Integrating Network Pharmacology and In Vitro Experiments for Assessing the Anti-Tumor Effects of Phyllanthus Urinaria L Anti-neoplastic Decoction in Hepatocellular Carcinoma.

Background: In China, traditional Chinese medicine (TCM) is an important part of the comprehensive treatment of hepatocellular carcinoma (HCC), and Chinese herb formulas with the effect of "yiqi jianpi jiedu huayu" (replenishing qi, strengthening spleen, and removing toxicity and blood stasis) are the common and efficient treatments for HCC. However, the mechanism of these formulas in treating HCC remain unclear. Objective: In this paper, our goal is to explore the potential mechanism of Phyllanthus urinaria L anti-neoplastic decoction (PAD), the representative formula of "yiqi jianpi jiedu huayu", in treating HCC. Design: The research team performed the network pharmacology and in vitro experiment (preparation of PAD aqueous extract, cell cultures and MTT assay, cell apoptosis assay, wound healing assay, transwell assays, western blot). Setting: The study took place in the Department of Hepatology, the Fourth Clinical Medical College of Guangzhou University of Chinese Medicine (Shenzhen Traditional Chinese Medicine Hospital), China. Outcome Measures: The active components and targets of PAD and HCC targets were screened by five Chinese herbs and two disease databases respectively. The network pharmacology was utilized to construct the relationship network between PAD and HCC, and the mechanism was predicted by pathway enrichment analysis. The experiment was performed to verify the intervention effect of PAD on HCC and phosphoinositide 3-kinase/protein kinase B (PI3K/Akt) pathway. Results: The relationship network between PAD and HCC suggested that PAD mainly regulated the potential therapeutic targets of HCC by key active components such as quercetin, luteolin, calycosin, wogonin, and pinocembrin. Pathway analysis demonstrated PAD could play an anti-HCC effect via multiple pathways (e.g., PI3K/Akt). Results of the experiment showed that PAD could effectively inhibit the proliferation and migration of HCC cells, and promote HCC cells apoptosis in a concentration-dependent behavior. Additionally, PAD could decrease the protein expression of phosphorylated PI3K/Akt. Conclusion: PAD mainly exerts an anti-HCC effect through multiple active components represented by quercetin and multiple pathways represented by the PI3K/Akt pathway. This study provided an experimental basis for the clinical application of PAD.

An herbal formulation "Shugan Xiaozhi decoction" ameliorates methionine/choline deficiency-induced nonalcoholic steatohepatitis through regulating inflammation and apoptosis-related pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Shugan Xiaozhi (SGXZ) decoction is a traditional Chinese medicine used for treating nonalcoholic steatohepatitis (NASH). It has been used clinically for over 20 years and proved to be effective; however, the molecular mechanism underlying the effects of SGXZ decoction remains unclear. AIM OF THE STUDY: We analyzed the chemical components, core targets, and molecular mechanisms of SGXZ decoction to improve NASH through network pharmacology and in vivo experiments. MATERIALS AND METHODS: The chemical components, core targets, and related signaling pathways of SGXZ decoction intervention in NASH were predicted using network pharmacology. Molecular docking was performed to verify chemical components and their core targets. The results were validated in the NASH model treated with SGXZ decoction. Mouse liver function was assessed by measuring ALT and AST levels. TC and TG levels were determined to evaluate lipid metabolism, and lipid deposition was assessed via oil red O staining. Mouse liver damage was determined via microscopy following hematoxylin and eosin staining. Liver fibrosis was assessed via Masson staining. Western blot (WB) and immunohistochemical (IHC) analyses were performed to detect inflammation and the expression of apoptosis-related proteins, including IL-1ß, IL-6, IL-18, TNF-α, MCP1, p53, FAS, Caspase-8, Caspase-3, Caspase-9, Bax, Bid, Cytochrome c, Bcl-2, and Bcl-XL. In addition, WB and IHC were used to assess protein expression associated with the TLR4/MyD88/NF-κB pathway. RESULTS: Quercetin, luteolin, kaempferol, naringenin, and nobiletin in SGXZ decoction were effective chemical components in improving NASH, and TNF-α, IL-6, and IL-1ß were the major core targets. Molecular docking indicated that these chemical components and major core targets might interact. KEGG pathway analysis showed that the pathways affected by SGXZ decoction, primarily including apoptosis and TLR4/NF-κB signaling pathways, interfere with NASH. In vivo experiments indicated that SGXZ decoction considerably ameliorated liver damage, fibrosis, and lipid metabolism disorder in MCD-induced NASH mouse models. In addition, WB and IHC verified the underlying molecular mechanisms of SGXZ decoction as predicted via network pharmacology. SGXZ decoction inhibited the activation of apoptosis-related pathways in MCD-induced NASH mice. Moreover, SGXZ decoction suppressed the activation of TLR4/MyD88/NF-κB pathway in MCD-induced NASH mice. CONCLUSION: SGXZ decoction can treat NASH through multiple targets and pathways. These findings provide new insights into the effective treatment of NASH using SGXZ decoction.

From gut to liver: unveiling the differences of intestinal microbiota in NAFL and NASH patients.

Non-alcoholic fatty liver disease (NAFLD) is increasingly recognized for its global prevalence and potential progression to more severe liver diseases such as non-alcoholic steatohepatitis (NASH). The gut microbiota plays a pivotal role in the pathogenesis of NAFLD, yet the detailed characteristics and ecological alterations of gut microbial communities during the progression from non-alcoholic fatty liver (NAFL) to NASH remain poorly understood. Methods: In this study, we conducted a comparative analysis of gut microbiota composition in individuals with NAFL and NASH to elucidate differences and characteristics. We utilized 16S rRNA sequencing to compare the intestinal gut microbiota among a healthy control group (65 cases), NAFL group (64 cases), and NASH group (53 cases). Random forest machine learning and database validation methods were employed to analyze the data. Results: Our findings indicate a significant decrease in the diversity of intestinal flora during the progression of NAFLD (p < 0.05). At the phylum level, high abundances of Bacteroidetes and Fusobacteria were observed in both NAFL and NASH patients, whereas Firmicutes were less abundant. At the genus level, a significant decrease in Prevotella expression was seen in the NAFL group (AUC 0.738), whereas an increase in the combination of Megamonas and Fusobacterium was noted in the NASH group (AUC 0.769). Furthermore, KEGG pathway analysis highlighted significant disturbances in various types of glucose metabolism pathways in the NASH group compared to the NAFL group, as well as notably compromised flavonoid and flavonol biosynthesis functions. The study uncovers distinct microbiota characteristics and microecological changes within the gut during the transition from NAFL to NASH, providing insights that could facilitate the discovery of novel biomarkers and therapeutic targets for NAFLD.

Polyphyllin I attenuates the invasion and metastasis via downregulating GRP78 in drug-resistant hepatocellular carcinoma cells.

Drug resistance to chemotherapy agents presents a major obstacle to the effective treatment of hepatocellular carcinoma (HCC), a common type of liver cancer. Increasing evidence indicates a link between drug resistance and the recurrence of HCC. Polyphyllin I (PPI), a promising pharmaceutical candidate, has shown potential therapeutic advantages in the treatment of sorafenib-resistant hepatocellular carcinoma (SR-HCC cells). In this study, we sought to investigate the mechanism underlying the inhibitory effect of PPI on the invasion and metastasis of SR-HCC cells. Our in vitro studies included scratch wound-healing migration assays and transwell assays to examine PPI's effect on HCC cell migration and invasion. Flow cytometry was employed to analyze the accumulation or efflux of chemotherapy drugs. The results of these experiments demonstrated that PPI increased the susceptibility of HCC to sorafenib while inhibiting SR-HCC cell growth, migration, and invasion. Molecular docking analysis revealed that PPI exhibited a higher binding affinity with GRP78. Western blot analysis and immunofluorescence experiments showed that PPI reduced the expression of GRP78, E-cadherin, N-cadherin, Vimentin, and ABCG2 in SR-HCC cells. Interference with and overproduction of GRP78 in vitro impacted the proliferation, migration, invasion, and metastasis of HCC cells. Further examination revealed that PPI hindered the expression of GRP78 protein, resulting in a suppressive effect on SR-HCC cell migration and invasion. Histological examination of tumor tissue substantiated that administering PPI via gavage to HepG2/S xenograft nude mice inhibited tumor growth and significantly reduced tumor size, as evidenced by xenograft experiments involving nude mice. Hematoxylin and eosin (HE) staining of tumor tissue specimens, along with immunohistochemistry (IHC), were conducted to evaluate the expression levels of Ki67, GRP78, N-cadherin, Vimentin, and ABCG2. The results indicated that PPI administration decreased the levels of proteins associated with metastasis and markers of drug resistance in tumor tissues, impeding tumor growth and spread. Overall, our findings demonstrated that PPI effectively suppressed the viability, proliferation, invasion, and metastasis of SR-HCC cells both in vitro and in vivo by modulating GRP78 activity. These findings provide new insights into the mechanism of PPI inhibition of SR-HCC cell invasion and metastasis, highlighting PPI as a potential treatment option for sorafenib-resistant HCC.

A classical herbal formula alleviates high-fat diet induced nonalcoholic steatohepatitis (NASH) via targeting mitophagy to rehabilitate dysfunctional mitochondria, validated by UPLC-HRMS identification combined with in vivo experiment.

BACKGROUND: Nonalcoholic steatohepatitis (NASH) has caused a significant burden on public health care systems, the economy and society. However, there has still been no officially approved pharmacotherapy for NASH. It has been suggested that oxidative stress and mitochondrial dysfunction play vital roles in NASH pathological progression. Shugan Xiaozhi (SG) formula, as a kind of classical herbal formula, was shown to attenuate NASH. PURPOSE: This study aimed to explore the potential mechanisms of SG formula treating NASH. STUDY DESIGN AND METHODS: Ultra-high-performance liquid chromatography-high resolution mass spectrometry combined with bioinformatics analysis was applied to explore the therapeutic targets and main components of SG formula. Moreover, in vivo NASH model was utilized to confirmed the therapeutic effects of SG formula. Molecular docking analysis and further validation experiments were conducted to verify the results of bioinformatics analysis. RESULTS: The in vivo experiments confirmed SG formula significantly attenuated hepatic pathological progression and relieved oxidative stress in high-fat diet (HFD) induced - NASH model. Ultra-high-performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS) combined with bioinformatics analysis expounded the components of SG formula and revealed the mitochondrial regulation mechanism of SG formula treating NASH. Further in vivo experiments validated that SG formula could alleviate oxidative stress by rehabilitating the structure and function of mitochondria, which was strongly related to regulating mitophagy. CONCLUSION: In summary, this study demonstrated that SG formula, which could attenuate NASH by regulating mitochondria and might be a potential pharmacotherapy for NASH.

MAT1A Suppression by the CTBP1/HDAC1/HDAC2 Transcriptional Complex Induces Immune Escape and Reduces Ferroptosis in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) remains a significant health burden globally due to its high prevalence and morbidity. C-terminal-binding protein 1 (CTBP1) is a transcriptional corepressor that modulates gene transcription by interacting with transcription factors or chromatin-modifying enzymes. High CTBP1 expression has been associated with the progression of various human cancers. In this study, bioinformatics analysis suggested the existence of a CTBP1/histone deacetylase 1 (HDAC1)/HDAC2 transcriptional complex that regulates the expression of methionine adenosyltransferase 1A (MAT1A), whose loss has been associated with ferroptosis suppression and HCC development. Thus, this study aims to investigate the interactions between the CTBP1/HDAC1/HDAC2 complex and MAT1A and their roles in HCC progression. First, high expression of CTBP1 was observed in HCC tissues and cells, where it promoted HCC cell proliferation and mobility while inhibiting cell apoptosis. CTBP1 interacted with HDAC1 and HDAC2 to suppress the MAT1A transcription, and silencing of either HDAC1 or HDAC2 or overexpression of MAT1A led to the inhibition of cancer cell malignancy. In addition, MAT1A overexpression resulted in increased S-adenosylmethionine levels, which promoted ferroptosis of HCC cells directly or indirectly by increasing CD8+ T-cell cytotoxicity and interferon-γ production. In vivo, MAT1A overexpression suppressed growth of CTBP1-induced xenograft tumors in mice while enhancing immune activity and inducing ferroptosis. However, treatment with ferrostatin-1, a ferroptosis inhibitor, blocked the tumor-suppressive effects of MAT1A. Collectively, this study reveals that the CTBP1/HDAC1/HDAC2 complex-induced MAT1A suppression is liked to immune escape and reduced ferroptosis of HCC cells.

Chinese herbal formula ruangan granule enhances the efficacy of entecavir to reverse advanced liver fibrosis/early cirrhosis in patients with chronic HBV infection: A multicenter, randomized clinical trial.

BACKGROUND: Nucleotide analogs treatment can reverse liver fibrosis in chronic hepatitis B (CHB). However, it has limited effect on fibrosis resolution in patients with CHB, particularly in preventing progression to hepatocellular carcinoma (HCC). Ruangan granule (RG), a Chinese herbal formula, has proven to produce a therapeutic effect against liver fibrosis in animal experiment. Thus, we aimed to evaluate the effect of our Chinese herbal formula (RG) combined with entecavir (ETV) to reverse advanced liver fibrosis/early cirrhosis from CHB. METHODS: A total of 240 CHB patients with histologically confirmed advanced liver fibrosis/early cirrhosis from 12 centers were randomly and blindly allocated to consume either ETV (0.5 mg/day) plus RG (2 times/day) or control (ETV) for 48 weeks (wk) treatment. Changes in histopathology, serology and imageology were observed. Liver fibrosis reversion, defined as a reduction in the Knodell HAI score by ≥ 2 points and Ishak score by ≥ 1 grade, was assessed. RESULTS: The rate of fibrosis regression and inflammation remission after 48 wk of treatment in histopathology was significantly higher in the ETV + RG group (38.73% vs. 23.94%, P = 0.031). The ultrasonic semiquantitative scores decreased by ≥ 2 points and were 41 (28.87%) and 15 (21.13%) in the ETV+RG and ETV groups, respectively (P = 0.026). The ETV+RG group had a significantly lower Fibrosis-4 score (FIB-4) index (P = 0.028). There was a significant difference between the ETV+RG and ETV groups in the liver function normalization rate (P < 0.01). Moreover, ETV plus RG combination treatment further reduced the risk of HCC in median 55-month follow-up (P < 0.01). CONCLUSIONS: This study illustrates that the Chinese herbal formula RG with ETV can improve advanced liver fibrosis/early cirrhosis regression in patients with CHB, further reducing the risk of HCC.

Integrating network analysis and experimental validation to reveal the mitophagy-associated mechanism of Yiqi Huoxue (YQHX) prescription in the treatment of myocardial ischemia/reperfusion injury.

Myocardial ischemia/reperfusion (I/R) injury is the main cause of increasing postischemic heart failure and currently there is no definite treatment for myocardial I/R injury. It has been suggested that oxidative stress-induced mitochondrial dysfunction plays an important role in the pathological development of myocardial I/R. In this study, Yiqi Huoxue (YQHX) prescription, as a kind of Chinese herbal formula, was developed and shown to alleviate I/R injury. Network analysis combined with ultrahigh-performance liquid chromatography-high resolution mass spectrometry expounded the active components of YQHX and revealed the mitophagy-regulation mechanism of YQHX treating I/R injury. In vivo experiments confirmed YQHX significantly alleviated I/R myocardial injury and relieved oxidative stress. In vitro experiments validated that YQHX could relieve hypoxia/reoxygenation injury and attenuate oxidative stress via improving the structure and function of mitochondria, which was strongly related to regulating mitophagy. In summary, this study demonstrated that YQHX, which could alleviate I/R injury via targeting mitophagy, might be a potential therapeutic strategy for myocardial I/R injury.

Anticancer Effect of Polyphyllin I in Suppressing Stem Cell-Like Properties of Hepatocellular Carcinoma via the AKT/GSK-3ß/ß-Catenin Signaling Pathway.

Polyphyllin I (PPI), also called Chong Lou saponin I, is a steroidal saponin isolated from the rhizome of Paris polyphylla. PPI has been demonstrated to have strong anticancer activity. However, its effect on the stemness of liver cancer stem cells (LCSCs) is not completely understood. Herein, we aimed to investigate the effect of PPI on the stem cell-like features of LCSCs and hepatocellular carcinoma (HCC). LCSCs were enriched in a serum-free medium and treated with PPI, sorafenib (Sora), or PPI and Sora. Several endpoints, including spheroid formation and differentiation, cell proliferation, surface markers of LCSCs, PPI binding targets, and stemness-associated protein expression, were evaluated. Immunofluorescence staining, quantitative real-time polymerase chain reaction, siRNA transfection, and coimmunoprecipitation ubiquitination assays were conducted for in-depth mechanistic studies. Evaluation of in vivo antitumor efficacy demonstrated that PPI effectively inhibited the proliferation of liver cancer cells and the self-renewal and differentiation of LCSCs. Flow cytometry indicated that PPI suppressed the expression of the stem cell surface markers EpCAM and CD13. Molecular docking showed a high affinity between PPI and proteins of the Wnt/ß-catenin signaling pathway, including AKT, GSK-3ß, and ß-catenin, with the binding energies of -5.51, -5.32, and -5.40 kcal/mol, respectively, which suggested that PPI might regulate the Wnt/ß-catenin signaling pathway to affect the stem cell-like properties of HCC. Further ex vivo experiments implied that PPI activated the AKT/GSK-3ß-mediated ubiquitin proteasomal degradation of ß-catenin and subsequently attenuated the prooncogenic effect of LCSCs. Finally, the anticancer property of PPI was confirmed in vivo. It was found that PPI inhibited the tumor growth in an HCC cell line xenograft model. Taken together, molecular docking analysis and experimental data highlighted the novel function of PPI in suppressing the stem cell-like characteristics of LCSCs via the AKT/GSK-3ß/ß-catenin signaling pathway.

Inhibition of inflammatory liver injury by the HMGB1-A box through HMGB1/TLR-4/NF-κB signaling in an acute liver failure mouse model.

We aimed to investigate the preventive effect of high mobility group box 1 (HMGB1)-A box and the mechanism by which it alleviates inflammatory injury in acute liver failure (ALF) by inhibiting the extracellular release of HMGB1. BALB/c mice were intraperitoneally (i.p.) administered LPS/D-GalN to establish an ALF mouse model. HMGB1-A box was administered (i.p.) 1 h before establishing the ALF mouse model. The levels of extracellularly released HMGB1, TLR-4/NF-κB signaling molecules, the proinflammatory cytokines TNF-α, IL-1ß, and IL-6 and COX-2 were measured in the liver tissue and/or serum by Immunohistochemistry, Western blotting and Enzyme-linked immunosorbent assay (ELISA). The levels of extracellularly released HMGB1, TLR-4/NF-κB signaling molecules and proinflammatory cytokines were measured in Huh7 cells as well as LPS- and/or HMGB1-A box treatment by confocal microscopy, Western blotting and ELISA. In the ALF mouse model, the levels of HMGB1 were significantly increased both in the liver and serum, TLR-4/NF-κB signaling molecules and proinflammatory cytokines also was upregulated. Notably, HMGB1-A box could reverse these changes. HMGB1-A box could also cause these changes in LPS-induced Huh7 cells. HMGB1-A box played a protective role by inhibiting inflammatory liver injury via the regulation of HMGB1/TLR-4/NF-κB signaling in the LPS/D-GaIN-induced ALF mouse model, which may be related to inhibiting the extracellular release of HMGB1.

Intervention of Shugan Xiaozhi Decoction on Nonalcoholic Fatty Liver Disease via Mediating Gut-Liver Axis.

Nonalcoholic fatty liver disease (NAFLD) is a chronic liver disease with an increasing incidence rate but few therapies. Shugan Xiaozhi decoction (SX) has demonstrated beneficial effects in treating NAFLD with an unclear mechanism. This study was aimed at investigating the therapeutic mechanism of SX on high-fat diet-induced NAFLD rats via the gut-liver axis. Hepatic steatosis and integrity of intestinal mucosa in NAFLD rats were assessed by histopathological staining. The level of lipid and inflammation were estimated by enzyme-linked immunosorbent assay. Western Blotting was used to detect apolipoprotein (apo) B48 expression. 16S rRNA analysis was used to measure the changes of gut microbial composition after SX treatment. The expressions of zona occludens 1 protein (ZO-1), occludin, and secretory immunoglobulin A (sIgA) in the colon were detected by immunostaining to investigate the intestinal barrier function. Our study found that SX reduced hepatic steatosis, the levels of alanine aminotransferase, aspartate aminotransferase, total cholesterol, and triglyceride and apoB48 expression but increased peroxisome proliferator activated receptor α (PPARα) level. Moreover, SX altered the diversity of gut microbiota, upregulating the relative abundance of f_Prevotellaceae, while downregulating f_Bacteroidales_ S24-7, f_Lachnospiraceae, f_Ruminococcaceae, f_Erysipelotrichaceae, and f_Desulfovibrionaceae. By increasing the expression of ZO-1 and occludin and decreasing the level of proinflammatory factors, including sIgA, lipopolysaccharide, tumor necrosis factor-α, interleukin-1ß, monocyte chemotactic protein-1, and transforming growth factor-ß1, SX improved intestinal mucosal integrity and barrier function. Our study illustrated that the gut-liver axis was a potential way for SX to ameliorate NAFLD, that is, by regulating the expression of PPARα, apoB48, and modulating gut microbiota to protect the intestinal barrier function, and thus alleviate lipid deposition and inflammatory response in the liver.

Effects of Xinkeshu tablets on coronary heart disease patients combined with anxiety and depression symptoms after percutaneous coronary intervention: A meta-analysis.

BACKGROUND: Xinkeshu tablets (XKS), a well-known Chinese patent drug, have been administered to coronary heart disease (CHD) patients with anxiety and depression after percutaneous coronary intervention (PCI). PURPOSE: This meta-analysis aimed to systematically evaluate the clinical effects of XKS for treating CHD patients with anxiety and depression after PCI. METHODS: Randomized controlled trials (RCTs) about XKS alone or combined with conventional drugs for the treatment of CHD patients with anxiety and depression after PCI were retrieved from 7 databases (MEDLINE, EMBASE, the Cochrane Library, China National Knowledge Infrastructure (CNKI), Chinese Scientific Journals Database (VIP) Database, Chinese Biomedical Database (CBM) and Wanfang Database) through November 2021. First, the studies were reviewed and screened by two independent assessors according to the eligibility criteria. Second, the methodological quality of the eligible studies was evaluated based on the Cochrane Collaboration's tool for assessing the risk of bias. Subsequently, meta-analysis was performed by using RevMan 5.4 software, and publication bias was evaluated by Stata 12.0 software. Finally, the Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach was applied to rate the quality of the evidence. RESULTS: In total, 11 clinical RCTs involving 1000 patients were included in this study. This meta-analysis found that compared with conventional treatment alone, XKS combined with conventional treatment significantly improved the anxiety scale scores (SMD = -1.97, 95% CI -3.13 to -0.82; p = 0.0008; I2 = 98%), the depression scores (SMD = -2.80, 95% CI -4.49 to -1.10; p = 0.001; I2 = 98%), the scores on the Medical Outcomes Study 36 Item Short Form Health Survey (SF36) (MD = 11.22, 95% CI 4.19 to 18.26; p =0.002; I2 = 95%) and the blood lipid levels of total cholesterol (TC) (MD = -0.38, 95% CI -0.62 to -0.13; p = 0.003; I2 = 0%) and triglyceride (TG) (MD = -0.31, 95% CI -0.46 to -0.17; p < 0.0001; I2 =  0%). CONCLUSION: The current evidence suggests that XKS might benefit CHD patients experiencing anxiety and depression after PCI by helping to improve their depression symptoms, TC and TG blood lipid levels. However, due to insufficient methodological quality of the studies, several risks of bias and inadequate reporting of the clinical data, more rigorous, multicenter, sufficient-sample and double-blind randomized clinical trials are warranted.

Effectiveness and safety of Shugan Jianpi (SGJP) formula in the treatment of nonalcoholic steatohepatitis (NASH): A protocol for systematic review and meta-analysis of randomized controlled trials.

BACKGROUND: It is known that nonalcoholic steatohepatitis (NASH) has been more and more popular in clinical practice. Apart from lifestyle modification, pharmacological therapy treating NASH has still been limited and insufficient. A growing number of studies demonstrated that Shugan Jianpi (SGJP) formula, as a kind of Chinese herbal medicine prescription, could improve blood lipid indexes, liver function, and other clinical measures in NASH patients. Nevertheless, there still has been a lack of study to systematically assess the efficacy and safety of SGJP formula treating NASH. Therefore, it is necessary to conduct this systematic review and meta-analysis. METHODS: A systematic literature search for articles up to December 2021 will be performed in following electronic databases: MEDLINE, Embase, the Cochrane Library, China National Knowledge Infrastructure, Chinese Scientific Journals Database Database, Chinese Biomedical Database, Chinese Biomedical Literature Service System, and Wanfang Database. Inclusion criteria are randomized controlled trials of SGJP formula applied on NASH patients. The primary outcome measures will be liver function, blood lipid indexes, ultrasound, or radiological imaging examination. The safety outcome measures will be adverse events and kidney function. RevMan 5.3 software will be used for data synthesis, sensitivity analysis, subgroup analysis, and risk of bias assessment. A funnel plot will be developed to evaluate reporting bias. Stata 12.0 will be used for meta-regression and Egger tests. We will use the Grading of Recommendations Assessment, Development and Evaluation system to assess the quality of evidence.Discussion: This study will provide a high-quality synthesis of the efficacy and safety of SGJP for NASH patients. ETHICS AND DISSEMINATION: This systematic review does not require ethics approval and will be submitted to a peer-reviewed journal. TRIAL REGISTRATION NUMBER: PROSPERO CRD42021259097.

Construction of a genome instability-derived lncRNA-based risk scoring system for the prognosis of hepatocellular carcinoma.

Emerging evidence revealed the critical roles of long non-coding RNAs (lncRNAs) in maintaining genomic instability. However, genome instability-associated lncRNAs (GILncRNAs) and their performance in clinical prognostic significance in hepatocellular carcinoma (HCC) are rarely reported. Our study constructed a computational framework integrating somatic mutation information and lncRNA expression profiles of HCC genome and we identified 88 GILncRNAs of HCC. Function enrichment analysis revealed that GILncRNAs were involved in various metabolism processes and genome instability of cancer. A genome instability-derived lncRNA-based gene signature (GILncSig) was constructed using training set data. The performance of GILncSig for outcome prediction was validated in testing set and The Cancer Genome Atlas (TCGA) set. The multivariate cox regression analysis and stratification analysis demonstrated GILncSig could serve as an independent prognostic factor for the overall survival of HCC patients. The time-dependent Receiver Operating Characteristic (ROC) curve illustrated GILncSig outperformed two recently published lncRNA signatures for overall survival prediction. The combination of GILncSig and tumor protein p53 (TP53) mutation status exhibited better prognostic performance in survival evaluation compared to TP53 mutation status alone. AC145343.1 was further validated to be a risk factor for HCC in vitro among GILncSig. Overall, our study provided a novel approach for identification of genome instability-associated lncRNAs and established an independent risk score system for outcome prediction of HCC patients, which provided a new insight for exploring in-depth mechanism and potential therapy strategy.

Traditional Chinese medicine in the treatment of nonalcoholic steatohepatitis.

Nonalcoholic steatohepatitis (NASH) is a common chronic liver disease in clinical practice. It has been considered that NASH is one of the main causes of chronic liver disease, cirrhosis and carcinoma. The mechanism of the NASH progression is complex, including lipid metabolism dysfunction, insulin resistance, oxidative stress, inflammation, apoptosis, fibrosis and gut microbiota dysbiosis. Except for lifestyle modification and bariatric surgery, there has been no pharmacological therapy that is being officially approved in NASH treatment. Traditional Chinese medicine (TCM), as a conventional and effective therapeutic strategy, has been proved to be beneficial in treating NASH in numbers of studies. In the light of this, TCM may provide a potential therapy for treating NASH. In this review, we summarized the associated mechanisms of action TCM treating NASH in preclinical studies and systematically analysis the effectiveness of TCM treating NASH in current clinical trials.

A Non-invasive Model for Predicting Liver Inflammation in Chronic Hepatitis B Patients With Normal Serum Alanine Aminotransferase Levels.

Background and Aims: Chronic hepatitis B (CHB) patients with normal alanine aminotransferase (ALT) levels are at risk of disease progression. Currently, liver biopsy is suggested to identify this population. We aimed to establish a non-invasive diagnostic model to identify patients with significant liver inflammation. Method: A total of 504 CHB patients who had undergone liver biopsy with normal ALT levels were randomized into a training set (n = 310) and a validation set (n = 194). Independent variables were analyzed by stepwise logistic regression analysis. After the predictive model for diagnosing significant inflammation (Scheuer's system, G ≥ 2) was established, a nomogram was generated. Discrimination and calibration aspects of the model were measured using the area under the receiver operating characteristic curve (AUC) and assessment of a calibration curve. Clinical significance was evaluated by decision curve analysis (DCA). Result: The model was composed of 4 variables: aspartate aminotransferase (AST) levels, γ-glutamyl transpeptidase (GGT) levels, hepatitis B surface antigen (HBsAg) levels, and platelet (PLT) counts. Good discrimination and calibration of the model were observed in the training and validation sets (AUC = 0.87 and 0.86, respectively). The best cutoff point for the model was 0.12, where the specificity was 83.43%, the sensitivity was 77.42%, and the positive likelihood and negative likelihood ratios were 4.67 and 0.27, respectively. The model's predictive capability was superior to that of each single indicator. Conclusion: This study provides a non-invasive approach for predicting significant liver inflammation in CHB patients with normal ALT. Nomograms may help to identify target patients to allow timely initiation of antiviral treatment.

Autophagic Inhibition of Caveolin-1 by Compound Phyllanthus urinaria L. Activates Ubiquitination and Proteasome Degradation of ß-catenin to Suppress Metastasis of Hepatitis B-Associated Hepatocellular Carcinoma.

Compound Phyllanthus urinaria L. (CP) is a traditional Chinese medicine (TCM) formula for cancer treatment in the clinic, particularly during progression of hepatitis B-associated hepatocellular carcinoma (HBV-associated HCC). Nevertheless, its anti-metastatic action and mechanisms are not well elucidated. In this study, CP was found to exert remarkable inhibitory effects on the proliferation, migration and invasion of HBV-associated HCC cells. The following network and biological analyses predicted that CP mainly targeted Caveolin-1 (Cav-1) to induce anti-metastatic effects, and Wnt/ß-catenin pathway was one of the core mechanisms of CP action against HBV-associated HCC. Further experimental validation implied that Cav-1 overexpression promoted metastasis of HBV-associated HCC by stabilizing ß-catenin, while CP administration induced autophagic degradation of Cav-1, activated the Akt/GSK3ß-mediated proteasome degradation of ß-catenin via ubiquitination activation, and subsequently attenuated the metastasis-promoting effect of Cav-1. In addition, the anti-cancer and anti-metastatic action of CP was further confirmed by in vivo and ex vivo experiments. It was found that CP inhibited the tumor growth and metastasis of HBV-associated HCC in both mice liver cancer xenograft and zebrafish xenotransplantation models. Taken together, our study not only highlights the novel function of CP formula in suppressing metastasis of HBV-associated HCC, but it also addresses the critical role of Cav-1 in mediating Akt/GSK3ß/ß-catenin axis to control the late-phase of cancer progression.

Investigating the mechanisms of Modified Xiaoyaosan (tiaogan-liqi prescription) in suppressing the progression of atherosclerosis, by means of integrative pharmacology and experimental validation.

Atherosclerosis (AS)-related diseases remain among the leading causes of death worldwide. Modified Xiaoyaosan (also called Tiaogan-Liqi prescription, TGLQ), a traditional Chinese medical formulation, has been widely applied in the treatment of AS-related diseases. The aim of this study was to investigate the underlying pharmacological mechanisms of TGLQ in acting on AS. A total of 548 chemical compounds contained in TGLQ, and 969 putative targets, were collected from the Computation Platform for Integrative Pharmacology of Traditional Chinese Medicine, while 1005 therapeutic targets for the treatment of AS were obtained from the DisGeNET, TTD and CTD databases. Moreover, the 63 key targets were screened by the intersection of the targets above, and by network topological analysis. Further functional enrichment analysis showed that the key targets were significantly associated with regulation of the immune system and inflammation, improvement of lipid and glucose metabolism, regulation of the neuroendocrine system and anti-thrombosis effect. The in vivo experiments confirmed that TGLQ could reduce plasma lipid profiles and plasma inflammatory cytokines, and also inhibit AS plaque formation, within the AS model ApoE-/- mice. The in vitro experiments validated the hypothesis that TGLQ could significantly reduce intracellular lipid accumulation, suppress the production of inflammatory cytokines of macrophages induced by oxidized-LDL, and inhibit the protein expression of heat shock protein 90 and toll-like receptor 4. This study identified a list of key targets of TGLQ in the treatment of AS by applying an integrative pharmacology approach, which was validated by in vivo and in vitro experimentation.

Long-term follow-up of cumulative incidence of hepatocellular carcinoma in hepatitis B virus patients without antiviral therapy.

BACKGROUND: China has a high prevalence of hepatitis B virus (HBV), but most chronic hepatitis B (CHB) patients do not receive standardized antiviral therapy. There are few relevant reports addressing the outcomes of the large number of CHB patients who do not receive antiviral therapy. AIM: To observe the outcomes of long-term follow-up of patients with CHB without antiviral treatment. METHODS: This study included 362 patients with CHB and 96 with hepatitis B cirrhosis without antiviral treatment and with only liver protection and anti-inflammatory treatment from 1993 to 1998. The median follow-up times were 10 and 7 years, respectively. A total of 203 CHB and 129 hepatitis B cirrhosis patients receiving antiviral therapy were selected as the control groups. The median follow-up times were 8 and 7 years, respectively. Kaplan-Meier curves were used to analyze the cumulative incidence of hepatocellular carcinoma (HCC), and the Cox regression model was used to analyze the risk factors for HCC. RESULTS: Among the patients in the non-antiviral group, 16.9% had spontaneous decreases in HBV DNA to undetectable levels, and 32.8% showed hepatitis B e antigen (HBeAg) seroconversion. In the antiviral group, 87.2% of patients had undetectable HBV DNA, and 52% showed HBeAg seroconversion. Among CHB and hepatitis B cirrhosis patients, the cumulative incidence rates of HCC were 14.9% and 53.1%, respectively, in the non-antiviral group and were 10.7% and 31.9%, respectively, in the antiviral group. There was no difference between the two groups regarding the CHB patients (P = 0.842), but there was a difference between the groups regarding the hepatitis B cirrhosis patients (P = 0.026). The cumulative incidence rates of HCC were 1.6% and 22.3% (P = 0.022) in the groups with and without spontaneous HBeAg seroconversion, respectively. The incidence rates of HCC among patients with and without spontaneous declines in HBV DNA to undetectable levels were 1.6% and 19.1%, respectively (P = 0.051). There was no difference in the cumulative incidence of HCC between the two groups regarding the patients with drug-resistant CHB (P = 0.119), but there was a significant difference between the two groups regarding the patients with cirrhosis (P = 0.004). The Cox regression model was used for regression of the corrected REACH-B score, which showed that alanine aminotransferase > 400 U/L, history of diabetes, and family history of liver cancer were risk factors for HCC among men aged > 40 years (P < 0.05). Multifactorial analysis showed that a family history of HCC among men was a risk factor for HCC. CONCLUSION: Antiviral therapy and non-antiviral therapy with liver protection and anti-inflammatory therapy can reduce the risk of HCC. Antiviral therapy may mask the spontaneous serological response of some patients during CHB. Therefore, the effect of early antiviral therapy on reducing the incidence of HCC cannot be overestimated.

Bioinformatics Analyses of Potential miRNA-mRNA Regulatory Axis in HBV-related Hepatocellular Carcinoma.

Aims: We aimed to explore the crucial miRNA-mRNA axis through bioinformatics analysis and provide evidences for the development of pathophysiological mechanisms and new therapies for HBV-related HCC. Methods: MiRNA (GSE76903) and mRNA (GSE77509) dataset were used to screen differentially expressed miRNAs (DE-miRNAs) and differentially expressed mRNAs (DE-mRNAs) using R software. Overlapping genes between DE-mRNAs and target genes of DE-miRNAs were identified as candidate genes. Hub genes were obtained via cytohubba analysis. The expression at protein and mRNA levels and prognostic value of hub genes were evaluated based on The Cancer Genome Atlas (TCGA) data. Key miRNA-mRNA axes were constructed according to predicted miRNA-mRNA pairs. MiRNA expression and prognostic role were respectively identified using starBase v3.0 and Kaplan-Meier plotter database. Real-time PCR was performed to verify the expression of crucial miRNAs and mRNAs. Coexpression of crucial miRNA and mRNA were analyzed using starBase v3.0. Results: CDK1, CCNB1, CKS2 and CCNE1 were screened as hub genes, which were significantly upregulated at protein and mRNA levels. These up-regulated hub genes were also significantly associated with poor prognosis. Hsa-mir-195-5p/CDK1, hsa-mir-5589-3p/CCNB1 and hsa-let-7c-3p/CKS2 were screened as critical miRNA-mRNA axes. Critical miRNAs were decreased in HCC, which indicates unfavourable prognosis. QPCR results showed that crucial miRNAs were decreased, whereas critical mRNAs were increased in HBV-related HCC. A reverse relationship between miRNA and mRNA in crucial axis was further verified. Conclusion: This study identified several miRNA-mRNA axes in HBV-related HCC. Hsa-mir-195-5p/CDK1, hsa-mir-5589-3p/CCNB1 and hsa-let-7c-3p/CKS2 might serve as potential prognostic biomarkers and therapeutic targets for HBV-related HCC.