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Dysregulations of epithelial-immune interactions frequently culminate in chronic inflammatory diseases of the skin, lungs, kidneys, and gastrointestinal tract. Yet, the intraepithelial processes that initiate and perpetuate inflammation in these organs [...]

Introduction End-stage liver disease is the leading cause of mortality in digestive diseases, with liver transplantation as the sole curative intervention (1). The scarcity of available donor livers has led [...], Steatotic donor livers are becoming more and more common in liver transplantation. However, the current use of steatotic grafts is less acceptable than normal grafts due to their higher susceptibility to ischemia/reperfusion (I/R) injury. To investigate the mechanism underlying the susceptibility of steatotic liver to I/R injury, we detected cell death markers and inflammation in clinical donor livers and animal models. We found that caspase-8-mediated hepatic apoptosis is activated in steatotic liver I/R injury. However, ablation of caspase-8 only slightly mitigated steatotic liver I/R injury without affecting inflammation. We further demonstrated that RIPK1 kinase induces both caspase-8-mediated apoptosis and cell deathindependent inflammation. Inhibition of RIPK1 kinase significantly protects against steatotic liver I/R injury by alleviating both hepatic apoptosis and inflammation. Additionally, we found that RIPK1 activation is induced by Z-DNA binding protein 1 (ZBP1) but not the canonical TNF-a pathway during steatotic liver I/R injury. Deletion of ZBP1 substantially decreases the steatotic liver I/R injury. Mechanistically, ZBP1 is amplified by palmitic acid-activated JNK pathway in steatotic livers. Upon I/R injury, excessive reactive oxygen species trigger ZBP1 activation by inducing its aggregation independent of the Z- nucleic acids sensing action in steatotic livers, leading to the kinase activation of RIPK1 and the subsequent aggravation of liver injury. Thus, ZBP1-mediated RIPK1-driven apoptosis and inflammation exacerbate steatotic liver I/R injury, which could be targeted to protect steatotic donor livers during transplantation.

Introduction Targeting of the immune system has provided clinical benefit for individuals with advanced solid tumors, including non-small cell lung cancer (NSCLC) (1). Currently approved immune-checkpoint inhibitors (ICIs) include monoclonal [...], BACKGROUND. Precise stratification of patients with non-small cell lung cancer (NSCLC) is needed for appropriate application of PD-1/PD-L1 blockade therapy. METHODS. We measured soluble forms of the immune-checkpoint molecules PD-L1, PD-1, and CTLA-4 in plasma of patients with advanced NSCLC before PD-1/PD-L1 blockade. A prospective biomarker-finding trial (cohort A) included 50 previously treated patients who received nivolumab. A retrospective observational study was performed for patients treated with any PD-1/PD-L1 blockade therapy (cohorts B and C), cytotoxic chemotherapy (cohort D), or targeted therapy (cohort E). Plasma samples from all patients were assayed for soluble immune-checkpoint molecules with a highly sensitive chemiluminescence-based assay. RESULTS. Nonresponsiveness to PD-1/PD-L1 blockade therapy was associated with higher concentrations of these soluble immune factors among patients with immune-reactive (hot) tumors. Such an association was not apparent for patients treated with cytotoxic chemotherapy or targeted therapy. Integrative analysis of tumor size, PD-L1 expression in tumor tissue (tPD-L1), and gene expression in tumor tissue and peripheral [CD8.sup.+] T cells revealed that high concentrations of the 3 soluble immune factors were associated with hyper or terminal exhaustion of antitumor immunity. The combination of soluble PD-L1 (sPD-L1) and sCTLA-4 efficiently discriminated responsiveness to PD-1/PD-L1 blockade among patients with immune-reactive tumors. CONCLUSION. Combinations of soluble immune factors might be able to identify patients unlikely to respond to PD- 1/PD-L1 blockade as a result of terminal exhaustion of antitumor immunity. Our data suggest that such a combination better predicts, along with tPD-L1, for the response of patients with NSCLC. TRIAL REGISTRATION. UMIN000019674. FUNDING. This study was funded by Ono Pharmaceutical Co. Ltd. and Sysmex Corporation.

Introduction Upon infection, host cells become virus production factories, leading to accumulation of large quantities of viral proteins. When viral protein accumulation overwhelms cellular degradative capabilities, toxic protein aggregates form, [...], Virophagy, the selective autophagosomal engulfment and lysosomal degradation of viral components, is crucial for neuronal cell survival and antiviral immunity. However, the mechanisms leading to viral antigen recognition and capture by autophagic machinery remain poorly understood. Here, we identified cyclin- dependent kinase-like 5 (CDKL5), known to function in neurodevelopment, as an essential regulator of virophagy. Loss-of- function mutations in CDKL5 are associated with a severe neurodevelopmental encephalopathy. We found that deletion of CDKL5 or expression of a clinically relevant pathogenic mutant of CDKL5 reduced virophagy of Sindbis virus (SINV), a neurotropic RNA virus, and increased intracellular accumulation of SINV capsid protein aggregates and cellular cytotoxicity. Cdkl5- knockout mice displayed increased viral antigen accumulation and neuronal cell death after SINV infection and enhanced lethality after infection with several neurotropic viruses. Mechanistic studies demonstrated that CDKL5 directly binds the canonical selective autophagy receptor p62 and phosphorylates p62 at T269/S272 to promote its interaction with viral capsid aggregates. We found that CDKL5-mediated phosphorylation of p62 facilitated the formation of large p62 inclusion bodies that captured viral capsids to initiate capsid targeting to autophagic machinery. Overall, these findings identify a cell-autonomous innate immune mechanism for autophagy activation to clear intracellular toxic viral protein aggregates during infection.

Radiation's impact on the tumor and immune microenvironment Radiation therapy (RT) remains one of the most effective oncologic treatments available, whether used in a primary, adjunctive, or palliative fashion. Its [...], Radiation therapy (RT) remains one of the most effective and utilized oncologic treatments available. While it can directly yield tumor cell death, its impact on the immune microenvironment is more complex, promoting either an antitumor response or, conversely, a tumor-promoting state. TGF-[beta], induced by RT, yields a more immunosuppressive environment, including potentially blunting response to immune-checkpoint blockade. In this issue of the JCI, Wang and colleagues demonstrate that RT reduced expression of bone morphogenetic protein and activin membrane-bound inhibitor (BAMBI), a TGF-[beta] pseudoreceptor. Limiting this effect, or increasing BAMBI, improved RT-induced tumor cell killing, tumor response, and antitumor immune effects. This realization points to a pathway of potential clinical translation.

Podocyte detachment and cell death Podocytes are postmitotic visceral epithelial cells that cover the glomerular basement membrane. Collectively with the glomerular endothelial cells and basement membrane, podocytes constitute the kidney [...], Profilin1 belongs to a family of small monomeric actin-binding proteins with diverse roles in fundamental actin-dependent cellular processes required for cell survival. Podocytes are postmitotic visceral epithelial cells critical for the structure and function of the kidney filtration barrier. There is emerging evidence that the actin-related mode of cell death known as mitotic catastrophe is an important pathway involved in podocyte loss. In this issue of the JCI, Tian, Pedigo, and colleagues demonstrate that profilinl deficiency in podocytes triggered cell cycle reentry, resulting in abortive cytokinesis with a loss in ribosomal RNA processing that leads to podocyte loss and glomerulosclerosis. This study demonstrates the essential role of actin dynamics in mediating this fundamental mode of podocyte cell death.

Introduction Cells burn sugars and fatty acids to harvest free energy for living, which, however, inevitably results in the production of reactive oxygen species (ROS), the harmful derivatives of molecular [...], Elevation of reactive oxygen species (ROS) levels is a general consequence of tumor cells' response to treatment and may cause tumor cell death. Mechanisms by which tumor cells clear fatal ROS, thereby rescuing redox balance and entering a chemoresistant state, remain unclear. Here, we show that cysteine sulfenylation by ROS confers on aryl hydrocarbon receptor (AHR) the ability to dissociate from the heat shock protein 90 complex but to bind to the PPP1R3 family member PPP1R3C of the glycogen complex in drug-treated tumor cells, thus activating glycogen phosphorylase to initiate glycogenolysis and the subsequent pentose phosphate pathway, leading to NADPH production for ROS clearance and chemoresistance formation. We found that basic ROS levels were higher in chemoresistant cells than in chemosensitive cells, guaranteeing the rapid induction of AHR sulfenylation for the clearance of excess ROS. These findings reveal that AHR can act as an ROS sensor to mediate chemoresistance, thus providing a potential strategy to reverse chemoresistance in patients with cancer.

Lysosomal inhibition as a therapeutic approach for cancer Lysosomes are membrane-bound vesicles containing hydrolytic enzymes that degrade diverse cellular contents, including proteins, carbohydrates, and lipids (1). In tumors, there is [...], Cancer cells rely on lysosome-dependent degradation to recycle nutrients that serve their energetic and biosynthetic needs. Despite great interest in repurposing the antimalarial hydroxychloroquine as a lysosomal inhibitor in clinical oncology trials, the mechanisms by which hydroxychloroquine and other lysosomal inhibitors induce tumor-cell cytotoxicity remain unclear. In this issue of theJCI, Bhardwaj et al. demonstrate that DC661, a dimeric form of chloroquine that inhibits palmitoyl-protein thioesterase 1 (PPT1), promoted lysosomal lipid peroxidation, resulting in lysosomal membrane permeabilization and tumor cell death. Remarkably, this lysosomal cell death pathway elicited cell-intrinsic immunogenicity and promoted T lymphocyte-mediated tumor cell clearance. The findings provide the mechanistic foundation for the potential combined use of immunotherapy and lysosomal inhibition in clinical trials.

Introduction Primary cilia are sensory organelles that protrude from the cell surface to detect extracellular signals that regulate cellular physiology. Cilia are highly dynamic microtubule-based organelles that are nucleated from [...], Defects in primary or motile cilia result in a variety of human pathologies, and retinal degeneration is frequently associated with these so-called ciliopathies. We found that homozygosity for a truncating variant in CEP162, a centrosome and microtubule-associated protein required for transition zone assembly during ciliogenesis and neuronal differentiation in the retina, caused late-onset retinitis pigmentosa in 2 unrelated families. The mutant CEP162E646R*5 protein was expressed and properly localized to the mitotic spindle, but it was missing from the basal body in primary and photoreceptor cilia. This impaired recruitment of transition zone components to the basal body and corresponded to complete loss of CEP162 function at the ciliary compartment, reflected by delayed formation of dysmorphic cilia. In contrast, shRNA knockdown of Cep162 in the developing mouse retina increased cell death, which was rescued by expression of CEP162-E646R*5, indicating that the mutant retains its role for retinal neurogenesis. Human retinal degeneration thus resulted from specific loss of the ciliary function of CEP162.

Introduction Nonspecific activation of memory [CD8.sup.+] T cells is a common feature of acute and chronic human viral diseases (1-7). Bystander-activated [CD8.sup.+] T cells respond in a TCR-independent manner and [...], Accumulation of activated immune cells results in nonspecific hepatocyte killing in chronic hepatitis B (CHB), leading to fibrosis and cirrhosis. This study aims to understand the underlying mechanisms in humans and to define whether these are driven by widespread activation or a subpopulation of immune cells. We enrolled CHB patients with active liver damage to receive antiviral therapy and performed longitudinal liver sampling using fine-needle aspiration to investigate mechanisms of CHB pathogenesis in the human liver. Single-cell sequencing of total liver cells revealed a distinct liver-resident, polyclonal [CD8.sup.+] T cell population that was enriched at baseline and displayed a highly activated immune signature during liver damage. Cytokine combinations, identified by in silico prediction of ligand-receptor interaction, induced the activated phenotype in healthy liver [CD8.sup.+] T cells, resulting in nonspecific Fas ligand-mediated killing of target cells. These results define a [CD8.sup.+] T cell population in the human liver that can drive pathogenesis and a key pathway involved in their function in CHB patients.

How kidney aging relates to podocytes and PD-1 According to the latest data from the WHO, while our life expectancy continues to increase in the past decade, healthy-adjusted life expectancy [...], Understanding the loss of kidney function resulting from kidney aging has become an emerging research focus that will facilitate the future development of antisenolytic treatments. In this issue of the JCI, Pippin et al. first identified PD-1 upregulation in the aged mouse podocyte via unbiased RNA-seq analysis. Overexpression of PD-1 in immortalized mouse podocytes induced cell death and a senescence-associated secretory phenotype, suggesting the pathological role of PD-1 upregulation in aged podocytes. Blocking PD-1 signaling via a neutralizing anti-PD-1 antibody reversed the aged phenotype in the aged mice and ameliorated proteinuria in an experimental focal segmental glomerulosclerosis (FSGS) mouse model. These findings highlight the role of PD-1 signaling in kidney aging and its therapeutic potential for human clinical trials.

Introduction The actin cytoskeleton coordinates a wide variety of cell functions ranging from cell division and differentiation to migration. The ability to reorganize the actin network in response to both [...], CD8 cytotoxic T lymphocytes (CTLs) rely on rapid reorganization of the branched F-actin network to drive the polarized secretion of lytic granules, initiating target cell death during the adaptive immune response. Branched F-actin is generated by the nucleation factor actin-related protein 2/3 (Arp2/3) complex. Patients with mutations in the actin-related protein complex 1B (ARPC1B) subunit of Arp2/3 show combined immunodeficiency, with symptoms of immune dysregulation, including recurrent viral infections and reduced [CD8.sup.+] T cell count. Here, we show that loss of ARPC1B led to loss of CTL cytotoxicity, with the defect arising at 2 different levels. First, ARPC1B is required for lamellipodia formation, cell migration, and actin reorganization across the immune synapse. Second, we found that ARPC1B is indispensable for the maintenance of TCR, CD8, and GLUT1 membrane proteins at the plasma membrane of CTLs, as recycling via the retromer and WASH complexes was impaired in the absence of ARPC1B. Loss of TCR, CD8, and GLUT1 gave rise to defects in T cell signaling and proliferation upon antigen stimulation of ARPC1B-deficient CTLs, leading to a progressive loss of [CD8.sup.+] T cells. This triggered an activation-induced immunodeficiency of CTL activity in ARPC1B-deficient patients, which could explain the susceptibility to severe and prolonged viral infections.

Introduction Worldwide, more than 250 million humans suffer from chronic hepatitis B (CHB), which results in an estimated 880,000 deaths per year caused by secondary complications like liver cirrhosis or [...], T cell therapy is a promising means to treat chronic hepatitis B virus (HBV) infection and HBV-associated hepatocellular carcinoma. T cells engineered to express an HBV-specific T cell receptor (TCR) may cure an HBV infection upon adoptive transfer. We investigated the therapeutic potential and safety of T cells stably expressing high-affinity HBV envelope- or core-specific TCRs recognizing European and Asian HLA-A2 subtypes. Both CD8* and [CD4.SUP.+] T cells from healthy donors and patients with chronic hepatitis B became polyfunctional effector cells when grafted with HBV-specific TCRs and eliminated HBV from infected HepG2-NTCP cell cultures. A single transfer of TCR-grafted T cells into HBV-infected, humanized mice controlled HBV infection, and virological markers declined by 4 to 5 log or below the detection limit. Engineered T cells specifically cleared infected hepatocytes without damaging noninfected cells when, as in a typical clinical setting, only a minority of hepatocytes were infected. Cell death was compensated by hepatocyte proliferation, and alanine amino transferase levels peaking between days 5 and 7 normalized again thereafter. Cotreatment with the entry inhibitor myrcludex B ensured long-term control of HBV infection. Thus, T cells stably transduced with highly functional TCRs have the potential to mediate clearance of HBV-infected cells, causing limited liver injury.

Introduction Ferroptosis is a cell death program that is executed via activation of a selective enzymatic process of lipid peroxidation (LPO) catalyzed by 15-lipoxygenases (15LOXes), dioxygenases for polyunsaturated fatty acids [...], Ferroptosis is a death program executed via selective oxidation of arachidonic acid-phosphatidylethanolamines (AA-PE) by 15-lipoxygenases. In mammalian cells and tissues, ferroptosis has been pathogenically associated with brain, kidney, and liver injury/diseases. We discovered that a prokaryotic bacterium, Pseudomonas aeruginosa, that does not contain AA-PE can express lipoxygenase (pLoxA), oxidize host AA-PE to 15-hydroperoxy-AA- PE (15-HOO-AA-PE), and trigger ferroptosis in human bronchial epithelial cells. Induction of ferroptosis by clinical P. aeruginosa isolates from patients with persistent lower respiratory tract infections was dependent on the level and enzymatic activity of pLoxA. Redox phospholipidomics revealed elevated levels of oxidized AA-PE in airway tissues from patients with cystic fibrosis (CF) but not with emphysema or CF without P. aeruginosa. We believe that the evolutionarily conserved mechanism of pLoxA-driven ferroptosis may represent a potential therapeutic target against P. aeruginosa- associated diseases such as CF and persistent lower respiratory tract infections.

Introduction Systemic lupus erythematosus (SLE) is an autoimmune syndrome of unclear etiology that predominantly affects women and disproportionately afflicts minorities (1). Lupus derives its name from the Latin word for [...], The etiopathogenesis of systemic lupus erythematosus (SLE), a clinically heterogeneous multisystemic syndrome that derives its name from the initial characterization of facial lesions that resemble the bite of a wolf, is considered a complex, multifactorial interplay between underlying genetic susceptibility factors and the environment. Prominent pathogenic factors include the induction of aberrant cell death pathways coupled with defective cell death clearance mechanisms that promote excessive externalization of modified cellular and nuclear debris with subsequent loss of tolerance to a wide variety of autoantigens and innate and adaptive immune dysregulation. While abnormalities in adaptive immunity are well recognized and are key to the pathogenesis of SLE, recent findings have emphasized fundamental roles of the innate immune system in the initiation and propagation of autoimmunity and the development of organ damage in this disease. This Review focuses on recent discoveries regarding the role of components of the innate immune system, specifically neutrophils and interferons, in promoting various aspects of lupus pathogenesis, with potential implications for novel therapeutic strategies.

Introduction Aminoglycosides are a widely used and successful class of antibiotics (1, 2). Despite their potent antimicrobial efficiency, all aminoglycoside antibiotics currently approved for use by the FDA are toxic [...], Exposure to aminoglycoside antibiotics can lead to the generation of toxic levels of reactive oxygen species (ROS) within mechanosensory hair cells of the inner ear that have been implicated in hearing and balance disorders. Better understanding of the origin of aminoglycoside-induced ROS could focus the development of therapies aimed at preventing this event. In this work, we used the zebrafish lateral line system to monitor the dynamic behavior of mitochondrial and cytoplasmic oxidation occurring within the same dying hair cell following exposure to aminoglycosides. The increased oxidation observed in both mitochondria and cytoplasm of dying hair cells was highly correlated with mitochondrial calcium uptake. Application of the mitochondrial uniporter inhibitor Ru360 reduced mitochondrial and cytoplasmic oxidation, suggesting that mitochondrial calcium drives ROS generation during aminoglycoside-induced hair cell death. Furthermore, targeting mitochondria with free radical scavengers conferred superior protection against aminoglycoside exposure compared with identical, untargeted scavengers. Our findings suggest that targeted therapies aimed at preventing mitochondrial oxidation have therapeutic potential to ameliorate the toxic effects of aminoglycoside exposure.

Introduction Chimeric antigen receptors (CARs) are synthetic receptors that retarget T cells to tumor surface antigens (1, 2). First-generation receptors typically link an antibody-derived tumor-binding element to either CD3θ or [...], Following immune attack, solid tumors upregulate coinhibitory ligands that bind to inhibitory receptors on T cells. This adaptive resistance compromises the efficacy of chimeric antigen receptor (CAR) T cell therapies, which redirect T cells to solid tumors. Here, we investigated whether programmed death-1-mediated (PD-1-mediated) T cell exhaustion affects mesothelin- targeted CAR T cells and explored cell-intrinsic strategies to overcome inhibition of CAR T cells. Using an orthotopic mouse model of pleural mesothelioma, we determined that relatively high doses of both CD28- and 4-1BB-based second-generation CAR T cells achieved tumor eradication. CAR-mediated CD28 and 4-1BB costimulation resulted in similar levels of T cell persistence in animals treated with low T cell doses; however, PD-1 upregulation within the tumor microenvironment inhibited T cell function. At lower doses, 4-1BB CAR T cells retained their cytotoxic and cytokine secretion functions longer than CD28 CAR T cells. The prolonged function of 4-1BB CAR T cells correlated with improved survival. PD-1/PD-1 ligand [PD-L1] pathway interference, through PD-1 antibody checkpoint blockade, cell-intrinsic PD-1 shRNA blockade, or a PD-1 dominant negative receptor, restored the effector function of CD28 CAR T cells. These findings provide mechanistic insights into human CAR T cell exhaustion in solid tumors and suggest that PD-1/PD-L1 blockade may be an effective strategy for improving the potency of CAR T cell therapies.