Multiple Cell Systems Research Articles

STEM CELLS HOMEOSTASIS AND AGING

Abstract Adult (tissue) stem cells support tissue homeostasis and repair throughout the life of an individual. Numerous changes occur with age that result in altered stem cell behavior and reduced tissue maintenance and regeneration. Changes can be cell autonomous including changes in cell cycle progression, decreased bioenergetic efficiency, increased DNA damage, and epigenetic alterations. In addition, poorly understood changes to the local and systemic environments occur that result in decreased stem cell activity or alterations in commitment or differentiation potential. This symposium will cover the impact of aging on multiple adult stem cell systems, including hematopoietic stem cells (HSCs), intestinal stem cells (ISCs), and neural stem cells.

Thermal runaway behavior of nickel–manganese–cobalt 18650 lithium-ion cells induced by internal and external heating failures

Lithium-ion cells can fail in thermal runaway in a number of ways. Thermally-induced failures can be initiated by either external heating or an internal failure which leads to internal heating. Internal failures such as dendrite growth can lead to an internal short circuit (ISC) which subsequently causes internal heating. Internal failures are a common cause for lithium-ion battery thermal runaway that leads to fire incidents. Preventing and mitigating thermal runaway failure is of great interest in battery fire safety research and development. To evaluate safety measures in a repeatable manner, there is a need for a simple and repeatable technique to reliably fail a given cell, while minimally affecting the pack prior to cell failure. This paper introduces an internal failure mechanism targeted on a single (or multiple) 18650 cell(s) to trigger thermal runaway by (1) modifying commercially acquired 18650s to accommodate a mini-heater cartridge, (2) performing cycling tests on the modified cells and comparing their performance to that of the as-received (pristine) cells in a custom-built failure tube environment, and (3) experimentally examining the characteristics (temperature, flux-time product, voltage drop, and mass loss) of internally heated and failed cells in comparison to externally heated and failed cells. The results show that the modification procedures do not alter a cell’s performance and its charge capacity. Failure tube tests show that the thermal runaway behavior and damage to the modified cells are representative of ”real-world” cell failures. The internally heated cells had different venting behavior relative to externally heated cells, which affects mass loss profiles. When inserting a single, modified cell into a battery package (laptop power bank) and triggering it to cause thermal runaway, failure propagation to the surrounding cells can be characterized. Overall, this study introduces a realistic cell internal failure approach that can benefit the development of battery safety techniques in a variety of applications. The internal-heater failure approach is proven to be easily prepared with standard laboratory apparatus without affecting the battery performance and produces repeatable failure results for single and multiple cell systems.

Biomechanics of cells and subcellular components: A comprehensive review of computational models and applications.

Cells are a fundamental structural, functional and biological unit for all living organisms. Up till now, considerable efforts have been made to study the responses of single cells and subcellular components to an external load, and understand the biophysics underlying cell rheology, mechanotransduction and cell functions using experimental and in silico approaches. In the last decade, computational simulation has become increasingly attractive due to its critical role in interpreting experimental data, analysing complex cellular/subcellular structures, facilitating diagnostic designs and therapeutic techniques, and developing biomimetic materials. Despite the significant progress, developing comprehensive and accurate models of living cells remains a grand challenge in the 21st century. To understand current state of the art, this review summarises and classifies the vast array of computational biomechanical models for cells. The article covers the cellular components at multi-spatial levels, that is, protein polymers, subcellular components, whole cells and the systems with scale beyond a cell. In addition to the comprehensive review of the topic, this article also provides new insights into the future prospects of developing integrated, active and high-fidelity cell models that are multiscale, multi-physics and multi-disciplinary in nature. This review will be beneficial for the researchers in modelling the biomechanics of subcellular components, cells and multiple cell systems and understanding the cell functions and biological processes from the perspective of cell mechanics.

Metabolomics reveals synergy between Ag and g-C3N4 in Ag/g-C3N4 composite photocatalysts: a unique feature among Ag-doped biocidal materials.

The silver/graphitic carbon nitride (Ag/g-C3N4) composite system exerts biocidal activity against the pathogenic bacterium Escherichia coli 1337-H that is stronger than that of well-known silver and titanium oxide (TiO2)-based composites. However, whether the Ag/g-C3N4 composite system has biocidal properties that the parent components do or do not have as separate chemical entities and whether they differ from those in Ag/TiO2 composite photocatalysts have not been clarified. We investigated the chemical (cooperative charge handling and electronic properties) and biological (metabolic) effects exerted by the addition of Ag to g-C3N4 and to TiO2. In this work, we undertook metabolome-wide analysis by liquid chromatography-electrospray ionization-quadrupole-time of flight-mass spectrometry to compare the metabolite profiles of untreated E. coli 1337-H cells or those subjected to disinfection with Ag, g-C3N4, 2Ag/g-C3N4, TiO2 and 2Ag/TiO2. While Ag or g-C3N4 moderately affected microbial metabolism according to the mean of the altered metabolites, multiple cell systems contributing to rapid cell death were immediately affected by the light-triggered radical species produced when Ag and g-C3N4 were as xAg/g-C3N4. The effects include drastically reduced production of small metabolites essential for detoxifying reactive oxygen species and those that regulate DNA replication fidelity, cell morphology and energy status. These biological consequences were different from those caused by Ag/TiO2-based biocides, demonstrating the uniqueness of the Ag/g-C3N4 system. Our results support the idea that the unique Ag/g-C3N4 biocidal properties are based on synergistic action and reveal new directions for designing future photocatalysts for use in disinfection and microbial control.

Regulation of NF-κB, FHC and SOD2 in response to oxidative stress in the freeze tolerant wood frog, Rana sylvatica

The wood frog, Rana sylvatica, is the primary model animal used for studying vertebrate freeze tolerance. Freeze tolerance adaptations by wood frogs are mediated by a set of well-tuned regulatory controls at the molecular level, starting from cell signal transduction and gene expression events that are ultimately reflected in protective responses by multiple cell systems. Previous studies provided excellent presumptive evidence for the involvement of the NF-κB transcription factor in freeze tolerance. The present study of the NF-κB pathway focussed on freezing time points, 4 h frozen and 24 h frozen for liver and skeletal muscle in wood frog. The total protein levels of the major NF-κB subunits p50 and p65, its inhibitor, p-IκB, and downstream targets, ferritin heavy chain (FHC) and manganese superoxide dismutase (MnSOD) were quantified using western blots. Results showed a significant increase in the levels of NF-κB subunits and its downstream targets during freezing. Nuclear distributions of NF-κB subunits and transcript levels of FHC were analysed to delve deeper into the pathway. Results obtained from nuclear distribution analysis were consistent with the total protein levels showing increased levels of p50 and p65 during 24 h freezing conditions compared to controls but no change in phospho-p65 levels. Further, FHC transcript levels increased in 24 h frozen liver but did not change in frozen muscles. These findings suggest the activation of NF-κB antioxidant defenses in wood frogs during freezing in potential anticipation of high oxidative stress during reperfusion during thawing.

RNA-seq Profiling and Co-expression Network Analysis of Long Noncoding RNAs and mRNAs Reveal Novel Pathogenesis of Noise-induced Hidden Hearing Loss

Noise-induced hidden hearing loss (NIHHL), one of the family of conditions described as noise-induced hearing loss (NIHL), is characterized by synaptopathy following moderate noise exposure that causes only temporary threshold elevation. Long noncoding RNAs (lncRNAs) mediate several essential regulatory functions in a wide range of biological processes and diseases, but their roles in NIHHL remain largely unknown. In order to determine the potential roles of these lncRNAs in the pathogenesis of NIHHL, we first evaluated their expression in NIHHL mice model and mapped possible regulatory functions and targets using RNA-sequencing (RNA-seq). In total, we identified 133 lncRNAs and 522 mRNAs that were significantly dysregulated in the NIHHL model. Gene Ontology (GO) showed that these lncRNAs were involved in multiple cell components and systems including synapses and the nervous and sensory systems. In addition, a lncRNA–mRNA network was constructed to identify core regulatory lncRNAs and transcription factors. KEGG analysis was also used to identify the potential pathways being affected in NIHHL. These analyses allowed us to identify the guanine nucleotide binding protein alpha stimulating (GNAS) gene as a key transcription factor and the adrenergic signaling pathway as a key pathway in the regulation of NIHHL pathogenesis. Our study is the first, to our knowledge, to isolate a lncRNA mediated regulatory pathway associated with NIHHL pathogenesis; these observations may provide fresh insight into the pathogenesis of NIHHL and may pave the way for therapeutic intervention in the future.

BMP signaling mediates glioma stem cell quiescence and confers treatment resistance in glioblastoma

Despite advances in therapy, glioblastoma remains an incurable disease with a dismal prognosis. Recent studies have implicated cancer stem cells within glioblastoma (glioma stem cells, GSCs) as mediators of therapeutic resistance and tumor progression. In this study, we investigated the role of the transforming growth factor-β (TGF-β) superfamily, which has been found to play an integral role in the maintenance of stem cell homeostasis within multiple stem cell systems, as a mediator of stem-like cells in glioblastoma. We find that BMP and TGF-β signaling define divergent molecular and functional identities in glioblastoma, and mark relatively quiescent and proliferative GSCs, respectively. Treatment of GSCs with BMP inhibits cell proliferation, but does not abrogate their stem-ness, as measured by self-renewal and tumorigencity. Further, BMP pathway activation confers relative resistance to radiation and temozolomide chemotherapy. Our findings define a quiescent cancer stem cell population in glioblastoma that may be a cellular reservoir for tumor recurrence following cytotoxic therapy.

Hemoglobin Expression and Function in Human Corneal Epithelium

The corneal epithelium forms the outer layer of the cornea. It provides mechanical protection, prevents fluid loss and forms a barrier to invasive pathogens. Though hemoglobin expression has been extensively studied in erythroid cells, recent work in multiple cell systems has documented hemoglobin expression in cells of non-erythroid origin. However, the function of hemoglobin in non-erythroid cells has yet to be established. The hypothesis that hemoglobin is expressed in corneal epithelium and that it functions to protect against oxidative stress will be examined in the present work. Hemoglobin expression and function was examined by immunocytochemistry and western blot analysis. Native human corneal explants and an immortalized human corneal epithelial cell line were examined. Expression of hemoglobin beta and delta chains was demonstrated at the protein level in both native and cell culture preparations. Hexamethylene bisacetamide is a known inducer of hemoglobin beta chain expression in murine erythroleukemia cells. HMBA did function to increase beta chain expression in cultured corneal epithelium as well. Beta chain immunolocalization was primarily cytoplasmic, while the delta chain localized to both cytoplasmic and membrane domains. Oxidative stress, from hydrogen peroxide exposure, was shown to upregulate delta chain expression. In conclusion, hemoglobin chains are expressed in corneal epithelium and could function to protect against oxidative stress. This is relevant given that exposure to light and high oxygen tensions render the cornea particularly susceptible to oxidative damage.

Hemoglobin Expression and Function in Human Corneal Epithelium

The corneal epithelium forms the outer layer of the cornea. It provides mechanical protection, prevents fluid loss and forms a barrier to invasive pathogens. Though hemoglobin expression has been extensively studied in erythroid cells, recent work in multiple cell systems has documented hemoglobin expression in cells of non-erythroid origin. However, the function of hemoglobin in non-erythroid cells has yet to be established. The hypothesis that hemoglobin is expressed in corneal epithelium and that it functions to protect against oxidative stress will be examined in the present work. Hemoglobin expression and function was examined by immunocytochemistry and western blot analysis. Native human corneal explants and an immortalized human corneal epithelial cell line were examined. Expression of hemoglobin beta and delta chains was demonstrated at the protein level in both native and cell culture preparations. Hexamethylene bisacetamide is a known inducer of hemoglobin beta chain expression in murine erythroleukemia cells. HMBA did function to increase beta chain expression in cultured corneal epithelium as well. Beta chain immunolocalization was primarily cytoplasmic, while the delta chain localized to both cytoplasmic and membrane domains. Oxidative stress, from hydrogen peroxide exposure, was shown to upregulate delta chain expression. In conclusion, hemoglobin chains are expressed in corneal epithelium and could function to protect against oxidative stress. This is relevant given that exposure to light and high oxygen tensions render the cornea particularly susceptible to oxidative damage.

1026 - JAK/STAT SIGNALLING, STEM CELL SUBVERSION AND MYELOID MALIGNANCIES

Our research focuses on the mechanisms whereby blood stem cells are subverted during the genesis of haematological malignancies. We have increasingly concentrated on JAK/STAT signalling which is dysregulated in many cancers and plays a key role in multiple stem cell systems. In particular we have explored the molecular and cellular pathogenesis of a group of pre-leukaemic disorders, the myeloproliferative neoplasms (MPNs), in studies which have spanned basic, translational and clinical research. These disorders are experimentally tractable and provide a model for the earliest stages of tumorigenesis, inaccessible in other cancers. In work which transformed MPN diagnosis and catalysed development of therapeutic JAK inhibitors, we and others identified phenotypic driver mutations in JAK2 and CALR which activate the JAK/STAT pathway and are present in most MPN patients. We are employing a variety of genomic approaches to explore MPN biology and improve patient management. In parallel we are investigating the functional consequences of JAK2 and CALR mutations in work which has led to unexpected insights into cancer biology as well as normal cytokine signalling. Recent highlights include: (i) first demonstration in any cancer that mutation order affects stem and progenitor behaviour, thus influencing clinical presentation, disease outcome and response to therapy (Ortmann et al NEJM 2015); (ii) description of unexpected non-canonical mechanisms of JAK/STAT signalling (Dawson et al Nature 2009; Dawson et al Cell Reports 2013); Park et al EMBO J 2016); (iii) a new MPN classification based on causal biological mechanisms and the development of personalised predictions tailored to individual patients (Grinfeld et al NEJM 2018); (iv) the use of somatic mutations as clonal markers to reconstruct the cellular ancestry of human haematopoiesis as a single phylogenetic tree with the fertilized egg at its root (Lee-Six et al Nature 2018). This latter approach is broadly applicable and allows interrogation of human stem cell dynamics in young and old, in health and disease.

CiliaCarta: An integrated and validated compendium of ciliary genes.

The cilium is an essential organelle at the surface of mammalian cells whose dysfunction causes a wide range of genetic diseases collectively called ciliopathies. The current rate at which new ciliopathy genes are identified suggests that many ciliary components remain undiscovered. We generated and rigorously analyzed genomic, proteomic, transcriptomic and evolutionary data and systematically integrated these using Bayesian statistics into a predictive score for ciliary function. This resulted in 285 candidate ciliary genes. We generated independent experimental evidence of ciliary associations for 24 out of 36 analyzed candidate proteins using multiple cell and animal model systems (mouse, zebrafish and nematode) and techniques. For example, we show that OSCP1, which has previously been implicated in two distinct non-ciliary processes, causes ciliogenic and ciliopathy-associated tissue phenotypes when depleted in zebrafish. The candidate list forms the basis of CiliaCarta, a comprehensive ciliary compendium covering 956 genes. The resource can be used to objectively prioritize candidate genes in whole exome or genome sequencing of ciliopathy patients and can be accessed at http://bioinformatics.bio.uu.nl/john/syscilia/ciliacarta/.

An injectable continuous stratified structurally and functionally biomimetic construct for enhancing osteochondral regeneration

Osteochondral regeneration with the formation of hyaline cartilage and subchondral bone as well as the integration between the newly formed tissues with the host tissue still remains a great challenge. In this study, a construct containing an injectable continuous stratified scaffold and multiple cell systems was designed for enhancing osteochondral regeneration. Briefly, an injectable sodium alginate(SA)/bioglass (BG) composite hydrogel containing bone marrow stem cells (BMSCs) (SA/BG + BMSCs) was used for subchondral bone regeneration and an injectable thermosensitive SA/agarose (AG) composite hydrogel with co-culture of BMSCs and articular chondrocytes (ACs) (SA/AG + ACs/BMSCs) was applied for articular cartilage regeneration. The continuous SA phase and the stratified structure enable the scaffold to mimic the natural osteochondral structure. In addition, the SA/BG + BMSCs hydrogel could enhance the osteoblast differentiation of BMSCs by upregulating their alkaline phosphatase and collagen I gene expressions, and the SA/AG + ACs/BMSCs hydrogel could promote the chondrocyte differentiation of BMSCs by upregulating their Acan and collagen II gene expressions, which indicated that this stratified scaffold could mimic the natural osteochondral function. Furthermore, after the stratified construct was injected into a rat osteochondral defect model, obvious neonatal articular cartilage tissues and subchondral bone tissues with regular surface and highly integration with normal tissues could be observed. This structural and functional biomimetic construct, together with its proper swelling ratio, could not only stimulate the hyaline cartilage and subchondral bone regeneration in an entire osteochondral unit but also promote the integration between the newly formed tissues and the host tissue.

A new conceptual framework for the therapy by optimized multidimensional pulses of therapeutic activity. The case of multiple myeloma model

We developed simulation methodology to assess eventual therapeutic efficiency of exogenous multiparametric changes in a four-component cellular system described by the system of ordinary differential equations. The method is numerically implemented to simulate the temporal behavior of a cellular system of multiple myeloma cells. The problem is conceived as an inverse optimization task where the alternative temporal changes of selected parameters of the ordinary differential equations represent candidate solutions and the objective function quantifies the goals of the therapy. The system under study consists of two main cellular components, tumor cells and their cellular environment, respectively. The subset of model parameters closely related to the environment is substituted by exogenous time dependencies - therapeutic pulses combining continuous functions and discrete parameters subordinated thereafter to the optimization. Synergistic interaction of temporal parametric changes has been observed and quantified whereby two or more dynamic parameters show effects that absent if either parameter is stimulated alone. We expect that the theoretical insight into unstable tumor growth provided by the sensitivity and optimization studies could, eventually, help in designing combination therapies.

Deiodinases and stem cells: an intimate relationship.

Thyroid hormone is a major determinant of tissue functions in vivo. The deiodinase family controls the tissue-specific activation or inactivation of intracellular thyroid hormones. Precise control of the T3-dependent transcriptional program is required by multiple cell systems, including the stem cell. In this context, the identification of a close connection between thyroid hormones and different signal pathways involved in the control of stem cell functions suggested that the deiodinases may play a role in the definition of stem cell biology and physiology. Stem cells have an unlimited self-renewal capacity and the potential to differentiate into different types of mature cells. Deciphering how all these events are achieved, how the T3 signal is controlled and integrated in stem cells and their niches, and how it can impact on them is essentially unknown and represents a challenge for coming years. In this review, I will explore the role played by the deiodinases in the modulation of the TH signal in stem cells of adult tissues, namely muscle and intestine, and how their actions control the delicate balance among self-renewal, proliferation and differentiation. Elucidation of the molecular mechanisms presiding thyroid hormone action in stem cells may reveal therapeutic potential, for example in the fields of regenerative diseases and cancer.

Bacterial Sphingomyelinase is a State-Dependent Inhibitor of the Cystic Fibrosis Transmembrane conductance Regulator (CFTR)

Sphingomyelinase C (SMase) inhibits CFTR chloride channel activity in multiple cell systems, an effect that could exacerbate disease in CF and COPD patients. The mechanism by which sphingomyelin catalysis inhibits CFTR is not known but evidence suggests that it occurs independently of CFTR’s regulatory “R” domain. In this study we utilized the Xenopus oocyte expression system to shed light on how CFTR channel activity is reduced by SMase. We found that the pathway leading to inhibition is not membrane delimited and that inhibited CFTR channels remain at the cell membrane, indicative of a novel silencing mechanism. Consistent with an effect on CFTR gating behavior, we found that altering gating kinetics influenced the sensitivity to inhibition by SMase. Specifically, increasing channel activity by introducing the mutation K1250A or pretreating with the CFTR potentiator VX-770 (Ivacaftor) imparted resistance to inhibition. In primary bronchial epithelial cells, we found that basolateral, but not apical, application of SMase leads to a redistribution of sphingomyelin and a reduction in forskolin- and VX-770-stimulated currents. Taken together, these data suggest that SMase inhibits CFTR channel function by locking channels into a closed state and that endogenous CFTR in HBEs is affected by SMase activity.

MiRNA-520f Reverses Epithelial-to-Mesenchymal Transition by Targeting ADAM9 and TGFBR2.

Reversing epithelial-to-mesenchymal transition (EMT) in cancer cells has been widely considered as an approach to combat cancer progression and therapeutic resistance, but a limited number of broadly comprehensive investigations of miRNAs involved in this process have been conducted. In this study, we screened a library of 1120 miRNA for their ability to transcriptionally activate the E-cadherin gene CDH1 in a promoter reporter assay as a measure of EMT reversal. By this approach, we defined miR-520f as a novel EMT-reversing miRNA. miR-520f expression was sufficient to restore endogenous levels of E-cadherin in cancer cell lines exhibiting strong or intermediate mesenchymal phenotypes. In parallel, miR-520f inhibited invasive behavior in multiple cancer cell systems and reduced metastasis in an experimental mouse model of lung metastasis. Mechanistically, miR-520f inhibited tumor cell invasion by directly targeting ADAM9, the TGFβ receptor TGFBR2 and the EMT inducers ZEB1, ZEB2, and the snail transcriptional repressor SNAI2, each crucial factors in mediating EMT. Collectively, our results show that miR-520f exerts anti-invasive and antimetastatic effects in vitro and in vivo, warranting further study in clinical settings. Cancer Res; 77(8); 2008-17. ©2017 AACR.

Interferon epsilon promotes HIV restriction at multiple steps of viral replication.

Interferon epsilon (IFNɛ) is a type I IFN that is expressed constitutively in the female reproductive tract (FRT), and contributes to protection in models of sexually transmitted infections. Using multiple cell systems, including reporter cell lines and activated peripheral blood lymphocytes (PBLs), we show that recombinant IFNɛ impairs HIV infection at stage(s) post HIV entry and up to the translation of viral proteins. Consistent with this, IFNɛ upregulated a number of host cell restriction factors that block HIV at these stages of the replication cycle. The potency of IFNɛ induction of these HIV restriction factors was comparable to conventional type I IFNs, namely IFNα and IFNβ. IFNɛ also significantly reduced the infectivity of progeny virion particles likely by inducing expression of HIV restriction factors, such as IFITM3, which act at that stage of infection. Thus, our data demonstrate that human IFNɛ suppresses HIV replication at multiple stages of infection.

Novel Hits in the Correction of ΔF508-Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Protein: Synthesis, Pharmacological, and ADME Evaluation of Tetrahydropyrido[4,3-d]pyrimidines for the Potential Treatment of Cystic Fibrosis.

Cystic fibrosis (CF) is a lethal genetic disease caused by mutations of the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) with a prevalence of the ΔF508 mutation. Whereas the detailed mechanisms underlying disease have yet to be fully elucidated, recent breakthroughs in clinical trials have demonstrated that CFTR dysfunction can be corrected by drug-like molecules. On the basis of this success, a screening campaign was carried out, seeking new drug-like compounds able to rescue ΔF508-CFTR that led to the discovery of a novel series of correctors based on a tetrahydropyrido[4,3-d]pyrimidine core. These molecules proved to be soluble, cell-permeable, and active in a disease relevant functional-assay. The series was then further optimized with emphasis on biological data from multiple cell systems while keeping physicochemical properties under strict control. The pharmacological and ADME profile of this corrector series hold promise for the development of more efficacious compounds to be explored for therapeutic use in CF.

Rapid combinatorial screening of inkjet-printed alkyl-ammonium cations in perovskite solar cells

Herein we demonstrate that efficient perovskite solar cells can be fabricated by a desktop multi-channel inkjet printer. In this approach, different alkyl-ammonium counterions were loaded separately into the printer cartridges and deposited onto PbI2 coated substrates. Structural as well as electronic properties of perovskite solar cells can be tuned by varying the composition ratio of multiple cations in-situ through the RGB color codes of the multi-channel inkjet printer. This roll-to-roll compatible technique provides rapid and facile screening of multiple and mixed counterion perovskite solar cell systems with high reproducibility, potentially leading to possibilities of high-throughput screening technique in future perovskite solar cell fabrication.

Activation of TLR3/interferon signaling pathway by bluetongue virus results in HIV inhibition in macrophages.

Bluetongue virus (BTV), a nonenveloped double-stranded RNA virus, is a potent inducer of type Ι interferons in multiple cell systems. In this study, we report that BTV16 treatment of primary human macrophages induced both type I and III IFN expression, resulting in the production of multiple antiviral factors, including myxovirus resistance protein A, 2',5'-oligoadenylate synthetase, and the IFN-stimulated gene 56. Additionally, BTV-treated macrophages expressed increased HIV restriction factors (apolipoprotein B mRNA-editing enzyme catalytic polypeptide 3 G/F/H) and CC chemokines (macrophage inflammatory protein 1-α, macrophage inflammatory protein 1-β, regulated on activation of normal T cell expressed and secreted), the ligands for HIV entry coreceptor CC chemokine receptor type 5. BTV16 also induced the expression of tetherin, which restricts HIV release from infected cells. Furthermore, TLR3 signaling of macrophages by BTV16 resulted in the induction of several anti-HIV microRNAs (miRNA-28, -29a, -125b, -150, -223, and -382). More importantly, the induction of antiviral responses by BTV resulted in significant suppression of HIV in macrophages. These findings demonstrate the potential of BTV-mediated TLR3 activation in macrophage innate immunity against HIV.