Peripheral helper T (T _PH ) cells can play pathogenic roles in human autoimmune diseases. T _PH cells are proposed to be the major B cell helpers in inflamed joints in rheumatoid arthritis (RA), but whether and how T _PH cells are engaged in tissue inflammation remains unclear. We demonstrate that T _PH cells comprise two subsets in RA: stem-like T _PH (S-T _PH ) and effector T _PH (E-T _PH ) cells. These two subsets differed in transcriptome, epigenome, B cell helper capacity, spatial localization, and cell interactions. S-T _PH cells displayed self-renewal capacity and were mainly found within tertiary lymphoid structures (TLSs) in synovial tissue together with B cells. S-T _PH cells potently induced B cells to produce immunoglobulins. By contrast, E-T _PH cells expressed effector molecules and colocalized with proinflammatory macrophages and CD8 ⁺ T cells outside TLSs. S-T _PH cells could differentiate into E-T _PH cells upon TCR stimulation and coculture with B cells. Collectively, our study shows that S-T _PH cells play a central role in promoting T _PH responses by undergoing self-renewal and seeding E-T _PH cells.

Human CD4 ⁺ T cells play a central role in the pathogenesis of autoimmune diseases, but their immunoregulatory mechanisms driving pathogenesis remain to be elucidated. We show that human T peripheral helper cells (T _PH cells) regulate peripheral immune responses via insulin-like growth factor–like family member 2 (IGFL2), an inflammatory factor found exclusively in primates. Single-cell RNA sequencing of seropositive rheumatoid arthritis (RA) synovium showed that IGFL2 is specifically expressed by CD4 ⁺ T cells, predominantly T _PH cells. IGFL2 promotes transforming growth factor–β–induced CXCL13 production in CD4 ⁺ T cells, activates nuclear factor κB signaling, and induces monocyte gene signatures like those of pathogenic macrophages. CRISPR-Cas9 knockout of IGFL2 in synovial T _PH cells suppressed this gene signature in cocultured monocytes. Blood IGFL2 protein levels correlated with RA disease severity and could be used as a potential biomarker. These findings highlight the involvement of IGFL2 in RA pathogenesis, emphasizing how human T _PH cells regulate local immune responses via IGFL2.

RNF213 encodes a unique protein with AAA+ ATPase and E3 ubiquitin ligase activities that are critical for its diverse roles, which range from involvement in human vasculopathies, such as Moyamoya disease, to ubiquitination of viral and bacterial pathogens. Nevertheless, its primary functions in human signaling remain unclear due to the limited identification of direct substrates. Here, we investigated the interaction between RNF213 and caveolin-1 (Cav-1), a small scaffolding protein vital for caveolae formation and the regulation of a plethora of cellular processes. Cav-1 specifically binds within the two functional AAA+ domains of RNF213 in an ATP-dependent manner, highlighting the influence of cellular energy status on this interaction. Consequently, RNF213 ubiquitinates Cav-1 at several N-terminal lysine residues through K48 and K63 linkages, although several Moyamoya disease-associated RNF213 mutations greatly reduce this polyubiquitination. Moreover, RNF213 activity inhibits phosphorylation of a key regulatory residue of Cav-1, as RNF213 knockdown under oxidative stress markedly enhances Cav-1 Tyr14 phosphorylation and modifies nitric oxide bioavailability in endothelial cells. Collectively, our results indicate that RNF213 functions as a molecular switch modulating Cav-1 signaling based on RNF213 functionality and cellular conditions. These findings offer new insights into vascular pathogenesis and the vast signal pathways along the RNF213–Cav-1 axis.

CD4 ⁺ tissue-resident memory T (T _RM ) cells contribute to host defense and to the pathogenesis of chronic inflammatory diseases, but the molecules that direct their differentiation are unknown. We found that the transcription factor hepatic leukemia factor (HLF) could direct the tissue residency program and function of CD4 ⁺ T _RM cells. HLF simultaneously up-regulated tissue retention receptors, down-regulated tissue egress receptors, and promoted proinflammatory CD4 ⁺ T _RM cells by inducing Bhlhe40 , and all of these processes were associated with changes in chromatin accessibility. Genetic deletion of Hlf inhibited CD4 ⁺ T _RM cell generation and ameliorated airway tissue inflammation in vivo. HLF ⁺ CD4 ⁺ T _RM cells isolated from inflamed airway tissue in humans had a tissue residency signature and expressed inflammatory cytokines. We conclude that HLF may act as a central regulator of proinflammatory CD4 ⁺ T _RM cell development and function.

Studies using mouse models of viral infection have shown that viral antigen-reactive regulatory T cells (Tregs) play multiple context-dependent roles, including attenuating the immune response against the pathogen and limiting inflammatory damage. By contrast, evidence for the presence and function of viral antigen-specific Tregs in humans remains very limited. Here, we investigated whether BNT162b2 mRNA COVID-19 vaccination activates spike (S)-reactive Tregs in healthy adults. We performed an integrated analysis combining single-cell RNA sequencing (scRNA-seq), T-cell receptor repertoire profiling, and flow cytometry of activation-induced marker (AIM)-expressing S-reactive CD4+ T cells isolated from peripheral blood mononuclear cells cultured with trimeric S protein. We found that CD39+ cells coexpressing CTLA4 and TIGIT within the S-reactive AIM+ CD4+ T-cell population were preferentially enriched for Tregs relative to their CD39− counterparts. In subjects vaccinated with the BNT162b2 mRNA vaccine, the frequencies of CD39+CTLA4+ and CD39+TIGIT+ S-reactive Treg-like cells were significantly increased following vaccination. Using scRNA-seq, we identified two FoxP3+ Treg clusters among S-reactive CD4+ T cells: one characterized by IFN-activated gene signatures and another by high HLA class II, CD39, and CTLA4 expression. Both subsets were transcriptionally activated following vaccination, with upregulation of genes associated with T-cell activation and inflammatory cytokines, including TNF and IFNG. Consistent with these findings, flow cytometry detected increased proportions of CD39+ and FoxP3+ S-reactive Tregs expressing IFN-γ and/or TNF-α following vaccination. Collectively, these findings support a model in which BNT162b2 mRNA vaccination activates both an S-reactive effector T-cell response and a Treg response, suggesting Treg coinduction may be critical for balancing antiviral immunity with immunoregulation.

Primary sclerosing cholangitis (PSC) is a chronic autoimmune cholestatic liver disease characterized by progressive inflammation and fibrosis, but the role of neutrophils in PSC remains poorly understood. To define neutrophil states in the PSC liver microenvironment, we performed single-cell RNA sequencing of leukocytes isolated from liver perfusate obtained from healthy donor grafts and explanted PSC livers. We identified a marked expansion of transcriptionally diverse neutrophil subclusters in PSC liver perfusate. Importantly, PSC livers were enriched for four neutrophil subclusters: RHOB-high, immediate early gene-high, heat shock protein-high, and CXCL8-high liver neutrophil-like subclusters. These PSC-associated neutrophil subclusters showed increased activity of the TNF and IL-17 signaling pathways, consistent with current views of PSC pathogenesis. They also displayed activation of inflammatory transcriptional regulators, including CEBPB, JUN, FOS, and RELA. Pseudotime analysis suggested that RHOB and immediate early gene subsets were derived from immature neutrophils, whereas liver neutrophil-like and heat shock protein-high subsets exhibited more mature and tissue-adapted features. Together, these data reveal previously unrecognized heterogeneity of liver neutrophils in PSC and identify inflammatory neutrophil programs that may contribute to sustained hepatic inflammation and represent potential therapeutic targets.