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Identification of Cyclin-Dependent Kinase 1 as a Novel Regulator of Type I Interferon Signaling in Systemic Lupus Erythematosus

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ARTHRITIS & RHEUMATOLOGY 2016年第5期68卷 1222-1232页

作者： Wu, Lingling Qin, Yuting Xia, Shiwei Dai, Min Han, Xiao Wu, Yanfang Zhang, Xiaoyan Ma, Jianyang Wang, Yan Tang, Yuanjia Liu, Zheng Zhu, Wei Jallal, Bahija Yao, Yihong Qu, Bo Shen, Nan Shanghai Jiao Tong Univ Sch Med Renji Hosp Shanghai Inst Rheumatol 145 Shan Dong Middle Rd Shanghai 200001 Peoples R China Shanghai Jiao Tong Univ Sch Med Shanghai Inst Biol Sci Inst Hlth Sci Shanghai 200030 Peoples R China Chinese Acad Sci Shanghai Peoples R China Medimmune Inc Gaithersburg MD 20878 USA Cincinnati Childrens Hosp Med Ctr Ctr Autoimmune Genom & Etiol Cincinnati OH 45229 USA Shanghai Jiao Tong Univ Sch Med Collaborat Innovat Ctr Translat Med Shanghai 200030 Peoples R China Shanghai Jiao Tong Univ Sch Med Renji Hosp State Key Lab Oncogenes & Related GenesShanghai Shanghai 200030 Peoples R China

Objective. Type I interferon (IFN) signaling is regarded as a central pathogenic pathway in systemic lupus erythematosus (SLE). Specific inhibition of this pathway is a core area for the development of new therapies for SLE. This study was undertaken to clarify the pathogenic mechanism involved and to identify new therapeutic targets, using a high-throughput screening plat-form to determine novel regulators that contribute to the overactivation of the type I IFN signaling pathway in SLE. Methods. A high-throughput IFN-stimulated response element (ISRE)-luciferase assay was used to screen for candidate genes that regulate the IFN signaling pathway. Western blotting was used to confirm the regulatory function of CDK1. SYBR Green quantitative reverse transcriptase-polymerase chain reaction was used to detect the expression of individual IFN-stimulated genes (ISGs). The differential expression of CDK1 and ISGs in SLE patients and healthy controls was analyzed using RNA sequencing data and a microarray. Results. The high-throughput ISRE-luciferase assay revealed that CDK1 enhanced type I IFN signaling. Consistent with this finding, CDK1 promoted the type I IFN-induced phosphorylation of STAT-1 and the up-regulated expression of ISGs. CDK1 expression was elevated in peripheral blood mononuclear cells (PBMCs) and kidney biopsy specimens from SLE patients and correlated positively with their IFN scores. A CDK1 inhibitor reduced the expression of ISGs in PBMCs from SLE patients and in renal cells from mice with lupus. Conclusion. Our findings indicate that CDK1 is a positive regulator of the IFN signaling pathway. The overexpression of CDK1 might contribute to the abnormally amplified type I IFN signaling in SLE, and the inhibition of CDK1 could be used to down-regulate type I IFN signaling in SLE.

关键词： ANIMAL experimentation CELL culture CELLULAR signal transduction GENE expression INTERFERONS MICE PHOSPHORYLATION POLYMERASE chain reaction PROTEIN kinases FINANCING of research STATISTICS SYSTEMIC lupus erythematosus T-test (Statistics) WESTERN immunoblotting DATA analysis DATA analysis software microarray technology DESCRIPTIVE statistics

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In Vivo Therapeutic Success of MicroRNA-155 Antagomir in a Mouse Model of Lupus Alveolar Hemorrhage

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ARTHRITIS & RHEUMATOLOGY 2016年第4期68卷 953-964页

作者： Zhou, Shiyu Wang, Yan Meng, Yao Xiao, Chunyuan Liu, Zheng Brohawn, Philip Higgs, Brandon W. Jallal, Bahija Jia, Qian Qu, Bo Huang, Xinfang Tang, Yuanjia Yao, Yihong Harley, John B. Shen, Nan Chinese Acad Sci Shanghai Inst Biol Sci Inst Hlth Sci Shanghai Peoples R China Shanghai Jiao Tong Univ Sch Med Shanghai 200030 Peoples R China Renji Hosp Shanghai Peoples R China MedImmune Gaithersburg MD USA Univ Cincinnati Cincinnati Childrens Hosp Med Ctr Coll Med Cincinnati OH USA Cincinnati VA Med Ctr Cincinnati OH USA Chinese Acad Sci Renji Hosp Inst Hlth Sci Shanghai Inst Biol Sci Shanghai Peoples R China Shanghai Jiao Tong Univ Sch Med Shanghai 200030 Peoples R China Cincinnati Childrens Hosp Med Ctr Cincinnati OH 45229 USA

ObjectiveDiffuse alveolar hemorrhage (DAH) is a rare but life-threatening complication of systemic lupus erythematosus (SLE). Pristane-treated B6 mice develop severe DAH within 2 weeks of treatment. MicroRNA-155 (miR-155) is a pleiotropic microRNA that plays a crucial role in the regulation of immune responses. Recent studies have revealed a pathogenic role of miR-155 in various autoimmune disorders. The purpose of this study was to examine the role of miR-155 in the development of DAH in pristane-induced lupus using miR-155-knockout (miR-155(-/-)) mice and miR-155 antagomir to silence miR-155. MethodsDAH was induced by an intraperitoneal injection of 0.5 ml of pristane. MicroRNA-155 antagomir was administered intravenously to silence miR-155 expression. Lung tissues were collected for RNA extraction and were embedded in paraffin for sectioning. Gene expression profiling data were analyzed using Ingenuity Pathway Analysis. Real-time quantitative polymerase chain reaction analysis was used for single-gene validation. Luciferase reporter assay and argonaute 2 immunoprecipitation were performed for target validation. ResultsMicroRNA-155 expression was significantly increased during the development of DAH. Disease progression was reduced in miR-155(-/-) mice as well as by in vivo silencing of miR-155 using a miR-155 antagomir. MicroRNA-155 silencing dampened pristane-induced ectopic activation of multiple inflammatory pathways and reduced the expression of proinflammatory cytokines. Several negative regulators of NF-B signaling were inhibited by pristane and were reactivated in miR-155(-/-) mice. In particular, the antiinflammatory factor peroxisome proliferator-activated receptor was identified as a direct target of miR-155. ConclusionMicroRNA-155 promotes pristane-induced lung inflammation. It contributes to ectopic activation of NF-B signaling pathways by targeting multiple negative regulators. MicroRNA-155 antagomir may be a promising therapeutic strategy for treati

关键词： ANIMAL experimentation CELL culture CELLULAR signal transduction HEMORRHAGE LUNG diseases MICE POLYMERASE chain reaction FINANCING of research RNA SYSTEMIC lupus erythematosus T-test (Statistics) DATA analysis software microarray technology GENE expression profiling DESCRIPTIVE statistics ONE-way analysis of variance IN vivo studies DISEASE complications

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Digital droplet PCR on disk

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LAB ON A CHIP 2016年第1期16卷 208-216页

作者： Schuler, Friedrich Trotter, Martin Geltman, Marcel Schwemmer, Frank Wadle, Simon Dominguez-Garrido, Elena Lopez, Maria Cervera-Acedo, Cristina Santibanez, Paula von Stetten, Felix Zengerle, Roland Paust, Nils Hahn Schickard D-79110 Freiburg Germany Univ Freiburg IMTEK Dept Microsyst Engn Lab MEMS Applicat D-79110 Freiburg Germany Univ Freiburg BIOSS Ctr Biol Signalling Studies D-79106 Freiburg Germany Rioja Salud Fdn Mol Diagnost Lab La Rioja Spain

Existing systems for digital droplet PCR (ddPCR) either suffer from low integration or are difficult to introduce to mass fabrication. Here we present an integrated system that is compatible to mass fabrication and combines emulsification, PCR, and fluorescence readout in a single chamber within a disposable cartridge (disk). Droplets are generated by injecting the sample into fluorinated oil via centrifugal step emulsification. The resulting emulsion is aligned in the PCR and readout zone by capillary action. During thermocycling, gas bubbles generated by degassing are removed by capillary driven transport through tapered regions in the PCR chamber. Thereby, the positioning of the emulsion within the readout zone of the PCR chamber is ensured at any time and no bubbles are present during readout. Manual handling of the disk solely requires pipetting of oil and PCR mix into the inlet structures, placing the disk into the thermocycler and subsequently into a microarray scanner. The functionality of the ddPCR process chain is demonstrated by quantitative detection of the cystic fibrosis causing mutation ***508del, which is of interest for non-invasive prenatal testing (NIPT). The mutation was detected in a concentration range spanning four orders of magnitude. We envision that this work will lay the base for the development of highly integrated sample-to-digital-answer PCR systems that can be employed in routine clinical diagnosis.

关键词： POLYMERASE chain reaction DIGITAL technology INK cartridges THERMOCYCLING CYSTIC fibrosis -- Diagnosis microarray technology

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Prion-like Aggregation of Mitochondrial Antiviral Signaling Protein in Lupus Patients Is Associated With Increased Levels of Type I Interferon

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ARTHRITIS & RHEUMATOLOGY 2016年第11期68卷 2697-2707页

作者： Shao, Wen-Hai Shu, Daniel H. Zhen, Yuxuan Hilliard, Brendan Priest, Stephen O. Cesaroni, Matteo Ting, Jenny P. -Y. Cohen, Philip L. Temple Univ Lewis Katz Sch Med Philadelphia PA 19122 USA Univ N Carolina Chapel Hill NC USA

ObjectiveIncreased levels of type I interferon (IFN) and type I IFN-regulated genes are found in patients with systemic lupus erythematosus (SLE) and may be central to its pathogenesis. Mitochondrial antiviral signaling protein (MAVS) is a key regulator of type I IFN that undergoes a dramatic prion-like aggregation and self propagates the activation signal from viral RNA to amplify downstream IFN production. We undertook this study to determine whether such MAVS aggregates might play a role in the sustained increased production of type I IFN in SLE. MethodsPeripheral blood mononuclear cells were isolated and mitochondrial extracts were prepared. MAVS aggregation was detected by semidenatured agarose gel electrophoresis and confirmed by immunofluorescence staining. MAVS-associated signaling proteins were analyzed by Western blotting. MAVS aggregation-associated gene expression signature was analyzed by microarray. ResultsIn blood cells from 22 of 67 SLE patients, essentially all MAVS was in a high molecular weight aggregated form. None of 6 rheumatoid arthritis patients and only 3 of 33 healthy controls had abnormal MAVS. Compared to MAVS aggregate-negative patients, MAVS aggregate-positive SLE patients had significantly higher serum levels of IFN and significantly increased levels of autoantibodies against Sm and U1 RNP. Gene array data revealed a characteristic gene expression pattern in these patients, with altered expression of genes involved in IFN signaling and membrane trafficking. ConclusionPersistent MAVS aggregates may lead to increased type I IFN production and result in unmitigated signals leading to autoimmunity.

关键词： ACADEMIC medical centers CELL physiology CELLULAR signal transduction CHI-squared test ENZYME-linked immunosorbent assay FLUORESCENT antibody technique GENE expression INTERFERONS PULSED-field gel electrophoresis FINANCING of research SYSTEMIC lupus erythematosus T-test (Statistics) WESTERN immunoblotting microarray technology GENE expression profiling SIGNAL peptides DESCRIPTIVE statistics MANN Whitney U Test

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Multiplex cell microarrays for high-throughput screening

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LAB ON A CHIP 2016年第22期16卷 4248-4262页

作者： Berthuy, Ophelie I. Muldur, Sinan K. Rossi, Francois Colpo, Pascal Blum, Loic J. Marquette, Christophe A. Univ Lyon 1 Univ Lyon CNRS INSACPE LyonICBMSUMR 5246 43Bd 11 Novembre 1918 F-69622 Villeurbanne France Europeen Commiss Joint Res Ctr Inst Heath & Consumer Protect Ispra VA Italy

microarray technology was developed in the early 1990s to measure the transcription levels of thousands of genes in parallel. The basic premise of high-density arraying has since been expanded to create cell microarrays. Cells on chip are powerful experimental tools for high-throughput and multiplex screening of samples or cellular functions. Miniaturization increases assay throughput while reducing both reagent consumption and cell population heterogeneity effect, making these systems attractive for a wide range of assays, from drug discovery to toxicology, stem cell research and therapy. It is usual to functionalize the surface of a substrate to design cell microarrays. One form of cell microarrays, the transfected cell microarray, wherein plasmid DNA or siRNA spotted on the surface of a substrate is reverse-transfected locally into adherent cells, has become a standard tool for parallel cell-based analysis. With the advent of technology, cells can also be directly spotted onto functionalized surfaces using robotic fluid-dispensing devices or printed directly on bio-ink material. We are providing herein an overview of the latest developments in optical cell microarrays allowing high-throughput and high-content analysis.

关键词： microarray technology GENETIC transcription MULTICELLULAR life TISSUE arrays CYTOLOGY

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Cellular events during scar-free skin regeneration in the spiny mouse, Acomys

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WOUND REPAIR AND REGENERATION 2016年第1期24卷 75-88页

作者： Brant, Jason O. Yoon, Jung H. Polvadore, Trey Barbazuk, William Brad Maden, Malcolm Univ Florida Dept Biol Rm 410 Canc Genet Res Ctr2033 Mowry Rd Gainesville FL 32610 USA Univ Florida UF Genet Inst Rm 410 Canc Genet Res Ctr2033 Mowry Rd Gainesville FL 32610 USA

In contrast to the lab mouse, Mus musculus, several species of spiny mouse, Acomys, can regenerate epidermis, dermis, hairs, sebaceous glands with smooth muscle erector pili muscles and skeletal muscle of the panniculus carnonsus after full thickness skin wounding. Here, we have compared the responses of these scarring and nonscarring organisms concentrating on the immune cells and wound cytokines, cell proliferation, and the collagenous components of the wound bed and scar. The blood of Acomys is very neutropenic but there are greater numbers of mast cells in the Acomys wound than the Mus wound. Most importantly there are no F4/80 macrophages in the Acomys wound and many proinflammatory cytokines are either absent or in very low levels which we suggest may be primarily responsible for the excellent regenerative properties of the skin of this species. There is little difference in cell proliferation in the two species either in the epidermis or mesenchymal tissues but the cell density and matrix composition of the wound is very different. In Mus there are 8 collagens which are up-regulated at least 5-fold in the wound creating a strongly trichrome-positive matrix whereas in Acomys there are very few collagens present and the matrix shows only light trichrome staining. The major component of the Mus matrix is collagen XII which is up-regulated between 10 and 30-fold after wounding. These results suggest that in the Acomys wound the absence of many cytokines resulting in the lack of macrophages is responsible for the failure to up-regulate fibrotic collagens, a situation which permits a regenerative response within the skin rather than the generation of a scar.

关键词： EPIDERMIS PHYSIOLOGY EPITHELIUM IMMUNE system ANIMAL experimentation BLOOD cell count CELL physiology COLLAGEN COMPARATIVE studies CONNECTIVE tissues CYTOKINES GRANULATION tissue HISTOLOGICAL technique MACROPHAGES MAST cells MICE NEUTROPHILS POLYMERASE chain reaction PROBABILITY theory REGENERATION (Biology) FINANCING of research SCARS SKIN physiology SPLEEN STAINS & staining (Microscopy) T-test (Statistics) WOUND healing TRAUMATOLOGY diagnosis PROTEOMICS FIBROSIS REVERSE transcriptase polymerase chain reaction microarray technology DESCRIPTIVE statistics LEUKOCYTE count LYMPHOCYTE count IN vivo studies METABOLISM

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rCGH: a comprehensive array-based genomic profile platform for precision medicine

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BIOINFORMATICS 2016年第9期32卷 1402-1404页

作者： Commo, Frederic Guinney, Justin Ferte, Charles Bot, Brian Lefebvre, Celine Soria, Jean-Charles Andre, Fabrice Univ Paris 11 Gustave Roussy INSERM U981 F-94805 Villejuif France Sage Bionetworks Seattle WA 98109 USA Inst Gustave Roussy Dept Med Oncol Rue Camille Desmoulins F-94805 Villejuif France

We present rCGH, a comprehensive array-based comparative genomic hybridization analysis workflow, integrating computational improvements and functionalities specifically designed for precision medicine. rCGH supports the major microarray platforms, ensures a full traceability and facilitates profiles interpretation and decision-making through sharable interactive visualizations.

关键词： HEALTH of cancer patients GENOMICS MEDICAL research microarray technology BIOINFORMATICS

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Short Stature and Macrodactyly in a 13-Year-Old Female

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CLINICAL PEDIATRICS 2016年第11期55卷 1071-1074页

作者： Henry, Rohan K. Chaudhari, Monika Bowden, Sasigarn A. Ohio State Univ Coll Med Nationwide Childrens Hosp Columbus OH 43210 USA

The article presents a case study of a 13-year-old Caucasian female, who was presented for a growth and pubertal evaluation. The patient had minimal breast development and distinct characteristics including left foot macrodactyly, color blindness, and scalp hemangioma. It mentions that in addition to short stature, the patient was diagnosed for macrodactyly with the help of microarray technique. It treveals that microarray should be considered early for detecting Turner syndrome in a female.

关键词： TOE abnormalities TURNER'S syndrome DIAGNOSIS GROWTH disorders KARYOTYPES FAMILY medical history microarray technology ABNORMALITIES in the anatomical extremities GENETIC aspects

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Shared Governance Embedded in Population Ethics Can Enhance Health Equity Research at Both Micro and Macro Levels

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AMERICAN JOURNAL OF BIOETHICS 2016年第10期16卷 64-66页

作者： Sodeke, Stephen O. Yates, Clayton C. Tuskegee Univ Natl Ctr Bioeth Tuskegee AL 36088 USA Tuskegee Univ Tuskegee AL 36088 USA

Click to increase image sizeClick to decrease image size

关键词： RESEARCH -- Evaluation BIOETHICS CONSORTIA DIGNITY ETHICS HEALTH services accessibility HEALTH status indicators EMPLOYEE participation in management POPULATION PROFESSIONAL associations WORK microarray technology

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densityCut: an efficient and versatile topological approach for automatic clustering of biological data

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BIOINFORMATICS 2016年第17期32卷 2567-2576页

作者： Ding, Jiarui Shah, Sohrab Condon, Anne Univ British Columbia Dept Comp Sci Vancouver BC V6T 1Z4 Canada BC Canc Res Ctr Dept Mol Oncol Vancouver BC V5Z 1L3 Canada

Motivation: Many biological data processing problems can be formalized as clustering problems to partition data points into sensible and biologically interpretable groups. Results: This article introduces densityCut, a novel density-based clustering algorithm, which is both time-and space-efficient and proceeds as follows: densityCut first roughly estimates the densities of data points from a K-nearest neighbour graph and then refines the densities via a random walk. A cluster consists of points falling into the basin of attraction of an estimated mode of the underlining density function. A post-processing step merges clusters and generates a hierarchical cluster tree. The number of clusters is selected from the most stable clustering in the hierarchical cluster tree. Experimental results on ten synthetic benchmark datasets and two microarray gene expression datasets demonstrate that densityCut performs better than state-of-the-art algorithms for clustering biological datasets. For applications, we focus on the recent cancer mutation clustering and single cell data analyses, namely to cluster variant allele frequencies of somatic mutations to reveal clonal architectures of individual tumours, to cluster single-cell gene expression data to uncover cell population compositions, and to cluster single-cell mass cytometry data to detect communities of cells of the same functional states or types. densityCut performs better than competing algorithms and is scalable to large datasets.

关键词： BIOLOGICAL databases COMPUTERS in biology DOCUMENT clustering microarray technology DNA microarrays GENE expression

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