Biology subjects

Zhao, L.

Publications and source records attributed to Zhao, L..

403 records · Page 23Linked to original sources

bioRxiv2017

Harnessing the Cross-talk between Tumor Cells and Tumor-associated Macrophages with a Nano-drug for modulation of Glioblastoma Immune Microenvironment

Glioblastoma (GBM) is the most frequent and malignant brain tumor with a high mortality rate. The presence of a large population of macrophages (M{varphi}) in the tumor microenvironment is a prominent feature of GBM and these so-called tumor-associated M{varphi} (TAM) closely interact with the GBM cells to promote the survival, progression and therapy resistance of the GBM. Various therapeutic strategies have been devised either targeting the GBM cells or the TAM but few have addressed the cross-talks between the two cell populations. The present study was carried out to explore the possibility of exploiting the cross-talks between the GBM cells (GC) and TAM for modulation of the GBM microenvironment through using Nano-DOX, a drug composite based on nanodiamonds bearing doxorubicin. In the in vitro work on human cell models, Nano-DOX-loaded TAM were first shown to be viable and able to infiltrate three-dimensional GC spheroids and release cargo drug therein. GC were then demonstrated to encourage Nano-DOX-loaded TAM to unload Nano-DOX back into GC which consequently emitted damage-associated molecular patterns (DAMPs) that are powerful immunostimulatory agents as well as indicators of cell damage. Nano-DOX was next proven to be a more potent inducer of GC DAMPs emission than doxorubicin. As a result, Nano-DOX-damaged GC exhibited an enhanced ability to attract both TAM and Nano-DOX-loaded TAM. Most remarkably, Nano-DOX-damaged GC reprogrammed the TAM from a pro-GBM phenotype to an anti-GBM phenotype that suppressed GC growth. Finally, the in vivo relevance of the in vitro findings was tested in animal study. Mice bearing orthotopic human GBM xenografts were intravenously injected with Nano-DOX-loaded mouse TAM which were found releasing drug in the GBM xenografts 24 h after injection. GC damage was evidenced by the induction of DAMPs emission within the xenografts and a shift of TAM phenotype was detected as well. Taken together, our results demonstrate a novel way with therapeutic potential to harness the cross-talk between GBM cells and TAM for modulation of the tumor immune microenvironment.\n\nAbbreviationsATP, adenosine triphosphate; BBB, blood-brain barrier; BCA, bicinchoninic acid; BMDM, bone marrow derived macrophages; CD, cluster of differentiation; CFSE, 5(6)-carboxyfluorescein diacetate, succinimidyl ester; CM, conditioned culture medium; CNS, central nervous system; CRT, calreticulin; DAMPs, damage-associated molecular patterns; DAB, diaminobenzidine; DOX, doxorubicin; ECL, enhanced chemiluminescence; ELISA, enzyme-linked immunosorbent assay; HMGB1, high mobility group protein B1; HSP90, heat shock protein 90; FACS, flow cytometry; GBM, glioblastoma; Guanylate Binding Protein 5 (GBP5); GC, glioblastoma cells; IHC, immunohistochemical; IL, interleukin; M{varphi}, macrophages; mBMDM, mouse BMDM; mBMDM2, Type-2 mBMDM; M1, Type-1 Mo; M2, Type-2 Mo; Nano-DOX, ND-PG-RGD-DOX; ND, nanodiamonds; Nano-DOX-mBMDM, Nano-DOX-loaded mouse BMDM; NGCM, Nano-DOX-treated-GC-conditioned medium; PBS, phosphate buffered saline; PG, polyglycerol; PMA, phorbol 12-myristate 13-acetate; PVDF, polyvinylidene fluoride; RGD, tripeptide of L-arginine, glycine and L-aspartic acid; RM, regular culture medium; SD, standard deviation; TAM, tumor-associated M{varphi}; TBST, Tris Buffered Saline with Tween(R) 20.\n\nGraphic abstract\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=153 SRC=\"FIGDIR/small/170282_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (111K):\norg.highwire.dtl.DTLVardef@180aea2org.highwire.dtl.DTLVardef@14922f7org.highwire.dtl.DTLVardef@96a696org.highwire.dtl.DTLVardef@92f050_HPS_FORMAT_FIGEXP M_FIG C_FIG

pharmacology and toxicology↗

bioRxiv2017

Resequencing the Escherichia coli genome by GenoCare single molecule sequencing platform

Next generation sequencing (NGS) has revolutionized life sciences research. Recently, a new class of third-generation sequencing platforms has arrived to meet increasing demands in the clinic, capable of directly measuring DNA and RNA sequences at the single-molecule level without amplification. Here, we use the new GenoCare single molecule sequencing platform from Direct Genomics to resequence the E. coli genome and show comparable performance to the Illumina MiSeq system. Our platform detects single-molecule fluorescence by total internal reflection microscopy, with sequencing-by-synthesis chemistry. With a consensus sequence of 99.71% nucleotide identity to that of the Illumina MiSeq systems, GenoCare was determined to be a reliable platform for single-molecule sequencing, with strong potential for clinical applications.

genomics↗

bioRxiv2017

Identification of Rab18 as an Essential Host Factor for BKPyV Infection Using a Whole Genome RNA Interference Screen

BK polyomavirus (BKPyV) is a human pathogen first isolated in 1971. BKPyV infection is ubiquitous in the human population, with over 80% of adults worldwide being seropositive for BKPyV. BKPyV infection is usually asymptomatic; however, BKPyV reactivation in immunosuppressed transplant patients causes two diseases, polyomavirus-associated nephropathy and hemorrhagic cystitis. To establish a successful infection in its host cells, BKPyV must travel in retrograde transport vesicles to reach the nuclei. To make this happen, BKPyV requires the cooperation of host cell proteins. To further identify host factors associated with BKPyV entry and intracellular trafficking, we performed a whole-genome siRNA screen on BKPyV infection of primary human renal proximal tubular epithelial cells. The results revealed the importance of the Ras-related protein Rab18 and syntaxin 18 for BKPyV infection. Our subsequent experiments implicated additional factors that interact with this pathway, and suggest a more detailed model of the intracellular trafficking process, indicating that BKPyV reaches the ER lumen through a retrograde transport pathway between the late endosome and the ER.

microbiology↗

bioRxiv2017

Near-Atomic Resolution Structure Determination in Over-Focus with Volta Phase Plate by Cs-corrected Cryo-EM

Volta phase plate (VPP) is a recently developed transmission electron microscope (TEM) apparatus that can significantly enhance the image contrast of biological samples in cryo-electron microscopy (cryo-EM) therefore impose the possibility to solve structures of relatively small macromolecules at high resolution. In this work, we performed theoretical analysis and found that using phase plate on objective lens spherical aberration (Cs)-corrected TEM may gain some interesting optical properties, including the over-focus imaging of macromolecules. We subsequently evaluated the imaging strategy of frozen-hydrated apo-ferritin with VPP on a Cs-corrected TEM and obtained the structure of apo-ferritin at near atomic resolution from both under- and over-focused dataset, illustrating the feasibility and new potential of combining VPP with Cs-corrected TEM for high resolution cryo-EM.\n\nHighlightsThe successful combination of volta phase plate and Cs-corrector in single particle cryo-EM.\n\nNear-atomic structure determined from over-focused images by cryo-EM. VPP-Cs-corrector coupled EM provides interesting optical properties.\n\nIn BriefWe took the unique advantage of the optical system by combining the volta phase plate and Cs-corrector in a modern TEM to collect high resolution micrographs of frozen-hydrated apo-ferritin in over-focus imaging conditions and determined the structure of apo-ferritin at 3.0 Angstrom resolution.

biophysics↗

bioRxiv2017

Single Molecule Sequencing Of M13 Virus Genome Without Amplification

Third generation sequencing is a direct measurement of DNA/RNA sequences at the single molecule level without amplification. In this study, we report sequencing of the genome of the M13 virus by a new single molecule sequencing platform. Our platform detects single molecule fluorescence by the total internal reflection microscope technique, with sequencing-by-synthesis chemistry. We sequenced the genome of M13 to a depth of 316x and 100% coverage. The consensus sequence accuracy is 100%. We demonstrated that single molecule sequencing has no significant GC bias.

genomics↗

bioRxiv2017

LncRNA-TUG1/EZH2 Axis Promotes Cell Proliferation, Migration And The EMT Phenotype Formation Through Sponging miR-382

Pancreatic carcinoma (PC) is the one of the most common and malignant cancer in the world. Despite many effort have been made in recent years, the survival rate of PC still remains unsatisfied. Therefore, investigating the mechanisms underlying the progression of PC might facilitate the development of novel treatments that improve patient prognosis. LncRNA Taurine Up-regulated Gene 1 (TUG1) was initially identified as a transcript up - regulated by taurine, siRNA - based depletion of TUG1 suppresses mouse retinal development, and the abnormal expression of TUG1 has been reported in many cancers. However, the biological role and molecular mechanism of TUG1 in pancreatic carcinoma (PC) still needs to be further investigated. In the current study, the expression of TUG1 in the PC cell lines and tissues was measured by quantitative real-time PCR (qRT-PCR), and loss-of-function and gain-of-function approaches were applied to investigate the function of TUG1 in PC cell. Online database analysis tools showed that miR-382 could interact with TUG1 and we found an inverse correlation between TUG1 and miR-382 in PC specimens. Moreover, dual luciferase reporter assay, RNA-binding protein immunoprecipitation (RIP) and applied biotin-avidin pulldown system further provide evidence that TUG1 directly targeted miR-382 by binding with microRNA binding site harboring in the TUG1 sequence. Furthermore, gene expression array analysis using clinical samples and RT-qPCR proposed that EZH2 was a target of miR-382 in PC. Collectively, these findings revealed that TUG1 functions as an oncogenic lncRNA that promotes tumor progression at least partially through function as an endogenous sponge by competing for miR-382 binding to regulate the miRNA target EZH2.

cancer biology↗

bioRxiv2016

Predicting gene expression level in E. coli from mRNA sequence information

MotivationThe accurate characterization of the translational mechanism is crucial for enhancing our understanding of the relationship between genotype and phenotype. In particular, predicting the impact of the genetic variants on gene expression will allow to optimize specific pathways and functions for engineering new biological systems. In this context, the development of accurate methods for predicting translation efficiency from the nucleotide sequence is a key challenge in computational biology.\n\nMethodsIn this work we present PGExpress, a binary classifier to discriminate between mRNA sequences with low and high translation efficiency in E. coli. PGExpress algorithm takes as input 12 features corresponding to RNA folding and anti-Shine-Dalgarno hybridization free energies. The method was trained on a set of 1,772 sequence variants (WT-High) of 137 essential E. coli genes. For each gene, we considered 13 sequence variants of the first 33 nucleotides encoding for the same amino acids followed by the superfolder GFP. Each gene variant is represented sequence blocks that include the Ribosome Binding Site (RBS), the first 33 nucleotides of the coding region (C33), the remaining part of the coding region (CC), and their combinations.\n\nResultsOur logistic regression-based tool (PGExpress) was trained using a 20-fold gene-based cross-validation procedure on the WT-High dataset. In this test PGExpress achieved an overall accuracy of 74%, a Matthews correlation coefficient 0.49 and an Area Under the Receiver Operating Characteristic Curve (AUC) of 0.81. Tested on 3 sets of sequences with different Ribosome Binding Sites, PGExpress reaches similar AUC. Finally, we validated our method by performing in-house experiments on five newly generated mRNA sequence variants. The predictions of the expression level of the new variants are in agreement with our experimental results in E. coli.\n\nAvailabilityhttp://folding.biofold.org/pgexpress\n\nContactmarkus.kollmann@hhu.de, emidio.capriotti@unibo.it

bioinformatics↗