Author Correction: Molecular architecture of lineage allocation and tissue organization in early mouse embryo.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Author Correction: Molecular architecture of lineage allocation and tissue organization in early mouse embryo.

An Amendment to this paper has been published and can be accessed via a link at the top of the paper.

Transcription factor SlWRKY50 enhances cold tolerance in tomato by activating the jasmonic acid signaling.

Tomato (Solanum lycopersicum) is a cold-sensitive crop but frequently experiences low-temperature stimuli. However, tomato responses to cold stress are still poorly understood. Our previous studies have shown that using wild tomato (Solanum habrochaites) as rootstock can significantly enhance the cold resistance of grafted seedlings, in which a high concentration of jasmonic acids (JAs) in scions exerts an important role, but the mechanism of JA accumulation remains unclear. Herein, we discovered that tomato SlWRKY50, a Group II WRKY transcription factor that is cold inducible, responds to cold stimuli and plays a key role in JA biosynthesis. SlWRKY50 directly bound to the promoter of tomato allene oxide synthase gene (SlAOS), and overexpressing SlWRKY50 improved tomato chilling resistance, which led to higher levels of Fv/Fm, antioxidative enzymes, SlAOS expression, and JA accumulation. SlWRKY50-silenced plants, however, exhibited an opposite trend. Moreover, diethyldithiocarbamate acid (a JA biosynthesis inhibitor) foliar treatment drastically reduced the cold tolerance of SlWRKY50-overexpression plants to wild-type levels. Importantly, SlMYC2, the key regulator of the JA signaling pathway, can control SlWRKY50 expression. Overall, our research indicates that SlWRKY50 promotes cold tolerance by controlling JA biosynthesis and that JA signaling mediates SlWRKY50 expression via transcriptional activation by SlMYC2. Thus, this contributes to the genetic knowledge necessary for developing cold-resistant tomato varieties.

Molecular architecture of lineage allocation and tissue organization in early mouse embryo.

During post-implantation development of the mouse embryo, descendants of the inner cell mass in the early epiblast transit from the naive to primed pluripotent state1. Concurrently, germ layers are formed and cell lineages are specified, leading to the establishment of the blueprint for embryogenesis. Fate-mapping and lineage-analysis studies have revealed that cells in different regions of the germ layers acquire location-specific cell fates during gastrulation2-5. The regionalization of cell fates preceding the formation of the basic body plan-the mechanisms of which are instrumental for understanding embryonic programming and stem-cell-based translational study-is conserved in vertebrate embryos6-8. However, a genome-wide molecular annotation of lineage segregation and tissue architecture of the post-implantation embryo has yet to be undertaken. Here we report a spatially resolved transcriptome of cell populations at defined positions in the germ layers during development from pre- to late-gastrulation stages. This spatiotemporal transcriptome provides high-resolution digitized in situ gene-expression profiles, reveals the molecular genealogy of tissue lineages and defines the continuum of pluripotency states in time and space. The transcriptome further identifies the networks of molecular determinants that drive lineage specification and tissue patterning, supports a role of Hippo-Yap signalling in germ-layer development and reveals the contribution of visceral endoderm to the endoderm in the early mouse embryo.

LINC00115 promotes chemoresistant breast cancer stem-like cell stemness and metastasis through SETDB1/PLK3/HIF1α signaling.

BACKGROUND: Cancer stem-like cell is a key barrier for therapeutic resistance and metastasis in various cancers, including breast cancer, yet the underlying mechanisms are still elusive. Through a genome-wide lncRNA expression profiling, we identified that LINC00115 is robustly upregulated in chemoresistant breast cancer stem-like cells (BCSCs). METHODS: LncRNA microarray assay was performed to document abundance changes of lncRNAs in paclitaxel (PTX)-resistant MDA-MB-231 BCSC (ALDH+) and non-BCSC (ALDH-). RNA pull-down and RNA immunoprecipitation (RIP) assays were performed to determine the binding proteins of LINC00115. The clinical significance of the LINC00115 pathway was examined in TNBC metastatic lymph node tissues. The biological function of LINC00115 was investigated through gain- and loss-of-function studies. The molecular mechanism was explored through RNA sequencing, mass spectrometry, and the CRISPR/Cas9-knockout system. The therapeutic potential of LINC00115 was examined through xenograft animal models. RESULTS: LINC00115 functions as a scaffold lncRNA to link SETDB1 and PLK3, leading to enhanced SETDB1 methylation of PLK3 at both K106 and K200 in drug-resistant BCSC. PLK3 methylation decreases PLK3 phosphorylation of HIF1α and thereby increases HIF1α stability. HIF1α, in turn, upregulates ALKBH5 to reduce m6A modification of LINC00115, resulting in attenuated degradation of YTHDF2-dependent m6A-modified RNA and enhanced LINC00115 stability. Thus, this positive feedback loop provokes BCSC phenotypes and enhances chemoresistance and metastasis in triple-negative breast cancer. SETDB1 inhibitor TTD-IN with LINC00115 ASO sensitizes PTX-resistant cell response to chemotherapy in a xenograft animal model. Correlative expression of LINC00115, methylation PLK3, SETDB1, and HIF1α are prognostic for clinical triple-negative breast cancers. CONCLUSIONS: Our findings uncover LINC00115 as a critical regulator of BCSC and highlight targeting LINC00115 and SETDB1 as a potential therapeutic strategy for chemotherapeutic resistant breast cancer.

ZBTB7A interferes with the RPL5-P53 feedback loop and reduces endoplasmic reticulum stress-induced apoptosis of pancreatic cancer cells.

Endoplasmic reticulum (ER) stress is a primary mechanism leading to cell apoptosis, making it of great research interests in cancer management. This study delves into the function of ribosomal protein L5 (RPL5) in ER stress within pancreatic cancer (PCa) cells and investigates its regulatory mechanisms. Bioinformatics predictions pinpointed RPL5 as an ER stress-related gene exhibiting diminished expression in PCa. Indeed, RPL5 was found to be poorly expressed in PCa tissues and cells, with this reduced expression correlating with an unfavorable prognosis. Moreover, RPL5 overexpression led to heightened levels of p-PERK, p-eIF2α, and CHOP, bolstering the proapoptotic effect of Tunicamycin, an ER stress activator, on PCa cells. Additionally, the RPL5 overexpression curbed cell proliferation, migration, and invasion. Tunicamycin enhanced the binding between RPL5 and murine double minute 2 (MDM2), thus suppressing MDM2-mediated ubiquitination and degradation of P53. Consequently, P53 augmentation intensified ER stress, which further enhanced the binding between RPL5 and MDM2 through PERK-dependent eIF2α phosphorylation, thereby establishing a positive feedback loop. Zinc finger and BTB domain containing 7A (ZBTB7A), conspicuously overexpressed in PCa samples, repressed RPL5 transcription, thereby reducing P53 expression. Silencing of ZBTB7A heightened ER stress and subdued the malignant attributes of PCa cells, effects counteracted upon RPL5 silencing. Analogous outcomes were recapitulated in vivo employing a xenograft tumor mouse model, where ZBTB7A silencing dampened the tumorigenic potential of PCa cells, an effect reversed by additional RPL5 silencing. In conclusion, this study suggests that ZBTB7A represses RPL5 transcription, thus impeding the RPL5-P53 feedback loop and mitigating ER-induced apoptosis in PCa cells.

Gut microbial metabolite butyrate improves anticancer therapy by regulating intracellular calcium homeostasis.

BACKGROUND AND AIMS: Gut microbiota are recognized to be important for anticancer therapy, yet the underlying mechanism is not clear. Here, through the analysis of clinical samples, we identify the mechanism by which the gut microbial metabolite butyrate inhibits HCC and then explore new strategies for HCC treatment. APPROACH AND RESULTS: In our study, we demonstrate that gut microbial metabolite butyrate improves anticancer therapy efficacy by regulating intracellular calcium homeostasis. Using liquid chromatography-mass spectrometry analysis, we found that butyrate metabolism is activated in HCC patients compared with healthy individuals. Butyrate levels are lower in the plasma of HCC patients by gas chromatography-mass spectrometry (GC-MS) analysis. Butyrate supplementation or depletion of short-chain Acyl-CoA dehydrogenase (SCAD) gene (ACADS), encoding a key enzyme for butyrate metabolism, significantly inhibits HCC proliferation and metastasis. The profiling analysis of genes upregulated by butyrate supplementation or ACADS knockdown reveals that calcium signaling pathway is activated, leading to dysregulation of intracellular calcium homeostasis and production of reactive oxygen species. Butyrate supplementation improves the therapy efficacy of a tyrosine kinase inhibitor sorafenib. On the basis of these findings, we developed butyrate and sorafenib coencapsulated mPEG-PLGA-PLL nanoparticles coated with anti-GPC3 antibody (BS@PEAL-GPC3) to prolong the retention time of drugs and enhance drug targeting, leading to high anticancer efficacy. BS@PEAL-GPC3 nanoparticles significantly reduce HCC progression. In addition, BS@PEAL-GPC3 nanoparticles display excellent HCC targeting with excellent safety. CONCLUSIONS: In conclusion, our findings provide new insight into the mechanism by which the gut microbial metabolites inhibit HCC progression, suggesting a translatable therapeutics approach to enhance the clinical targeted therapeutic efficacy.

Regulation of cohesin-mediated chromosome folding by PDS5 in mammals.

Cohesin regulates sister chromatid cohesion but also contributes to chromosome folding by promoting the formation of chromatin loops, a process mediated by loop extrusion. Although PDS5 regulates cohesin dynamics on chromatin, the exact function of PDS5 in cohesin-mediated chromatin looping remains unclear. Two paralogs of PDS5 exist in vertebrates, PDS5A and PDS5B. Here we show that PDS5A and PDS5B co-localize with RAD21 and CTCF at loop anchors. Rapid PDS5A or PDS5B degradation in liver cancer cells using an inducible degron system reduces chromatin loops and increases loop size. RAD21 enrichment at loop anchors is decreased upon depletion of PDS5A or PDS5B. PDS5B loss also reduces CTCF signals at loop anchors and has a stronger effect on loop enlargement compared with PDS5A. Co-depletion of PDS5A and PDS5B reduces RAD21 levels at loop anchors although the amount of cohesin on chromatin is increased. Our study provides insight into how PDS5 proteins regulate cohesin-mediated chromatin looping.

USP7-mediated JUND suppresses RCAN2 transcription and elevates NFATC1 to enhance stem cell property in colorectal cancer.

Cancer stem cells (CSCs) encompass a subset of highly aggressive tumor cells that are involved in tumor initiation and progression. This study investigates the function of regulator of calcineurin 2 (RCAN2) in the stem cell property in colorectal cancer (CRC). By analyzing four GEO datasets, we obtained RCAN2 as a stemness-related gene in CRC. RCAN2 was poorly expressed in CRC tissues and cells, especially in CSCs. RCAN2 restoration reduced calcineurin activity and promoted phosphorylation and degradation of nuclear factor of activated T cells 1 (NFATC1) protein, leading to reduced stemness of CSCs. JunD proto-oncogene (JUND), whose protein level was increased in CRC samples and CRC stem cells, bound to RCAN2 and suppressed its transcription. The abundant ubiquitin specific peptidase 7 (USP7) in CSCs enhanced JUND protein stability through deubiquitination modification. Lentivirus-mediated knockdown of USP7 or JUND also blocked the calcineurin-NFATC1 signaling and reduced the protein levels of stemness-related proteins. Moreover, the USP7 knockdown weakened the colony/sphere formation ability as well as the tumorigenicity of CSCs, and it reduced the CSC content in xenograft tumors. However, further restoration of JUND rescued the stemness of the CSCs. Overall, this study demonstrates that USP7-mediated JUND suppresses RCAN2 transcription and activates NFATC1 to enhance stem cell property in CRC. 1. RCAN2 is poorly expressed in CRC tissues and cells and especially in CSCs. 2. RCAN2 reduces stemness of CSCs by blocking calcineurin-NFATC1 signal transduction. 3. JUND binds to RCAN2 promoter to suppresses RCAN2 transcription. 4. USP7 enhances JUND protein stability via deubiquitination modification. 5. Downregulation of USP7 or JUND restores RCAN2 level and suppresses stemness of CSCs.

Widespread roles of enhancer-like transposable elements in cell identity and long-range genomic interactions.

A few families of transposable elements (TEs) have been shown to evolve into cis-regulatory elements (CREs). Here, to extend these studies to all classes of TEs in the human genome, we identified widespread enhancer-like repeats (ELRs) and find that ELRs reliably mark cell identities, are enriched for lineage-specific master transcription factor binding sites, and are mostly primate-specific. In particular, elements of MIR and L2 TE families whose abundance co-evolved across chordate genomes, are found as ELRs in most human cell types examined. MIR and L2 elements frequently share long-range intra-chromosomal interactions and binding of physically interacting transcription factors. We validated that eight L2 and nine MIR elements function as enhancers in reporter assays, and among 20 MIR-L2 pairings, one MIR repressed and one boosted the enhancer activity of L2 elements. Our results reveal a previously unappreciated co-evolution and interaction between two TE families in shaping regulatory networks.

Mechanisms of linezolid resistance in Staphylococcus capitis with the novel mutation C2128T in the 23S rRNA gene in China.

PURPOSE: The objective of this study was to investigate the molecular characteristics and potential resistance mechanisms of linezolid-resistant (LZR) Staphylococcus capitis isolates from a tertiary hospital in China. METHODS: S. capitis isolates were obtained from clinical patient specimens; three of the isolates came from blood cultures and one from the hydrothorax. The agar dilution and E-test methods were used to identify antibiotic resistance. The chloramphenicol-florfenicol resistance (cfr) gene carrier status of the strains was determined by PCR. Whole-genome sequencing (WGS) was used to identify point mutations and L3, L4, and L22 mutations and to study the genetic environment of the cfr gene and the relationships between strains. RESULTS: The 4 isolates obtained in this study were all linezolid-resistant Staphylococcus strains. A similar of susceptibility profile pattern was observed in all four S. capitis strains, each of which exhibited a multidrug-resistant phenotype. A potentially novel mutation, C2128T, was identified, and the cfr genes of S. capitis strains were all positive. Additionally, the same mutations (C2128T and G2600T) were identified in all 23S rRNA sequences of the isolates, whereas mutations were lacking in the L3, L4, and L22 ribosomal proteins. The genetic environments surrounding cfr were identical in all four isolates. A schematic diagram of the phylogenetic tree showed that they were closely related to AYP1020, CR01, and TW2795, and a total of seven drug resistance genes were identified in these strains. CONCLUSIONS: The study indicated that the resistance of the Staphylococcus capitis strains to linezolid was caused by multiple mechanisms, and a potential novel mutation, C2128T, that may have an impact on bacterial resistance was identified.

Antimicrobial lanostane triterpenoids from the fruiting bodies of Ganoderma applanatum.

A chemical investigation on the 90% EtOH extract of the fruiting bodies of Ganoderma applanatum led to the isolation of three new lanostane triterpenoids, designated as 25-methoxy-11-oxo-ganoderiol D (1), 3-oxo-25-methoxy-24,26-dihydroxy-lanosta-7,9(11)-diene (2), and 3ß-acetyloxy-lucidone H (3). Structural elucidation of all the compounds were performed by spectral methods such as 1 D and 2 D (1H-1H COSY, HMQC, and HMBC) NMR spectroscopy. All the triterpenoids were in vitro evaluated for their antimicrobial activities against six pathogenic microorganisms. Compounds 1 and 2 exhibited some activities against three Gram positive bacteria with MIC values less than 60 µg/ml.

Accurate loop calling for 3D genomic data with cLoops.

MOTIVATION: Sequencing-based 3D genome mapping technologies can identify loops formed by interactions between regulatory elements hundreds of kilobases apart. Existing loop-calling tools are mostly restricted to a single data type, with accuracy dependent on a predefined resolution contact matrix or called peaks, and can have prohibitive hardware costs. RESULTS: Here, we introduce cLoops ('see loops') to address these limitations. cLoops is based on the clustering algorithm cDBSCAN that directly analyzes the paired-end tags (PETs) to find candidate loops and uses a permuted local background to estimate statistical significance. These two data-type-independent processes enable loops to be reliably identified for both sharp and broad peak data, including but not limited to ChIA-PET, Hi-C, HiChIP and Trac-looping data. Loops identified by cLoops showed much less distance-dependent bias and higher enrichment relative to local regions than existing tools. Altogether, cLoops improves accuracy of detecting of 3D-genomic loops from sequencing data, is versatile, flexible, efficient, and has modest hardware requirements. AVAILABILITY AND IMPLEMENTATION: cLoops with documentation and example data are freely available at: https://github.com/YaqiangCao/cLoops. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Inter-nucleosomal communication between histone modifications for nucleosome phasing.

Combinatorial effects of epigenetic modifications on transcription activity have been proposed as "histone codes". However, it is unclear whether there also exist inter-nucleosomal communications among epigenetic modifications at single nucleosome level, and if so, what functional roles they play. Meanwhile, how clear nucleosome patterns, such as nucleosome phasing and depletion, are formed at functional regions remains an intriguing enigma. To address these questions, we developed a Bayesian network model for interactions among different histone modifications across neighboring nucleosomes, based on the framework of dynamic Bayesian network (DBN). From this model, we found that robust inter-nucleosomal interactions exist around transcription start site (TSS), transcription termination sites (TTS) or around CTCF binding sites; and these inter-nucleosomal interactions are often involved in transcription regulation. In addition to these general principles, DBN also uncovered a novel specific epigenetic interaction between H2A.Z and H4K20me1 on neighboring nucleosomes, involved in nucleosome free region (NFR) and nucleosome phasing establishment or maintenance. The level of negative correlation between neighboring H2A.Z and H4K20me1 strongly correlate with the size of NFR and the strength of nucleosome phasing around TSS. Our study revealed inter-nucleosomal communications as important players in signal propagation, chromatin remodeling and transcription regulation.

An improved MLVF method and its comparison with traditional MLVF, spa typing, MLST/SCCmec and PFGE for the typing of methicillin-resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) has become an important nosocomial pathogen, causing considerable morbidity and mortality. During the last 20 years, a variety of genotyping methods have been introduced for screening the prevalence of MRSA. In this study, we developed and evaluated an improved approach capillary gel electrophoresis based multilocus variable-number tandem-repeat fingerprinting (CGE/MLVF) for rapid MRSA typing. A total of 42 well-characterized strains and 116 non-repetitive clinical MRSA isolates collected from six hospitals in northeast China between 2009 and 2010 were tested. The results obtained by CGE/MLVF against clinical isolates were compared with traditional MLVF, spa typing, Multilocus sequence typing/ staphylococcal cassette chromosome mec (MLST/SCCmec) and pulse field gel electrophoresis (PFGE). The discriminatory power estimated by Simpson's index of diversity was 0.855 (28 types), 0.855 (28 patterns), 0.623 (11 types), 0.517 (8 types) and 0.854 (28 patterns) for CGE/MLVF, traditional MLVF, spa typing, MLST/SCCmec and PFGE, respectively. All methods tested showed a satisfied concordance in clonal complex level calculated by adjusted Rand's coefficient. CGE/MLVF showed better reproducibility and accuracy than traditional MLVF and PFGE methods. In addition, the CGE/MLVF has potential to produce portable results. In conclusion, CGE/MLVF is a rapid and easy to use MRSA typing method with lower cost, good reproducibility and high discriminatory power for monitoring the outbreak and clonal spread of MRSA isolates.

KAP1 stabilizes MYCN mRNA and promotes neuroblastoma tumorigenicity by protecting the RNA m6A reader YTHDC1 protein degradation.

BACKGROUND: Neuroblastoma (NB) patients with amplified MYCN often face a grim prognosis and are resistant to existing therapies, yet MYCN protein is considered undruggable. KAP1 (also named TRIM28) plays a crucial role in multiple biological activities. This study aimed to investigate the relationship between KAP1 and MYCN in NB. METHODS: Transcriptome analyses and luciferase reporter assay identified that KAP1 was a downstream target of MYCN. The effects of KAP1 on cancer cell proliferation and colony formation were explored using the loss-of-function assays in vitro and in vivo. RNA stability detection was used to examine the influence of KAP1 on MYCN expression. The mechanisms of KAP1 to maintain MYCN mRNA stabilization were mainly investigated by mass spectrum, immunoprecipitation, RIP-qPCR, and western blotting. In addition, a xenograft mouse model was used to reveal the antitumor effect of STM2457 on NB. RESULTS: Here we identified KAP1 as a critical regulator of MYCN mRNA stability by protecting the RNA N6-methyladenosine (m6A) reader YTHDC1 protein degradation. KAP1 was highly expressed in clinical MYCN-amplified NB and was upregulated by MYCN. Reciprocally, KAP1 knockdown reduced MYCN mRNA stability and inhibited MYCN-amplified NB progression. Mechanistically, KAP1 regulated the stability of MYCN mRNA in an m6A-dependent manner. KAP1 formed a complex with YTHDC1 and RNA m6A writer METTL3 to regulate m6A-modified MYCN mRNA stability. KAP1 depletion decreased YTHDC1 protein stability and promoted MYCN mRNA degradation. Inhibiting MYCN mRNA m6A modification synergized with chemotherapy to restrain tumor progression in MYCN-amplified NB. CONCLUSIONS: Our research demonstrates that KAP1, transcriptionally activated by MYCN, forms a complex with YTHDC1 and METTL3, which in turn maintain the stabilization of MYCN mRNA in an m6A-dependent manner. Targeting m6A modification by STM2457, a small-molecule inhibitor of METTL3, could downregulate MYCN expression and attenuate tumor proliferation. This finding provides a new alternative putative therapeutic strategy for MYCN-amplified NB.

Pumpkin CmoDREB2A enhances salt tolerance of grafted cucumber through interaction with CmoNAC1 to regulate H2O2 and ABA signaling and K+/Na+ homeostasis.

Pumpkin CmoNAC1 enhances salt tolerance in grafted cucumbers. However, the potential interactions with other proteins that may co-regulate salt tolerance alongside CmoNAC1 have yet to be explored. In this study, we identified pumpkin CmoDREB2A as a pivotal transcription factor that interacts synergistically with CmoNAC1 in the co-regulation of salt tolerance. Both transcription factors were observed to bind to each other's promoters, forming a positive regulatory loop of their transcription. Knockout of CmoDREB2A in the root resulted in reduced salt tolerance in grafted cucumbers, whereas overexpression demonstrated the opposite effect. Multiple assays in our study provided evidence of the protein interaction between CmoDREB2A and CmoNAC1. Exploiting this interaction, CmoDREB2A facilitated the binding of CmoNAC1 to the promoters of CmoRBOHD1, CmoNCED6, CmoAKT1;2, and CmoHKT1;1, inducing H2O2 and ABA synthesis and increasing the K+/Na+ ratio in grafted cucumbers under salt stress. Additionally, CmoNAC1 also promoted the binding of CmoDREB2A to CmoHAK5;1/CmoHAK5;2 promoters, further contributing to the K+/Na+ homeostasis. In summary, these findings reveal a crucial mechanism of CmoNAC1 and CmoDREB2A forming a complex enhancing salt tolerance in grafted cucumbers.

TRIM24 Cooperates with Ras Mutation to Drive Glioma Progression through snoRNA Recruitment of PHAX and DNA-PKcs.

The factors driving glioma progression remain poorly understood. Here, the epigenetic regulator TRIM24 is identified as a driver of glioma progression, where TRIM24 overexpression promotes HRasV12 anaplastic astrocytoma (AA) progression into epithelioid GBM (Ep-GBM)-like tumors. Co-transfection of TRIM24 with HRasV12 also induces Ep-GBM-like transformation of human neural stem cells (hNSCs) with tumor protein p53 gene (TP53) knockdown. Furthermore, TRIM24 is highly expressed in clinical Ep-GBM specimens. Using single-cell RNA-sequencing (scRNA-Seq), the authors show that TRIM24 overexpression impacts both intratumoral heterogeneity and the tumor microenvironment. Mechanically, HRasV12 activates phosphorylated adaptor for RNA export (PHAX) and upregulates U3 small nucleolar RNAs (U3 snoRNAs) to recruit Ku-dependent DNA-dependent protein kinase catalytic subunit (DNA-PKcs). Overexpressed TRIM24 is also recruited by PHAX to U3 snoRNAs, thereby facilitating DNA-PKcs phosphorylation of TRIM24 at S767/768 residues. Phosphorylated TRIM24 induces epigenome and transcription factor network reprogramming and promotes Ep-GBM-like transformation. Targeting DNA-PKcs with the small molecule inhibitor NU7441 synergizes with temozolomide to reduce Ep-GBM tumorigenicity and prolong animal survival. These findings provide new insights into the epigenetic regulation of Ep-GBM-like transformation and suggest a potential therapeutic strategy for patients with Ep-GBM.

Myeloid Deletion of Cdc42 Protects Liver From Hepatic Ischemia-Reperfusion Injury via Inhibiting Macrophage-Mediated Inflammation in Mice.

BACKGROUND & AIMS: Hepatic ischemia-reperfusion injury (HIRI) often occurs in liver surgery, such as partial hepatectomy and liver transplantation, in which myeloid macrophage-mediated inflammation plays a critical role. Cell division cycle 42 (Cdc42) regulates cell migration, cytoskeleton rearrangement, and cell polarity. In this study, we explore the role of myeloid Cdc42 in HIRI. METHODS: Mouse HIRI models were established with 1-hour ischemia followed by 12-hour reperfusion in myeloid Cdc42 knockout (Cdc42mye) and Cdc42flox mice. Myeloid-derived macrophages were traced with RosamTmG fluorescent reporter under LyzCre-mediated excision. The experiments for serum or hepatic enzymic activities, histologic and immunologic analysis, gene expressions, flow cytometry analysis, and cytokine antibody array were performed. RESULTS: Myeloid deletion of Cdc42 significantly alleviated hepatic damages with the reduction of hepatic necrosis and inflammation, and reserved hepatic functions following HIRI in mice. Myeloid Cdc42 deficiency suppressed the infiltration of myeloid macrophages, reduced the secretion of proinflammatory cytokines, restrained M1 polarization, and promoted M2 polarization of myeloid macrophages in livers. In addition, inactivation of Cdc42 promoted M2 polarization via suppressing the phosphorylation of STAT1 and promoting phosphorylation of STAT3 and STAT6 in myeloid macrophages. Furthermore, pretreatment with Cdc42 inhibitor, ML141, also protected mice from hepatic ischemia-reperfusion injury. CONCLUSIONS: Inhibition or deletion of myeloid Cdc42 protects liver from HIRI via restraining the infiltration of myeloid macrophages, suppressing proinflammatory response, and promoting M2 polarization in macrophages.

Icariin potentiates the antitumor activity of gemcitabine in gallbladder cancer by suppressing NF-κB.

AIM: Gemcitabine has been increasingly prescribed for the treatment of gallbladder cancer. However, the response rate is low. The aim of this study is to determine whether icariin, a flavonoid isolated from Epimedi herba, could potentiate the antitumor activity of gemcitabine in gallbladder cancer. METHODS: Human gallbladder carcinoma cell lines GBC-SD and SGC-996 were tested. Cell proliferation and apoptosis were analyzed using MTT assay and flow cytometry, respectively. The expression of apoptosis- and proliferation-related molecules was detected with Western blotting. Caspase-3 activity was analyzed using colorimetric assay, and NF-κB activity was measured with ELISA. A gallbladder cancer xenograft model was established in female BALB/c (nu/nu) mice. The mice were intraperitoneally administered gemcitabine (125 mg/kg) in combination with icariin (40 mg/kg) for 2 weeks. RESULTS: Icariin (40-160 µg/mL) dose-dependently suppressed cell proliferation and induced apoptosis in both GBC-SD and SGC-996 cells, with SGC-996 cells being less sensitive to the drug. Icariin (40 µg/mL) significantly enhanced the antitumor activity of gemcitabine (0.5 µmol/L) in both GBC-SD and SGC-996 cells. The mice bearing gallbladder cancer xenograft treated with gemcitabine in combination with icariin exhibited significantly smaller tumor size than the mice treated with either drug alone. In GBC-SD cells, icariin significantly inhibited both the constitutive and gemcitabine-induced NF-κB activity, enhanced caspase-3 activity, induced G(0)-G(1) phase arrest, and suppressed the expression of Bcl-2, Bcl-xL and surviving proteins. CONCLUSION: Icariin, by suppressing NF-κB activity, exerts antitumor activity, and potentiates the antitumor activity of gemcitabine in gallbladder cancer. Combined administration of gemcitabine and icariin may offer a better therapeutic option for the patients with gallbladder cancer.