High concentration hydrogen attenuates sepsis-induced acute kidney injury by promoting mitochondrial biogenesis and fusion.

Sepsis is a major cause of mortality among critical patients. Acute kidney injury (AKI) is the common complication in patients with sepsis, characterized by rapid deterioration of renal function. The purpose of this study is to assess the impact of inhaling high concentration hydrogen on septic mice with AKI and to examine the involvement of mitochondria in this process. High concentration hydrogen does not cause hypoxia and can alleviate AKI and improve 7-day survival in septic mice. Inflammatory factors are markedly elevated in the serum and renal tissues in CLP group which are dramatically down-regulated by hydrogen. The activities of both antioxidant enzymes are significantly reduced after CLP, whereas hydrogen markedly increases the activities of SOD and CAT. MMP is found to be significantly lower in CLP group whereas this effect is reversed by hydrogen. The trend of ATP content in renal tissues corresponded with that of MMP. There is a substantial downregulation of PGC-1α, Nrf2, and TFAM protein in CLP group. Drp1 expression is significantly higher in CLP group compared to Sham group, while the opposite trend is observed for MFN2. Hydrogen can reverse these changes. Inhalation of high concentration hydrogen can improve acute kidney injury, 7-day survival, inflammatory response and oxidative stress in septic mice. The mechanism may be related to inhibit renal mitochondrial fission and promote mitochondrial fusion and biogenesis.

CD94 deficiency or blockade unleashes the anti-tumor immunity in mice and humanized murine models.

NKG2 family members have emerged as promising targets in tumor immunotherapy. CD94 can dimerize with both inhibitory and activating NKG2 proteins, while the overall effect and value of targeting CD94 on anti-tumor immunity are unclear. Here, it is shown that the expression of CD94 is upregulated on tumor-infiltrating natural killer (NK) cells and CD8+ T cells, and is related to their exhausted characteristics. Tumor-bearing CD94 knockout (CD94-KO) mice exhibit delayed tumor growth, decreased lung metastases, and prolonged survival. Single cell RNA-seq reveals a remodeled tumor microenvironment in CD94-KO mice, with a reduction in immunosuppressive cells and an increase in anti-tumor immune cells. Moreover, NK cells and CD8+ T cells become proliferative and strongly tumoricidal in CD94-KO mice, thus contributing to the tumor inhibition effect of CD94 deficiency. Treatment with a humanized anti-CD94 blocking antibody (h15C10) alone, in tumor-bearing humanized mouse, delays tumor progression, and improves the therapeutic efficacy of PD-L1 blockade through combination therapy. Our study indicates that CD94 may work as a candidate target in checkpoint immunotherapy.

Intracellular checkpoints for NK cell cancer immunotherapy.

Natural killer (NK) cells are key innate immune lymphocytes, which play important roles against tumors. However, tumor-infiltrating NK cells are always hypofunctional/exhaustive. On the one hand, this state is contributed by context-dependent interactions between inhibitory NK cell checkpoint receptors and their ligands, which usually vary in different tumor types and stages during tumor development. On the other hand, the inhibitory functions of intracellular checkpoint molecules of NK cells are more similar across different tumor types, representing common mechanisms limiting the potential of NK cell therapy. In this review, representative NK cell intracellular checkpoint molecules in different aspects of NK cell biology were reviewed, and therapeutic potentials were discussed by targeting these molecules to promote antitumor NK cell therapy.

Interleukin-34-orchestrated tumor-associated macrophage reprogramming is required for tumor immune escape driven by p53 inactivation.

As the most frequent genetic alteration in cancer, more than half of human cancers have p53 mutations that cause transcriptional inactivation. However, how p53 modulates the immune landscape to create a niche for immune escape remains elusive. We found that cancer stem cells (CSCs) established an interleukin-34 (IL-34)-orchestrated niche to promote tumorigenesis in p53-inactivated liver cancer. Mechanistically, we discovered that Il34 is a gene transcriptionally repressed by p53, and p53 loss resulted in IL-34 secretion by CSCs. IL-34 induced CD36-mediated elevations in fatty acid oxidative metabolism to drive M2-like polarization of foam-like tumor-associated macrophages (TAMs). These IL-34-orchestrated TAMs suppressed CD8+ T cell-mediated antitumor immunity to promote immune escape. Blockade of the IL-34-CD36 axis elicited antitumor immunity and synergized with anti-PD-1 immunotherapy, leading to a complete response. Our findings reveal the underlying mechanism of p53 modulation of the tumor immune microenvironment and provide a potential target for immunotherapy of cancer with p53 inactivation.

The E3 Ubiquitin Ligase FBXO38 Maintains the Antitumor Function of Natural Killer Cells by Sustaining IL15R Signaling.

Natural killer (NK) cells are the main innate antitumor effector cells but their function is often constrained in the tumor microenvironment. It has been reported that the E3 ligase FBXO38 accelerates PD-1 degradation in tumor-infiltrating T cells to unleash their cytotoxic function. In this study, we found that the transcriptional levels of FBXO38 in intratumoral NK cells of patients with cancer and tumor-bearing mice were significantly lower than in peritumoral NK cells. Conditional knockout of FBXO38 in NK cells accelerated tumor growth and increased tumor metastasis. FBXO38 deficiency resulted in impaired proliferation and survival of tumor-infiltrating NK (TINK) cells. Mechanistically, FBXO38 deficiency enhanced TGF-ß signaling, including elevating expression of Smad2 and Smad3, which suppressed expression of the transcription factor Eomes and further reduced expression of surface IL15Rß and IL15Rγc on NK cells. Consequently, FBXO38 deficiency led to TINK cell hyporesponsiveness to IL15. Consistent with these observations, FBXO38 mRNA expression was positively correlated with the proliferation of TINK cells in multiple human tumors. To study the therapeutic potential of FBXO38, mice bearing human tumors were treated with FBXO38 overexpressed human primary NK cells and showed a significant reduction in tumor size and prolonged survival. In conclusion, our results suggest that FBXO38 sustains NK-cell expansion and survival to promote antitumor immunity and have potential therapeutic implications as they suggest FBXO38 could be harnessed to enhance NK cell-based cancer immunotherapy.

Hepatocyte-derived FGL1 accelerates liver metastasis and tumor growth by inhibiting CD8+ T and NK cells.

Fibrinogen-like protein 1 (FGL1) contributes to the proliferation and metabolism of hepatocytes; however, as a major ligand of the immune checkpoint, its role in the liver regional immune microenvironment is poorly understood. Hepatocytes specifically and highly expressed FGL1 under normal physiological conditions. Increases in hepatic CD8+ T and NK cell numbers and functions were found in Fgl1-deficient (Fgl1-/-) mice, but not in the spleen or lymph node, similar to findings in anti-FGL1 mAb-treated wild-type mice. Furthermore, Fgl1 deficiency or anti-FGL1 mAb blockade restrained liver metastasis and slowed the growth of orthotopic tumors, with significantly prolonged survival of tumor-bearing mice. Tumor-infiltrating hepatic CD8+ T and NK cells upregulated the expression of lymphocyte activation gene-3 (LAG-3) and exhibited stronger antitumor activities after anti-FGL1 treatment. The antitumor efficacy of FGL1 blockade depended on cytotoxic T lymphocytes and NK cells, demonstrated by using a cell-deficient mouse model and cell transfer in vivo. In vitro, FGL1 directly inhibited hepatic T and NK cells related to the receptor LAG-3. In conclusion, hepatocyte-derived FGL1 played critical immunoregulatory roles in the liver and contributed to liver metastasis and tumor growth by inhibiting CD8+ T and NK cell functions via the receptor LAG-3, providing a new strategy for liver cancer immunotherapy.

Targeting myeloid-derived suppressor cells promotes antiparasitic T-cell immunity and enhances the efficacy of PD-1 blockade (15 words).

Immune exhaustion corresponds to a loss of effector function of T cells that associates with cancer or chronic infection. Here, our objective was to decipher the mechanisms involved in the immune suppression of myeloid-derived suppressor cells (MDSCs) and to explore the potential to target these cells for immunotherapy to enhance checkpoint blockade efficacy in a chronic parasite infection. We demonstrated that programmed cell-death-1 (PD-1) expression was significantly upregulated and associated with T-cell dysfunction in advanced alveolar echinococcosis (AE) patients and in Echinococcus multilocularis-infected mice. PD-1 blockade ex vivo failed to reverse AE patients' peripheral blood T-cell dysfunction. PD-1/PD-L1 blockade or PD-1 deficiency had no significant effects on metacestode in mouse model. This was due to the inhibitory capacities of immunosuppressive granulocytic MDSCs (G-MDSCs), especially in the liver surrounding the parasite pseudotumor. MDSCs suppressed T-cell function in vitro in an indoleamine 2, 3 dioxygenase 1 (IDO1)-dependent manner. Although depleting MDSCs alone restored T-cell effector functions and led to some limitation of disease progression in E. multilocularis-infected mice, combination with PD-1 blockade was better to induce antiparasitic efficacy. Our findings provide preclinical evidence in support of targeting MDSC or combining such an approach with checkpoint blockade in patients with advanced AE. (200 words).

Reconstituted CD74+ NK cells trigger chronic graft versus host disease after allogeneic bone marrow transplantation.

Chronic graft-versus-host disease (cGVHD) is the most common long-term complication after allogeneic hematopoietic stem cell transplantation (allo-HSCT). The patients with pulmonary cGVHD in particular have a very poor prognosis. NK cells are the first reconstituted lymphocyte subset after allo-HSCT; however, the impact of reconstituted NK cells on cGVHD is unclear. Here, we found allogeneic recipients showed obvious pulmonary cGVHD. Surprisingly, deletion of reconstituted NK cells resulted in maximal relief of pulmonary cGVHD. Mechanistically, reconstituted NK cells with donor profiles modulated the pulmonary inflammatory microenvironment to trigger cGVHD. Reconstituted NK cells secreted IFN-γ and TNF-α to induce CXCL10 production by epithelial cells, which recruited macrophages and CD4+ T cells to the lungs. Then macrophages and CD4+ T cells were activated by the inflammatory microenvironment, thereby mediating lung injury. Through assessment of differences in cellular energy, we found that CD74+ NK cells with high mitochondrial potential and pro-inflammatory activity triggered pulmonary cGVHD. Furthermore, targeted elimination of CD74+ NK cells using the anti-CD74 antibody significantly alleviated pulmonary cGVHD but preserved the CD74- NK cells to exert graft-versus-leukemia (GVL) effects. Data from human samples corroborated our findings in mouse models. Collectively, our results reveal that reconstituted CD74+ NK cells trigger pulmonary cGVHD and suggest that administration of CD74 antibody was a potential therapeutic for patients with cGVHD.

Pulmonary tumor thrombotic microangiopathy: Two case reports and literature review.

RATIONALE: Pulmonary tumor thrombotic microangiopathy (PTTM) is a rare but serious complication in patients with malignancy; its main manifestation includes acute pulmonary hypertension with severe respiratory distress. More than 200 cases have been reported since it was first identified in 1990. PTTM accounts for approximately 0.9% to 3.3% of deaths due to malignancy, but only a minority of patients are diagnosed ante-mortem, with most patients having a definitive diagnosis after autopsy. PATIENT CONCERNS: Two middle-aged women both died within a short period of time due to progressive dyspnea and severe pulmonary hypertension. DIAGNOSES: One patient was definitively confirmed as a gastrointestinal malignant tumor by liver puncture biopsy pathology. Ultimately, the clinical diagnosis was pulmonary tumor thrombotic microangiopathy. INTERVENTIONS: The patient was treated symptomatically with oxygen, diuresis, and anticoagulation, while a liver puncture was perfected to clarify the cause. OUTCOMES: Two cases of middle-aged female patients with rapidly progressive pulmonary hypertension and respiratory failure resulted in death with malignant neoplasm. LESSONS: PTTM has a rapid onset and a high morbidity and mortality rate. Our clinicians need to be more aware of the need for timely diagnosis through a targeted clinical approach, leading to more targeted treatment and a better prognosis.

Galectin-3-ITGB1 Signaling Mediates Interleukin 10 Production of Hepatic Conventional Natural Killer Cells in Hepatitis B Virus Transgenic Mice and Correlates with Hepatocellular Carcinoma Progression in Patients.

BACKGROUND AND AIMS: The outcomes of HBV infections are related to complex immune imbalances; however, the precise mechanisms by which HBV induces immune dysfunction are not well understood. METHODS: HBV transgenic (HBs-Tg) mice were used to investigate intrahepatic NK cells in two distinct subsets: conventional NK (cNK) and liver-resident NK (LrNK) cells during a chronic HBV infection. RESULTS: The cNK cells, but not the LrNK cells, were primarily responsible for the increase in the number of bulk NK cells in the livers of ageing HBs-Tg mice. The hepatic cNK cells showed a stronger ability to produce IL-10, coupled with a higher expression of CD69, TIGIT and PD-L1, and lower NKG2D expression in ageing HBs-Tg mice. A lower mitochondrial mass and membrane potential, and less polarized localization were observed in the hepatic cNK cells compared with the splenic cNK cells in the HBs-Tg mice. The enhanced galectin-3 (Gal-3) secreted from HBsAg+ hepatocytes accounted for the IL-10 production of hepatic cNK cells via ITGB1 signaling. For humans, LGALS3 and ITGB1 expression is positively correlated with IL-10 expression, and negatively correlated with the poor clinical progression of HCC. CONCLUSIONS: Gal-3-ITGB1 signaling shapes hepatic cNK cells but not LrNK cells during a chronic HBV infection, which may correlate with HCC progression.

Pyroptotic T cell-derived active IL-16 has a driving function in ovarian endometriosis development.

Endometriosis, affecting 6%-10% of women, often leads to pain and infertility and its underlying inflammatory mechanisms are poorly understood. We established endometriosis models in wild-type and IL16KO mice, revealing the driver function of IL-16 in initiating endometriosis-related inflammation. Using an in vitro system, we confirmed iron overload-induced GSDME-mediated pyroptosis as a key trigger for IL-16 activation and release. In addition, our research led to the development of Z30702029, a compound inhibiting GSDME-NTD-mediated pyroptosis, which shows promise as a therapeutic intervention for endometriosis. Importantly, our findings extend beyond endometriosis, highlighting GSDME-mediated pyroptosis as a broader pathway for IL-16 release and offering insights into potential treatments for various inflammatory conditions.

Blockade of CD300A enhances the ability of human NK cells to lyse hematologic malignancies.

OBJECTIVE: The human cluster of differentiation (CD)300A, a type-I transmembrane protein with immunoreceptor tyrosine-based inhibitory motifs, was investigated as a potential immune checkpoint for human natural killer (NK) cells targeting hematologic malignancies (HMs). METHODS: We implemented a stimulation system involving the CD300A ligand, phosphatidylserine (PS), exposed to the outer surface of malignant cells. Additionally, we utilized CD300A overexpression, a CD300A blocking system, and a xenotransplantation model to evaluate the impact of CD300A on NK cell efficacy against HMs in in vitro and in vivo settings. Furthermore, we explored the association between CD300A and HM progression in patients. RESULTS: Our findings indicated that PS hampers the function of NK cells. Increased CD300A expression inhibited HM lysis by NK cells. CD300A overexpression shortened the survival of HM-xenografted mice by impairing transplanted NK cells. Blocking PS-CD300A signals with antibodies significantly amplified the expression of lysis function-related proteins and effector cytokines in NK cells, thereby augmenting the ability to lyse HMs. Clinically, heightened CD300A expression correlated with shorter survival and an "exhausted" phenotype of intratumoral NK cells in patients with HMs or solid tumors. CONCLUSIONS: These results propose CD300A as a potential target for invigorating NK cell-based treatments against HMs.

Smart-sight: Video-based waste characterization for RDF-3 production.

A material recovery facility (MRF) can transform municipal solid waste (MSW) into a valued commodity called refuse-derived fuel (RDF) as a promising solution to waste-to-energy conversion. The quality of the produced RDF significantly relies on the composition of in-feed waste and waste characterization method applied for auditing purposes, a process that is both time-consuming and fraught with potential hazards. This study focuses to enhance the workflow of the waste characterization process at an MRF. A solution named Smart Sight is proposed to detect and classify waste based on videos recorded after processing MSW through a mechanical sorting line consisting of bag breakers and trommel screens. A comprehensive dataset is created encompassing thirteen mixed waste classes from single and multi-family streams. The dataset is preprocessed with motion compensation techniques and frame differencing methods to extract and refine valuable frames. A one-stage YOLO detector model is then trained over the dataset. The experimental results show that the proposed method works efficiently at detecting and classifying waste objects in indoor MRF environments. Accuracy, precision, recall, and F1 score related to the proposed solution are found to be 0.70, 0.762, 0.69 and 0.72, respectively, with a mAP@0.5 of 0.716. The proposed approach is validated using data collected from local MRF by comparing the estimated waste composition values of the proposed solution with laboratory results obtained through current standardized industrial practices. Comparison reveals that waste characterization estimation obtained is consistent with the laboratory results, inferring that Smart-Sight is a viable tool for estimating waste composition.

RORα is required for expansion and memory maintenance of ILC1s via a lymph node-liver axis.

Type 1 innate lymphoid cells (ILC1s) possess adaptive immune features, which confer antigen-specific memory responses against haptens and viruses. However, the transcriptional regulation of memory ILC1 responses is currently not known. We show that retinoic acid receptor-related orphan receptor alpha (RORα) has high expression in memory ILC1s in murine contact hypersensitivity (CHS) models. RORα deficiency diminishes ILC1-mediated CHS responses significantly but has no effect on memory T cell-mediated CHS responses. During sensitization, RORα promotes sensitized-ILC1 expansion by suppressing expression of cell-cycle repressors in draining lymph nodes. RORα programs gene-expression patterns related to cell survival and is required for the long-term maintenance of memory ILC1s in the liver. Our findings reveal RORα to be a key transcriptional factor for sensitized-ILC1 expansion and long-term maintenance of memory ILC1s.

Corrigendum: Bispecific NK-cell engager targeting BCMA elicits stronger antitumor effects and produces less proinflammatory cytokines than T-cell engager.

[This corrects the article DOI: 10.3389/fimmu.2023.1113303.].

A pan-cancer single-cell panorama of human natural killer cells.

Natural killer (NK) cells play indispensable roles in innate immune responses against tumor progression. To depict their phenotypic and functional diversities in the tumor microenvironment, we perform integrative single-cell RNA sequencing analyses on NK cells from 716 patients with cancer, covering 24 cancer types. We observed heterogeneity in NK cell composition in a tumor-type-specific manner. Notably, we have identified a group of tumor-associated NK cells that are enriched in tumors, show impaired anti-tumor functions, and are associated with unfavorable prognosis and resistance to immunotherapy. Specific myeloid cell subpopulations, in particular LAMP3+ dendritic cells, appear to mediate the regulation of NK cell anti-tumor immunity. Our study provides insights into NK-cell-based cancer immunity and highlights potential clinical utilities of NK cell subsets as therapeutic targets.

Gasdermin D licenses MHCII induction to maintain food tolerance in small intestine.

The intestinal epithelial cells (IECs) constitute the primary barrier between host cells and numerous foreign antigens; it is unclear how IECs induce the protective immunity against pathogens while maintaining the immune tolerance to food. Here, we found IECs accumulate a less recognized 13-kD N-terminal fragment of GSDMD that is cleaved by caspase-3/7 in response to dietary antigens. Unlike the 30-kD GSDMD cleavage fragment that executes pyroptosis, the IEC-accumulated GSDMD cleavage fragment translocates to the nucleus and induces the transcription of CIITA and MHCII molecules, which in turn induces the Tr1 cells in upper small intestine. Mice treated with a caspase-3/7 inhibitor, mice with GSDMD mutation resistant to caspase-3/7 cleavage, mice with MHCII deficiency in IECs, and mice with Tr1 deficiency all displayed a disrupted food tolerance phenotype. Our study supports that differential cleavage of GSDMD can be understood as a regulatory hub controlling immunity versus tolerance in the small intestine.

Bispecific NK-cell engager targeting BCMA elicits stronger antitumor effects and produces less proinflammatory cytokines than T-cell engager.

Bispecific antibodies have attracted more attention in recent years for the treatment of tumors, in which most of them target CD3, which mediates the killing of tumor cells by T cells. However, T-cell engager may cause serious side effects, including neurotoxicity and cytokine release syndrome. More safe treatments are still needed to address unmet medical needs, and NK cell-based immunotherapy is a safer and more effective way to treat tumors. Our study developed two IgG-like bispecific antibodies with the same configuration: BT1 (BCMA×CD3) attracted T cells and tumor cells, while BK1 (BCMA×CD16) attracted NK cells and tumor cells. Our study showed that BK1 mediated NK cell activation and upregulated the expression of CD69, CD107a, IFN-γ and TNF. In addition, BK1 elicited a stronger antitumor effect than BT1 both in vitro and in vivo. Combinatorial treatment (BK1+BT1) showed a stronger antitumor effect than either treatment alone, as indicated by in vitro experiments and in vivo murine models. More importantly, BK1 induced fewer proinflammatory cytokines than BT1 both in vitro and in vivo. Surprisingly, BK1 reduced cytokine production in the combinatorial treatment, suggesting the indispensable role of NK cells in the control of cytokine secretion by T cells. In conclusion, our study compared NK-cell engagers and T-cell engagers targeting BCMA. The results indicated that NK-cell engagers were more effective with less proinflammatory cytokine production. Furthermore, the use of NK-cell engagers in combinatorial treatment helped to reduce cytokine secretion by T cells, suggesting a bright future for NK-cell engagers in clinical settings.

Tumors evade immune cytotoxicity by altering the surface topology of NK cells.

The highly variable response rates to immunotherapies underscore our limited knowledge about how tumors can manipulate immune cells. Here the membrane topology of natural killer (NK) cells from patients with liver cancer showed that intratumoral NK cells have fewer membrane protrusions compared with liver NK cells outside tumors and with peripheral NK cells. Dysregulation of these protrusions prevented intratumoral NK cells from recognizing tumor cells, from forming lytic immunological synapses and from killing tumor cells. The membranes of intratumoral NK cells have altered sphingomyelin (SM) content and dysregulated serine metabolism in tumors contributed to the decrease in SM levels of intratumoral NK cells. Inhibition of SM biosynthesis in peripheral NK cells phenocopied the disrupted membrane topology and cytotoxicity of the intratumoral NK cells. Targeting sphingomyelinase confers powerful antitumor efficacy, both as a monotherapy and as a combination therapy with checkpoint blockade.