The histone demethylase JMJD1C regulates CPS1 expression and promotes the proliferation of paroxysmal nocturnal haemoglobinuria clones through cell metabolic reprogramming.

Paroxysmal nocturnal haemoglobinuria (PNH) is a disorder resulting from erythrocyte membrane deficiencies caused by PIG-A gene mutations. While current treatments alleviate symptoms, they fail to address the underlying cause of the disease-the pathogenic PNH clones. In this study, we found that the expression of carbamoyl phosphate synthetase 1 (CPS1) was downregulated in PNH clones, and the level of CPS1 was negatively correlated with the proportion of PNH clones. Using PIG-A knockout K562 (K562 KO) cells, we demonstrated that CPS1 knockdown increased cell proliferation and altered cell metabolism, suggesting that CPS1 participates in PNH clonal proliferation through metabolic reprogramming. Furthermore, we observed an increase in the expression levels of the histone demethylase JMJD1C in PNH clones, and JMJD1C expression was negatively correlated with CPS1 expression. Knocking down JMJD1C in K562 KO cells upregulated CPS1 and H3K36me3 expression, decreased cell proliferation and increased cell apoptosis. Chromatin immunoprecipitation analysis further demonstrated that H3K36me3 regulated CPS1 expression. Finally, we demonstrated that histone demethylase inhibitor JIB-04 can suppressed K562 KO cell proliferation and reduced the proportion of PNH clones in PNH mice. In conclusion, aberrant regulation of the JMJD1C-H3K36me3-CPS1 axis contributes to PNH clonal proliferation. Targeting JMJD1C with a specific inhibitor unveils a potential strategy for treating PNH patients.

PMI-controlled mannose metabolism and glycosylation determines tissue tolerance and virus fitness.

Host survival depends on the elimination of virus and mitigation of tissue damage. Herein, we report the modulation of D-mannose flux rewires the virus-triggered immunometabolic response cascade and reduces tissue damage. Safe and inexpensive D-mannose can compete with glucose for the same transporter and hexokinase. Such competitions suppress glycolysis, reduce mitochondrial reactive-oxygen-species and succinate-mediated hypoxia-inducible factor-1α, and thus reduce virus-induced proinflammatory cytokine production. The combinatorial treatment by D-mannose and antiviral monotherapy exhibits in vivo synergy despite delayed antiviral treatment in mouse model of virus infections. Phosphomannose isomerase (PMI) knockout cells are viable, whereas addition of D-mannose to the PMI knockout cells blocks cell proliferation, indicating that PMI activity determines the beneficial effect of D-mannose. PMI inhibition suppress a panel of virus replication via affecting host and viral surface protein glycosylation. However, D-mannose does not suppress PMI activity or virus fitness. Taken together, PMI-centered therapeutic strategy clears virus infection while D-mannose treatment reprograms glycolysis for control of collateral damage.

An engineered Sox17 induces somatic to neural stem cell fate transitions independently from pluripotency reprogramming.

Advanced strategies to interconvert cell types provide promising avenues to model cellular pathologies and to develop therapies for neurological disorders. Yet, methods to directly transdifferentiate somatic cells into multipotent induced neural stem cells (iNSCs) are slow and inefficient, and it is unclear whether cells pass through a pluripotent state with full epigenetic reset. We report iNSC reprogramming from embryonic and aged mouse fibroblasts as well as from human blood using an engineered Sox17 (eSox17FNV). eSox17FNV efficiently drives iNSC reprogramming while Sox2 or Sox17 fail. eSox17FNV acquires the capacity to bind different protein partners on regulatory DNA to scan the genome more efficiently and has a more potent transactivation domain than Sox2. Lineage tracing and time-resolved transcriptomics show that emerging iNSCs do not transit through a pluripotent state. Our work distinguishes lineage from pluripotency reprogramming with the potential to generate more authentic cell models for aging-associated neurodegenerative diseases.

Chronic type I interferon signaling promotes lipid-peroxidation-driven terminal CD8+ T cell exhaustion and curtails anti-PD-1 efficacy.

Identifying signals that govern the differentiation of tumor-infiltrating CD8+ T cells (CD8+ TILs) toward exhaustion can improve current therapeutic approaches for cancer. Here, we show that type I interferons (IFN-Is) act as environmental cues, enhancing terminal CD8+ T cell exhaustion in tumors. We find enrichment of IFN-I-stimulated genes (ISGs) within exhausted CD8+ T cells (Tex cells) in patients across various cancer types, with heightened ISG levels correlating with poor response to immune checkpoint blockade (ICB) therapy. In preclinical models, CD8+ TILs devoid of IFN-I signaling develop less exhaustion features, provide better tumor control, and show greater response to ICB-mediated rejuvenation. Mechanistically, chronic IFN-I stimulation perturbs lipid metabolism and redox balance in Tex cells, leading to aberrant lipid accumulation and elevated oxidative stress. Collectively, these defects promote lipid peroxidation, which potentiates metabolic and functional exhaustion of Tex cells. Thus, cell-intrinsic IFN-I signaling regulates the extent of CD8+ TIL exhaustion and has important implications for immunotherapy.

Comparing the B and T cell-mediated immune responses in patients with type 2 diabetes receiving mRNA or inactivated COVID-19 vaccines.

Acquiring protective immunity through vaccination is essential, especially for patients with type 2 diabetes who are vulnerable for adverse clinical outcomes during coronavirus disease 2019 (COVID-19) infection. Type 2 diabetes (T2D) is associated with immune dysfunction. Here, we evaluated the impact of T2D on the immunological responses induced by mRNA (BNT162b2) and inactivated (CoronaVac) vaccines, the two most commonly used COVID-19 vaccines. The study consisted of two parts. In Part 1, the sera titres of IgG antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) alpha receptor binding domain (RBD), their neutralizing capacity, and antigen-specific CD4+T and CD8+T cell responses at 3-6 months after vaccination were compared between BNT162b2 (n=60) and CoronaVac (n=50) vaccinees with or without T2D. Part 2 was a time-course study investigating the initial B and T cell responses induced by BNT162b2 among vaccinees (n=16) with or without T2D. Our data showed that T2D impaired both cellular and humoral immune responses induced by CoronaVac. For BNT162b2, T2D patients displayed a reduction in CD4+T-helper 1 (Th1) differentiation following their first dose. However, this initial defect was rectified by the second dose of BNT162b2, resulting in comparable levels of memory CD4+ and CD8+T cells, anti-RBD IgG, and neutralizing antibodies with healthy individuals at 3-6 months after vaccination. Hence, T2D influences the effectiveness of COVID-19 vaccines depending on their platform. Our findings provide a potential mechanism for the susceptibility of developing adverse outcomes observed in COVID-19 patients with T2D and received either CoronaVac or just one dose of BNT162b2.

Autoantibodies in systemic lupus erythematosus: From immunopathology to therapeutic target.

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by multiple organ inflammatory damage and wide spectrum of autoantibodies. The autoantibodies, especially anti-dsDNA and anti-Sm autoantibodies are highly specific to SLE, and participate in the immune complex formation and inflammatory damage on multiple end-organs such as kidney, skin, and central nervous system (CNS). However, the underlying mechanisms of autoantibody-induced tissue damage and systemic inflammation are still not fully understood. Single cell analysis of autoreactive B cells and monoclonal antibody screening from patients with active SLE has improved our understanding on the origin of autoreactive B cells and the antigen targets of the pathogenic autoantibodies. B cell depletion therapies have been widely studied in the clinics, but the development of more specific therapies against the pathogenic B cell subset and autoantibodies with improved efficacy and safety still remain a big challenge. A more comprehensive autoantibody profiling combined with functional characterization of autoantibodies in diseases development will shed new insights on the etiology and pathogenesis of SLE and guide a specific treatment to individual SLE patients.

Bacillus Calmette-Guérin-induced trained immunity protects against SARS-CoV-2 challenge in K18-hACE2 mice.

SARS-CoV-2 has been confirmed in over 450 million confirmed cases since 2019. Although several vaccines have been certified by the WHO and people are being vaccinated on a global scale, it has been reported that multiple SARS-CoV-2 variants can escape neutralization by antibodies, resulting in vaccine breakthrough infections. Bacillus Calmette-Guérin (BCG) is known to induce heterologous protection based on trained immune responses. Here, we investigated whether BCG-induced trained immunity protected against SARS-CoV-2 in the K18-hACE2 mouse model. Our data demonstrate that i.v. BCG (BCG-i.v.) vaccination induces robust trained innate immune responses and provides protection against WT SARS-CoV-2, as well as the B.1.617.1 and B.1.617.2 variants. Further studies suggest that myeloid cell differentiation and activation of the glycolysis pathway are associated with BCG-induced training immunity in K18-hACE2 mice. Overall, our study provides the experimental evidence that establishes a causal relationship between BCG-i.v. vaccination and protection against SARS-CoV-2 challenge.

Intranasal administration of a single dose of a candidate live attenuated vaccine derived from an NSP16-deficient SARS-CoV-2 strain confers sterilizing immunity in animals.

Live attenuated vaccines might elicit mucosal and sterilizing immunity against SARS-CoV-2 that the existing mRNA, adenoviral vector and inactivated vaccines fail to induce. Here, we describe a candidate live attenuated vaccine strain of SARS-CoV-2 in which the NSP16 gene, which encodes 2'-O-methyltransferase, is catalytically disrupted by a point mutation. This virus, designated d16, was severely attenuated in hamsters and transgenic mice, causing only asymptomatic and nonpathogenic infection. A single dose of d16 administered intranasally resulted in sterilizing immunity in both the upper and lower respiratory tracts of hamsters, thus preventing viral spread in a contact-based transmission model. It also robustly stimulated humoral and cell-mediated immune responses, thus conferring full protection against lethal challenge with SARS-CoV-2 in a transgenic mouse model. The neutralizing antibodies elicited by d16 effectively cross-reacted with several SARS-CoV-2 variants. Secretory immunoglobulin A was detected in the blood and nasal wash of vaccinated mice. Our work provides proof-of-principle evidence for harnessing NSP16-deficient SARS-CoV-2 for the development of live attenuated vaccines and paves the way for further preclinical studies of d16 as a prototypic vaccine strain, to which new features might be introduced to improve safety, transmissibility, immunogenicity and efficacy.

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

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.

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

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.

Complement receptor 3 mediates renal protection in experimental C3 glomerulopathy.

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.

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

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.

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

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.

Intranasal peptide-induced tolerance and linked suppression: consequences of complement deficiency.

A role for complement, particularly the classical pathway, in the regulation of immune responses is well documented. Deficiencies in C1q or C4 predispose to autoimmunity, while deficiency in C3 affects the suppression of contact sensitization and generation of oral tolerance. Complement components including C3 have been shown to be required for both B-cell and T-cell priming. The mechanisms whereby complement can mediate these diverse regulatory effects are poorly understood. Our previous work, using the mouse minor histocompatibility (HY) model of skin graft rejection, showed that both C1q and C3 were required for the induction of tolerance following intranasal peptide administration. By comparing tolerance induction in wild-type C57BL/6 and C1q-, C3-, C4- and C5-deficient C57BL/6 female mice, we show here that the classical pathway components including C3 are required for tolerance induction, whereas C5 plays no role. C3-deficient mice failed to generate a functional regulatory T (Treg) -dendritic cell (DC) tolerogenic loop required for tolerance induction. This was related to the inability of C3-deficient DC to up-regulate the arginine-consuming enzyme, inducible nitric oxide synthase (Nos-2), in the presence of antigen-specific Treg cells and peptide, leading to reduced Treg cell generation. Our findings demonstrate that the classical pathway and C3 play a critical role in the peptide-mediated induction of tolerance to HY by modulating DC function.

C1q as a unique player in angiogenesis with therapeutic implication in wound healing.

We have previously shown that C1q is expressed on endothelial cells (ECs) of newly formed decidual tissue. Here we demonstrate that C1q is deposited in wound-healing skin in the absence of C4 and C3 and that C1q mRNA is locally expressed as revealed by real-time PCR and in situ hybridization. C1q was found to induce permeability of the EC monolayer, to stimulate EC proliferation and migration, and to promote tube formation and sprouting of new vessels in a rat aortic ring assay. Using a murine model of wound healing we observed that vessel formation was defective in C1qa(-/-) mice and was restored to normal after local application of C1q. The mean vessel density of wound-healing tissue and the healed wound area were significantly increased in C1q-treated rats. On the basis of these results we suggest that C1q may represent a valuable therapeutic agent that can be used to treat chronic ulcers or other pathological conditions in which angiogenesis is impaired, such as myocardial ischemia.

IL-10-producing regulatory B cells induced by IL-33 (Breg(IL-33)) effectively attenuate mucosal inflammatory responses in the gut.

Regulatory B cells (Breg) have attracted increasing attention for their roles in maintaining peripheral tolerance. Interleukin 33 (IL-33) is a recently identified IL-1 family member, which leads a double-life with both pro- and anti-inflammatory properties. We report here that peritoneal injection of IL-33 exacerbated inflammatory bowel disease in IL-10-deficient (IL-10(-/-)) mice, whereas IL-33-treated IL-10-sufficient (wild type) mice were protected from the disease induction. A phenotypically unconventional subset(s) (CD19(+)CD25(+)CD1d(hi)IgM(hi)CD5(-)CD23(-)Tim-1(-)) of IL-10 producing Breg-like cells (Breg(IL-33)) was identified responsible for the protection. We demonstrated further that Breg(IL-33) isolated from these mice could suppress immune effector cell expansion and functions and, upon adoptive transfer, effectively blocked the development of spontaneous colitis in IL-10(-/-) mice. Our findings indicate an essential protective role, hence therapeutic potential, of Breg(IL-33) against mucosal inflammatory disorders in the gut.

Integrin CD11b positively regulates TLR4-induced signalling pathways in dendritic cells but not in macrophages.

Tuned and distinct responses of macrophages and dendritic cells to Toll-like receptor 4 (TLR4) activation induced by lipopolysaccharide (LPS) underpin the balance between innate and adaptive immunity. However, the molecule(s) that confer these cell-type-specific LPS-induced effects remain poorly understood. Here we report that the integrin α(M) (CD11b) positively regulates LPS-induced signalling pathways selectively in myeloid dendritic cells but not in macrophages. In dendritic cells, which express lower levels of CD14 and TLR4 than macrophages, CD11b promotes MyD88-dependent and MyD88-independent signalling pathways. In particular, in dendritic cells CD11b facilitates LPS-induced TLR4 endocytosis and is required for the subsequent signalling in the endosomes. Consistent with this, CD11b deficiency dampens dendritic cell-mediated TLR4-triggered responses in vivo leading to impaired T-cell activation. Thus, by modulating the trafficking and signalling functions of TLR4 in a cell-type-specific manner CD11b fine tunes the balance between adaptive and innate immune responses initiated by LPS.