Publications by authors named "Dallas Jones"

45 Publications

Human innate lymphoid cell precursors express CD48 that modulates ILC differentiation through 2B4 signaling.

Sci Immunol 2020 11;5(53)

Department of Pediatric, Division of Children's Cancer and Blood Disorders, University of Colorado and Children's Hospital of Colorado, Research Complex 1, North Tower, 12800 E. 19th Ave., Mail Stop 8302, Room P18-4108, Aurora, CO 80045, USA.

Innate lymphoid cells (ILCs) develop from common lymphoid progenitors (CLPs), which further differentiate into the common ILC progenitor (CILP) that can give rise to both ILCs and natural killer (NK) cells. Murine ILC intermediates have recently been characterized, but the human counterparts and their developmental trajectories have not yet been identified, largely due to the lack of homologous surface receptors in both organisms. Here, we show that human CILPs (CD34CD117α4β7Lin) acquire CD48 and CD52, which define NK progenitors (NKPs) and ILC precursors (ILCPs). Two distinct NK cell subsets were generated in vitro from CD34CD117α4β7LinCD48CD52 and CD34CD117α4β7LinCD48CD52 NKPs, respectively. Independent of NKPs, ILCPs exist in the CD34CD117α4β7LinCD48CD52 subset and give rise to ILC1s, ILC2s, and NCR ILC3s, whereas CD34CD117α4β7LinCD48CD52 ILCPs give rise to a distinct subset of ILC3s that have lymphoid tissue inducer (LTi)-like properties. In addition, CD48-expressing CD34CD117α4β7Lin precursors give rise to tissue-associated ILCs in vivo. We also observed that the interaction of 2B4 with CD48 induced differentiation of ILC2s, and together, these findings show that expression of CD48 by human ILCPs modulates ILC differentiation.
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http://dx.doi.org/10.1126/sciimmunol.aay4218DOI Listing
November 2020

Biomechanical and Bone Material Properties of Schnurri-3 Null Mice.

JBMR Plus 2019 Nov 11;3(11):e10226. Epub 2019 Sep 11.

1st Medical Department Hanusch Hospital Ludwig Boltzmann Institute of Osteology at the Hanusch Hospital of WGKK and AUVA Trauma Centre Meidling Vienna Austria.

is an essential regulator of postnatal skeletal remodeling. Shn3-deficient mice (Shn3-/-) have high bone mass; however, their bone mechanical and material properties have not been investigated to date. We performed three-point bending of femora, compression tests of L3 vertebrae. We also measured intrinsic material properties, including bone mineralization density distribution (BMDD) and osteocyte lacunae section (OLS) characteristics by quantitative backscatter electron imaging, as well as collagen cross-linking by Fourier transform infrared microspectroscopy of femora from Shn3-/- and WT mice at different ages (6 weeks, 4 months, and 18 months). Moreover, computer modeling was performed for the interpretation of the BMDD outcomes. Femora and L3 vertebrae from Shn3-/- aged 6 weeks revealed increased ultimate force (2.2- and 3.2-fold, < .01, respectively). Mineralized bone volume at the distal femoral metaphysis was about twofold (at 6 weeks) to eightfold (at 4 and 18 months of age) in Shn3-/- ( < .001). Compared with WT, the average degree of trabecular bone mineralization was similar at 6 weeks, but increased at 4 and 18 months of age (+12.6% and +7.7%, < .01, respectively) in Shn3-/-. The analysis of OLS characteristics revealed a higher OLS area for Shn3-/- versus WT at all ages (+16%, +23%, +21%, respectively, < .01). The collagen cross-link ratio was similar between groups. We conclude that femora and vertebrae from Shn3-/- had higher ultimate force in mechanical testing. Computer modeling demonstrated that in cases of highly increased bone volume, the average degree of bone matrix mineralization can be higher than in WT bone, which was actually measured in the older Shn3-/- groups. The area of 2D osteocyte lacunae sections was also increased in Shn3-deficiency, which could only partly be explained by larger remnant areas of primary cortical bone. © 2019 The Authors. published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.
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http://dx.doi.org/10.1002/jbm4.10226DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6874182PMC
November 2019

Varicella zoster virus productively infects human peripheral blood mononuclear cells to modulate expression of immunoinhibitory proteins and blocking PD-L1 enhances virus-specific CD8+ T cell effector function.

PLoS Pathog 2019 03 14;15(3):e1007650. Epub 2019 Mar 14.

Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado, United States of America.

Varicella zoster virus (VZV) is a lymphotropic alpha-herpesvirinae subfamily member that produces varicella on primary infection and causes zoster, vascular disease and vision loss upon reactivation from latency. VZV-infected peripheral blood mononuclear cells (PBMCs) disseminate virus to distal organs to produce clinical disease. To assess immune evasion strategies elicited by VZV that may contribute to dissemination of infection, human PBMCs and VZV-specific CD8+ T cells (V-CD8+) were mock- or VZV-infected and analyzed for immunoinhibitory protein PD-1, PD-L1, PD-L2, CTLA-4, LAG-3 and TIM-3 expression using flow cytometry. All VZV-infected PBMCs (monocytes, NK, NKT, B cells, CD4+ and CD8+ T cells) and V-CD8+ showed significant elevations in PD-L1 expression compared to uninfected cells. VZV induced PD-L2 expression in B cells and V-CD8+. Only VZV-infected CD8+ T cells, NKT cells and V-CD8+ upregulated PD-1 expression, the immunoinhibitory receptor for PD-L1/PD-L2. VZV induced CTLA-4 expression only in V-CD8+ and no significant changes in LAG-3 or TIM-3 expression were observed in V-CD8+ or PBMC T cells. To test whether PD-L1, PD-L2 or CTLA-4 regulates V-CD8+ effector function, autologous PBMCs were VZV-infected and co-cultured with V-CD8+ cells in the presence of blocking antibodies against PD-L1, PD-L2 or CTLA-4; ELISAs revealed significant elevations in IFNγ only upon blocking of PD-L1. Together, these results identified additional immune cells that are permissive to VZV infection (monocytes, B cells and NKT cells); along with a novel mechanism for inhibiting CD8+ T cell effector function through induction of PD-L1 expression.
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http://dx.doi.org/10.1371/journal.ppat.1007650DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6435197PMC
March 2019

Varicella Zoster Virus Alters Expression of Cell Adhesion Proteins in Human Perineurial Cells via Interleukin 6.

J Infect Dis 2019 09;220(9):1453-1461

Department of Neurology, University of Colorado School of Medicine, Aurora.

Background: In temporal arteries (TAs) from patients with giant cell arteritis, varicella zoster virus (VZV) is seen in perineurial cells that surround adventitial nerve bundles and form the peripheral nerve-extrafascicular tissue barrier (perineurium). We hypothesized that during VZV reactivation from ganglia, virus travels transaxonally and disrupts the perineurium to infect surrounding cells.

Methods: Mock- and VZV-infected primary human perineurial cells (HPNCs) were examined for alterations in claudin-1, E-cadherin, and N-cadherin. Conditioned supernatant was analyzed for a soluble factor(s) mediating these alterations and for the ability to increase cell migration. To corroborate in vitro findings, a VZV-infected TA was examined.

Results: In VZV-infected HPNCs, claudin-1 redistributed to the nucleus; E-cadherin was lost and N-cadherin gained, with similar changes seen in VZV-infected perineurial cells in a TA. VZV-conditioned supernatant contained increased interleukin 6 (IL-6) that induced E-cadherin loss and N-cadherin gain and increased cell migration when added to uninfected HPNCs; anti-IL-6 receptor antibody prevented these changes.

Conclusions: IL-6 secreted from VZV-infected HPNCs facilitated changes in E- and N-cadherin expression and cell migration, reminiscent of an epithelial-to-mesenchymal cell transition, potentially contributing to loss of perineurial cell barrier integrity and viral spread. Importantly, an anti-IL-6 receptor antibody prevented virus-induced perineurial cell disruption.
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http://dx.doi.org/10.1093/infdis/jiz095DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6761973PMC
September 2019

Varicella Zoster Virus Induces Differential Cell-Type Specific Responses in Human Corneal Epithelial Cells and Keratocytes.

Invest Ophthalmol Vis Sci 2019 02;60(2):704-711

Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado, United States.

Purpose: While VZV DNA and antigen have been detected in acute and chronic VZV keratitis, it is unclear whether productive infection of corneal cells is ongoing or whether residual, noninfectious VZV antigens elicit inflammation. Herein, we examined VZV-infected primary human corneal epithelial cells (HCECs) and keratocytes (HKs) to elucidate the pathogenesis of VZV keratitis.

Methods: HCECs and HKs were mock- or VZV infected. Seven days later, cells were examined for morphology, proinflammatory cytokine and matrix metalloproteinase (MMP) release, ability to recruit peripheral blood mononuclear cells (PBMCs) and neutrophils, and MMP substrate cleavage.

Results: Both cell types synthesized infectious virus. VZV-infected HCECs proliferated, whereas VZV-infected HKs died. Compared to mock-infected cells, VZV-infected HCECs secreted significantly more IL-6, IL-8, IL-10, and IL-12p70 that were confirmed at the transcript level, and MMP-1 and MMP-9; conditioned supernatant attracted PBMCs and neutrophils and cleaved MMP substrates. In contrast, VZV-infected HKs suppressed cytokine secretion except for IL-8, which attracted neutrophils, and suppressed MMP release and substrate cleavage.

Conclusions: Overall, VZV-infected HCECs recapitulate findings of VZV keratitis with respect to epithelial cell proliferation, pseudodendrite formation and creation of a proinflammatory environment, providing an in vitro model for VZV infection of corneal epithelial cells. Furthermore, the proliferation and persistence of VZV-infected HCECs suggest that these cells may serve as viral reservoirs if immune clearance is incomplete. Finally, the finding that VZV-infected HKs die and suppress most proinflammatory cytokines and MMPs may explain the widespread death of these cells with unchecked viral spread due to ineffective recruitment of PBMCs.
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http://dx.doi.org/10.1167/iovs.18-25801DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6383726PMC
February 2019

Varicella zoster virus differentially alters morphology and suppresses proinflammatory cytokines in primary human spinal cord and hippocampal astrocytes.

J Neuroinflammation 2018 Nov 15;15(1):318. Epub 2018 Nov 15.

Department of Neurology, University of Colorado School of Medicine, 4200 E. 19th Avenue, Mail Stop B182, Aurora, CO, 80045, USA.

Background: Varicella zoster virus (VZV) is a ubiquitous alphaherpesvirus that produces varicella and zoster. VZV can infect multiple cell types in the spinal cord and brain, including astrocytes, producing myelopathy and encephalopathy. While studies of VZV-astrocyte interactions are sparse, a recent report showed that quiescent primary human spinal cord astrocytes (qHA-sps) did not appear activated morphologically during VZV infection. Since astrocytes play a critical role in host defenses during viral infections of the central nervous system, we examined the cytokine responses of qHA-sps and quiescent primary human hippocampal astrocytes (qHA-hps) to VZV infection in vitro, as well as the ability of conditioned supernatant to recruit immune cells.

Methods: At 3 days post-infection, mock- and VZV-infected qHA-sps and qHA-hps were examined for morphological changes by immunofluorescence antibody assay using antibodies directed against glial fibrillary acidic protein and VZV. Conditioned supernatants were analyzed for proinflammatory cytokines [interleukin (IL)-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, interferon-gamma, and tumor necrosis factor-α] using the Meso Scale Discovery multiplex ELISA platform. Finally, the ability of conditioned supernatants to attract peripheral blood mononuclear cells (PBMCs) was determined using a chemotaxis assay. Quiescent primary human perineurial cells (qHPNCs) served as a control for VZV-induced cytokine production and PBMC migration. To confirm that the astrocytes have the ability to increase cytokine secretion, qHA-sps and qHA-hps were treated with IL-1β and examined for morphological changes and IL-6 secretion.

Results: VZV-infected qHA-sps displayed extensive cellular processes, whereas VZV-infected qHA-hps became swollen and clustered together. Astrocytes had the capacity to secrete IL-6 in response to IL-1β. Compared to mock-infected cells, VZV-infected qHA-sps showed significantly reduced secretion of IL-2, IL-4, IL-6, IL-12p70, and IL-13, while VZV-infected qHA-hps showed significantly reduced IL-8 secretion. In contrast, levels of all 10 cytokines examined were significantly increased in VZV-infected qHPNCs. Consistent with these results, conditioned supernatant from VZV-infected qHPNCs, but not that from VZV-infected qHA-sps and qHA-hps, recruited PBMCs.

Conclusions: VZV-infected qHA-sps and qHA-hps have distinct morphological alterations and patterns of proinflammatory cytokine suppression that could contribute to ineffective viral clearance in VZV myelopathy and encephalopathy, respectively.
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http://dx.doi.org/10.1186/s12974-018-1360-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6236967PMC
November 2018

Nanoparticle uptake by circulating leukocytes: A major barrier to tumor delivery.

J Control Release 2018 09 17;286:85-93. Epub 2018 Jul 17.

Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Anschutz Medical Campus, Aurora, CO, United States. Electronic address:

Decades of research into improving drug delivery to tumors has documented uptake of particulate delivery systems by resident macrophages in the lung, liver, and spleen, and correlated short circulation times with reduced tumor accumulation. An implicit assumption in these studies is that nanoparticles present in the blood are available for distribution to the tumor. This study documents significant levels of lipoplex uptake by circulating leukocytes, and its effect on distribution to the tumor and other organs. In agreement with previous studies, PEGylation dramatically extends circulation times and enhances tumor delivery. However, our studies suggest that this relationship is not straightforward, and that particle sequestration by leukocytes can significantly alter biodistribution, especially with non-PEGylated nanoparticle formulations. We conclude that leukocyte uptake should be considered in biodistribution studies, and that delivery to these circulating cells may present opportunities for treating viral infections and leukemia.
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http://dx.doi.org/10.1016/j.jconrel.2018.07.031DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6936323PMC
September 2018

Varicella Zoster Virus Induces Nuclear Translocation of the Neurokinin-1 Receptor, Promoting Lamellipodia Formation and Viral Spread in Spinal Astrocytes.

J Infect Dis 2018 09;218(8):1324-1335

Department of Neurology, University of Colorado School of Medicine, Aurora.

Background: Varicella zoster virus (VZV) can present as a myelopathy with spinal astrocyte infection. Recent studies support a role for the neurokinin-1 receptor (NK-1R) in virus infections, as well as for cytoskeletal alterations that may promote viral spread. Thus, we examined the role of NK-1R in VZV-infected primary human spinal astrocytes (HA-sps) to shed light on the pathogenesis of VZV myelopathy.

Methods: Mock- and VZV-infected HA-sps were examined for substance P (subP) production, NK-1R localization, morphological changes, and viral spread in the presence or absence of the NK-1R antagonists aprepitant and rolapitant.

Results: VZV infection of HA-sps induced nuclear localization of full-length and truncated NK-1R in the absence of the endogenous ligand, subP, and was associated with extensive lamellipodia formation and viral spread that was inhibited by NK-1R antagonists.

Conclusions: We have identified a novel, subP-independent, proviral function of nuclear NK-1R associated with lamellipodia formation and viral spread that is distinct from subP-induced NK-1R cell membrane/cytoplasmic localization without lamellipodia formation. These results suggest that binding of a putative viral ligand to NK-1R produces a dramatically different NK-1R downstream effect than binding of subP. Finally, the NK-1R antagonists aprepitant and rolapitant provide promising alternatives to nucleoside analogs in treating VZV infections, including myelopathy.
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http://dx.doi.org/10.1093/infdis/jiy297DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6129113PMC
September 2018

Reply to 'Dissecting the role of miR-140 and its host gene'.

Nat Cell Biol 2018 05;20(5):519-520

Abide Therapeutics, San Diego, CA, USA.

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http://dx.doi.org/10.1038/s41556-018-0076-5DOI Listing
May 2018

Varicella zoster virus-infected cerebrovascular cells produce a proinflammatory environment.

Neurol Neuroimmunol Neuroinflamm 2017 Sep 13;4(5):e382. Epub 2017 Jul 13.

Department of Neurology (D.J., M.A.N.), Department of Medicine (C.P.N., B.E.P.), and Department of Pediatrics (K.S.), University of Colorado School of Medicine, Aurora.

Objective: To test whether varicella zoster virus (VZV) infection of human brain vascular cells and of lung fibroblasts directly increases proinflammatory cytokine levels, consistent with VZV as a causative agent in intracerebral VZV vasculopathy and giant-cell arteritis (GCA).

Methods: Conditioned supernatant from mock- and VZV-infected human brain vascular adventitial fibroblasts (HBVAFs), human perineurial cells (HPNCs), human brain vascular smooth muscle cells (HBVSMCs), and human fetal lung fibroblasts (HFLs) were collected at 72 hours postinfection and analyzed for levels of 30 proinflammatory cytokines using the Meso Scale Discovery Multiplex ELISA platform.

Results: Compared with mock infection, VZV infection led to significantly increased levels of the following: interleukin-8 (IL-8) in all cell lines examined; IL-6 in HBVAFs, HPNCs, and HFLs, with no change in HBVSMCs; and vascular endothelial growth factor A in HBVAFs, HBVSMCs, and HFLs, with a significant decrease in HPNCs. Other cytokines, including IL-2, IL-4, IL-15, IL-16, TGF-b, Eotaxin-1, Eotaxin-3, IP-10, MCP-1, and granulocyte macrophage colony-stimulating factor, were also significantly altered upon VZV infection in a cell type-specific manner.

Conclusions: VZV infection of vascular cells can directly produce a proinflammatory environment that may potentially lead to prolonged arterial wall inflammation and vasculitis. The VZV-mediated increase in IL-8 and IL-6 is consistent with that seen in the CSF of patients with intracerebral VZV vasculopathy, and the VZV-mediated increase in IL-6 is consistent with the cytokine's elevated levels in temporal arteries and plasma of patients with GCA.
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http://dx.doi.org/10.1212/NXI.0000000000000382DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5682918PMC
September 2017

Targeted Genome Sequencing Reveals Varicella-Zoster Virus Open Reading Frame 12 Deletion.

J Virol 2017 10 27;91(20). Epub 2017 Sep 27.

Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado, USA.

The neurotropic herpesvirus varicella-zoster virus (VZV) establishes a lifelong latent infection in humans following primary infection. The low abundance of VZV nucleic acids in human neurons has hindered an understanding of the mechanisms that regulate viral gene transcription during latency. To overcome this critical barrier, we optimized a targeted capture protocol to enrich VZV DNA and cDNA prior to whole-genome/transcriptome sequence analysis. Since the VZV genome is remarkably stable, it was surprising to detect that VZV32, a VZV laboratory strain with no discernible growth defect in tissue culture, contained a 2,158-bp deletion in open reading frame (ORF) 12. Consequently, ORF 12 and 13 protein expression was abolished and Akt phosphorylation was inhibited. The discovery of the ORF 12 deletion, revealed through targeted genome sequencing analysis, points to the need to authenticate the VZV genome when the virus is propagated in tissue culture. Viruses isolated from clinical samples often undergo genetic modifications when cultured in the laboratory. Historically, VZV is among the most genetically stable herpesviruses, a notion supported by more than 60 complete genome sequences from multiple isolates and following multiple passages. However, application of enrichment protocols to targeted genome sequencing revealed the unexpected deletion of a significant portion of VZV ORF 12 following propagation in cultured human fibroblast cells. While the enrichment protocol did not introduce bias in either the virus genome or transcriptome, the findings indicate the need for authentication of VZV by sequencing when the virus is propagated in tissue culture.
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http://dx.doi.org/10.1128/JVI.01141-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5625521PMC
October 2017

A human tissue-based functional assay platform to evaluate the immune function impact of small molecule inhibitors that target the immune system.

PLoS One 2017 18;12(7):e0180870. Epub 2017 Jul 18.

Oncology & Immunology Discovery, Merck Research Laboratories, Boston, MA, United States of America.

While the immune system is essential for the maintenance of the homeostasis, health and survival of humans, aberrant immune responses can lead to chronic inflammatory and autoimmune disorders. Pharmacological modulation of drug targets in the immune system to ameliorate disease also carry a risk of immunosuppression that could lead to adverse outcomes. Therefore, it is important to understand the 'immune fingerprint' of novel therapeutics as they relate to current and, clinically used immunological therapies to better understand their potential therapeutic benefit as well as immunosuppressive ability that might lead to adverse events such as infection risks and cancer. Since the mechanistic investigation of pharmacological modulators in a drug discovery setting is largely compound- and mechanism-centric but not comprehensive in terms of immune system impact, we developed a human tissue based functional assay platform to evaluate the impact of pharmacological modulators on a range of innate and adaptive immune functions. Here, we demonstrate that it is possible to generate a qualitative and quantitative immune system impact of pharmacological modulators, which might help better understand and predict the benefit-risk profiles of these compounds in the treatment of immune disorders.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0180870PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5515432PMC
September 2017

Varicella zoster virus vasculopathy: The expanding clinical spectrum and pathogenesis.

J Neuroimmunol 2017 07 18;308:112-117. Epub 2017 Mar 18.

Department of Neurology, University of Colorado School of Medicine, Aurora, CO 80045, USA. Electronic address:

Varicella zoster virus (VZV) is a ubiquitous, human alphaherpesvirus that produces varicella on primary infection then becomes latent in ganglionic neurons along the entire neuraxis. In elderly and immunocompromised individuals, VZV reactivates and travels along nerve fibers peripherally resulting in zoster. However, VZV can also spread centrally and infect cerebral and extracranial arteries (VZV vasculopathy) to produce transient ischemic attacks, stroke, aneurysm, sinus thrombosis and giant cell arteritis, as well as granulomatous aortitis. The mechanisms of virus-induced pathological vascular remodeling are not fully elucidated; however, recent studies suggest that inflammation and dysregulation of programmed death ligand-1 play a significant role.
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http://dx.doi.org/10.1016/j.jneuroim.2017.03.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5489071PMC
July 2017

Inhibition of RORγT Skews TCRα Gene Rearrangement and Limits T Cell Repertoire Diversity.

Cell Rep 2016 12;17(12):3206-3218

Merck Research Laboratories, 901 California Avenue, Palo Alto, CA 94304, USA. Electronic address:

Recent studies have elucidated the molecular mechanism of RORγT transcriptional regulation of Th17 differentiation and function. RORγT was initially identified as a transcription factor required for thymopoiesis by maintaining survival of CD4CD8 (DP) thymocytes. While RORγ antagonists are currently being developed to treat autoimmunity, it remains unclear how RORγT inhibition may impact thymocyte development. In this study, we show that in addition to regulating DP thymocytes survival, RORγT also controls genes that regulate thymocyte migration, proliferation, and T cell receptor (TCR)α selection. Strikingly, pharmacological inhibition of RORγ skews TCRα gene rearrangement, limits T cell repertoire diversity, and inhibits development of autoimmune encephalomyelitis. Thus, targeting RORγT not only inhibits Th17 cell development and function but also fundamentally alters thymic-emigrant recognition of self and foreign antigens. The analysis of RORγ inhibitors has allowed us to gain a broader perspective of the diverse function of RORγT and its impact on T cell biology.
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http://dx.doi.org/10.1016/j.celrep.2016.11.073DOI Listing
December 2016

Varicella-Zoster Virus Downregulates Programmed Death Ligand 1 and Major Histocompatibility Complex Class I in Human Brain Vascular Adventitial Fibroblasts, Perineurial Cells, and Lung Fibroblasts.

J Virol 2016 Dec 14;90(23):10527-10534. Epub 2016 Nov 14.

Department of Neurology, University of Colorado School of Medicine, Aurora, Colorado, USA

Varicella-zoster virus (VZV) vasculopathy produces stroke, giant cell arteritis, and granulomatous aortitis, and it develops after virus reactivates from ganglia and spreads transaxonally to arterial adventitia, resulting in persistent inflammation and pathological vascular remodeling. The mechanism(s) by which inflammatory cells persist in VZV-infected arteries is unknown; however, virus-induced dysregulation of programmed death ligand 1 (PD-L1) may play a role. Specifically, PD-L1 can be expressed on virtually all nucleated cells and suppresses the immune system by interacting with the programmed cell death protein receptor 1, found exclusively on immune cells; thus, downregulation of PD-L1 may promote inflammation, as seen in some autoimmune diseases. Both flow cytometry and immunofluorescence analyses to test whether VZV infection of adventitial cells downregulates PD-L1 showed decreased PD-L1 expression in VZV-infected compared to mock-infected human brain vascular adventitial fibroblasts (HBVAFs), perineural cells (HPNCs), and fetal lung fibroblasts (HFLs) at 72 h postinfection. Quantitative RT-PCR analyses showed no change in PD-L1 transcript levels between mock- and VZV-infected cells, indicating a posttranscriptional mechanism for VZV-mediated downregulation of PD-L1. Flow cytometry analyses showed decreased major histocompatibility complex class I (MHC-I) expression in VZV-infected cells and adjacent uninfected cells compared to mock-infected cells. These data suggest that reduced PD-L1 expression in VZV-infected adventitial cells contribute to persistent vascular inflammation observed in virus-infected arteries from patients with VZV vasculopathy, while downregulation of MHC-I prevents viral clearance.

Importance: Here, we provide the first demonstration that VZV downregulates PD-L1 expression in infected HBVAFs, HPNCs, and HFLs, which, together with the noted VZV-mediated downregulation of MHC-I, might foster persistent inflammation in vessels, leading to pathological vascular remodeling during VZV vasculopathy and persistent inflammation in infected lungs to promote subsequent infection of T cells and hematogenous virus spread. Identification of a potential mechanism by which persistent inflammation in the absence of effective viral clearance occurs in VZV vasculopathy and VZV infection of the lung is a step toward targeted therapy of VZV-induced disease.
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http://dx.doi.org/10.1128/JVI.01546-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5110195PMC
December 2016

Integrating metabolomics and transcriptomics data to discover a biocatalyst that can generate the amine precursors for alkamide biosynthesis.

Plant J 2016 12 27;88(5):775-793. Epub 2016 Sep 27.

The Roy J. Carver Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, IA, USA.

The Echinacea genus is exemplary of over 30 plant families that produce a set of bioactive amides, called alkamides. The Echinacea alkamides may be assembled from two distinct moieties, a branched-chain amine that is acylated with a novel polyunsaturated fatty acid. In this study we identified the potential enzymological source of the amine moiety as a pyridoxal phosphate-dependent decarboxylating enzyme that uses branched-chain amino acids as substrate. This identification was based on a correlative analysis of the transcriptomes and metabolomes of 36 different E. purpurea tissues and organs, which expressed distinct alkamide profiles. Although no correlation was found between the accumulation patterns of the alkamides and their putative metabolic precursors (i.e., fatty acids and branched-chain amino acids), isotope labeling analyses supported the transformation of valine and isoleucine to isobutylamine and 2-methylbutylamine as reactions of alkamide biosynthesis. Sequence homology identified the pyridoxal phosphate-dependent decarboxylase-like proteins in the translated proteome of E. purpurea. These sequences were prioritized for direct characterization by correlating their transcript levels with alkamide accumulation patterns in different organs and tissues, and this multi-pronged approach led to the identification and characterization of a branched-chain amino acid decarboxylase, which would appear to be responsible for generating the amine moieties of naturally occurring alkamides.
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http://dx.doi.org/10.1111/tpj.13295DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5195896PMC
December 2016

A Clade-Specific Arabidopsis Gene Connects Primary Metabolism and Senescence.

Front Plant Sci 2016 12;7:983. Epub 2016 Jul 12.

Department of Genetics, Development and Cell Biology, Iowa State University, AmesIA, USA; Center for Metabolic Biology, Iowa State University, AmesIA, USA.

Nearly immobile, plants have evolved new components to be able to respond to changing environments. One example is Qua Quine Starch (QQS, AT3G30720), an Arabidopsis thaliana-specific orphan gene that integrates primary metabolism with adaptation to environment changes. SAQR (Senescence-Associated and QQS-Related, AT1G64360), is unique to a clade within the family Brassicaceae; as such, the gene may have arisen about 20 million years ago. SAQR is up-regulated in QQS RNAi mutant and in the apx1 mutant under light-induced oxidative stress. SAQR plays a role in carbon allocation: overexpression lines of SAQR have significantly decreased starch content; conversely, in a saqr T-DNA knockout (KO) line, starch accumulation is increased. Meta-analysis of public microarray data indicates that SAQR expression is correlated with expression of a subset of genes involved in senescence, defense, and stress responses. SAQR promoter::GUS expression analysis reveals that SAQR expression increases after leaf expansion and photosynthetic capacity have peaked, just prior to visible natural senescence. SAQR is expressed predominantly within leaf and cotyledon vasculature, increasing in intensity as natural senescence continues, and then decreasing prior to death. In contrast, under experimentally induced senescence, SAQR expression increases in vasculature of cotyledons but not in true leaves. In SAQR KO line, the transcript level of the dirigent-like disease resistance gene (AT1G22900) is increased, while that of the Early Light Induced Protein 1 gene (ELIP1, AT3G22840) is decreased. Taken together, these data indicate that SAQR may function in the QQS network, playing a role in integration of primary metabolism with adaptation to internal and environmental changes, specifically those that affect the process of senescence.
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http://dx.doi.org/10.3389/fpls.2016.00983DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4940393PMC
July 2016

Proinflammatory cytokines and matrix metalloproteinases in CSF of patients with VZV vasculopathy.

Neurol Neuroimmunol Neuroinflamm 2016 Aug 13;3(4):e246. Epub 2016 Jun 13.

Departments of Neurology (D.J., E.A., S.S., D.G., M.A.N.) and Immunology and Microbiology (D.G.), University of Colorado School of Medicine, Aurora.

Objective: To determine the levels of proinflammatory cytokines and matrix metalloproteinases (MMPs) in the CSF of patients with virologically verified varicella zoster virus (VZV) vasculopathy.

Methods: CSF from 30 patients with virologically verified VZV vasculopathy was analyzed for levels of proinflammatory cytokines and MMPs using the Meso Scale Discovery multiplex ELISA platform. Positive CNS inflammatory disease controls were provided by CSF from 30 patients with multiple sclerosis. Negative controls were provided by CSF from 20 healthy controls.

Results: Compared to multiple sclerosis CSF and CSF from healthy controls, levels of interleukin (IL)-8, IL-6, and MMP-2 were significantly elevated in VZV vasculopathy CSF.

Conclusions: CSF of patients with VZV vasculopathy revealed a unique profile of elevated proinflammatory cytokines, IL-8 and IL-6, along with elevated MMP-2. The relevance of these cytokines to the pathogenesis of VZV vasculopathy requires further study.
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http://dx.doi.org/10.1212/NXI.0000000000000246DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4907802PMC
August 2016

Measurement uncertainty for clinical laboratories - a revision of the concept.

Clin Chem Lab Med 2016 Aug;54(8):1303-7

The uncertainty of a measurement result is a fundamental concept in metrology indicating the range within the "true" value of a measurement should lie. Although not commonly reported with results, the calculation of measurement uncertainty (MU) has become common in routine clinical laboratories. Interpretation of numerical pathology results is made by comparison with data from other measurements. As MU is aimed at assisting with result interpretation, it should be related to the specific comparison being made. There are three basic type of comparators: a previous result from the same patient, a population reference interval, or a clinical decision point. For each comparison, the "true" value is that which would have been obtained from the instrument used to make the comparator measurements if it was measured without uncertainty. The MU is the range of likely deviations from this true value due to the method used to produce the result under interpretation. For patient monitoring, if the two measurements were made on the same analyzer, the uncertainty is the imprecision of the assay over the relevant time frame. In comparing with a manufacturer-specific reference interval, the MU is deviation from the manufacturer's master calibrator. For clinical decision points produced with the assays traceable to international references, the MU is related to deviation from that reference standard. For optimal use of MU in the clinical laboratory, it may be necessary to consider the use of the test result and the concept of a single MU for each result may need to be revised.
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http://dx.doi.org/10.1515/cclm-2016-0311DOI Listing
August 2016

QQS orphan gene regulates carbon and nitrogen partitioning across species via NF-YC interactions.

Proc Natl Acad Sci U S A 2015 Nov 9;112(47):14734-9. Epub 2015 Nov 9.

Department of Genetics, Development and Cell Biology, Iowa State University, Ames, IA 50011; Center for Metabolic Biology, Iowa State University, Ames, IA 50011;

The allocation of carbon and nitrogen resources to the synthesis of plant proteins, carbohydrates, and lipids is complex and under the control of many genes; much remains to be understood about this process. QQS (Qua-Quine Starch; At3g30720), an orphan gene unique to Arabidopsis thaliana, regulates metabolic processes affecting carbon and nitrogen partitioning among proteins and carbohydrates, modulating leaf and seed composition in Arabidopsis and soybean. Here the universality of QQS function in modulating carbon and nitrogen allocation is exemplified by a series of transgenic experiments. We show that ectopic expression of QQS increases soybean protein independent of the genetic background and original protein content of the cultivar. Furthermore, transgenic QQS expression increases the protein content of maize, a C4 species (a species that uses 4-carbon photosynthesis), and rice, a protein-poor agronomic crop, both highly divergent from Arabidopsis. We determine that QQS protein binds to the transcriptional regulator AtNF-YC4 (Arabidopsis nuclear factor Y, subunit C4). Overexpression of AtNF-YC4 in Arabidopsis mimics the QQS-overexpression phenotype, increasing protein and decreasing starch levels. NF-YC, a component of the NF-Y complex, is conserved across eukaryotes. The NF-YC4 homologs of soybean, rice, and maize also bind to QQS, which provides an explanation of how QQS can act in species where it does not occur endogenously. These findings are, to our knowledge, the first insight into the mechanism of action of QQS in modulating carbon and nitrogen allocation across species. They have major implications for the emergence and function of orphan genes, and identify a nontransgenic strategy for modulating protein levels in crop species, a trait of great agronomic significance.
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http://dx.doi.org/10.1073/pnas.1514670112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4664325PMC
November 2015

Effect of renal function on serum cardiac troponin T--Population and individual effects.

Clin Biochem 2015 Aug 11;48(12):807-10. Epub 2015 May 11.

Department of Chemical Pathology, SydPath, St Vincent's Hospital, Sydney, Victoria St, Darlinghurst, NSW 2010, Australia; Faculty of Medicine, University of New South Wales, Randwick, NSW 2031, Australia.

Background: Elevations of serum cardiac troponin T (cTnT) have been described in patients with end stage chronic kidney disease (CKD) although the mechanism is unknown, whether from increased production or decreased clearance. Less is known about cTnT in short term changes in renal function and in lesser degrees of renal impairment.

Objectives: This study aimed to investigate the effect of renal function changes on cTnT within individuals and characterise the distribution of cTnT according to renal function in the population.

Design And Methods: A hospital laboratory database extract was performed for paired creatinine and cTnT results. cTnT was compared with estimated glomerular filtration rate (eGFR) at the population level. In individuals who had undergone repeat testing, changes in cTnT were compared with corresponding changes in creatinine.

Results: At the population level, 17,113 cTnT and creatinine measurements from 10,418 patients demonstrated rising cTnT with falling eGFR, with no eGFR threshold for this effect. Of these, 3108 pairs of results were obtained from patients who had undergone repeat testing. The median retesting interval was 15 h (interquartile range: 7-25 h). Within individuals, the magnitude of changes in cTnT approximated 33% of changes in creatinine.

Conclusions: At the population level, moderate reductions in GFR (30-59 mL/min/1.73 m(2)) corresponded to a median cTnT above the 14 ng/L upper reference limit. The modest association between changes in cTnT and creatinine within individuals in the short term further highlights the need for caution when interpreting troponin elevations in this setting.
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http://dx.doi.org/10.1016/j.clinbiochem.2015.05.004DOI Listing
August 2015

Analytical performance specifications for EQA schemes - need for harmonisation.

Clin Chem Lab Med 2015 May;53(6):919-24

External Quality Assurance (EQA) is a vital tool in laboratory medicine to assess individual laboratory analytical performance and also the differences between the results from different laboratories. This information is also useful for professional bodies and manufacturers as part of post-market surveillance. The process involves the measurement of one or more samples by many laboratories and then assessment of the results. Individual results are generally assessed by how far they lie from a target, which may be established using reference methods or a median of some or all of the submitted results. The distance of a result from the target is compared with analytical performance specifications in order to assess the analytical quality. One of the uses of the Stockholm hierarchy of performance goals is to set the performance specifications for analysis of EQA results. Fifteen years after the Stockholm consensus meeting, EQA analytical performance specifications appear to still vary widely between EQA providers. This can be due to a range of factors, including the rationale for setting the criteria, the expected response to a failure to meet the specified performance, the clinical meaning behind meeting the specifications, and the possible need for further analytical improvements. There are also differences in the models chosen to set the criteria, usually either state of the art or biological variation, and then differences in how these are applied. While harmonisation of EQA performance specifications may be some time off, all EQA providers should define the nature of their specifications and the basis for their selection and make this information available to customers.
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http://dx.doi.org/10.1515/cclm-2014-1268DOI Listing
May 2015

Tetraspanin CD9 and ectonucleotidase CD73 identify an osteochondroprogenitor population with elevated osteogenic properties.

Development 2015 Feb 6;142(3):438-43. Epub 2015 Jan 6.

Abide Therapeutics, San Diego, CA 92121, USA.

Cell-based bone regeneration strategies offer promise for traumatic bone injuries, congenital defects, non-union fractures and other skeletal pathologies. Postnatal bone remodeling and fracture healing provide evidence that an osteochondroprogenitor cell is present in adult life that can differentiate to remodel or repair the fractured bone. However, cell-based skeletal repair in the clinic is still in its infancy, mostly due to poor characterization of progenitor cells and lack of knowledge about their in vivo behavior. Here, we took a combined approach of high-throughput screening, flow-based cell sorting and in vivo transplantation to isolate markers that identify osteochondroprogenitor cells. We show that the presence of tetraspanin CD9 enriches for osteochondroprogenitors within CD105(+) mesenchymal cells and that these cells readily form bone upon transplantation. In addition, we have used Thy1.2 and the ectonucleotidase CD73 to identify subsets within the CD9(+) population that lead to endochondral or intramembranous-like bone formation. Utilization of this unique cell surface phenotype to enrich for osteochondroprogenitor cells will allow for further characterization of the molecular mechanisms that regulate their osteogenic properties.
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http://dx.doi.org/10.1242/dev.113571DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4302994PMC
February 2015

Schnurri-3 regulates ERK downstream of WNT signaling in osteoblasts.

J Clin Invest 2013 Sep 15;123(9):4010-22. Epub 2013 Aug 15.

Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, New York 10065, USA.

Mice deficient in Schnurri-3 (SHN3; also known as HIVEP3) display increased bone formation, but harnessing this observation for therapeutic benefit requires an improved understanding of how SHN3 functions in osteoblasts. Here we identified SHN3 as a dampener of ERK activity that functions in part downstream of WNT signaling in osteoblasts. A D-domain motif within SHN3 mediated the interaction with and inhibition of ERK activity and osteoblast differentiation, and knockin of a mutation in Shn3 that abolishes this interaction resulted in aberrant ERK activation and consequent osteoblast hyperactivity in vivo. Additionally, in vivo genetic interaction studies demonstrated that crossing to Lrp5(-/-) mice partially rescued the osteosclerotic phenotype of Shn3(-/-) mice; mechanistically, this corresponded to the ability of SHN3 to inhibit ERK-mediated suppression of GSK3β. Inducible knockdown of Shn3 in adult mice resulted in a high-bone mass phenotype, providing evidence that transient blockade of these pathways in adults holds promise as a therapy for osteoporosis.
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http://dx.doi.org/10.1172/JCI69443DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3754267PMC
September 2013

The microtubule-associated protein DCAMKL1 regulates osteoblast function via repression of Runx2.

J Exp Med 2013 Aug 5;210(9):1793-806. Epub 2013 Aug 5.

State Key Laboratory of Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.

Osteoblasts are responsible for the formation and mineralization of the skeleton. To identify novel regulators of osteoblast differentiation, we conducted an unbiased forward genetic screen using a lentiviral-based shRNA library. This functional genomics analysis led to the identification of the microtubule-associated protein DCAMKL1 (Doublecortin-like and CAM kinase-like 1) as a novel regulator of osteogenesis. Mice with a targeted disruption of Dcamkl1 displayed elevated bone mass secondary to increased bone formation by osteoblasts. Molecular experiments demonstrated that DCAMKL1 represses osteoblast activation by antagonizing Runx2, the master transcription factor in osteoblasts. Key elements of the cleidocranial dysplasia phenotype observed in Runx2(+/-) mice are reversed by the introduction of a Dcamkl1-null allele. Our results establish a genetic linkage between these two proteins in vivo and demonstrate that DCAMKL1 is a physiologically relevant regulator of anabolic bone formation.
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http://dx.doi.org/10.1084/jem.20111790DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3754873PMC
August 2013

Control of bone resorption in mice by Schnurri-3.

Proc Natl Acad Sci U S A 2012 May 9;109(21):8173-8. Epub 2012 May 9.

Department of Immunology and Infectious Disease, Harvard School of Public Health, Boston, MA 02115, USA.

Mice lacking the large zinc finger protein Schnurri-3 (Shn3) display increased bone mass, in part, attributable to augmented osteoblastic bone formation. Here, we show that in addition to regulating bone formation, Shn3 indirectly controls bone resorption by osteoclasts in vivo. Although Shn3 plays no cell-intrinsic role in osteoclasts, Shn3-deficient animals show decreased serum markers of bone turnover. Mesenchymal cells lacking Shn3 are defective in promoting osteoclastogenesis in response to selective stimuli, likely attributable to reduced expression of the key osteoclastogenic factor receptor activator of nuclear factor-κB ligand. The bone phenotype of Shn3-deficient mice becomes more pronounced with age, and mice lacking Shn3 are completely resistant to disuse osteopenia, a process that requires functional osteoclasts. Finally, selective deletion of Shn3 in the mesenchymal lineage recapitulates the high bone mass phenotype of global Shn3 KO mice, including reduced osteoclastic bone catabolism in vivo, indicating that Shn3 expression in mesenchymal cells directly controls osteoblastic bone formation and indirectly regulates osteoclastic bone resorption.
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http://dx.doi.org/10.1073/pnas.1205848109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3361406PMC
May 2012

MLK3 regulates bone development downstream of the faciogenital dysplasia protein FGD1 in mice.

J Clin Invest 2011 Nov 3;121(11):4383-92. Epub 2011 Oct 3.

Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, Massachusetts, USA.

Mutations in human FYVE, RhoGEF, and PH domain-containing 1 (FGD1) cause faciogenital dysplasia (FGDY; also known as Aarskog syndrome), an X-linked disorder that affects multiple skeletal structures. FGD1 encodes a guanine nucleotide exchange factor (GEF) that specifically activates the Rho GTPase CDC42. However, the mechanisms by which mutations in FGD1 affect skeletal development are unknown. Here, we describe what we believe to be a novel signaling pathway in osteoblasts initiated by FGD1 that involves the MAP3K mixed-lineage kinase 3 (MLK3). We observed that MLK3 functions downstream of FGD1 to regulate ERK and p38 MAPK, which in turn phosphorylate and activate the master regulator of osteoblast differentiation, Runx2. Mutations in FGD1 found in individuals with FGDY ablated its ability to activate MLK3. Consistent with our description of this pathway and the phenotype of patients with FGD1 mutations, mice with a targeted deletion of Mlk3 displayed multiple skeletal defects, including dental abnormalities, deficient calvarial mineralization, and reduced bone mass. Furthermore, mice with knockin of a mutant Mlk3 allele that is resistant to activation by FGD1/CDC42 displayed similar skeletal defects, demonstrating that activation of MLK3 specifically by FGD1/CDC42 is important for skeletal mineralization. Thus, our results provide a putative biochemical mechanism for the skeletal defects in human FGDY and suggest that modulating MAPK signaling may benefit these patients.
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http://dx.doi.org/10.1172/JCI59041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3204846PMC
November 2011

Osteoimmunology at the nexus of arthritis, osteoporosis, cancer, and infection.

J Clin Invest 2011 Jul 1;121(7):2534-42. Epub 2011 Jul 1.

Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, Massachusetts 02115, USA.

Over the past decade and a half, the biomedical community has uncovered a previously unappreciated reciprocal relationship between cells of the immune and skeletal systems. Work in this field, which has been termed "osteoimmunology," has resulted in the development of clinical therapeutics for seemingly disparate diseases linked by the common themes of inflammation and bone remodeling. Here, the important concepts and discoveries in osteoimmunology are discussed in the context of the diseases bridging these two organ systems, including arthritis, osteoporosis, cancer, and infection, and the targeted treatments used by clinicians to combat them.
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http://dx.doi.org/10.1172/JCI46262DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3223839PMC
July 2011

Dampening of death pathways by schnurri-2 is essential for T-cell development.

Nature 2011 Apr;472(7341):105-9

Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, Massachusetts 02115, USA.

Generation of a diverse and self-tolerant T-cell repertoire requires appropriate interpretation of T-cell antigen receptor (TCR) signals by CD4(+ ) CD8(+) double-positive thymocytes. Thymocyte cell fate is dictated by the nature of TCR-major-histocompatibility-complex (MHC)-peptide interactions, with signals of higher strength leading to death (negative selection) and signals of intermediate strength leading to differentiation (positive selection). Molecules that regulate T-cell development by modulating TCR signal strength have been described but components that specifically define the boundaries between positive and negative selection remain unknown. Here we show in mice that repression of TCR-induced death pathways is critical for proper interpretation of positive selecting signals in vivo, and identify schnurri-2 (Shn2; also known as Hivep2) as a crucial death dampener. Our results indicate that Shn2(-/-) double-positive thymocytes inappropriately undergo negative selection in response to positive selecting signals, thus leading to disrupted T-cell development. Shn2(-/-) double-positive thymocytes are more sensitive to TCR-induced death in vitro and die in response to positive selection interactions in vivo. However, Shn2-deficient thymocytes can be positively selected when TCR-induced death is genetically ablated. Shn2 levels increase after TCR stimulation, indicating that integration of multiple TCR-MHC-peptide interactions may fine-tune the death threshold. Mechanistically, Shn2 functions downstream of TCR proximal signalling compenents to dampen Bax activation and the mitochondrial death pathway. Our findings uncover a critical regulator of T-cell development that controls the balance between death and differentiation.
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http://dx.doi.org/10.1038/nature09848DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3077958PMC
April 2011

The E3 ubiquitin ligase Wwp2 regulates craniofacial development through mono-ubiquitylation of Goosecoid.

Nat Cell Biol 2011 Jan 19;13(1):59-65. Epub 2010 Dec 19.

Department of Immunology and Infectious Disease, Harvard School of Public Health, Boston, Massachusetts 0211, USA.

Craniofacial anomalies (CFAs) are the most frequently occurring human congenital disease, and a major cause of infant mortality and childhood morbidity. Although CFAs seems to arise from a combination of genetic factors and environmental influences, the underlying gene defects and pathophysiological mechanisms for most CFAs are currently unknown. Here we reveal a role for the E3 ubiquitin ligase Wwp2 in regulating craniofacial patterning. Mice deficient in Wwp2 develop malformations of the craniofacial region. Wwp2 is present in cartilage where its expression is controlled by Sox9. Our studies demonstrate that Wwp2 influences craniofacial patterning through its interactions with Goosecoid (Gsc), a paired-like homeobox transcription factor that has an important role in craniofacial development. We show that Wwp2-associated Gsc is a transcriptional activator of the key cartilage regulatory protein Sox6. Wwp2 interacts with Gsc to facilitate its mono-ubiquitylation, a post-translational modification required for optimal transcriptional activation of Gsc. Our results identify for the first time a physiological pathway regulated by Wwp2 in vivo, and also a unique non-proteolytic mechanism through which Wwp2 controls craniofacial development.
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http://dx.doi.org/10.1038/ncb2134DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3059716PMC
January 2011