Liver Extracellular Vesicles and Particles Enriched ß-Sitosterol Effectively Promote Liver Regeneration in Mice.

Background: The liver's regenerative capacity allows it to repair itself after injury. Extracellular vesicles and particles (EVPs) in the liver's interstitial space are crucial for signal transduction, metabolism, and immune regulation. Understanding the role and mechanism of liver-derived EVPs in regeneration is significant, particularly after partial hepatectomy, where the mechanisms remain unclear. Methods: A 70% hepatectomy model was established in mice, and EVPs were isolated and characterized using electron microscopy, nanocharacterization, and Western blot analysis. Combined metabolomic and transcriptomic analyses revealed ß-sitosterol enrichment in EVPs and activation of the Hedgehog signaling pathway during regeneration. The role of ß-sitosterol in EVPs on the Hedgehog pathway and its targets were identified using qRT-PCR, Western blot analysis. The regulation of carnitine synthesis by this pathway was determined using a dual luciferase assay. The effect of a ß-sitosterol diet on liver regeneration was verified in mice. Results: After 70% hepatectomy, the liver successfully regenerated without liver failure or death. At 24 hours post-surgery, tissue staining showed transient regeneration-associated steatosis (TRAS), with increased Ki67 positivity at 48 hours. EVPs displayed a spherical lipid bilayer structure with particle sizes of 70-130 nm. CD9, CD63, and CD81 in liver-derived EVPs were confirmed. Transcriptomic and metabolomic analyses showed EVPs supplementation significantly promoted carnitine synthesis and fatty acid oxidation. Tissue staining confirmed accelerated TRAS resolution and enhanced liver regeneration with EVP supplementation. Mass spectrometry identified ß-sitosterol in EVPs, which binds to Smo protein, activating the Hedgehog pathway. This led to the nuclear transport of Gli3, stimulating Setd5 transcription and inducing carnitine synthesis, thereby accelerating fatty acid oxidation. Mice with increased ß-sitosterol intake showed faster TRAS resolution and liver regeneration compared to controls. Conclusion: Liver-derived EVPs promote regeneration after partial hepatectomy. ß-sitosterol from EVPs accelerates fatty acid oxidation and promotes liver regeneration by activating Hedgehog signaling pathway.

Erratum: A positive feedback loop of CENPU/E2F6/E2F1 facilitates proliferation and metastasis via ubiquitination of E2F6 in hepatocellular carcinoma: Erratum.

[This corrects the article DOI: 10.7150/ijbs.69495.].

METTL5 stabilizes c-Myc by facilitating USP5 translation to reprogram glucose metabolism and promote hepatocellular carcinoma progression.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most prevalent cancers in the world, with a high likelihood of metastasis and a dismal prognosis. The reprogramming of glucose metabolism is critical in the development of HCC. The Warburg effect has recently been confirmed to occur in a variety of cancers, including HCC. However, little is known about the molecular biological mechanisms underlying the Warburg effect in HCC cells. In this study, we sought to better understand how methyltransferase 5, N6-adenosine (METTL5) controls the development of HCC and the Warburg effect. METHODS: In the current study, quantitative real-time polymerase chain reaction and Western blotting were used to detect the expression of METTL5 in HCC tissues and cell lines. Several different cell models and animal models were established to determine the role of METTL5 in glucose metabolism reprogramming and the underlying molecular mechanism of HCC. Glutathione-S-transferase pulldown, coimmunoprecipitation, RNA sequencing, non-targeted metabolomics, polysome profiling, and luciferase reporter assays were performed to investigate the molecular mechanisms of METTL5 in HCC cells. RESULTS: We discovered that METTL5 drove glucose metabolic reprogramming to promote the proliferation and metastasis of HCC. Mechanistically, upregulation of METTL5 promoted c-Myc stability and thus activated its downstream glycolytic genes lactate dehydrogenase A (LDHA), enolase 1 (ENO1), triosephosphate isomerase 1 (TPI1), solute carrier family 2 member 1 (SLC2A1), and pyruvate kinase M2 (PKM2). The c-Box and ubiquitin binding domain (UBA) regions of ubiquitin specific peptidase 5 (USP5) binded to c-Myc protein and inhibited K48-linked polyubiquitination of c-Myc. Further study revealed that METTL5 controled the USP5 translation process, which in turn regulated the ubiquitination of c-Myc. Furthermore, we identified cAMP responsive element binding protein 1 (CREB1)/P300 as a critical transcriptional regulator of METTL5 that promoted the transcription of METTL5 in HCC. In patient-derived tumor xenograft (PDX) models, adenovirus-mediated knockout of METTL5 had a good antitumor effect and prolonged the survival of PDX-bearing mice. CONCLUSIONS: These findings point to a novel mechanism by which CREB1/P300-METTL5-USP5-c-Myc controls abnormal glucose metabolism and promotes tumor growth, suggesting that METTL5 is a potential therapeutic target and prognostic biomarker for HCC.

Multi-omics analysis revealed the role of extracellular vesicles in hepatobiliary & pancreatic tumor.

Liquid biopsy is rapidly growing into a hot research field due to its unique advantages of minimal invasiveness, and extracellular vesicle (EVs) are also expected to become an important pillar in the diagnostic technology system as a newly discovered active substance carrier. More and more research has highlighted the important contribution of EVs in the progress of tumor. Molecular changes during disease progression could be detected in EVs. However, the diagnostic applications of EVs are not generally understood. Combined with the characteristics of hepatobiliary and pancreatic tumor, we summarized the recent developments in various omics analysis of EVs. Furtherly, we explored the role of EVs in the early diagnosis of hepatobiliary and pancreatic tumors by multi-omics analysis.

A positive feedback loop of CENPU/E2F6/E2F1 facilitates proliferation and metastasis via ubiquitination of E2F6 in hepatocellular carcinoma.

Centromere protein U (CENPU), a centromere-binding protein required for cellular mitosis, has been reported to be closely associated with carcinogenesis in multiple malignancies; however, the role of CENPU in hepatocellular carcinoma (HCC) is still unclear. Herein, we investigated its biological role and molecular mechanism in the development of HCC. High CENPU expression in HCC tissue was observed and correlated positively with a poor prognosis in HCC patients. CENPU knockdown inhibited the proliferation, metastasis, and G1/S transition of HCC cells in vivo and in vitro, while ectopic expression of CENPU exerted the opposite effects. Mechanistically, CENPU physically interacted with E2F6 and promoted its ubiquitin-mediated degradation, thus affecting the transcription level of E2F1 and further accelerating the G1/S transition to promote HCC cell proliferation. E2F1 directly binds to the CENPU promoter and increases the transcription of CENPU, thereby forming a positive regulatory loop. Collectively, our findings indicate a crucial role for CENPU in E2F1-mediated signalling for cell cycle progression and reveal a role for CENPU as a predictive biomarker and therapeutic target for HCC patients.

A novel lncRNA RP11-386G11.10 reprograms lipid metabolism to promote hepatocellular carcinoma progression.

OBJECTIVE: Emerging studies suggest that long non-coding RNAs (lncRNAs) play crucial roles in hepatocellular carcinoma (HCC). A rapidly increasing number of studies have shown that metabolic changes including lipid metabolic reprogramming play a significant role in the progression of HCC. But it remains to be elucidated how lncRNAs affect tumor cell metabolism. METHODS: Through analysis and screening of The Cancer Genome Atlas-Liver Hepatocellular Carcinoma (TCGA-LIHC) dataset, we found a novel lncRNA RP11-386G11.10 was overexpressed, related to prognosis, conserved and non-protein-coding in HCC and related to poor prognosis. Then, CCK-8, colony formation, Transwell invasion, wound healing assays were performed and nude mouse subcutaneous tumour formation and lung metastasis models were established to explore the effect of RP11-386G11.10 on HCC tumour growth and metastasis. Chromatography-mass spectrometry (GC-MS) and Nile red staining detected the effect of RP11-386G11.10 on lipid metabolism in HCC. Mechanistically, we clarified the RP11-386G11.10/miR-345-3p/HNRNPU signalling pathway through dual luciferase reporter, RNA immunoprecipitation (RIP) and chromatin immunoprecipitation (ChIP) assays and identified ZBTB7A as a transcription factor of RP11-386G11.10. RESULTS: RP11-386G11.10 was overexpressed in HCC and positively correlated with tumour size, TNM stage, and poor prognosis in HCC patients. RP11-386G11.10 promoted the proliferation and metastasis of HCC cells in vitro and in vivo. Mechanistically, RP11-386G11.10 acted as a competing endogenous RNA (ceRNA) for miR-345-3p to regulate the expression of HNRNPU and its downstream lipogenic enzymes, leading to lipid accumulation in HCC cells and promoting their growth and metastasis. In addition, we identified ZBTB7A as a transcription factor of RP11-386G11.10. Moreover, HNRNPU promoted the expression of ZBTB7A in HCC cells, thereby increasing the transcriptional activity of RP11-386G11.10, and forming a positive feedback loop, ultimately leading continuous lipid accumulation, growth and metastasis in HCC cells. CONCLUSIONS: Our results indicated that the lncRNA RP11-386G11.10 was a novel oncogenic lncRNA that was strongly correlated with the poor prognosis of HCC. The ZBTB7A-RP11-386G11.10-HNRNPU positive feedback loop promoted the progression of HCC by regulating lipid anabolism. RP11-386G11.10 may become a new diagnostic and prognostic biomarker and therapy target for HCC.