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Abstract
BACKGROUND. Recessive dystrophic epidermolysis bullosa (RDEB) is a rare, devastating, and life-threatening inherited skin fragility disorder due to a lack of functional type VII collagen, for which no effective therapy exists. ABCB5-positive dermal mesenchymal stem cells (ABCB5+ MSCs) possess immunomodulatory capacities, a favorable skin homing potential and the ability to secrete type VII collagen. In a COL7A1–/– mouse model of RDEB, treatment with ABCB5+ MSCs markedly extended the animals’ lifespans. METHODS. In this international, multicentric, single-arm, phase I/IIa clinical trial, 16 patients (aged 4–36 years) enrolled into four age cohorts received three intravenous infusions of 2×106 ABCB5+ MSCs/kg on days 0, 17 and 35. Patients were followed up for 12 weeks regarding efficacy and 12 months regarding safety. RESULTS. At 12 weeks, statistically significant median (IQR) reductions in the Epidermolysis Bullosa Disease Activity and Scarring Index activity (EBDASI activity) score of 13.0% (2.9%-30%; P = 0.049) and the Instrument for Scoring Clinical Outcome of Research for Epidermolysis Bullosa clinician (iscorEB c) score of 18.2% (4.1%-41.7%; P = 0.037) were observed. Reductions in itch and pain numerical rating scale scores were greatest on day 35, amounting to 37.5% (0.0%-42.9%; P = 0.033) and 25.0% (-8.4%-46.4%; P = 0.168), respectively. Three adverse events were considered related to the cell product, one mild lymphadenopathy and two hypersensitivity reactions. The latter two were serious but resolved without sequelae shortly after withdrawal of treatment. CONCLUSION. This trial demonstrates good tolerability, manageable safety and potential efficacy of intravenous ABCB5+ MSCs as a readily available disease-modifying therapy for RDEB and provides a rationale for further clinical evaluation. TRIAL REGISTRATION. clinicaltrials.gov NCT03529877; EudraCT 2018-001009-98 FUNDING. The trial was sponsored by RHEACELL GmbH & Co. KG, Heidelberg, Germany. Contributions by NY Frank and MH Frank to this work were supported by the National Institutes of Health (NIH)/National Eye Institute (NEI) grants RO1EY025794 and R24EY028767.
Authors
Dimitra Kiritsi, Kathrin Dieter, Elke Niebergall-Roth, Silvia Fluhr, Cristina Daniele, Jasmina Esterlechner, Samar Sadeghi, Seda Ballikaya, Leoni Erdinger, Franziska Schauer, Stella Gewert, Martin Laimer, Johann W. Bauer, Alain Hovnanian, Giovanna Zambruno, May El Hachem, Emmanuelle Bourrat, Maria Papanikolaou, Gabriela Petrof, Sophie Kitzmüller, Christen L. Ebens, Markus H. Frank, Natasha Y. Frank, Christoph Ganss, Anna E. Martinez, John A. McGrath, Jakub Tolar, Mark A. Kluth
Abstract
Lung allograft rejection results in the accumulation of low molecular weight hyaluronic acid (LMW-HA), which further propagates inflammation and tissue injury. We have previously shown that therapeutic lymphangiogenesis in a murine model of lung allograft rejection reduced tissue LMW-HA and was associated with improved transplant outcomes. Herein we investigated the use of 4-Methylumbelliferone, a known inhibitor of HA synthesis, to alleviate acute allograft rejection in a murine model of lung transplantation. We found that treating mice with 4MU from day 20-30 post-transplant was sufficient to significantly improve outcomes, characterized by a reduction in T-cell mediated lung inflammation, LMW-HA content, and improved pathology scores. In vitro, 4MU directly attenuated activation, proliferation, and differentiation of naïve CD4+ T-cells into Th1 cells. As 4MU has already been demonstrated to be safe for human use, we believe examining 4MU for the treatment of acute lung allograft rejection may be of clinical significance.
Authors
Jewel Imani, Kaifeng Liu, Ye Cui, Jean-Pierre Assaker, Junwen Han, Auyon J. Ghosh, Julie Ng, Shikshya Shrestha, Anthony M. Lamattina, Pierce H. Louis, Anne Hentschel, Anthony J. Esposito, Ivan O. Rosas, Xiaoli Liu, Mark A. Perrella, Jamil Azzi, Gary Visner, Souheil El-Chemaly
Abstract
Patients with acute leukemia who are unable to achieve complete remission prior to allogeneic hematopoietic stem cell transplantation (SCT) have dismal outcomes with relapse rates well in excess of 60%. Haplo-identical SCT (haplo-SCT) may allow enhanced graft-versus-leukemia (GVL) effects by virtue of HLA class I/II donor-host disparities but typically requires intensive immune-suppression with post-transplant cyclophosphamide (PT-Cy) to prevent lethal graft-versus-host disease (GVHD). Here we demonstrate in preclinical models that glucocorticoid administration from day -1 to +5 inhibits alloantigen presentation by professional recipient antigen presenting cells in the gastrointestinal tract and prevents donor T-cell priming and subsequent expansion therein. In contrast, direct glucocorticoid signaling of donor T-cells promotes chemokine and integrin signatures permissive of preferential circulation and migration into the bone marrow, promoting donor T-cell residency. This results in significant reductions in GVHD whilst promoting potent GVL effects (relapse in recipients receiving glucocorticoids, vehicle or PT-Cy was 12%, 56% and 100% respectively). Intriguingly, patients with acute myeloid leukemia not in remission that received unmanipulated haplo-SCT and peri-transplant glucocorticoids also had an unexpectedly low relapse rate at 1 year (32%: 95% CI, 18%-47%) with high overall survival at 3 years (58%: 95% CI, 38-74%). These data highlight a potentially simple and effective approach to prevent relapse in patients with otherwise incurable leukemia that could be studied in prospective randomized trials.
Authors
Takayuki Inoue, Motoko Koyama, Katsuji Kaida, Kazuhiro Ikegame, Kathleen S. Ensbey, Luke Samson, Shuichiro Takahashi, Ping Zhang, Simone A. Minnie, Satoshi Maruyama, Shinichi Ishii, Takashi Daimon, Takahiro Fukuda, Hirohisa Nakamae, Takahide Ara, Yumiko Maruyama, Ken Ishiyama, Tatsuo Ichinohe, Yoshiko Atsuta, Bruce R. Blazar, Scott N. Furlan, Hiroyasu Ogawa, Geoffrey R. Hill
Abstract
Genetic variants in Granulin (GRN), which encodes the secreted glycoprotein Progranulin (PGRN), are associated with several neurodegenerative diseases including frontotemporal lobar degeneration, neuronal ceroid lipofuscinosis, and Alzheimer’s disease. These genetic alterations manifest in pathological changes due to a reduction of PGRN expression; therefore, identifying factors that can modulate PGRN levels in vivo would enhance our understanding of PGRN in neurodegeneration, and could reveal novel potential therapeutic targets. Here, we report that modulation of the endocytosis-lysosomal pathway via reduction of Nemo-like kinase (Nlk) in microglia, and not neurons, can alter total brain Pgrn levels in mice. We demonstrate that Nlk reduction promotes Pgrn degradation by enhancing its trafficking through endocytosis-lysosomal pathway, specifically in microglia. Furthermore, genetic interaction studies in mice showed that Nlk heterozygosity in Grn haploinsufficient mice further reduces Pgrn levels and induces neuropathological phenotypes associated with PGRN deficiency. Our results reveal a new mechanism for Pgrn level regulation in the brain through the active catabolism by microglia and provide insights into the pathophysiology of PGRN-associated diseases.
Authors
Tingting Dong, Leon Tejwani, Youngseob Jung, Hiroshi Kokubu, Kimberly Luttik, Terri M. Driessen, Janghoo Lim
Abstract
Influenza A virus (IAV) and SARS-CoV-2 are pandemic viruses causing millions of deaths, yet their clinical manifestations are distinctly different. With the hypothesis that upper airway immune and epithelial cells responses are also distinct, we performed single-cell RNA-sequencing (scRNA-Seq) on nasal wash cells freshly collected from adults with either acute COVID-19 or influenza or from healthy controls. We focused on major cell types and subtypes in a subset of donor samples. Nasal wash cells are enriched for macrophages and neutrophils for both influenza and COVID-19 compared to healthy controls. Hillock-like epithelial cells, M2-like macrophages, and age-dependent B cells are enriched in COVID-19 samples. A global decrease in interferon (IFN)-associated transcripts in neutrophils, macrophages, and epithelial cells is apparent in COVID-19 compared to influenza. The innate immune response to SARS-CoV-2 appears to be maintained in macrophages, despite evidence for limited epithelial immune sensing. Cell-to-cell interaction analyses reveal a decrease in epithelial interactions in COVID-19 and highlight differences in macrophage-macrophage interactions for COVID-19 and influenza. Our study demonstrates that scRNA-Seq can define host and viral transcriptional activity at the site of infection and reveal distinct local epithelial and immune cell responses for COVID-19 and influenza that may contribute to their divergent disease courses.
Authors
Kevin M. Gao, Alan G. Derr, Zhiru Guo, Kerstin Nundel, Ann Marshak-Rothstein, Robert W. Finberg, Jennifer P. Wang
Abstract
A diet high in fermentable oligosaccharides, disaccharides, monosaccharides, and polyols (FODMAPs) (HFM) induces gastrointestinal symptoms in patients with irritable bowel syndrome (IBS) and a diet low in FODMAPs (LFM) improves symptoms in up to 60% of IBS patients. However, the mechanism by which FODMAPs affect IBS symptoms is unclear. We showed that mice fed on an HFM diet have mast cell activation and colonic barrier loss. Using mast cell-deficient mice with/without mast cell reconstitution, we showed that HFM-mediated colonic barrier loss is dependent on TLR4-dependent mast cell activation. In in vitro studies, we demonstrated IBS fecal supernatant stimulates mast cell significantly more compared to fecal supernatant from healthy controls. This effect of IBS fecal supernatant on mast cell stimulation is ameliorated in absence of TLR4 receptor and after an LFM diet. Translating these findings into IBS patients, we found an LFM diet improves colonic barrier function and reduces mast cell activation while decreasing fecal LPS levels. Our findings indicate that a HFM diet causes mast cell activation via LPS which in turn leads to colonic barrier loss and an LFM diet reverses these pathophysiologic mucosal changes.
Authors
Prashant Singh, Gintautas Grabauskas, Shi-Yi Zhou, Jun Gao, Yawen Zhang, Chung Owyang
Abstract
Sarcomas contain a subpopulation of tumor propagating cells (TPCs) with enhanced tumor-initiating and self-renewal properties. However, it is unclear whether the TPC phenotype in sarcomas is stable or a dynamic cell state that can derive from non-tumor propagating cells (non-TPCs). In this study, we utilized a mouse model of undifferentiated pleomorphic sarcoma (UPS) to trace the lineage relationship between sarcoma side population (SP) cells that are enriched for TPCs and non-side population (non-SP) cells. By co-transplanting SP and non-SP cells expressing different endogenous fluorescent reporters, we show that non-SP cells can give rise to SP cells with enhanced tumor propagating potential in-vivo. Lineage trajectory analysis using single-cell RNA sequencing from SP and non-SP cells supports the notion that non-SP cells can assume the SP cell phenotype de novo. To test the effect of eradicating SP cells on tumor growth and self-renewal, we generated mouse sarcomas in which the Diphtheria Toxin Receptor (DTR) is expressed in the SP cells and their progeny. Ablation of the SP population using diphtheria toxin (DT) did not impede tumor growth or self-renewal. Together, we show that sarcoma SP is a dynamic cell state and targeting TPCs alone is insufficient to eliminate tumor progression.
Authors
Yuning Jackie Tang, Vijitha Puviindran, Yu Xiang, Yasuhito Yahara, Hongyuan Zhang, Puviindran Nadesan, Yarui Diao, David G. Kirsch, Benjamin A. Alman
Abstract
BACKGROUND. Little is known about the autoreactive B cells in anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). We aimed to investigate tolerance checkpoints of circulating antigen-specific proteinase 3 (PR3+) B cells. METHODS. Multicolor flow cytometry in combination with bioinformatics and functional in vitro studies were performed on baseline samples of peripheral blood mononuclear cells from 154 well-characterized participants of the RAVE trial (NCT00104299) with severely active PR3-AAV and myeloperoxidase (MPO)-AAV, and 27 healthy controls (HCs). Clinical data and outcomes from the trial were correlated with PR3+ B cells (total and subsets). RESULTS. The frequency of PR3+ B cells among circulating B cells was higher in PR3-AAV (4.77% [3.98%-6.01%]), than in MPO-AAV (3.16% [2.51%-5.22%]), and in AAV compared to HCs (1.67% [1.27%-2.16%], p<0.001 for all comparisons), implying a defective central tolerance checkpoint in patients. Only PBMC from PR3-AAV contained PR3+ B cells capable of secreting PR3-ANCA IgG in vitro, proving to be functionally distinct from those of MPO-AAV and HCs. Unsupervised clustering identified subtle subsets of atypical autoreactive PR3+ memory B cells accumulating through the maturation process in PR3-AAV patients. PR3+ B cells were enriched in the memory B cell compartment of PR3-AAV, and were associated with higher serum CXCL13 levels, suggesting an increased germinal center activity. PR3+ B cells correlated with systemic inflammation (C-reactive protein and erythrocyte sedimentation rate, p<0.05) and complete remission (p<0.001). CONCLUSIONS. This study suggests the presence of defective central antigen-independent and peripheral antigen-dependent checkpoints in patients in PR3-AAV, elucidating the selection process of autoreactive B cells.
Authors
Alvise Berti, Sophie Hillion, Amber M. Hummel, Young Min Son, Nedra Chriti, Tobias Peikert, Eva M. Carmona, Wayel H. Abdulahad, Peter Heeringa, Kristina M. Harris, E. William St. Clair, Paul Brunetta, Fernando Fervenza, Carol Langford, Cees Kallenberg, Peter Merkel, Paul A. Monach, Philip Seo, Robert F. Spiera, John H. Stone, Guido Grandi, Jie Sun, Jacques-Olivier Pers, Ulrich Specks, Divi Cornec
Abstract
HIV infection in the human gastrointestinal (GI) tract is thought to be central to HIV progression, but knowledge of this interaction is primarily limited to cohorts within westernized countries. Here, we present a large cohort recruited from high HIV endemic areas in South Africa and found that people living with HIV (PLWH) presented at a younger age for investigation in the GI clinic. We identified severe CD4 T-cell depletion in the GI tract, which was greater in the small intestine than in the large intestine and not correlated with years on ART or plasma viremia. HIV-p24 staining showed persistent viral expression, particularly in the colon, despite full suppression of plasma viremia. Quantification of mucosal ARV drugs revealed no differences in drug peneration between the duodemum and colon. Plasma markers of gut barrier breakdown and immune activation were elevated irrespective of HIV, but peripheral T-cell activation was inversely correlated with loss of gut CD4 T-cells in PLWH alone. T-cell activation is a strong predictor of HIV progression and independent of plasma viral load, implying that the irreversible loss of GI CD4 T-cells is a key event in the HIV pathogensis of PLWH in South Africa, yet the underlying mechanisms remain unknown.
Authors
Osaretin E. Asowata, Alveera Singh, Abigail Ngoepe, Nicholas Herbert, Rabiah Fardoos, Kavidha Reddy, Yenzekile Zungu, Faith Nene, Ntombifuthi Mthabela, Dirhona Ramjit, Farina Karim, Katya Govender, Thumbi Ndung’u, J. Zachary Porterfield, John H. Adamson, Fusi G. Madela, Vukani T. Manzini, Frank Anderson, Alasdair Leslie, Henrik N. Kløverpris
Abstract
Mounting evidence points to alterations in mitochondrial metabolism in renal cell carcinoma (RCC). However, the mechanisms that regulate the TCA cycle in RCC remain uncharacterized. Here, we demonstrate that loss of TCA cycle enzyme expression is retained in RCC metastatic tissues. Moreover, proteomic analysis demonstrates that reduced TCA cycle enzyme expression is far more pronounced in RCC relative to other tumor types. Loss of TCA cycle enzyme expression is correlated with reduced expression of the transcription factor peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) which is also lost in RCC tissues. PGC-1α re-expression in RCC cells restores the expression of TCA cycle enzymes in vitro and in vivo and leads to enhanced glucose carbon incorporation into TCA cycle intermediates. Mechanistically, TGF-β signaling, in concert with histone deacetylase 7 (HDAC7), suppresses TCA cycle enzyme expression. Our studies show that pharmacologic inhibition of TGF-β restores the expression of TCA cycle enzymes and suppresses tumor growth in an orthotopic model of RCC. Taken together, this investigation reveals a novel role for the TGF-β/HDAC7 axis in global suppression of TCA cycle enzymes in RCC and provides new insight into the molecular basis of altered mitochondrial metabolism in this malignancy.
Authors
Hyeyoung Nam, Anirban Kundu, Suman Karki, Garrett J. Brinkley, Darshan S. Chandrashekar, Richard L. Kirkman, Juan Liu, Maria V. Liberti, Jason W. Locasale, Tanecia Mitchell, Sooryanarayana Varambally, Sunil Sudarshan
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