Interleukin 21 Drives a Hypermetabolic State and CD4+ T-Cell-Associated Pathogenicity in Chronic Intestinal Inflammation.

BACKGROUND & AIMS: Incapacitated regulatory T cells (Tregs) contribute to immune-mediated diseases. Inflammatory Tregs are evident during human inflammatory bowel disease; however, mechanisms driving the development of these cells and their function are not well understood. Therefore, we investigated the role of cellular metabolism in Tregs relevant to gut homeostasis. METHODS: Using human Tregs, we performed mitochondrial ultrastructural studies via electron microscopy and confocal imaging, biochemical and protein analyses using proximity ligation assay, immunoblotting, mass cytometry and fluorescence-activated cell sorting, metabolomics, gene expression analysis, and real-time metabolic profiling utilizing the Seahorse XF analyzer. We used a Crohn's disease single-cell RNA sequencing dataset to infer the therapeutic relevance of targeting metabolic pathways in inflammatory Tregs. We examined the superior functionality of genetically modified Tregs in CD4+ T-cell-induced murine colitis models. RESULTS: Mitochondria-endoplasmic reticulum appositions, known to mediate pyruvate entry into mitochondria via voltage-dependent anion channel 1 (VDAC1), are abundant in Tregs. VDAC1 inhibition perturbed pyruvate metabolism, eliciting sensitization to other inflammatory signals reversible by membrane-permeable methyl pyruvate supplementation. Notably, interleukin (IL) 21 diminished mitochondria-endoplasmic reticulum appositions, resulting in enhanced enzymatic function of glycogen synthase kinase 3 ß, a putative negative regulator of VDAC1, and a hypermetabolic state that amplified Treg inflammatory response. Methyl pyruvate and glycogen synthase kinase 3 ß pharmacologic inhibitor (LY2090314) reversed IL21-induced metabolic rewiring and inflammatory state. Moreover, IL21-induced metabolic genes in Tregs in vitro were enriched in human Crohn's disease intestinal Tregs. Adoptively transferred Il21r-/- Tregs efficiently rescued murine colitis in contrast to wild-type Tregs. CONCLUSIONS: IL21 triggers metabolic dysfunction associated with Treg inflammatory response. Inhibiting IL21-induced metabolism in Tregs may mitigate CD4+ T-cell-driven chronic intestinal inflammation.

Interleukin-21 Drives a Hypermetabolic State and CD4+ T Cell-associated Pathogenicity in Chronic Intestinal Inflammation.

BACKGROUND & AIMS: Incapacitated regulatory T cells (Tregs) contribute to immune-mediated diseases. Inflammatory Tregs are evident during human inflammatory bowel disease (IBD); however, mechanisms driving the development of these cells and their function are not well understood. Therefore, we investigated the role of cellular metabolism in Tregs relevant to gut homeostasis. METHODS: Using human Tregs, we performed mitochondrial ultrastructural studies via electron microscopy and confocal imaging, biochemical and protein analyses using proximity ligation assay, immunoblotting, mass cytometry and fluorescence-activated cell sorting, metabolomics, gene expression analysis, and real-time metabolic profiling utilizing Seahorse XF analyzer. We utilized Crohn's disease single-cell RNA sequencing dataset to infer therapeutic relevance of targeting metabolic pathways in inflammatory Tregs. We examined the superior functionality of genetically-modified Tregs in CD4+ T cell-induced murine colitis models. RESULTS: Mitochondria-endoplasmic reticulum (ER) appositions, known to mediate pyruvate entry into mitochondria via VDAC1, are abundant in Tregs. VDAC1 inhibition perturbed pyruvate metabolism, eliciting sensitization to other inflammatory signals reversible by membrane-permeable methyl pyruvate (MePyr) supplementation. Notably, IL-21 diminished mitochondria-ER appositions, resulting in enhanced enzymatic function of glycogen synthase kinase 3 ß (GSK3ß), a putative negative regulator of VDAC1, and a hypermetabolic state that amplified Treg inflammatory response. MePyr and GSK3ß pharmacologic inhibitor (LY2090314) reversed IL-21-induced metabolic rewiring and inflammatory state. Moreover, IL-21-induced metabolic genes in Tregs in vitro were enriched in human Crohn's disease intestinal Tregs. Adoptively transferred Il21r-/- Tregs efficiently rescued murine colitis in contrast to wild-type Tregs. CONCLUSIONS: IL-21 triggers metabolic dysfunction associated with Treg inflammatory response. Inhibiting IL-21-induced metabolism in Tregs may mitigate CD4+ T cell-driven chronic intestinal inflammation.

Matrix-induced autologous chondrocyte implantation (MACI) in the knee: clinical outcomes and challenges.

PURPOSE: Matrix-induced autologous chondrocyte implantation (MACI) has demonstrated effectiveness in treating isolated cartilage defects of the knee but medium- and long-term evidence and information on the management of postoperative complications or partially successful cases are sparse. This study hypothesised that MACI is effective for up to 5 years and that patients with posttreatment problems may go on to obtain clinical benefit from other interventions. METHODS: A follow-on, prospective case series of patients recruited into a previous controlled, randomised, prospective study or newly enroled. Patients were followed up 6, 12, 24 and 60 months after surgery. Outcome measures were Tegner (activity levels) and Lysholm (pain, stability, gait, clinical symptoms) scores. Zone-specific subgroups were analysed 6, 12 and 24 months postoperatively. RESULTS: Sixty-five patients were treated with MACI. Median Tegner score improved from II to IV at 12 months; an improvement maintained to 60 months. Mean Lysholm score improved from 28.5 to 76.6 points (±19.8) at 24 months, settling back to 75.5 points after 5 years (p > 0.0001). No significant differences were identified in the zone-specific analysis. Posttreatment issues (N = 12/18.5 %) were resolved with microfracture, debridement, OATS or bone grafting. CONCLUSIONS: MACI is safe and effective in the majority of patients. Patients in whom treatment is only partially successful can go on to obtain clinical benefit from other cartilage repair options. This study adds to the clinical evidence on the MACI procedure, offers insight into likely treatment outcomes, and highlights MACI's usefulness as part of an armamentarium of surgical approaches to the treatment of isolated knee defects. LEVEL OF EVIDENCE: Prospective case control study with no control group, Level III.