Abstract
The Wnt/β-catenin signaling pathway plays an important role in renal development and is reexpressed in the injured kidney and other organs. β-Catenin signaling is protective in acute kidney injury (AKI) through actions on the proximal tubule, but the current dogma is that Wnt/β-catenin signaling promotes fibrosis and development of chronic kidney disease (CKD). As the role of proximal tubular β-catenin signaling in CKD remains unclear, we genetically stabilized (i.e., activated) β-catenin specifically in murine proximal tubules. Mice with increased tubular β-catenin signaling were protected in 2 murine models of AKI to CKD progression. Oxidative stress, a common feature of CKD, reduced the conventional T cell factor/lymphoid enhancer factor–dependent β-catenin signaling and augmented FoxO3-dependent activity in proximal tubule cells in vitro and in vivo. The protective effect of proximal tubular β-catenin in renal injury required the presence of FoxO3 in vivo. Furthermore, we identified cystathionine γ-lyase as a potentially novel transcriptional target of β-catenin/FoxO3 interactions in the proximal tubule. Thus, our studies overturned the conventional dogma about β-catenin signaling and CKD by showing a protective effect of proximal tubule β-catenin in CKD and identified a potentially new transcriptional target of β-catenin/FoxO3 signaling that has therapeutic potential for CKD.
Authors
Stellor Nlandu-Khodo, Yosuke Osaki, Lauren Scarfe, Haichun Yang, Melanie Phillips-Mignemi, Jane Tonello, Kenyi Saito-Diaz, Surekha Neelisetty, Alla Ivanova, Tessa Huffstater, Robert McMahon, M. Mark Taketo, Mark deCaestecker, Balakuntalam Kasinath, Raymond C. Harris, Ethan Lee, Leslie S. Gewin
Abstract
Although aging represents the most important epidemiologic risk factor for fibrotic disease, the reasons for this are incompletely understood. Excess collagen deposition in tissues is the sine qua non of tissue fibrosis and can be viewed as an imbalance between collagen production and collagen degradation. Yet we still lack a detailed understanding of the changes that take place during development, maturation, and aging in extracellular matrix (ECM) dynamics. Resolution of fibrosis is impaired in aging, and this impairment may explain why age is the most important risk factor for fibrotic diseases, such as idiopathic pulmonary fibrosis. However, ECM dynamics and impaired resolution of fibrosis in aging remain understudied. Here we show that cell-mediated collagen uptake and degradation are diminished in aged animals and this finding correlates with downregulation of the collagen endocytic receptor mannose receptor, C-type 2 (Mrc2). We identify myeloid zinc finger-1 as a potentially novel transcriptional regulator of Mrc2, and both this transcription factor and Mrc2 are downregulated in multiple tissues and organisms in an age-dependent manner. Thus, cell-mediated degradation of collagen is an essential process that promotes resolution of fibrosis, and impairment in this process contributes to age-related fibrosis.
Authors
Michael J. Podolsky, Christopher D. Yang, Carlos Lizama Valenzuela, Ritwik Datta, Steven K. Huang, Stephen L. Nishimura, Sarah L. Dallas, Paul J. Wolters, Claude Jourdan Le Saux, Kamran Atabai
Abstract
Efficient adeno-associated virus–mediated (AAV-mediated) gene delivery remains a significant obstacle to effective retinal gene therapies. Here, we apply directed evolution — guided by deep sequencing and followed by direct in vivo secondary selection of high-performing vectors with a GFP-barcoded library — to create AAV viral capsids with the capability to deliver genes to the outer retina in primates. A replication-incompetent library, produced via providing rep in trans, was created to mitigate risk of AAV propagation. Six rounds of in vivo selection with this library in primates — involving intravitreal library administration, recovery of genomes from outer retina, and extensive next-generation sequencing of each round — resulted in vectors with redirected tropism to the outer retina and increased gene delivery efficiency to retinal cells. These viral vectors expand the toolbox of vectors available for primate retina, and they may enable less invasive delivery of therapeutic genes to patients, potentially offering retina-wide infection at a similar dosage to vectors currently in clinical use.
Authors
Leah C. Byrne, Timothy P. Day, Meike Visel, Jennifer A. Strazzeri, Cécile Fortuny, Deniz Dalkara, William H. Merigan, David V. Schaffer, John G. Flannery
Abstract
The role CD4+ T cells play in tumor immunity is less well appreciated than the cytotoxic role of CD8+ T cells. Despite clear evidence for CD4+ T cell dependency across multiple immunotherapies, the mechanisms by which CD4+ T cells infiltrate tumors remain poorly understood. Prior studies by our group have shown in a mouse model of pancreatic cancer that systemic activation of the cell surface TNF superfamily member CD40 drives T cell infiltration into tumors and, in combination with immune checkpoint blockade, leads to durable tumor regressions and cures that depend on both CD8+ and CD4+ T cells. Here, we used single-cell transcriptomics to examine the tumor microenvironment following treatment with agonist CD40 antibody with or without immune checkpoint blockade. We show that intratumor myeloid cells produce the chemokine CCL5 in response to CD40 agonist and that CCL5 mediates an influx of CD4+ T cells into the tumor microenvironment. Disruption of CCL5 genetically or pharmacologically mitigates the influx of CD4+ but not CD8+ T cells into tumors and blunts the therapeutic efficacy of immunotherapy. These findings highlight a previously unappreciated role for CCL5 in selectively mediating CD4+ T cell tumor infiltration in response to effective immunotherapy.
Authors
Austin P. Huffman, Jeffrey H. Lin, Samuel I. Kim, Katelyn T. Byrne, Robert H. Vonderheide
Abstract
Discovery strategies commonly focus on the identification of chemical libraries or natural products, but the modulation of endogenous ligands offers a much better therapeutic strategy due to their low adverse potential. Recently, we found that hexadecanamide (Hex) is present in hippocampal nuclei of normal mice as an endogenous ligand of PPARα. This study underlines the importance of Hex in inducing the expression of brain-derived neurotrophic factor (BDNF) from hippocampal neurons via PPARα. The level of Hex was lower in the hippocampi of 5XFAD mice as compared with that in non-Tg mice. Oral administration of Hex increased the level of this molecule in the hippocampus to stimulate BDNF and its downstream plasticity-associated molecules, promote synaptic functions in the hippocampus, and improve memory and learning in 5XFAD mice. However, oral Hex remained unable to stimulate hippocampal plasticity and improve cognitive behaviors in 5XFADPparα-null and 5XFADPparα-ΔHippo mice, indicating an essential role of hippocampal PPARα in Hex-mediated improvement in hippocampal functions. This is the first demonstration to our knowledge of protection of hippocampal functions by oral administration of a hippocampus-based drug, suggesting that Hex may be explored for therapeutic intervention in AD.
Authors
Dhruv Patel, Avik Roy, Sumita Raha, Madhuchhanda Kundu, Frank J. Gonzalez, Kalipada Pahan
Abstract
The ability of Mycobacterium tuberculosis to form serpentine cords is intrinsically related to its virulence, but specifically how M. tuberculosis cording contributes to pathogenesis remains obscure. Here, we show that several M. tuberculosis clinical isolates form intracellular cords in primary human lymphatic endothelial cells (hLECs) in vitro and in the lymph nodes of patients with tuberculosis. We identified via RNA-Seq a transcriptional program that activated, in infected-hLECs, cell survival and cytosolic surveillance of pathogens pathways. Consistent with this, cytosolic access was required for intracellular M. tuberculosis cording. Mycobacteria lacking ESX-1 type VII secretion system or phthiocerol dimycocerosates expression, which failed to access the cytosol, were indeed unable to form cords within hLECs. Finally, we show that M. tuberculosis cording is a size-dependent mechanism used by the pathogen to avoid its recognition by cytosolic sensors and evade either resting or IFN-γ–induced hLEC immunity. These results explain the long-standing association between M. tuberculosis cording and virulence and how virulent mycobacteria use intracellular cording as strategy to successfully adapt and persist in the lymphatic tracts.
Authors
Thomas R. Lerner, Christophe J. Queval, Rachel P. Lai, Matthew R.G. Russell, Antony Fearns, Daniel J. Greenwood, Lucy Collinson, Robert J. Wilkinson, Maximiliano G. Gutierrez
Abstract
The incidence of type 1 diabetes (T1D) has been increasing among children and adolescents, in which environmental factors, including gut microbiota, play an important role. However, the underlying mechanisms are yet to be determined. Here, we show that patients with newly diagnosed T1D displayed not only a distinct profile of gut microbiota associated with decreased short-chain fatty acids (SCFAs) production, but also an altered IgA-mediated immunity compared with healthy control subjects. Using germ-free NOD mice, we demonstrate that gut microbiota from patients with T1D promoted different IgA-mediated immune responses compared with healthy control gut microbiota. Treatment with the SCFA, acetate, reduced gut bacteria–induced IgA response accompanied by decreased severity of insulitis in NOD mice. We believe our study provides new insights into the functional effects of gut microbiota on inducing IgA immune response in T1D, suggesting that SCFAs might be potential therapeutic agents in T1D prevention and/or treatment.
Authors
Juan Huang, James A. Pearson, Jian Peng, Youjia Hu, Sha Sha, Yanpeng Xing, Gan Huang, Xia Li, Fang Hu, Zhiguo Xie, Yang Xiao, Shuoming Luo, Chen Chao, F. Susan Wong, Zhiguang Zhou, Li Wen
Abstract
The impact of transient ischemic-hypoxemic insults on the developing fetal brain is poorly understood despite evidence suggesting an association with neurodevelopmental disorders such as schizophrenia and autism. To address this, we designed an aberrant uterine hypercontractility paradigm with oxytocin to better assess the consequences of acute, but transient, placental ischemia-hypoxemia in term pregnant rats. Using MRI, we confirmed that oxytocin-induced aberrant uterine hypercontractility substantially compromised uteroplacental perfusion. This was supported by the observation of oxidative stress and increased lactate concentration in the fetal brain. Genes related to oxidative stress pathways were significantly upregulated in male, but not female, offspring 1 hour after oxytocin-induced placental ischemia-hypoxemia. Persistent upregulation of select mitochondrial electron transport chain complex proteins in the anterior cingulate cortex of adolescent male offspring suggested that this sex-specific effect was enduring. Functionally, offspring exposed to oxytocin-induced uterine hypercontractility showed male-specific abnormalities in social behavior with associated region-specific changes in gene expression and functional cortical connectivity. Our findings, therefore, indicate that even transient but severe placental ischemia-hypoxemia could be detrimental to the developing brain and point to a possible mitochondrial link between intrauterine asphyxia and neurodevelopmental disorders.
Authors
Arvind Palanisamy, Tusar Giri, Jia Jiang, Annie Bice, James D. Quirk, Sara B. Conyers, Susan E. Maloney, Nandini Raghuraman, Adam Q. Bauer, Joel R. Garbow, David F. Wozniak
Abstract
A complete understanding of human immune responses to Ebola virus infection is limited by the availability of specimens and the requirement for biosafety level 4 (BSL-4) containment. In an effort to bridge this gap, we evaluated cryopreserved PBMCs from 4 patients who survived Ebola virus disease (EVD) using an established mass cytometry antibody panel to characterize various cell populations during both the acute and convalescent phases. Acute loss of nonclassical monocytes and myeloid DCs, especially CD1c+ DCs, was noted. Classical monocyte proliferation and CD38 upregulation on plasmacytoid DCs coincided with declining viral load. Unsupervised analysis of cell abundance demonstrated acute declines in monocytic, NK, and T cell populations, but some populations, many of myeloid origin, increased in abundance during the acute phase, suggesting emergency hematopoiesis. Despite cell losses during the acute phase, upregulation of Ki-67 correlated with recovery of cell populations over time. These data provide insights into the human immune response during EVD.
Authors
Anita K. McElroy, Rama S. Akondy, David R. Mcllwain, Han Chen, Zach Bjornson-Hooper, Nilanjan Mukherjee, Aneesh K. Mehta, Garry Nolan, Stuart T. Nichol, Christina F. Spiropoulou
Abstract
Sepsis survivors suffer from increased vulnerability to infections, and lymphopenia presumably contributes to this problem. The mechanisms of the recovery of memory CD4+ T cells after sepsis remain elusive. We used the cecal ligation and puncture mouse model of sepsis to study the restoration of the memory CD4+ T cells during recovery from sepsis. Then, adoptive transfer of antigen-specific naive CD4+ T cells followed by immunization and BrdU labeling were performed to trace the proliferation and migration of memory CD4+ T cells. We revealed that the bone marrow (BM) is the primary site of CD4+ memory T cell homing and proliferation after sepsis-induced lymphopenia. Of interest, BM CD4+ T cells had a higher basal proliferation rate in comparison with splenic T cells. These cells also show features of resident memory T cells yet have the capacity to migrate outside the BM niche and engraft secondary lymphoid organs. The BM niche also sustains viability and functionality of CD4+ T cells. We also identified IL-7 as the major inducer of proliferation of the BM memory CD4+ T cells and showed that recombinant IL-7 improves the recovery of these cells. Taken together, we provide data on the mechanism and location of memory CD4+ T cell proliferation during recovery from septic lymphopenia, which are of relevance in studying immunostimulatory therapies in sepsis.
Authors
Tomasz Skirecki, Patrycja Swacha, Grażyna Hoser, Jakub Golab, Dominika Nowis, Ewa Kozłowska
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