Targeting PKM2 improves the gemcitabine sensitivity of intrahepatic cholangiocarcinoma cells via inhibiting ß-catenin signaling pathway.

Gemcitabine is considered the standard first-line chemotherapeutic agent for patients with intrahepatic cholangiocarcinoma (ICC). However, its therapeutic efficacy is hampered by the development of chemoresistance. Pyruvate kinase M2 (PKM2), a crucial mediator of the final step in glycolysis, has been implicated in the origination and advancement of diverse malignancies. Its expression is increased in many tumor types and this may correlate with increased drug sensitivity. However, the specific effect of PKM2 on the gemcitabine sensitivity in ICC remains to be elucidated. In this research, we aimed to elucidate the role and functional significance of PKM2 in ICC, as well as the heightened susceptibility of ICC cells to gemcitabine by targeting PKM2 and the underlying molecular mechanisms. Immunohistochemical and immunofluorescence analyses revealed elevated expression of PKM2 in both tumor cells and macrophages in human ICC tissues. Reducing PKM2 levels significantly restrained the proliferation of tumor cells, impeded cell cycle advance, induced programmed cell death, and suppressed metastasis. In addition, knockdown or pharmacological inhibition of PKM2 could enhance the response of ICC cells to gemcitabine in vitro. Interestingly, conditioned medium co-culture system suggested that conditioned medium from M2 macrophages increased gemcitabine sensitivity of ICC cells. However, silencing PKM2 or pharmacological inhibition of PKM2 in M2 macrophages did not ameliorate the gemcitabine resistance mediated by M2 macrophages derived conditioned medium. Mechanistically, downregulation of PKM2 repressed the expression of ß-catenin and its downstream transcriptional targets, thereby hindering the propagation of ß-catenin signaling cascade. Finally, the results of the subcutaneous xenograft experiment in nude mice provided compelling evidence of a synergistic interaction between PKM2-IN-1 and gemcitabine in vivo. In summary, we reported that PKM2 may function as an advantageous target for increasing the sensitivity of ICC to gemcitabine treatment. Targeting PKM2 improves the gemcitabine sensitivity of ICC cells via inhibiting ß-catenin signaling pathway.

Glycolysis in Chronic Liver Diseases: Mechanistic Insights and Therapeutic Opportunities.

Chronic liver diseases (CLDs) cover a spectrum of liver diseases, ranging from nonalcoholic fatty liver disease to liver cancer, representing a growing epidemic worldwide with high unmet medical needs. Glycolysis is a conservative and rigorous process that converts glucose into pyruvate and sustains cells with the energy and intermediate products required for diverse biological activities. However, abnormalities in glycolytic flux during CLD development accelerate the disease progression. Aerobic glycolysis is a hallmark of liver cancer and is responsible for a broad range of oncogenic functions including proliferation, invasion, metastasis, angiogenesis, immune escape, and drug resistance. Recently, the non-neoplastic role of aerobic glycolysis in immune activation and inflammatory disorders, especially CLD, has attracted increasing attention. Several key mediators of aerobic glycolysis, including HIF-1α and pyruvate kinase M2 (PKM2), are upregulated during steatohepatitis and liver fibrosis. The pharmacological inhibition or ablation of PKM2 effectively attenuates hepatic inflammation and CLD progression. In this review, we particularly focused on the glycolytic and non-glycolytic roles of PKM2 in the progression of CLD, highlighting the translational potential of a glycolysis-centric therapeutic approach in combating CLD.

Enhanced Expression of ARK5 in Hepatic Stellate Cell and Hepatocyte Synergistically Promote Liver Fibrosis.

AMPK-related protein kinase 5 (ARK5) is involved in a broad spectrum of physiological and cell events, and aberrant expression of ARK5 has been observed in a wide variety of solid tumors, including liver cancer. However, the role of ARK5 in liver fibrosis remains largely unexplored. We found that ARK5 expression was elevated in mouse fibrotic livers, and showed a positive correlation with the progression of liver fibrosis. ARK5 was highly expressed not only in activated hepatic stellate cells (HSCs), but also in hepatocytes. In HSCs, ARK5 prevents the degradation of transforming growth factor ß type I receptor (TßRI) and mothers against decapentaplegic homolog 4 (Smad4) proteins by inhibiting the expression of Smad ubiquitin regulatory factor 2 (Smurf2), thus maintaining the continuous transduction of the transforming growth factor ß (TGF-ß) signaling pathway, which is essential for cell activation, proliferation and survival. In hepatocytes, ARK5 induces the occurrence of epithelial-mesenchymal transition (EMT), and also promotes the secretion of inflammatory factors. Inflammatory factors, in turn, further enhance the activation of HSCs and deepen the degree of liver fibrosis. Notably, we demonstrated in a mouse model that targeting ARK5 with the selective inhibitor HTH-01-015 attenuates CCl4-induced liver fibrosis in mice. Taken together, the results indicate that ARK5 is a critical driver of liver fibrosis, and promotes liver fibrosis by synergy between HSCs and hepatocytes.

NUSAP1, a novel stemness-related protein, promotes early recurrence of hepatocellular carcinoma.

Early recurrence (within 2 years after resection) is the primary cause of poor outcomes among hepatocellular carcinoma (HCC) patients, and liver cancer stem cells are the main contributors to postsurgical HCC recurrence. Nucleolar and spindle-associated protein 1 (NUSAP1) has been reported to be involved in tumor progression. We investigated the function and clinical value of NUSAP1 in early recurrence of HCC. Data from public datasets and our cohort were used to assess the association between NUSAP1 expression and early HCC recurrence. Gain- and loss-of-function experiments were carried out in vivo and in vitro. The predictive effect of NUSAP1 on early HCC recurrence was further evaluated by a validation cohort. We found that elevated NUSAP1 expression in HCC specimens was correlated with poor outcome, especially in cases with postoperative early recurrence. Functional studies indicated that NUSAP1 significantly promotes HCC progression. A postsurgical recurrence murine model further revealed that upregulated NUSAP1 dramatically increased the likelihood of HCC early recurrence. RNA sequencing data revealed that the gene sets of cancer stemness and the signal transducer and activator of transcription 3 (STAT3) pathway were enriched by NUSAP1 overexpression. Mechanistically, NUSAP1 enhanced cancer stemness through stimulating STAT3 nuclear translocation and activation through receptor of activated protein C kinase 1 (RACK1). In a validation cohort with 112 HCC patients, NUSAP1 effectively predicted HCC early recurrence. Our results indicated that NUSAP1 promotes early recurrence of HCC by sustaining cancer stemness and could serve as a valuable predictive indicator for postsurgical intervention in HCC patients.

Microskeletal stiffness promotes aortic aneurysm by sustaining pathological vascular smooth muscle cell mechanosensation via Piezo1.

Mechanical overload of the vascular wall is a pathological hallmark of life-threatening abdominal aortic aneurysms (AAA). However, how this mechanical stress resonates at the unicellular level of vascular smooth muscle cells (VSMC) is undefined. Here we show defective mechano-phenotype signatures of VSMC in AAA measured with ultrasound tweezers-based micromechanical system and single-cell RNA sequencing technique. Theoretical modelling predicts that cytoskeleton alterations fuel cell membrane tension of VSMC, thereby modulating their mechanoallostatic responses which are validated by live micromechanical measurements. Mechanistically, VSMC gradually adopt a mechanically solid-like state by upregulating cytoskeleton crosslinker, α-actinin2, in the presence of AAA-promoting signal, Netrin-1, thereby directly powering the activity of mechanosensory ion channel Piezo1. Inhibition of Piezo1 prevents mice from developing AAA by alleviating pathological vascular remodeling. Our findings demonstrate that deviations of mechanosensation behaviors of VSMC is detrimental for AAA and identifies Piezo1 as a novel culprit of mechanically fatigued aorta in AAA.

Lung-derived HMGB1 is detrimental for vascular remodeling of metabolically imbalanced arterial macrophages.

Pulmonary disease increases the risk of developing abdominal aortic aneurysms (AAA). However, the mechanism underlying the pathological dialogue between the lungs and aorta is undefined. Here, we find that inflicting acute lung injury (ALI) to mice doubles their incidence of AAA and accelerates macrophage-driven proteolytic damage of the aortic wall. ALI-induced HMGB1 leaks and is captured by arterial macrophages thereby altering their mitochondrial metabolism through RIPK3. RIPK3 promotes mitochondrial fission leading to elevated oxidative stress via DRP1. This triggers MMP12 to lyse arterial matrix, thereby stimulating AAA. Administration of recombinant HMGB1 to WT, but not Ripk3-/- mice, recapitulates ALI-induced proteolytic collapse of arterial architecture. Deletion of RIPK3 in myeloid cells, DRP1 or MMP12 suppression in ALI-inflicted mice repress arterial stress and brake MMP12 release by transmural macrophages thereby maintaining a strengthened arterial framework refractory to AAA. Our results establish an inter-organ circuitry that alerts arterial macrophages to regulate vascular remodeling.