Nicotine restores olfactory function by activation of prok2R/Akt/FoxO3a axis in Parkinson's disease.

BACKGROUND: Olfactory dysfunction occurs frequently in Parkinson's disease (PD). In this study, we aimed to explore the potential biomarkers and underlying molecular pathways of nicotine for the treatment of olfactory dysfunction in 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-induced PD mice. METHODS: MPTP was introduced into C57BL/6 male mice to generate a PD model. Regarding in vivo experiments, we performed behavioral tests to estimate the protective effects of nicotine in MPTP-induced PD mice. RNA sequencing and traditional molecular methods were used to identify molecules, pathways, and biological processes in the olfactory bulb of PD mouse models. Then, in vitro experiments were conducted to evaluate whether nicotine can activate the prok2R/Akt/FoxO3a signaling pathway in both HEK293T cell lines and primary olfactory neurons treated with 1-methyl-4-phenylpyridinium (MPP+). Next, prok2R overexpression (prok2R+) and knockdown (prok2R-) were introduced with lentivirus, and the Akt/FoxO3a signaling pathway was further explored. Finally, the damaging effects of MPP+ were evaluated in prok2R overexpression (prok2R+) HEK293T cell lines. RESULTS: Nicotine intervention significantly alleviated olfactory and motor dysfunctions in mice with PD. The prok2R/Akt/FoxO3a signaling pathway was activated after nicotine treatment. Consequently, apoptosis of olfactory sensory neurons was significantly reduced. Furthermore, prok2R+ and prok2R- HEK293T cell lines exhibited upregulation and downregulation of the Akt/FoxO3a signaling pathway, respectively. Additionally, prok2R+ HEK293T cells were resistant to MPP+-induced apoptosis. CONCLUSIONS: This study showed the effectiveness and underlying mechanisms of nicotine in improving hyposmia in PD mice. These improvements were correlated with reduced apoptosis of olfactory sensory neurons via activated prok2R/Akt/FoxO3a axis. These results explained the potential protective functions of nicotine in PD patients.

Facts and Hopes on Chimeric Cytokine Agents for Cancer Immunotherapy.

Cytokines are key mediators of immune responses that can modulate the antitumor activity of immune cells. Cytokines have been explored as a promising cancer immunotherapy. However, there are several challenges to cytokine therapy, especially a lack of tumor targeting, resulting in high toxicity and limited efficacy. To overcome these limitations, novel approaches have been developed to engineer cytokines with improved properties, such as chimeric cytokines. Chimeric cytokines are fusion proteins that combine different cytokine domains or link cytokines to antibodies (immunocytokines) or other molecules that can target specific receptors or cells. Chimeric cytokines can enhance the selectivity and stability of cytokines, leading to reduced toxicity and improved efficacy. In this review, we focus on two promising cytokines, IL2 and IL15, and summarize the current advances and challenges of chimeric cytokine design and application for cancer immunotherapy. Most of the current approaches focus on increasing the potency of cytokines, but another important goal is to reduce toxicity. Cytokine engineering is promising for cancer immunotherapy as it can enhance tumor targeting while minimizing adverse effects.

Bacteria-derived nanovesicles enhance tumour vaccination by trained immunity.

Trained immunity enhances the responsiveness of immune cells to subsequent infections or vaccinations. Here we demonstrate that pre-vaccination with bacteria-derived outer-membrane vesicles, which contain large amounts of pathogen-associated molecular patterns, can be used to potentiate, and enhance, tumour vaccination by trained immunity. Intraperitoneal administration of these outer-membrane vesicles to mice activates inflammasome signalling pathways and induces interleukin-1ß secretion. The elevated interleukin-1ß increases the generation of antigen-presenting cell progenitors. This results in increased immune response when tumour antigens are delivered, and increases tumour-antigen-specific T-cell activation. This trained immunity increased protection from tumour challenge in two distinct cancer models.

Seawater Corrosion Resistance of Duplex Stainless Steel and the Axial Compressive Stiffness of Its Reinforced Concrete Columns.

In order to explore the corrosion resistance of duplex stainless steel under seawater corrosion and the compressive stiffness of its reinforced concrete columns, this study first performed seawater corrosion resistance tests on HRB400 ordinary steel rebar and S32205 duplex stainless steel rebar. The effect of the corrosion product film on the corrosion behavior was investigated through polarization curve tests and electrochemical impedance spectroscopy tests. The results showed that the corrosion rate of S32205 duplex stainless steel in a seawater environment was approximately 1/15 that of the HRB400 ordinary steel rebar. The anodic polarization curve of duplex stainless steel rebars exhibited a greater slope than that of carbon steel rebars. In the simulated seawater environment, the corrosion rate of these two kinds of steel bars showed different trends. The corrosion rate of ordinary steel bar HRB400 first decreased and then increased, while that of duplex stainless steel S2205 increased steadily. Furthermore, 18 short concrete columns reinforced with ordinary and duplex stainless steel rebars were subjected to the axial compression test and stiffness analysis; the stiffness of the short columns was calculated from the test data. The theoretical values agreed with the test values, with a stiffness calculation error of less than 5%.

Clinical and preclinical evaluation of miR-144-5p as a key target for major depressive disorder.

BACKGROUND: Neuronal abnormalities are closely associated with major depressive disorder (MDD). Available evidence suggests a role for microRNAs (miRNAs) in regulating the expression of genes involved in MDD. Hence, miRNAs that can be potential therapeutic targets need to be identified. METHODS: A mouse model of chronic unpredictable stress (CUS) was used to evaluate the function of miRNAs in MDD. miR-144-5p was screened from the hippocampi of CUS mice based on sequencing results. Adenovirus-associated vectors were used to overexpress or knockdown miR-144-5p in mice. BpV(pic) and LY294002 were used to determine the relationship between miR-144-5p target genes PTEN and TLR4 in neuronal impairment caused by miR-144-5p deficiency. Western blotting, immunofluorescence, ELISA immunosorbent assay, and Golgi staining were used to detect neuronal abnormalities. Serum samples from healthy individuals and patients with MDD were used to detect miR-144-5p levels in the serum and serum exosomes using qRT-PCR. RESULTS: miR-144-5p expression was significantly decreased within the hippocampal dentate gyrus (DG) of CUS mice. Upregulation of miR-144-5p in the DG ameliorated depression-like behavior in CUS mice and attenuated neuronal abnormalities by directly targeting PTEN and TLR4 expression. Furthermore, miR-144-5p knockdown in normal mice led to depression-like behavior via inducing neuronal abnormalities, including abnormal neurogenesis, neuronal apoptosis, altered synaptic plasticity, and neuroinflammation. miR-144-5p deficiency-mediated neuronal impairment was mediated by PI3K/Akt/FoxO1 signaling. Furthermore, miR-144-5p levels were downregulated in the sera of patients with MDD and associated with depressive symptoms. Consistently, serum exosome-derived miR-144-5p levels were decreased in patients with MDD. CONCLUSION: miR-144-5p plays a vital role in regulating neuronal abnormalities in depression. Our findings provide translational evidence that miR-144-5p is a new potential therapeutic target for MDD.

GBP2 promotes clear cell renal cell carcinoma progression through immune infiltration and regulation of PD­L1 expression via STAT1 signaling.

Guanylate­binding protein 2 (GBP2) has been widely studied in cancer, however, its potential role in clear cell renal cell carcinoma (ccRCC) is not fully elucidated. The present study aimed to explore the effect of GBP2 on tumor progression and its possible underlying molecular mechanisms in ccRCC. The Cancer Genome Atlas, Gene Expression Omnibus, Cancer Cell Line Encyclopedia databases, and several bioinformatics analysis tools, such as Gene Expression Profiling Interactive Analysis 2, Kaplan­Meier plotter, UALCAN, LinkedOmics, Metascape, GeneMANIA and Tumor Immune Estimation Resource, were used to characterize the functional relationship between GBP2 and ccRCC. Focusing on the association between GBP2 and programmed death ligand 1 (PD­L1) in vitro, the regulatory mechanism was investigated by knockdown and overexpression of GBP2 in Caki­1 and 786­O cells using reverse transcription­quantitative PCR, western blotting and co­immunoprecipitation techniques. The results indicated that GBP2 was commonly upregulated in ccRCC, correlating with worse prognosis. In addition, GBP2 expression levels were positively associated with different patterns of immune cell infiltration, suggesting that the GBP2 gene regulates PD­L1 expression via the signal transducer and activator of transcription 1 (STAT1) pathway. The present study suggested that GBP2 regulates tumor immune infiltration and promotes tumor immune escape through PD­L1 expression, revealing a potential immunotherapeutic target for ccRCC.

IL-2 delivery by engineered mesenchymal stem cells re-invigorates CD8+ T cells to overcome immunotherapy resistance in cancer.

Immune checkpoint blockade (ICB)-based immunotherapy depends on functional tumour-infiltrating lymphocytes (TILs), but essential cytokines are less understood. Here we uncover an essential role of endogenous IL-2 for ICB responsiveness and the correlation between insufficient IL-2 signalling and T-cell exhaustion as tumours progress. To determine if exogenous IL-2 in the tumour microenvironment can overcome ICB resistance, we engineered mesenchymal stem cells (MSCs) to successfully deliver IL-2 mutein dimer (SIL2-EMSC) to TILs. While MSCs have been used to suppress inflammation, SIL2-EMSCs elicit anti-tumour immunity and overcome ICB resistance without toxicity. Mechanistically, SIL2-EMSCs activate and expand pre-existing CD8+ TILs, sufficient for tumour control and induction of systemic anti-tumour effects. Furthermore, engineered MSCs create synergy of innate and adaptive immunity. The therapeutic benefits of SIL2-EMSCs were also observed in humanized mouse models. Overall, engineered MSCs rejuvenate CD8+ TILs and thus potentiate ICB and chemotherapy.

MCPIP1 alleviated alcohol-induced immune dysfunction via the MAPK/ERK signaling pathway.

RATIONALE: In recent years, monocyte chemotactic protein-induced protein 1 (MCPIP1) has been reported to control inflammation via IL-10. OBJECTIVES: The aims of this study were to determine (1) whether MCPIP1 can repair damage to the immune system after alcohol use and (2) whether MCPIP1 can repair the immune function impaired by alcohol use through the MAPK/ERK signaling pathway. Our results will inform the treatment of immune dysfunction caused by alcohol consumption. METHODS: Scrambled shRNA or MCPIP-1-shRNA carried by the lentiviral vector (50µl each at 1×108TU/ml) was injected retrogradely through the pancreatic duct to pretreat male C57BL/6 mice. Five days after the injection, mice were exposed to intragastric ethanol infusion (5g/kg, 25% ethanol w/v) daily or vehicle for 10 days. RESULTS: MCPIP-1 protein was increased in the pancreas after alcohol exposure. MCPIP-1 shRNA specifically decreased MCPIP-1 protein expression and mRNA level in the pancreas. Specific knockdown of MCPIP-1 exacerbates pancreatic necrosis, interstitial edema, and inflammatory infiltrates after alcohol exposure. Meanwhile, specific knockdown of MCPIP-1 also increased pancreatic pro-inflammatory cytokine (IL-6 and IL-1ß), chemokine MCP-1, and chemokine receptor 2 (CCR2) after alcohol exposure. What's more, p-JNK and p-ERK in the pancreas were all similarly increased in response to pancreas-specific knockdown of MCPIP-1 during alcohol exposure. CONCLUSIONS: Taken together, the results above suggested that MCPIP1 repairs the immune function impaired by alcohol use via stimulating JNK and ERK pathways. Our results will inform the treatment of immune dysfunction caused by alcohol consumption.

Chemokine (C-C motif) receptor 2 is associated with the pathological grade and inflammatory response in IgAN children.

BACKGROUND: Chemokine (C-C motif) receptor 2 (CCR2) is involved in important physiological and pathological processes, such as inflammation and autoimmune diseases. Abnormal immune and inflammatory responses play a critical role in the development and progression of IgA nephritis (IgAN). However, the role of CCR2 in IgAN is unknown. METHODS: Fifteen IgAN children who were diagnosed by kidney biopsy provided kidney biopsy tissue, blood and urine samples, and age-matched healthy control subjects (blood donators n = 12; tissue donators n = 8) were included. Immunohistochemical analysis was used to detect the expression of CCR2, MCP-1, IL-6, IL-17, and TNF-α in the kidney tissues. Relative optical density (OD) was calculated by Image J software, and the correlation between CCR2 expression and pathological grade in IgAN children was analyzed. RESULTS: The expression of CCR2 significantly increased in mesangial cells of children with IgAN compared to that in control group (P < 0.001), especially in IgAN patients with Lee's grade III to IV (P < 0.001). Interestingly, CCR2 expression was positively correlated with Lee's grade (r = 0.9152, P = 0.0001) in IgAN children. The expression levels of inflammatory factors were markedly increased in IgAN children, and importantly CCR2 expression was positively correlated with it's expression level. CONCLUSIONS: The results suggest that CCR2 signaling might be involved in pathological process and inflammatory responses of children IgAN, and could potentially be an intervention target in children IgAN.

MCPIP1 promotes cell proliferation, migration and angiogenesis of glioma via VEGFA-mediated ERK pathway.

Glioma is the most common primary malignant intracranial tumor in the population, and is often associated with abundant angiogenesis. However, how angiogenesis is regulated during glioma progression is still poorly understood. Data mining of cancer patient database shows that MCPIP1 is positively correlated with VEGFA expression and negatively with survival. In this study, we report that overexpressed MCPIP1 in glioma cells is a boost of angiogenesis. Mechanistically, MCPIP1 upregulates the expression of VEGFA in glioma, and promote the secretion of VEGFA to the surroundings, which could stimulate angiogenesis through ERK pathway. Blocking VEGFA expression and secretion inhibited MCPIP1-mediated angiogenesis and glioma progression in vitro and xenograft models. Collectively, these results identify a critical role for MCPIP1 in angiogenesis and glioma progression by regulating the VEGFA-mediated ERK pathway, suggesting that targeting MCPIP1 may be a potential glioma-selective therapeutic strategy.

Selective delivery of low-affinity IL-2 to PD-1+ T cells rejuvenates antitumor immunity with reduced toxicity.

PD-1 signaling on T cells is the major pathway that limits T cell immunity, but the efficacy of anti-PD-1 therapy has been limited to a small proportion of patients with advanced cancers. We fortuitously observed that anti-PD-1 therapy depends on IL-2 signaling, which raises the possibility that a lack of IL-2 limits anti-PD-1-induced effector T cell expansion. To selectively deliver IL-2 to PD-1+CD8+ tumor-infiltrating lymphocytes (TILs), we engineered a low-affinity IL-2 paired with anti-PD-1 (PD-1-laIL-2), which reduced affinity to peripheral Treg cells but enhanced avidity to PD-1+CD8+ TILs. PD-1-laIL-2 exerted better tumor control and lower toxicity than single or mixed treatments. Mechanistically, PD-1-laIL-2 could effectively expand dysfunctional and tumor-specific CD8+ T cells. Furthermore, we discovered that presumably dysfunctional PD-1+TIM3+ TILs are the dominant tumor-specific T cells responding to PD-1-laIL-2. Collectively, these results highlight that PD-1-laIL-2 can target and reactivate tumor-specific TILs for tumor regression as a unique strategy with stronger efficacy and lower toxicity.

Dual targeting of CTLA-4 and CD47 on Treg cells promotes immunity against solid tumors.

Blockade of CD47, the "do not eat me" signal, has limited effects in solid tumors despite its potent antitumor effects in hematopoietic malignancies. Taking advantage of the high expression of cytotoxic T lymphocyte-associated protein 4 (CTLA-4) on Treg cells and abundant Fc receptor-expressing active phagocytes inside the tumor microenvironment (TME), we designed and tested a heterodimer combining an anti-CTLA-4 antibody, which targets Treg cells, with the CD47 ligand, signal regulatory protein α (SIRPα), to selectively block CD47 on intratumoral Treg cells. We hypothesized that heterodimer treatment would increase antibody-dependent cellular phagocytosis of the targeted Treg cells. We found that anti-CTLA-4×SIRPα preferentially depleted ICOShigh immunosuppressive Treg cells in the TME and enhanced immunity against solid tumors, including MC38 and CT26 murine colon cancers. Mechanistically, we found that CD47 expression on Treg cells limited anti-CTLA-4-mediated depletion and Fc on the heterodimer-enhanced depletion. Furthermore, anti-human CTLA-4×SIRPα depleted tumor Treg cells and exhibits less toxicity than anti-human CTLA-4 in a humanized mouse model. Collectively, these results demonstrate that simultaneously modulating both "eat me" and do not eat me signals induces Treg cell depletion inside the TME and may be an effective strategy for treating solid tumors.

Do proinflammatory cytokines play a role in clozapine-associated glycometabolism disorders?

RATIONALE AND OBJECTIVE: Clozapine (CLZ) is the most effective drug for treatment-resistant schizophrenia but is associated with many side effects, including glycometabolism disorders. Immunological mechanisms may be involved in the development of clozapine side effects. Research relating the immunomodulatory effects of clozapine and its early markers to clinically relevant adverse events is needed to reduce the harmful side effects of clozapine. This study aimed to investigate the role of proinflammatory cytokines in clozapine-associated glycometabolism disorders. METHODS: We measured the effect of a range of doses of clozapine on glycometabolism-related parameters and proinflammatory cytokines levels in mice peripheral blood. We also examined the differences between these indicators in the peripheral blood of clozapine-treated schizophrenia patients and healthy controls. Furthermore, we detected proinflammatory cytokines expression in mice pancreatic tissue. RESULTS: Following clozapine administration, glucagon significantly decreased in mouse serum, and proinflammatory cytokine IL-ß levels markedly increased. Clozapine reliably increased proinflammatory cytokines (IL-1ß, IL-6, and TNF-α) expression in murine pancreatic tissue. Compared with healthy controls, clozapine-treated patients' BMI, blood glucose, and proinflammatory cytokines (IL-1ß, IL-6, and TNF-α) increased significantly. In clozapine-treated patients, a higher clozapine daily dosage was associated with higher levels of the proinflammatory cytokines IL-1ß and IL-6, and a significant positive correlation was observed between blood glucose levels and the proinflammatory cytokines IL-6 and TNF-α. CONCLUSION: Findings from animal experiments and clinical trials have shown clear evidence that clozapine has a regulatory effect on immune-related proinflammatory cytokines and influences glycometabolism indicators.

DNA Sensing in Mismatch Repair-Deficient Tumor Cells Is Essential for Anti-tumor Immunity.

Increased neoantigens in hypermutated cancers with DNA mismatch repair deficiency (dMMR) are proposed as the major contributor to the high objective response rate in anti-PD-1 therapy. However, the mechanism of drug resistance is not fully understood. Using tumor models defective in the MMR gene Mlh1 (dMLH1), we show that dMLH1 tumor cells accumulate cytosolic DNA and produce IFN-ß in a cGAS-STING-dependent manner, which renders dMLH1 tumors slowly progressive and highly sensitive to checkpoint blockade. In neoantigen-fixed models, dMLH1 tumors potently induce T cell priming and lose resistance to checkpoint therapy independent of tumor mutational burden. Accordingly, loss of STING or cGAS in tumor cells decreases tumor infiltration of T cells and endows resistance to checkpoint blockade. Clinically, downregulation of cGAS/STING in human dMMR cancers correlates with poor prognosis. We conclude that DNA sensing within tumor cells is essential for dMMR-triggered anti-tumor immunity. This study provides new mechanisms and biomarkers for anti-dMMR-cancer immunotherapy.

ZMYND8 Expression in Breast Cancer Cells Blocks T-Lymphocyte Surveillance to Promote Tumor Growth.

Emerging studies indicate that DNA damage in cancer cells triggers antitumor immunity, but its intrinsic regulatory mechanism in breast cancer cells remains poorly understood. Here, we show that ZMYND8 is upregulated and inhibits micronucleus formation and DNA damage in breast cancer cells. Loss of ZMYND8 triggered activation of the DNA sensor cyclic guanosine monophosphate-adenosine monophosphate synthase in micronuclei, leading to further activation of the downstream signaling effectors stimulator of IFN genes and NF-κB, but not TANK-binding kinase 1 and IFN regulatory factor 3, thereby inducing the expression of IFNß and IFN-stimulated genes (ISG) in breast cancer cells in vitro and tumors in vivo. ZMYND8 knockout (KO) in breast cancer cells promoted infiltration of CD4+ and CD8+ T cells, leading to tumor inhibition in syngeneic mouse models, which was significantly attenuated by treatment of anti-CD4/CD8-depleting antibodies or anti-IFNAR1 antibody and in immunodeficient Rag1 KO mice. In human breast tumors, ZMYND8 was negatively correlated with ISGs, CD4, CD8A, CD8B, and the tumor-lymphocyte infiltration phenotype. Collectively, these findings demonstrate that maintenance of genome stability by ZMYND8 causes breast cancer cells to evade cytotoxic T-lymphocyte surveillance, which leads to tumor growth. SIGNIFICANCE: These findings show that ZMYND8 is a new negative and intrinsic regulator of the innate immune response in breast tumor cells, and ZMYND8 may be a possible target for antitumor immunotherapy.

Author Correction: Tumor cells suppress radiation-induced immunity by hijacking caspase 9 signaling.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Telomere Stress Potentiates STING-Dependent Anti-tumor Immunity.

Telomerase is an attractive target for anti-tumor therapy as it is almost universally expressed in cancer cells. Here, we show that treatment with a telomere-targeting drug, 6-thio-2'-deoxyguanosine (6-thio-dG), leads to tumor regression through innate and adaptive immune-dependent responses in syngeneic and humanized mouse models of telomerase-expressing cancers. 6-thio-dG treatment causes telomere-associated DNA damages that are sensed by dendritic cells (DCs) and activates the host cytosolic DNA sensing STING/interferon I pathway, resulting in enhanced cross-priming capacity of DCs and tumor-specific CD8+ T cell activation. Moreover, 6-thio-dG overcomes resistance to checkpoint blockade in advanced cancer models. Our results unveil how telomere stress increases innate sensing and adaptive anti-tumor immunity and provide strong rationales for combining telomere-targeting therapy with immunotherapy.

Tumor cells suppress radiation-induced immunity by hijacking caspase 9 signaling.

High-dose radiation activates caspases in tumor cells to produce abundant DNA fragments for DNA sensing in antigen-presenting cells, but the intrinsic DNA sensing in tumor cells after radiation is rather limited. Here we demonstrate that irradiated tumor cells hijack caspase 9 signaling to suppress intrinsic DNA sensing. Instead of apoptotic genomic DNA, tumor-derived mitochondrial DNA triggers intrinsic DNA sensing. Specifically, loss of mitochondrial DNA sensing in Casp9-/- tumors abolishes the enhanced therapeutic effect of radiation. We demonstrated that combining emricasan, a pan-caspase inhibitor, with radiation generates synergistic therapeutic effects. Moreover, loss of CASP9 signaling in tumor cells led to adaptive resistance by upregulating programmed death-ligand 1 (PD-L1) and resulted in tumor relapse. Additional anti-PD-L1 blockade can further overcome this acquired immune resistance. Therefore, combining radiation with a caspase inhibitor and anti-PD-L1 can effectively control tumors by sequentially blocking both intrinsic and extrinsic inhibitory signaling.

Author Correction: A next-generation tumor-targeting IL-2 preferentially promotes tumor-infiltrating CD8+ T-cell response and effective tumor control.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Hypofractionated EGFR tyrosine kinase inhibitor limits tumor relapse through triggering innate and adaptive immunity.

Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) are a first-line therapy for rapidly killing tumors such as those associated with non-small cell lung cancer by blocking oncogenic receptor signaling, but tumor relapse often occurs. Here, we have observed that hypofractionated EGFR TKI treatment (HypoTKI) is more potent than standard hyperfractionated EGFR TKI treatment (HyperTKI), and its antitumor effect associated with preventing tumor relapse depends on T cells. HypoTKI triggers greater innate sensing for type I IFN and CXCL10 production through the Myd88 signaling pathway to enhance tumor-specific T cell infiltration and reactivation. We also demonstrate that timely programmed cell death ligand-1 (PD-L1) blockade can synergize with HypoTKI to control advanced large tumors and effectively limit tumor relapse without severe side effects. Our study provides evidence for exploring the potential of a proper combination of EGFR TKIs and immunotherapy as a first-line treatment for treating EGFR-driven tumors.