Inhibition of apoptosis by oncogenic hepatitis B virus X protein: Implications for the treatment of hepatocellular carcinoma

Hepatitis B virus X protein(HBx) plays an important role in the development of hepatocellular carcinoma(HCC). In addition, hepatoma upregulated protein(HURP) is a cellular oncogene that is upregulated in a majority of HCC cases. We highlight here recent findings demonstrating a link between HBx, HURP and anti-apoptosis effects observed in cisplatin-treated HCC cells. We observed that Hep3B cells overexpressing HBx display increased HURP mRNA and protein levels, and show resistance to cisplatin-induced apoptosis. Knockdown of HURP in HBx-expressing cells reverses this effect, and sensitizes cells to cisplatin. The anti-apoptotic effect of HBx requires activation of the p38/MAPK pathway as well as expression of SATB1, survivin and HURP. Furthermore, silencing of HURP using short-hairpin RNA promotes accumulation of p53 and reduces cell proliferation in SK-Hep-1 cells(p53^(+/–)), whereas these effects are not observed in p53-mutant Mahlavu cells. Similarly, HURP silencing does not affect the proliferation of H1299 lung carcinoma cells or Hep3 B HCC cells which lack p53. Silencing of HURP sensitizes SK-Hep-1 cells to cisplatin. While HURP overexpression promotes p53 ubiquitination and degradation by the proteasome, HURP silencing reverses these effects. Inoculation of SK-Hep-1 cancer cells in which HURP has been silenced produces smaller tumors than control in nude mice. Besides, gankyrin, a positive regulator of the E3 ubiquitin ligase MDM2, is upregulated following HURP expression, and silencing of gankyrin reduces HURP-mediated downregulation of p53. In addition, we observed a positive correlation between HURP and gankyrin protein levels in HCC patients(r^2 = 0.778; n = 9). These findings suggest a role for the viral protein HBx and the host protein HURP in preventing p53-mediated apoptosis during cancer progression and establishment of chemoresistance.

Expression characteristics and diagnostic value of annexin A2 in hepatocellular carcinoma

AIM： TO investigate the characteristics and diagnostic value of annexin A2 （ANXA2） expression in cancerous tissues and sera of patients with hepatitis B virus （HBV）-related hepatocellular carcinoma （HCC）. METHODS： Levels of liver ANXA2 gene transcription or protein expression were analyzed in HCC-, their self- controlled precancerous-, and distant cancerous- tissues from 30 HCC. Serum levels of ANXA2 expression in 115 patients with HCC, 25 with metastatic liver can cer, 35 with chronic hepatitis, 28 with acute hepatitis, 38 with cirrhosis, and 30 healthy controls were deter- mined. Clinicopathological characteristics of circulating ANXA2 expression were analyzed, and its diagnostic efficiency and clinical values in HCC were evaluated. RESULTS： ANXA2 expression was localized in both cell membrane and cytoplasm in HCC tissue, mainly in the cytoplasm of matched adjacent cancerous tissue, and there was almost no positive staining in matched distant cancerous tissue. Abnormal expression of liver ANXA2 was present in HCC tissues compared with self-con- trolled adjacent- and distant-cancerous tissues at pro- tein or mRNA level. Circulating ANXA2 in HCC patients was significantly higher than that of other liver diseases （P 〈 0.01） except metastatic liver cancer. If the diag- nostic cutoff value of ANXA2 level was more than 18 ng/ mL, the incidence of serum ANXA2 was 86.96% in the HCC group, 80% in the metastatic liver cancer group, 31.58% in the liver cirrhosis group, none in the chronic hepatitis or acute hepatitis or normal control group, respectively. Serum ANXA2 expression in HCC patients was correlated with HBV infection （27.38 ± 5.67 ng/mL vs 18.58 ± 7.83 ng/mL, P 〈 0.01）, extrahepatic metas- tasis （26.11±5.43 ng/mL ys 22.79 ± 5.64 ng/mL, P 〈 0.01）, and portal vein thrombus （26.03 ± 5.99 ng/mL vs 23.06 ± 5.03 ng/mL, P 〈 0.01）, and was significantly higher （P 〈 0.01） in the moderately- （26.19±5.34 ng/ mL） or the poorly- differentiated group （27.05 ± 5.13 ng/mL） than in the well differentiated group （20.43 ± 4.97 ng/mL）, and in the tumor node metastasis stages Ⅲ-Ⅳ（P 〈 0.01） than in stages Ⅰ-Ⅱ. ANXA2 was not correlated with patient sex, age, size or α-fetoprotein （AFP） level. Area under the receiver operating charac- teristic curve for the whole range of sensitivities and specificities was 0.796 for ANXA2 and 0.782 for AFP. Combining detection of serum ANXA2 and AFP substan- tially improved the diagnostic efficiency （96.52%） and the neclative predictive value （＇96.61%） for HCC.of ANXA2 expression has good diagnostic potential for HCC diagnosis.

Mechanoresponsive Gene Upregulation by Force Depends on H3K9Demethylation

All living cells in a human body are made of the same DNA molecule but cells in different tissues express different genes and proteins.How the transcription process is controlled and regulated is largely unknown.Specifically,mechanical forces are increasingly recognized to play critical roles in cell and tissue functions.However,what controls force-induced gene transcription is elusive.Recently we have reported that a local surface force transfers from integrins to the cytoskeleton and the link of nucleoskeleton and the cytoskeleton(LINC)into the nucleus and deforms chromatin directly to induce rapid activation of transgene DHFR.Here we show that endogenous mechanoresponsive genes egr-1 and Cav1 are rapidly upregulated and their upregulation depends on stress angles relative to the cell long axis,suggesting direct impact of these genes by force.Demethylation of histone 3 at lysine 9(H3K9)trimethylation(H3K9me3)at nuclear interiors(euchromatin)is necessary for force-induced transcription upregulation.Our findings suggest that force-rapid upregulation of mechanoresponsive genes by force depends on H3K9me3 demethylation.

A Simulation for Flavivirus Infection Decoy Responses

In this work, we discuss the development of simulation code for a model of the cross-reactive adaptive immune response seen in flavivirus infections. The model specifically addresses flavivirus pathogen virulence in G0?vs G1?cell states. The MHC-I upregulation of resting cells (G0 state) allows the T-cells generated for flavivirus peptide antigens to attack healthy cells also. The cells in G1?state are not upregulated as much and so virus hides in them and hence is propagated upon rupture. Hence, this type of model is referred to as a decoy model because the immune system is decoyed into preferentially recognizing the upregulated cells while the virus actively propagates in another small, but important, cell population. We show that the generic assumption of upregulation via a model which includes the?G0/G1?differential upregulation leads to immunopathological consequences. We outline the details behind the simulation code decisions and provide some theoretical justification for our model of collateral damage and upregulation.

Using a Collateral Damage Model to Explain Survival Data in West Nile Virus Infections

Simulation code for a model of the adaptive immune response seen in flavivirus infections is used to explain the immunopathological consequences seen in West Nile Virus virus (WNV) infections. We use a model that specifically handles the differences in how the virus infects resting cells, the G0 state, versus dividing cells, the G1 state, which includes vastly increased MHC-I upregulation for resting cells over dividing cells. The simulation suggests how the infection progresses in a one host model and the results shed insight into the unusual survival curve data obtained for this infection: there is an increase in health even though viral load has increased.

Upregulation of GLE1 and LCP2 Genes in H5N1 Influenza Virus Infected Patients

Previous study showed that the Gle1 RNA export mediator-like (Gle1l) gene and the lymphocyte cytosolic protein 2 (Lcp2) gene were upregulated in response to influenza virus A/Puerto Rico/8/1934 (H1N1) in a mouse mode. To determine whether these two genes were upregulated in humans after influenza A virus infection, nasopharyngeal swabs were collected from eleven patients with flu-like symptoms for viral RNA extraction and PCR amplification. Sequencing analysis revealed that nucleoprotein (NP) gene fragments amplified from nasopharyngeal swabs of four patients shared the highest similarity with the NP gene from avian influenza A (H5N1) virus (A/ goose/Shantou/753/2002). Peripheral blood samples were then collected from four patients for quantitative analysis of GLE1 and LCP2 gene expression. Our results demonstrated that both GLE1 and LCP2 genes were upregulated in H5N1 influenza A virus infected patients, suggesting that upregulation of GLE1 and LCP2 genes may be important for the host defense against influenza A viruses.

Cryptotanshinone attenuates isoprenaline-induced cardiac fibrosis in the mouse associated with upregulation and activation of matrix metalloproteinase-2

Objectives The impairment of matrix metallopro- teinase-2(MMP-2)has been associated with the development of cardiac fibrosis.Although the Chinese herb Salvia miltior-rhiza has been widely used in patients with cardiovascular disorders,the mechanisms involved have not been elucidated. The purpose of the present study was to determine whether the administration of cryptotanshinone,an active ingredient of Salvia miltiorrhiza,could prevent the cardiac fibrosis induced by isoprenaline and to investigate the underlying mechanisms. Methods and Results Male C57BL/6 mice were submitted to receive daily injection of 0.9%saline,3 mg/kg isoprenaline, or isoprenaline plus 20 mg/kg cryptotanshinone by gastric gavage for 2 weeks.Herein,we demonstrate that cryptotanshinone can significantly ameliorate the isoprenaline-induced cardiac fibrosis,which was associated with marked up-regulation and activation of MMP-2 in ventricular myocardium. Additionally,we demonstrate that cryptotanshinone can dose-dependently upregulate and activate MMP-2 in cultured cardiac fibroblast.Moreover,incubation with cryptotanshinone also can prevent isoprenaline-induced downregulation and inactivation of MMP-2 in cultured cardiac fibroblast. Conclusions Taken together,our data suggest that cryptotanshinone may become a novel and potent antifibrotic agent. The present findings might further our understanding of the role of MMP-2 in cardiac fibrosis and antifibrotic mechanisms of cryptotanshinone.

A novel alkaline phosphatase positive granule in rat neutrophils

The localization of the alkaline phosphatase (ALPase) activity in isolated rat neutrophils was ultrastructurally observed to further study the function and mechanism of the neutrophils in defending the body against invading microbes. Methods: The rat neutrophils were isolated from the whole blood and peritoneum. The ALPase cytochemical and ultrastructural observation were performed in the research. Results: It was found that ALPase-positive granules were localized in both rat blood and peritoneal neutrophils,which were different from the traditional azurophil and the special granules, and located in cytoplasm in the forms of some novel vacuoles and slender rod-shaped structures. These structures were easily upregulated by phorbol 12-myristate 13-acetate (PMA ) and/or N-formyl--fen--phe (fMLP) treatment, and shifted toward the surface of the cells. The upregulation was easily observed 2. 5 min after the stimulation with PMA. Most of the alkaline phosphatase-positive granules fused with each other at first, and some of them were connected with the cell surface plasma membrane at 15 min after the stimulation with PMA or fMLP. Conclusion: A novel alkaline phosphatase - positive granule exists in isolated rat neutrophils and may originate from the Golgi complexes of the neutrophils. The activity of the granules relates to at least activation of protein kinase C.

通过USP7结合分子胶上调P53蛋白

Molecular glues are typically small chemical molecules that act at the interface between a target protein and degradation machinery to trigger ternary complex formation.Identifying molecular glues is challenging.There is a scarcity of target-specific upregulating molecular glues,which are highly anticipated for numerous targets,including P53.P53 is degraded in proteasomes through polyubiquitination by specific E3 ligases,whereas deubiquitinases(DUBs)remove polyubiquitination conjugates to counteract these E3ligases.Thus,small-molecular glues that enhance P53 anchoring to DUBs may stabilize P53 through deubiquitination.Here,using small-molecule microarray-based technology and unbiased screening,we identified three potential molecular glues that may tether P53 to the DUB,USP7,and elevate the P53 level.Among the molecular glues,bromocriptine(BC)is an FDA-approved drug with the most robust effects.BC was further verified to increase P53 stability via the predicted molecular glue mechanism engaging USP7.Consistent with P53 upregulation in cancer cells,BC was shown to inhibit the proliferation of cancer cells in vitro and suppress tumor growth in a xenograft model.In summary,we established a potential screening platform and identified potential molecular glues upregulating P53.Similar strategies could be applied to the identification of other types of molecular glues that may benefit drug discovery and chemical biology studies.

Efficient and multiplex gene upregulation in plants through CRISPR-Cas-mediated knockin of enhancers

Geneupregulation through genome editing is important for plant research and breeding.Targeted insertion of short transcriptional enhancers(STEs)into gene promoters may offer a universal solution akin to transgene-mediated overexpression while avoiding the drawbacks associated with transgenesis.Here,we introduce an“in locus activation”technique in rice that leverages well-characterized STEs for refined,heritable,and multiplexed gene upregulation.To address the scarcity of potent enhancers,we developed a large-scale mining approach and discovered a suite of STEs that are capable of enhancing gene expression in rice protoplasts.The in locus integration of these STEs into eight rice genes resulted in substantial tran-scriptional upregulation in the edited plants,with up to 869.1-fold increases in their transcript levels.Em-ploying a variety of STEs,we achieved delicate control of gene expression,enabling the fine-tuning of key phenotypic traits such as plant height.Our approach also enabled efficient multiplexed gene upregu-lation,with up to four genes activated simultaneously,significantly enhancing the nicotinamide mononucleotide metabolic pathway.Importantly,heritability studies from the To to T3 generations confirmed the stable and heritable nature of STE-driven gene activation.Collectively,our work demon-strates that coupled with STE mining,leveraging genome editing for in locus activation and gene upregu-lation holds great promise to be widely adopted in fundamental plant research and crop breeding.

Upregulation of MiR-1280 Expression in Non-small Cell Lung Cancer Tissues

Background：Non-small cell lung cancer （NSCLC） is a prolific and high-mortality disease with few effective treatments.Although the detection and surgical techniques for NSCLC continue to advance,the survival rate for the patients with NSCLC remains poor.Enhanced predictive biomarkers such as microRNAs （miRNAs） are needed at the time of diagnosis to better tailor therapies for patients.This study focused on the expression ofmiR-1280 in NSCLC tissues and distal normal tissues in order to explore the association between miR-1280 expression and NSCLC.Methods：A total of 72 newly diagnosed primary NSCLC patients were enrolled in this study.Quantitative real-time polymerase chain reaction （PCR） was performed to identify the expression level ofmiR-1280 in the NSCLC tissues and distal normal tissues of these patients.Results：The miR-1280 expression was significantly higher in the NSCLC tissues （0.084 ± 0.099） than distal normal tissues （0.014 ± 0.015,P =0.009）.In 54 patients （75%）,the miR-1280 expression in the NSCLC tissues was upregulated （2-△△ct 〉 2）,and no case showed a downregulation of miR-1280 expression.Conclusions：The expression level ofmiR-1280 could be regarded as a biomarker for NSCLC.

Upregulation of Divalent Metal Transporter 1 (DMT1) Is Involved in Amyloid Precursor Protein Processing and Aβ Generation

The amyloid beta precursor protein (APP) and its pathogenic byproduct β-amyloid peptide (Aβ) play central roles in the pathogenesis of Alzheimer’s disease (AD). Reduction in

Mitogen-activated protein kinase signal pathways play an important role in right ventricular hypertrophy of tetralogy of Fallot

Background Tetralogy of Fallot （TOF） is the most common malformation of children with an incidence of approximately 10% of congenital heart disease patients. There can be a wide spectrum to the severity of the anatomic defects, which include ventricular septal defect, aortic override, right ventricular outflow tract obstruction, and right ventricular hypertrophy. We examined the relationship between right ventricular hypertrophy in patients with TOF and the gene expression of factors in the mitogen-activated protein kinase （MAPK） signal pathway. Methods To gain insight into the characteristic gene（s） involved in molecular mechanisms of right ventricular hypertrophy in TOF, differential mRNA and micro RNA expression profiles were assessed using expression-based micro array technology on right ventricular biopsies from young TOF patients who underwent primary correction and on normal heart tissue. We then analyzed the gene expression of the MAPK signal pathway using reverse transcription-polymerase chain reaction （RT-PCR） in normals and TOF patients. Results Using the micro RNA chip V3.0 and human whole genome oligonucleotide microarray VI.0 to detect the gene expression, we found 1068 genes showing altered expression of at least two-fold in TOF patients compared to the normal hearts, and 47 micro RNAs that showed a significant difference of at least two-fold in TOF patients. We then analyzed these mRNAs and micro RNAs by target gene predicting software Microcosm Targets version 5.0, and determined those mRNA highly relevant to the right ventricular hypertrophy by RT-PCR method. There were obvious differences in the gene expression of factors in the MAPK signal pathway when using RT-PCR, which was consistent to the results of the cDNA microarray.Conclusion The upregulation of genes in the MAPK signal pathway may be the key events that contribute to right ventricular hypertrophy and stunted angiogenesis in patients with TOF.

NF-κB,a hot topic in biochemical and medical studies in China

In 1986, nuclear factor (NF)-κB was first discovered in the laboratory of Nobel Prize laureate David Baltimore via its interaction with a sequence (5′-GGGACTTTCC-3′) in the immunoglobulin light-chain enhancer in B

New insights into different surfactants’impacts on sludge fermentation:Focusing on the particular metabolic processes and microbial genetic traits

Surfactants were expected to exhibit positive effects on the waste activated sludge(WAS)disposal.However,the systematic comparison of different categories of surfactants on the WAS fermentation and the functional mechanisms,especially microbial metabolic traits,have not yet been precisely explored.This study revealed the positive effects of different surfactants on the volatile fatty acid(VFA)production,which followed the order of alkyl polysaccharides(APG)>sodium dodecylbenzene sulfonate(SDBS)>hexadecyl trimethyl ammonium bromide(HTAB).Mechanistic exploration found that the presence of different surfactants improved solubilization and hydrolysis steps,and then contributed to the subsequent acidification with different efficiencies.The functional microorganisms associated with VFA generation were enriched in surfactant-conditioned reactors.Metagenomic analysis further indicated that the key genes involved in the particular process of VFA generation were over-expressed.The simultaneous bioavailable substrate improvement,functional bacterial enrichment,and metabolic activity upregulation induced by different surfactants jointly contributed to VFA promotion during WAS fermentation.This study could provide a comprehensive realization of surfactants’impacts on theWAS fermentation process,and more importantly,it reminded the public to discern the distinct interplaying effects induced by different chemicals in regulating the WAS disposal and resource recovery.

Upregulation of PIP3-Dependent Rac Exchanger 1(P-Rex1) Promotes Prostate Cancer Metastasis

Excessive activation of G-protein coupled receptor (GPCR) and receptor tyrosine kinase (RTK) pathways has been linked to prostate cancer metastasis. Rac activation

The long noncoding RNA lnc-ob1 facilitates bone formation by upregulating Osterix in osteoblasts

With the support by the National Natural Science Foundation of China,the research team directed by Prof.Wang XiaoGang(王晓刚)at the Beijing Advanced Innovation Center for Big Data-Based Precision Medicine,Beihang University,recently reported that lncRNA regulates osteoblast activity and bone formation in mice by up-regulating the osteogenic transcription factor Osterix,which was published in Nature Metabolism(2019,1:485—496).

Toward fine-tuned metabolic networks in industrial microorganisms

There are numerous microorganisms in nature capable of synthesizing diverse useful compounds;however,these natural microorganisms are generally inefficient in the production of target products on an industrial scale,relative to either chemical synthesis or extraction methods.To achieve industrial production of useful compounds,these natural microorganisms must undergo a certain degree of mutation or effective fine-tuning strategies.This review describes how to achieve an ideal metabolic fine-tuned process,including static control strategies and dynamic control strategies.The static control strategies mainly focus on various matabolic engineering strategies,including protein engineering,upregulation/downregulation,and combinatrorial control of these metabolic engineering strategies,to enhance the flexibility of their application in fine-tuned metabolic metworks.Then,we focus on the dynamic control strategies for fine-tuned metabolic metworks.The design principles derived would guide us to construct microbial cell factories for various useful compounds.

X chromosome inactivation and active X upregulation in therian mammals: facts, questions, and hypotheses

X chromosome inactivation is a mechanism that modulates the expression of X-linked genes in eutherian females(XX).Ohno proposed that to achieve a proper balance between X-linked and autosomal genes,those on the active X should also undergo a 2-fold upregula-tion.Although some support for Ohno's hypothesis has been provided through the years,recent genomic studies testing this hypoth-esis have brought contradictory results and fueled debate.Thus far,there are as many results in favor as against Ohno's hypothesis,depending on the nature of the datasets and the various assumptions and thresholds involved in the analyses.However,they have con-firmed the importance of dosage balance between X-linked and autosomal genes involved in stoichiometric relationships.These facts as well as questions and hypotheses are discussed below.

Notch signaling inhibitor and anti-PD-L1 antibody combination therapies decelerate tumor progression in pancreatic cancer

Objective:Pancreatic cancer(PC)is a highly lethal malignancy with an immunosuppressive environment.Yet,current immune checkpoint inhibitor monotherapies have shown limited efficacy in PC,prompting the need for combination therapies.Herein,we hypothesized that combinations of Notch signaling inhibitor and anti-ligand programmed death-ligand 1(PD-L1)antibody immunotherapy would show synergistic efficacy.Methods:The baseline expression of PD-L1 and HES1 was measured in PC cell lines,single-cell RNA-seq data of PC(GSA:CRA001160),and cBioPortal databases.In an in vitro study,MIA PaCa2 and SW1990 were used to explore the mechanism between Notch signaling and PD-L1.To study the effects in vivo,a subcutaneous tumor model was established using Pan02 cells treated with either anti-PD-L1 monoclonal antibody and/or Notch inhibitor DAPT.The study performed involving human samples was approved by the Ethics Committee of Peking Union Medical College Hospital(approval No.S-K460,approval date:April 23,2018).Animal studies were approved by the Animal Research Ethics Committee of Peking Union Medical College Hospital(approval No.XHDW-2019-049,approval date:November 28,2019).Results:The Notch signaling inhibitor upregulated PD-L1 expression in PC tumor cells both in vitro and in vivo.Notch effector HES1 knockdown produced PD-L1 upregulation in both MIA PaCa2 and SW1990 cells.Combined DAPT and anti-PD-L1 antibody treatment of Pan02 subcutaneous tumor model resulted in significantly reduced tumor weights compared to that with monotherapy,as well as significantly reduced Ki67 than that in the monotherapy group and control group.Flow cytometry analysis revealed significantly increased CD8+T cell infiltration in tumors of the combination group compared with those of the monotherapy group.Conclusion:Notch signaling blockade might enhance the antitumor effect of anti-PD-L1 therapy in PC.