MEF2C regulates NK cell effector functions through control of lipid metabolism.

Natural killer (NK) cells are a critical first line of defense against viral infection. Rare mutations in a small subset of transcription factors can result in decreased NK cell numbers and function in humans, with an associated increased susceptibility to viral infection. However, our understanding of the specific transcription factors governing mature human NK cell function is limited. Here we use a non-viral CRISPR-Cas9 knockout screen targeting genes encoding 31 transcription factors differentially expressed during human NK cell development. We identify myocyte enhancer factor 2C (MEF2C) as a master regulator of human NK cell functionality ex vivo. MEF2C-haploinsufficient patients and mice displayed defects in NK cell development and effector function, with an increased susceptibility to viral infection. Mechanistically, MEF2C was required for an interleukin (IL)-2- and IL-15-mediated increase in lipid content through regulation of sterol regulatory element-binding protein (SREBP) pathways. Supplementation with oleic acid restored MEF2C-deficient and MEF2C-haploinsufficient patient NK cell cytotoxic function. Therefore, MEF2C is a critical orchestrator of NK cell antiviral immunity by regulating SREBP-mediated lipid metabolism.

The X-linked epigenetic regulator UTX controls NK cell-intrinsic sex differences.

Viral infection outcomes are sex biased, with males generally more susceptible than females. Paradoxically, the numbers of antiviral natural killer (NK) cells are increased in males. We demonstrate that while numbers of NK cells are increased in male mice, they display decreased effector function compared to females in mice and humans. These differences were not solely dependent on gonadal hormones, because they persisted in gonadectomized mice. Kdm6a (which encodes the protein UTX), an epigenetic regulator that escapes X inactivation, was lower in male NK cells, while NK cell-intrinsic UTX deficiency in female mice increased NK cell numbers and reduced effector responses. Furthermore, mice with NK cell-intrinsic UTX deficiency showed increased lethality to mouse cytomegalovirus. Integrative multi-omics analysis revealed a critical role for UTX in regulating chromatin accessibility and gene expression critical for NK cell homeostasis and effector function. Collectively, these data implicate UTX as a critical molecular determinant of sex differences in NK cells.

The transcription factor Fli1 restricts the formation of memory precursor NK cells during viral infection.

Natural killer (NK) cells are innate lymphocytes that possess traits of adaptive immunity, such as memory formation. However, the molecular mechanisms by which NK cells persist to form memory cells are not well understood. Using single-cell RNA sequencing, we identified two distinct effector NK cell (NKeff) populations following mouse cytomegalovirus infection. Ly6C- memory precursor (MP) NK cells showed enhanced survival during the contraction phase in a Bcl2-dependent manner, and differentiated into Ly6C+ memory NK cells. MP NK cells exhibited distinct transcriptional and epigenetic signatures compared with Ly6C+ NKeff cells, with a core epigenetic signature shared with MP CD8+ T cells enriched in ETS1 and Fli1 DNA-binding motifs. Fli1 was induced by STAT5 signaling ex vivo, and increased levels of the pro-apoptotic factor Bim in early effector NK cells following viral infection. These results suggest that a NK cell-intrinsic checkpoint controlled by the transcription factor Fli1 limits MP NK formation by regulating early effector NK cell fitness during viral infection.

Single-cell sequencing of human white adipose tissue identifies new cell states in health and obesity.

White adipose tissue (WAT) is an essential regulator of energy storage and systemic metabolic homeostasis. Regulatory networks consisting of immune and structural cells are necessary to maintain WAT metabolism, which can become impaired during obesity in mammals. Using single-cell transcriptomics and flow cytometry, we unveil a large-scale comprehensive cellular census of the stromal vascular fraction of healthy lean and obese human WAT. We report new subsets and developmental trajectories of adipose-resident innate lymphoid cells, dendritic cells and monocyte-derived macrophage populations that accumulate in obese WAT. Analysis of cell-cell ligand-receptor interactions and obesity-enriched signaling pathways revealed a switch from immunoregulatory mechanisms in lean WAT to inflammatory networks in obese WAT. These results provide a detailed and unbiased cellular landscape of homeostatic and inflammatory circuits in healthy human WAT.

Regulation of systemic metabolism by tissue-resident immune cell circuits.

Recent studies have demonstrated that tissue homeostasis and metabolic function are dependent on distinct tissue-resident immune cells that form functional cell circuits with structural cells. Within these cell circuits, immune cells integrate cues from dietary contents and commensal microbes in addition to endocrine and neuronal signals present in the tissue microenvironment to regulate structural cell metabolism. These tissue-resident immune circuits can become dysregulated during inflammation and dietary overnutrition, contributing to metabolic diseases. Here, we review the evidence describing key cellular networks within and between the liver, gastrointestinal tract, and adipose tissue that control systemic metabolism and how these cell circuits become dysregulated during certain metabolic diseases. We also identify open questions in the field that have the potential to enhance our understanding of metabolic health and disease.

Infection induces tissue-resident memory NK cells that safeguard tissue health.

Tissue health is dictated by the capacity to respond to perturbations and then return to homeostasis. Mechanisms that initiate, maintain, and regulate immune responses in tissues are therefore essential. Adaptive immunity plays a key role in these responses, with memory and tissue residency being cardinal features. A corresponding role for innate cells is unknown. Here, we have identified a population of innate lymphocytes that we term tissue-resident memory-like natural killer (NKRM) cells. In response to murine cytomegalovirus infection, we show that circulating NK cells were recruited in a CX3CR1-dependent manner to the salivary glands where they formed NKRM cells, a long-lived, tissue-resident population that prevented autoimmunity via TRAIL-dependent elimination of CD4+ T cells. Thus, NK cells develop adaptive-like features, including long-term residency in non-lymphoid tissues, to modulate inflammation, restore immune equilibrium, and preserve tissue health. Modulating the functions of NKRM cells may provide additional strategies to treat inflammatory and autoimmune diseases.

ILC1 Confer Early Host Protection at Initial Sites of Viral Infection.

Infection is restrained by the concerted activation of tissue-resident and circulating immune cells. Whether tissue-resident lymphocytes confer early antiviral immunity at local sites of primary infection prior to the initiation of circulating responses is not well understood. Furthermore, the kinetics of initial antiviral responses at sites of infection remain unclear. Here, we show that tissue-resident type 1 innate lymphoid cells (ILC1) serve an essential early role in host immunity through rapid production of interferon (IFN)-γ following viral infection. Ablation of Zfp683-dependent liver ILC1 lead to increased viral load in the presence of intact adaptive and innate immune cells critical for mouse cytomegalovirus (MCMV) clearance. Swift production of interleukin (IL)-12 by tissue-resident XCR1+ conventional dendritic cells (cDC1) promoted ILC1 production of IFN-γ in a STAT4-dependent manner to limit early viral burden. Thus, ILC1 contribute an essential role in viral immunosurveillance at sites of initial infection in response to local cDC1-derived proinflammatory cytokines.

Mouse cytomegalovirus-experienced ILC1s acquire a memory response dependent on the viral glycoprotein m12.

Innate lymphoid cells (ILCs) are tissue-resident sentinels that are essential for early host protection from pathogens at initial sites of infection. However, whether pathogen-derived antigens directly modulate the responses of tissue-resident ILCs has remained unclear. In the present study, it was found that liver-resident type 1 ILCs (ILC1s) expanded locally and persisted after the resolution of infection with mouse cytomegalovirus (MCMV). ILC1s acquired stable transcriptional, epigenetic and phenotypic changes a month after the resolution of MCMV infection, and showed an enhanced protective effector response to secondary challenge with MCMV consistent with a memory lymphocyte response. Memory ILC1 responses were dependent on the MCMV-encoded glycoprotein m12, and were independent of bystander activation by proinflammatory cytokines after heterologous infection. Thus, liver ILC1s acquire adaptive features in an MCMV-specific manner.

Type 1 Innate Lymphoid Cells Protect Mice from Acute Liver Injury via Interferon-γ Secretion for Upregulating Bcl-xL Expression in Hepatocytes.

Although type 1 innate lymphoid cells (ILC1s) have been originally found as liver-resident ILCs, their pathophysiological role in the liver remains poorly investigated. Here, we demonstrated that carbon tetrachloride (CCl4) injection into mice activated ILC1s, but not natural killer (NK) cells, in the liver. Activated ILC1s produced interferon-γ (IFN-γ) and protected mice from CCl4-induced acute liver injury. IFN-γ released from activated ILC1s promoted the survival of hepatocytes through upregulation of Bcl-xL. An activating NK receptor, DNAM-1, was required for the optimal activation and IFN-γ production of liver ILC1s. Extracellular adenosine triphosphate accelerated interleukin-12-driven IFN-γ production by liver ILC1s. These findings suggest that ILC1s are critical for tissue protection during acute liver injury.

No time to die: Epigenetic regulation of natural killer cell survival.

NK cells are short-lived innate lymphocytes that can mediate antigen-independent responses to infection and cancer. However, studies from the past two decades have shown that NK cells can acquire transcriptional and epigenetic modifications during inflammation that result in increased survival and lifespan. These findings blur the lines between the innate and adaptive arms of the immune system, and suggest that the homeostatic mechanisms that govern the persistence of innate immune cells are malleable. Indeed, recent studies have shown that NK cells undergo continuous and strictly regulated adaptations controlling their survival during development, tissue residency, and following inflammation. In this review, we summarize our current understanding of the critical factors regulating NK cell survival throughout their lifespan, with a specific emphasis on the epigenetic modifications that regulate the survival of NK cells in various contexts. A precise understanding of the molecular mechanisms that govern NK cell survival will be important to enhance therapies for cancer and infectious diseases.

Adipose-Resident Group 1 Innate Lymphoid Cells Promote Obesity-Associated Insulin Resistance.

Innate lymphoid cells (ILCs) function to protect epithelial barriers against pathogens and maintain tissue homeostasis in both barrier and non-barrier tissues. Here, utilizing Eomes reporter mice, we identify a subset of adipose group 1 ILC (ILC1) and demonstrate a role for these cells in metabolic disease. Adipose ILC1s were dependent on the transcription factors Nfil3 and T-bet but phenotypically and functionally distinct from adipose mature natural killer (NK) and immature NK cells. Analysis of parabiotic mice revealed that adipose ILC1s maintained long-term tissue residency. Diet-induced obesity drove early production of interleukin (IL)-12 in adipose tissue depots and led to the selective proliferation and accumulation of adipose-resident ILC1s in a manner dependent on the IL-12 receptor and STAT4. ILC1-derived interferon-γ was necessary and sufficient to drive proinflammatory macrophage polarization to promote obesity-associated insulin resistance. Thus, adipose-resident ILC1s contribute to obesity-related pathology in response to dysregulated local proinflammatory cytokine production.

Cold versus hot snare endoscopic mucosal resection for large (≥15 mm) flat non-pedunculated colorectal polyps: a randomised controlled trial.

BACKGROUND AND AIMS: Conventional hot snare endoscopic mucosal resection (H-EMR) is effective for the management of large (≥20 mm) non-pedunculated colon polyps (LNPCPs) however, electrocautery-related complications may incur significant morbidity. With a superior safety profile, cold snare EMR (C-EMR) of LNPCPs is an attractive alternative however evidence is lacking. We conducted a randomised trial to compare the efficacy and safety of C-EMR to H-EMR. METHODS: Flat, 15-50 mm adenomatous LNPCPs were prospectively enrolled and randomly assigned to C-EMR or H-EMR with margin thermal ablation at a single tertiary centre. The primary outcome was endoscopically visible and/or histologically confirmed recurrence at 6 months surveillance colonoscopy. Secondary outcomes were clinically significant post-EMR bleeding (CSPEB), delayed perforation and technical success. RESULTS: 177 LNPCPs in 177 patients were randomised to C-EMR arm (n=87) or H-EMR (n=90). Treatment groups were equivalent for technical success 86/87 (98.9%) C-EMR versus H-EMR 90/90 (100%); p=0.31. Recurrence was significantly greater in C-EMR (16/87, 18.4% vs 1/90, 1.1%; relative risk (RR) 16.6, 95% CI 2.24 to 122; p<0.001).Delayed perforation (1/90 (1.1%) vs 0; p=0.32) only occurred in the H-EMR group. CSPEB was significantly greater in the H-EMR arm (7/90 (7.8%) vs 1/87 (1.1%); RR 6.77, 95% CI 0.85 to 53.9; p=0.034). CONCLUSION: Compared with H-EMR, C-EMR for flat, adenomatous LNPCPs, demonstrates superior safety with equivalent technical success. However, endoscopic recurrence is significantly greater for cold snare resection and is currently a limitation of the technique. TRIAL REGISTRATION NUMBER: NCT04138030.

Leukemia inhibitory factor protects against graft-versus-host disease while preserving graft-versus-leukemia activity.

Graft-versus-host disease (GVHD) remains a major complication after allogeneic hematopoietic stem cell transplantation, a widely used therapy for hematologic malignancies and blood disorders. Here, we report an unexpected role of cytokine leukemia inhibitory factor (LIF) in protecting against GVHD development. Administrating recombinant LIF protein (rLIF) protects mice from GVHD-induced tissue damage and lethality without compromising the graft-versus-leukemia activity, which is crucial to prevent tumor relapse. We found that rLIF decreases the infiltration and activation of donor immune cells and protects intestinal stem cells to ameliorate GVHD. Mechanistically, rLIF downregulates IL-12-p40 expression in recipient dendritic cells after irradiation through activating STAT1 signaling, which results in decreased major histocompatibility complex II levels on intestinal epithelial cells and decreased donor T-cell activation and infiltration. This study reveals a previously unidentified protective role of LIF for GVHD-induced tissue pathology and provides a potential effective therapeutic strategy to limit tissue pathology without compromising antileukemic efficacy.

Natural Killer Cell Memory.

Natural killer (NK) cells have historically been considered short-lived cytolytic cells that can rapidly respond against pathogens and tumors in an antigen-independent manner and then undergo cell death. Recently, however, NK cells have been shown to possess traits of adaptive immunity and can acquire immunological memory in a manner similar to that of T and B cells. In this review, we discuss evidence of NK cell memory and the mechanisms involved in the generation and survival of these innate lymphocytes.

BNIP3- and BNIP3L-Mediated Mitophagy Promotes the Generation of Natural Killer Cell Memory.

Natural killer (NK) cells are innate lymphocytes that possess traits of adaptive immunity, such as clonal expansion, contraction, and generation of long-lived "memory" cells, processes poorly understood at the molecular level. Here, we found that as proliferating NK cells accumulated dysfunctional mitochondria during viral infection, a protective mitophagy pathway was induced during the contraction phase to promote their survival in a reactive oxygen species (ROS)-dependent manner. Inhibition of mechanistic target of rapamycin (mTOR) or activation of AMP-activated protein kinase (AMPK) during the contraction-to-memory phase transition of the antiviral response increased autophagic activity and enhanced memory NK cell numbers through an Atg3-dependent mechanism. Furthermore, we demonstrated a temporally regulated role for mitophagy-inducing proteins BCL2/adenovirus E1B 19-kDa interacting protein 3 (BNIP3) and BNIP3-like (BNIP3L) in the generation of robust NK cell memory. Thus, our study reveals the functional importance of mitophagy during the dynamic response of these cytolytic innate lymphocytes.

Long-term outcomes after endoscopic submucosal dissection for relative indication early gastric cancer in nonsurgical candidates.

BACKGROUND AND AIMS: Endoscopic submucosal dissection (ESD) is effective in treating early gastric cancer (EGC). Its role in patients with comorbidities along with more advanced disease is unknown. We sought to evaluate this in a large Western cohort. METHODS: Consecutive patients who underwent ESD for EGC in a single tertiary Western endoscopy center over 10 years were prospectively analyzed. The primary outcomes were long-term overall survival (OS) and disease-free survival (DFS) up to 5 years. Secondary outcomes were efficacy and serious adverse events (SAEs). RESULTS: ESD for 157 cases of EGC in 149 patients was performed in an elderly and comorbid cohort with a mean age of 73.7 years and age-adjusted Charlson Comorbidity Index of 4.2. Over a median follow-up of 51.6 months, no significant differences were found in 5-year OS (88.9% vs 77.9%, P = .290) and DFS (83.2% vs 75.1%, P = .593) between absolute indication (AI) EGC and relative indication (RI) EGC. The AI EGC cohort achieved higher en-bloc (96.3% vs 87.5%, P = .069) and R0 resection rates (93.6% vs 62.5%, P < .001) when compared with RI EGC. No significant differences were found in SAEs (7.3% vs 12.5%, P = .363). No mortality or surgical resection ensued from adverse events from ESD. CONCLUSIONS: ESD safely confers DFS in poor surgical candidates with RI EGC in a large Western cohort. Patients who are elderly and/or with comorbidities or decline surgical resection may benefit from ESD and avoid the risks of surgery and its long-term sequelae. (Clinical trial registration number: NCT02306707.).

Hybrid resection versus conventional resection for laterally spreading lesions of the papilla.

BACKGROUND AND AIMS: Although conventional hot snare resection (CR) of laterally spreading lesions of the major papilla (LSL-Ps) is effective, it can be associated with delayed bleeding in upward of 25% of cases. Given the excellent safety profile of cold snare polypectomy in the colorectum, we investigated the efficacy and safety of a novel hybrid resection (HR) technique for LSL-P management, consisting of hot snare papillectomy plus cold snare resection of the laterally spreading component. METHODS: A prospective cohort of patients underwent HR in a tertiary referral center over 60 months until December 2022. This cohort was compared with a historical cohort of patients who underwent CR at the same institution over 120 months until August 2017. The primary outcomes were recurrence and bleeding. RESULTS: Twenty patients underwent HR (14 female; mean age 65.2 ± 12.2 years). Median lesion size was 30 mm (interquartile range, 25.0-47.5 mm). Recurrent or residual adenoma (RRA) was greater with HR (58.8% [n = 10] vs 29.8% [n = 14]; P = .034). The odds ratio for recurrence was 3.6 times (95% CI, 1.2-11.0) higher with HR (P = .027). RRA was multifocal in 4 (40%) and had a composite RRA volume >10 mm in 7 (70%). The median number of procedures required to treat RRA was higher with HR (4 vs 1, P = .002). There was no difference between CR and HR for intraprocedural bleeding (41.1% [n = 23] vs 25% [n = 5]; P = .587) or delayed bleeding (25.0% vs 10.0%, P = .211). There were no perforations. CONCLUSIONS: The novel HR technique for LSL-P management is associated with a high rate of RRA that is recalcitrant to treatment, without mitigating the risk of intraprocedural or delayed bleeding. Therefore, CR should remain the mainstay management option for treating patients with an LSL-P. (Clinical trial registration number: NCT02306603.).

Endoscopic resection of large anastomotic polyps is safe and effective.

BACKGROUND: Large (≥20mm) adenomatous anastomotic polyps (LAAPs) are uncommon. Data pertaining to their prevalence, characteristics, and the efficacy of endoscopic resection (ER) are absent. A safe and effective strategy for ER would reduce morbidity and healthcare costs. METHODS: Large nonpedunculated colorectal polyps of ≥20mm (LNPCPs) referred for ER were prospectively studied. Multiple data points were recorded including anastomotic location, polyp morphology, resection modality, complications, and technical success. RESULTS: Over 7 years until November 2022, 2629 lesions were referred. Of these, 10 (0.4%) were LAAPs (median size 35 mm [interquartile range (IQR) 30-40mm]). All LAAPs were removed by piecemeal endoscopic mucosal resection (EMR), most (n=9; 90%) in combination with cold-forceps avulsion with adjuvant snare-tip soft coagulation (CAST). On comparison of the LAAP group with the conventional LNPCP group, CAST was more commonly used (90% vs. 9%; P<0.001) and deep mural injury (DMI) type II was more frequent (40% vs. 11%, P=0.003); however, significant DMI (III-V) did not occur. At 6 month (IQR 5.25-6 months) surveillance, there was no recurrence in any of the 10 cases. There were no serious adverse events. CONCLUSIONS: LAAPs present unique challenges owing to their location overlying an anastomosis. Despite these challenges they can be safely and effectively managed endoscopically without recurrence at endoscopic follow-up.