Role of MIC levels and 23S rRNA mutation sites to clarithromycin in 14-day clarithromycin bismuth quadruple therapy for Helicobacter pylori eradication: A prospective trial in Beijing.

Background: Rising clarithromycin resistance undermines Helicobacter pylori (H. pylori) treatment efficacy. We aimed to determine clarithromycin's minimum inhibitory concentration (MIC) levels and identify specific mutation sites in the 23S ribosomal subunit (23S rRNA) that predict treatment outcomes in a 14-day regimen of clarithromycin bismuth quadruple therapy (amoxicillin 1g, clarithromycin 500 mg, rabeprazole 10 mg, and colloidal bismuth pectin 200 mg). Materials and methods: We included adult H. pylori patients who hadn't previously undergone clarithromycin-based treatment, either as initial or rescue therapy. Exclusions were made for penicillin allergy, recent use of related medications, severe illnesses, or inability to cooperate. Patients underwent a 14-day clarithromycin bismuth quadruple therapy. Gastric mucosa specimens were obtained during endoscopy before eradication. MIC against amoxicillin and clarithromycin was determined using the E-test method. The receiver operating characteristic (ROC) curve helped to find the optimal clarithromycin resistance MIC breakpoint. Genetic sequences of H. pylori 23S rRNA were identified through Sanger Sequencing. (ChiCTR2200061476). Results: Out of 196 patients recruited, 92 met the inclusion criteria for the per-protocol (PP) population. The overall intention-to-treat (ITT) eradication rate was 80.00 % (84/105), while the modified intention-to-treat (MITT) and PP eradication rates were 90.32 % (84/93) and 91.30 % (84/92) respectively. No amoxicillin resistance was observed, but clarithromycin resistance rates were 36.19 % (38/105), 35.48 % (33/93), and 34.78 % (33/92) in the ITT, MITT, and PP populations respectively. Compared with the traditional clarithromycin resistance breakpoint of 0.25 µg/mL, a MIC threshold of 12 µg/mL predicted better eradication. Among 173 mutations on 152 sites in the 23S rRNA gene, only the 2143A > G mutation could predict eradication outcomes (p < 0.000). Conclusions: Interpretation of elevated MIC values is crucial in susceptibility testing, rather than a binary "susceptible" or "resistant" classification. The 2143A > G mutation has limited specificity in predicting eradication outcomes, necessitating further investigation into additional mutation sites associated with clarithromycin resistance.

Targeting the IRE1α-XBP1s axis confers selective vulnerability in hepatocellular carcinoma with activated Wnt signaling.

Liver-specific Ern1 knockout impairs tumor progression in mouse models of hepatocellular carcinoma (HCC). However, the mechanistic role of IRE1α in human HCC remains unclear. In this study, we show that XBP1s, the major downstream effector of IRE1α, is required for HCC cell survival both in vitro and in vivo. Mechanistically, XBP1s transactivates LEF1, a key co-factor of ß-catenin, by binding to its promoter. Moreover, XBP1s physically interacts with LEF1, forming a transcriptional complex that enhances classical Wnt signaling. Consistently, the activities of XBP1s and LEF1 are strongly correlated in human HCC and with disease prognosis. Notably, selective inhibition of XBP1 splicing using an IRE1α inhibitor significantly repressed the viability of tumor explants as well as the growth of tumor xenografts derived from patients with distinct Wnt/LEF1 activities. Finally, machine learning algorithms developed a powerful prognostic signature based on the activities of XBP1s/LEF1. In summary, our study uncovers a key mechanistic role for the IRE1α-XBP1s pathway in human HCC. Targeting this axis could provide a promising therapeutic strategy for HCC with hyperactivated Wnt/LEF1 signaling.

Adelmidrol ameliorates liver ischemia-reperfusion injury through activating Nrf2 signaling pathway.

Liver ischemia/reperfusion (I/R) injury commonly occurs after various liver surgeries. Adelmidrol, an N- palmitoylethanolamide analog, has anti-inflammatory, anti-oxidant, and anti-injury properties. To investigate whether adelmidrol could reduce liver I/R injury, we established a mouse of liver I/R injury and an AML12 cell hypoxia-reoxygenation model to perform experiments using multiple indicators. Serum ALT and AST levels, and H&E staining were used to measure liver damage; MDA content, superoxide dismutase and glutathione activities, and dihydroethidium staining were used to measure oxidative stress; mRNA expression levels of tumor necrosis factor-α, interleukin (IL)-1ß, IL-6, MCP-1, and Ly6G staining were used to measure inflammatory response; and protein expression of Bax, Bcl-2, C-caspase3, and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling staining were used to measure apoptosis. The experimental results showed that adelmidrol reduced liver I/R injury. In addition, adelmidrol pretreatment elevated AML12 cell activity and reduced I/R-and H/R-induced apoptosis, inflammatory injury, and oxidative stress. ML385, an inhibitor of nuclear factor erythroid2-related factor 2 (Nrf2), reverses liver I/R injury attenuated by adelmidrol. These results suggest that adelmidrol ameliorates liver I/R injury by activating the Nrf2 signaling pathway.

Arachidonic acid metabolism CYP450 pathway is deregulated in hepatocellular carcinoma and associated with microvascular invasion.

Arachidonic acid metabolism plays a crucial role in the development and progression of inflammatory and metabolic liver diseases. However, its role in hepatocellular carcinoma (HCC) remains unclear. In this study, we investigated the expression of key genes involved in the arachidonic acid metabolism pathway in HCC using a combination of bioinformatics, proteomics and immunohistochemistry analyses. Through a comprehensive analysis of publicly available datasets, clinical HCC tissues, and tissue microarrays, we compared the expression of hepatic arachidonic acid metabolic genes. We observed significant downregulation of cytochrome P450 (CYP450) pathway genes at both the messenger RNA and protein levels in HCC tissues compared to normal liver tissues. Furthermore, we observed a strong correlation between the deregulation of the arachidonic acid metabolism CYP450 pathway and the pathological features and prognosis of HCC. Specifically, the expression of CYP2C8/9/18/19 was significantly correlated with pathological grade (r = -.484, p < .0001), vascular invasion (r = -.402, p < .0001), aspartate transaminase (r = -.246, p = .025), gamma-glutamyl transpeptidase (r = -.252, p = .022), alkaline phosphatase (r = -.342, p = .002), alpha-fetoprotein (r = -.311, p = .004) and carbohydrate antigen 19-9 (r = -.227, p = .047). Moreover, we discovered a significant association between CYP450 pathway activity and vascular invasion in HCC. Collectively, these data indicate that arachidonic acid CYP450 metabolic pathway deregulation is implicated in HCC progression and may be a potential predictive factor for early recurrence in patients with HCC.

Air-ventilated normothermic machine perfusion alleviates hepatic injury from DCD rat through CYP1A2.

BACKGROUND: Normothermic machine perfusion (NMP) could provide protection to organs from donation after circulatory death (DCD) before transplantation, and its molecular mechanism remains unclear. Our previous study discovered that the air-ventilated NMP confers a better DCD liver recovery than oxygen-ventilated NMP. The purpose in the current study was to investigate the protective mechanism of air-ventilated NMP in a rat model of DCD liver by metabolomics, and to select biomarker to predict liver function recovery. MATERIALS AND METHODS: Peroxisome proliferator activator receptor-α (PPARα) agonist or antagonist was administered via the perfusion circuit in the air-ventilated NMP. Perfusate samples were taken for measurements of aminotransferases using standard biochemical methods, tumor necrosis factor-alpha and interleukin-6. Liver biopsies were allocated for detection of metabolomics, PPARα and cytochrome P450 1A2 (CYP1A2). RESULTS: Metabolomics analysis revealed the significant increased γ-linolenic acid and decreased adrenic acid during the air-ventilated NMP, indicating linoleic acid metabolism pathway was associated with a better DCD liver recovery; as a major enzyme involved in linolenic acid metabolism, CYP1A2 was found correlated with a less inflammation and better liver function with the air-ventilated NMP; PPARα agonist could increase CYP1A2 expression and activity, decrease inflammation response, and improve liver function with the air-ventilated NMP, while PPARα antagonist played the opposite. CONCLUSION: Air-ventilated NMP confers a better liver recovery from DCD rats through the activated linoleic acid metabolism and CYP1A2 upregulation; CYP1A2 expression and activity might function as biomarker to predict DCD liver function recovery with NMP.

Global Trends of Gastrointestinal Endoscopy Anesthesia/Sedation: A Bibliometric Study (from 2001 to 2022).

Background: Gastrointestinal (GI) endoscopy becomes more and more common now in order to diagnose and treat GI diseases, and anesthesia/sedation plays an important role. We aim to discuss the developmental trends and evaluate the research hotspots using bibliometric methods for GI endoscopy anesthesia/sedation in the past two decades. Methods: The original and review articles published from 2001 to December 2022 related to GI endoscopy anesthesia/sedation were extracted from the Web of Science database. Four different softwares (CiteSpace, VOSviewer, and Bibliometrix, Online Analysis Platform of Literature Metrology (Bibliometric)) were used for this comprehensive analysis. Results: According to our retrieval strategy, we found a total of 3154 related literatures. Original research articles were 2855, and reviews were 299. There has been a substantial increase in the research on GI endoscopy anesthesia/sedation in recent 22 years. These publications have been cited 66,418 times, with a mean of 21.04 citations per publication. The US maintained a leading position in global research, with the largest number of publications (29.94%), and China ranked second (19.92%). Keyword burst and concurrence showed that conscious sedation, colonoscopy and midazolam were the most frequently occurring keywords. Conclusion: Our research found that GI endoscopy anesthesia/sedation was in a period of rapid development and demonstrated the improvement of medical instruments and surgical options that had significantly contributed to the field of GI endoscopy anesthesia/sedation. The US dominates this field, and the selection and dosage of sedative regimens have always been the foci of disease research to improve comfort and safety, while adverse events and risks arouse attention gradually. In the past 20 years, hotspots mainly focus on upper gastrointestinal endoscopy, gastroscopy, and esophagogastroduodenoscopy. These data would provide future directions for clinicians and researchers regarding GI endoscopy anesthesia/sedation.

Mitogen activated protein kinase phosphatase 5 alleviates liver ischemia-reperfusion injury by inhibiting TAK1/JNK/p38 pathway.

Mitogen activated protein kinase phosphatase 5 (MKP5) is a member of the MKP family and has been implicated in diverse biological and pathological conditions. However, it is unknown what role MKP5 plays in liver ischemia/reperfusion (I/R) injury. In the present study, we used MKP5 global knockout (KO) and MKP5 overexpressing mice to establish a liver I/R injury model in vivo, and MKP5 knockdown or MKP5 overexpressing HepG2 cells to establish a hypoxia-reoxygenation (H/R) model in vitro. In this study we demonstrated that protein expression of MKP5 was significantly downregulated in liver tissue of mice after I/R injury, and HepG2 cells subjected to H/R injury. MKP5 KO or knockdown significantly increased liver injury, as demonstrated by elevated serum transaminases, hepatocyte necrosis, infiltrating inflammatory cells, secretion of pro-inflammatory cytokines, apoptosis, oxidative stress. Conversely, MKP5 overexpression significantly attenuated liver and cell injury. Furthermore, we showed that MKP5 exerted its protective effect by inhibiting c-Jun N-terminal kinase (JNK)/p38 activity, and its action was dependent on Transforming growth factor-ß-activated kinase 1 (TAK1) activity. According to our results, MKP5 inhibited the TAK1/JNK/p38 pathway to protect liver from I/R injury. Our study identifies a novel target for the diagnosis and treatment of liver I/R injury.

m6 A reader YTHDF3 triggers the progression of hepatocellular carcinoma through the YTHDF3/m6 A-EGFR/STAT3 axis and EMT.

Hepatocellular carcinoma (HCC) is the sixth most common cancer and the fourth leading cause of tumor-related deaths worldwide. N6-methyladenosine (m6 A) mediates RNA metabolism in tumor biology. However, the regulatory role of YTHDF3, an m6 A reader, in HCC progression and its underlying mechanisms remains unclear. Therefore, this study aims to investigate the oncogenic effect of YTHDF3 on HCC progression via the epigenetic regulation of m6 A-modified mRNAs. The expression levels of YTHDF3 in HCC tissues and matched adjacent liver tissues were detected using western blot analysis, immunohistochemistry, and quantitative real-time polymerase chain reaction. The function of YTHDF3 in HCC progression and its underlying mechanisms have been studied both in vitro and in vivo. YTHDF3 expression was significantly higher in HCC tissues than in paracancerous liver tissues. YTHDF3 was also significantly upregulated in HCC with microvascular invasion (MVI) compared to that in HCC without MVI. YTHDF3 overexpression facilitated the proliferation, invasion, and migration of HCC cells both in vitro and in vivo. However, the YTHDF3 knockdown resulted in an inverse trend. Mechanistically, YTHDF3 enhanced the translation and stability of the m6 A-modified epidermal growth factor receptor (EGFR) mRNA, which activated the downstream EGFR/signal transducer and activator of transcription 3 (STAT3) and epithelial-mesenchymal transition (EMT) oncogenic pathways. YTHDF3 enhanced the stability and translation of m6 A-modified EGFR mRNA and stimulated HCC progression via the YTHDF3/m6 A-EGFR/STAT3 and EMT pathways. These findings reveal that YTHDF3 plays a significant role in regulating HCC progression, suggesting a promising and novel target for HCC treatment.

LATS-regulated nuclear-cytoplasmic translocation of SREBP2 inhibits hepatocellular carcinoma cell migration and invasion via epithelial-mesenchymal transition.

Abnormal cholesterol synthesis plays a crucial role in the development of hepatocellular carcinoma (HCC). Sterol regulatory element-binding protein 2 (SREBP2) is involved in cholesterol synthesis by translocating to the nucleus where it stimulates the transcription of genes encoding enzymes involved in the cholesterol synthesis pathway. However, the function and regulatory mechanism of SREBP2 in HCC remain unclear. In this study, we aimed to gain a better understanding of the effects of SREBP2 and its functional mechanism in HCC. In 20 HCC patients, we demonstrated that SREBP2 was highly expressed in HCC specimens, relative to their peritumoral tissue, and that higher expression correlated positively with a poor prognosis in these patients. Moreover, higher SREBP2 levels in the nucleus enhanced the occurrence of microvascular invasion, whereas inhibition of SREBP2 nuclear translocation by fatostatin markedly suppressed the migration and invasion of HCC cells via the epithelial-mesenchymal transition (EMT) process. The effects of SREBP2 were subject to functional activity of large tumor suppressor kinase (LATS), whereas inhibition of LATS promoted nuclear translocation of SREBP2, as observed in hepatoma cells and a subset of subcutaneous tumor samples from nude mice. In conclusion, SREBP2 enhances the invasion and metastasis of HCC cells by promoting EMT, which can be strengthened by the repression of LATS. Therefore, SREBP2 may serve as a novel therapeutic target for HCC.

Charged multivesicular body protein 2B ameliorates biliary injury in the liver from donation after cardiac death rats via autophagy with air-oxygenated normothermic machine perfusion.

Normothermic machine perfusion (NMP) could provide a curative treatment to reduce biliary injury in donation after cardiac death (DCD) donor livers; however, the underlying mechanisms remain poorly understood. In a rat model, our study compared air-oxygenated NMP to hyperoxygenated NMP and found that air-oxygenated NMP improved DCD functional recovery. Here, we found that the charged multivesicular body protein 2B (CHMP2B) expression was substantially elevated in the intrahepatic biliary duct endothelium of the cold-preserved rat DCD liver after air-oxygenated NMP or in biliary endothelial cells under hypoxia/physoxia. CHMP2B knockout (CHMP2B-/-) rat livers showed increased biliary injury after air-oxygenated NMP, indicated by decreased bile production and bilirubin level, elevated biliary levels of lactate dehydrogenase and gamma-glutamyl transferase. Mechanically, we demonstrated that CHMP2B was transcriptionally regulated by Kruppel-like transcription factor 6 (KLF6) and alleviated biliary injury through decreasing autophagy. Collectively, our results suggested that air-oxygenated NMP regulates CHMP2B expression through the KLF6, which reduces biliary injury by inhibiting autophagy. Targeting the KLF6-CHMP2B autophagy axis may provide a solution to reducing biliary injury in DCD livers undergoing NMP.

The alteration of intestinal mucosal α-synuclein expression and mucosal microbiota in Parkinson's disease.

Parkinson's disease (PD) is the second most common neurodegenerative disease but still lacks a preclinical strategy to identify it. The diagnostic value of intestinal mucosal α-synuclein (αSyn) in PD has not drawn a uniform conclusion. The relationship between the alteration of intestinal mucosal αSyn expression and mucosal microbiota is unclear. Nineteen PD patients and twenty-two healthy controls were enrolled in our study from whom were collected, using gastrointestinal endoscopes, duodenal and sigmoid mucosal samples for biopsy. Multiplex immunohistochemistry was performed to detect total, phosphorylate, and oligomer α-synuclein. Next-generation 16S rRNA amplicon sequencing was applied for taxonomic analysis. The results implied that oligomer α-synuclein (OSyn) in sigmoid mucosa of PD patients was transferred from the intestinal epithelial cell membrane to the cytoplasm, acinar lumen, and stroma. Its distribution feature was significantly different between the two groups, especially the ratio of OSyn/αSyn. The microbiota composition in mucosa also differed. The relative abundances of Kiloniellales, Flavobacteriaceae, and CAG56 were lower, while those of Proteobacteria, Gammaproteobacteria, Burkholderiales, Burkholdriaceae, Oxalobacteraceae, Ralstonia, Massilla, and Lactoccus were higher in duodenal mucosa of PD patients. The relative abundances of Thermoactinomycetales and Thermoactinomycetaceae were lower, while those of Prevotellaceae and Bifidobacterium longum were higher in patients' sigmoid mucosa. Further, the OSyn/αSyn level was positively correlated with the relative abundances of Proteobacteria, Gammaproteobacteria, Burkholderiales, Pseudomonadales, Burkholderiaceae, and Ralstonia in the duodenal mucosa, while it was negatively correlated with the Chao1 index and observed operational taxonomic units of microbiota in sigmoid mucosa. The intestinal mucosal microbiota composition of PD patients altered with the relative abundances of proinflammatory bacteria in the duodenal mucosa increased. The ratio of the OSyn/αSyn level in the sigmoid mucosa indicated a potential diagnostic value for PD, which also correlated with mucosal microbiota diversity and composition. KEY POINTS: • The distribution of OSyn in sigmoid mucosa differed between PD patients and healthy controls. • Significant alterations in the microbiome were found in PD patients' gut mucosa. • OSyn/αSyn level in sigmoid mucosa indicated a potential diagnostic value for PD.

FXR1 promotes proliferation, invasion and migration of hepatocellular carcinoma in vitro and in vivo.

Hepatocellular carcinoma (HCC) is a common malignancy that is associated with a poor prognosis. The extensively studied TGF-ß pathway is mediated by SMAD proteins. FXR1, a protein-coding gene belonging to the fragile X-related (FXR) family, is involved in the TGF-ß pathway. Previous studies have shown that FXR1 promotes the proliferation, invasion, and migration of colorectal cancer cells. The aim of the present study was to explore the effects of FXR1 on HCC via the TGF-ß/SMAD signaling pathway. Immunohistochemical analysis was used to detect the expression of FXR1 in HCC and normal tissues. Western blotting was used to detect protein expression levels in the HCC cell lines, cell migration and invasion were assessed using Transwell assays, and cell proliferation was assessed using a colony formation assay. The ability of the liver cancer cells to grow in vivo was investigated using a nude mouse tumor-bearing model. The results showed that FXR1 expression was upregulated in HCC tissues compared with normal tissues. Knockdown of FXR1 resulted in reduced expression of SMAD2/3 and EMT-related proteins in HCC cells. In addition, FXR1 knockdown inhibited the proliferation, migration, and invasion of HCC cells. FXR1 knockdown also reversed the promoting effect of TGF-ß on the invasive ability of HCC cells. Knockdown of SMAD2/3 reversed the increase in HCC cell invasion induced by FXR1 overexpression. Finally, upregulated FXR1 expression was associated with a poorer prognosis in patients with HCC. In conclusion, FXR1 promoted HCC proliferation, migration, and invasion through the regulation of SMAD2/3.

Evaluation of commonly used cardiovascular drugs in inhibiting vonoprazan metabolism in vitro and in vivo.

As a novel acid-suppressing drug, vonoprazan shows the potential to replace traditional proton-pump inhibitors. With its widespread use, some adverse effects that require further study have emerged due to drug-drug interactions. Our study is the first experiment that evaluated the drug-drug interactions of eleven common cardiovascular drugs that inhibit vonoprazan metabolism in vitro and in vivo. Rat liver microsome incubation and molecular simulation docking were applied to explore the inhibition mechanism. Amlodipine and nifedipine showed inhibitory effects on vonoprazan metabolism in both rat and human liver microsomes in the first evaluation part in vitro. The inhibition mechanism analysis results demonstrated that amlodipine and nifedipine might inhibit the metabolism of vonoprazan by a mixed type of competitive and non-competitive inhibition. However, the pharmacokinetic data of the vonoprazan prototype revealed that amlodipine affected vonoprazan in vivo while nifedipine did not. Thus, more attention should be paid when amlodipine is prescribed with vonoprazan. Furthermore, the changes in its carboxylic acid metabolites MI hinted at a complex situation. Molecular simulation suggested the CYP2B6 enzyme may contribute more to this than CYP3A4, and further inhibitory experiments preliminarily verified this speculation. In conclusion, the use of vonoprazan with cardiovascular drugs, especially amlodipine, should receive particular attention in clinical prescriptions.

PD-1/PD-L1 inhibition promotes hepatic regeneration in small-for-size liver following extended hepatectomy.

BACKGROUND: Small-for-size syndrome following liver surgery is characterized by compromised liver regeneration. Liver macrophages play key roles in initiating liver regeneration, and modulation of the immune microenvironment through macrophages may accelerate liver regeneration. In our current study, we aimed to explore the involvement of innate immunity after extended hepatectomy in rats and humans, and to test the effect of immunity modulation on small-for-size liver regeneration in rats. METHODS: Serum programmed cell death protein ligand 1 (PD-L1) was measured after major hepatectomy and minor hepatectomy in humans and rats. Liver regeneration in rats was assessed using liver-to-body weight ratio and kinetic growth rate, antigen Ki67 and proliferating cell nuclear antigen (PCNA), and macrophage polarization was assessed by inducible nitric oxide synthase (iNOS), cluster of differentiation protein 163 (CD163) expression by immunohistochemistry (IHC) and iNOS/CD163 ratio. Rat hepatocyte BRL or human hepatocyte LO2 were co-cultured with rat bone marrow-derived macrophages or human macrophages THP-1. BMS-1 or Nivolumab were used to block programmed cell death protein 1 (PD-1)/PD-L1 in vitro and in vivo. RESULTS: PD-L1 expressions were significantly higher following major hepatectomy compared to minor resection in both humans and rats; compromised liver regeneration after extended hepatectomy in rats was associated with PD-L1 upregulation and M2 macrophage polarization. M1 macrophages increased proliferation of hepatocytes through interleukin-6 (IL-6), and M2 macrophages decreased hepatocyte proliferation; blocking PD-1/PD-L1 reversed the effect of M2 macrophages on the survival of hepatocytes in vitro and promoted liver growth in rats through M1 macrophage polarization. CONCLUSION: Compromised hepatic regeneration following extended hepatectomy is characterized by M2 macrophage polarization and upregulated PD-L1 expression. Blocking PD-1/PD-L1 may enhance small-for-size liver regeneration by inducing M1 macrophage polarization.

Necroptosis throws novel insights on patient classification and treatment strategies for hepatocellular carcinoma.

Introduction: Necroptosis is a novel pattern of immunogenic cell death and has triggered an emerging wave in antitumor therapy. More evidence has suggested the potential associations between necroptosis and intra-tumoral heterogeneity. Currently, the underlying role of necroptosis remains elusive in hepatocellular carcinoma (HCC) at antitumor immunity and inter-tumoral heterogeneity. Methods: This study enrolled a total of 728 HCC patients and 139 immunotherapy patients from eight public datasets. The consensus clustering approach was employed to depict tumor heterogeneity of cancer necroptosis. Subsequently, our study further decoded the heterogeneous clinical outcomes, genomic landscape, biological behaviors, and immune characteristics in necroptosis subtypes. For each patient, providing curative clinical recommendations and developing potential therapeutic drugs were used to promote precise medicine. Results: With the use of the weighted gene coexpression network analysis (WGCNA) algorithm, necroptosis-associated long non-coding RNAs (lncRNAs) (NALRs) were identified in HCC. Based on the NALR expression, two heterogeneous subtypes were decoded with distinct clinical outcomes. Compared to patients in C1, patients in C2 harbored superior pathological stage and presented more unfavorable overall survival and recurrence-free survival. Then, the robustness and reproducibility of necroptosis subtypes were further validated via the nearest template prediction (NTP) approach and classical immune phenotypes. Through comprehensive explorations, C1 was characterized by enriched immune-inflammatory and abundant immune infiltration, while C2 possessed elevated proliferative and metabolic activities and highly genomic instability. Moreover, our results indicated that C1 was more prone to obtain desirable benefits from immunotherapy. For patients in C2, numerous underlying therapeutic agents were developed, which might produce significant efficacy. Conclusion: This study identified two necroptosis subtypes with distinct characteristics, decoding the tumor heterogeneity. For an individualized patient, our work tailored corresponding treatment strategies to improve clinical management.

CD8+ T cell trajectory subtypes decode tumor heterogeneity and provide treatment recommendations for hepatocellular carcinoma.

Introduction: Mounting evidence has revealed that the interactions and dynamic alterations among immune cells are critical in shaping the tumor microenvironment and ultimately map onto heterogeneous clinical outcomes. Currently, the underlying clinical significance of immune cell evolutions remains largely unexplored in hepatocellular carcinoma (HCC). Methods: A total of 3,817 immune cells and 1,750 HCC patients of 15 independent public datasets were retrieved. The Seurat and Monocle algorithms were used to depict T cell evolution, and nonnegative matrix factorization (NMF) was further applied to identify the molecular classification. Subsequently, the prognosis, biological characteristics, genomic variations, and immune landscape among distinct clusters were decoded. The clinical efficacy of multiple treatment approaches was further investigated. Results: According to trajectory gene expression, three heterogeneous clusters with different clinical outcomes were identified. C2, with a more advanced pathological stage, presented the most dismal prognosis relative to C1 and C3. Eight independent external cohorts validated the robustness and reproducibility of the three clusters. Further explorations elucidated C1 to be characterized as lipid metabolic HCC, and C2 was referred to as cell-proliferative HCC, whereas C3 was defined as immune inflammatory HCC. Moreover, C2 also displayed the most conspicuous genomic instability, and C3 was deemed as "immune-hot", having abundant immune cells and an elevated expression of immune checkpoints. The assessments of therapeutic intervention suggested that patients in C1 were suitable for transcatheter arterial chemoembolization treatment, and patients in C2 were sensitive to tyrosine kinase inhibitors, while patients in C3 were more responsive to immunotherapy. We also identified numerous underlying therapeutic agents, which might be conducive to clinical transformation in the future. Conclusions: Our study developed three clusters with distinct characteristics based on immune cell evolutions. For specifically stratified patients, we proposed individualized treatment strategies to improve the clinical outcomes and facilitate the clinical management.

Significance of Liver Zonation in Hepatocellular Carcinoma.

Liver zonation is fundamental to normal liver function, and numerous studies have investigated the microstructure of normal liver lobules. However, only a few studies have explored the zonation signature in hepatocellular carcinoma (HCC). In this study, we investigated the significance of liver zonation in HCC with the help of single-cell RNA sequencing (scRNA-seq) and multicolor immunofluorescence staining. Liver zonation-related genes were extracted from the literature, and a three-gene model was established for HCC prognosis. The model reliability was validated using bulk RNA and single-cell RNA-level data, and the underlying biological mechanism was revealed by a functional enrichment analysis. The results showed that the signaling pathways of high-risk groups were similar to those of perivenous zones in the normal liver, indicating the possible regulating role of hypoxia in HCC zonation. Furthermore, the co-staining results showed that the low-grade tumors lost their zonation features whereas the high-grade tumors lost the expression of zonation-related genes, which supported the results obtained from the sequencing data.

Effects of Simvastatin on the Metabolism of Vonoprazan in Rats Both in vitro and in vivo.

Purpose: To study the potential drug-drug interactions between simvastatin and vonoprazan and to provide the scientific basis for rational use of them in clinical practice. Methods: An incubation system was established with rat liver microsomes, and the main metabolite of vonoprazan M-I was detected by UPLC-MS/MS. The IC50 value of simvastatin was then calculated and its inhibitory mechanism against vonoprazan was also analyzed. Twelve SD rats were randomly divided into 2 groups, then they were given simvastatin or saline for 2 weeks continuously. On the day of the experiment, both groups were intragastrically administered with vonoprazan once, followed by the collection of blood at different time points. Then the plasma concentration of vonoprazan and M-I in rats were detected by UPLC-MS/MS. Results: In vitro experiments revealed that simvastatin could inhibit the metabolism of vonoprazan, and its inhibition type belonged to the mixed non-competitive and competitive inhibition model. In vivo experiments in rats demonstrated that the area under concentration time curve (AUC) of vonoprazan was decreased but the clearance (CLz/F) of it was increased in the simvastatin administrated group, as compared to those of the control group. However, M-I in simvastatin treated group exhibited the higher AUC and lower CLz/F values compared to those in the control group. These data indicated that multiple doses of simvastatin administration could reduce the plasma concentration of vonoprazan and accelerate its metabolic rate in rats. Conclusion: Simvastatin could inhibit the metabolism of vonoprazan in vitro but multiple doses of simvastatin exhibited the opposite effect In vivo. Altogether, our data indicated that an interaction existed between simvastatin and vonoprazan and additional cares might be taken when they were co-administrated in clinic.

Identification of Context-Specific Fitness Genes Associated With Metabolic Rearrangements for Prognosis and Potential Treatment Targets for Liver Cancer.

Liver cancer is the most frequent fatal malignancy. Furthermore, there is a lack of effective therapeutics for this cancer type. To construct a prognostic model for potential beneficiary screens and identify novel treatment targets, we used an adaptive daisy model (ADaM) to identify context-specific fitness genes from the CRISPR-Cas9 screens database, DepMap. Functional analysis and prognostic significance were assessed using data from TCGA and ICGC cohorts, while drug sensitivity analysis was performed using data from the Liver Cancer Model Repository (LIMORE). Finally, a 25-gene prognostic model was established. Patients were then divided into high- and low-risk groups; the high-risk group had a higher stemness index and shorter overall survival time than the low-risk group. The C-index, time-dependent ROC curves, and multivariate Cox regression analysis confirmed the excellent prognostic ability of this model. Functional enrichment analysis revealed the importance of metabolic rearrangements and serine/threonine kinase activity, which could be targeted by trametinib and is the key pathway in regulating liver cancer cell viability. In conclusion, the present study provides a prognostic model for patients with liver cancer and might help in the exploration of novel therapeutic targets to ultimately improve patient outcomes.

Cytochrome P450 2E1 predicts liver functional recovery from donation after circulatory death using air-ventilated normothermic machine perfusion.

The optimal oxygen concentration is unclear for normothermic machine perfusion (NMP) of livers from donation after circulatory death (DCD). Our purposes were to investigate the effect of air-ventilated NMP on the DCD liver, analyze the underlying mechanism and select the targets to predict liver functional recovery with NMP. NMP was performed using the NMP system with either air ventilation or oxygen ventilation for 2 h in the rat liver following warm ischemia and cold-storage preservation. Proteomics and metabolomics were used to reveal the significant molecular networks. The bioinformation analysis was validated by administering peroxisome proliferator activator receptor-γ (PPARγ) antagonist and agonist via perfusion circuit in the air-ventilated NMP. Results showed that air-ventilated NMP conferred a better functional recovery and a less inflammatory response in the rat DCD liver; integrated proteomics and metabolomics analysis indicated that intrahepatic docosapentaenoic acid downregulation and upregulation of cytochrome P450 2E1 (CYP2E1) expression and activity were associated with DCD liver functional recovery with air-ventilated NMP; PPARγ antagonist worsened liver function under air-oxygenated NMP whereas PPARγ agonist played the opposite role. In conclusion, air-ventilated NMP confers a better liver function from DCD rats through the DAP-PPARγ-CYP2E1 axis; CYP2E1 activity provides a biomarker of liver functional recovery from DCD.