Journal of immunology (Baltimore, Md. : 1950)
Silk, JD;Abbott, RJM;Adams, KJ;Bennett, AD;Brett, S;Cornforth, TV;Crossland, KL;Figueroa, DJ;Jing, J;O'Connor, C;Pachnio, A;Patasic, L;Peredo, CE;Quattrini, A;Quinn, LL;Rust, AG;Saini, M;Sanderson, JP;Steiner, D;Tavano, B;Viswanathan, P;Wiedermann, GE;Wong, R;Jakobsen, BK;Britten, CM;Gerry, AB;Brewer, JE;
PMID: 34853077 | DOI: 10.4049/jimmunol.2001357
Adoptive T cell therapy with T cells expressing affinity-enhanced TCRs has shown promising results in phase 1/2 clinical trials for solid and hematological tumors. However, depth and durability of responses to adoptive T cell therapy can suffer from an inhibitory tumor microenvironment. A common immune-suppressive agent is TGF-β, which is secreted by tumor cells and cells recruited to the tumor. We investigated whether human T cells could be engineered to be resistant to inhibition by TGF-β. Truncating the intracellular signaling domain from TGF-β receptor (TGFβR) II produces a dominant-negative receptor (dnTGFβRII) that dimerizes with endogenous TGFβRI to form a receptor that can bind TGF-β but cannot signal. We previously generated specific peptide enhanced affinity receptor TCRs recognizing the HLA-A*02-restricted peptides New York esophageal squamous cell carcinoma 1 (NY-ESO-1)157-165/l-Ag family member-1A (TCR: GSK3377794, formerly NY-ESO-1c259) and melanoma Ag gene A10254-262 (TCR: ADP-A2M10, formerly melanoma Ag gene A10c796). In this article, we show that exogenous TGF-β inhibited in vitro proliferation and effector functions of human T cells expressing these first-generation high-affinity TCRs, whereas inhibition was reduced or abolished in the case of second-generation TCRs coexpressed with dnTGFβRII (e.g., GSK3845097). TGF-β isoforms and a panel of TGF-β-associated genes are overexpressed in a range of cancer indications in which NY-ESO-1 is commonly expressed, particularly in synovial sarcoma. As an example, immunohistochemistry/RNAscope identified TGF-β-positive cells close to T cells in tumor nests and stroma, which had low frequencies of cells expressing IFN-γ in a non-small cell lung cancer setting. Coexpression of dnTGFβRII may therefore improve the efficacy of TCR-transduced T cells.
Veterinary Immunology and Immunopathology
Albarrak SM, Waters WR, Stabel JR, Hostetter JM.
PMID: - | DOI: 10.1016/j.vetimm.2018.05.003
In the present study, we evaluated expression of IFN-γ, IL-17, TNF-α, IL-10 and TGF-β by mucosal cells, including WC1+ γδ T cells, in ileal tissues taken from non-infected cattle and cattle naturally infected with Mycobacterium avium subsp paratuberculosis (MAP). Infected cattle were either in the subclinical or clinical stage of infection. We hypothesized that the cytokine profile of the WC1+ γδ T cell subset would be different between subclinical and clinical cattle. Our data indicate a significant increase in the numbers of WC1+ γδ T cells expressing IL-10 in clinical cattle compared to subclinical and non-infected cattle. We observed a significant increase in TGF-β expression by non-WC1+ cells in clinically infected cattle. Expression of IFN-γ, IL-17 and TNF-α in mucosal cells, including the WC1+ γδ T cell subset, was identified in all examined groups. However, our data indicate that the stage of infection did not significantly influence expression of these proinflammatory cytokines. This study demonstrates changes in the cytokine mRNA expression profile of mucosal cells in the ileum, and specifically WC1+ γδ T cells, as cattle progress to the clinical disease. The change is characterized by an increase in expression of anti-inflammatory cytokines.
Scola, L;Bongiorno, MR;Forte, GI;Aiello, A;Accardi, G;Scrimali, C;Spina, R;Lio, D;Candore, G;
PMID: 35886018 | DOI: 10.3390/genes13071235
Differential genetically determined expression of transforming growth factor-β (TGF-β pathway and of vascular endothelial growth factor-A (VEGF-A) might modulate the molecular "milieu" involved in the etio-pathogenesis of non-melanoma skin cancer (NMSC). We have evaluated the frequency of some functionally relevant SNPs of TGF-β and VEGF-A genes in 70 NMSC patients and 161 healthy controls, typed for TGF-β1 rs1800471, TGF-β2 rs900, TGF-βR1 rs334348 and rs334349, TGF-βR2 rs4522809 and VEGF-A rs3025039 SNPs. TGF-βR2 rs1800629G allele and related genotypes were found to be associated with a possible protective role against NMSC, whereas VEGF-A rs3025039T was associated with an increased risk. To evaluate the effect of genotype combinations on NMSC susceptibility, we determined the frequencies of 31 pseudo-haplotypes due to non-random linkage among alleles of loci not lying on the same chromosome. Two pseudo-haplotypes that imply a minor allele of TGF-βR2 or minor allele of VEGF-A SNPs combined with major alleles of the other SNPs were, respectively, associated with a protective effect, and susceptibility to NMSC. In addition, a pseudo-haplotype involving minor alleles of TGF-β2 rs900, TGF-βR1 rs334348 and rs4522809 SNPs might be a susceptibility marker for NMSC. In conclusion, our data suggest that a complex interplay among the genetic polymorphisms of TGF-β, TGF-β receptors and VEGF-A genes might influence the net effect of genetic background of the patients on NMSC development. This might be relevant in the risk evaluation, diagnosis and treatment of NMSC.
Liu, Y;Guerrero-Juarez, C;Xiao, F;Shettigar, N;Ramos, R;Kuan, C;Lin, Y;de Jesus Martinez Lomeli, L;Park, J;Oh, J;Liu, R;Lin, S;Tartaglia, M;Yang, R;Yu, Z;Nie, Q;Li, J;Plikus, M;
| DOI: 10.1016/j.devcel.2022.06.005
Hair follicle stem cells are regulated by dermal papilla fibroblasts, their principal signaling niche. Overactivation of Hedgehog signaling in the niche dramatically accelerates hair growth and induces follicle multiplication in mice. On single-cell RNA sequencing, dermal papilla fibroblasts increase heterogeneity to include new Wnt5ahigh states. Transcriptionally, mutant fibroblasts activate regulatory networks for Gli1, Alx3, Ebf1, Hoxc8, Sox18, and Zfp239. These networks jointly upregulate secreted factors for multiple hair morphogenesis and hair-growth-related pathways. Among these is non-conventional TGF-β ligand Scube3. We show that in normal mouse skin, Scube3 is expressed only in dermal papillae of growing, but not in resting follicles. SCUBE3 protein microinjection is sufficient to induce new hair growth, and pharmacological TGF-β inhibition rescues mutant hair hyper-activation phenotype. Moreover, dermal-papilla-enriched expression of SCUBE3 and its growth-activating effect are partially conserved in human scalp hair follicles. Thus, Hedgehog regulates mesenchymal niche function in the hair follicle via SCUBE3/TGF-β mechanism.
Snail enhances arginine synthesis by inhibiting ubiquitination-mediated degradation of ASS1
Jia, H;Yang, Y;Li, M;Chu, Y;Song, H;Zhang, J;Zhang, D;Zhang, Q;Xu, Y;Wang, J;Xu, H;Zou, X;Peng, H;Hou, Z;
PMID: 34184805 | DOI: 10.15252/embr.202051780
Snail is a dedicated transcriptional repressor and acts as a master inducer of EMT and metastasis, yet the underlying signaling cascades triggered by Snail still remain elusive. Here, we report that Snail promotes colorectal cancer (CRC) migration by preventing non-coding RNA LOC113230-mediated degradation of argininosuccinate synthase 1 (ASS1). LOC113230 is a novel Snail target gene, and Snail binds to the functional E-boxes within its proximal promoter to repress its expression in response to TGF-β induction. Ectopic expression of LOC113230 potently suppresses CRC cell growth, migration, and lung metastasis in xenograft experiments. Mechanistically, LOC113230 acts as a scaffold to facilitate recruiting LRPPRC and the TRAF2 E3 ubiquitin ligase to ASS1, resulting in enhanced ubiquitination and degradation of ASS1 and decreased arginine synthesis. Moreover, elevated ASS1 expression is essential for CRC growth and migration. Collectively, these findings suggest that TGF-β and Snail promote arginine synthesis via inhibiting LOC113230-mediated LRPPRC/TRAF2/ASS1 complex assembly and this complex can serve as potential target for the development of new therapeutic approaches to treat CRC.
Airway basal stem cells reutilize the embryonic proliferation regulator, Tgfβ-Id2 axis, for tissue regeneration
Kiyokawa, H;Yamaoka, A;Matsuoka, C;Tokuhara, T;Abe, T;Morimoto, M;
PMID: 34129836 | DOI: 10.1016/j.devcel.2021.05.016
During development, quiescent airway basal stem cells are derived from proliferative primordial progenitors through the cell-cycle slowdown. In contrast, basal cells contribute to adult tissue regeneration by shifting from slow cycling to proliferating and subsequently back to slow cycling. Although sustained proliferation results in tumorigenesis, the molecular mechanisms regulating these transitions remain unknown. Using temporal single-cell transcriptomics of developing murine airway progenitors and genetic validation experiments, we found that TGF-β signaling decelerated cell cycle by inhibiting Id2 and contributed to slow-cycling basal cell specification during development. In adult tissue regeneration, reduced TGF-β signaling restored Id2 expression and initiated regeneration. Id2 overexpression and Tgfbr2 knockout enhanced epithelial proliferation; however, persistent Id2 expression drove basal cell hyperplasia that resembled a precancerous state. Together, the TGF-β-Id2 axis commonly regulates the proliferation transitions in basal cells during development and regeneration, and its fine-tuning is critical for normal regeneration while avoiding basal cell hyperplasia.
Preliminary RNA-microarray analysis of long non-coding RNA expression in abnormally invasive placenta
Zhang, H;Wu, S;Ye, S;Ma, H;Liu, Z;
PMID: 33235622 | DOI: 10.3892/etm.2020.9445
Long non-coding RNAs (lncRNAs) are reported to have important roles in placental development and function, but the role of lncRNAs in abnormally invasive placenta (AIP) remains elusive. In the present study, the differential expression profiles of lncRNAs were analyzed to identify novel targets for further study of AIP. A total of 10 lncRNAs were chosen for validation by reverse transcription-quantitative PCR. To further determine the functions of dysregulated lncRNAs and their corresponding mRNAs, functional enrichment analysis, coexpression analysis were performed. A total of 329 lncRNAs and 179 mRNAs were identified to be differently expressed between the invasive and control group. Gene ontology analysis revealed that the 10 most significantly enriched functions included upregulated mRNAs and the most significantly enriched term was related to the proteinaceous extracellular matrix (ECM). In the pathway analysis, the two most significantly enriched pathways were the TGF-β signaling pathway for upregulated mRNAs and the pentose phosphate pathway for downregulated mRNAs. Furthermore, for certain dysregulated lncRNAs, their associated mRNAs were also dysregulated. Of note, BMP and activin membrane-bound inhibitor and TGF-β-induced, as the target genes of the TGF-β pathway, were indicated to be closely related to the ECM and invasive placental cells. Their nearby lncRNAs G008916 and vault RNA2-1 were also significantly dysregulated. In conclusion, significant lncRNAs with the potential to serve as biomarkers for AIP were identified.
Palmer MV, Thacker TC, Waters WR.
PMID: 27902779 | DOI: 10.1371/journal.pone.0167471
The hallmark lesion of tuberculosis in humans and animals is the granuloma. The granuloma represents a distinct host cellular immune response composed of epithelioid macrophages, lymphocytes, and multinucleated giant cells, often surrounding a caseous necrotic core. Within the granuloma, host-pathogen interactions determine disease outcome. Factors within the granulomas such as cytokines and chemokines drive cell recruitment, activity, function and ultimately the success or failure of the host's ability to control infection. Hence, an understanding of the granuloma-level cytokine response is necessary to understand tuberculosis pathogenesis. In-situ cytokine expression patterns were measured using a novel in situ hybridization assay, known as RNAScope® in granulomas of the lungs, tracheobronchial lymph nodes and caudal mediastinal lymph nodes of cattle experimentally infected with Mycobacterium bovis via aerosol exposure. In spite of microscopic morphological similarities, significant differences were seen between late stage granulomas of the lung compared to those of the tracheobronchial lymph nodes for IL-17A, IFN-γ, TGF-β, IL10 and IL-22 but not for TNF-α. Additionally, significant differences were noted between granulomas from two different thoracic lymph nodes that both receive afferent lymphatics from the lungs (i.e., tracheobronchial and caudal mediastinal lymph nodes) for TNF-α, IL-17A, IFN-γ, TGF-β and IL-10 but not for IL-22. These findings show that granuloma morphology alone is not a reliable indicator of granuloma function as granulomas of similar morphologies can have disparate cytokine expression patterns. Moreover, anatomically distinct lymph nodes (tracheobronchial vs caudal mediastinal) differ in cytokine expression patterns even when both receive afferent lymphatics from a lung containing tuberculoid granulomas. These findings show that selection of tissue and anatomic location are critical factors in assessing host immune response to M. bovis and should be considered carefully.
J Comp Pathol. 2015 Jul 16.
Palmer MV, Thacker TC, Waters WR.
PMID: 26189773 | DOI: 10.1016/j.jcpa.2015.06.004.
Mycobacterium bovis is the cause of tuberculosis in most animal species including cattle and is a serious zoonotic pathogen. In man, M. bovis infection can result in disease clinically indistinguishable from that caused by Mycobacterium tuberculosis, the cause of most human tuberculosis. Regardless of host, the typical lesion induced by M. bovis or M. tuberculosis is the tuberculoid granuloma. Tuberculoid granulomas are dynamic structures reflecting the interface between host and pathogen and, therefore, pass through various morphological stages (I to IV). Using a novel in-situ hybridization assay, transcription of various cytokine and chemokine genes was examined qualitatively and quantitatively using image analysis. In experimentally infected cattle, pulmonary granulomas of all stages were examined 150 days after aerosol exposure to M. bovis. Expression of mRNA encoding tumour necrosis factor (TNF)-α, transforming growth factor-β, interferon (IFN)-γ, interleukin (IL)-17A, IL-16, IL-10, CXCL9 and CXCL10 did not differ significantly between granulomas of different stages. However, relative expression of the various cytokines was characteristic of a Th1 response, with high TNF-α and IFN-γ expression and low IL-10 expression. Expression of IL-16 and the chemokines CXCL9 and CXCL10 was high, suggestive of granulomas actively involved in T-cell chemotaxis.
The Journal of clinical investigation
Lai, YJ;Tsai, FC;Chang, GJ;Chang, SH;Huang, CC;Chen, WJ;Yeh, YH;
PMID: 35775491 | DOI: 10.1172/JCI142548
Atrial fibrosis is an essential contributor to atrial fibrillation (AF). It remains unclear whether atrial endocardial endothelial cells (AEECs) that undergo endothelial-mesenchymal transition (EndMT) are among the sources of atrial fibroblasts. We studied human atria, TGF-β-treated human AEECs, cardiac-specific TGF-β-transgenic mice, and heart failure rabbits to identify the underlying mechanism of EndMT in atrial fibrosis. Using isolated AEECs, we found that miR-181b was induced in TGF-β-treated AEECs, which decreased semaphorin 3A (Sema3A) and increased EndMT markers, and these effects could be reversed by a miR-181b antagomir. Experiments in which Sema3A was increased by a peptide or decreased by a siRNA in AEECs revealed a mechanistic link between Sema3A and LIM-kinase 1/phosphorylated cofilin (LIMK/p-cofilin) signaling and suggested that Sema3A is upstream of LIMK in regulating actin remodeling through p-cofilin. Administration of the miR-181b antagomir or recombinant Sema3A to TGF-β-transgenic mice evoked increased Sema3A, reduced EndMT markers, and significantly decreased atrial fibrosis and AF vulnerability. Our study provides a mechanistic link between the induction of EndMT by TGF-β via miR-181b/Sema3A/LIMK/p-cofilin signaling to atrial fibrosis. Blocking miR-181b and increasing Sema3A are potential strategies for AF therapeutic intervention.
Veterinary Immunology and Immunopathology
Palmer MV , Thacker TC, Waters WR.
PMID: - | DOI: 10.1016/j.vetimm.2016.08.015
Regardless of host, pathogenic mycobacteria of the Mycobacterium tuberculosiscomplex such as Mycobacterium bovis, induce a characteristic lesion known as agranuloma, tubercle or tuberculoid granuloma. Granulomas represent a distinct host response to chronic antigenic stimuli, such as foreign bodies, certain bacterial components, or persistent pathogens such as M. bovis. Granulomas are composed of specific cell types including epithelioid macrophages, lymphocytes and a morphologically distinctive cell type, the multinucleated giant cell. Multinucleated giant cells are formed by the fusion of multiple macrophages; however, their function remains unclear. In humans, giant cells in tuberculous granulomas have been shown to express various cytokines, chemokines and enzymes important to the formation and maintenance of the granuloma. The objective of this study was to quantitatively assess multinucleated giant cell cytokine expression in bovine tuberculoid granulomas; focusing on cytokines of suspected relevance to bovine tuberculosis. Using calves experimentally infected with M. bovis, in situ cytokine expression was quantitatively assessed using RNAScope® for the following cytokines TNF-α, IFN-γ, TGF-β, IL-17A and IL-10. Multinucleated giant cells in bovine tuberculoid granulomas expressed all examined cytokines to varying degrees, with differential expression of TGF-β, IL-17A and IL-10 in giant cells from early versus late stage granulomas. There was a modest, positive correlation between the level of cytokine expression and cell size or number of nuclei. These results suggest that multinucleated giant cells are active participants within bovine tuberculoid granulomas, contributing to the cytokine milieu necessary to form and maintain granulomas.
Metastasis-associated macrophages constrain antitumor capability of natural killer cells in the metastatic site at least partially by membrane bound transforming growth factor β
Journal for immunotherapy of cancer
Brownlie, D;Doughty-Shenton, D;Yh Soong, D;Nixon, C;O Carragher, N;M Carlin, L;Kitamura, T;
PMID: 33472858 | DOI: 10.1136/jitc-2020-001740
Metastatic breast cancer is a leading cause of cancer-related death in women worldwide. Infusion of natural killer (NK) cells is an emerging immunotherapy for such malignant tumors, although elimination of the immunosuppressive tumor environment is required to improve its efficacy. The effects of this "metastatic" tumor environment on NK cells, however, remain largely unknown. Previous studies, including our own, have demonstrated that metastasis-associated macrophages (MAMs) are one of the most abundant immune cell types in the metastatic tumor niche in mouse models of metastatic breast cancer. We thus investigated the effects of MAMs on antitumor functions of NK cells in the metastatic tumor microenvironment. MAMs were isolated from the tumor-bearing lung of C57BL/6 mice intravenously injected with E0771-LG mouse mammary tumor cells. The effects of MAMs on NK cell cytotoxicity towards E0771-LG cells were evaluated in vitro by real-time fluorescence microscopy. The effects of MAM depletion on NK cell activation, maturation, and accumulation in the metastatic lung were evaluated by flow cytometry (CD69, CD11b, CD27) and in situ hybridization (Ncr1) using colony-stimulating factor 1 (CSF-1) receptor conditional knockout (Csf1r-cKO) mice. Finally, metastatic tumor loads in the chest region of mice were determined by bioluminescence imaging in order to evaluate the effect of MAM depletion on therapeutic efficacy of endogenous and adoptively transferred NK cells in suppressing metastatic tumor growth. MAMs isolated from the metastatic lung suppressed NK cell-induced tumor cell apoptosis in vitro via membrane-bound transforming growth factor β (TGF-β) dependent mechanisms. In the tumor-challenged mice, depletion of MAMs increased the percentage of activated (CD69+) and mature (CD11b+CD27-) NK cells and the number of Ncr1+ NK cells as well as NK cell-mediated tumor rejection in the metastatic site. Moreover, MAM depletion or TGF-β receptor antagonist treatment significantly enhanced the therapeutic efficacy of NK cell infusion in suppressing early metastatic tumor outgrowth. This study demonstrates that MAMs are a main negative regulator of NK cell function within the metastatic tumor niche, and MAM targeting is an attractive strategy to improve NK cell-based immunotherapy for metastatic breast cancer.
