Brahma-Related Gene 1 Inhibition Prevents Liver Fibrosis and Cholangiocarcinoma by Attenuating Progenitor Expansion.

Intrahepatic cholangiocarcinoma (iCCA) is closely correlated with hepatic progenitor cell (HPC) expansion and liver fibrosis. Brahma-related gene 1 (Brg1), an enzymatic subunit of the switch/sucrose nonfermentable complex that is critical in stem cell maintenance and tumor promotion, is prominently up-regulated in both HPCs and iCCA; however, its role in this correlation remains undefined. A retrospective cohort study indicated that high Brg1 expression suggests poor prognosis in patients with iCCA. In chronically injured livers induced by a 0.1% 3,5-diethoxycarbonyl-1,4-dihydrocollidine diet or bile duct ligation surgery, HPCs were dramatically activated, as indicated by their enhanced expression of Brg1 and a subset of stem cell markers; however, Brg1 ablation in HPCs strongly suppressed HPC expansion and liver fibrosis. Furthermore, in a chemically induced iCCA model, inhibition of Brg1 by a specific inhibitor or inducible gene ablation markedly improved histology and suppressed iCCA growth. Mechanistically, in addition to transcriptionally promoting both Wnt receptor genes and target genes, Brg1 was found to bind to the β-catenin/transcription factor 4 transcription complex, suggesting a possible approach for regulation of Wnt/β-catenin signaling. We have demonstrated the function of Brg1 in promoting HPC expansion, liver cirrhosis, and, ultimately, iCCA development in chronically injured livers, which is largely dependent on Wnt/β-catenin signaling. Our data suggest that therapies targeting Brg1-expressing HPCs are promising for the treatment of liver cirrhosis and iCCA.

Zhou Y ，Chen Y ，Zhang X ，Xu Q ，Wu Z ，Cao X ，Shao M ，Shu Y ，Lv T ，Lu C ，Xie M ，Wen T ，Yang J ，Shi Y ，Bu H ... -  《-》

Loss of Numb promotes hepatic progenitor expansion and intrahepatic cholangiocarcinoma by enhancing Notch signaling.

Numb, a stem cell fate determinant, acts as a tumor suppressor and is closely related to a wide variety of malignancies. Intrahepatic cholangiocarcinoma (iCCA) originates from hepatic progenitors (HPCs); however, the role of Numb in HPC malignant transformation and iCCA development is still unclear. A retrospective cohort study indicated that Numb was frequently decreased in tumor tissues and suggests poor prognosis in iCCA patients. Consistently, in a chemically induced iCCA mouse model, Numb was downregulated in tumor cells compared to normal cholangiocytes. In diet-induced chronic liver injury mouse models, Numb ablation significantly promoted histological impairment, HPC expansion, and tumorigenesis. Similarly, Numb silencing in cultured iCCA cells enhanced cell spheroid growth, invasion, metastasis, and the expression of stem cell markers. Mechanistically, Numb was found to bind to the Notch intracellular domain (NICD), and Numb ablation promoted Notch signaling; this effect was reversed when Notch signaling was blocked by γ-secretase inhibitor treatment. Our results suggested that loss of Numb plays an important role in promoting HPC expansion, HPC malignant transformation, and, ultimately, iCCA development in chronically injured livers. Therapies targeting suppressed Numb are promising for the treatment of iCCA.

Shu Y ，Xu Q ，Xu Y ，Tao Q ，Shao M ，Cao X ，Chen Y ，Wu Z ，Chen M ，Zhou Y ，Zhou P ，Shi Y ，Bu H ... -  《Cell Death & Disease》

Brahma-related gene-1 promotes tubular senescence and renal fibrosis through Wnt/β-catenin/autophagy axis.

Although accelerated cellular senescence is closely related to the progression of chronic kidney disease (CKD) and renal fibrosis, the underlying mechanisms remain largely unknown. Here, we reported that tubular aberrant expression of Brahma-related gene 1 (BRG1), an enzymatic subunit of the SWItch/Sucrose Non-Fermentable complex, is critically involved in tubular senescence and renal fibrosis. BRG1 was significantly up-regulated in the kidneys, predominantly in tubular epithelial cells, of both CKD patients and unilateral ureteral obstruction (UUO) mice. In vivo, shRNA-mediated knockdown of BRG1 significantly ameliorated renal fibrosis, improved tubular senescence, and inhibited UUO-induced activation of Wnt/β-catenin pathway. In mouse renal tubular epithelial cells (mTECs) and primary renal tubular cells, inhibition of BRG1 diminished transforming growth factor-β1 (TGF-β1)-induced cellular senescence and fibrotic responses. Correspondingly, ectopic expression of BRG1 in mTECs or normal kidneys increased p16INK4a, p19ARF, and p21 expression and senescence-associated β-galactosidase (SA-β-gal) activity, indicating accelerated tubular senescence. Additionally, BRG1-mediated pro-fibrotic responses were largely abolished by small interfering RNA (siRNA)-mediated p16INK4a silencing in vitro or continuous senolytic treatment with ABT-263 in vivo. Moreover, BRG1 activated the Wnt/β-catenin pathway, which further inhibited autophagy. Pharmacologic inhibition of the Wnt/β-catenin pathway (ICG-001) or rapamycin (RAPA)-mediated activation of autophagy effectively blocked BRG1-induced tubular senescence and fibrotic responses, while bafilomycin A1 (Baf A1)-mediated inhibition of autophagy abolished the effects of ICG-001. Further, BRG1 altered the secretome of senescent tubular cells, which promoted proliferation and activation of fibroblasts. Taken together, our results indicate that BRG1 induces tubular senescence by inhibiting autophagy via the Wnt/β-catenin pathway, which ultimately contributes to the development of renal fibrosis.

Gong W ，Luo C ，Peng F ，Xiao J ，Zeng Y ，Yin B ，Chen X ，Li S ，He X ，Liu Y ，Cao H ，Xu J ，Long H ... -  《-》

Hepatitis B virus X (HBx) induces tumorigenicity of hepatic progenitor cells in 3,5-diethoxycarbonyl-1,4-dihydrocollidine-treated HBx transgenic mice.

Hepatitis B virus X (HBx) protein is implicated in hepatitis B virus (HBV)-associated liver carcinogenesis. However, it remains unclear whether HBx-expressing hepatic progenitor cells (HPCs) are attributed to liver tumor formation. In this study, by using HBx transgenic mice and a 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC)-induced liver injury model, the relationship between HBx expression and tumorigenicity of HPCs was analyzed. Compared with control mice, an elevated number of EpCAM(+) cells with characteristics of HPCs was observed in HBx mice after 1 month and 4 months of DDC diet feeding. All HBx transgenic mice developed liver tumors characterized by histological features of both hepatocellular carcinoma (HCC) and cholangiocarcinoma after 7 months of DDC feeding. Notably, EpCAM(+) HPCs isolated from premalignant HBx mice exposed to a DDC diet for 4 months formed subcutaneous mixed-lineage tumors (four out of six) in nonobese diabetic/severe-combined immunodeficient (NOD/SCID) mice, and none of the cells from wildtype (WT) induced tumor, indicating that HBx may induce malignant transformation of HPCs that contributes to tumorigenesis. We also found higher titers of circulating interleukin (IL)-6, activities of IL-6/STAT3, and Wnt/β-catenin signaling pathways in HBx transgenic mice, suggesting HBx may induce intrinsic changes in HPCs by way of the above signaling that enables HPCs with tumorigenicity potential. Finally, clinical evidence showed that high HBx expression in human HBV-related HCC was statistically associated with expansion of EpCAM(+) or OV6(+) tumor cells and aggressive clinicopathologic features. HBx induces intrinsic cellular transformation promoting the expansion and tumorigenicity of HPCs in DDC-treated mice, which may be a possible origin for liver cancer induced by chronic hepatitis infection.

Wang C ，Yang W ，Yan HX ，Luo T ，Zhang J ，Tang L ，Wu FQ ，Zhang HL ，Yu LX ，Zheng LY ，Li YQ ，Dong W ，He YQ ，Liu Q ，Zou SS ，Lin Y ，Hu L ，Li Z ，Wu MC ，Wang HY ... -  《-》

β-Catenin Sustains and Is Required for YES-associated Protein Oncogenic Activity in Cholangiocarcinoma.

YES-associated protein (YAP) aberrant activation is implicated in intrahepatic cholangiocarcinoma (iCCA). Transcriptional enhanced associate domain (TEAD)-mediated transcriptional regulation is the primary signaling event downstream of YAP. The role of Wnt/β-Catenin signaling in cholangiocarcinogenesis remains undetermined. Here, we investigated the possible molecular interplay between YAP and β-Catenin cascades in iCCA. Activated AKT (Myr-Akt) was coexpressed with YAP (YapS127A) or Tead2VP16 via hydrodynamic tail vein injection into mouse livers. Tumor growth was monitored, and liver tissues were collected and analyzed using histopathologic and molecular analysis. YAP, β-Catenin, and TEAD interaction in iCCAs was investigated through coimmunoprecipitation. Conditional Ctnnb1 knockout mice were used to determine β-Catenin function in murine iCCA models. RNA sequencing was performed to analyze the genes regulated by YAP and/or β-Catenin. Immunostaining of total and nonphosphorylated/activated β-Catenin staining was performed in mouse and human iCCAs. We discovered that TEAD factors are required for YAP-dependent iCCA development. However, transcriptional activation of TEADs did not fully recapitulate YAP's activities in promoting cholangiocarcinogenesis. Notably, β-Catenin physically interacted with YAP in human and mouse iCCA. Ctnnb1 ablation strongly suppressed human iCCA cell growth and Yap-dependent cholangiocarcinogenesis. Furthermore, RNA-sequencing analysis revealed that YAP/ transcriptional coactivator with PDZ-binding motif (TAZ) regulate a set of genes significantly overlapping with those controlled by β-Catenin. Importantly, activated/nonphosphorylated β-Catenin was detected in more than 80% of human iCCAs. YAP induces cholangiocarcinogenesis via TEAD-dependent transcriptional activation and interaction with β-Catenin. β-Catenin binds to YAP in iCCA and is required for YAP full transcriptional activity, revealing the functional crosstalk between YAP and β-Catenin pathways in cholangiocarcinogenesis.

Zhang Y ，Xu H ，Cui G ，Liang B ，Chen X ，Ko S ，Affo S ，Song X ，Liao Y ，Feng J ，Wang P ，Wang H ，Xu M ，Wang J ，Pes GM ，Ribback S ，Zeng Y ，Singhi A ，Schwabe RF ，Monga SP ，Evert M ，Tang L ，Calvisi DF ，Chen X ... -  《-》