Publications by authors named "Shigeyuki Shichino"

24 Publications

  • Page 1 of 1

Interleukin-11-expressing fibroblasts have a unique gene signature correlated with poor prognosis of colorectal cancer.

Nat Commun 2021 04 16;12(1):2281. Epub 2021 Apr 16.

Department of Biochemistry, Toho University School of Medicine, Tokyo, Japan.

Interleukin (IL)-11 is a member of the IL-6 family of cytokines and is involved in multiple cellular responses, including tumor development. However, the origin and functions of IL-11-producing (IL-11) cells are not fully understood. To characterize IL-11 cells in vivo, we generate Il11 reporter mice. IL-11 cells appear in the colon in murine tumor and acute colitis models. Il11ra1 or Il11 deletion attenuates the development of colitis-associated colorectal cancer. IL-11 cells express fibroblast markers and genes associated with cell proliferation and tissue repair. IL-11 induces the activation of colonic fibroblasts and epithelial cells through phosphorylation of STAT3. Human cancer database analysis reveals that the expression of genes enriched in IL-11 fibroblasts is elevated in human colorectal cancer and correlated with reduced recurrence-free survival. IL-11 fibroblasts activate both tumor cells and fibroblasts via secretion of IL-11, thereby constituting a feed-forward loop between tumor cells and fibroblasts in the tumor microenvironment.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-021-22450-3DOI Listing
April 2021

Transient Depletion of CD4 Cells Induces Remodeling of the TCR Repertoire in Gastrointestinal Cancer.

Cancer Immunol Res 2021 Mar 5. Epub 2021 Mar 5.

Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.

Antibody-mediated transient depletion of CD4 cells enhances the expansion of tumor-reactive CD8 T cells and exhibits robust antitumor effects in preclinical and clinical studies. To investigate the clonal T-cell responses following transient CD4 cell depletion in patients with cancer, we conducted a temporal analysis of the T-cell receptor (TCR) repertoire in the first-in-human clinical trial of IT1208, a defucosylated humanized monoclonal anti-CD4. Transient depletion of CD4 cells promoted replacement of T-cell clones among CD4 and CD8 T cells in the blood. This replacement of the TCR repertoire was associated with the extent of CD4 T-cell depletion and an increase in CD8 T-cell count in the blood. Next, we focused on T-cell clones overlapping between the blood and tumor in order to track tumor-associated T-cell clones in the blood. The total frequency of blood-tumor overlapping clones tended to increase in patients receiving a depleting dose of anti-CD4, which was accompanied by the replacement of overlapping clones. The greater expansion of CD8 overlapping clones was commonly observed in the patients who achieved tumor shrinkage. These results suggested that the clonal replacement of the TCR repertoire induced by transient CD4 cell depletion was accompanied by the expansion of tumor-reactive T-cell clones that mediated antitumor responses. Our findings propose beneficial remodeling of the TCR repertoire following transient CD4 cell depletion and provide novel insight into the antitumor effects of monoclonal anti-CD4 treatment in patients with cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/2326-6066.CIR-20-0989DOI Listing
March 2021

Generation of a p16 Reporter Mouse and Its Use to Characterize and Target p16 Cells In Vivo.

Cell Metab 2020 Nov 18;32(5):814-828.e6. Epub 2020 Sep 18.

Division of Molecular Regulation of Inflammatory and Immune Diseases, Research Institute of Biomedical Sciences, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-0022, Japan.

Cell senescence plays a key role in age-associated organ dysfunction, but the in vivo pathogenesis is largely unclear. Here, we generated a p16-Cre-tdTomato mouse model to analyze the in vivo characteristics of p16 cells at a single-cell level. We found tdTomato-positive p16 cells detectable in all organs, which were enriched with age. We also found that these cells failed to proliferate and had half-lives ranging from 2.6 to 4.2 months, depending on the tissue examined. Single-cell transcriptomics in the liver and kidneys revealed that p16 cells were present in various cell types, though most dominant in hepatic endothelium and in renal proximal and distal tubule epithelia, and that these cells exhibited heterogeneous senescence-associated phenotypes. Further, elimination of p16 cells ameliorated nonalcoholic steatohepatitis-related hepatic lipidosis and immune cell infiltration. Our new mouse model and single-cell analysis provide a powerful resource to enable the discovery of previously unidentified senescence functions in vivo.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cmet.2020.09.006DOI Listing
November 2020

The increased frequency of methicillin-resistant Staphylococcus aureus with low MIC of beta-lactam antibiotics isolated from hospitalized patients.

J Infect Chemother 2020 Jun 22;26(6):604-610. Epub 2020 Feb 22.

Department of Nephrology and Laboratory Medicine, Division of Blood Purification, Kanazawa University, Kanazawa, Japan.

Methicillin-resistant Staphylococcus aureus (MRSA) causes severe infectious diseases and can be life-threatening in healthcare-settings. MRSA is classified into health-care associated (HA)-MRSA strains and community acquired (CA)-MRSA strains based on genotype and phenotype. CA-MRSA has been reported to show the lower minimal inhibitory concentration (MIC) of some antibiotics as compared to HA-MRSA. Recently, the prevalence of CA-MRSA has been increased in worldwide. CA-MRSA is isolated not only from the healthy individuals in a community but also from the patients in healthcare settings. However, the changing trend in frequency of HA-MRSA and CA-MRSA in the hospital setting is not clear. Therefore, we analyzed the trend of MIC to speculate the frequency of HA-MRSA and CA-MRSA in the facility. Moreover, gene mutations were evaluated on resistant gene loci with next generation sequencer. The frequency of strains with low MIC of beta-lactam antibiotics was gradually increased in isolated MRSA strains from the hospitalized patients. Whole genome analysis revealed the frequency of gene mutation was also decreased in some resistant loci, such as blaZ and blaR1. These findings highlight the changing trend of MRSA strains isolated from hospitalized patients.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jiac.2020.01.016DOI Listing
June 2020

Collagen adhesion gene is associated with bloodstream infections caused by methicillin-resistant Staphylococcus aureus.

Int J Infect Dis 2020 Feb 15;91:22-31. Epub 2019 Nov 15.

Department of Molecular Preventive Medicine, University of Tokyo, Tokyo, Japan; Division of Molecular Regulation of Inflammatory and Immune Diseases, Research Institute of Biomedical Sciences, Tokyo University of Science, Noda, Japan.

Objectives: Methicillin-resistant Staphylococcus aureus (MRSA) causes hospital- and community-acquired infections. It is not clear whether genetic characteristics of the bacteria contribute to disease pathogenesis in MRSA infection. We hypothesized that whole genome analysis of MRSA strains could reveal the key gene loci and/or the gene mutations that affect clinical manifestations of MRSA infection.

Methods: Whole genome sequences (WGS) of MRSA of 154 strains were analyzed with respect to clinical manifestations and data. Further, we evaluated the association between clinical manifestations in MRSA infection and genomic information.

Results: WGS revealed gene mutations that correlated with clinical manifestations of MRSA infection. Moreover, 12 mutations were selected as important mutations by Random Forest analysis. Cluster analysis revealed strains associated with a high frequency of bloodstream infection (BSI). Twenty seven out of 34 strains in this cluster caused BSI. These strains were all positive for collagen adhesion gene (cna) and have mutations in the locus, those were selected by Random Forest analysis. Univariate and multivariate analysis revealed that these gene mutations were the predictor for the incidence of BSI. Interestingly, mutant CNA protein showed lower attachment ability to collagen, suggesting that the mutant protein might contribute to the dissemination of bacteria.

Conclusions: These findings suggest that the bacterial genotype affects the clinical characteristics of MRSA infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ijid.2019.11.003DOI Listing
February 2020

Products of Chemoenzymatic Synthesis Representing MUC1 Tandem Repeat Unit with T-, ST- or STn-antigen Revealed Distinct Specificities of Anti-MUC1 Antibodies.

Sci Rep 2019 11 12;9(1):16641. Epub 2019 Nov 12.

Division of Glycobiologics, Intractable Disease Research Center, Graduate School of Medicine, Juntendo University, 2-1-1 Hongo, Bunkyo-ku, Tokyo, 113-8421, Japan.

Anti-mucin1 (MUC1) antibodies have long been used clinically in cancer diagnosis and therapy and specific bindings of some of them are known to be dependent on the differential glycosylation of MUC1. However, a systematic comparison of the binding specificities of anti-MUC1 antibodies was not previously conducted. Here, a total of 20 glycopeptides including the tandem repeat unit of MUC1, APPAHGVTSAPDTRPAPGSTAPPAHGV with GalNAc (Tn-antigen), Galβ1-3GalNAc (T-antigen), NeuAcα2-3Galβ1-3GalNAc (sialyl-T-antigen), or NeuAcα2-6GalNAc (sialyl-Tn-antigen) at each threonine or serine residue were prepared by a combination of chemical glycopeptide synthesis and enzymatic extension of carbohydrate chains. These glycopeptides were tested by the enzyme-linked immunosorbent assay (ELISA) for their capacity to bind 13 monoclonal antibodies (mAbs) known to be specific for MUC1. The results indicated that anti-MUC1 mAbs have diverse specificities but can be classified into a few characteristic groups based on their binding pattern toward glycopeptides in some cases having a specific glycan at unique glycosylation sites. Because the clinical significance of some of these antibodies was already established, the structural features identified by these antibodies as revealed in the present study should provide useful information relevant to their further clinical use and the biological understanding of MUC1.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-019-53052-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6851390PMC
November 2019

First-in-human phase 1 study of IT1208, a defucosylated humanized anti-CD4 depleting antibody, in patients with advanced solid tumors.

J Immunother Cancer 2019 07 24;7(1):195. Epub 2019 Jul 24.

Department of Experimental Therapeutics, National Cancer Center Hospital East, 6-5-1 Kashiwanoha, Kashiwa, Chiba, 277-8577, Japan.

Background: Transient CD4 T cell depletion led to the proliferation of tumor-specific CD8 T cells in the draining lymph node and increased infiltration of PD-1CD8 T cells into the tumor, which resulted in strong anti-tumor effects in tumor-bearing mice. This is a first-in-human study of IT1208, a defucosylated humanized anti-CD4 monoclonal antibody, engineered to exert potent antibody-dependent cellular cytotoxicity.

Methods: Patients with advanced solid tumors were treated with intravenous IT1208 at doses of 0.1 or 1.0 mg/kg. The first patient in each cohort received a single administration, and the other patients received two administrations of IT1208 on days 1 and 8.

Results: Eleven patients were enrolled in the 0.1 mg/kg (n = 4) and 1.0 mg/kg cohorts (n = 7). Grade 1 or 2 infusion-related reactions was observed in all patients. Decreased CD4 T cells in peripheral blood due to IT1208 were observed in all patients and especially in those receiving two administrations of 1.0 mg/kg. CD8 T cells increased on day 29 compared with baseline in most patients, resulting in remarkably decreased CD4/8 ratios. One microsatellite-stable colon cancer patient achieved durable partial response showing increased infiltration of both CD4 and CD8 T cells into tumors after IT1208 administration. Moreover, transcriptomic profiling of the liver metastasis of the patient revealed upregulation of the expression of interferon-stimulated genes, T cell activation-related genes, and antigen presentation-related genes after IT1208 administration. Two additional patients with gastric or esophageal cancer achieved stable disease lasting at least 3 months.

Conclusions: IT1208 monotherapy successfully depleted CD4 T cells with a manageable safety profile and encouraging preliminary efficacy signals, which warrants further investigations, especially in combination with immune checkpoint inhibitors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s40425-019-0677-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6657210PMC
July 2019

In vitro expansion of endogenous human alveolar epithelial type II cells in fibroblast-free spheroid culture.

Biochem Biophys Res Commun 2019 08 6;515(4):579-585. Epub 2019 Jun 6.

Division of Molecular Regulation of Inflammatory and Immune Diseases, Research Institute of Biomedical Sciences, Tokyo University of Science, Noda, 278-0022, Japan. Electronic address:

Alveolar epithelial type II cells (AEC2) are stem cells of the alveoli and play crucial roles in maintaining lung homeostasis and the pathogenesis of lung diseases. We recently reported on an organoid culture system for endogenous murine AEC2. Despite advances in generation of human induced pluripotent stem cell-derived AEC2, in vitro expansion of endogenous human AEC2 has not been reported and genetic manipulation of human AEC2 has been difficult. Here, we show that endogenous human AEC2 could be cultured and passaged using a three-dimensional culture system with a specific combination of signal ligands and inhibitors. The culture system was suitable for retroviral gene transduction into AEC2. Transduction of pulmonary fibrosis-associated mutant surfactant protein C (SFPTC) into AEC2 revealed characteristic transcriptional traits similar to those of patients with idiopathic pulmonary fibrosis. Our culture system will be a useful tool for investigating human AEC2 functions in vitro.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bbrc.2019.05.187DOI Listing
August 2019

Gli signaling pathway modulates fibroblast activation and facilitates scar formation in pulmonary fibrosis.

Biochem Biophys Res Commun 2019 06 8;514(3):684-690. Epub 2019 May 8.

Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan; Division of Molecular Regulation of Inflammatory and Immune Diseases, Research Institute of Biomedical Sciences, Tokyo University of Science, Noda, Japan; Japan Agency for Medical Research and Development-CREST Program, Tokyo, Japan. Electronic address:

Pulmonary fibrosis is characterized by progressive and irreversible scarring of alveoli, which causes reduction of surface epithelial area and eventually respiratory failure. The precise mechanism of alveolar scarring is poorly understood. In this study, we explored transcriptional signatures of activated fibroblasts in alveolar airspaces by using intratracheal transfer in bleomycin-induced lung fibrosis. Lung fibroblasts transferred into injured alveoli upregulated genes related to translation and metabolism in the first two days, and upregulated genes related to extracellular matrix (ECM) production between day 2 and 7. Upstream analysis of these upregulated genes suggested possible contribution of hypoxia-inducible factors 1a (Hif1a) to fibroblast activation in the first two days, and possible contribution of kruppel-like factor 4 (Klf4) and glioma-associated oncogene (Gli) transcription factors to fibroblast activation in the following profibrotic phase. Fibroblasts purified based on high Acta2 expression after intratracheal transfer were also characterized by ECM production and upstream regulation by Klf4 and Gli proteins. Pharmacological inhibition of Gli proteins by GANT61 in bleomycin-induced lung fibrosis altered the pattern of scarring characterized by dilated airspaces and smaller fibroblast clusters. Activated fibroblasts isolated from GANT61-treated mice showed decreased migration capacity, suggesting that Gli signaling inhibition attenuated fibroblast activation. In conclusion, we revealed transcriptional signatures and possible upstream regulators of activated fibroblasts in injured alveolar airspaces. The altered scar formation by Gli signaling inhibition supports that activated fibroblasts in alveolar airspaces may play a critical role in scar formation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bbrc.2019.05.011DOI Listing
June 2019

Engraftment and proliferation potential of embryonic lung tissue cells in irradiated mice with emphysema.

Sci Rep 2019 03 6;9(1):3657. Epub 2019 Mar 6.

Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, 113-0033, Japan.

Recently, there has been increasing interest in stem cell transplantation therapy, to treat chronic respiratory diseases, using lung epithelial cells or alveolospheres derived from endogenous lung progenitor cells. However, optimal transplantation strategy of these cells has not been addressed. To gain insight into the optimization of stem cell transplantation therapy, we investigated whether lung cell engraftment potential differ among different developmental stages. After preconditioning with irradiation and elastase to induce lung damage, we infused embryonic day 15.5 (E15.5) CAG-EGFP whole lung cells, and confirmed the engraftment of epithelial cells, endothelial cells, and mesenchymal cells. The number of EGFP-positive epithelial cells increased from day 7 to 28 after infusion. Among epithelial cells derived from E13.5, E15.5, E18.5, P7, P14, and P56 mice, E15.5 cells demonstrated the most efficient engraftment. In vitro, E15.5 epithelial cells showed high proliferation potential. Transcriptome analyses of sorted epithelial cells from E13.5, E15.5, E18.5, P14, and P56 mice revealed that cell cycle and cell-cell adhesion genes were highly enriched in E15.5 epithelial cells. Our findings suggest that cell therapy for lung diseases might be most effective when epithelial cells with transcriptional traits similar to those of E15.5 epithelial cells are used.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-019-40237-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6403395PMC
March 2019

TCR Repertoire Analysis Reveals Mobilization of Novel CD8 T Cell Clones Into the Cancer-Immunity Cycle Following Anti-CD4 Antibody Administration.

Front Immunol 2018 24;9:3185. Epub 2019 Jan 24.

Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.

Depletion of CD4 cells using an anti-CD4 monoclonal antibody (anti-CD4 mAb) induces the expansion of tumor-reactive CD8 T cells and strong antitumor effects in several murine tumor models. However, it is not known whether the anti-CD4 mAb treatment activates a particular or a broad spectrum of tumor-reactive CD8 T cell clones. To investigate the changes in the TCR repertoire induced by the anti-CD4 mAb treatment, we performed unbiased high-throughput TCR sequencing in a B16F10 mouse subcutaneous melanoma model. By Inter-Organ Clone Tracking analysis, we demonstrated that anti-CD4 mAb treatment increased the diversity and combined frequency of CD8 T cell clones that overlapped among the tumor, draining lymph node (dLN), and peripheral blood repertoires. Interestingly, the anti-CD4 mAb treatment-induced expansion of overlapping clones occurred mainly in the dLN rather than in the tumor. Overall, the Inter-Organ Clone Tracking analysis revealed that anti-CD4 mAb treatment enhances the mobilization of a wide variety of tumor-reactive CD8 T cell clones into the Cancer-Immunity Cycle and thus induces a robust antitumor immune response in mice.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fimmu.2018.03185DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6353793PMC
October 2019

Combined treatment with HMGN1 and anti-CD4 depleting antibody reverses T cell exhaustion and exerts robust anti-tumor effects in mice.

J Immunother Cancer 2019 01 29;7(1):21. Epub 2019 Jan 29.

Division of Molecular Regulation of Inflammatory and Immune Diseases, Research Institute for Biomedical Sciences, Tokyo University of Science, Chiba, Japan.

Background: Transient depletion of CD4 T cells results in tumor suppression and survival benefit in murine models; however, the tumor progression and recurrence still occur over more long-term monitoring of mice. Thus, we explored an additional strategy to enhance endogenous immune responses by an alarmin, high mobility group nucleosome binding protein 1 (HMGN1).

Methods: The anti-tumor effects of HMGN1, anti-CD4 depleting antibody, and their combined treatment were monitored in the Colon26 or the B16F10 subcutaneous murine models. The tumor-infiltrating CD8 T cell proliferation, differentiation, exhaustion, and its gene expression were determined by flow cytometry, transcriptome analysis, and quantitative real-time PCR.

Results: Our results show that a systemic administration of low doses of HMGN1 with an anti-CD4 depleting antibody (HMGN1/αCD4) promoted expansion of CD8 T cell populations (e.g. CD137 PD-1 and CD44 PD-1), recruited CCR7 migratory dendritic cells to the tumor, and reduced co-inhibitory molecules (e.g. PD-1, LAG-3, and TIM-3) to counteract CD8 T cell exhaustion.

Conclusion: The HMGN1/αCD4 treatment expanded effector CD8 T cells and prolonged their anti-tumor activities by rescuing them from exhaustion, thus resulting in tumor regression and even rejection in long-term monitored mice.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s40425-019-0503-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6352494PMC
January 2019

Mesenchymal-Epithelial Interactome Analysis Reveals Essential Factors Required for Fibroblast-Free Alveolosphere Formation.

iScience 2019 Jan 26;11:318-333. Epub 2018 Dec 26.

Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo 113-0033, Japan; Division of Molecular Regulation of Inflammatory and Immune Diseases, Research Institute of Biomedical Sciences, Tokyo University of Science, Noda 278-0022, Japan. Electronic address:

Lung epithelial cells and fibroblasts are key cell populations in lung development. Fibroblasts support type 2 alveolar epithelial cells (AEC2) in the developing and mature lung. However, fibroblast-AEC2 interactions have not been clearly described. We addressed this in the present study by time course serial analysis of gene expression sequencing (SAGE-seq) of epithelial cells and fibroblasts of developing and mature murine lungs. We identified lung fibroblast-epithelial interactions that potentially regulate alveologenesis and are mediated by fibroblast-expressed ligands and epithelial cell surface receptors. In the epithelial-fibroblast co-culture alveolosphere formation assay, single intervention against fibroblast-expressed ligand or associated signaling cascades promoted or inhibited alveolosphere growth. Adding the ligand-associated molecules fibroblast growth factor 7 and Notch ligand and inhibitors of bone morphogenetic protein 4, transforming growth factor β, and glycogen synthase kinase-3β to the culture medium enabled fibroblast-free alveolosphere formation. The results revealed the essential factors regulating fibroblast-AEC2 interactions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.isci.2018.12.022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6329323PMC
January 2019

Transcriptome network analysis identifies protective role of the LXR/SREBP-1c axis in murine pulmonary fibrosis.

JCI Insight 2019 Jan 10;4(1). Epub 2019 Jan 10.

Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.

Pulmonary fibrosis (PF) is an intractable disorder with a poor prognosis. Although lung fibroblasts play a central role in PF, the key regulatory molecules involved in this process remain unknown. To address this issue, we performed a time-course transcriptome analysis on lung fibroblasts of bleomycin- and silica-treated murine lungs. We found gene modules whose expression kinetics were associated with the progression of PF and human idiopathic PF (IPF). Upstream analysis of a transcriptome network helped in identifying 55 hub transcription factors that were highly connected with PF-associated gene modules. Of these hubs, the expression of Srebf1 decreased in line with progression of PF and human IPF, suggesting its suppressive role in fibroblast activation. Consistently, adoptive transfer and genetic modification studies revealed that the hub transcription factor SREBP-1c suppressed PF-associated gene expression changes in lung fibroblasts and PF pathology in vivo. Moreover, therapeutic pharmacological activation of LXR, an SREBP-1c activator, suppressed the Srebf1-dependent activation of fibroblasts and progression of PF. Thus, SREBP-1c acts as a protective hub of lung fibroblast activation in PF. Collectively, the findings of the current study may prove to be valuable in the development of effective therapeutic strategies for PF.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1172/jci.insight.122163DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6485671PMC
January 2019

Lung fibroblasts express a miR-19a-19b-20a sub-cluster to suppress TGF-β-associated fibroblast activation in murine pulmonary fibrosis.

Sci Rep 2018 11 9;8(1):16642. Epub 2018 Nov 9.

Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.

Lung fibroblasts play a pivotal role in pulmonary fibrosis, a devastating lung disease, by producing extracellular matrix. MicroRNAs (miRNAs) suppress numerous genes post-transcriptionally; however, the roles of miRNAs in activated fibroblasts in fibrotic lungs remain poorly understood. To elucidate these roles, we performed global miRNA-expression profiling of fibroblasts from bleomycin- and silica-induced fibrotic lungs and investigated the functions of miRNAs in activated lung fibroblasts. Clustering analysis of global miRNA-expression data identified miRNA signatures exhibiting increased expression during fibrosis progression. Among these signatures, we found that a miR-19a-19b-20a sub-cluster suppressed TGF-β-induced activation of fibroblasts in vitro. Moreover, to elucidate whether fibroblast-specific intervention against the sub-cluster modulates pathogenic activation of fibroblasts in fibrotic lungs, we intratracheally transferred the sub-cluster-overexpressing fibroblasts into bleomycin-treated lungs. Global transcriptome analysis of the intratracheally transferred fibroblasts revealed that the sub-cluster not only downregulated expression of TGF-β-associated pro-fibrotic genes, including Acta2, Col1a1, Ctgf, and Serpine1, but also upregulated expression of the anti-fibrotic genes Dcn, Igfbp5, and Mmp3 in activated lung fibroblasts. Collectively, these findings indicated that upregulation of the miR-19a-19b-20a sub-cluster expression in lung fibroblasts counteracted TGF-β-associated pathogenic activation of fibroblasts in murine pulmonary fibrosis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-018-34839-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6226532PMC
November 2018

Reduction of Proliferating Olfactory Cells and Low Expression of Extracellular Matrix Genes Are Hallmarks of the Aged Olfactory Mucosa.

Front Aging Neurosci 2018 27;10:86. Epub 2018 Mar 27.

Department of Otolaryngology, The University of Tokyo, Tokyo, Japan.

The incidence of olfactory impairment increases with age; however, the detailed molecular and cellular mechanisms underlying this increase are yet to be determined. We examined the influence of aging on olfactory receptor neurons (ORNs), which are maintained by a unique stem cell system, from olfactory progenitor cells to mature ORNs, by histological comparisons of the physiological status of the olfactory epithelium between young adult and aged mice. Furthermore, we clarified the expression of genes encoding inflammatory cytokines, neurotrophins, growth factors, and extracellular matrix proteins to reveal the molecular mechanisms underlying olfactory impairment caused by aging. The numbers of mature and immature ORNs, but not olfactory progenitors, decreased in the aged olfactory epithelium, with a concurrent reduction in Ki-67-positive proliferating cells. Transcriptome analyses revealed an increase in , encoding a component of senescence-associated secretary phenotypes (SASP), and a decrease in , encoding a growth factor for ORNs, in the aged nasal mucosa. Interestingly, expression levels of several extracellular matrix genes, including , decreased in the aged nasal mucosa. Consistent with the transcriptional changes, the number of -GFP-positive cells decreased in the aged lamina propria. Our data suggest that reduction in ORN number and cell proliferation, reduced extracellular matrix gene expression, and increased SASP contribute to olfactory impairment during aging.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fnagi.2018.00086DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5880952PMC
March 2018

Long-Lasting Graft-Derived Donor T Cells Contribute to the Pathogenesis of Chronic Graft-versus-Host Disease in Mice.

Front Immunol 2017 18;8:1842. Epub 2017 Dec 18.

Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.

Chronic graft-versus-host disease (cGVHD) is a major complication in long-term survivors of allogeneic hematopoietic stem cell transplantation (allo-HSCT). Graft-derived T cells (T) have been implicated in the induction of cGVHD; however, the extent of their contribution to the pathogenesis of cGVHD remains unclear. Using a mouse model of cGVHD, we demonstrate that T predominate over hematopoietic stem cell-derived T cells generated (T) in cGVHD-affected organs such as the liver and lung even at day 63 after allo-HSCT. Persisting T, in particular CD8 T, not only displayed an exhausted or senescent phenotype but also contained a substantial proportion of cells that had the potential to proliferate and produce inflammatory cytokines. Host antigens indirectly presented by donor HSC-derived hematopoietic cells were involved in the maintenance of T in the reconstituted host. Selective depletion of T in the chronic phase of disease resulted in the expansion of T and thus neither the survival nor histopathology of cGVHD was ameliorated. On the other hand, T depletion caused activation of T and resulted in a lethal T-mediated exacerbation of GVHD. The findings presented here clarify the pathological role of long-lasting T in cGVHD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fimmu.2017.01842DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5741650PMC
December 2017

Simian Immunodeficiency Virus Targeting of CXCR3 CD4 T Cells in Secondary Lymphoid Organs Is Associated with Robust CXCL10 Expression in Monocyte/Macrophage Subsets.

J Virol 2017 07 9;91(13). Epub 2017 Jun 9.

AIDS Research Center, National Institute of Infectious Diseases, Tokyo, Japan

Glycosylation of Env defines pathogenic properties of simian immunodeficiency virus (SIV). We previously demonstrated that pathogenic SIVmac239 and a live-attenuated, quintuple deglycosylated Env mutant (Δ5G) virus target CD4 T cells residing in different tissues during acute infection. SIVmac239 and Δ5G preferentially infected distinct CD4 T cells in secondary lymphoid organs (SLOs) and within the lamina propria of the small intestine, respectively (C. Sugimoto et al., J Virol 86:9323-9336, 2012, https://doi.org/10.1128/JVI.00948-12). Here, we studied the host responses relevant to SIV targeting of CXCR3 CCR5 CD4 T cells in SLOs. Genome-wide transcriptome analyses revealed that Th1-polarized inflammatory responses, defined by expression of CXCR3 chemokines, were distinctly induced in the SIVmac239-infected animals. Consistent with robust expression of CXCL10, CXCR3 T cells were depleted from blood in the SIVmac239-infected animals. We also discovered that elevation of CXCL10 expression in blood and SLOs was secondary to the induction of CD14 CD16 monocytes and MAC387 macrophages, respectively. Since the significantly higher levels of SIV infection in SLOs occurred with a massive accumulation of infiltrated MAC387 macrophages, T cells, dendritic cells (DCs), and residential macrophages near high endothelial venules, the results highlight critical roles of innate/inflammatory responses in SIVmac239 infection. Restricted infection in SLOs by Δ5G also suggests that glycosylation of Env modulates innate/inflammatory responses elicited by cells of monocyte/macrophage/DC lineages. We previously demonstrated that a pathogenic SIVmac239 virus and a live-attenuated, deglycosylated mutant Δ5G virus infected distinct CD4 T cell subsets in SLOs and the small intestine, respectively (C. Sugimoto et al., J Virol 86:9323-9336, 2012, https://doi.org/10.1128/JVI.00948-12). Accordingly, infections with SIVmac239, but not with Δ5G, deplete CXCR3 CCR5 CD4 T (Th1) cells during the primary infection, thereby compromising the cellular immune response. Thus, we hypothesized that distinct host responses are elicited by the infections with two different viruses. We found that SIVmac239 induced distinctly higher levels of inflammatory Th1 responses than Δ5G. In particular, SIVmac239 infection elicited robust expression of CXCL10, a chemokine for CXCR3 cells, in CD14 CD16 monocytes and MAC387 macrophages recently infiltrated in SLOs. In contrast, Δ5G infection elicited only modest inflammatory responses. These results suggest that the glycosylation of Env modulates the inflammatory/Th1 responses through the monocyte/macrophage subsets and elicits marked differences in SIV infection and clinical outcomes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/JVI.00439-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5469254PMC
July 2017

Reduced supply of monocyte-derived macrophages leads to a transition from nodular to diffuse lesions and tissue cell activation in silica-induced pulmonary fibrosis in mice.

Am J Pathol 2015 Nov 9;185(11):2923-38. Epub 2015 Oct 9.

Department of Molecular Preventive Medicine, Graduate School of Medicine, the University of Tokyo, Tokyo, Japan; Japan Science and Technology Agency, Core Research for Evolutional Science and Technology (CREST), Tokyo, Japan. Electronic address:

Pulmonary fibrosis (PF) is an intractable disorder with a poor prognosis. Lung macrophages have been reported to regulate both progression and remission of bleomycin-induced diffuse PF. However, it remains unclear how macrophages contribute to silica-induced progressive nodular PF and the associated tissue cell responses in vivo. We found that lack of monocyte-derived macrophages results in the formation of diffuse PF after silica instillation. We found that the proportion and the number of monocyte-derived macrophages were persistently higher in silica-induced progressive PF compared with bleomycin-induced PF. Surprisingly, in Ccr2(-/-) mice, in which monocyte-derived macrophage infiltration is impaired, silica administration induced diffuse PF with loose nodule formation and greater activation of tissue cells. In the diffuse lesions, the distribution of epithelial cells, distribution of myofibroblasts, and architecture of the basement membrane were disrupted. Consistent with the development of diffuse lesions, genes that were differentially expressed in CD45(-) tissue cells from the lung of wild-type and Ccr2(-/-) mice were highly enriched in human diffuse, progressive PF. In gene ontology network analyses, many of these genes were associated with tissue remodeling and included genes not previously associated with PF, such as Mmp14, Thbs2, and Fgfr4. Overall, these results indicate that monocyte-derived macrophages prevent transition from nodular to diffuse silica-induced PF, potentially by regulating tissue cell responses.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ajpath.2015.07.013DOI Listing
November 2015

Intratracheal cell transfer demonstrates the profibrotic potential of resident fibroblasts in pulmonary fibrosis.

Am J Pathol 2015 Nov 9;185(11):2939-48. Epub 2015 Oct 9.

Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan; Japan Agency for Medical Research and Development-Core Research for Evolutional Science and Technology (AMED-CREST) Program, Tokyo, Japan. Electronic address:

Pulmonary fibrosis is a devastating disease for which there are few effective therapies. Activated fibroblasts form subepithelial clusters known as fibroblastic foci, which are characterized by excessive collagen deposition. The origin of activated fibroblasts is controversial and needs to be clarified to understand their pathogenicity. Here, using an intratracheal adoptive cell transfer method, we show that resident fibroblasts in alveolar walls have the highest profibrotic potential. By using collagen I(α)2-green fluorescent protein and neural/glial antigen 2-DsRed fluorescent reporter mice, we identified resident fibroblasts and pericytes in the alveolar walls based on surface marker expression and ultrastructural characteristics. In the early phase of bleomycin-induced pulmonary fibrosis, activated fibroblasts migrated into epithelium-denuded alveolar airspaces. Purified resident fibroblasts delivered into injured alveoli by an intratracheal route showed similar activated signatures as activated fibroblasts and formed fibroblastic foci. Neither pericytes nor epithelial cells had the same profibrotic potential. Transferred resident fibroblasts highly up-regulated profibrotic genes including α-smooth muscle actin and were a significant source of collagen deposition. These data provide insights into the cellular mechanisms of fibrogenesis and show intratracheal cell transfer to be a useful tool for exploring novel therapeutic targets against pulmonary fibrosis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ajpath.2015.07.022DOI Listing
November 2015

Robust Antitumor Effects of Combined Anti-CD4-Depleting Antibody and Anti-PD-1/PD-L1 Immune Checkpoint Antibody Treatment in Mice.

Cancer Immunol Res 2015 Jun 20;3(6):631-40. Epub 2015 Feb 20.

Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.

Depletion of CD4(+) cells in tumor-bearing mice has strong antitumor effects. However, the mechanisms underlying these effects and the therapeutic benefits of CD4(+) cell depletion relative to other immunotherapies have not been fully evaluated. Here, we investigated the antitumor effects of an anti-CD4-depleting mAb as a monotherapy or in combination with immune checkpoint mAbs. In B16F10, Colon 26, or Lewis lung carcinoma subcutaneous tumor models, administration of the anti-CD4 mAb alone had strong antitumor effects that were superior to those elicited by CD25(+) Treg depletion or other immune checkpoint mAbs, and which were completely reversed by CD8(+) cell depletion. CD4(+) cell depletion led to the proliferation of tumor-specific CD8(+) T cells in the draining lymph node and increased infiltration of PD-1(+)CD8(+) T cells into the tumor, with a shift toward type I immunity within the tumor. Combination treatment with the anti-CD4 mAb and immune checkpoint mAbs, particularly anti-PD-1 or anti-PD-L1 mAbs, synergistically suppressed tumor growth and greatly prolonged survival. To our knowledge, this work represents the first report of robust synergy between anti-CD4 and anti-PD-1 or anti-PD-L1 mAb therapies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/2326-6066.CIR-14-0190DOI Listing
June 2015

Lymph node stromal cells negatively regulate antigen-specific CD4+ T cell responses.

J Immunol 2014 Aug 14;193(4):1636-44. Epub 2014 Jul 14.

Department of Molecular Preventive Medicine, Graduate School of Medicine, University of Tokyo, Tokyo 113-0033, Japan; Japan Science and Technology Agency, Tokyo 102-8666, Japan;

Lymph node (LN) stromal cells (LNSCs) form the functional structure of LNs and play an important role in lymphocyte survival and the maintenance of immune tolerance. Despite their broad spectrum of function, little is known about LNSC responses during microbial infection. In this study, we demonstrate that LNSC subsets display distinct kinetics following vaccinia virus infection. In particular, compared with the expansion of other LNSC subsets and the total LN cell population, the expansion of fibroblastic reticular cells (FRCs) was delayed and sustained by noncirculating progenitor cells. Notably, newly generated FRCs were preferentially located in perivascular areas. Viral clearance in reactive LNs preceded the onset of FRC expansion, raising the possibility that viral infection in LNs may have a negative impact on the differentiation of FRCs. We also found that MHC class II expression was upregulated in all LNSC subsets until day 10 postinfection. Genetic ablation of radioresistant stromal cell-mediated Ag presentation resulted in slower contraction of Ag-specific CD4(+) T cells. We propose that activated LNSCs acquire enhanced Ag-presentation capacity, serving as an extrinsic brake system for CD4(+) T cell responses. Disrupted function and homeostasis of LNSCs may contribute to immune deregulation in the context of chronic viral infection, autoimmunity, and graft-versus-host disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.4049/jimmunol.1302946DOI Listing
August 2014

Tracking of intertissue migration reveals the origins of tumor-infiltrating monocytes.

Proc Natl Acad Sci U S A 2014 May 13;111(21):7771-6. Epub 2014 May 13.

Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan;

Myeloid cells such as monocytes and monocyte-derived macrophages promote tumor progression. Recent reports suggest that extramedullary hematopoiesis sustains a sizable reservoir of tumor-infiltrating monocytes in the spleen. However, the influence of the spleen on tumor development and the extent to which spleen monocytes populate the tumor relative to bone marrow (BM) monocytes remain controversial. Here, we used mice expressing the photoconvertible protein Kikume Green-Red to track the redistribution of monocytes from the BM and spleen, and mice expressing fluorescent ubiquitination-based cell-cycle indicator proteins to monitor active hematopoiesis in these tissues. In mice bearing late-stage tumors, the BM, besides being the major site of monocyte production, supplied the expansion of the spleen reservoir, replacing 9% of spleen monocytes every hour. Deployment of monocytes was equally rapid from the BM and the spleen. However, BM monocytes were younger than those in the spleen and were 2.7 times more likely to migrate into the tumor from the circulation. Partly as a result of this intrinsic difference in migration potential, spleen monocytes made only a minor contribution to the tumor-infiltrating monocyte population. At least 27% of tumor monocytes had traveled from the BM in the last 24 h, compared with only 2% from the spleen. These observations highlight the importance of the BM as the primary hematopoietic tissue and monocyte reservoir in tumor-bearing mice, despite the changes that occur in the spleen monocyte reservoir during tumor development.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.1402914111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4040600PMC
May 2014

Qualitative rather than quantitative changes are hallmarks of fibroblasts in bleomycin-induced pulmonary fibrosis.

Am J Pathol 2013 Sep 22;183(3):758-73. Epub 2013 Jul 22.

Department of Molecular Preventive Medicine, Graduate School of Medicine, University of Tokyo, Tokyo, Japan.

Pulmonary fibrosis is characterized by accumulation of activated fibroblasts that produce excessive amounts of extracellular matrix components such as collagen type I. However, the dynamics and activation signatures of fibroblasts during fibrogenesis remain poorly understood, especially in vivo. We examined changes in lung tissue cell populations and in the phenotype of activated fibroblasts after acute injury in a model of bleomycin-induced pulmonary fibrosis. Despite clustering of collagen type I-producing fibroblasts in fibrotic regions, flow cytometry-based quantitative analysis of whole lungs revealed that the number of fibroblasts in the lungs remained constant. At the peak of inflammation, fibroblast proliferation and apoptosis were both increased, suggesting that the clustering was not merely a result of proliferation, but also of fibroblast migration from nearby alveolar walls. Parabiosis experiments demonstrated that fibroblasts were not supplied from the circulation. Comprehensive gene expression analysis of freshly isolated fibroblasts revealed a detailed activation signature associated with fibrogenesis, including changes in genes responsible for migration and extracellular matrix construction. The Spp1 gene, which encodes osteopontin, was highly up-regulated and was an identifying characteristic of activated fibroblasts present at the sites of remodeling. Osteopontin may serve as a useful marker of profibrotic fibroblasts. These results provide insights into the cellular and molecular mechanisms underlying pulmonary fibrosis and provide a foundation for development of specific antifibrotic therapies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ajpath.2013.06.005DOI Listing
September 2013