RGS19 activates the MYH9/ß-catenin/c-Myc positive feedback loop in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most common fatal cancers worldwide, and the identification of novel treatment targets and prognostic biomarkers is urgently needed because of its unsatisfactory prognosis. Regulator of G-protein signaling 19 (RGS19) is a multifunctional protein that regulates the progression of various cancers. However, the specific function of RGS19 in HCC remains unclear. The expression of RGS19 was determined in clinical HCC samples. Functional and molecular biology experiments involving RGS19 were performed to explore the potential mechanisms of RGS19 in HCC. The results showed that the expression of RGS19 is upregulated in HCC tissues and is significantly associated with poor prognosis in HCC patients. RGS19 promotes the proliferation and metastasis of HCC cells in vitro and in vivo. Mechanistically, RGS19, via its RGS domain, stabilizes the MYH9 protein by directly inhibiting the interaction of MYH9 with STUB1, which has been identified as an E3 ligase of MYH9. Moreover, RGS19 activates ß-catenin/c-Myc signaling via MYH9, and RGS19 is also a transcriptional target gene of c-Myc. A positive feedback loop formed by RGS19, MYH9, and the ß-catenin/c-Myc axis was found in HCC. In conclusion, our research revealed that competition between RGS19 and STUB1 is a critical mechanism of MYH9 regulation and that the RGS19/MYH9/ß-catenin/c-Myc feedback loop may represent a promising strategy for HCC therapy.

A comprehensive analysis of m6A/m7G/m5C/m1A-related gene expression and immune infiltration in liver ischemia-reperfusion injury by integrating bioinformatics and machine learning algorithms.

BACKGROUND: Liver ischemia-reperfusion injury (LIRI) is closely associated with immune infiltration, which commonly occurs after liver surgery, especially liver transplantation. Therefore, it is crucial to identify the genes responsible for LIRI and develop effective therapeutic strategies that target immune response. Methylation modifications in mRNA play various crucial roles in different diseases. This study aimed to identify potential methylation-related markers in patients with LIRI and evaluate the corresponding immune infiltration. METHODS: Two Gene Expression Omnibus datasets containing human liver transplantation data (GSE12720 and GSE151648) were downloaded for integrated analysis. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses were conducted to investigate the functional enrichment of differentially expressed genes (DEGs). Differentially expressed methylation-related genes (DEMRGs) were identified by overlapping DEG sets and 65 genes related to N6-methyladenosine (m6A), 7-methylguanine (m7G), 5-methylcytosine (m5C), and N1-methyladenosine (m1A). To evaluate the relationship between DEMRGs, a protein-protein interaction (PPI) network was utilized. The core DEMRGs were screened using three machine learning algorithms: least absolute shrinkage and selection operator, random forest, and support vector machine-recursive feature elimination. After verifying the diagnostic efficacy using the receiver operating characteristic curve, we validated the expression of the core DEMRGs in clinical samples and performed relative cell biology experiments. Additionally, the immune status of LIRI was comprehensively assessed using the single sample gene set enrichment analysis algorithm. The upstream microRNA and transcription factors of the core DEMRGs were also predicted. RESULTS: In total, 2165 upregulated and 3191 downregulated DEGs were identified, mainly enriched in LIRI-related pathways. The intersection of DEGs and methylation-related genes yielded 28 DEMRGs, showing high interaction in the PPI network. Additionally, the core DEMRGs YTHDC1, METTL3, WTAP, and NUDT3 demonstrated satisfactory diagnostic efficacy and significant differential expression and corresponding function based on cell biology experiments. Furthermore, immune infiltration analyses indicated that several immune cells correlated with all core DEMRGs in the LIRI process to varying extents. CONCLUSIONS: We identified core DEMRGs (YTHDC1, METTL3, WTAP, and NUDT3) associated with immune infiltration in LIRI through bioinformatics and validated them experimentally. This study may provide potential methylation-related gene targets for LIRI immunotherapy.

TMEM147 aggravates the progression of HCC by modulating cholesterol homeostasis, suppressing ferroptosis, and promoting the M2 polarization of tumor-associated macrophages.

BACKGROUND: The endoplasmic reticulum (ER) regulates critical processes, including lipid synthesis, which are affected by transmembrane proteins localized in the ER membrane. One such protein, transmembrane protein 147 (TMEM147), has recently been implicated for its role in hepatocellular carcinoma (HCC) tumorigenesis; however, the mechanisms remain unclear. We investigated the role of TMEM147 in HCC and the underlying mechanisms. METHODS: TMEM147 expression was examined in human HCC cells and adjacent non-tumorous tissues using quantitative reverse transcription-polymerase chain reaction, western blotting, and immunohistochemistry. In vitro and in vivo studies were conducted to investigate the impact of TMEM147 on the progression of HCC. Proteins interacting with TMEM147 were identified via RNA-seq, immunoprecipitation, and mass spectrometry analyses. Lipidomic analysis and enzyme-linked immunosorbent assay (ELISA) were employed to determine and analyze cholesterol and 27-hydroxycholesterol (27HC) contents. Extensive experimental techniques were used to study ferroptosis in HCC cells. The fatty acid content of macrophages affected by TMEM147 was quantified using ELISA. Macrophage phenotypes were determined using immunofluorescence assay and flow cytometric analysis. RESULTS: TMEM147 mRNA and protein levels were increased in HCC cells, and the increased TMEM147 expression was associated with a poor survival. TMEM147 promoted tumor cell proliferation and metastases in vitro and in vivo. The protein was found to interact with the key enzyme 7-dehydrocholesterol reductase (DHCR7), which affected cellular cholesterol homeostasis and increased the extracellular levels of 27HC in HCC cells. TMEM147 also promoted the expression of DHCR7 by enhancing the activity of signal transducer and activator of transcription 2. 27HC expression upregulated glutathione peroxidase 4 in HCC, leading to ferroptosis resistance and promotion of HCC proliferation. HCC cell-derived 27HC expression increased the lipid metabolism in macrophages and activated peroxisome proliferator-activated receptor-γ signaling, thereby activating M2 macrophage polarization and promoting HCC cell invasion and migration. CONCLUSIONS: Our results indicate that TMEM147 confers ferroptosis resistance and M2 macrophage polarization, which are primarily dependent on the upregulation of cellular cholesterol homeostasis and 27HC secretion, leading to cancer growth and metastasis. These findings suggest that the TMEM147/STAT2/DHCR7/27HC axis in the tumor microenvironment may serve as a promising therapeutic target for HCC.

RNF125 attenuates hepatocellular carcinoma progression by downregulating SRSF1-ERK pathway.

Hepatocellular carcinoma (HCC) is one of the most deadly malignant cancers worldwide. Research into the crucial genes responsible for maintaining the aggressive behaviour of cancer cells is important for the clinical treatment of HCC. The purpose of this study was to determine whether the E3 ubiquitin ligase Ring Finger Protein 125 (RNF125) plays a role in the proliferation and metastasis of HCC. RNF125 expression in human HCC samples and cell lines was investigated using TCGA dataset mining, qRT‒PCR, western blot, and immunohistochemistry assays. In addition, 80 patients with HCC were studied for the clinical value of RNF125. Furthermore, the molecular mechanism by which RNF125 contributes to hepatocellular carcinoma progression was determined with mass spectrometry (MS), coimmunoprecipitation (Co-IP), dual-luciferase reporter assays, and ubiquitin ladder assays. We found that RNF125 was markedly downregulated in HCC tumour tissues, which was associated with a poor prognosis for patients with HCC. Moreover, the overexpression of RNF125 inhibited HCC proliferation and metastasis both in vitro and in vivo, whereas the knockdown of RNF125 exerted antithetical effects. Mechanistically, mass spectrometry analysis revealed a protein interaction between RNF125 and SRSF1, and RNF125 accelerated the proteasome-mediated degradation of SRSF1, which impeded HCC progression by inhibiting the ERK signalling pathway. Furthermore, RNF125 was detected to be the downstream target of miR-103a-3p. In this study, we identified that RNF125 is a tumour suppressor in HCC and inhibits HCC progression by inhibiting the SRSF1/ERK pathway. These findings provide a promising treatment target for HCC.

HLTF promotes hepatocellular carcinoma progression by enhancing SRSF1 stability and activating ERK/MAPK pathway.

Helicase-like transcription factor (HLTF) has been found to be involved in the progression of several tumors, but the role of HLTF in hepatocellular carcinoma (HCC) progression has not been studied. Here, our study explored the underlying mechanism of HLTF in HCC progression for the first time. Database analysis and clinical sample examination indicated that HLTF was upregulated in HCC tissues and was related to poor clinicopathological features in patients. Upregulation of HLTF accelerated the growth and metastasis of HCC cells both in vitro and in vivo. Bioinformatics analysis and subsequent experiments revealed that ERK/MAPK signaling pathway activation was vital to HLTF-mediated proliferation and metastasis in HCC cells. Moreover, HLTF was demonstrated to interact with SRSF1 and contribute to its protein stability to activate the ERK/MAPK signaling pathway and enhance HCC growth and metastasis. In addition, miR-511-5p was expressed at a low level in HCC tissues, was negatively correlated HLTF, and regulated HLTF expression. Our study shows that HLTF plays an oncogenic role in HCC progression and provides a novel biomarker and therapeutic target for the diagnosis and treatment of HCC.

Comprehensive analysis of the prognostic value and functions of prefoldins in hepatocellular carcinoma.

Prefoldins (PFDNs), a group of proteins known to be associated with cytoskeletal rearrangement, are involved in tumor progression in various cancer types. However, little is known about the roles of PFDNs in hepatocellular carcinoma (HCC). Herein, we investigated the transcriptional and survival data of PFDNs from The Cancer Genome Atlas (TCGA) database. Gene Ontology (GO), Gene Set Enrichment Analysis (GSEA), and single-sample gene set enrichment analysis (ssGSEA) were used to evaluate the potential functions of PFDN1/2/3/4. We also detected the expression of PFDN1/2/3/4 via immunohistochemistry (IHC), Western blotting, and real-time PCR in our clinical samples. We found that the PFDN family showed elevated expression in HCC tissues, while only PFDN1/2/3/4 were found to be significantly correlated with poor prognosis of patients with HCC in the TCGA database. Further investigation was associated with PFDN1-4. We found that the expression of PFDN1/2/3/4 was significantly associated with advanced clinicopathologic features. Apart from the TCGA database, IHC, real-time PCR, and immunoblotting identified the overexpression of PFDN1/2/3/4 in HCC tissues and HCC cell lines. Taken together, these results indicated that PFDN1/2/3/4 might be novel prognostic biomarkers and treatment targets for patients with HCC.

LncRNA Hnf4αos exacerbates liver ischemia/reperfusion injury in mice via Hnf4αos/Hnf4α duplex-mediated PGC1α suppression.

LncRNAs are involved in the pathophysiologic processes of multiple diseases, but little is known about their functions in hepatic ischemia/reperfusion injury (HIRI). As a novel lncRNA, the pathogenetic significance of hepatic nuclear factor 4 alpha, opposite strand (Hnf4αos) in hepatic I/R injury remains unclear. Here, differentially expressed Hnf4αos and Hnf4α antisense RNA 1 (Hnf4α-as1) were identified in liver tissues from mouse ischemia/reperfusion models and patients who underwent liver resection surgery. Hnf4αos deficiency in Hnf4αos-KO mice led to improved liver function, alleviated the inflammatory response and reduced cell death. Mechanistically, we found a regulatory role of Hnf4αos-KO in ROS metabolism through PGC1α upregulation. Hnf4αos also promoted the stability of Hnf4α mRNA through an RNA/RNA duplex, leading to the transcriptional activation of miR-23a and miR-23a depletion was required for PGC1α function in hepatoprotective effects on HIRI. Together, our findings reveal that Hnf4αos elevation in HIRI leads to severe liver damage via Hnf4αos/Hnf4α/miR-23a axis-mediated PGC1α inhibition.

Identification of CFHR4 as a Potential Prognosis Biomarker Associated With lmmune Infiltrates in Hepatocellular Carcinoma.

Background: Complement factor H-related 4 (CFHR4) is a protein-coding gene that plays an essential role in multiple diseases. However, the prognostic value of CFHR4 in hepatocellular carcinoma (HCC) is unknown. Methods: Using multiple databases, we investigated CFHR4 expression levels in HCC and multiple cancers. The relationship between CFHR4 expression levels and clinicopathological variables was further analyzed. Various potential biological functions and regulatory pathways of CFHR4 in HCC were identified by performing a Gene Ontology (GO) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and Gene Set Enrichment Analysis (GSEA). Single-sample gene set enrichment analysis (ssGSEA) was performed to confirm the correlation between CFHR4 expression and immune cell infiltration. The correlations between CFHR4 expression levels in HCC and N6-methyladenosine (m6A) modifications and the competing endogenous RNA (ceRNA) regulatory networks were confirmed in TCGA cohort. Results: CFHR4 expression levels were significantly decreased in HCC tissues. Low CFHR4 expression in HCC tissues was significantly correlated with the patients' sex, race, age, TNM stage, pathological stage, tumor status, residual tumor, histologic grade and alpha fetal protein (AFP) level. GO and KEGG analyses revealed that differentially expressed genes related to CFHR4 may be involved in the synaptic membrane, transmembrane transporter complex, gated channel activity, chemical carcinogenesis, retinol metabolism, calcium signaling pathway, PPAR signaling pathway, insulin and gastric acid secretion. GSEA revealed that the FCGR-activated reaction, PLK1 pathway, ATR pathway, MCM pathway, cascade reactions of PI3K and FGFR1, reactant-mediated MAPK activation and FOXM1 pathway were significantly enriched in HCC with low CFHR4 expression. Moreover, CFHR4 expression was inversely correlated the levels of infiltrating Th2 cells, NK CD56bright cells and Tfh cells. In contrast, we observed positive correlations with the levels of infiltrating DCs, neutrophils, Th17 cells and mast cells. CFHR4 expression showed a strong correlation with various immunomarker groups in HCC. In addition, high CFHR4 expression significantly prolonged the overall survival (OS), disease-specific survival (DSS) and progression-free interval (PFI). We observed a substantial correlation between the expression of CFHR4 and multiple N6-methyladenosine genes in HCC and constructed potential CFHR4-related ceRNA regulatory networks. Conclusions: CFHR4 might be a potential therapeutic target for improving the HCC prognosis and is closely related to immune cell infiltration.

NNMT promotes the progression of intrahepatic cholangiocarcinoma by regulating aerobic glycolysis via the EGFR-STAT3 axis.

Nicotinamide N-methyltransferase (NNMT), a member of the N-methyltransferase family, plays an important role in tumorigenesis. However, its expression and biological functions in intrahepatic cholangiocarcinoma (iCCA) remain to be established. In our study, we identified NNMT as an oncogene in iCCA and provided mechanistic insights into the roles of NNMT in iCCA progression. High NNMT expression in iCCA tissues was identified using western blotting and immunohistochemistry (IHC). We identified a significantly higher NNMT expression level in human iCCA tissues than that in adjacent normal tissues. Increased NNMT expression promoted iCCA cell proliferation and metastasis in vitro and in vivo. Mechanistically, NNMT inhibited the level of histone methylation in iCCA cells by consuming the methyl donor S-adenosyl methionine (SAM), thereby promoting the expression of epidermal growth factor receptor (EGFR). EGFR may activate the aerobic glycolysis pathway in iCCA cells by activating the STAT3 signaling pathway. In conclusion, we identified NNMT as an oncogene in iCCA and provided mechanistic insights into the roles of NNMT in iCCA progression.

Modified Laparoscopic Percutaneous Extraperitoneal Closure Using a Sledge-Shaped Needle for Inguinal Hernia and Hydrocele in 1199 Pediatric Patients.

Background: The objective of the study is to determine the safety and efficiency of the modified laparoscopic percutaneous extraperitoneal closure (LPEC) to treat pediatric patients with inguinal hernia or hydrocele. Methods: From January 2014 to July 2018, the patients with inguinal hernia or hydrocele who were operated on using modified LPEC were included. We modified LPEC with a sledge-shaped needle and reinforcement flag. By means of medial umbilical fold, the reinforcement surgery should be performed on the patients with huge internal rings (diameter >1.5 cm). Operative time, complication rate, incidence of reinforcement, and contralateral patent processus vaginalis were described between inguinal hernia and hydrocele. Results: In this study, 764 patients with inguinal hernia and 435 patients with hydrocele were successfully performed by the modified LPEC. The 383 (50.1%) patients with inguinal hernia and 266 (61.1%) patients with hydrocele were identified with a contralateral patency of internal ring and underwent simultaneous prophylactic surgery. During surgical procedures, the medial umbilical fold reinforcement (inguinal hernia/hydrocele = 50/1) was performed on 51 patients. Hernia recurrence occurred in 2 cases. All patients had a good cosmetic appearance without additional dissection. Conclusion: Modified LPEC using a sledge-shaped needle and applying the medial umbilical fold reinforcement is a safe and effective surgical procedure.