Publications by authors named "Marina Botto"

123 Publications

Type I interferons affect the metabolic fitness of CD8 T cells from patients with systemic lupus erythematosus.

Nat Commun 2021 03 31;12(1):1980. Epub 2021 Mar 31.

Department of Immunology and Inflammation, Centre for Inflammatory Disease, Imperial College London, London, UK.

The majority of patients with systemic lupus erythematosus (SLE) have high expression of type I IFN-stimulated genes. Mitochondrial abnormalities have also been reported, but the contribution of type I IFN exposure to these changes is unknown. Here, we show downregulation of mitochondria-derived genes and mitochondria-associated metabolic pathways in IFN-High patients from transcriptomic analysis of CD4 and CD8 T cells. CD8 T cells from these patients have enlarged mitochondria and lower spare respiratory capacity associated with increased cell death upon rechallenge with TCR stimulation. These mitochondrial abnormalities can be phenocopied by exposing CD8 T cells from healthy volunteers to type I IFN and TCR stimulation. Mechanistically these 'SLE-like' conditions increase CD8 T cell NAD+ consumption resulting in impaired mitochondrial respiration and reduced cell viability, both of which can be rectified by NAD+ supplementation. Our data suggest that type I IFN exposure contributes to SLE pathogenesis by promoting CD8 T cell death via metabolic rewiring.
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http://dx.doi.org/10.1038/s41467-021-22312-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8012390PMC
March 2021

Longitudinal proteomic profiling of dialysis patients with COVID-19 reveals markers of severity and predictors of death.

Elife 2021 Mar 11;10. Epub 2021 Mar 11.

Department of Immunology and Inflammation, Imperial College London, London, United Kingdom.

End-stage kidney disease (ESKD) patients are at high risk of severe COVID-19. We measured 436 circulating proteins in serial blood samples from hospitalised and non-hospitalised ESKD patients with COVID-19 (n=256 samples from 55 patients). Comparison to 51 non-infected patients revealed 221 differentially expressed proteins, with consistent results in a separate subcohort of 46 COVID-19 patients. 203 proteins were associated with clinical severity, including IL6, markers of monocyte recruitment (e.g. CCL2, CCL7), neutrophil activation (e.g. proteinase-3) and epithelial injury (e.g. KRT19). Machine learning identified predictors of severity including IL18BP, CTSD, GDF15, and KRT19. Survival analysis with joint models revealed 69 predictors of death. Longitudinal modelling with linear mixed models uncovered 32 proteins displaying different temporal profiles in severe versus non-severe disease, including integrins and adhesion molecules. These data implicate epithelial damage, innate immune activation, and leucocyte-endothelial interactions in the pathology of severe COVID-19 and provide a resource for identifying drug targets.
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http://dx.doi.org/10.7554/eLife.64827DOI Listing
March 2021

PD-1 blockade improves Kupffer cell bacterial clearance in acute liver injury.

J Clin Invest 2021 Feb;131(4)

Department of Metabolism, Digestion and Reproduction, Section of Hepatology and Gastroenterology, and.

Patients with acute liver failure (ALF) have systemic innate immune suppression and increased susceptibility to infections. Programmed cell death 1 (PD-1) expression by macrophages has been associated with immune suppression during sepsis and cancer. We therefore examined the role of the programmed cell death 1/programmed death ligand 1 (PD-1/PD-L1) pathway in regulating Kupffer cell (KC) inflammatory and antimicrobial responses in acetaminophen-induced (APAP-induced) acute liver injury. Using intravital imaging and flow cytometry, we found impaired KC bacterial clearance and systemic bacterial dissemination in mice with liver injury. We detected increased PD-1 and PD-L1 expression in KCs and lymphocyte subsets, respectively, during injury resolution. Gene expression profiling of PD-1+ KCs revealed an immune-suppressive profile and reduced pathogen responses. Compared with WT mice, PD-1-deficient mice and anti-PD-1-treated mice with liver injury showed improved KC bacterial clearance, a reduced tissue bacterial load, and protection from sepsis. Blood samples from patients with ALF revealed enhanced PD-1 and PD-L1 expression by monocytes and lymphocytes, respectively, and that soluble PD-L1 plasma levels could predict outcomes and sepsis. PD-1 in vitro blockade restored monocyte functionality. Our study describes a role for the PD-1/PD-L1 axis in suppressing KC and monocyte antimicrobial responses after liver injury and identifies anti-PD-1 immunotherapy as a strategy to reduce infection susceptibility in ALF.
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http://dx.doi.org/10.1172/JCI140196DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7880414PMC
February 2021

Temporal changes in complement activation in haemodialysis patients with COVID-19 as a predictor of disease progression.

Clin Kidney J 2020 Oct 2;13(5):889-896. Epub 2020 Oct 2.

Centre for Inflammatory Disease, Imperial College London, London, UK.

Background: Complement activation may play a pathogenic role in patients with severe coronavirus disease 2019 (COVID-19) by contributing to tissue inflammation and microvascular thrombosis.

Methods: Serial samples were collected from patients receiving maintenance haemodialysis (HD). Thirty-nine patients had confirmed COVID-19 and 10 patients had no evidence of COVID-19. Plasma C5a and C3a levels were measured using enzyme-linked immunosorbent assay.

Results: We identified elevated levels of plasma C3a and C5a in HD patients with severe COVID-19 compared with controls. Serial sampling identified that C5a levels were elevated prior to clinical deterioration in patients who developed severe disease. C3a more closely mirrored both clinical and biochemical disease severity.

Conclusions: Our findings suggest that activation of complement plays a role in the pathogenesis of COVID-19, leading to endothelial injury and lung damage. C5a may be an earlier biomarker of disease severity than conventional parameters such as C-reactive protein and this warrants further investigation in dedicated biomarker studies. Our data support the testing of complement inhibition as a therapeutic strategy for patients with severe COVID-19.
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http://dx.doi.org/10.1093/ckj/sfaa192DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7577776PMC
October 2020

C3 Drives Inflammatory Skin Carcinogenesis Independently of C5.

J Invest Dermatol 2021 Feb 16;141(2):404-414.e6. Epub 2020 Jul 16.

Centre for Inflammatory Disease, Department of Immunology and Inflammation, Imperial College London, United Kingdom.

Nonmelanoma skin cancer such as cutaneous squamous cell carcinoma (cSCC) is the most common form of cancer and can occur as a consequence of DNA damage to the epithelium by UVR or chemical carcinogens. There is growing evidence that the complement system is involved in cancer immune surveillance; however, its role in cSCC remains unclear. Here, we show that complement genes are expressed in tissue from patients with cSCC, and C3 activation fragments are present in cSCC biopsies, indicating complement activation. Using a range of complement-deficient mice in a two-stage mouse model of chemically-induced cSCC, where a subclinical dose of 7,12-dimethylbenz[a]anthracene causes oncogenic mutations in epithelial cells and 12-O-tetradecanoylphorbol-13-acetate promotes the outgrowth of these cells, we found that C3-deficient mice displayed a significantly reduced tumor burden, whereas an opposite phenotype was observed in mice lacking C5aR1, C5aR2, and C3a receptor. In addition, in mice unable to form the membrane attack complex, the tumor progression was unaltered. C3 deficiency did not affect the cancer response to 7,12-dimethylbenz[a]anthracene treatment alone but reduced the epidermal hyperplasia during 12-O-tetradecanoylphorbol-13-acetate-induced inflammation. Collectively, these data indicate that C3 drives tumorigenesis during chronic skin inflammation, independently of the downstream generation of C5a or membrane attack complex.
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http://dx.doi.org/10.1016/j.jid.2020.06.025DOI Listing
February 2021

Th1 responses in vivo require cell-specific provision of OX40L dictated by environmental cues.

Nat Commun 2020 07 9;11(1):3421. Epub 2020 Jul 9.

Institute of Immunology and Immunotherapy, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK.

The OX40-OX40L pathway provides crucial co-stimulatory signals for CD4 T cell responses, however the precise cellular interactions critical for OX40L provision in vivo and when these occur, remains unclear. Here, we demonstrate that provision of OX40L by dendritic cells (DCs), but not T cells, B cells nor group 3 innate lymphoid cells (ILC3s), is critical specifically for the effector Th1 response to an acute systemic infection with Listeria monocytogenes (Lm). OX40L expression by DCs is regulated by cross-talk with NK cells, with IFNγ signalling to the DC to enhance OX40L in a mechanism conserved in both mouse and human DCs. Strikingly, DC expression of OX40L is redundant in a chronic intestinal Th1 response and expression by ILC3s is necessary. Collectively these data reveal tissue specific compartmentalisation of the cellular provision of OX40L and define a mechanism controlling DC expression of OX40L in vivo.
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http://dx.doi.org/10.1038/s41467-020-17293-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7347572PMC
July 2020

Complement alone drives efficacy of a chimeric antigonococcal monoclonal antibody.

PLoS Biol 2019 06 19;17(6):e3000323. Epub 2019 Jun 19.

Division of Infectious Diseases and Immunology, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America.

Multidrug-resistant Neisseria gonorrhoeae is a global health problem. Monoclonal antibody (mAb) 2C7 recognizes a gonococcal lipooligosaccharide epitope that is expressed by >95% of clinical isolates and hastens gonococcal vaginal clearance in mice. Chimeric mAb 2C7 (human immunoglobulin G1 [IgG1]) with an E430G Fc modification that enhances Fc:Fc interactions and hexamerization following surface-target binding and increases complement activation (HexaBody technology) showed significantly greater C1q engagement and C4 and C3 deposition compared to mAb 2C7 with wild-type Fc. Greater complement activation by 2C7-E430G Fc translated to increased bactericidal activity in vitro and, consequently, enhanced efficacy in mice, compared with "Fc-unmodified" chimeric 2C7. Gonococci bind the complement inhibitors factor H (FH) and C4b-binding protein (C4BP) in a human-specific manner, which dampens antibody (Ab)-mediated complement-dependent killing. The variant 2C7-E430G Fc overcame the barrier posed by these inhibitors in human FH/C4BP transgenic mice, for which a single 1 μg intravenous dose cleared established infection. Chlamydia frequently coexists with and exacerbates gonorrhea; 2C7-E430G Fc also proved effective against gonorrhea in gonorrhea/chlamydia-coinfected mice. Complement activation alone was necessary and sufficient for 2C7 function, evidenced by the fact that (1) "complement-inactive" Fc modifications that engaged Fc gamma receptor (FcγR) rendered 2C7 ineffective, nonetheless; (2) 2C7 was nonfunctional in C1q-/- mice, when C5 function was blocked, or in C9-/- mice; and (3) 2C7 remained effective in neutrophil-depleted mice and in mice treated with PMX205, a C5a receptor (C5aR1) inhibitor. We highlight the importance of complement activation for antigonococcal Ab function in the genital tract. Elucidating the correlates of protection against gonorrhea will inform the development of Ab-based gonococcal vaccines and immunotherapeutics.
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http://dx.doi.org/10.1371/journal.pbio.3000323DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6602280PMC
June 2019

Tumor Cells Hijack Macrophage-Produced Complement C1q to Promote Tumor Growth.

Cancer Immunol Res 2019 07 4;7(7):1091-1105. Epub 2019 Jun 4.

Sorbonne Paris Cite, Cordeliers Research Center, University Paris Descartes Paris 5, Paris, France.

Clear-cell renal cell carcinoma (ccRCC) possesses an unmet medical need, particularly at the metastatic stage, when surgery is ineffective. Complement is a key factor in tissue inflammation, favoring cancer progression through the production of complement component 5a (C5a). However, the activation pathways that generate C5a in tumors remain obscure. By data mining, we identified ccRCC as a cancer type expressing concomitantly high expression of the components that are part of the classical complement pathway. To understand how the complement cascade is activated in ccRCC and impacts patients' clinical outcome, primary tumors from three patient cohorts ( = 106, 154, and 43), ccRCC cell lines, and tumor models in complement-deficient mice were used. High densities of cells producing classical complement pathway components C1q and C4 and the presence of C4 activation fragment deposits in primary tumors correlated with poor prognosis. The orchestrated production of C1q by tumor-associated macrophages (TAM) and C1r, C1s, C4, and C3 by tumor cells associated with IgG deposits, led to C1 complex assembly, and complement activation. Accordingly, mice deficient in C1q, C4, or C3 displayed decreased tumor growth. However, the ccRCC tumors infiltrated with high densities of C1q-producing TAMs exhibited an immunosuppressed microenvironment, characterized by high expression of immune checkpoints (i.e., PD-1, Lag-3, PD-L1, and PD-L2). Our data have identified the classical complement pathway as a key inflammatory mechanism activated by the cooperation between tumor cells and TAMs, favoring cancer progression, and highlight potential therapeutic targets to restore an efficient immune reaction to cancer.
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http://dx.doi.org/10.1158/2326-6066.CIR-18-0891DOI Listing
July 2019

Human Factor H Domains 6 and 7 Fused to IgG1 Fc Are Immunotherapeutic against .

J Immunol 2018 11 28;201(9):2700-2709. Epub 2018 Sep 28.

Division of Infectious Diseases and Immunology, Department of Medicine, University of Massachusetts Medical School, Worcester, MA 01605;

Novel therapeutics against multidrug-resistant are urgently needed. Gonococcal lipooligosaccharide often expresses lacto--neotetraose (LNnT), which becomes sialylated in vivo, enhancing factor H (FH) binding and contributing to the organism's ability to resist killing by complement. We previously showed that FH domains 18-20 (with a D-to-G mutation at position 1119 in domain 19) fused to Fc (FHD1119G/Fc) displayed complement-dependent bactericidal activity in vitro and attenuated gonococcal vaginal colonization of mice. Gonococcal lipooligosaccharide phase variation can result in loss of LNnT expression. Loss of sialylated LNnT, although associated with a considerable fitness cost, could decrease efficacy of FHD1119G/Fc. Similar to , gonococci also bind FH domains 6 and 7 through Neisserial surface protein A (NspA). In this study, we show that a fusion protein comprising FH domains 6 and 7 fused to human IgG1 Fc (FH6,7/Fc) bound to 15 wild-type antimicrobial resistant isolates of and to each of six gonococcal deletion mutants. FH6,7/Fc mediated complement-dependent killing of 8 of the 15 wild-type gonococcal isolates and effectively reduced the duration and burden of vaginal colonization of three gonococcal strains tested in wild-type mice, including two strains that resisted complement-dependent killing but on which FH6,7/Fc enhanced C3 deposition. FH/Fc lost efficacy when Fc was mutated to abrogate C1q binding and in mice, highlighting the requirement of the classical pathway for its activity. Targeting gonococci with FH6,7/Fc provides an additional immunotherapeutic approach against multidrug-resistant gonorrhea.
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http://dx.doi.org/10.4049/jimmunol.1701666DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6200640PMC
November 2018

Epithelial damage and tissue γδ T cells promote a unique tumor-protective IgE response.

Nat Immunol 2018 08 16;19(8):859-870. Epub 2018 Jul 16.

Department of Medicine, Imperial College London, London, UK.

IgE is an ancient and conserved immunoglobulin isotype with potent immunological function. Nevertheless, the regulation of IgE responses remains an enigma, and evidence of a role for IgE in host defense is limited. Here we report that topical exposure to a common environmental DNA-damaging xenobiotic initiated stress surveillance by γδTCR intraepithelial lymphocytes that resulted in class switching to IgE in B cells and the accumulation of autoreactive IgE. High-throughput antibody sequencing revealed that γδ T cells shaped the IgE repertoire by supporting specific variable-diversity-joining (VDJ) rearrangements with unique characteristics of the complementarity-determining region CDRH3. This endogenous IgE response, via the IgE receptor FcεRI, provided protection against epithelial carcinogenesis, and expression of the gene encoding FcεRI in human squamous-cell carcinoma correlated with good disease prognosis. These data indicate a joint role for immunosurveillance by T cells and by B cells in epithelial tissues and suggest that IgE is part of the host defense against epithelial damage and tumor development.
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http://dx.doi.org/10.1038/s41590-018-0161-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6071860PMC
August 2018

CD93 regulates central nervous system inflammation in two mouse models of autoimmune encephalomyelitis.

Immunology 2018 11 11;155(3):346-355. Epub 2018 Jul 11.

BIIG, Brain Inflammation and Immunity Group, Cardiff University School of Medicine, Cardiff, UK.

Microglia and non-professional immune cells (endothelial cells, neurons) participate in the recognition and removal of pathogens and tissue debris in the injured central nervous system through major pro-inflammatory processes. However, the mechanisms involved in regulating these responses remain ill-characterized. We herein show that CD93, also known as complement C1qRp/AA4 stem cell marker, has an important role in the regulation of inflammatory processes. The role of CD93 was evaluated in two models of neuroinflammation. We used the MOG-experimental autoimmune encephalomyelitis (EAE) model and the antibody-dependent EAE (ADEAE), which were induced in wild-type and CD93 knockout mice. We found that CD93 was highly expressed by neurons, endothelial cells and microglia (ramified > amoeboid). Astrocytes and oligodendrocytes did not to express CD93. We further observed that CD93-deficient (CD93 ) mice presented a more robust brain and spinal cord inflammation in EAE and ADEAE. Encephalitis in CD93 was characterized by increased numbers of infiltrating M1 macrophages (CD11c CD206 ) and amoeboid microglia exhibiting a more activated phenotype (Tomato Lectin Cox2 ). Damage to and leakage through the blood-brain barrier was increased in CD93 animals and was associated with a more robust neuronal injury when compared with wild-type EAE mice. We propose that CD93 is an important neuro-immune regulator to control central nervous system inflammation.
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http://dx.doi.org/10.1111/imm.12974DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6187214PMC
November 2018

Tissue-Restricted Adaptive Type 2 Immunity Is Orchestrated by Expression of the Costimulatory Molecule OX40L on Group 2 Innate Lymphoid Cells.

Immunity 2018 06 12;48(6):1195-1207.e6. Epub 2018 Jun 12.

MRC Laboratory of Molecular Biology, Cambridge CB2 0QH, UK. Electronic address:

The local regulation of type 2 immunity relies on dialog between the epithelium and the innate and adaptive immune cells. Here we found that alarmin-induced expression of the co-stimulatory molecule OX40L on group 2 innate lymphoid cells (ILC2s) provided tissue-restricted T cell co-stimulation that was indispensable for Th2 and regulatory T (Treg) cell responses in the lung and adipose tissue. Interleukin (IL)-33 administration resulted in organ-specific surface expression of OX40L on ILC2s and the concomitant expansion of Th2 and Treg cells, which was abolished upon deletion of OX40L on ILC2s (Il7raTnfsf4 mice). Moreover, Il7raTnfsf4 mice failed to mount effective Th2 and Treg cell responses and corresponding adaptive type 2 pulmonary inflammation arising from Nippostrongylus brasiliensis infection or allergen exposure. Thus, the increased expression of OX40L in response to IL-33 acts as a licensing signal in the orchestration of tissue-specific adaptive type 2 immunity, without which this response fails to establish.
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http://dx.doi.org/10.1016/j.immuni.2018.05.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6015114PMC
June 2018

C1q restrains autoimmunity and viral infection by regulating CD8 T cell metabolism.

Science 2018 05;360(6388):558-563

Faculty of Medicine, Imperial College London, London W12 ONN, UK.

Deficiency of C1q, the initiator of the complement classical pathway, is associated with the development of systemic lupus erythematosus (SLE). Explaining this association in terms of abnormalities in the classical pathway alone remains problematic because C3 deficiency does not predispose to SLE. Here, using a mouse model of SLE, we demonstrate that C1q, but not C3, restrains the response to self-antigens by modulating the mitochondrial metabolism of CD8 T cells, which can themselves propagate autoimmunity. C1q deficiency also triggers an exuberant effector CD8 T cell response to chronic viral infection leading to lethal immunopathology. These data establish a link between C1q and CD8 T cell metabolism and may explain how C1q protects against lupus, with implications for the role of viral infections in the perpetuation of autoimmunity.
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http://dx.doi.org/10.1126/science.aao4555DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6545171PMC
May 2018

Hyposialylated IgG activates endothelial IgG receptor FcγRIIB to promote obesity-induced insulin resistance.

J Clin Invest 2018 01 27;128(1):309-322. Epub 2017 Nov 27.

Center for Pulmonary and Vascular Biology, Department of Pediatrics, University of Texas Southwestern Medical Center, Dallas, Texas, USA.

Type 2 diabetes mellitus (T2DM) is a common complication of obesity. Here, we have shown that activation of the IgG receptor FcγRIIB in endothelium by hyposialylated IgG plays an important role in obesity-induced insulin resistance. Despite becoming obese on a high-fat diet (HFD), mice lacking FcγRIIB globally or selectively in endothelium were protected from insulin resistance as a result of the preservation of insulin delivery to skeletal muscle and resulting maintenance of muscle glucose disposal. IgG transfer in IgG-deficient mice implicated IgG as the pathogenetic ligand for endothelial FcγRIIB in obesity-induced insulin resistance. Moreover, IgG transferred from patients with T2DM but not from metabolically healthy subjects caused insulin resistance in IgG-deficient mice via FcγRIIB, indicating that similar processes may be operative in T2DM in humans. Mechanistically, the activation of FcγRIIB by IgG from obese mice impaired endothelial cell insulin transcytosis in culture and in vivo. These effects were attributed to hyposialylation of the Fc glycan, and IgG from T2DM patients was also hyposialylated. In HFD-fed mice, supplementation with the sialic acid precursor N-acetyl-D-mannosamine restored IgG sialylation and preserved insulin sensitivity without affecting weight gain. Thus, IgG sialylation and endothelial FcγRIIB may represent promising therapeutic targets to sever the link between obesity and T2DM.
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http://dx.doi.org/10.1172/JCI89333DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5749535PMC
January 2018

B cell OX40L supports T follicular helper cell development and contributes to SLE pathogenesis.

Ann Rheum Dis 2017 Dec 17;76(12):2095-2103. Epub 2017 Aug 17.

Division of Medical and Molecular Genetics and Immunology, Infection and Inflammatory Disease, King's College London, London, UK.

Objectives: (encodes OX40L) is a susceptibility locus for systemic lupus erythematosus (SLE). Risk alleles increase expression in cell lines, but the mechanism linking this effect to disease is unclear, and the OX40L-expressing cell types mediating the risk are not clearly established. Blockade of OX40L has been demonstrated to reduce disease severity in several models of autoimmunity, but not in SLE. We sought to investigate its potential therapeutic role in lupus.

Methods: We used a conditional knockout mouse system to investigate the function of OX40L on B and T lymphocytes in systemic autoimmunity.

Results: Physiologically, OX40L on both B and T cells contributed to the humoral immune response, but B cell OX40L supported the secondary humoral response and antibody affinity maturation. Our data also indicated that loss of B cell OX40L impeded the generation of splenic T follicular helper cells. We further show that in two models of SLE-a spontaneous congenic model and the H2-IA graft-versus-host-induced model-loss of B cell OX40L ameliorates the autoimmune phenotype. This improvement was, in each case, accompanied by a decline in T follicular helper cell numbers. Importantly, the germline knockout did not exhibit a markedly different phenotype from the B cell knockout in these models.

Conclusions: These findings contribute to a model in which genetically determined increased OX40L expression promotes human SLE by several mechanisms, contingent on its cellular expression. The improvement in pathology in two models of systemic autoimmunity indicates that OX40L is an excellent therapeutic target in SLE.
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http://dx.doi.org/10.1136/annrheumdis-2017-211499DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5705841PMC
December 2017

Multi-functional mechanisms of immune evasion by the streptococcal complement inhibitor C5a peptidase.

PLoS Pathog 2017 Aug 14;13(8):e1006493. Epub 2017 Aug 14.

Faculty of Medicine, Imperial College London, London, United Kingdom.

The complement cascade is crucial for clearance and control of invading pathogens, and as such is a key target for pathogen mediated host modulation. C3 is the central molecule of the complement cascade, and plays a vital role in opsonization of bacteria and recruitment of neutrophils to the site of infection. Streptococcal species have evolved multiple mechanisms to disrupt complement-mediated innate immunity, among which ScpA (C5a peptidase), a C5a inactivating enzyme, is widely conserved. Here we demonstrate for the first time that pyogenic streptococcal species are capable of cleaving C3, and identify C3 and C3a as novel substrates for the streptococcal ScpA, which are functionally inactivated as a result of cleavage 7 amino acids upstream of the natural C3 convertase. Cleavage of C3a by ScpA resulted in disruption of human neutrophil activation, phagocytosis and chemotaxis, while cleavage of C3 generated abnormally-sized C3a and C3b moieties with impaired function, in particular reducing C3 deposition on the bacterial surface. Despite clear effects on human complement, expression of ScpA reduced clearance of group A streptococci in vivo in wildtype and C5 deficient mice, and promoted systemic bacterial dissemination in mice that lacked both C3 and C5, suggesting an additional complement-independent role for ScpA in streptococcal pathogenesis. ScpA was shown to mediate streptococcal adhesion to both human epithelial and endothelial cells, consistent with a role in promoting bacterial invasion within the host. Taken together, these data show that ScpA is a multi-functional virulence factor with both complement-dependent and independent roles in streptococcal pathogenesis.
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http://dx.doi.org/10.1371/journal.ppat.1006493DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5555575PMC
August 2017

Complement C1q is hydroxylated by collagen prolyl 4 hydroxylase and is sensitive to off-target inhibition by prolyl hydroxylase domain inhibitors that stabilize hypoxia-inducible factor.

Kidney Int 2017 10 12;92(4):900-908. Epub 2017 May 12.

School of Clinical Medicine, Cambridge Institute for Medical Research, Cambridge Biomedical Campus, University of Cambridge, Cambridge, UK. Electronic address:

Complement C1q is part of the C1 macromolecular complex that mediates the classical complement activation pathway: a major arm of innate immune defense. C1q is composed of A, B, and C chains that require post-translational prolyl 4-hydroxylation of their N-terminal collagen-like domain to enable the formation of the functional triple helical multimers. The prolyl 4-hydroxylase(s) that hydroxylate C1q have not previously been identified. Recognized prolyl 4-hydroxylases include collagen prolyl-4-hydroxylases (CP4H) and the more recently described prolyl hydroxylase domain (PHD) enzymes that act as oxygen sensors regulating hypoxia-inducible factor (HIF). We show that several small-molecule prolyl hydroxylase inhibitors that activate HIF also potently suppress C1q secretion by human macrophages. However, reducing oxygenation to a level that activates HIF does not compromise C1q hydroxylation. In vitro studies showed that a C1q A chain peptide is not a substrate for PHD2 but is a substrate for CP4H1. Circulating levels of C1q did not differ between wild-type mice or mice with genetic deficits in PHD enzymes, but were reduced by prolyl hydroxylase inhibitors. Thus, C1q is hydroxylated by CP4H, but not the structurally related PHD hydroxylases. Hence, reduction of C1q levels may be an important off-target side effect of small molecule PHD inhibitors developed as treatments for renal anemia.
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http://dx.doi.org/10.1016/j.kint.2017.03.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5612014PMC
October 2017

Altered expression of signalling lymphocyte activation molecule receptors in T-cells from lupus nephritis patients-a potential biomarker of disease activity.

Rheumatology (Oxford) 2017 07;56(7):1206-1216

Department of Medicine, Imperial College London, Centre for Complement and Inflammation Research.

Objectives: The aim was to investigate whether the signalling lymphocyte activation molecule (SLAM) signalling pathways contribute to LN and whether SLAM receptors could be valuable biomarkers of disease activity.

Methods: Peripheral blood mononuclear cells from 30National Research Ethics Service SLE patients with biopsy-proven LN were analysed by flow cytometry. Clinical measures of disease activity were assessed. The expression of the SLAM family receptors on T-cell subpopulations [CD4, CD8 and double negative (DN) T cells] was measured and compared between lupus patients with active renal disease and those in remission.

Results: The frequency of CD8 T cells expressing SLAMF3, SLAMF5 and SLAMF7 was significantly lower in LN patients who were in remission. In contrast, these subsets were similar in patients with active renal disease and in healthy individuals. Patients with active nephritis had an increased percentage of circulating monocytes, consistent with a potential role played by these cells in glomerular inflammation. Changes in the frequency of DN T cells positive for SLAMF2, SLAMF4 and SLAMF7 were observed in lupus patients irrespective of the disease activity. We detected alterations in the cellular expression of the SLAM family receptors, but these changes were less obvious and did not reveal any specific pattern. The percentage of DN T cells expressing SLAMF6 could predict the clinical response to B-cell depletion in patients with LN.

Conclusion: Our study demonstrates altered expression of the SLAM family receptors in SLE T lymphocytes. This is consistent with the importance of the SLAM-associated pathways in lupus pathogenesis.
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http://dx.doi.org/10.1093/rheumatology/kex078DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5850773PMC
July 2017

Complement C3 Exacerbates Imiquimod-Induced Skin Inflammation and Psoriasiform Dermatitis.

J Invest Dermatol 2017 03 19;137(3):760-763. Epub 2016 Nov 19.

Centre for Complement and Inflammation Research, Department of Medicine, Imperial College London, London, UK. Electronic address:

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http://dx.doi.org/10.1016/j.jid.2016.11.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5319416PMC
March 2017

Effect of irradiation/bone marrow transplantation on alveolar epithelial type II cells is aggravated in surfactant protein D deficient mice.

Histochem Cell Biol 2017 Jan 26;147(1):49-61. Epub 2016 Aug 26.

Child Health, Clinical and Experimental Sciences, Faculty of Medicine, University of Southampton, Southampton General Hospital, Southampton, SO16 6YD, UK.

Irradiation followed by bone marrow transplantation (BM-Tx) is a frequent therapeutic intervention causing pathology to the lung. Although alveolar epithelial type II (AE2) cells are essential for lung function and are damaged by irradiation, the long-term consequences of irradiation and BM-Tx are not well characterized. In addition, it is unknown whether surfactant protein D (SP-D) influences the response of AE2 cells to the injurious events. Therefore, wildtype (WT) and SP-D mice were subjected to a myeloablative whole body irradiation dose of 8 Gy and subsequent BM-Tx and compared with age- and sex-matched untreated controls. AE2 cell changes were investigated quantitatively by design-based stereology. Compared with WT, untreated SP-D mice showed a higher number of larger sized AE2 cells and a greater amount of surfactant-storing lamellar bodies. Irradiation and BM-Tx induced hyperplasia and hypertrophy in WT and SP-D mice as well as the formation of giant lamellar bodies. The experimentally induced alterations were more severe in the SP-D than in the WT mice, particularly with respect to the surfactant-storing lamellar bodies which were sometimes extremely enlarged in SP-D mice. In conclusion, irradiation and BM-Tx have profound long-term effects on AE2 cells and their lamellar bodies. These data may explain some of the clinical pulmonary consequences of this procedure. The data should also be taken into account when BM-Tx is used as an experimental procedure to investigate the impact of bone marrow-derived cells for the phenotype of a specific genotype in the mouse.
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http://dx.doi.org/10.1007/s00418-016-1479-7DOI Listing
January 2017

Bacillus anthracis Spore Surface Protein BclA Mediates Complement Factor H Binding to Spores and Promotes Spore Persistence.

PLoS Pathog 2016 06 15;12(6):e1005678. Epub 2016 Jun 15.

Center for Infectious and Inflammatory Diseases, Institute of Biosciences and Technology, Texas A&M Health Science Center, Houston, Texas, United States of America.

Spores of Bacillus anthracis, the causative agent of anthrax, are known to persist in the host lungs for prolonged periods of time, however the underlying mechanism is poorly understood. In this study, we demonstrated that BclA, a major surface protein of B. anthracis spores, mediated direct binding of complement factor H (CFH) to spores. The surface bound CFH retained its regulatory cofactor activity resulting in C3 degradation and inhibition of downstream complement activation. By comparing results from wild type C57BL/6 mice and complement deficient mice, we further showed that BclA significantly contributed to spore persistence in the mouse lungs and dampened antibody responses to spores in a complement C3-dependent manner. In addition, prior exposure to BclA deletion spores (ΔbclA) provided significant protection against lethal challenges by B. anthracis, whereas the isogenic parent spores did not, indicating that BclA may also impair protective immunity. These results describe for the first time an immune inhibition mechanism of B. anthracis mediated by BclA and CFH that promotes spore persistence in vivo. The findings also suggested an important role of complement in persistent infections and thus have broad implications.
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http://dx.doi.org/10.1371/journal.ppat.1005678DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4909234PMC
June 2016

Complement receptor 3 mediates renal protection in experimental C3 glomerulopathy.

Kidney Int 2016 Apr 3;89(4):823-32. Epub 2016 Feb 3.

Centre for Complement and Inflammation Research, Imperial College, London, UK. Electronic address:

C3 glomerulopathy is a complement-mediated renal disease that is frequently associated with abnormalities in regulation of the complement alternative pathway. Mice with deficiency of factor H (Cfh(-/-)), a negative alternative pathway regulator, are an established experimental model of C3 glomerulopathy in which complement C3 fragments including iC3b accumulate along the glomerular basement membrane. Here we show that deficiency of complement receptor 3 (CR3), the main receptor for iC3b, enhances the severity of spontaneous renal disease in Cfh(-/-) mice. This effect was found to be dependent on CR3 expression on bone marrow-derived cells. CR3 also mediated renal protection outside the setting of factor H deficiency, as shown by the development of enhanced renal injury in CR3-deficient mice during accelerated nephrotoxic nephritis. The iC3b-CR3 interaction downregulated the proinflammatory cytokine response of both murine and human macrophages to lipopolysaccharide stimulation in vitro, suggesting that the protective effect of CR3 on glomerular injury was mediated via modulation of macrophage-derived proinflammatory cytokines. Thus, CR3 has a protective role in glomerulonephritis and suggests that pharmacologic potentiation of the macrophage CR3 interaction with iC3b could be therapeutically beneficial.
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http://dx.doi.org/10.1016/j.kint.2015.11.024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4869622PMC
April 2016

C1q acts in the tumour microenvironment as a cancer-promoting factor independently of complement activation.

Nat Commun 2016 Feb 1;7:10346. Epub 2016 Feb 1.

Istituto di Ricovero e Cura a Carattere Scientifico, Istituto Auxologico Italiano, Milan 20145, Italy.

Complement C1q is the activator of the classical pathway. However, it is now recognized that C1q can exert functions unrelated to complement activation. Here we show that C1q, but not C4, is expressed in the stroma and vascular endothelium of several human malignant tumours. Compared with wild-type (WT) or C3- or C5-deficient mice, C1q-deficient (C1qa(-/-)) mice bearing a syngeneic B16 melanoma exhibit a slower tumour growth and prolonged survival. This effect is not attributable to differences in the tumour-infiltrating immune cells. Tumours developing in WT mice display early deposition of C1q, higher vascular density and an increase in the number of lung metastases compared with C1qa(-/-) mice. Bone marrow (BM) chimeras between C1qa(-/-) and WT mice identify non-BM-derived cells as the main local source of C1q that can promote cancer cell adhesion, migration and proliferation. Together these findings support a role for locally synthesized C1q in promoting tumour growth.
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http://dx.doi.org/10.1038/ncomms10346DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4740357PMC
February 2016

C1q Modulates the Response to TLR7 Stimulation by Pristane-Primed Macrophages: Implications for Pristane-Induced Lupus.

J Immunol 2016 Feb 15;196(4):1488-94. Epub 2016 Jan 15.

Centre for Complement and Inflammation Research, Division of Immunology and Inflammation, Department of Medicine, Imperial College London, London W12 0NN, United Kingdom;

The complement component C1q is known to play a controversial role in the pathogenesis of systemic lupus erythematosus, but the underlying mechanisms remain poorly understood. Intraperitoneal injection of pristane induces a lupus-like syndrome whose pathogenesis implicates the secretion of type I IFN by CD11b(+) Ly6C(high) inflammatory monocytes in a TLR7-dependent fashion. C1q was also shown to influence the secretion of IFN-α. In this study, we explored whether C1q deficiency could affect pristane-induced lupus. Surprisingly, C1qa(-/-) mice developed lower titers of circulating Abs and milder arthritis compared with the controls. In keeping with the clinical scores, 2 wk after pristane injection the peritoneal recruitment of CD11b(+) Ly6C(high) inflammatory monocytes in C1qa(-/-) mice was impaired. Furthermore, C1q-deficient pristane-primed resident peritoneal macrophages secreted significantly less CCL3, CCL2, CXCL1, and IL-6 when stimulated in vitro with TLR7 ligand. Replenishing C1q in vivo during the pristane-priming phase rectified this defect. Conversely, pristane-primed macrophages from C3-deficient mice did not show impaired cytokine production. These findings demonstrate that C1q deficiency impairs the TLR7-dependent chemokine production by pristane-primed peritoneal macrophages and suggest that C1q, and not C3, is involved in the handling of pristane by phagocytic cells, which is required to trigger disease in this model.
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http://dx.doi.org/10.4049/jimmunol.1401009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4745139PMC
February 2016

Partial Complement Factor H Deficiency Associates with C3 Glomerulopathy and Thrombotic Microangiopathy.

J Am Soc Nephrol 2016 05 15;27(5):1334-42. Epub 2015 Sep 15.

Centre for Complement and Inflammation Research, Imperial College, London, United Kingdom

The complement-mediated renal diseases C3 glomerulopathy (C3G) and atypical hemolytic uremic syndrome (aHUS) strongly associate with inherited and acquired abnormalities in the regulation of the complement alternative pathway (AP). The major negative regulator of the AP is the plasma protein complement factor H (FH). Abnormalities in FH result in uncontrolled activation of C3 through the AP and associate with susceptibility to both C3G and aHUS. Although previously developed FH-deficient animal models have provided important insights into the mechanisms underlying susceptibility to these unique phenotypes, these models do not entirely reproduce the clinical observations. FH is predominantly synthesized in the liver. We generated mice with hepatocyte-specific FH deficiency and showed that these animals have reduced plasma FH levels with secondary reduction in plasma C3. Unlike mice with complete FH deficiency, hepatocyte-specific FH-deficient animals developed neither plasma C5 depletion nor accumulation of C3 along the glomerular basement membrane. In contrast, subtotal FH deficiency associated with mesangial C3 accumulation consistent with C3G. Although there was no evidence of spontaneous thrombotic microangiopathy, the hepatocyte-specific FH-deficient animals developed severe C5-dependent thrombotic microangiopathy after induction of complement activation within the kidney by accelerated serum nephrotoxic nephritis. Taken together, our data indicate that subtotal FH deficiency can give rise to either spontaneous C3G or aHUS after a complement-activating trigger within the kidney and that the latter is C5 dependent.
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http://dx.doi.org/10.1681/ASN.2015030295DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4849824PMC
May 2016

Triglyceride-Rich Lipoproteins Modulate the Distribution and Extravasation of Ly6C/Gr1(low) Monocytes.

Cell Rep 2015 Sep 3;12(11):1802-15. Epub 2015 Sep 3.

Centre for Complement and Inflammation Research, Division of Immunology and Inflammation, Department of Medicine, Imperial College London, Du Cane Road, London W12 ONN, UK. Electronic address:

Monocytes are heterogeneous effector cells involved in the maintenance and restoration of tissue integrity. However, their response to hyperlipidemia remains poorly understood. Here, we report that in the presence of elevated levels of triglyceride-rich lipoproteins, induced by administration of poloxamer 407, the blood numbers of non-classical Ly6C/Gr1(low) monocytes drop, while the number of bone marrow progenitors remains similar. We observed an increased crawling and retention of the Gr1(low) monocytes at the endothelial interface and a marked accumulation of CD68(+) macrophages in several organs. Hypertriglyceridemia was accompanied by an increased expression of tissue, and plasma CCL4 and blood Gr1(low) monocyte depletion involved a pertussis-toxin-sensitive receptor axis. Collectively, these findings demonstrate that a triglyceride-rich environment can alter blood monocyte distribution, promoting the extravasation of Gr1(low) cells. The behavior of these cells in response to dyslipidemia highlights the significant impact that high levels of triglyceride-rich lipoproteins may have on innate immune cells.
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http://dx.doi.org/10.1016/j.celrep.2015.08.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4590546PMC
September 2015

The paradoxical roles of C1q and C3 in autoimmunity.

Immunobiology 2016 06 11;221(6):719-25. Epub 2015 May 11.

Centre for Complement and Inflammation Research, Department of Medicine, Imperial College London, London, UK. Electronic address:

In this review we will focus on the links between complement and autoimmune diseases and will highlight how animal models have provided insights into the manner by which C1q and C3 act to modulate both adaptive and innate immune responses. In particular we will highlight how C1q may not only act as initiator of the classical complement pathway, but can also mediate multiple immune responses in a complement activation independent manner.
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http://dx.doi.org/10.1016/j.imbio.2015.05.001DOI Listing
June 2016

Complement C1q-induced activation of β-catenin signalling causes hypertensive arterial remodelling.

Nat Commun 2015 Feb 26;6:6241. Epub 2015 Feb 26.

1] Department of Cardiovascular Medicine, The University of Tokyo Graduate School of Medicine, Tokyo 113-8655, Japan [2] CREST, Japan Science and Technology Agency, Tokyo 102-0075, Japan [3] Department of Cardiovascular Medicine, Osaka University Graduate School of Medicine, Osaka 565-0871, Japan.

Hypertension induces structural remodelling of arteries, which leads to arteriosclerosis and end-organ damage. Hyperplasia of vascular smooth muscle cells (VSMCs) and infiltration of immune cells are the hallmark of hypertensive arterial remodelling. However, the precise molecular mechanisms of arterial remodelling remain elusive. We have recently reported that complement C1q activates β-catenin signalling independent of Wnts. Here, we show a critical role of complement C1-induced activation of β-catenin signalling in hypertensive arterial remodelling. Activation of β-catenin and proliferation of VSMCs were observed after blood-pressure elevation, which were prevented by genetic and chemical inhibition of β-catenin signalling. Macrophage depletion and C1qa gene deletion attenuated the hypertension-induced β-catenin signalling, proliferation of VSMCs and pathological arterial remodelling. Our findings unveil the link between complement C1 and arterial remodelling and suggest that C1-induced activation of β-catenin signalling becomes a novel therapeutic target to prevent arteriosclerosis in patients with hypertension.
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http://dx.doi.org/10.1038/ncomms7241DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4351572PMC
February 2015