recruitment through a cyclooxygenase-2-dependent mechanism. Taken together, our results suggest COX-2 as a potential therapeutic target in the treatment of liver fibrosis.
Disclosure
The authors disclose no conflicts.
