Free Radical Scavenging Effect and Immunomodulatory Activity of Total Saponins Extract of Ginseng Fibrous Roots.

Ginseng (Panax ginseng C.A. Mey) is known for its rich saponin compounds and tonic effects. To better utilize the medicinal value of ginseng, this study investigated the extraction process, components, free radical scavenging ability, and immunomodulatory activity of total saponins of ginseng fibrous roots. The response surface methodology was employed to optimize the extraction process of total saponins, and Q-Orbitrap high-resolution liquid chromatography-mass spectrometry (LC-MS) was used to identify the chemical constituents in the total saponins extract of ginseng fibrous roots (GRS). The results showed that the optimal extraction process was achieved with an ethanol concentration of 68%, a material-solvent ratio of 1:25 mL/g, and an extraction time of 20 min, yielding a total saponin content of 6.34% under these conditions. The extract contained four terpenoid compounds and four polyphenolic compounds. GRS exhibited considerable scavenging activity against DPPH and ABTS radicals, with IC50 values of 0.893 and 0.210 mg/mL, respectively. Moreover, GRS restored immune suppression in mice by increasing white blood cell, red blood cell, and neutrophil counts, and improving the lymphocyte. It also promoted immune system recovery, as evidenced by elevated serum levels of IL-2, IFN-γ, TNF-α, and IL-1ß in mice. GRS is a natural compound with promising potential for developing antioxidants and immunomodulatory foods.

In Vitro Anti-Tumor and Hypoglycemic Effects of Total Flavonoids from Willow Buds.

Salix babylonica L. is a species of willow tree that is widely cultivated worldwide as an ornamental plant, but its medicinal resources have not yet been reasonably developed or utilized. Herein, we extracted and purified the total flavonoids from willow buds (PTFW) for component analysis in order to evaluate their in vitro anti-tumor and hypoglycemic activities. Through Q-Orbitrap LC-MS/MS analysis, a total of 10 flavonoid compounds were identified (including flavones, flavan-3-ols, and flavonols). The inhibitory effects of PTFW on the proliferation of cervical cancer HeLa cells, colon cancer HT-29 cells, and breast cancer MCF7 cells were evaluated using an MTT assay. Moreover, the hypoglycemic activity of PTFW was determined by investigating the inhibitory effects of PTFW on α-amylase and α-glucosidase. The results indicated that PTFW significantly suppressed the proliferation of HeLa cells, HT-29 cells, and MCF7 cells, with IC50 values of 1.432, 0.3476, and 2.297 mg/mL, respectively. PTFW, at different concentrations, had certain inhibitory effects on α-amylase and α-glucosidase, with IC50 values of 2.94 mg/mL and 1.87 mg/mL, respectively. In conclusion, PTFW at different doses exhibits anti-proliferation effects on all three types of cancer cells, particularly on HT-29 cells, and also shows significant hypoglycemic effects. Willow buds have the potential to be used in functional food and pharmaceutical industries.

Control status of ambulatory blood pressure and its relationship with arterial stiffness in the China nationwide registry of treated hypertensive patients: the REACTION-ABP study.

The control rate of ambulatory blood pressure (BP) is unclear in Chinese hypertensive patients, and whether it would be associated with the ambulatory arterial stiffness indices is also unknown. From June 2018 until December 2022, 4408 treated hypertensive patients (52.8% men, average age 58.2 years) from 77 hospitals in China were registered. Ambulatory BPs were measured with validated monitors and analyzed with a web-based standardized Shuoyun system ( www.shuoyun.com.cn ). The BP control rate was the highest in the office (65.7%), moderate in the daytime (45.0%), low in the morning (34.1%), and the lowest in the nighttime (27.6%, P < 0.001). Only 21.0% had their 24 h BP perfectly controlled. The stepwise regression analyses identified that the factors associated with an imperfect 24 h BP control included male sex, smoking and drinking habits, a higher body mass index, serum total cholesterol and triglycerides, and the use of several specific types of antihypertensive drugs. After adjustment for the above-mentioned factors, the 24 h pulse pressure (PP) and its components, the elastic and stiffening PPs, were all significantly associated with an uncontrolled office and ambulatory BP status with the standardized odds ratios ranging from 1.09 to 4.68 (P < 0.05). The ambulatory arterial stiffness index (AASI) was only associated with an uncontrolled nighttime and 24 h BP status. In conclusion, the control rates of 24 h ambulatory BP, especially that in the nighttime and morning time windows, were low in Chinese hypertensive patients, which might be associated with arterial stiffness in addition to other common risk factors.

Targeting NAD+ metabolism of hepatocellular carcinoma cells by lenvatinib promotes M2 macrophages reverse polarization, suppressing the HCC progression.

BACKGROUND: Lowered nicotinamide adenine dinucleotide (NAD+) levels in tumor cells drive tumor hyperprogression during immunotherapy, and its restoration activates immune cells. However, the effect of lenvatinib, a first-line treatment for unresectable hepatocellular carcinoma (HCC), on NAD+ metabolism in HCC cells, and the metabolite crosstalk between HCC and immune cells after targeting NAD+ metabolism of HCC cells remain unelucidated. METHODS: Liquid chromatography-tandem mass spectrometry (LC-MS/MS) and ultra-high-performance liquid chromatography multiple reaction monitoring-mass spectrometry (UHPLC-MRM-MS) were used to detect and validate differential metabolites. RNA sequencing was used to explore mRNA expression in macrophages and HCC cells. HCC mouse models were used to validate the effects of lenvatinib on immune cells and NAD+ metabolism. The macrophage properties were elucidated using cell proliferation, apoptosis, and co-culture assays. In silico structural analysis and interaction assays were used to determine whether lenvatinib targets tet methylcytosine dioxygenase 2 (TET2). Flow cytometry was performed to assess changes in immune cells. RESULTS: Lenvatinib targeted TET2 to synthesize and increase NAD+ levels, thereby inhibiting decomposition in HCC cells. NAD+ salvage increased lenvatinib-induced apoptosis of HCC cells. Lenvatinib also induced CD8+ T cells and M1 macrophages infiltration in vivo. And lenvatinib suppressed niacinamide, 5-Hydroxy-L-tryptophan and quinoline secretion of HCC cells, and increased hypoxanthine secretion, which contributed to proliferation, migration and polarization function of macrophages. Consequently, lenvatinib targeted NAD+ metabolism and elevated HCC-derived hypoxanthine to enhance the macrophages polarization from M2 to M1. Glycosaminoglycan binding disorder and positive regulation of cytosolic calcium ion concentration were characteristic features of the reverse polarization. CONCLUSIONS: Targeting HCC cells NAD+ metabolism by lenvatinib-TET2 pathway drives metabolite crosstalk, leading to M2 macrophages reverse polarization, thereby suppressing HCC progression. Collectively, these novel insights highlight the role of lenvatinib or its combination therapies as promising therapeutic alternatives for HCC patients with low NAD+ levels or high TET2 levels.

HBV Enhances Sorafenib Resistance in Hepatocellular Carcinoma by Reducing Ferroptosis via SRSF2-Mediated Abnormal PCLAF Splicing.

Hepatocellular carcinoma (HCC) is one of the most lethal human cancers. Hepatitis B virus (HBV) infection accounts for nearly 50% of HCC cases. Recent studies indicate that HBV infection induces resistance to sorafenib, the first-line systemic treatment for advanced HCC for more than a decade, from 2007 to 2020. Our previous research shows that variant 1 (tv1) of proliferating cell nuclear antigen clamp-associated factor (PCLAF), overexpressed in HCC, protects against doxorubicin-induced apoptosis. However, there are no reports on the relevance of PCLAF in sorafenib resistance in HBV-related HCC. In this article, we found that PCLAF levels were higher in HBV-related HCC than in non-virus-related HCC using bioinformatics analysis. Immunohistochemistry (IHC) staining of clinical samples and the splicing reporter minigene assay using HCC cells revealed that PCLAF tv1 was elevated by HBV. Furthermore, HBV promoted the splicing of PCLAF tv1 by downregulating serine/arginine-rich splicing factor 2 (SRSF2), which hindered the inclusion of PCLAF exon 3 through a putative cis-element (116-123), "GATTCCTG". The CCK-8 assay showed that HBV decreased cell susceptibility to sorafenib through SRSF2/PCLAF tv1. HBV reduced ferroptosis by decreasing intracellular Fe2+ levels and activating GPX4 expression via the SRSF2/PCLAF tv1 axis, according to a mechanism study. Suppressed ferroptosis, on the other hand, contributed to HBV-mediated sorafenib resistance through SRSF2/PCLAF tv1. These data suggested that HBV regulated PCLAF abnormal alternative splicing by suppressing SRSF2. HBV caused sorafenib resistance by reducing ferroptosis via the SRSF2/PCLAF tv1 axis. As a result, the SRSF2/PCLAF tv1 axis may be a prospective molecular therapeutic target in HBV-related HCC, as well as a predictor of sorafenib resistance. The inhibition of the SRSF2/PCLAF tv1 axis may be crucial in the emergence of systemic chemotherapy resistance in HBV-associated HCC.

Effects of ultraviolet radiation on microorganism and nitrogen metabolism in sewage under plateau background.

The experiments were conducted in the Tibetan plateau environment, and the sewage treatment conditions were designed with ultraviolet (UV) irradiation for 5 min, 10 min, 30 min, and 180 min. The Illumina MiSeq high-throughput sequencing technology was used to analyze the microbiological and metabolomic patterns of the plateau sewage treatment at the experimental scale, and then the response mechanisms of microbial and nitrogen metabolism in sewage treatment were explored. The abundance of metabolism at the first level and global and overview maps at the second level were higher in the plateau environment than in other regions. The KEGG pathway shows the effect of UV on nitrogen metabolism and its aptitude to improving or inhibit it. The two main nitrogen removal processes are nitrification and dissimilatory nitrate reduction. This study reveals the response of activated sludge to UV radiation in a plateau environment from microbiological and metabolomic perspectives, providing ideas and perspectives for the study of water treatment system methods, as well as laying a valuable theoretical foundation for the enhancement of plateau sewage treatment capacity.

Quercetin modulates the liver metabolic profile in a chronic unpredictable mild stress rat model based on metabolomics technology.

Depression is the most prevalent psychiatric disease, and its pathogenesis is still unclear. Currently, studies on the pathogenesis of depression are mainly focused on the brain. The liver can modulate brain function via the liver-brain axis, indicating that the liver plays an important role in the development of depression. This study aims to explore the protective effect of quercetin against chronic unpredictable mild stress (CUMS)-induced metabolic changes and the corresponding mechanisms in the rat liver based on untargeted metabolomics technology. In this study, 96 male rats were divided into six groups: control, different doses of quercetin (10 mg per kg bw or 50 mg per kg bw), CUMS, and CUMS + different doses of quercetin. After 8 weeks of CUMS modeling, the liver samples were collected for metabolomics analysis. A total of 17 altered metabolites were identified, including D-glutamic acid, S-adenosylmethionine, lithocholylglycine, L-homocystine, prostaglandin PGE2, leukotriene E4, cholic acid, 5-methyltetrahydrofolic acid, taurochenodeoxycholic acid, S-adenosylhomocysteine, deoxycholic acid, folic acid, L-methionine, leukotriene C5, estriol-17-glucuronide, PE, and PC, indicating that methionine metabolism, bile acid metabolism, and phosphatidylcholine biosynthesis are the major pathways involved in CUMS-induced hepatic metabolic disorders. Hepatic methylation damage may play a role in the pathophysiology of depression, as evidenced by the first discovery of the abnormality of hepatic methionine metabolism. Abnormal changes in hepatic bile acids may provide stronger evidence for depression pathogenesis involving the microbiota-gut-brain axis, suggesting that the liver is involved in depression development and may be a treatment target. The quercetin treatment alleviated the CUMS-induced liver metabolism disorder, suggesting that quercetin may protect against depression by regulating liver metabolism.

Domesticated HERV-W env contributes to the activation of the small conductance Ca2+-activated K+ type 2 channels via decreased 5-HT4 receptor in recent-onset schizophrenia.

The human endogenous retroviruses type W family envelope (HERV-W env) gene is located on chromosome 7q21-22. Our previous studies show that HERV-W env is elevated in schizophrenia and HERV-W env can increase calcium influx. Additionally, the 5-HTergic system and particularly 5-hydroxytryptamine (5-HT) receptors play a prominent role in the pathogenesis and treatment of schizophrenia. 5-hydroxytryptamine receptor 4 (5-HT4R) agonist can block calcium channels. However, the underlying relationship between HERV-W env and 5-HT4R in the etiology of schizophrenia has not been revealed. Here, we used enzyme-linked immunosorbent assay to detect the concentration of HERV-W env and 5-HT4R in the plasma of patients with schizophrenia and we found that there were decreased levels of 5-HT4R and a negative correlation between 5-HT4R and HERV-W env in schizophrenia. Overexpression of HERV-W env decreased the transcription and protein levels of 5-HT4R but increased small conductance Ca2+-activated K+ type 2 channels (SK2) expression levels. Further studies revealed that HERV-W env could interact with 5-HT4R. Additionally, luciferase assay showed that an essential region (-364 to -176 from the transcription start site) in the SK2 promoter was required for HERV-W env-induced SK2 expression. Importantly, 5-HT4R participated in the regulation of SK2 expression and promoter activity. Electrophysiological recordings suggested that HERV-W env could increase SK2 channel currents and the increase of SK2 currents was inhibited by 5-HT4R. In conclusion, HERV-W env could activate SK2 channels via decreased 5-HT4R, which might exhibit a novel mechanism for HERV-W env to influence neuronal activity in schizophrenia.

Blue light attenuates TGF-ß2-induced epithelial-mesenchymal transition in human lens epithelial cells via autophagy impairment.

BACKGROUND: Pathogenesis of posterior capsular opacification (PCO) was related to pathological epithelial-mesenchymal transition (EMT) of lens epithelial cells (LECs). It has been reported that blue light could have an effect on EMT. This study aims to elucidate the role and potential mechanism of autophagy in EMT after blue light exposure in LECs. METHODS: HLE-B3 cells were treated with TGF-ß2 with different concentration and time to induce EMT as a model of PCO in vitro. Cells were exposed to blue light with or without TGF-ß2. The expression levels of EMT-associated markers were analyzed by qRT-PCR, western blotting and cell migration ability was determined by transwell migration assay and wound healing assay. The expressions of autophagy-related proteins were analyzed by western blotting, immunofluorescence and transmission electron microscopy. Rapamycin and chloroquine were utilized in cells for autophagy activation and inhibition. RESULTS: TGF-ß2 induced autophagy activation during EMT progression in HLE-B3 cells in a dose- and time-dependent manner. Blue light exposure inhibited TGF-ß2-induced EMT characterized by inhibited expression of EMT related markers and reduced migration capacity. Meanwhile, blue light exposure impaired autophagy activated by TGF-ß2. Furthermore, Autophagy activation with rapamycin rescued EMT attenuated by blue light. Autophagy inhibition with chloroquine reduced TGF-ß2-induced EMT in HLE-B3 cells. CONCLUSION: Blue light exposure had inhibited effects on TGF-ß2-induced EMT in LECs through autophagy impairment, which provides a new insight on prevention and treatment of PCO.

Isolated nocturnal hypertension in relation to host and environmental factors and clock genes.

Isolated nocturnal hypertension (INH) is a special type of out-of-office hypertension. Its determinants and pathophysiology remain unclear. In a nested case-control study, we intend to investigate the host, environmental, and genetic factors in relation to INH. Among 2030 outpatients screened from December 2008 till June 2015, 128 patients with INH were identified, and then 128 normotensives were matched according to sex and age. INH was an elevated nocturnal blood pressure (BP ≥120/70 mmHg) in the presence of a normal daytime BP (< 135/85 mmHg). Host factors included age, sex, body mass index, smoking and drinking, sleep time and duration, heart rate, serum lipids, and serum creatinine. Environmental cues encompassed season, ambient temperature, atmospheric pressure, humidity, and wind speed, and genetic cues 29 single-nucleotide polymorphisms (SNPs) in 12 clock genes. Daytime and nighttime BPs averaged 124.9/80.7  and 114.5/73.7 mmHg, respectively, in the INH patients and 121.0/76.5 and 101.8/63.3 mmHg in the normotensive controls. Stepwise logistic regression analyses revealed that INH was associated with nighttime heart rate (P = .0018), sleep duration (P = .0499), and relative humidity (P = .0747). The odds ratios (95% CI) for each 10 beats/min faster nighttime heart rate and 10% lower relative humidity were 1.82 (1.25-2.65) and 0.82 (0.67-1.00), respectively. Irrespective of the genetic models, no significant association was observed between INH and the SNPs (P ≥ .054). In conclusion, INH was associated with host and environmental factors rather than genetic markers.

Astragaloside-IV alleviates high glucose-induced ferroptosis in retinal pigment epithelial cells by disrupting the expression of miR-138-5p/Sirt1/Nrf2.

Astragaloside-IV (AS-IV) (C41H68O14) is a high-purity natural product extracted from Astragalus, which has demonstrated biological activities. However, the effect of AS-IV on retinal pigment epithelial (RPE) cells in diabetic retinopathy (DR) remains unclear. In this study, high glucose (HG) was shown to promote ARPE-19 RPE cell death, increase the contents of reactive oxygen species (ROS) and oxidized glutathione (GSSG), and enhance lipid peroxidation density of mitochondrial membrane. In contrast, AS-IV decreased glutathione (GSH) content, mitochondria size and ridge. Addition of iron death inhibitor Ferrostatin-1 (Fer-1) to RPE cells decreased cell dead rate, thus indicating that HG-induced mitochondrial damage occurred due to ferroptosis. AS-IV alleviated HG-induced RPE cell damage. Furthermore, HG decreased levels of silent information regulator 1 (Sirt1) and nuclear factor (erythroid-derived 2)-like 2 (Nrf2) in the nucleus of RPE cells; AS-IV could alleviate these effects and increased expression of glutathione peroxidase 4 (GPX4), glutamate cysteine ligase (GCLM) and glutamate cysteine ligase catalytic subunit (GCLC), which are Nrf2 downstream genes. Mechanistically, AS-IV was shown to alleviate the effects of HG by increasing mir-138-5p expression in RPE cells and promoting expression of Sirt1 and Nrf2 in the nucleus. Transfection of mir-138-5p agonist inhibited the regulatory effects of AS-IV on Sirt1 and Nrf2, accompanied by decreased GPX4, GCLM and GCLC levels, and restoration of ferroptosis-related changes. Collectively, HG increased ferroptosis rate in RPE cells. In addition, AS-IV inhibited miR-138-5p expression, subsequently increasing Sirt1/Nrf2 activity and cellular antioxidant capacity to alleviate ferroptosis, resulting decreased cell death, which potentially inhibits the DR pathological process.

UBE2T-mediated Akt ubiquitination and Akt/ß-catenin activation promotes hepatocellular carcinoma development by increasing pyrimidine metabolism.

The oncogene protein ubiquitin-conjugating enzyme E2T (UBE2T) is reported to be upregulated in hepatocellular carcinoma (HCC) and correlated with poor clinical outcomes of HCC patients. However, the underlying mechanism by which UBE2T exerts its oncogenic function in HCC remains largely unexplored. In this study, in vitro and in vivo experiments suggested that UBE2T promoted HCC development including proliferation and metastasis. GSEA analysis indicated that UBE2T was positively correlated with pyrimidine metabolism, and LC/MS-MS metabolomics profiling revealed that the key products of pyrimidine metabolism were significantly increased in UBE2T-overexpressing cells. UBE2T overexpression led to the upregulation of several key enzymes catalyzing de novo pyrimidine synthesis, including CAD, DHODH, and UMPS. Moreover, the utilization of leflunomide, a clinically approved DHODH inhibitor, blocked the effect of UBE2T in promoting HCC progression. Mechanistically, UBE2T increased Akt K63-mediated ubiquitination and Akt/ß-catenin signaling pathway activation. The disruption of UBE2T-mediated ubiquitination on Akt, including E2-enzyme-deficient mutation (C86A) of UBE2T and ubiquitination-site-deficient mutation (K8/14 R) of Akt impaired UBE2T's effect in upregulating CAD, DHODH, and UMPS. Importantly, we demonstrated that UBE2T was positively correlated with p-Akt, ß-catenin, CAD, DHODH, and UMPS in HCC tumor tissues. In summary, our study indicates that UBE2T increases pyrimidine metabolism by promoting Akt K63-linked ubiquitination, thus contributing to HCC development. This work provides a novel insight into HCC development and a potential therapeutic strategy for HCC patients.

Hepatic Insulin Resistance Is Not Pathway Selective in Humans With Nonalcoholic Fatty Liver Disease.

OBJECTIVE: Both glucose and triglyceride production are increased in type 2 diabetes and nonalcoholic fatty liver disease (NAFLD). For decades, the leading hypothesis to explain these paradoxical observations has been selective hepatic insulin resistance wherein insulin drives de novo lipogenesis (DNL) while failing to suppress glucose production. Here, we aimed to test this hypothesis in humans. RESEARCH DESIGN AND METHODS: We recruited obese subjects who met criteria for bariatric surgery with (n = 16) or without (n = 15) NAFLD and assessed 1) insulin-mediated regulation of hepatic and peripheral glucose metabolism using hyperinsulinemic-euglycemic clamps with [6,6-2H2]glucose, 2) fasting and carbohydrate-driven hepatic DNL using deuterated water (2H2O), and 3) hepatocellular insulin signaling in liver biopsy samples collected during bariatric surgery. RESULTS: Compared with subjects without NAFLD, those with NAFLD demonstrated impaired insulin-mediated suppression of glucose production and attenuated-not increased-glucose-stimulated/high-insulin lipogenesis. Fructose-stimulated/low-insulin lipogenesis was intact. Hepatocellular insulin signaling, assessed for the first time in humans, exhibited a proximal block in insulin-resistant subjects: Signaling was attenuated from the level of the insulin receptor through both glucose and lipogenesis pathways. The carbohydrate-regulated lipogenic transcription factor ChREBP was increased in subjects with NAFLD. CONCLUSIONS: Acute increases in lipogenesis in humans with NAFLD are not explained by altered molecular regulation of lipogenesis through a paradoxical increase in lipogenic insulin action; rather, increases in lipogenic substrate availability may be the key.

Treatment of Masked Hypertension with a Chinese Herbal Formula: A Randomized, Placebo-Controlled Trial.

BACKGROUND: Masked hypertension is associated with adverse cardiovascular outcomes. Nonetheless, no randomized controlled trials exist in the treatment of masked hypertension. The aim of this randomized, placebo-controlled trial was to investigate the efficacy and safety of blood pressure (BP)-lowering treatment with a Chinese herbal formula, gastrodia-uncaria granules, in patients with masked hypertension. METHODS: Patients with an office BP of <140/90 mm Hg and daytime ambulatory BP of 135 to 150 mm Hg systolic or 85 to 95 mm Hg diastolic were randomly assigned 1:1 to the treatment of gastrodia-uncaria granules or placebo 5 to 10 g twice daily for 4 weeks. The primary efficacy variable was the change in daytime ambulatory BP. RESULTS: At baseline, office and daytime BP of the 251 participants (mean age, 50.4 years; 53.4% men; mean body mass index 24.5 kg/m2; and 2.8%, 1.6%, and 30.7% with cardiovascular disease, diabetes, and smoking, respectively) averaged 129/82 and 135/89 mm Hg, respectively. In the intention-to-treat analysis, daytime systolic/diastolic BP was reduced by 5.44/3.39 and 2.91/1.60 mm Hg in the gastrodia-uncaria granules and placebo groups, respectively. The between-group difference in BP reductions was significant for the daytime (2.52/1.79 mm Hg; P≤0.025) and 24-hour BP (2.33/1.49 mm Hg; P≤0.012), but not for the clinic and nighttime BPs (P≥0.162). The per-protocol analysis in 229 patients produced similar results. Only 1 adverse event (sleepiness during the day) was reported, and no serious adverse event occurred. CONCLUSIONS: BP-lowering treatment with Chinese traditional medicine gastrodia-uncaria granules is efficacious for patients with masked hypertension. Registration: URL: https://www.clinicaltrials.gov; Unique identifier: NCT02156024.

Reciprocal regulation of HIF-1α and Uroplakin 1A promotes glycolysis and proliferation in Hepatocellular Carcinoma.

Uroplakin 1A (UPK1A) has recently been found dysregulation in many cancers. However, the functions of UPK1A and its underlying mechanisms in hepatocellular carcinoma (HCC) remain poorly understand. In the present study, we found that UPK1A was highly expressed in HCC tumor tissues compared with adjacent non-tumor tissues. Datasets from the Cancer Genome Atlas project (TCGA) and Gene expression Omnibus confirmed that UPK1A was highly expressed in HCC. High expression of UPK1A predicted poor overall survival (OS) in patients with HCC. Univariate and multivariate analysis showed that UPK1A was a significant and independent prognostic predictor for OS of patients with HCC. Functionally, silencing UPK1A suppressed HCC cell glycolysis and proliferation. Mechanistically, hypoxia-inducible factor 1α (HIF-1α) directly bound to the hypoxia response elements (HRE) of UPK1A promoter region, which led to the up-regulation of UPK1A under hypoxia. Furthermore, downregulation of UPK1A reduced key enzyme of glycolysis via regulating HIF-1α. Taken together, these data indicates the existence of a positive feedback loop between HIF-1α and UPK1A that modulates glycolysis and proliferation under hypoxia in HCC cells.

Long noncoding RNA UPK1A-AS1 indicates poor prognosis of hepatocellular carcinoma and promotes cell proliferation through interaction with EZH2.

BACKGROUND: Dysregulation of long non-coding RNAs (lncRNAs) is responsible for cancer initiation and development, positioning lncRNAs as not only biomarkers but also promising therapeutic targets for cancer treatment. A growing number of lncRNAs have been reported in hepatocellular carcinoma (HCC), but their functional and mechanistic roles remain unclear. METHODS: Gene Set Enrichment Analysis was used to investigate the molecular mechanism of UPK1A antisense RNA 1 (UPK1A-AS1). Cell Counting Kit-8 assays, EdU assays, flow cytometry, western blotting, and xenograft assays were used to confirm the role of UPK1A-AS1 in the proliferation of HCC cells in vitro and in vivo. Bioinformatics analyses and quantitative polymerase chain reaction (qRT-PCR) were performed to explore the interplay between UPK1A-AS1 and enhancer of zeste homologue 2 (EZH2). RNA immunoprecipitation (RIP), RNA pull-down assays, western blotting, and qRT-PCR were conducted to confirm the interaction between UPK1A-AS1 and EZH2. The interaction between UPK1A-AS1 and miR-138-5p was examined by luciferase reporter and RIP assays. Finally, the expression level and prognosis value of UPK1A-AS1 in HCC were analyzed using RNA sequencing data from The Cancer Genome Atlas datasets. RESULTS: We showed that UPK1A-AS1, a newly identified lncRNA, promoted cellular proliferation and tumor growth by accelerating cell cycle progression. Cell cycle-related genes, including CCND1, CDK2, CDK4, CCNB1, and CCNB2, were significantly upregulated in HCC cells overexpressing UPK1A-AS1. Furthermore, overexpression of UPK1A-AS1 could protect HCC cells from cis-platinum toxicity. Mechanistically, UPK1A-AS1 interacted with EZH2 to mediate its nuclear translocation and reinforce its binding to SUZ12, leading to increased H27K3 trimethylation. Targeting EZH2 with specific small interfering RNA impaired the UPK1A-AS1-mediated upregulation of proliferation and cell cycle progression-related genes. Moreover, miR-138-5p was identified as a direct target of UPK1A-AS1. Additionally, UPK1A-AS1 was significantly upregulated in HCC, and the upregulation of UPK1A-AS1 predicted poor prognosis for patients with HCC. CONCLUSIONS: Our study revealed that UPK1A-AS1 promotes HCC development by accelerating cell cycle progression through interaction with EZH2 and sponging of miR-138-5p, suggesting that UPK1A-AS1 possesses substantial potential as a novel biomarker for HCC prognosis and therapy.

HRG switches TNFR1-mediated cell survival to apoptosis in Hepatocellular Carcinoma.

Background: Tumor necrosis factor receptor 1 (TNFR1) signaling plays a pleiotropic role in the development of hepatocellular carcinoma (HCC). The formation of TNFR1-complex I supports cell survival while TNFR1-complex II leads to apoptosis, and the underlying mechanisms of the transformation of these TNFR1 complexes in HCC remain poorly defined. Methods: The interaction protein of TNFR1 was identified by GST pulldown assay, immunoprecipitation and mass spectrometry. In vitro and in vivo assay were performed to explore the biological features and mechanisms underlying the regulation of TNFR1 signals by histidine-rich glycoprotein (HRG). Data from the public databases and HCC samples were utilized to analyze the expression and clinical relevance of HRG. Results: HRG directly interacted with TNFR1 and stabilized TNFR1 protein by decreasing the Lys(K)-48 ubiquitination mediated-degradation. The formation of TNFR1-complex II was prompted by HRG overexpression via upregulating Lys(K)-63 ubiquitination of TNFR1. Besides, overexpression of HRG suppressed expression of pro-survival genes by impairing the activation of NF-κB signaling in the presence of TNFR1. Moreover, downregulation of HRG was a result of feedback inhibition of NF-κB activation in HCC. In line with the pro-apoptotic switch of TNFR1 signaling after HRG induction, overexpression of HRG inhibited cell proliferation and increased apoptosis in HCC. Conclusions: Our findings illustrate a crucial role for HRG in suppressing HCC via inclining TNFR1 to a pro-apoptotic cellular phenotype. Restoring HRG expression in HCC tissues might be a promising pharmacological approach to blocking tumor progression by shifting cellular fate from cell survival to apoptosis.

MiR-424 Functions as Potential Diagnostic and Prognostic Biomarker in Melanoma.

BACKGROUND: Melanoma is one of the most aggressive and lethal skin cancers worldwide. To our knowledge, no specific or sensitive biomarkers have been clinically used to diagnose or predict melanoma prognosis. MicroRNAs (miRNAs) have been shown to regulate oncogenesis and tumor development in various cancers including melanoma. The aim of present study was to determine the clinical value of miR-424 in melanoma. METHODS: First, we examined the expression levels of miR-424 in tissue and serum samples of melanoma patients using real-time quantitative polymerase chain reaction (RT-qPCR) analysis. Then, receiver operating characteristic curves were used to determine the diagnostic value of miR-424. Furthermore, the chi-square test was used to analyze the relationship between the expression of miR-424 and the clinical characteristics of the patients. Finally, the Kaplan-Meier survival analysis was applied to validate the prognostic value of miR-424 in melanoma. RESULTS: The results demonstrated that miR-424 expression was remarkably increased in tissues and serum of pa-tients with melanoma. Moreover, results of ROC analysis showed both tissue and serum expression of miR-424 can serve as diagnostic biomarker for melanoma. Meanwhile, miR-424 expression was significantly associated with tumor thickness (p = 0.031), metastasis (p = 0.010) and tumor stage (p = 0.005) and ulceration (p < 0.001). Finally, patients with higher miR-424 expression have shown decreased overall survival and disease-free survival than those with low miR-424 expression, implying that high miR-424 expression will contribute to poor prognosis of melanoma. CONCLUSIONS: MiR-424 may function as a diagnostic and prognostic biomarker for melanoma.

OGT suppresses S6K1-mediated macrophage inflammation and metabolic disturbance.

Enhanced inflammation is believed to contribute to overnutrition-induced metabolic disturbance. Nutrient flux has also been shown to be essential for immune cell activation. Here, we report an unexpected role of nutrient-sensing O-linked ß-N-acetylglucosamine (O-GlcNAc) signaling in suppressing macrophage proinflammatory activation and preventing diet-induced metabolic dysfunction. Overnutrition stimulates an increase in O-GlcNAc signaling in macrophages. O-GlcNAc signaling is down-regulated during macrophage proinflammatory activation. Suppressing O-GlcNAc signaling by O-GlcNAc transferase (OGT) knockout enhances macrophage proinflammatory polarization, promotes adipose tissue inflammation and lipolysis, increases lipid accumulation in peripheral tissues, and exacerbates tissue-specific and whole-body insulin resistance in high-fat-diet-induced obese mice. OGT inhibits macrophage proinflammatory activation by catalyzing ribosomal protein S6 kinase beta-1 (S6K1) O-GlcNAcylation and suppressing S6K1 phosphorylation and mTORC1 signaling. These findings thus identify macrophage O-GlcNAc signaling as a homeostatic mechanism maintaining whole-body metabolism under overnutrition.