Publications by authors named "Nripesh Prasad"

41 Publications

VAP-A and its binding partner CERT drive biogenesis of RNA-containing extracellular vesicles at ER membrane contact sites.

Dev Cell 2022 04 13;57(8):974-994.e8. Epub 2022 Apr 13.

Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN, USA; Department of Pathology, Microbiology and Immunology, Vanderbilt University Medical Center, Nashville, TN, USA. Electronic address:

RNA transfer via extracellular vesicles (EVs) influences cell phenotypes; however, lack of information regarding biogenesis of RNA-containing EVs has limited progress in the field. Here, we identify endoplasmic reticulum membrane contact sites (ER MCSs) as platforms for the generation of RNA-containing EVs. We identify a subpopulation of small EVs that is highly enriched in RNA and regulated by the ER MCS linker protein VAP-A. Functionally, VAP-A-regulated EVs are critical for miR-100 transfer between cells and in vivo tumor formation. Lipid analysis of VAP-A-knockdown EVs revealed reductions in the EV biogenesis lipid ceramide. Knockdown of the VAP-A-binding ceramide transfer protein CERT led to similar defects in EV RNA content. Imaging experiments revealed that VAP-A promotes luminal filling of multivesicular bodies (MVBs), CERT localizes to MVBs, and the ceramide-generating enzyme neutral sphingomyelinase 2 colocalizes with VAP-A-positive ER. We propose that ceramide transfer via VAP-A-CERT linkages drives the biogenesis of a select RNA-containing EV population.
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http://dx.doi.org/10.1016/j.devcel.2022.03.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9075344PMC
April 2022

Immune Activity and Response Differences of Oncolytic Viral Therapy in Recurrent Glioblastoma: Gene Expression Analyses of a Phase IB Study.

Clin Cancer Res 2022 02;28(3):498-506

Department of Neurosurgery, Heersink School of Medicine, The University of Alabama at Birmingham, Birmingham, Alabama.

Purpose: Previously, clinical trials of experimental virotherapy for recurrent glioblastoma multiforme (GBM) demonstrated that inoculation with a conditionally replication-competent Δγ34.5 oncolytic herpes simplex virus (oHSV), G207, was safe. Following the initial safety study, a phase Ib trial enrolled 6 adult patients diagnosed with GBM recurrence from which tumor tissue was banked for future studies.

Patients And Methods: Here, we analyzed tumor RNA sequencing (RNA-seq) data obtained from pre- and posttreatment (collected 2 or 5 days after G207 injection) biopsies from the phase Ib study patients.

Results: Using a Spearman rank-order correlation analysis, we identified approximately 500 genes whose expression pattern correlated with survival duration. Many of these genes were enriched for the intrinsic IFN-mediated antiviral and adaptive immune functional responses, including immune cell chemotaxis and antigen presentation to T-cells. Furthermore, we show that the expression of several T-cell-related genes was highest in the patient with the longest survival after G207 inoculation.

Conclusions: Our data support that the oHSV-induced type I IFN production and the subsequent recruitment of an adaptive immune response differed between enrolled patients and showed association with survival duration in patients with recurrent malignant glioma after treatment with an early generation oHSV.
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http://dx.doi.org/10.1158/1078-0432.CCR-21-2636DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8846434PMC
February 2022

Integrated Analysis of the Pancreas and Islets Reveals Unexpected Findings in Human Male With Type 1 Diabetes.

J Endocr Soc 2021 Dec 29;5(12):bvab162. Epub 2021 Oct 29.

Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN, USA.

Clinical and pathologic heterogeneity in type 1 diabetes is increasingly being recognized. Findings in the islets and pancreas of a 22-year-old male with 8 years of type 1 diabetes were discordant with expected results and clinical history (islet autoantibodies negative, hemoglobin A1c 11.9%) and led to comprehensive investigation to define the functional, molecular, genetic, and architectural features of the islets and pancreas to understand the cause of the donor's diabetes. Examination of the donor's pancreatic tissue found substantial but reduced β-cell mass with some islets devoid of β cells (29.3% of 311 islets) while other islets had many β cells. Surprisingly, isolated islets from the donor pancreas had substantial insulin secretion, which is uncommon for type 1 diabetes of this duration. Targeted and whole-genome sequencing and analysis did not uncover monogenic causes of diabetes but did identify high-risk human leukocyte antigen haplotypes and a genetic risk score suggestive of type 1 diabetes. Further review of pancreatic tissue found islet inflammation and some previously described α-cell molecular features seen in type 1 diabetes. By integrating analysis of isolated islets, histological evaluation of the pancreas, and genetic information, we concluded that the donor's clinical insulin deficiency was most likely the result autoimmune-mediated β-cell loss but that the constellation of findings was not typical for type 1 diabetes. This report highlights the pathologic and functional heterogeneity that can be present in type 1 diabetes.
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http://dx.doi.org/10.1210/jendso/bvab162DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8633619PMC
December 2021

Identification of canine circulating miRNAs as tumor biospecific markers using Next-Generation Sequencing and Q-RT-PCR.

Biochem Biophys Rep 2021 Dec 19;28:101106. Epub 2021 Aug 19.

Scott-Ritchey Research Center, College of Veterinary Medicine, Auburn University, USA.

Delay in cancer diagnosis often results in metastasis and an inability to successfully treat the tumor. The use of broadly cancer-specific biomarkers at an early stage may improve cancer treatment and staging. This study has explored circulatory exosomal miRNAs as potential diagnostic biomarkers to identify cancer patients. Secretory exosomal miRNAs were isolated from 13 canine cancer cell lines (lymphoma, mast cell tumor, histiocytic cell line, osteosarcoma, melanoma, and breast tumor) and were sequenced by Next-Generation sequencing (NGS). We have identified 6 miRNAs (cfa-miR-9, -1841, -1306, -345, -132, and -26b) by NGS that were elevated in all cancer cell types. The miRNAs identified by NGS were then examined by Q-RT-PCR. The PCR data demonstrated similar expression patterns to those seen with NGS but provided fold differences that were much lower than those seen for NGS. Cfa-miR-9 was found to be the most consistently elevated miRNA in NGS and PCR, making it the most likely miRNA to prove diagnostic. In this study, we have demonstrated that it is possible to identify exosomal miRNAs with elevated secretion across multiple tumor types that could be used as circulatory diagnostic biomarkers for liquid biopsy in the future.
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http://dx.doi.org/10.1016/j.bbrep.2021.101106DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8379617PMC
December 2021

Combinatorial transcription factor profiles predict mature and functional human islet α and β cells.

JCI Insight 2021 09 22;6(18). Epub 2021 Sep 22.

Division of Diabetes, Endocrinology and Metabolism, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee, USA.

Islet-enriched transcription factors (TFs) exert broad control over cellular processes in pancreatic α and β cells, and changes in their expression are associated with developmental state and diabetes. However, the implications of heterogeneity in TF expression across islet cell populations are not well understood. To define this TF heterogeneity and its consequences for cellular function, we profiled more than 40,000 cells from normal human islets by single-cell RNA-Seq and stratified α and β cells based on combinatorial TF expression. Subpopulations of islet cells coexpressing ARX/MAFB (α cells) and MAFA/MAFB (β cells) exhibited greater expression of key genes related to glucose sensing and hormone secretion relative to subpopulations expressing only one or neither TF. Moreover, all subpopulations were identified in native pancreatic tissue from multiple donors. By Patch-Seq, MAFA/MAFB-coexpressing β cells showed enhanced electrophysiological activity. Thus, these results indicate that combinatorial TF expression in islet α and β cells predicts highly functional, mature subpopulations.
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http://dx.doi.org/10.1172/jci.insight.151621DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8492318PMC
September 2021

Coordinated interactions between endothelial cells and macrophages in the islet microenvironment promote β cell regeneration.

NPJ Regen Med 2021 Apr 6;6(1):22. Epub 2021 Apr 6.

Department of Medicine, Division of Diabetes, Endocrinology, and Metabolism, Vanderbilt University Medical Center, Nashville, TN, USA.

Endogenous β cell regeneration could alleviate diabetes, but proliferative stimuli within the islet microenvironment are incompletely understood. We previously found that β cell recovery following hypervascularization-induced β cell loss involves interactions with endothelial cells (ECs) and macrophages (MΦs). Here we show that proliferative ECs modulate MΦ infiltration and phenotype during β cell loss, and recruited MΦs are essential for β cell recovery. Furthermore, VEGFR2 inactivation in quiescent ECs accelerates islet vascular regression during β cell recovery and leads to increased β cell proliferation without changes in MΦ phenotype or number. Transcriptome analysis of β cells, ECs, and MΦs reveals that β cell proliferation coincides with elevated expression of extracellular matrix remodeling molecules and growth factors likely driving activation of proliferative signaling pathways in β cells. Collectively, these findings suggest a new β cell regeneration paradigm whereby coordinated interactions between intra-islet MΦs, ECs, and extracellular matrix mediate β cell self-renewal.
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http://dx.doi.org/10.1038/s41536-021-00129-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8024255PMC
April 2021

Investigational Assay for Haplotype Phasing of the Huntingtin Gene.

Mol Ther Methods Clin Dev 2020 Dec 11;19:162-173. Epub 2020 Sep 11.

Wave Life Sciences Ltd., Cambridge, MA 02138, USA.

Novel treatments for Huntington's disease (HD), a progressive neurodegenerative disorder, include selective targeting of the mutant allele of the huntingtin gene (m) carrying the abnormally expanded disease-causing cytosine-adenine-guanine (CAG) repeat. WVE-120101 and WVE-120102 are investigational stereopure antisense oligonucleotides that enable selective suppression of m by targeting single-nucleotide polymorphisms (SNPs) that are in haplotype phase with the CAG repeat expansion. Recently developed long-read sequencing technologies can capture CAG expansions and distant SNPs of interest and potentially facilitate haplotype-based identification of patients for clinical trials of oligonucleotide therapies. However, improved methods are needed to phase SNPs with CAG repeat expansions directly and reliably without need for familial genotype/haplotype data. Our haplotype phasing method uses single-molecule real-time sequencing and a custom algorithm to determine with confidence bases at SNPs on mutant alleles, even without familial data. Herein, we summarize this methodology and validate the approach using patient-derived samples with known phasing results. Comparison of experimentally measured CAG repeat lengths, heterozygosity, and phasing with previously determined results showed improved performance. Our methodology enables the haplotype phasing of SNPs of interest and the disease-causing, expanded CAG repeat of the huntingtin gene, enabling accurate identification of patients with HD eligible for allele-selective clinical studies.
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http://dx.doi.org/10.1016/j.omtm.2020.09.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7648085PMC
December 2020

Circulating microRNA as biomarkers of canine mammary carcinoma in dogs.

J Vet Intern Med 2020 May 27;34(3):1282-1290. Epub 2020 Apr 27.

Department of Pathobiology, College of Veterinary Medicine, AURIC-Auburn University Research Initiative in Cancer, Auburn University, Auburn, Alabama, United States.

Background: Differentiating benign from canine malignant mammary tumors requires invasive surgical biopsy. Circulating microRNAs (miRNA) may represent promising minimally invasive cancer biomarkers in people and animals.

Objectives: To evaluate the serum mRNA profile between dogs with and without mammary carcinoma, and to determine if any of these markers have prognostic significance.

Animals: Ten healthy client-owned female dogs (5 intact, 5 spayed) and 10 dogs with histologically confirmed mammary carcinoma were included; 9 were client-owned, whereas 1 was a research colony dog.

Methods: Retrospective study. Serum miRNA was evaluated by RNA deep-sequencing (RNAseq) and digital droplet PCR (dPCR).Expression of candidate biomarkers miR-18a, miR-19b, miR-29b, miR-34c, miR-122, miR-125a, and miR-181a was compared with clinical characteristics, including grade, metastasis, and survival.

Results: 452 unique serum miRNAs were detected by RNAseq. Sixty-five individual miRNAs were differentially expressed (>±1.5-fold) and statistically significant between groups. Serum miR-19b (P = .003) and miR-125a (P < .001) were significantly higher in the mammary carcinoma group by dPCR. Both had high accuracy based on receiver operator characteristic area under the curve (0.930 for miR-125a; 0.880 for miR-19b). Circulating miR-18a by RNAseq was significantly higher in mammary carcinoma dogs with histologic evidence of lymphatic invasion (P = 0.03). There was no significant association with any miRNA and survival or inflammatory status.

Conclusions And Clinical Importance: Circulating miRNAs are differentially expressed in dogs with mammary carcinoma. Serum miR-19b and miR-18a represent candidate biomarkers for diagnosis and prognosis, respectively.
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http://dx.doi.org/10.1111/jvim.15764DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7255679PMC
May 2020

Differential Expression of miRNAs in Hypoxia ("HypoxamiRs") in Three Canine High-Grade Glioma Cell Lines.

Front Vet Sci 2020 28;7:104. Epub 2020 Feb 28.

Department of Pathobiology, Auburn University, Auburn, AL, United States.

Dogs with spontaneous high-grade gliomas increasingly are being proposed as useful large animal pre-clinical models for the human disease. Hypoxia is a critical microenvironmental condition that is common in both canine and human high-grade gliomas and drives increased angiogenesis, chemo- and radioresistance, and acquisition of a stem-like phenotype. Some of this effect is mediated by the hypoxia-induced expression of microRNAs, small (~22 nucleotides long), non-coding RNAs that can modulate gene expression through interference with mRNA translation. Using an model with three canine high-grade glioma cell lines (J3T, SDT3G, and G06A) exposed to 72 h of 1.5% oxygen vs. standard 20% oxygen, we examined the global "hypoxamiR" profile using small RNA-Seq and performed pathway analysis for targeted genes using both Panther and NetworkAnalyst. Important pathways include many that are well-established as being important in glioma biology, general cancer biology, hypoxia, angiogenesis, immunology, and stem-ness, among others. This work provides the first examination of the effect of hypoxia on miRNA expression in the context of canine glioma, and highlights important similarities with the human disease.
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http://dx.doi.org/10.3389/fvets.2020.00104DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7093022PMC
February 2020

Tacrolimus- and sirolimus-induced human β cell dysfunction is reversible and preventable.

JCI Insight 2020 01 16;5(1). Epub 2020 Jan 16.

Division of Diabetes, Endocrinology and Metabolism, Department of Medicine, and.

Posttransplantation diabetes mellitus (PTDM) is a common and significant complication related to immunosuppressive agents required to prevent organ or cell transplant rejection. To elucidate the effects of 2 commonly used agents, the calcineurin inhibitor tacrolimus (TAC) and the mTOR inhibitor sirolimus (SIR), on islet function and test whether these effects could be reversed or prevented, we investigated human islets transplanted into immunodeficient mice treated with TAC or SIR at clinically relevant levels. Both TAC and SIR impaired insulin secretion in fasted and/or stimulated conditions. Treatment with TAC or SIR increased amyloid deposition and islet macrophages, disrupted insulin granule formation, and induced broad transcriptional dysregulation related to peptide processing, ion/calcium flux, and the extracellular matrix; however, it did not affect regulation of β cell mass. Interestingly, these β cell abnormalities reversed after withdrawal of drug treatment. Furthermore, cotreatment with a GLP-1 receptor agonist completely prevented TAC-induced β cell dysfunction and partially prevented SIR-induced β cell dysfunction. These results highlight the importance of both calcineurin and mTOR signaling in normal human β cell function in vivo and suggest that modulation of these pathways may prevent or ameliorate PTDM.
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http://dx.doi.org/10.1172/jci.insight.130770DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7030815PMC
January 2020

Decrease in MiR-148a Expression During Initiation of Chief Cell Transdifferentiation.

Cell Mol Gastroenterol Hepatol 2020 29;9(1):61-78. Epub 2019 Aug 29.

Nashville VA Medical Center, Nashville, Tennessee; Department of Surgery, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, Tennessee; Department of Cell and Developmental Biology, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, Tennessee; Epithelial Biology Center, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, Tennessee. Electronic address:

Gastric chief cells differentiate from mucous neck cells and develop their mature state at the base of oxyntic glands with expression of secretory zymogen granules. After parietal cell loss, chief cells transdifferentiate into mucous cell metaplasia, designated spasmolytic polypeptide-expressing metaplasia (SPEM), which is considered a candidate precursor of gastric cancer. We examined the range of microRNA (miRNA) expression in chief cells and identified miRNAs involved in chief cell transdifferentiation into SPEM. Among them, miR-148a was strongly and specifically expressed in chief cells and significantly decreased during the process of chief cell transdifferentiation. Interestingly, suppression of miR-148a in a conditionally immortalized chief cell line induced up-regulation of CD44 variant 9 (CD44v9), one of the transcripts expressed at an early stage of SPEM development, and DNA methyltransferase 1 (Dnmt1), an established target of miR-148a. Immunostaining analyses showed that Dnmt1 was up-regulated in SPEM cells as well as in chief cells before the emergence of SPEM in mouse models of acute oxyntic atrophy using either DMP-777 or L635. In the cascade of events that leads to transdifferentiation, miR-148a was down-regulated after acute oxyntic atrophy either in xCT knockout mice or after sulfasalazine inhibition of xCT. These findings suggest that the alteration of miR-148a expression is an early event in the process of chief cell transdifferentiation into SPEM.
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http://dx.doi.org/10.1016/j.jcmgh.2019.08.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6881610PMC
May 2021

AS03-Adjuvanted H5N1 Avian Influenza Vaccine Modulates Early Innate Immune Signatures in Human Peripheral Blood Mononuclear Cells.

J Infect Dis 2019 05;219(11):1786-1798

Vanderbilt Vaccine Research Program, Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, Tennessee.

Background: Adjuvant System 03 (AS03) markedly enhances responses to influenza A/H5N1 vaccines, but the mechanisms of this enhancement are incompletely understood.

Methods: Using ribonucleic acid sequencing on peripheral blood mononuclear cells (PBMCs) from AS03-adjuvanted and unadjuvanted inactivated H5N1 vaccine recipients, we identified differentially expressed genes, enriched pathways, and genes that correlated with serologic responses. We compared bulk PBMC findings with our previously published assessments of flow-sorted immune cell types.

Results: AS03-adjuvanted vaccine induced the strongest differential signals on day 1 postvaccination, activating multiple innate immune pathways including interferon and JAK-STAT signaling, Fcγ receptor (FcγR)-mediated phagocytosis, and antigen processing and presentation. Changes in signal transduction and immunoglobulin genes predicted peak hemagglutinin inhibition (HAI) titers. Compared with individual immune cell types, activated PBMC genes and pathways were most similar to innate immune cells. However, several pathways were unique to PBMCs, and several pathways identified in individual cell types were absent in PBMCs.

Conclusions: Transcriptomic analysis of PBMCs after AS03-adjuvanted H5N1 vaccination revealed early activation of innate immune signaling, including a 5- to 8-fold upregulation of FcγR1A/1B/1C genes. Several early gene responses were correlated with HAI titer, indicating links with the adaptive immune response. Although PBMCs and cell-specific results shared key innate immune signals, unique signals were identified by both approaches.
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http://dx.doi.org/10.1093/infdis/jiy721DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6500554PMC
May 2019

Human islets expressing HNF1A variant have defective β cell transcriptional regulatory networks.

J Clin Invest 2019 01 3;129(1):246-251. Epub 2018 Dec 3.

Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, Tennessee, USA.

Using an integrated approach to characterize the pancreatic tissue and isolated islets from a 33-year-old with 17 years of type 1 diabetes (T1D), we found that donor islets contained β cells without insulitis and lacked glucose-stimulated insulin secretion despite a normal insulin response to cAMP-evoked stimulation. With these unexpected findings for T1D, we sequenced the donor DNA and found a pathogenic heterozygous variant in the gene encoding hepatocyte nuclear factor-1α (HNF1A). In one of the first studies of human pancreatic islets with a disease-causing HNF1A variant associated with the most common form of monogenic diabetes, we found that HNF1A dysfunction leads to insulin-insufficient diabetes reminiscent of T1D by impacting the regulatory processes critical for glucose-stimulated insulin secretion and suggest a rationale for a therapeutic alternative to current treatment.
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http://dx.doi.org/10.1172/JCI121994DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6307934PMC
January 2019

Ectonucleoside Triphosphate Diphosphohydrolase-3 Antibody Targets Adult Human Pancreatic β Cells for In Vitro and In Vivo Analysis.

Cell Metab 2019 03 15;29(3):745-754.e4. Epub 2018 Nov 15.

Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN 37240, USA; Department of Medicine, Division of Diabetes, Endocrinology, and Metabolism, Vanderbilt University Medical Center, Nashville, TN 37232, USA; VA Tennessee Valley Healthcare, Nashville, TN 37212, USA. Electronic address:

Identification of cell-surface markers specific to human pancreatic β cells would allow in vivo analysis and imaging. Here we introduce a biomarker, ectonucleoside triphosphate diphosphohydrolase-3 (NTPDase3), that is expressed on the cell surface of essentially all adult human β cells, including those from individuals with type 1 or type 2 diabetes. NTPDase3 is expressed dynamically during postnatal human pancreas development, appearing first in acinar cells at birth, but several months later its expression declines in acinar cells while concurrently emerging in islet β cells. Given its specificity and membrane localization, we utilized an NTPDase3 antibody for purification of live human β cells as confirmed by transcriptional profiling, and, in addition, for in vivo imaging of transplanted human β cells. Thus, NTPDase3 is a cell-surface biomarker of adult human β cells, and the antibody directed to this protein should be a useful new reagent for β cell sorting, in vivo imaging, and targeting.
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http://dx.doi.org/10.1016/j.cmet.2018.10.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6402969PMC
March 2019

Diverse Long RNAs Are Differentially Sorted into Extracellular Vesicles Secreted by Colorectal Cancer Cells.

Cell Rep 2018 10;25(3):715-725.e4

Department of Biological Sciences, Vanderbilt University Medical Center, Nashville, TN 37235, USA. Electronic address:

The regulation and functional roles of secreted coding and long noncoding RNAs (lncRNAs; >200 nt) are largely unknown. We previously showed that mutant KRAS colorectal cancer (CRC) cells release extracellular vesicles (EVs) containing distinct proteomes, microRNAs (miRNAs), and circular RNAs. Here, we comprehensively identify diverse classes of CRC extracellular long RNAs secreted in EVs and demonstrate differential export of specific RNAs. Distinct noncoding RNAs, including antisense transcripts and transcripts derived from pseudogenes, are enriched in EVs compared to cellular profiles. We detected strong enrichment of Rab13 in mutant KRAS EVs and demonstrate functional delivery of Rab13 mRNA to recipient cells. To assay functional transfer of lncRNAs, we implemented a CRISPR/Cas9-based RNA-tracking system to monitor delivery to recipient cells. We show that gRNAs containing export signals from secreted RNAs can be transferred from donor to recipient cells. Our data support the existence of cellular mechanisms to selectively export diverse classes of RNA.
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http://dx.doi.org/10.1016/j.celrep.2018.09.054DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6248336PMC
October 2018

Malignant canine mammary epithelial cells shed exosomes containing differentially expressed microRNA that regulate oncogenic networks.

BMC Cancer 2018 Aug 20;18(1):832. Epub 2018 Aug 20.

Department of Pathobiology, College of Veterinary Medicine, Auburn University, 166 Greene Hall, Auburn, AL, 36849, USA.

Background: Breast (mammary) cancers in human (BC) and canine (CMT) patients share clinical, pathological, and molecular similarities that suggest dogs may be a useful translational model. Many cancers, including BC, shed exosomes that contain microRNAs (miRs) into the microenvironment and circulation, and these may represent biomarkers of metastasis and tumor phenotype.

Methods: Three normal canine mammary epithelial cell (CMEC) cultures and 5 CMT cell lines were grown in serum-free media. Exosomes were isolated from culture media by ultracentrifugation then profiled by transmission electron microscopy, dynamic light scattering, and Western blot. Exosomal small RNA was deep-sequenced on an Illumina HiSeq2500 sequencer and validated by qRT-PCR. In silico bioinformatic analysis was carried out to determine microRNA gene and pathway targets.

Results: CMEC and CMT cell lines shed round, "cup-shaped" exosomes approximately 150-200 nm, and were immunopositive for exosomal marker CD9. Deep-sequencing averaged ~ 15 million reads/sample. Three hundred thirty-eight unique miRs were detected, with 145 having > ±1.5-fold difference between one or more CMT and CMEC samples. Gene ontology analysis revealed that the upregulated miRs in this exosomal population regulate a number of relevant oncogenic networks. Several miRNAs including miR-18a, miR-19a and miR-181a were predicted in silico to target the canine estrogen receptor (ESR1α).

Conclusions: CMEC and CMT cells shed exosomes in vitro that contain differentially expressed miRs. CMT exosomal RNA expresses a limited number of miRs that are up-regulated relative to CMEC, and these are predicted to target biologically relevant hormone receptors and oncogenic pathways. These results may inform future studies of circulating exosomes and the utility of miRs as biomarkers of breast cancer in women and dogs.
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http://dx.doi.org/10.1186/s12885-018-4750-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6102898PMC
August 2018

Neonatal lactocrine deficiency affects the adult porcine endometrial transcriptome at pregnancy day 13.

Biol Reprod 2019 01;100(1):71-85

Department of Animal Sciences, Endocrinology and Animal Biosciences Program, Rutgers University, New Brunswick, New Jersey, USA.

Reproductive performance of female pigs that do not receive sufficient colostrum from birth is permanently impaired. Whether lactocrine deficiency, reflected by low serum immunoglobulin immunocrit (iCrit), affects patterns of endometrial gene expression during the periattachment period of early pregnancy is unknown. Here, objectives were to determine effects of low iCrit at birth on the adult endometrial transcriptome on pregnancy day (PxD) 13. On the first day of postnatal life, gilts were assigned to high or low iCrit groups. Adult high (n = 8) and low (n = 7) iCrit gilts were bred (PxD 0), and humanely slaughtered on PxD 13 when tissues and fluids were collected. The endometrial transcriptome was defined for each group using mRNAseq and microRNAseq. Reads were mapped to the Sus scrofa 11.1 genome build. Mature microRNAs were annotated using miRBase 21. Differential expression was defined based on fold change (≥ ±1.5). Lactocrine deficiency did not affect corpora lutea number, uterine horn length, uterine wet weight, conceptus recovery, or uterine luminal fluid estrogen content on PxD 13. However, mRNAseq revealed 1157 differentially expressed endometrial mRNAs in high versus low iCrit gilts. Differentially expressed genes had functions related to solute transport, endometrial receptivity, and immune response. Six differentially expressed endometrial microRNAs included five predicted to target 62 differentially expressed mRNAs, affecting similar biological processes. Thus, lactocrine deficiency on the first day of postnatal life can alter uterine developmental trajectory with lasting effects on endometrial responses to pregnancy as reflected at the level of the transcriptome on PxD 13.
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http://dx.doi.org/10.1093/biolre/ioy180DOI Listing
January 2019

Reversible deficits in apical transporter trafficking associated with deficiency in diacylglycerol acyltransferase.

Traffic 2018 11 21;19(11):879-892. Epub 2018 Sep 21.

Department of Surgery, Vanderbilt University School of Medicine, Nashville, Tennessee.

Deficiency in diacylglycerol acyltransferase (DGAT1) is a rare cause of neonatal diarrhea, without a known mechanism or in vitro model. A patient presenting at our institution at 7 weeks of life with failure to thrive and diarrhea was found by whole-exome sequencing to have a homozygous DGAT1 truncation mutation. Duodenal biopsies showed loss of DGAT1 and deficits in apical membrane transporters and junctional proteins in enterocytes. When placed on a very low-fat diet, the patient's diarrhea resolved with normalization of brush border transporter localization in endoscopic biopsies. DGAT1 knockdown in Caco2-BBe cells modeled the deficits in apical trafficking, with loss of apical DPPIV and junctional occludin. Elevation in cellular lipid levels, including diacylglycerol (DAG) and phospholipid metabolites of DAG, was documented by lipid analysis in DGAT1 knockdown cells. Culture of the DGAT1 knockdown cells in lipid-depleted media led to re-establishment of occludin and return of apical DPPIV. DGAT1 loss appears to elicit global changes in enterocyte polarized trafficking that could account for deficits in absorption seen in the patient. The in vitro modeling of this disease should allow for investigation of possible therapeutic targets.
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http://dx.doi.org/10.1111/tra.12608DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6191315PMC
November 2018

-GlcNAc transferase missense mutations linked to X-linked intellectual disability deregulate genes involved in cell fate determination and signaling.

J Biol Chem 2018 07 16;293(27):10810-10824. Epub 2018 May 16.

From the Department of Biochemistry and Molecular Biology, Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia 30602,

It is estimated that ∼1% of the world's population has intellectual disability, with males affected more often than females. is an X-linked gene encoding for the enzyme GlcNAc transferase (OGT), which carries out the reversible addition of -acetylglucosamine (GlcNAc) to Ser/Thr residues of its intracellular substrates. Three missense mutations in the tetratricopeptide (TPR) repeats of OGT have recently been reported to cause X-linked intellectual disability (XLID). Here, we report the discovery of two additional novel missense mutations (c.775 G>A, p.A259T, and c.1016 A>G, p.E339G) in the TPR domain of OGT that segregate with XLID in affected families. Characterization of all five of these XLID missense variants of OGT demonstrates modest declines in thermodynamic stability and/or activities of the variants. We engineered each of the mutations into a male human embryonic stem cell line using CRISPR/Cas9. Investigation of the global GlcNAc profile as well as OGT and GlcNAc hydrolase levels by Western blotting showed no gross changes in steady-state levels in the engineered lines. However, analyses of the differential transcriptomes of the OGT variant-expressing stem cells revealed shared deregulation of genes involved in cell fate determination and liver X receptor/retinoid X receptor signaling, which has been implicated in neuronal development. Thus, here we reveal two additional mutations encoding residues in the TPR regions of OGT that appear causal for XLID and provide evidence that the relatively stable and active TPR variants may share a common, unelucidated mechanism of altering gene expression profiles in human embryonic stem cells.
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http://dx.doi.org/10.1074/jbc.RA118.002583DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6036218PMC
July 2018

Cystic fibrosis-related diabetes is caused by islet loss and inflammation.

JCI Insight 2018 04 19;3(8). Epub 2018 Apr 19.

Department of Medicine, Division of Diabetes, Endocrinology, and Metabolism, Vanderbilt University Medical Center, Nashville, Tennessee, USA.

Cystic fibrosis-related (CF-related) diabetes (CFRD) is an increasingly common and devastating comorbidity of CF, affecting approximately 35% of adults with CF. However, the underlying causes of CFRD are unclear. Here, we examined cystic fibrosis transmembrane conductance regulator (CFTR) islet expression and whether the CFTR participates in islet endocrine cell function using murine models of β cell CFTR deletion and normal and CF human pancreas and islets. Specific deletion of CFTR from murine β cells did not affect β cell function. In human islets, CFTR mRNA was minimally expressed, and CFTR protein and electrical activity were not detected. Isolated CF/CFRD islets demonstrated appropriate insulin and glucagon secretion, with few changes in key islet-regulatory transcripts. Furthermore, approximately 65% of β cell area was lost in CF donors, compounded by pancreatic remodeling and immune infiltration of the islet. These results indicate that CFRD is caused by β cell loss and intraislet inflammation in the setting of a complex pleiotropic disease and not by intrinsic islet dysfunction from CFTR mutation.
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http://dx.doi.org/10.1172/jci.insight.98240DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5931120PMC
April 2018

Spaceflight Modifies Gene Expression in Response to Antibiotic Exposure and Reveals Role of Oxidative Stress Response.

Front Microbiol 2018 16;9:310. Epub 2018 Mar 16.

Department of Chemical and Biological Engineering, University of Colorado Boulder, Boulder, CO, United States.

Bacteria grown in space experiments under microgravity conditions have been found to undergo unique physiological responses, ranging from modified cell morphology and growth dynamics to a putative increased tolerance to antibiotics. A common theory for this behavior is the loss of gravity-driven convection processes in the orbital environment, resulting in both reduction of extracellular nutrient availability and the accumulation of bacterial byproducts near the cell. To further characterize the responses, this study investigated the transcriptomic response of to both microgravity and antibiotic concentration. was grown aboard International Space Station in the presence of increasing concentrations of the antibiotic gentamicin with identical ground controls conducted on Earth. Here we show that within 49 h of being cultured, adapted to grow at higher antibiotic concentrations in space compared to Earth, and demonstrated consistent changes in expression of 63 genes in response to an increase in drug concentration in both environments, including specific responses related to oxidative stress and starvation response. Additionally, we find 50 stress-response genes upregulated in response to the microgravity when compared directly to the equivalent concentration in the ground control. We conclude that the increased antibiotic tolerance in microgravity may be attributed not only to diminished transport processes, but also to a resultant antibiotic cross-resistance response conferred by an overlapping effect of stress response genes. Our data suggest that direct stresses of nutrient starvation and acid-shock conveyed by the microgravity environment can incidentally upregulate stress response pathways related to antibiotic stress and in doing so contribute to the increased antibiotic stress tolerance observed for bacteria in space experiments. These results provide insights into the ability of bacteria to adapt under extreme stress conditions and potential strategies to prevent antimicrobial-resistance in space and on Earth.
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http://dx.doi.org/10.3389/fmicb.2018.00310DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5865062PMC
March 2018

α Cell Function and Gene Expression Are Compromised in Type 1 Diabetes.

Cell Rep 2018 03;22(10):2667-2676

Department of Medicine, Division of Diabetes, Endocrinology and Metabolism, Vanderbilt University Medical Center, Nashville, TN, USA; Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN, USA; Department of Veterans Affairs, Tennessee Valley Healthcare System, Nashville, TN, USA. Electronic address:

Many patients with type 1 diabetes (T1D) have residual β cells producing small amounts of C-peptide long after disease onset but develop an inadequate glucagon response to hypoglycemia following T1D diagnosis. The features of these residual β cells and α cells in the islet endocrine compartment are largely unknown, due to the difficulty of comprehensive investigation. By studying the T1D pancreas and isolated islets, we show that remnant β cells appeared to maintain several aspects of regulated insulin secretion. However, the function of T1D α cells was markedly reduced, and these cells had alterations in transcription factors constituting α and β cell identity. In the native pancreas and after placing the T1D islets into a non-autoimmune, normoglycemic in vivo environment, there was no evidence of α-to-β cell conversion. These results suggest an explanation for the disordered T1D counterregulatory glucagon response to hypoglycemia.
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http://dx.doi.org/10.1016/j.celrep.2018.02.032DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6368357PMC
March 2018

Age-dependent human β cell proliferation induced by glucagon-like peptide 1 and calcineurin signaling.

J Clin Invest 2017 Oct 18;127(10):3835-3844. Epub 2017 Sep 18.

Division of Diabetes, Endocrinology and Metabolism, Department of Medicine, Vanderbilt University Medical Center, Nashville, Tennessee, USA.

Inadequate pancreatic β cell function underlies type 1 and type 2 diabetes mellitus. Strategies to expand functional cells have focused on discovering and controlling mechanisms that limit the proliferation of human β cells. Here, we developed an engraftment strategy to examine age-associated human islet cell replication competence and reveal mechanisms underlying age-dependent decline of β cell proliferation in human islets. We found that exendin-4 (Ex-4), an agonist of the glucagon-like peptide 1 receptor (GLP-1R), stimulates human β cell proliferation in juvenile but not adult islets. This age-dependent responsiveness does not reflect loss of GLP-1R signaling in adult islets, since Ex-4 treatment stimulated insulin secretion by both juvenile and adult human β cells. We show that the mitogenic effect of Ex-4 requires calcineurin/nuclear factor of activated T cells (NFAT) signaling. In juvenile islets, Ex-4 induced expression of calcineurin/NFAT signaling components as well as target genes for proliferation-promoting factors, including NFATC1, FOXM1, and CCNA1. By contrast, expression of these factors in adult islet β cells was not affected by Ex-4 exposure. These studies reveal age-dependent signaling mechanisms regulating human β cell proliferation, and identify elements that could be adapted for therapeutic expansion of human β cells.
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http://dx.doi.org/10.1172/JCI91761DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5617654PMC
October 2017

Interrupted Glucagon Signaling Reveals Hepatic α Cell Axis and Role for L-Glutamine in α Cell Proliferation.

Cell Metab 2017 Jun;25(6):1362-1373.e5

Division of Diabetes, Endocrinology, and Metabolism, Vanderbilt University Medical Center, Nashville, TN 37232, USA; Department of Molecular Physiology and Biophysics, Vanderbilt University Medical Center, Nashville, TN 37232, USA; VA Tennessee Valley Healthcare, Nashville, TN 37212, USA. Electronic address:

Decreasing glucagon action lowers the blood glucose and may be useful therapeutically for diabetes. However, interrupted glucagon signaling leads to α cell proliferation. To identify postulated hepatic-derived circulating factor(s) responsible for α cell proliferation, we used transcriptomics/proteomics/metabolomics in three models of interrupted glucagon signaling and found that proliferation of mouse, zebrafish, and human α cells was mTOR and FoxP transcription factor dependent. Changes in hepatic amino acid (AA) catabolism gene expression predicted the observed increase in circulating AAs. Mimicking these AA levels stimulated α cell proliferation in a newly developed in vitro assay with L-glutamine being a critical AA. α cell expression of the AA transporter Slc38a5 was markedly increased in mice with interrupted glucagon signaling and played a role in α cell proliferation. These results indicate a hepatic α islet cell axis where glucagon regulates serum AA availability and AAs, especially L-glutamine, regulate α cell proliferation and mass via mTOR-dependent nutrient sensing.
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http://dx.doi.org/10.1016/j.cmet.2017.05.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5572896PMC
June 2017

A high-throughput molecular data resource for cutaneous neurofibromas.

Sci Data 2017 04 11;4:170045. Epub 2017 Apr 11.

Children's Tumour Foundation, New York, New York 10005, USA.

Neurofibromatosis type 1 (NF1) is a genetic disorder with a range of clinical manifestations such as widespread growth of benign tumours called neurofibromas, pain, learning disorders, bone deformities, vascular abnormalities and even malignant tumours. With the establishment of the Children's Tumour Foundation biobank, neurofibroma samples can now be collected directly from patients to be analysed by the larger scientific community. This work describes a pilot study to characterize one class of neurofibroma, cutaneous neurofibromas, by molecularly profiling of ~40 cutaneous neurofibromas collected from 11 individual patients. Data collected from each tumour includes (1) SNP Arrays, (2) Whole genome sequencing (WGS) and (3) RNA-Sequencing. These data are now freely available for further analysis at http://www.synapse.org/cutaneousNF.
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http://dx.doi.org/10.1038/sdata.2017.45DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5387919PMC
April 2017

Defining age- and lactocrine-sensitive elements of the neonatal porcine uterine microRNA-mRNA interactome.

Biol Reprod 2017 02;96(2):327-340

Department of Animal Sciences, Endocrinology, and Animal Biosciences Program, Rutgers University, New Brunswick, New Jersey, USA.

Factors delivered to offspring in colostrum within 2 days of birth support neonatal porcine uterine development. The uterine mRNA transcriptome is affected by age and nursing during this period. Whether uterine microRNA (miRNA) expression is affected similarly is unknown. Objectives were to (1) determine effects of age and nursing on porcine uterine miRNA expression between birth and postnatal day (PND) 2 using miRNA sequencing (miRNAseq) and; (2) define affected miRNA–mRNA interactions and associated biological processes using integrated target prediction analysis. At birth (PND 0), gilts were euthanized, nursed ad libitum, or gavage-fed milk replacer for 48 h. Uteri were collected at birth or 50 h postnatal. MicroRNAseq data were validated using quantitative real-time PCR. Targets were predicted using an established mRNA database generated from the same tissues. For PND 2 versus PND 0 comparisons, 31 differentially expressed (DE) miRNAs were identified for nursed, and 42 DE miRNAs were identified for replacer-fed gilts. Six DE miRNAs were identified for nursed versus replacer-fed gilts on PND 2. Target prediction for inversely correlated DE miRNA–mRNA pairings indicated 20 miRNAs targeting 251 mRNAs in nursed, versus 29 miRNAs targeting 585 mRNAs in replacer-fed gilts for PND 2 versus PND 0 comparisons, and 5 miRNAs targeting 81 mRNAs for nursed versus replacer-fed gilts on PND 2. Biological processes predicted to be affected by age and nursing included cell-to-cell signaling, cell morphology, and tissue morphology. Results indicate novel age- and lactocrine-sensitive miRNA–mRNA relationships associated with porcine neonatal uterine development between birth and PND 2
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http://dx.doi.org/10.1093/biolre/iox001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5819844PMC
February 2017

A signalling cascade of IL-33 to IL-13 regulates metaplasia in the mouse stomach.

Gut 2018 05 14;67(5):805-817. Epub 2017 Feb 14.

Departments of Cell and Developmental Biology, Vanderbilt University, Nashville, Tennessee, USA.

Objective: Alternatively activated macrophages (M2) are associated with the progression of spasmolytic polypeptide-expressing metaplasia (SPEM) in the stomach. However, the precise mechanism(s) and critical mediators that induce SPEM are unknown.

Design: To determine candidate genes important in these processes, macrophages from the stomach corpus of mice with SPEM (DMP-777-treated) or advanced SPEM (L635-treated) were isolated and RNA sequenced. Effects on metaplasia development after acute parietal cell loss induced by L635 were evaluated in interleukin (IL)-33, IL-33 receptor (ST2) and IL-13 knockout (KO) mice.

Results: Profiling of metaplasia-associated macrophages in the stomach identified an M2a-polarised macrophage population. Expression of IL-33 was significantly upregulated in macrophages associated with advanced SPEM. L635 induced metaplasia in the stomachs of wild-type mice, but not in the stomachs of IL-33 and ST2 KO mice. While IL-5 and IL-9 were not required for metaplasia induction, IL-13 KO mice did not develop metaplasia in response to L635. Administration of IL-13 to ST2 KO mice re-established the induction of metaplasia following acute parietal cell loss.

Conclusions: Metaplasia induction and macrophage polarisation after parietal cell loss is coordinated through a cytokine signalling network of IL-33 and IL-13, linking a combined response to injury by both intrinsic mucosal mechanisms and infiltrating M2 macrophages.
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http://dx.doi.org/10.1136/gutjnl-2016-312779DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5681443PMC
May 2018

Cell-Based Systems Biology Analysis of Human AS03-Adjuvanted H5N1 Avian Influenza Vaccine Responses: A Phase I Randomized Controlled Trial.

PLoS One 2017 18;12(1):e0167488. Epub 2017 Jan 18.

Vanderbilt Vaccine Research Program, Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN, United States of America.

Background: Vaccine development for influenza A/H5N1 is an important public health priority, but H5N1 vaccines are less immunogenic than seasonal influenza vaccines. Adjuvant System 03 (AS03) markedly enhances immune responses to H5N1 vaccine antigens, but the underlying molecular mechanisms are incompletely understood.

Objective And Methods: We compared the safety (primary endpoint), immunogenicity (secondary), gene expression (tertiary) and cytokine responses (exploratory) between AS03-adjuvanted and unadjuvanted inactivated split-virus H5N1 influenza vaccines. In a double-blinded clinical trial, we randomized twenty adults aged 18-49 to receive two doses of either AS03-adjuvanted (n = 10) or unadjuvanted (n = 10) H5N1 vaccine 28 days apart. We used a systems biology approach to characterize and correlate changes in serum cytokines, antibody titers, and gene expression levels in six immune cell types at 1, 3, 7, and 28 days after the first vaccination.

Results: Both vaccines were well-tolerated. Nine of 10 subjects in the adjuvanted group and 0/10 in the unadjuvanted group exhibited seroprotection (hemagglutination inhibition antibody titer > 1:40) at day 56. Within 24 hours of AS03-adjuvanted vaccination, increased serum levels of IL-6 and IP-10 were noted. Interferon signaling and antigen processing and presentation-related gene responses were induced in dendritic cells, monocytes, and neutrophils. Upregulation of MHC class II antigen presentation-related genes was seen in neutrophils. Three days after AS03-adjuvanted vaccine, upregulation of genes involved in cell cycle and division was detected in NK cells and correlated with serum levels of IP-10. Early upregulation of interferon signaling-related genes was also found to predict seroprotection 56 days after first vaccination.

Conclusions: Using this cell-based systems approach, novel mechanisms of action for AS03-adjuvanted pandemic influenza vaccination were observed.

Trial Registration: ClinicalTrials.gov NCT01573312.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0167488PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5242433PMC
August 2017

Modulation of microRNA-mRNA Target Pairs by Human Papillomavirus 16 Oncoproteins.

mBio 2017 01 3;8(1). Epub 2017 Jan 3.

Department of Developmental, Molecular and Chemical Biology, Tufts University School of Medicine, Boston, Massachusetts, USA

The E6 and E7 proteins are the major oncogenic drivers encoded by high-risk human papillomaviruses (HPVs). While many aspects of the transforming activities of these proteins have been extensively studied, there are fewer studies that have investigated how HPV E6/E7 expression affects the expression of cellular noncoding RNAs. The goal of our study was to investigate HPV16 E6/E7 modulation of cellular microRNA (miR) levels and to determine the potential consequences for cellular gene expression. We performed deep sequencing of small and large cellular RNAs in primary undifferentiated cultures of human foreskin keratinocytes (HFKs) with stable expression of HPV16 E6/E7 or a control vector. After integration of the two data sets, we identified 51 differentially expressed cellular miRs associated with the modulation of 1,456 potential target mRNAs in HPV16 E6/E7-expressing HFKs. We discovered that the degree of differential miR expression in HFKs expressing HPV16 E6/E7 was not necessarily predictive of the number of corresponding mRNA targets or the potential impact on gene expression. Additional analyses of the identified miR-mRNA pairs suggest modulation of specific biological activities and biochemical pathways. Overall, our study supports the model that perturbation of cellular miR expression by HPV16 E6/E7 importantly contributes to the rewiring of cellular regulatory circuits by the high-risk HPV E6 and E7 proteins that contribute to oncogenic transformation.

Importance: High-risk human papillomaviruses (HPVs) are the causative agents of almost all cervical cancers and many other cancers, including anal, vaginal, vulvar, penile, and oropharyngeal cancers. Despite the availability of efficacious HPV vaccines, it is critical to determine how HPVs cause cancer, as many people remain unvaccinated and the vaccine does not prevent cancer development in individuals who are already infected. Two HPV proteins, E6 and E7, are the major drivers of cancer development, and much remains to be learned about how the expression of these viral proteins reprograms infected cells, ultimately resulting in cancer development. Small, noncoding human RNAs, termed microRNAs (miRs), regulate gene expression and have been implicated in almost all human cancers, including HPV-associated cancers. Our study provides a comprehensive analysis of how E6 and E7 alter the expression of human miRs and how this potentially impacts cellular gene expression, which may contribute to cancer development.
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http://dx.doi.org/10.1128/mBio.02170-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5210503PMC
January 2017

A comparison of microRNA expression profiles from splenic hemangiosarcoma, splenic nodular hyperplasia, and normal spleens of dogs.

BMC Vet Res 2016 Dec 3;12(1):272. Epub 2016 Dec 3.

Scott Ritchey Research Center, Auburn University College of Veterinary Medicine, Auburn University, Auburn, AL, USA.

Background: Splenic masses are common in older dogs; yet diagnosis preceding splenectomy and histopathology remains elusive. MicroRNAs (miRNAs) are short, non-coding RNAs that play a role in post-transcriptional regulation, and differential expression of miRNAs between normal and tumor tissue has been used to diagnose neoplastic diseases. The objective of this study was to determine differential expression of miRNAs by use of RNA-sequencing in canine spleens that were histologically confirmed as hemangiosarcoma, nodular hyperplasia, or normal.

Results: Twenty-two miRNAs were found to be differentially expressed in hemangiosarcoma samples (4 between hemangiosarcoma and both nodular hyperplasia and normal spleen and 18 between hemangiosarcoma and normal spleen only). In particular, mir-26a, mir-126, mir-139, mir-140, mir-150, mir-203, mir-424, mir-503, mir-505, mir-542, mir-30e, mir-33b, mir-365, mir-758, mir-22, and mir-452 are of interest in the pathogenesis of hemangiosarcoma.

Conclusions: Findings of this study confirm the hypothesis that miRNA expression profiles are different between canine splenic hemangiosarcoma, nodular hyperplasia, and normal spleens. A large portion of the differentially expressed miRNAs have roles in angiogenesis, with an additional group of miRNAs being dysregulated in vascular disease processes. Two other miRNAs have been implicated in cancer pathways such as PTEN and cell cycle checkpoints. The finding of multiple miRNAs with roles in angiogenesis and vascular disease is important, as hemangiosarcoma is a tumor of endothelial cells, which are driven by angiogenic stimuli. This study shows that miRNA dysregulation is a potential player in the pathogenesis of canine splenic hemangiosarcoma.
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http://dx.doi.org/10.1186/s12917-016-0903-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5135805PMC
December 2016
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