Architectural protein subclasses shape 3D organization of genomes during lineage commitment.

Understanding the topological configurations of chromatin may reveal valuable insights into how the genome and epigenome act in concert to control cell fate during development. Here, we generate high-resolution architecture maps across seven genomic loci in embryonic stem cells and neural progenitor cells. We observe a hierarchy of 3D interactions that undergo marked reorganization at the submegabase scale during differentiation. Distinct combinations of CCCTC-binding factor (CTCF), Mediator, and cohesin show widespread enrichment in chromatin interactions at different length scales. CTCF/cohesin anchor long-range constitutive interactions that might form the topological basis for invariant subdomains. Conversely, Mediator/cohesin bridge short-range enhancer-promoter interactions within and between larger subdomains. Knockdown of Smc1 or Med12 in embryonic stem cells results in disruption of spatial architecture and downregulation of genes found in cohesin-mediated interactions. We conclude that cell-type-specific chromatin organization occurs at the submegabase scale and that architectural proteins shape the genome in hierarchical length scales.

Two forms of loops generate the chromatin conformation of the immunoglobulin heavy-chain gene locus.

The immunoglobulin heavy-chain (IgH) gene locus undergoes radial repositioning within the nucleus and locus contraction in preparation for gene recombination. We demonstrate that IgH locus conformation involves two levels of chromosomal compaction. At the first level, the locus folds into several multilooped domains. One such domain at the 3' end of the locus requires an enhancer, Eµ; two other domains at the 5' end are Eµ independent. At the second level, these domains are brought into spatial proximity by Eµ-dependent interactions with specific sites within the V(H) region. Eµ is also required for radial repositioning of IgH alleles, indicating its essential role in large-scale chromosomal movements in developing lymphocytes. Our observations provide a comprehensive view of the conformation of IgH alleles in pro-B cells and the mechanisms by which it is established.

The SET oncoprotein promotes estrogen-induced transcription by facilitating establishment of active chromatin.

SET is a multifunctional histone-binding oncoprotein that regulates transcription by an unclear mechanism. Here we show that SET enhances estrogen-dependent transcription. SET knockdown abrogates transcription of estrogen-responsive genes and their enhancer RNAs. In response to 17ß-estradiol (E2), SET binds to the estrogen receptor α (ERα) and is recruited to ERα-bound enhancers and promoters at estrogen response elements (EREs). SET functions as a histone H2 chaperone that dynamically associates with H2A.Z via its acidic C-terminal domain and promotes H2A.Z incorporation, ERα, MLL1, and KDM3A loading and modulates histone methylation at EREs. SET depletion diminishes recruitment of condensin complexes to EREs and impairs E2-dependent enhancer-promoter looping. Thus, SET boosts E2-induced gene expression by establishing an active chromatin structure at ERα-bound enhancers and promoters, which is essential for transcriptional activation.

Heat-shock protein 90α protects NME1 against degradation and suppresses metastasis of breast cancer.

BACKGROUND: NME1 has been exploited as a potential translational target for decades. Substantial efforts have been made to upregulate the expression of NME1 and restore its anti-metastasis function in metastatic cancer. METHODS: Cycloheximide (CHX) chase assay was used to measure the steady-state protein stability of NME1 and HSP90α. The NME1-associating proteins were identified by immunoprecipitation combined with mass spectrometric analysis. Gene knockdown and overexpression were employed to examine the impact of HSP90AA1 on intracellular NME1 degradation. The motility and invasiveness of breast cancer cells were examined in vitro using wound healing and transwell invasion assays. The orthotopic spontaneous metastasis and intra-venous experimental metastasis assays were used to test the formation of metastasis in vivo, respectively. RESULTS: HSP90α interacts with NME1 and increases NME1 lifetime by impeding its ubiquitin-proteasome-mediated degradation. HSP90α overexpression significantly inhibits the metastatic potential of breast cancer cells in vitro and in vivo. A novel cell-permeable peptide, OPT22 successfully mimics the HSP90α function and prolongs the life span of endogenous NME1, resulting in reduced metastasis of breast cancer. CONCLUSION: These results not only reveal a new mechanism of NME1 degradation but also pave the way for the development of new and effective approaches to metastatic cancer therapy.

Identification of two galectin-4 proteins (PcGal4-L and PcGal4-L-CRD) and their function in AMP expression in Procambarus clarkii.

Galectins, a family of lectins that bind to ß-galactoside, possess conserved carbohydrate recognition domains (CRDs) and play a crucial role in recognizing and eliminating pathogens in invertebrates. Two galectin-4 genes (PcGal4) isoforms, named PcGal4-L and PcGal4-L-CRD, were cloned from the cDNA library of Procambarus clarkia in our study. PcGal4-L contains an open reading frame (ORF, 1089 bp), which encodes a protein consisting of 362 amino acids including a single CRD and six low complexity regions. The full-length cDNA of PcGal4-L-CRD contains a 483 bp ORF that encodes a protein of 160 amino acids, with a single CRD and a low-complexity region. The difference between the two PcGal4 isoforms is that PcGal4-L has 202 additional amino acids after the CRD compared to the PcGal4-L-CRD. These two isoforms are grouped together with other galectins from crustaceans through phylogenetic analysis. Further study revealed that total PcGal4 (including PcGal4-L and PcGal4-L-CRD) was primarily expressed in the muscle, gills and intestine. The mRNA levels of total PcGal4 in gills and hemocytes were significantly induced after challenge with Aeromonas hydrophila. Both recombinant PcGal4-L and its spliced isoform, PcGal4-L-CRD, could directly bind to lipopolysaccharides, peptidoglycan and five tested microorganisms, inducing a wide spectrum of microbial agglutination. The spliced isoform PcGal4-L-CRD showed a stronger binding ability than PcGal4-L. In addition, when the PcGal4 was knockdown, transcriptions of seven antimicrobial peptides (AMPs) genes (ALF5, ALF6, ALF8, CRU1, CRU2, CRU3 and CRU4) in gills and seven AMPs genes (ALF5, ALF6, ALF8, ALF9, CRU1, CRU3 and CRU4) in hemocytes were significantly decreased. Meanwhile, the survival rate of P. clarkii decreased in the PcGal4-dsRNA group. In summary, these results indicate that PcGal4 can mediate the innate immunity in P. clarkii by bacterial recognition and agglutination, as well as regulating AMP expression, thus recognition and understanding of the functions of galectin in crustaceans in immune resistance.

Oncoprotein SET dynamically regulates cellular stress response through nucleocytoplasmic transport in breast cancer.

SETß is the predominant isoform of oncoprotein SE translocation (SET) in various breast cancer cell lines. Interactome-transcriptome analysis has shown that SETß is intimately associated with cellular stress response. Among various exogenous stimuli, formaldehyde (FA) causes distinct biological effects in a dose-dependent manner. In response to FA at different concentrations, SET dynamically shuttles between the nucleus and cytoplasm, performing diverse biofunctions to restore homeostasis. At a low concentration, FA acts as an epidermal growth factor (EGF) and activates the HER2 receptor and downstream signaling pathways in HER2+ breast cancer cells, resulting in enhanced cell proliferation. Nucleocytoplasmic transport of SETß is controlled by the PI3K/PKCα/CK2α axis and depletion or blockade of the transport of SETß suppresses EGF-induced activation of AKT and ERK. SETß also inhibits not only stress-induced activation of p38 MAPK signaling pathway, but also assembly of stress granules by hindering formation of the G3BP1-RNA complex. Our findings suggest that SET functions as an important regulator which modulates cellular stress signaling pathways dynamically.

A structural hierarchy mediated by multiple nuclear factors establishes IgH locus conformation.

Conformation of antigen receptor gene loci spatially juxtaposes rearranging gene segments in the appropriate cell lineage and developmental stage. We describe a three-step pathway that establishes the structure of the 2.8-Mb immunoglobulin heavy chain gene (IgH) locus in pro-B cells. Each step uses a different transcription factor and leads to increasing levels of structural organization. CTCF mediates one level of compaction that folds the locus into several 250- to 400-kb subdomains, and Pax5 further compacts the 2-Mb region that encodes variable (VH) gene segments. The 5' and 3' domains are brought together by the transcription factor YY1 to establish the configuration within which gene recombination initiates. Such stepwise mechanisms may apply more generally to establish regulatory fine structure within megabase-sized topologically associated domains.

Dynamics of estrogen-induced ROS and DNA strand break generation in estrogen receptor α-positive breast cancer.

DNA repair machinery is involved in estrogen-dependent transactivation. Mounting evidence suggests that mechanisms underlying estrogen-induced DNA damage are complicated. To date estrogen-induced DNA oxidation and its impact on ERα-mediated transaction remains ambiguous. Herein, we found that the process of 17ß-estradiol (E2)-induced ROS production can be approximately divided into two phases according to responding time and generation mechanisms. The intracellular Ca2+ fluctuation and ERα-dependent transcription lead to temporospatially different oxidative DNA damage. Further, we demonstrate that DNA oxidation is dispensable for estrogen-responsive gene expression. Dynamics of estrogen-induced DNA strand break generation also show two-phase pattern and topoisomerase-mediated DNA stand breaks are essential in estrogen signaling. Collectively, our findings have provided new insights into oxidative DNA damage in estrogen signaling.

A hook wire dislodged into the subglottic area as a rare complication following computed tomography-guided hook wire localization: a case report.

BACKGROUND: Computed tomography-guided hook wire localization (CT-GHWL) was used to localize the small pulmonary nodules before video-assisted thoracic surgery (VATS). Its associated complications included hook wire dislodgement, pulmonary hemorrhage, and pneumothorax. This is the first report of a patient with a hook wire sliding into the subglottic area after CT-GHWL. CASE PRESENTATION: A 27-year-old female had productive cough for 8 days. A high-resolution CT scan showed a 12 mm part-solid nodule in the number 8 segment of the left lung. Prior to VATS, she received CT-GHWL to localize the nodule. During VATS, the hook wire unexpectedly slid away. A chest computed tomography was immediately performed and the sagittal reconstructed images showed the needle at the subglottic area. Finally, the needle was extracted by biopsy forceps under bronchoscope evaluation. The patient was eventually recovered and discharged. CONCLUSIONS: Dislodge of the hook wire into the subglottic area is an extremely rare but serious complication following CT-GHWL. Attention should be paid to securing the needle on the lung surface during VATS.

An improved cell line-derived xenograft humanized mouse model for evaluation of PD-1/PD-L1 blocker BMS202-induced immune responses in colorectal cancer.

The establishment of an in vivo mouse model mimicking human tumor-immune environments provides a promising platform for immunotherapy assessment, drug discovery and clinical decision guidance. To this end, we construct humanized NCG mice by transplanting human hCD34 + hematopoietic progenitors into non-obese diabetic (NOD) Cg- Prkdc scidIL2rg tm1Wjl /Sz (null; NCG) mice and monitoring the development of human hematopoietic and immune systems (Hu-NCG). The cell line-derived xenograft (CDX) Hu-NCG mouse models are set up to assess the outcome of immunotherapy mediated by the small molecule BMS202. As a PD-1/PD-L1 blocker, BMS202 shows satisfactory antitumour efficacy in the HCT116 and SW480 xenograft Hu-NCG mouse models. Mechanistically, BMS202 exerts antitumour efficacy by improving the tumor microenvironment and enhancing the infiltration of hCD8 + T cells and the release of hIFNγ in tumor tissue. Thus, tumor-bearing Hu-NCG mice are a suitable and important in vivo model for preclinical study, particularly in cancer immunotherapy.

Association of PD-L1 expression status with the efficacy of PD-1/PD-L1 inhibitors and overall survival in solid tumours: A systematic review and meta-analysis.

Whether PD-L1-positive patients derive more overall survival benefit from PD-1/PD-L1 inhibitors in the treatment of advanced solid tumours is unclear. We systematically searched the PubMed, Cochrane library and EMBASE databases from January 1, 1966 to March 1, 2019, to identify randomised controlled trials of PD-1/PD-L1 inhibitors (nivolumab, pembrolizumab, atezolizumab, durvalumab and avelumab) that had available hazard ratios (HRs) for death according to PD-L1 status. A random-effects model was used to calculate the pooled overall survival (OS) HR and 95% CI among PD-L1-positive and PD-L1-negative patients. An interaction test was performed to evaluate the heterogeneity between the two estimates. A total of 24 randomised trials, involving 12,966 participants, fulfilled the inclusion criteria. An OS benefit of PD-1/PD-L1 inhibitors was found in both PD-L1-positive patients (HR, 0.65; 95% CI, 0.60-0.70) and PD-L1-negative patients (HR, 0.82; 95% CI, 0.74-0.91) even at the minimum cut-off value of 1%. Significant differences in the efficacy of PD-1/PD-L1 inhibitors between PD-L1-positive and PD-L1-negative patients were noted at different cut-off values. Moreover, there was a positive dose-response relationship between PD-L1 positivity and OS benefit (HR for 1%, 0.58, [0.50, 0.67]; 5%, 0.52 [0.43, 0.64]; 10%, 0.50 [0.40, 0.63]). Subgroup analyses showed that these results were generally consistent, regardless of study design, line of treatment, treatment type, tumour type, PD-L1 staining cell type and median follow-up time. We demonstrated that PD-1/PD-L1 inhibitors significantly improved OS in both PD-L1 positive and PD-L1 negative patients compared to controls, but the magnitude of benefit was clinically PD-L1-dependent.

Nicaraven Attenuates Postoperative Systemic Inflammatory Responses-Induced Tumor Metastasis.

BACKGROUND: Inflammation has been demonstrated to promote cancer metastasis. Due to the well-known systemic inflammatory responses (SIR) after major surgery, it is critical to investigate and attenuate SIR-induced tumor metastasis of cancer patients suffering surgical procedures. METHODS: C57BL/6 mice were intravenously injected with Lewis lung cancer cells at 6, 24, and 72 h after the induction of intestinal ischemia/reperfusion (I/R) injury. We found that the number of tumor nodules significantly increased in lungs of mice injected with cancer cells at 6 h but not at 24 and 72 h after I/R injury. The administration of nicaraven 30 min before and 24 h after I/R injury effectively attenuated the enhanced tumor metastasis to lungs. Protein array showed the increase of various cytokines in plasma of mice at 6 h after I/R injury, but many of them were attenuated by the administration of nicaraven. Immunostaining indicated the increase of Ly6g-, CD206-, and CD11c-positive inflammatory cells in the lungs, but it was also attenuated by nicaraven administration. CONCLUSIONS: Postoperative SIR-induced tumor metastasis have been clearly evidenced in our experimental model, and the administration of nicaraven may ameliorate the SIR-induced tumor metastasis by suppressing inflammatory responses.

Comparison of outcomes of pedicled jejunal and colonic conduit for esophageal reconstruction.

BACKGROUND: At present, the gastric tube is the first choice for esophageal reconstruction after esophagectomy for various benign and malignant diseases. However, when the stomach is not available, a pedicled jejunum or colon is used to reconstruct the esophagus. The present study aimed to compare the postoperative outcomes and quality of life of patients receiving jejunal and colonic conduits. METHODS: In the present retrospective study, the clinical data of 71 patients with esophageal carcinoma, who received jejunal reconstruction (jejunum group, n = 34) and colonic reconstruction (colon group, n = 37) from 2005 to 2015, were compared. RESULTS: Compared with the colon group, the jejunum group had a lower incidence of postoperative anastomotic leakage, lesser duration of postoperative drainage, and faster recovery. Furthermore, the scores were better in the jejunum group than in the colon group, in terms of short-term overall quality of life, physical function and social relationships. Moreover, the jejunal group had a significantly lower frequency of pH < 4 simultaneous reflux time > 5 min (N45) and the longest reflux time (LT) at 24 weeks after surgery. CONCLUSION: In esophageal cancer, when gastric tube construction is not feasible, a pedicled jejunum may be preferred over a colonic conduit due to lower incidence of acid reflux, anastomotic leakage and higher postoperative short-term quality of life, and rapid postoperative recovery.

The prognostic value of PKM2 and its correlation with tumour cell PD-L1 in lung adenocarcinoma.

BACKGROUND: The prognostic value of PKM2 and its correlation with tumour cell PD-L1 in lung adenocarcinoma (LUAD) is unclear. METHODS: A total of 506 lung adenocarcinoma samples from The Cancer Genome Atlas (TCGA) dataset and 173 LUAD tumour tissues from Jiangxi Cancer Hospital were used to analyse the correlation between PKM2 and PD-L1 expression. We further established a stable LUAD cell line with PKM2 knockdown and confirmed the association via Western blotting and flow cytometry analysis. Moreover, the prognostic values of PKM2 and PD-L1 were evaluated by the Kaplan-Meier method and Cox proportional hazards models. RESULTS: Based on the above two large cohorts, we found that PKM2 was significantly positively associated with PD-L1 expression (r = 0.132, P = 0.003 and r = 0.287, P < 0.001, respectively). Subsequently, we found that PKM2 knockdown substantially inhibited PD-L1 expression in the A549 LUAD cell line. Moreover, survival analysis showed that higher expression of PKM2 was correlated with significantly shorter overall survival (OS) and disease-free survival (DFS) in lung adenocarcinoma patients (P < 0.001 and P = 0.050, respectively). Subgroup analysis showed that lung adenocarcinoma patients who expressed high PKM2 and PD-L1 levels experienced the poorest OS and DFS. Additionally, multivariate analysis suggested that high PKM2 and PD-L1 expression was an independent prognostic indicator for worse OS and DFS (HR = 1.462, P < 0.001 and HR = 1.436, P = 0.004, respectively). CONCLUSIONS: Our results demonstrated that PKM2 regulated PD-L1 expression and was associated with poor outcomes in lung adenocarcinoma patients.

Thoracic tumor resection combined with SVC replacement using autologous pericardium.

BACKGROUND: Invasion of the superior vena cava (SVC) by thoracic tumors and occurrence of SVC syndrome are often encountered in clinical practice; but the prognosis in these cases is poor. Replacement of the SVC with autologous pericardial tissue is rarely performed. In this study, we sought to investigate the postoperative outcomes of this rare procedure. METHODS: We performed a retrospective analysis of six patients who underwent SVC replacement using autologous pericardial tissue between October 2010 and November 2016. We collected data on the patients' pathological features, operative characteristics, postoperative outcomes, and survival. RESULTS: All six patients were male with an average age of 52 years (range, 18-62 years). Three of the patients had lung cancer, one had stage III thymoma, and two had germinoma. Four of the six patients had mild or moderate superior vena cava compression and no corresponding clinical symptoms. The other two patients had severe compression and obvious symptoms of SVC syndrome, with the typical swelling of the face, eyelids, and upper extremities. All six patients underwent complete tumor resection, with two of the lung cancer patients undergoing right lobectomy and one undergoing right pneumonectomy. With respect to the postoperative outcomes, one patient died, whereas the others did not develop any major complications. At the end of the follow-up period, five of the patients were alive and none of the patients had developed thrombosis in the grafts. CONCLUSIONS: Our findings indicated that SVC replacement with autologous pericardium is technically feasible and safe, with few postoperative complications and favorable long-term effects. Although it has some limitations, this method appears to be useful in achieving SVC reconstruction of moderate size. SVC replacement with autologous pericardium appears to have the potential for widespread clinical use.

Analysis of Volatile Compounds in Pears by HS-SPME-GC×GC-TOFMS.

Aroma plays an important role in fruit quality and varies among different fruit cultivars. In this study, a sensitive and accurate method based on headspace solid-phase microextraction (HS-SPME) coupled with comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry (GC×GC-TOFMS) was developed to comprehensively compare aroma components of five pear cultivars. In total, 241 volatile compounds were identified and the predominant volatile compounds were esters (101 compounds), followed by alcohols (20 compounds) and aldehydes (28 compounds). The longyuanyangli has the highest relative concentration (838.12 ng/g), while the Packham has the lowest (208.45 ng/g). This study provides a practical method for pear aroma analysis using SPME and GC×GC-TOFMS.

Robust Variable Selection and Estimation Based on Kernel Modal Regression.

Model-free variable selection has attracted increasing interest recently due to its flexibility in algorithmic design and outstanding performance in real-world applications. However, most of the existing statistical methods are formulated under the mean square error (MSE) criterion, and susceptible to non-Gaussian noise and outliers. As the MSE criterion requires the data to satisfy Gaussian noise condition, it potentially hampers the effectiveness of model-free methods in complex circumstances. To circumvent this issue, we present a new model-free variable selection algorithm by integrating kernel modal regression and gradient-based variable identification together. The derived modal regression estimator is related closely to information theoretic learning under the maximum correntropy criterion, and assures algorithmic robustness to complex noise by replacing learning of the conditional mean with the conditional mode. The gradient information of estimator offers a model-free metric to screen the key variables. In theory, we investigate the theoretical foundations of our new model on generalization-bound and variable selection consistency. In applications, the effectiveness of the proposed method is verified by data experiments.

Effects of γ-Irradiation on the Molecular Structures and Functions of Human Serum Albumin.

In this paper, we use spectroscopic methods (fluorescence spectroscopy, UV absorption spectroscopy, and circular dichroism (CD) spectroscopy) to elucidate the effects of reactive oxygen species generated by γ-irradiation on the molecular properties of human serum albumin (HSA). The results of fluorescence spectroscopy indicated that oxidation by γ-irradiation can lead to conformational changes of HSA. Data of CD spectra suggested that with the increase of radiation dose the percentage of α-helix in HSA has decreased. The determination of protein hydrophobicity showed that the effective hydrophobicity of HSA decreased up to 62% compared to the native HSA solution due to the exposure to the γ-irradiation. Furthermore, small changes in the esterase-like activity of HSA were introduced because of oxidation. The content of bityrosine increased markedly, suggesting that the oxidized HSA was aggregated. Moreover, there was no obvious change in the molecular properties of HSA with low γ-irradiation dose. Changes happened when the irradiation dose exceeded 200 Gy.

Photoluminescence properties of a new orange-red-emitting Sm(3+)-La3SbO7 phosphor.

The antimonate compound La3SbO7 has high chemical stability, lattice stiffness and thermal stability. Orange-red-emitting antimonate-based phosphors La3SbO7:xSm(3+) (x = 0.02, 0.05, 0.08, 0.10, 0.15, 0.20 and 0.25) were synthesized. The phase structure and photoluminescence properties of these phosphors were investigated. The emission spectrum obtained on excitation at 407 nm contained exclusively the characteristic emissions of Sm(3+) at 568, 608, 654 and 716 nm, which correspond to the transitions from (4)G5/2 to (6)H5/2, (6)H7/2, (6)H9/2 and (6)H11/2 of Sm(3+), respectively. The strongest emission was located at 608 nm due to the (4)G5/2→(6)H7/2 transition of Sm(3+), generating bright orange-red light. The critical quenching concentration of Sm(3+) in La3SbO7:Sm(3+) phosphor was determined as 10% and the energy transfer between Sm(3+) was found to be through an exchange interaction. The International Commission on Illumination chromaticity coordinates of the La3SbO7:0.10Sm(3+) phosphors are located in the orange-red region. The La3SbO7:Sm(3+) phosphors may be potentially used as red phosphors for white light-emitting diodes.

The potential benefits of nicaraven to protect against radiation-induced injury in hematopoietic stem/progenitor cells with relative low dose exposures.

Nicaraven, a hydroxyl radical-specific scavenger has been demonstrated to attenuate radiation injury in hematopoietic stem cells with 5Gy γ-ray exposures. We explored the effect and related mechanisms of nicaraven for protecting radiation injury induced by sequential exposures to a relatively lower dose γ-ray. C57BL/6 mice were given nicaraven or placebo within 30min before exposure to 50mGy γ-ray daily for 30days in sequences (cumulative dose of 1.5Gy). Mice were victimized 24h after the last radiation exposure, and the number, function and oxidative stress of hematopoietic stem cells were quantitatively estimated. We also compared the gene expression in these purified stem cells from mice received nicaraven and placebo treatment. Nicaraven increased the number of c-kit(+) stem/progenitor cells in bone marrow and peripheral blood, with a recovery rate around 60-90% of age-matched non-irradiated healthy mice. The potency of colony forming from hematopoietic stem/progenitor cells as indicator of function was completely protected with nicaraven treatment. Furthermore, nicaraven treatment changed the expression of many genes associated to DNA repair, inflammatory response, and immunomodulation in c-kit(+) stem/progenitor cells. Nicaraven effectively protected against damages of hematopoietic stem/progenitor cells induced by sequential exposures to a relatively low dose radiation, via complex mechanisms.