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Objective: Excessive lipid accumulation in adipocytes results in prevalence of obesity and metabolic syndrome. Curcumin (CUR), a naturally phenolic active ingredient, has been shown to have lipid-lowering effects. However, its underlying mechanisms have remained largely unknown. Therefore, the study aims to determine the effect of CUR on cellular lipid accumulation in porcine subcutaneous preadipocytes (PSPA) and to clarify novel mechanisms.Methods: The PSPA were cultured and treated with or without CUR. Both cell counting Kit-8 and lactate dehydrogenase release assays were used to examine cytotoxicity. Intracellular lipid contents were measured by oil-red-o staining extraction and triglyceride quantification. Apoptosis was determined by flow cytometry and the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-nick end labelling assay. Adipogenic and apoptosis genes were analyzed by quantitative polymerase chain reaction and Western blot.Results: The CUR dose-dependently reduced the proliferation and lipid accumulation of PSPA. Noncytotoxic doses of CUR (10 to 20 μM) significantly inhibited extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation and expression of adipogenic genes peroxisome proliferation-activity receptor-γ (PPAR-γ), CCAAT/enhancer binding protein-α, sterol regulatory element-binding protein-1c, adipocyte protein-2, glucose transporter-4 as well as key lipogenic enzymes fatty acid synthase and acetyl-CoA carboxylase, while ERK1/2 activation significantly reversed CUR-reduced lipid accumulation by increasing PPAR-γ. Furthermore, compared with differentiation induced media treated cells, higher dose of CUR (30 μM) significantly decreased the expression of AKT and B-cell lymphoma-2 (BCL2), while increased the expression of BCL-2-associated X (BAX) and the BAX/BCL-2 expression ratio, suggesting triggered apoptosis by inactivating AKT and increasing BAX/BCL-2 ratio and Caspase-3 expression. Moreover, AKT activation significantly rescued CUR inhibiting lipid accumulation via repressing apoptosis.Conclusion: These results demonstrate that CUR is capable of suppressing differentiation by inhibiting ERK1/2-PPAR-γ signaling pathway and triggering apoptosis via decreasing AKT and subsequently increasing BAX/BCL-2 ratio and Caspase-3, suggesting that CUR provides an important method for the reduction of porcine body fat, as well as the prevention and treatment of human obesity.

CCAAT/Enhancer Binding protein beta (C/EBPβ) is a transcriptional regulator involved in numerous physiological processes. Herein, we describe a role for C/EBPβ as a regulator of skeletal muscle stem cell function. In particular, C/EBPβ is expressed in muscle stem cells in healthy muscle where it inhibits myogenic differentiation. Downregulation of C/EBPβ expression at the protein and transcriptional level allows for differentiation. Persistence of C/EBPβ promotes stem cell self-renewal and C/EBPβ expression is required for mitotic quiescence in this cell population. As a critical regulator of skeletal muscle homeostasis, C/EBPβ expression is stimulated in pathological conditions such as cancer cachexia, which perturbs muscle regeneration and promotes myofiber atrophy in the context of systemic inflammation. C/EBPβ is also an important regulator of cytokine expression and immune response genes, a mechanism by which it can influence muscle stem cell function. In this viewpoint, we describe a role for C/EBPβ in muscle stem cells and propose a functional intersection between C/EBPβ and NF-kB action in the regulation of cancer cachexia.

Fatty acid binding protein 4 (FABP4) was crucial to fatty acid uptake and intracellular transport. However, the mechanisms regulating yak (Bos grunniens) FABP4 transcription were not determined. In the current study, predominant expression levels of yak FABP4 were identified in subcutaneous fat and longissimus dorsi muscles by quantitative real-time polymerase chain reactions (qPCR). The CCAAT/enhancer binding protein alpha (CEBPα) and myocyte enhancer factor 2A (MEF2A), as transcriptional activator or repressor in the promoter region of FABP4, were confirmed by both site-directed mutagenesis experiment and chromatin immunoprecipitation assay. Additionally, molecular mechanisms of CEBPɑ regulation were analyzed to explore the transcriptional regulatory property of FABP4, which indicated that transcriptional activity of CEBPɑ depended on CCAAT/ enhancer binding protein beta (CEBPβ) transcription factor. Our results demonstrated that CEBPβ binding directly to the promoter region drove CEBPɑ transcription, improving yak FABP4 transcriptional activity in adipocytes. This mechanism expanded the information on the transcriptional regulatory network of adipogenesis.

Human adipose-derived stem cells (hADSCs) and human umbilical vein endothelial cells (HUVECs) co-cultured in vitro are widely used in adipose tissue engineering but exhibit various limitations. Chemokine (C-C motif) ligand 2 (CCL2) has been proved essential during adipogenesis and angiogenesis in vivo. We examined whether adipogenesis and angiogenesis could also be directly promoted by CCL2 in vitro. Cells were cultured with 0, 10, 50, and 100 ng/ml CCL2. The effects of CCL2 on adipogenesis of hADSCs, and lipid accumulation in the positive control group (hADSCs), blank control group (hADSCs + HUVECs), and experimental group (hADSCs + HUVECs + CCL2) in the hADSC and HUVEC direct co-culture system were evaluated by Oil Red O staining. Angiogenesis in the presence of CCL2 was evaluated by Matrigel tube formation assay. Angiogenic- and adipogenic-associated gene and protein expression in the co-culture system were measured by Quantitative Real-time Polymerase Chain Reaction and western blotting, respectively. All concentrations of CCL2 promoted hADSC adipogenic differentiation and HUVEC tube formation (P

In the endoplasmic reticulum (ER), accumulation of abnormal proteins with malformed higher-order structures activates signaling pathways (inositol-requiring enzyme 1α (IRE1α)/X-box binding protein 1 (XBP-1) pathway, protein kinase RNA-activated-like endoplasmic reticulum kinase (PERK)/CCAAT/enhancer binding protein-homologous protein (CHOP) pathway and activating transcription factor 6α (ATF6α) pathway) that result in a cellular response suppressing the production of abnormal proteins or inducing apoptosis. These responses are collectively known as the unfolded protein response (UPR). Recently, it has been suggested that the UPR induced by saturated fatty acids in hepatocytes and pancreatic β cells is involved in the development of metabolic diseases such as diabetes. The effect of palmitate, a saturated fatty acid, on the UPR has also been investigated in adipocytes, which are associated with the development of metabolic disorders, but the results were inconclusive. Therefore, as the major saturated fatty acids present in the daily diet are palmitate and stearate, we examined the effects of these saturated fatty acids on UPR in adipocytes. Here, we show that saturated fatty acids caused limited activation of the UPR in adipocytes. Exposure to stearate for several hours elevated the ratio of spliced XBP-1 mRNA, and this effect was stronger than that of palmitate. Moreover, the phosphorylation level of IRE1α, upstream of XBP-1 and expression levels of its downstream targets such as DNAJB9 and Pdia6 were elevated in 3T3-L1 adipocytes exposed to stearate. On the other hand, stearate did not affect the phosphorylation of PERK, its activation of CHOP, or the cleavage of ATF6α. Thus, in adipocytes, exposure to stearate activates the UPR via the IRE1α/XBP-1 pathway, but not the PERK/CHOP and ATF6α pathway.

The objective of the study was to investigate the effect of nobiletin on adipogenesis in 3T3-L1 cells. Here, we found that nobiletin could promote adipogenesis in 3T3-L1 cells in the absence of adipogenic inducers such as insulin, 3-Isobutyl-1-methylxanthine and dexamethasone. In addition, Real time quantitative PCR showed that the expression of adipogenic genes such as CCAAT/enhancer binding proteins, peroxisome proliferator-activated receptors-γ and fatty acid-binding protein aP2 were up-regulated in 3T3-L1 cells in the presence of nobiletin. Next, we investigated the role of Akt in the effect of nobiletin on the adipogenesis in 3T3-L1 cells by LY294002 blocking PI3K/Akt pathway. The experimental results showed that the promotive effect of nobiletin on adipogenesis was attenuated, accompanied by the down-regulation of adipogenic genes expression, after the activity of Akt was inhibited, suggesting that Akt plays an important role in the effect of nobiletin on promoting adipogenesis in 3T3-L1 cells. Still, the inhibition of Akt pathway by LY294002 was attenuated in the presence of nobiletin. These findings provide a possibility that nobiletin may be a potential candidate agent for stimulating Akt pathway in 3T3-L1 cells.

The overexpansion of adipose tissues leads to obesity and eventually results in metabolic disorders. Garcinia cambogia (G. cambogia) has been used as an antiobesity supplement. However, the molecular mechanisms underlying the effects of G. cambogia on cellular processes have yet to be fully understood. Here, we discovered that G. cambogia attenuated the expression of CEBPB (CCAAT/enhancer binding protein (C/EBP), beta), an important adipogenic factor, suppressing its transcription in differentiated cells. In addition, G. cambogia inhibited macroautophagic/autophagic flux by decreasing autophagy-related gene expression and autophagosome formation. Notably, G. cambogia markedly elevated the expression of KLF3 (Kruppel-like factor 3 (basic)), a negative regulator of adipogenesis, by reducing SQSTM1/p62-mediated selective autophagic degradation. Furthermore, increased KLF3 induced by G. cambogia interacted with CTBP2 (C-terminal binding protein 2) to form a transcriptional repressor complex and inhibited Cebpa and Pparg transcription. Importantly, we found that RPS6KA1 and STAT3 were involved in the G. cambogia-mediated regulation of CEBPB and autophagic flux. In an obese animal model, G. cambogia reduced high-fat diet (HFD)-induced obesity by suppressing epididymal and inguinal subcutaneous white adipose tissue mass and adipocyte size, which were attributed to the regulation of targets that had been consistently identified in vitro. These findings provide new insight into the mechanism of G. cambogia-mediated regulation of adipogenesis and suggest molecular links to therapeutic targets for the treatment of obesity. Abbreviations: 3-MA: 3-methyladenine; ACTB: actin beta; ATG: autophagy-related; Baf: bafilomycin A(1); BECN1: beclin 1; CEBP: CCAAT/enhancer binding protein (C/EBP); CHX: cycloheximide; CREB: cAMP response element binding protein; CTBP: C-terminal binding protein; EGCG: (-)-epigallocatechin gallate; eWAT: epididymal white; G. cambogia: Garcinia cambogia; GFP: green fluorescent protein; H&E: hematoxylin and eosin; HFD: high-fat diet; iWAT: inguinal subcutaneous white; KLF: Kruppel-like factor; LAP: liver-enriched transcriptional activating proteins; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; ND: normal diet; PPARG: peroxisome proliferator activated receptor gamma; qPCR: quantitative real-time PCR; RFP: red fluorescent protein; RPS6KA1: ribosomal protein S6 kinase A1; siRNA: small-interfering RNA; SQSTM1/p62: sequestosome 1; STAT: signal transducer and activator of transcription; TEM: transmission electron microscopy

Tubulointerstitial inflammation plays an important role in the progression of diabetic nephropathy (DN), and tubular epithelial cells (TECs) are crucial promoters of the inflammatory cascade. Exchange protein activated by cAMP (Epac) has been shown to suppress the angiotensin II (Ang-II)-induced release of inflammatory cytokines in tubular cells. However, the role of Epac in TEC-mediated tubulointerstitial inflammation in DN remains unknown. We found that administering the Epac agonist 8-pCPT-2'-O-Me-cAMP (8-O-cAMP) to db/db mice inhibited tubulointerstitial inflammation characterized by macrophage infiltration and increased inflammatory cytokine release and consequently alleviated tubulointerstitial fibrosis in the kidney. Furthermore, 8-O-cAMP administration restored CCAAT/enhancer binding protein β (C/EBP-β) expression and further upregulated the expression of Suppressor of cytokine signaling 3 (SOCS3), while inhibiting p-STAT3, MCP-1, IL-6, and TNF-α expression in the kidney cortex in db/db mice. And in vitro study showed that macrophage migration and MCP-1 expression induced by high glucose (HG, 30 mM) were notably reduced by 8-O-cAMP in human renal proximal tubule epithelial (HK-2) cells. In addition, 8-O-cAMP treatment restored C/EBP-β expression in HK-2 cells and promoted C/EBP-β translocation to the nucleus, where it transcriptionally upregulated SOCS3 expression, subsequently inhibiting STAT3 phosphorylation. Under HG conditions, siRNA-mediated knockdown of C/EBP-β or SOCS3 in HK-2 cells partially blocked the inhibitory effect of Epac activation on the release of MCP-1. In contrast, SOCS3 overexpression inhibited HG-induced activation of STAT3 and MCP-1 expression in HK-2 cells. These findings indicate that Epac activation via 8-O-cAMP ameliorates tubulointerstitial inflammation in DN through the C/EBP-β/SOCS3/STAT3 pathway.

Neuroblastoma (NB) is the most common extracranial solid malignancy in children and its mortality rate is relatively high. However, driver genes of NB are not clearly identified. Using bioinformatics analysis, we determined the top 8 differentially expressed genes (DEGs) in NB, including GFAP, PAX6, FOXG1, GAD1, PTPRC, ISL1, GRM5, and GATA3. Insulin gene enhancer binding protein 1 (ISL1) is a LIM homeodomain transcription factor which has been found to be highly expressed in a variety of malignant tumors, but the function of ISL1 in NB has not been fully elucidated. We identified ISL1 as an oncogene in NB. ISL1 is preferentially upregulated in NB tissues compared with normal tissues. High ISL1 expression is significantly associated with poor outcome of NB patients. Knockdown of ISL1 markedly represses proliferation and induces cell apoptosis in vitro, and suppresses tumorigenicity in vivo, while overexpression of ISL1 has the opposite effects. Mechanistically, we demonstrate that ISL1 promotes cell proliferation and EMT transformation through PI3K/AKT signaling pathway by upregulating Aurora kinase A (AURKA), a serine-threonine kinase that is essential for the survival of NB cells. The blockade of AURKA attenuates the function of ISL1 overexpression in the regulation of cell proliferation and migration, Conclusively, this study showed that ISL1 targeted AURKA to facilitate the development of NB, which provided new insights into the tumorigenesis of NB. Thus, ISL1 may be a promising therapeutic target in the future.

Nonalcoholic fatty liver disease (NAFLD) is the leading cause of chronic liver disease globally. NAFLD is a consequence of fat accumulation in the liver leading to lipotoxicity. Increasing evidence has demonstrated the critical role of autophagy in NAFLD. This study uncovers the unexpected role of immune surveillance protein DDX58/Rig-1 (DExD/H box helicase 58) in activating macroautophagy/autophagy and protecting from lipotoxicity associated with NAFLD. Here we show for the first time that DDX58 protein is significantly reduced in nonalcoholic steatohepatitis (NASH) mouse model, an aggressive form of NAFLD characterized by inflammation and fibrosis of the liver. In addition to decreased expression of DDX58, we found that DDX58 activity can be attenuated by treatments with palmitic acid (PA), a saturated fatty acid. To investigate whether PA inhibition of DDX58 is harmful to the cell, we characterized DDX58 function in hepatocytes when exposed to high doses of PA in the presence and/or absence of DDX58. We show that siRNA knockdown of DDX58 promotes apoptosis. Importantly, we show that stable overexpression of DDX58 is protective against toxic levels of PA and stimulates autophagy. This study begins to demonstrate the regulation of the autophagy receptor protein SQSTM1/p62 through DDX58. DDX58 expression directly influences SQSTM1 mRNA and protein levels. This work proposes a model in which activating DDX58 increases an autophagic response and this aids in clearing toxic lipid inclusion bodies, which leads to inflammation and apoptosis. Activating a DDX58-induced autophagy response may be a strategy for treating NAFLD. Abbreviations:5ʹpppdsRNA: 5ʹ triphosphate double-stranded RNA; CDAHFD: choline-deficient, L-amino acid defined high-fat diet; CEBPB: CCAAT/enhancer binding protein (C/EBP), beta; CQ: chloroquine; DDX58/retinoic acid inducible gene 1/Rig-1: DExD/H box helicase 58; h: hours; IFIH1/MDA5: interferon induced with helicase C domain 1; IFNB/IFN-β: interferon beta 1, fibroblast; KO: knockout; MAVS: mitochondrial antiviral signaling protein; NAFLD: nonalcoholic fatty liver disease; NASH: nonalcoholic steatohepatitis; NFKB/NF-κB: nuclear factor of kappa light polypeptide gene enhancer in B cells; PA: palmitic acid; poly:IC: polyinosinic:polycytidylic acid; PRR: pattern recognition receptors; PSR: picrosirus red; RAP: rapamycin; RLR: RIG-I-like receptor; SQSTM1/p62: sequestosome 1; STING1: stimulator of interferon response cGAMP interactor 1; TBK1: TANK-binding kinase 1.

DNA cytosine methylation in mammals modulates gene expression and chromatin accessibility. It also impacts mutation rates, via spontaneous oxidative deamination of 5-methylcytosine (5mC) to thymine. In most cases the resulting T:G mismatches are repaired, following T excision by one of the thymine DNA glycosylases, TDG or MBD4. We found that C-to-T mutations are enriched in the binding sites of CCAAT/enhancer binding proteins (CEBP). Within a CEBP site, the presence of a T:G mismatch increased CEBPβ binding affinity by a factor of >60 relative to the normal C:G base pair. This enhanced binding to a mismatch inhibits its repair by both TDG and MBD4 in vitro. Furthermore, repair of the deamination product of unmethylated cytosine, which yields a U:G DNA mismatch that is normally repaired via uracil DNA glycosylase, is also inhibited by CEBPβ binding. Passage of a replication fork over either a T:G or U:G mismatch, before repair can occur, results in a C-to-T mutation in one of the daughter duplexes. Our study thus provides a plausible mechanism for accumulation of C-to-T human somatic mutations.

Background Although the morbidity and mortality rates associated with idiopathic pulmonary fibrosis (IPF) are high, there is still lack of powerful and precise therapeutic options for IPF. Object Through in vitro model, this study sought to determine whether binding of acetylated CCAAT/enhancer binding protein β (C/EBPβ) to alpha-smooth muscle actin (α-SMA) promoter could affect the activity of the latter as well as assess if it is essential for epithelial-to-mesenchymal transition (EMT) and extracellular matrix deposition in IPF. Methods The expression of EMT and C/EBPβ in A549 cells treated with transforming growth factor-beta (TGF-β) as pulmonary fibrotic model was detected by western blotting and qPCR. Collagen-I expression using ELISA was performed. The luciferase activity was used to examine the activity of C/EBPβ. Knockdown of C/EBPβ was performed by siRNA. We also investigated the effect of deacetylation of C/EBPβ on EMT using sirtuin 1 (SIRT1). The binding ability of C/EBPβ with α-SMA promoter was affirmed via chromatin immunoprecipitation (ChIP) and electrophoresis mobility shift assay (EMSA). The relationship between α-SMA and acetylated C/EBPβ was determined with co-immunoprecipitation (Co-IP). SiRNA-mediated knockdown of C/EBPβ in A549 cells attenuated TGF-β1-induced myofibroblast differentiation and ECM deposition. The extent of association between acetylated C/EBPβ and α-SMA promoter was dynamically monitored. Results It was confirmed that deacetylation of C/EBPβ in A549 cells successfully ameliorated TGF-β1-induced EMT, as shown by reduction in α-SMA expression and excessive collagen-I accumulation. Conclusion The EMT and fibrotic effect of TGF-β1 is dependent on acetylated C/EBPβ-mediated regulation of α-SMA gene activity. Thus, C/EBPβ acetylation may play a central role in pulmonary fibrosis.

Objective: Vanin1 (VNN1) is a pantetheinase that can catalyze the hydrolysis of pantetheine to produce pantothenic acid and cysteamine. Our previous studies showed that VNN1 is specifically expressed in chicken liver. In this study, we aimed to investigate the roles of peroxisome proliferators activated receptor α (PPARα) and miRNA-181a-5p in regulating VNN1 gene expression in chicken liver.Methods: 5′-RACE was performed to identify the transcription start site of chicken VNN1. JASPAR and TFSEARCH were used to analyze the potential transcription factor binding sites in the promoter region of chicken VNN1 and miRanda was used to search miRNA binding sites in 3′ untranslated region (3′UTR) of chicken VNN1. We used a knock-down strategy to manipulate PPARα (or miRNA-181a-5p) expression levels in vitro to further investigate its effect on VNN1 gene transcription. Luciferase reporter assays were used to explore the specific regions of VNN1 targeted by PPARα and miRNA-181a-5p.Results: Sequence analysis of the VNN1 promoter region revealed several transcription factor-binding sites, including hepatocyte nuclear factor 1α (HNF1α), PPARα, and CCAAT/enhancer binding protein α. GW7647 (a specific agonist of PPARα) increased the expression level of VNN1 mRNA in chicken primary hepatocytes, whereas knockdown of PPARα with siRNA increased VNN1 mRNA expression. Moreover, the predicted PPARα-binding site was confirmed to be necessary for PPARα regulation of VNN1 gene expression. In addition, the VNN1 3′UTR contains a sequence that is completely complementary to nucleotides 1 to 7 of miRNA-181a-5p. Overexpression of miR-181a-5p significantly decreased the expression level of VNN1 mRNA.Conclusion: This study demonstrates that PPARα is an important transcriptional activator of VNN1 gene expression and that miRNA-181a-5p acts as a negative regulator of VNN1 expression in chicken hepatocytes.

Background Gualou Guizhi Decoction (GLGZD) is commonly used to treat stroke. The present study investigated the potential roles of GLGZD on inflammation involving microRNA-155 (miR-155) in a model of ischemic stroke using middle cerebral artery occlusion (MCAO) rats. Methods Sprague-Dawley rats were randomly divided into three groups: Sham operated group, MCAO model group, and GLGZD treatment group. The ischemic model was established by 2 h left MCAO followed by reperfusion. Neurological deficits were evaluated with a modified Ashworth scale in each group. The changes in individual paw parameters were assessed by Catwalk gait analysis. Inflammatory cytokines were measured by enzyme linked immunosorbent assay (ELISA) and protein levels and gene expression related to inflammation were detected by Western blot and quantitative reverse transcription-PCR (qRTPCR) assays, respectively. The expression of inflammatory signaling proteins was additionally detected by immunohistochemistry. Results Treatment of MCAO rats with GLGZD improved neuronal defects and limb motivity. Additionally, GLGZD was able to inhibit miR-155 upregulation, resulting in down-regulation of miR155-targeted molecules in MCAO rats, including suppressor of cytokine signaling 1 (SOCS1), inhibitor of nuclear factor kappa-B kinase (IKK), mothers against decapentaplegic homolog 2 (SMAD2) and CCAAT/ enhancer binding protein beta (CEBPβ). Meanwhile, the production of anti-inflammatory cytokines was dramatically enhanced by GLGZD treatment when comparing with the MCAO model group. Conclusions In conclusion, GLGZD down-regulates miR-155, mediating subsequent neuroinflammation and resulting in neuroprotection which contributes to reduced spasticity after ischemic stroke.

The central region of the intervertebral disc (IVD) is rich in proteoglycans, leading to a hyperosmotic environment, which fluctuates with daily loading. The cells of the nucleus pulposus (NP cells) have adapted to this environment via the function of tonicity enhancer binding protein (TonEBP), and NP cells have been shown to express several water channels known as aquaporins (AQP). We have previously shown that AQP1 and 5 decrease during IVD degeneration. Here, the regulation of AQP1 and 5 by hyperosmotic conditions and the role of TonEBP in this regulation was investigated. AQP1 and 5 gene expression was upregulated by hyperosmotic conditions mimicking the osmolality of the healthy IVD, which was abrogated by TonEBP knockdown. Furthermore, AQP1 and 5 immunopositivity was significantly reduced in TonEBPΔ/Δ E17.5 mice when compared with wildtype controls, indicating in vivo expression of AQP1 and 5 is controlled at least in part by TonEBP. This hyperosmotic regulation of AQP1 and 5 could help to explain the decreased AQP1 and 5 expression during degeneration, when the osmolality of the NP decreases. Together this data suggests that TonEBP-regulated osmo-adaptation may be disrupted during IVD degeneration when the expression of both AQPs is reduced.

Endoplasmic reticulum stress is an important contributor to the cerebral ischemic injury. Sappanone A (SA), a kind of natural homoisoflavanone extracted from Caesalpinia sappan L, has been evidenced to exhibit anti-inflammatory and antioxidative properties. The present study aimed to investigate the potential neuroprotective effects of SA in cerebral ischemia-reperfusion injury. The potential neuroprotective effect of SA was tested in a rat model of middle cerebral artery occlusion (MCAO) allowing reperfusion and PC12 cell model of oxygen-glucose deprivation and reperfusion (OGD/R). Post-ischemic neuronal injury was evaluated by 2, 3, 5-triphenyltetrazolium chloride (TTC) and hematoxylin-eosin (H&E) staining. The levels of inflammatory factors and oxidative stress-related markers were detected using corresponding kits. Cell apoptosis was evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) or flow cytometry, and the expression of apoptosis-associated proteins was determined using western blot analysis. Subsequently, endoplasmic reticulum stress-related proteins were detected through western blot analysis, and CCAAT/enhancer binding protein (C/EBP) homologous protein (CHOP) was overexpressed to confirm the contribution of endoplasmic reticulum stress inhibition by SA to the neuroprotective effects post OGD/R. Results revealed that SA was effective in ameliorating cerebral infarction and pathological injuries post-reperfusion following MCAO, which is associated with reduced inflammation, oxidative stress, and cell apoptosis by SA in the brain. Consistently, these neuroprotective effects of SA post ischemia-reperfusion were also observed in a PC12 cell model of OGD/R. Importantly, endoplasmic reticulum stressors, including the CHOP, the 78 kDa glucose-regulated protein 78 (GRP78), and phosphorylated eukaryotic initiation factors 2α (EIF-2α), were significantly downregulated by SA, while CHOP overexpression attenuated the beneficial effects of SA on inflammation, oxidative stress, and apoptosis in OGD/R-induced PC12 cells. These results demonstrated that SA alleviates endoplasmic reticulum stress, ameliorating inflammation, oxidative stress, and apoptosis, and thereby serves as therapeutic potential for protection against cerebral ischemia-reperfusion injury in ischemic stroke.

Background & Aims Epigenetic regulation of gene expression plays a critical role in the development of liver cancer; however, the molecular mechanisms of epigenetic-driven liver cancers are not well understood. The aims of this study were to examine molecular mechanisms that cause the dedifferentiation of hepatocytes into cancer cells in aggressive hepatoblastoma and test if the inhibition of these mechanisms inhibits tumor growth. Methods We have analyzed CCAAT/Enhancer Binding Protein alpha (C/EBPα), Transcription factor Sp5, and histone deacetylase (HDAC)1 pathways from a large biobank of fresh hepatoblastoma (HBL) samples using high-pressure liquid chromatography–based examination of protein–protein complexes and have examined chromatin remodeling on the promoters of markers of hepatocytes and p21. The HDAC1 activity was inhibited in patient-derived xenograft models of HBL and in cultured hepatoblastoma cells and expression of HDAC1-dependent markers of hepatocytes was examined. Results Analyses of a biobank showed that a significant portion of HBL patients have increased levels of an oncogenic de-phosphorylated-S190-C/EBPα, Sp5, and HDAC1 compared with amounts of these proteins in adjacent regions. We found that the oncogenic de-phosphorylated-S190-C/EBPα is created in aggressive HBL by protein phosphatase 2A, which is increased within the nucleus and dephosphorylates C/EBPα at Ser190. C/EBPα–HDAC1 and Sp5–HDAC1 complexes are abundant in hepatocytes, which dedifferentiate into cancer cells. Studies in HBL cells have shown that C/EBPα–HDAC1 and Sp5–HDAC1 complexes reduce markers of hepatocytes and p21 via repression of their promoters. Pharmacologic inhibition of C/EBPα–HDAC1 and Sp5–HDAC1 complexes by Suberoylanilide hydroxamic acid (SAHA) and small interfering RNA–mediated inhibition of HDAC1 increase expression of hepatocyte markers, p21, and inhibit proliferation of cancer cells. Conclusions HDAC1-mediated repression of markers of hepatocytes is an essential step for the development of HBL, providing background for generation of therapies for aggressive HBL by targeting HDAC1 activities., Graphical abstract

Rosa davurica Pall. (RDP) fruits are popularly consumed as beverages and healthy food in China because of their various beneficial activities. In particular, flavonoids are one of the major active ingredients of RDP fruits with predominant pharmacological effects. However, the anti-obesity activities of flavonoids from RDP fruits and their regulation effect on the gut microbiota have not been determined. In the present study, the flavonoid-rich extracts (RDPF) were isolated from RDP fruits and their anti-obesity effects were investigated using a high-fat diet (HFD)-induced obese mouse model. The results showed that RDPF intervention significantly inhibited the body weight, liver weight, kidney weight and epididymal adipose tissue weight of HFD-fed mice without affecting the calorie intake. Plasma lipid levels were also significantly lowered by RDPF treatment. Histological examination showed that RDPF supplementation partially recovered HFD-induced hepatic steatosis in the liver. RDPF also prevented oxidative injury of the liver, as evidenced by the altered superoxide dismutase (SOD), catalase (CAT) and malondialdehyde (MDA) levels. The expression levels of CCAAT/enhancer binding protein α (C/EBPα), sterol regulatory element binding protein-1C (SREBP-1C), fatty acid synthase (FAS), acyl-coenzyme A oxidase 1 (ACOX1), peroxisome proliferator-activated receptor (PPARα) and CAT mRNA in the livers of mice were also regulated by RDPF administration. 16S rRNA gene sequence data further indicated that RDPF addition increased the microbial diversity and reshaped the community composition. Intriguingly, RDPF intervention did not exhibit inhibitory tendency toward the ratio of Firmicutes to Bacteroidetes, but markedly decreased the relative abundance of Erysipelotrichaceae. This study provided novel insights into the application of RDPF in the food industry.

Studies have shown that transcription factor activating enhancer binding protein 4 (TFAP4) plays a vital role in multiple types of cancer; however, the TFAP4 expression profile is still unknown, as is its value within the human pan‐cancer analysis. The present study comprehensively analysed TFAP4 expression patterns from 33 types of malignancies, along with the significance of TFAP4 for prognosis prediction and cancer immunity. TFAP4 displayed inconsistent levels of gene expression across the diverse cancer cell lines, and displayed abnormal expression within most malignant tumours, which closely corresponded to overall survival. More importantly, the TFAP4 level was also significantly related to the degree of tumour infiltration. TFAP4 was correlated using gene markers in tumour‐infiltrating immune cells and immune scores. TFAP4 expression was correlated with tumour mutation burden and microsatellite instability in different cancer types, and enrichment analyses identified TFAP4‐associated terms and pathways. The present study comprehensively analysed the expression of TFAP4 across 33 distinct types of cancers, which revealed that TFAP4 may possibly play a vital role during cancer formation and development. TFAP4 is related to differing degrees of immune infiltration within cancers, which suggests the potential of TFAP4 as an immunotherapy target in cancers. Our study demonstrated that TFAP4 plays an important role in tumorigenesis as a prognostic biomarker, which highlights the possibility of developing new targeted treatments.

The single-exon gene CEBPA encodes for CCAAT/enhancer binding protein-α, a transcription factor essential for granulocyte differentiation. In acute myeloid leukemia (AML), CEBPA mutations occur at ...

It is well-documented that lead (Pb) toxicity can affect almost all systems in living organisms. It can induce selective autophagy of mitochondria (mitophagy) by triggering reactive oxygen species production. Emerging evidence has suggested that Pb-induced autophagy can also be activated by the endoplasmic reticulum (ER) stress pathway. However, the interplay between ER stress and mitophagy remains to be elucidated. In this study, human embryonic kidney HEK293 cells were employed to investigate the role of ER stress in Pb-induced mitophagy. The results showed that the cell viability was decreased and cell damage was induced after exposure to Pb (0, 0.5, 1, 2, and 4 mM) for 24 h in a dose-dependent manner. Moreover, the expression of LC3-Ⅱ was significantly increased, and the expression of HSP60 was dramatically decreased after exposure to 1 mM and 2 mM Pb, indicating the induction of mitophagy following Pb exposure. Meanwhile, the expressions of activating transcription factor 6, inositol-requiring protein-1α, CCAAT/enhancer binding protein homologous protein, and glucose-regulated protein 78 were dramatically increased after Pb treatment, signifying the initiation of ER stress. Notably, the mitophagic effect was significantly compromised when ER stress was inhibited by 0.5 mM 4-phenylbutyrate, which was evidenced by lesser decreases in HSP60 expression and level of LC3-Ⅱ, suggesting Pb-induced mitophagy may be activated by the ER stress. Taken together, these findings provide a better understanding of Pb toxicity and suggest that Pb-induced ER stress may play a regulatory role in the upstream of mitophagy.

Senile osteoporosis (OP) is often concomitant with decreased autophagic activity. OPTN (optineurin), a macroautophagy/autophagy (hereinafter referred to as autophagy) receptor, is found to play a pivotal role in selective autophagy, coupling autophagy with bone metabolism. However, its role in osteogenesis is still mysterious. Herein, we identified Optn as a critical molecule of cell fate decision for bone marrow mesenchymal stem cells (MSCs), whose expression decreased in aged mice. Aged mice revealed osteoporotic bone loss, elevated senescence of MSCs, decreased osteogenesis, and enhanced adipogenesis, as well as optn(–)(/ –) mice. Importantly, restoring Optn by transplanting wild-type MSCs to optn(–)(/ –) mice or infecting optn(–)(/ –) mice with Optn-containing lentivirus rescued bone loss. The introduction of a loss-of-function mutant of Optn(K193R) failed to reestablish a bone-fat balance. We further identified FABP3 (fatty acid binding protein 3, muscle and heart) as a novel selective autophagy substrate of OPTN. FABP3 promoted adipogenesis and inhibited osteogenesis of MSCs. Knockdown of FABP3 alleviated bone loss in optn(–)(/ –) mice and aged mice. Our study revealed that reduced OPTN expression during aging might lead to OP due to a lack of FABP3 degradation via selective autophagy. FABP3 accumulation impaired osteogenesis of MSCs, leading to the occurrence of OP. Thus, reactivating OPTN or inhibiting FABP3 would open a new avenue to treat senile OP. Abbreviations: ADIPOQ: adiponectin, C1Q and collagen domain containing; ALPL: alkaline phosphatase, liver/bone/kidney; BGLAP/OC/osteocalcin: bone gamma carboxyglutamate protein; BFR/BS: bone formation rate/bone surface; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CDKN1A/p21: cyclin-dependent kinase inhibitor 1A; CDKN2A/p16: cyclin dependent kinase inhibitor 2A; CDKN2B/p15: cyclin dependent kinase inhibitor 2B; CEBPA: CCAAT/enhancer binding protein (C/EBP), alpha; COL1A1: collagen, type I, alpha 1; Ct. BV/TV: cortical bone volume fraction; Ct. Th: cortical thickness; Es. Pm: endocortical perimeter; FABP4/Ap2: fatty acid binding protein 4, adipocyte; H2AX: H2A.X variant histone; HE: hematoxylin and eosin; MAP1LC3B: microtubule-associated protein 1 light chain 3 beta; MAR: mineral apposition rate; MSCs: bone marrow mesenchymal stem cells; NBR1: NBR1, autophagy cargo receptor; OP: osteoporosis; OPTN: optineurin; PDB: Paget disease of bone; PPARG: peroxisome proliferator activated receptor gamma; Ps. Pm: periosteal perimeter; qRT-PCR: quantitative real-time PCR; γH2AX: Phosphorylation of the Serine residue of H2AX; ROS: reactive oxygen species; RUNX2: runt related transcription factor 2; SA-GLB1: senescence-associated (SA)-GLB1 (galactosidase, beta 1); SP7/Osx/Osterix: Sp7 transcription factor 7; SQSTM1/p62: sequestosome 1; TAX1BP1: Tax1 (human T cell leukemia virus type I) binding protein 1; Tb. BV/TV: trabecular bone volume fraction; Tb. N: trabecular number; Tb. Sp: trabecular separation; Tb. Th: trabecular thickness; μCT: micro computed tomography.

This study was conducted to compare the mRNA expression levels of myogenic-adipogenic makers in the skeletal muscle and adipocytes formation, body weight, rumen weight, and papilla length on Hanwoo calves at newborn and 6 months of age. Animals used three newborn Hanwoo calves (NC) and three Hanwoo calves 6 months of age (SC). Body weight and rumen weight were significantly increased in SC compared to NC (p < 0.01), and papilla length was longer about 10-fold in SC than NC. Adipocytes was possible to visually identify more adipocytes in SC compared to NC, and were mainly formed around the blood vessels. mRNA expression of myogenin, myosin heavy chain 1 and myosin heavy chain 2A in both longissimus dorsi (LD) and semimembranosus (SM) was found to increase with calves growth (p < 0.01), and it was confirmed that have higher levels of mRNA expression in SM than LD. In LD tissues, the mRNA expression of stearoyl-CoA desaturase (SCD, p < 0.03) and peroxisome proliferator activated receptor γ (PPARγ, p < 0.04) was significantly higher in SC than NC. In SM tissues, mRNA expression levels of SCD (p < 0.02) and CCAAT/enhancer binding protein β (C/EBPβ, p < 0.01) were higher in SC than NC, and also mRNA expression levels of PPARγ increased, but there was no significant difference. Thus, the calves period suggests that it is an important step in the development of the rumen and the myogenesis and adipogenesis.

Introduction: Minimal residual disease (MRD) assessment in acute myeloid leukemia (AML) cases is a complex, multi-modality process and, though much of its clinical implications at different points are extensively studied, it remains even now a challenging area. It is a disease the biology of which governs the modality of MRD assessment; in patients harboring specific molecular targets, high sensitivity techniques can be applied. On the other hand, relapse is considered as the leading cause of treatment failure in AML patients undergoing allogeneic hematopoietic stem cell transplantation (alloHSCT). Materials and methods: Since November 2018 until June 2020, 10 AML patients underwent matched unrelated donor (MUD) HSCT at the University Clinic of Hematology-Skopje, Republic of North Macedonia. Molecular markers were identified in a total of 4 patients; 3 patients expressed chimeric fusion transcripts; two RUNX-RUNX1T1 and one for CBFB-MYH11. One patient harbored mutation in the transcription factor CCAAT/enhancer binding protein α (CEBPA). Post-transplant MRD kinetics was evaluated by using quantitative polymerase chain reaction (RT-qPCR) or multiplex fluorescent-PCR every three months during the first two years after the transplantation. Results: MRD negativity was achieved in three pre-transplant MRD positive patients by the sixth month of HSCT. They sustained hematological and molecular remission for 19, 9 and 7 months, respectively. The fourth patient died due to transplant-related complications. Conclusion: According to our experience, when molecularly-defined AML patients undergo HSCT, regular MRD monitoring helps predict impending relapse and direct future treatment strategies.

This study was aimed to investigate the inhibitory effects of Porphyra dentata (P. dentata) extract on the adipogenesis of 3T3-L1 cells and evaluate its anti-obesity effect. The proliferation of 3T3-L1 cells and differentiation of adipocytes under treatment of P. dentata extract was examined by measuring the cell viability using alamarBlue assay and lipid droplets by Oil Red O staining. Results showed that P. dentata extract has no cytotoxicity effect and lipid droplets formation decreased in a concentration-dependent manner in 3T3-L1 cells. It has been confirmed that transcription factors affecting lipid accumulation and anti-adipogenic effects during cell differentiation are linked to P. dentata extract. We observed that P. dentata shows lowering the mRNA expression of peroxisome proliferator-activated receptor γ2 (PPARγ2), CCAAT/enhancer binding protein α (C/EBPα) that adipogenesis-associated key transcription factors and inhibiting adipogenesis in the early stages of differentiation. Treating the cells with P. dentata did not only suppressed PPARγ2 and C/EBPα but also significantly decreased the mRNA expression of adiponectin, Leptin, fatty acid synthase, adipocyte protein 2, and Acetyl-coA carboxylase 1. Overall, the P. dentata extract demonstrated inhibitory property in adipogenesis, which has a potential effect in anti-obesity in 3T3-L1 cells.

The three-dimensional configuration of the chromatin architecture is known to be crucial for alterations in the transcriptional network; however, the underlying mechanisms of epigenetic control of senescence-related gene expression by modulating the chromatin architecture remain unknown. Here, we demonstrate frequent chromosomal compartment switching during mouse embryonic fibroblasts (MEFs) replicative senescence as characterized by senescence-inactivated (SIAEs) and -activated enhancers (SAEs) in topologically associated domains (TADs). Mechanistically, SAEs are closely correlated with senescence-associated secretory phenotype (SASP) genes, which are a key transcriptional feature of an aging microenvironment that contributes to tumor progression, aging acceleration, and immunoinflammatory responses. Moreover, SAEs can positively regulate robust changes in SASP expression. The transcription factor CCAAT/enhancer binding protein α (C/EBPα) is capable of enhancing SAE activity, which accelerates the emergence of SAEs flanking SASPs and the secretion of downstream factors, contributing to the progression of senescence. Our results provide novel insight into the TAD-related control of SASP gene expression by revealing hierarchical roles of the chromatin architecture, transcription factors, and enhancer activity in the regulation of cellular senescence.

Interleukin enhancer-binding factor 2 (ILF2) forms a heterodimer with interleukin enhancer-binding factor 3 (ILF3) via double-stranded RNA-binding motif and zinc finger associated domain and thus regulates gene expression and cancer cell growth. However, how ILF2 is degraded in cells remains elusive. In this work, using stable isotope labeling by amino acids in cell culture (SILAC) quantitative proteomics, we find that ILF2 is downregulated in cells expressing cereblon (CRBN). Using affinity purification and immunoblotting analysis, we demonstrate that CRBN interacts with ILF2 and functions as a substrate receptor of the cullin-4 RING E3 ligase complex. Biochemical experiments disclose that CRBN expression reduces ILF2 protein level and this reduction is diminished when the proteasome is inhibited. Upon protein synthesis inhibition, the degradation of ILF2 is enhanced by CRBN. Moreover, CRBN promotes the ubiquitination of ILF2 and thus results in the ubiquitin-mediated proteasomal degradation. Analyses of previously identified post-translational modification sites and the crystal structure of ILF2 discover the potential ubiquitination sites on ILF2. Through mutagenesis and biochemical experiments, we further reveal that the K45R mutation completely abolishes the effect of CRBN on ILF2, suggesting that this is the key residue responsible for its ubiquitination. Taken together, we identify an E3 ligase that regulates ILF2 and uncover a molecular pathway for its degradation. This work might be helpful to elucidate the molecular mechanism by which CRBN regulates diverse cellular functions.

Background Breast cancer (BC) is a malignant tumor that occurs in the epithelial tissue of the breast gland. Long non-coding RNA (lncRNA) small nucleolar RNA host gene 3 (SNHG3) has been found to promote BC cell proliferation and invasion by regulating the microRNA (miR)-101/zinc-finger enhancer binding axis in BC. Herein, the objective of the present study is to evaluate the effect of lncRNA SNHG3 on BC cell proliferation and metastasis with the Notch signaling pathway. Methods Differentially expressed lncRNA in BC tissues and normal breast tissues was analyzed. SNHG3 si-RNA-1 and SNHG3 si-RNA-2 were constructed to detect the mechanism of SNHG3 interference in BC cell proliferation, viability, migration and invasion. Then, dual-luciferase reporter gene assay was utilized to verify the binding relation between SNHG3 and miR-154-3p as well as miR-154-3p and Notch2. Moreover, xenograft transplantation was applied to confirm the in vitro experiments. Results Highly expressed SNHG3 was observed in BC tissues. The growth of BC cells in vivo and in vitro was evidently repressed after silencing SNHG3. BC cell invasion and migration were inhibited by silencing SNHG3 in vitro. SNHG3 could act as a competing endogenous RNA of miR-154-3p and upregulate the Notch signaling pathway to promote BC cell development. Activation of the Notch signaling pathway can partly reverse the inhibition of cell activity induced by silencing SNHG3. Conclusion Our study demonstrated that interfered lncRNA SNHG3 promoted BC cell proliferation and metastasis by activating the Notch signaling pathway. This investigation may offer new insight for BC treatment.

The transcription factor nuclear factor kappa B (NF-κB) regulates the expression of many inflammatory genes that are overexpressed in a subset of people with schizophrenia. Transcriptional reduction in one NF-κB inhibitor, Human Immunodeficiency Virus Enhancer Binding Protein 2 (HIVEP2), is found in the brain of patients, aligning with evidence of NF-κB over-activity. Cellular co-expression of HIVEP2 and cytokine transcripts is a prerequisite for a direct effect of HIVEP2 on pro-inflammatory transcription, and we do not know if changes in HIVEP2 and markers of neuroinflammation are occurring in the same brain cell type. We performed in situ hybridisation on postmortem dorsolateral prefrontal cortex tissue to map and compare the expression of HIVEP2 and Serpin Family A Member 3 (SERPINA3), one of the most consistently increased inflammatory genes in schizophrenia, between schizophrenia patients and controls. We find that HIVEP2 expression is neuronal and is decreased in almost all grey matter cortical layers in schizophrenia patients with neuroinflammation, and that SERPINA3 is increased in the dorsolateral prefrontal cortex grey matter and white matter in the same group of patients. We are the first to map the upregulation of SERPINA3 to astrocytes and to some neurons, and find evidence to suggest that blood vessel-associated astrocytes are the main cellular source of SERPINA3 in the schizophrenia cortex. We show that a lack of HIVEP2 in mice does not cause astrocytic upregulation of Serpina3n but does induce its transcription in neurons. We speculate that HIVEP2 downregulation is not a direct cause of astrocytic pro-inflammatory cytokine synthesis in schizophrenia but may contribute to neuronally-mediated neuroinflammation.

Endoplasmic reticulum (ER) stress and subsequent apoptosis played vital role in liver injury and dysfunction. The aim of this study was to investigate the protective effect and mechanism of salidroside on hypoxia induced liver injury both in vivo and in vitro. Male SD rats were exposed to hypobaric chamber to simulate high altitude hypoxia model. High altitude hypoxia led to significant liver injury and apoptosis, increased the expression levels of p-JNK, BAX and ER stress markers. Salidroside treatment significantly inhibited hypoxia induced ER stress by decreasing the protein expression of glucose-regulated protein 78 (GRP78), CCAAT/enhancer binding protein homologous protein (CHOP) and phosphorylated inositol-requiring enzyme 1α (p-IRE1α). In addition, salidroside treatment also restrained the ER stress-mediated apoptotic pathway, as indicated by decreased pro-apoptotic proteins p-JNK, TRAF2, BAX, and cleaved caspase 9 and caspase 12, as well as upregulation of Bcl-2. Furthermore, in vitro study found that blocking IRE1α pathway using specific inhibitor STF-083010 subsequently reversed the protective effect of salidroside on liver apoptosis. Taken together, our findings revealed that salidroside exerts protective effects against hypoxia induced liver injury through inhibiting ER stress mediated apoptosis via IRE1α/JNK pathway.

Establishment of a physiologically relevant human hepatocyte‐like cell system for in vitro translational research has been hampered by the limited availability of cell models that accurately reflect human biology and the pathophysiology of human disease. Here we report a robust, reproducible, and scalable protocol for the generation of hepatic organoids from human induced pluripotent stem cells (hiPSCs) using short exposure to nonengineered matrices. These hepatic organoids follow defined stages of hepatic development and express higher levels of early (hepatocyte nuclear factor 4A [HNF4A], prospero‐related homeobox 1 [PROX1]) and mature hepatic and metabolic markers (albumin, asialoglycoprotein receptor 1 [ASGR1], CCAAT/enhancer binding protein α [C/EBPα]) than two‐dimensional (2D) hepatocyte‐like cells (HLCs) at day 20 of differentiation. We used this model to explore the biology of the pleiotropic TRIB1 (Tribbles‐1) gene associated with a number of metabolic traits, including nonalcoholic fatty liver disease and plasma lipids. We used genome editing to delete the TRIB1 gene in hiPSCs and compared TRIB1‐deleted iPSC‐HLCs to isogenic iPSC‐HLCs under both 2D culture and three‐dimensional (3D) organoid conditions. Under conventional 2D culture conditions, TRIB1‐deficient HLCs showed maturation defects, with decreased expression of late‐stage hepatic and lipogenesis markers. In contrast, when cultured as 3D hepatic organoids, the differentiation defects were rescued, and a clear lipid‐related phenotype was noted in the TRIB1‐deficient induced pluripotent stem cell HLCs. Conclusion: This work supports the potential of genome‐edited hiPSC‐derived hepatic 3D organoids in exploring human hepatocyte biology, including the functional interrogation of genes identified through human genetic investigation.

BACKGROUND Endoplasmic reticulum (ER) stress is an important mechanism in the progression of chronic and acute liver diseases, especially in the progression and recovery of liver fibrosis. Excessive and long-term ER stress induces apoptosis. ER stress-induced apoptosis is considered to be an important pathway in the development of liver fibrosis. Cyclooxygenase-2 (COX-2) induction is also closely related to ER stress. In our previous studies, we showed that celecoxib, a COX-2 inhibitor, improves liver fibrosis and portal hypertension. However, the role and mechanism of celecoxib in alleviating liver fibrosis remain unclear. AIM To investigate whether celecoxib alleviates liver fibrosis by inhibiting hepatocyte apoptosis via the ER stress response. METHODS Cirrhosis was induced by intraperitoneal injections of thioacetamide (TAA) for 16 wk (injection dose is 200 mg/kg per 3 d for the first 8 wk and 100 mg /kg per 3 d after 8 wk). Thirty-six male Sprague-Dawley rats were randomly divided into three groups, namely, control group, TAA group, and TAA + celecoxib group. In the last 8 wk, TAA-induced cirrhotic rats received celecoxib (20 mg/kg/day) or the vehicle by gastric gavage. After 16 wk, the rats were sacrificed, and serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), and albumin (ALB) were detected. The hepatic fibrosis areas were evaluated by Sirius red staining and the degree of fibrosis was assessed by measuring the level of hydroxyproline. ER stress levels were evaluated by detecting the marker proteins glucose-regulated protein 78 (GRP78), CCAAT/enhancer binding protein homologous protein (CHOP), PKR-like ER protein kinase (PERK), activating transcription factor 6 (ATF6), and inositol-requiring enzyme 1 alpha (IRE1α). Apoptosis levels were evaluated by detecting caspase-12 and caspase-3. RESULTS The serum ALT and AST levels in the liver were significantly reduced by celecoxib; however, the serum ALB had no significant changes. Celecoxib significantly reduced the degree of liver fibrosis and the levels of hydroxyproline (-38% and -25.7%, respectively, P < 0.01). Celecoxib ameliorated ER stress by reducing the level of GRP78 compared to the TAA group (P < 0.05). Consistently, after celecoxib administration, the upregulation of TAA-induced hepatic apoptosis markers (caspase-12 and caspase-3) and CHOP were significantly inhibited. In addition, after celecoxib treatment, the expression of key molecules associated with ER stress (PERK, ATF6, and IRE1) was decreased (P < 0.05). CONCLUSION Therapeutic administration of celecoxib effectively reduces hepatic apoptosis in TAA-induced cirrhotic rats. The mechanism of action may be attributed to the suppression of CHOP expression, which subsequently inhibits ER stress.

Background and aims A 42 year-old male with premature atherosclerosis, severe dyslipidemia and resistance to treatment with high dose statin and a recommended dose of a PCSK9 inhibitor, was found to have a duplication of the PCSK9 gene. However, the clinical phenotype, which included a more than 15-fold elevation in circulating PCSK9, was unexpected given that he had one additional gene copy. Methods Here we have carried out whole genome sequencing and transcriptional reporter assays to investigate the molecular mechanism leading to this unusual FH phenotype. Results The PCSK9 duplication was found to contain the full coding sequence but with an 829 bp shorter 3′-untranslated region (UTR) sequence. All possible rearrangements include a head-to-head fusion between a completely duplicated PCSK9 and a chromosomal region, normally situated ~80 kb away, that includes HNF4 and USF1 binding sites that could promote transcription of the PCSK9 gene. Transcriptional reporter assays demonstrated that a construct harboring the HNF4 binding site significantly increased the promoter activity by 2.5-fold with a smaller effect noted for a USF1 construct. Conclusions Here we describe, in a patient with resistant hypercholesterolemia, a novel PCSK9 gene rearrangement that enables upregulation of PCSK9 expression by allowing proximity to an active enhancer binding to HNF4A.

In this study, we tested the hypothesis that overexpression of miR-9 and miR-29a may contribute to DPN development and progression. We performed a meta-analysis of miR expression profile studies in human diabetes mellitus (DM) and the data suggested that miR-9 and miR-29a were highly expressed in patients with DM, which was further verified in serum samples collected from 30 patients diagnosed as DM. Besides, ISL1 was confirmed to be a target gene of miR-9 and miR-29a. Lentivirus-mediated forced expression of insulin gene enhancer binding protein-1 (ISL1) activated the sonic hedgehog (SHH) signaling pathway, increased motor nerve conduction velocity and threshold of nociception, and modulated expression of neurotrophic factors in sciatic nerves in rats with DM developed by intraperitoneal injection of 0.45% streptozotocin, suggesting that ISL1 could delay DM progression and promote neural regeneration and repair after sciatic nerve damage. However, lentivirus-mediated forced expression of miR-9 or miR-29a exacerbated DM and antagonized the beneficial effect of ISL1 on DPN. Collectively, this study revealed potential roles of miR-9 and miR-29a as contributors to DPN development through the SHH signaling pathway by binding to ISL1. Additionally, the results provided an experimental basis for the targeted intervention treatment of miR-9 and miR-29a.

To develop anti-tumor agents for lung cancer, we aim to characterize a herbal compound, daidzein-rich isoflavones aglycone (DRIA), in inhibiting the proliferation and NF-κB signaling pathway of lung cancer. MTT and colony formation assays were used to analyze the proliferation of lung cancer cells in presence of DRIA treatment, which showed that DRIA dose-dependently inhibited the proliferation and colony formation of lung cancer cells. Enzyme-linked immunosorbent assay revealed that interleukin-6 (IL6) and interleukin-8 (IL-8) levels were reduced by DRIA. p65-NFκB expression and activation, which was enhanced by TNF-α and C/EBPβ treatment, were attenuated by DRIA. Exogenous tumor necrosis factor-α (TNF-α) and CCAAT/enhancer binding protein (C/EBPβ) were used to enhance NF-κB signaling in cells, and the effects of DRIA in attenuating NF-κB signaling were assessed by analyzing p65-NFκB expression in mRNA and protein levels, using quantitative real-time PCR (qRT-PCR), western blot and immunofluorescence staining. immunohistochemical staining revealed that Ki-67 and p65-NF-κB levels in A594 tumor xenografts of A594 tumors were also reduced by DRIA treatment in mice. Our data indicates that DRIA is effective in inhibiting the proliferation and NFκB signaling of lung cancer both in vitro and in vivo.

Bronchopulmonary dysplasia (BPD) is a major cause of mortality and morbidity in premature infants, characterized by alveolar simplification, surfactant deficiency, and respiratory distress. In the present study, we have investigated the functional roles of sumoylated CCAAT/enhancer binding protein alpha (C/EBPα) in the BPD rat model. A significant increase in small ubiquitin‐like modifier 1 (SUMO1) and sumoylated C/EBPα protein levels were observed in BPD rats, and the levels of the sumoylated C/EBPα were associated with the pulmonary surfactant proteins (SPs). In order to confirm the role of sumoylated C/EBPα in BPD rats, SUMO1 was knocked down by lentiviral transfection of neonatal rat lungs with SUMO1‐RNAi‐LV. We found that the expression of C/EBPα and surfactant proteins increased following SUMO1 knockdown. Furthermore, the relatively low decrease in the levels of C/EBPα sumoylation was correlated with reduced glycogen consumption. Besides, co‐immunoprecipitation assays revealed that sumoylation is involved in the regulation of the interaction between C/EBPα and TGFβ2 in the lung. In conclusion, our findings indicate that sumoylation may act as a negative regulator of the C/EBPα‐mediated transactivation in BPD rats.

Pancreatic adenocarcinoma (PAAD) accounts for ~85% of all pancreatic cancer cases and is associated with a less favorable prognosis. Aberrant DNA methylation may influence the progression of PAAD by inducing abnormal gene expression. Methylation data of PAAD samples with prognosis information were obtained from The Cancer Genome Atlas (training set) and European Bioinformatics Institute Array Express databases (validation sets). Using the limma package, the differentially methylated genes in the training dataset were screened. Combined with the Weighted Gene Co-expression Network Analysis package, the co-methylated genes in key modules were identified. Then, a cor.test function in R software was applied to explore the functions of key the methylated genes. Correlation analyses of the expression levels and methylation levels of key methylated genes were performed, followed by identification of methylated genes associated with prognosis using Univariate Cox regression analysis. The optimal combination of prognosis related methylated genes was determined using a Cox-Proportional Hazards (Cox-PH) model. Subsequently, the risk score prognostic prediction system was constructed by combining the Cox-PH prognosis coefficients of the selected optimized genes. Based on the constructed risk score system, samples in all datasets were divided into high and low risk samples and the survival status was compared using survival curves. Furthermore, the correlation between independent prognostic factors and the risk score system was determined using the survival package. A total of 50 genes associated with prognosis of PAAD and a 12-gene optimal combination were obtained, including: CCAAT/enhancer binding protein α, histone cluster 1 H4E, STAM binding protein-like 1, phospholipase D3, centrosomal protein 55, ssDNA binding protein 4, glutamate AMPA receptor subunit 1, switch-associated protein 70, adenylate-cyclase activating polypeptide 1 receptor 1, yippee-like 3, homeobox C4 and insulin-like growth factor binding protein 1. Subsequently, a risk score prognostic prediction system of these 12 genes was constructed and validated. In addition, pathological N category, radiotherapy and risk status were identified as independent prognostic factors. Overall, the risk score prognostic prediction system constructed in the present study may be effective for predicting the prognosis of patients with PAAD.

St. Paul’s Wort (Siegesbeckia orientalis L.) confers anti-oxidative, anti-inflammatory, anti-allergic, anti-infertility, and immunosuppressive properties. Here, we elucidated whether high hydrostatic pressure extract of St. Paul’s Wort (SPW-HHPE) had anti-adipogenic activity. SPW-HHPE inhibited adipogenesis by reducing intracellular lipid accumulation. SPW-HHPE reduced the mRNA and protein expression of adipogenic regulatory factors [peroxisome proliferator-activated receptor gamma (PPARγ), CCAAT/enhancer binding protein alpha (C/EBPα), and sterol regulatory element binding protein-1c]. In addition, SPW-HHPE decreased the mRNA expression levels of lipogenic enzymes (fatty acid synthase and acetyl-CoA carboxylase) as well as adipocytokines (adiponectin and leptin). The inhibitory effect of SPW-HHPE on adipogenesis was mainly attributed to the enhancement of the Wnt/β-catenin signaling pathway. When β-catenin siRNA was transfected into 3T3-L1 adipocytes, the mRNA expression of PPARγ and C/EBPα was upregulated; however, their expression was attenuated by SPW-HHPE. These results suggest that SPW-HHPE suppresses adipogenesis by stimulating Wnt/β-catenin pathway.

Mulberry (Morus alba L.) fruits have long been used in traditional medicine and as edible berries in many countries. This study investigated the antiadipogenic effect of high hydrostatic pressure mulberry fruit extract (MFE) during 3T3-L1 adipocyte differentiation. MFE decreased lipid and triglyceride accumulation and glycerol-3-phosphate dehydrogenase activity. The mRNA expression levels of genes related to adipogenesis, such as the adipocyte protein 2, proliferator-activated receptor-γ, and CCAAT/enhancer binding protein-α, were suppressed by MFE. They also reduced microRNA (miR)-21 and miR-143 expression, which are involved in adipogenesis. In contrast, adenosine monophosphate-activated protein kinase (AMPK) activity was increased by MFE. These results suggested that MFE may suppress adipogenesis through modulating miR-21/143 expression and AMPK activity in 3T3-L1 adipocytes, which may be useful as antiobesity food agents.

Hazelnut proteins are an excellent source of bioactive peptides. Our previous study demonstrated that several novel peptides derived from Corylus heterophylla Fisch (C. heterophylla Fisch) have antioxidant and anti-inflammatory activities. In this study, we purified and identified anti-obesity peptides from hazelnut protein hydrolysates by chromatography and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Subsequently, we evaluated the inhibitory effect of the synthetic peptide on adipogenesis in 3T3-L1 adipocytes by Oil Red O staining, reverse transcription polymerase chain reaction (RT-PCR) and western blot. The results showed that a novel synthetic pentapeptide, Arg-Leu-Leu-Pro-His (RLLPH), derived from the C3 fraction, attenuated adipogenesis by downregulating the expression of several mRNAs related to adipogenesis, including peroxisome proliferator-activated receptor gamma (PPARγ), CCAAT/enhancer binding protein alpha (C/EBPα), adipocyte fatty acid-binding protein 2 (aP2), sterol regulatory element binding protein 1c (SREBP-1c), fatty acid synthase (FAS), acetyl-CoA carboxylase 1 (ACC1), and 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR). Moreover, RLLPH upregulated the levels of phosphorylated adenosine monophosphate-activated protein kinase (AMPK) and its substrate acetyl-CoA carboxylase (ACC) in 3T3-L1 adipocytes. A stability study revealed that RLLPH was relatively stable during pepsin digestion. These findings suggest that RLLPH has potential anti-obesity effects and may help combat metabolic diseases.

Changes in epigenetic DNA methylation are the most promising predictor of biological age and lifespan in humans, but whether methylation changes affect ageing is unresolved. Here, we discuss converging data, which indicate that one mode by which aberrant DNA methylation can affect ageing is via CCAAT/enhancer binding protein beta (C/EBPβ). This basic leucine-zipper (bZIP) transcription factor is controlled by the lifespan regulator mechanistic/mammalian target of rapamycin complex 1 (mTORC1) and plays an important role in energy homeostasis and adipose tissue differentiation. Emerging evidence indicates that access of C/EBPβ proteins to cognate binding sites is regulated by DNA demethylation via ten-eleven translocation (TET) methylcytosine dioxygenases and their adaptor proteins growth arrest and DNA damage-inducible protein 45 alpha (GADD45α) and inhibitor of growth 1 (ING1). We discuss the emerging causal nexus between C/EBPβ, energy metabolism, and DNA demethylation in organismal ageing.

Several studies have shown that Cl− channels regulate the differentiation of some cell types. Thus, we investigated the role of Cl− channels on adipocyte differentiation using adipose tissue-derived stem cells (ASCs) and Cl− channel blocker. We induced rabbit ASCs into adipocytes using Cl− channel blocker. The expression levels of adipocyte markers were no significant difference between the cells treated with a Cl− channel blocker NPPB and untreated cells. However, when the cells were treated with NPPB, lipid droplets (LDs) sizes decreased compared with the untreated control. Interestingly, the expression levels of Rab8a, which is known as a regulator of LD fusion, were also decreased in the cells treated with NPPB. Other Cl− channel blockers, DIDS and IAA-94, also inhibited large LDs formation and Rab8a expression. These results demonstrate that Cl− channels do not regulate the adipocyte differentiation, but do regulate the LDs formation via Rab8a expression. Abbreviations: ASCs: adipose tissue-derived stem cells; LDs: lipid droplets; RUNX2: runt-related transcription factor 2; CFTR: cystic fibrosis transmembrane conductance regulator; TG: triacylglycerol; FA: fatty acid; GLUT4: glucose transporter type 4; ER: endoplasmic reticulum; ADRP: adipose differentiation-related protein; TIP47: tail-interacting protein of 47 kD; HSL: hormone sensitive lipase; PBS: phosphate-buffered saline; DMEM: Dulbecco’s modified Eagle Medium; FBS: fetal bovine serum; SMA: smooth muscle actin; FAS: fatty acid synthase; ZONAB: ZO-1 associated nucleic acid binding protein; PPAR-γ: peroxisome proliferator-activated receptor-γ; C/EBPα: CCAAT/enhancer binding protein α; CE: cholesteryl ester; V-ATPase: vacuolar H+ ATPase.

Background: Long non-coding RNAs (lncRNAs) constitute an important component of the regulatory apparatus that controls stem cell pluripotency. However, the specific mechanisms utilized by these lncRNAs in the control of pluripotency are not fully characterized. Methods: We utilized a RNA reverse transcription-associated trap sequencing (RAT-seq) approach to profile the mouse genome-wide interaction targets for lncRNAs that are screened by RNA-seq. Results: We identified Peblr20 (Pou5F1 enhancer binding lncRNA 20) as a novel lncRNA that is associated with stem cell reprogramming. Peblr20 was differentially transcribed in fibroblasts compared to induced pluripotent stem cells (iPSCs). Notably, we found that Peblr20 utilized a trans mechanism to interact with the regulatory elements of multiple stemness genes. Using gain- and loss-of-function experiments, we showed that knockdown of Peblr20 caused iPSCs to exit from pluripotency, while overexpression of Peblr20 activated endogenous Pou5F1 expression. We further showed that Peblr20 promoted pluripotent reprogramming. Mechanistically, we demonstrated that Peblr20 activated endogenous Pou5F1 by binding to the Pou5F1 enhancer in trans, recruiting TET2 demethylase and activating the enhancer-transcribed RNAs. Conclusions: Our data reveal a novel epigenetic mechanism by which a lncRNA controls the fate of stem cells by trans-regulating the Pou5F1 enhancer RNA pathway. We demonstrate the potential for leveraging lncRNA biology to enhance the generation of stem cells for regenerative medicine.

Menin displays either tumor suppression or promotion functions in a context-dependent manner. Previously, we proposed that Menin acts as a tumor suppressor by inhibiting cell growth in pancreatic ductal adenocarcinoma (PDAC), whereas the relationship between the Menin expression and overall survival rate of PDAC patients has not been completely elucidated, indicating the complexity of Menin functions in PDAC progression. Here, we identify Menin as a promoter of epithelial-mesenchymal transition (EMT), which is largely associated with cell migration or metastasis, with modest activity in cell growth inhibition. Ectopic expression of Menin suppresses the expression of CCAAT/enhancer-binding protein beta (CEBPB) and epithelial-specific genes by histone deacetylation and further enhances the TGF-β signaling-related EMT process. We also demonstrate that CCAAT/enhancer binding protein (C/EBP) beta (C/EBPβ; encoded by CEBPB) acts downstream of Menin and TGF-β signaling for balancing growth inhibition and EMT, and C/EBPβ overexpression could restore the anti-cancer functions of Menin in pancreatic cancer by cooperatively activating CDKN2A/B genes and antagonizing EMT processes. Taken together, our results suggest that Menin functions as an oncogene for cancer metastasis upon C/EBPβ depletion or acts as a tumor suppressor by cooperation with C/EBPβ to activate CDKN2A transcription. Keywords: Menin, C/EBPβ, epithelial-mesenchymal transition, CDKN2A, pancreatic cancer

Organisms frequently encounter environments with nutrient shortages and their survival depends on changes in physiology and the ability to conserve resources. In bacteria, many physiological changes associated with starvation have been identified, but the underlying genetic components and regulatory networks that direct these physiological changes are often poorly defined. Here, we aimed to better define the gene regulatory networks that mediate the starvation response in Myxococcus xanthus, a bacterium that copes with starvation by producing fruiting bodies filled with dormant and stress-resistant spores. We focused on the direct promoter/gene targets of Nla28, a transcriptional activator/enhancer binding protein (EBP) that is important for early rounds of gene expression following starvation. Using expression profiling to identify genes that are downregulated in nla28 mutant cells and bioinformatics to identify the putative promoters of these genes, 12 potential promoter targets (37 genes) of Nla28 were identified. The results of in vitro promoter binding assays, coupled with in vitro and in vivo mutational analyses, suggested that the 12 promoters are in vivo targets of Nla28 and that Nla28 dimers use tandem, imperfect repeats of an 8-bp sequence for binding. Interestingly, nine of the Nla28 target promoters are intragenic, located in the protein coding sequence of an upstream gene or in the protein coding sequence of one gene within an operon (internal promoters). Based on mutational analyses, we concluded that the 12 Nla28 target loci contain at least one gene important for production of stress-resistant spores following starvation. Most of these loci contain genes predicted to be involved in regulatory or defense-related functions. Using the consensus Nla28 binding sequence, followed by bioinformatics and expression profiling, 58 additional promoters and 102 genes were tagged as potential Nla28 targets. Among these putative Nla28 targets, functions such as regulatory, metabolic and cell envelope biogenesis were commonly assigned to genes.

Obesity is a multifactorial chronic disease characterized by the excessive accumulation of fat in adipose tissue driven by hypertrophy and hyperplasia of adipocytes through adipogenesis. Adipogenesis plays a key role in the development of obesity and related metabolic disorders, which makes it potential target for the therapeutic approach to obesity. An increasing number of studies confirm the pleiotropic action of the combined treatment with metformin and statins, suggesting their anti-hypertensive, anti-inflammatory, and anti-adipogenic effect. The aim of this study was to analyze the effect of different doses of metformin (MET) and simvastatin (SIM) on the expression of key transcription factors of adipogenesis. Mouse 3T3-L1 preadipocytes were induced to differentiation in adipogenic medium with sustained MET and SIM treatment to assess the effect on adipogenesis. Nine days after initiating adipogenesis, the cells were prepared for further experiments, including Oil Red O staining, RT-PCR, Western blotting, and immunocytochemistry. Treating the cells with the combination of MET and SIM slightly reduced the intensity of Oil Red O staining compared with the control group, and down-regulated mRNA and protein expression of PPARγ, C/EBPα, and SREBP-1C. In conclusion, the inhibitory effect of MET and SIM on adipocyte differentiation, as indicated by decreased lipid accumulation, appears to be mediated through the down-regulation of adipogenic transcription factors, peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer binding pro-tein α (C/EBPα), and sterol regulatory element-binding protein 1 (SREBP-1C).

Patients with B-cell chronic lymphocytic leukemia (CLL) have an increased risk of severe infections due to disease- and treatment-related immunodeficiency. As a result, patients with hematologic malignancies have been given priority for primary COVID-19 vaccination. Unfortunately, many studies have suggested that patients with B-cell chronic lymphocytic leukemia (CLL) who have been fully vaccinated can develop severe and often fatal complications. Therefore, adjuvants that can induce mRNA vaccine efficacy are desperately needed for this category of patients with haematological malignancies. A recent, study by Oxford University scientists showed that leucine zipper transcription factor-like 1(LZTFL1), as a candidate causal gene and its enhancer the rs17713054 A risk allele was significantly responsible for the twofold increased risk of respiratory failure from COVID-19 associated with 3p21.31.By using sequence analysis, the risk allele generates a second CCAAT/enhancer binding protein beta (CEBPB) motif in the enhancer. Moreover, neither LZTFL1 variants found in T cells nor B cells are responsible for increasing death risk from COVID-19 infection according to oxford study. Here, we propose attestable hypothesis that trans retinoic acid could enhance the immune response in vaccinated patients with B-cell chronic lymphocytic leukemia (CLL) according to the recent findings of Oxford scientists by inducing the casual gene(LZTFL1) in CD4 T cells and inhibiting (CEBPB) motif.Conclusions Haematological malignancies (blood cancers) patients are more vulnerable to COVID-19 disuse severity and mortality. Un fortunately mRNA vaccine seem to be less effective with weak immune response and insufficient level of generated antibodies in this category of patients. Therefore we suggest all trans retinoic acid is a good candidate as mRNA COVID-19 vaccine adjuvant via inducing LZTFL1 gene in CD4Tcells and this activation could improve the immune response and increase the level of the generated antibodies. Moreover LZTFL1 gene in CD4 T cells is not associated with increasing risk of COVID-19 infection because of absence of its enhancer(The risk allele of the SNP, rs17713054 A) in immune cells according to oxford recent study. In addition to all trans retinoic acid could inhibit CCAAT/enhancer binding protein beta motif that is generated by The risk allele of the SNP, rs17713054 A

Glioma, which originates in the brain, is the most aggressive tumor of the central nervous system. It has been shown that microRNA (miRNA) controls the proliferation, migration and apoptosis of glioma cells. The objective of the present study was to measure microRNA-3148 (miR-3148) expression and investigate its impact on the pathogenetic mechanism of glioma. In the present study, reverse transcription-quantitative real-time PCR was employed to detect miR-3148 expression levels in glioma tissues and cell lines. Cell Counting Kit-8 assay, 5-ethynyl-2'-deoxyuridine assay, and Transwell migration assay were performed to assess the influence of miR-3148 on the malignant biological behavior of glioma cells. The biological functions of miR-3148 in glioma were examined via a xenograft tumor growth assay. Furthermore, the association between miR-3148 and DCUN1D1 was investigated via immunohistochemistry, dual-luciferase reporter assay and western blotting. It was observed that miR-3148 was expressed at low levels in glioma cells, and this represented a poor survival rate. In addition, an increased level of miR-3148 in cells and animal models inhibited glioma cell migration and proliferation. Moreover, miR-3148 decreased DCUN1D1 and curbed the nuclear factor κ enhancer binding protein (NF-κB) signaling pathway, thus decreasing the growth of glioma. Thus, miR-3148 is expressed within glioma tissues at low levels where it suppresses glioma by curbing the NF-κB pathway and lowering DCUN1D1.

Our previous work identified human immunodeficiency virus type I enhancer binding protein 1 (HIVEP1) as a putative driver of LPS-induced NF-κB signaling in humans in vivo. While HIVEP1 is known to interact with NF-ĸB binding DNA motifs, its function in mammalian cells is unknown. We report increased HIVEP1 mRNA expression in monocytes from patients with sepsis and monocytes stimulated by Toll-like receptor agonists and bacteria. In complementary overexpression and gene deletion experiments HIVEP1 was shown to inhibit NF-ĸB activity and induction of NF-ĸB responsive genes. RNA sequencing demonstrated profound transcriptomic changes in HIVEP1 deficient monocytic cells and transcription factor binding site analysis showed enrichment for κB site regions. HIVEP1 bound to the promoter regions of NF-ĸB responsive genes. Inhibition of cytokine production by HIVEP1 was confirmed in LPS-stimulated murine Hivep1-/- macrophages and HIVEP1 knockdown zebrafish exposed to the common sepsis pathogen Streptococcus pneumoniae. These results identify HIVEP1 as a negative regulator of NF-κB in monocytes/macrophages that inhibits proinflammatory reactions in response to bacterial agonists in vitro and in vivo., peer-reviewed