BACKGROUND: Regulatory dendritic cell (DCreg) subset exhibits a unique capacity for inducing immune tolerance among the variety subsets of dendritic cells (DCs) within gut-associated lymphoid tissues (GALTs). Fms-like tyrosine kinase 3 ligand (FLT3L) is involved in the differentiation of DCregs and the subsequent expansion of regulatory T-cells (Tregs) mediated by DCregs, though the precise mechanism remains poorly understood. This study aimed to explore the expansion mechanism of Treg induced by DCreg and the role of FLT3L in this process. METHODS: DCregs were distinguished from other DC subsets isolated from GALTs of BALB/c mice through a mixed lymphocyte reaction assay. The functions and mechanisms by which FLT3L promoted Treg expansion via DCregs were investigated in vitro through co-culture experiments involving DCregs and either CD4 + CD25 - T-cells or CD4 + CD25 + T-cells. Additionally, an in vivo experiment was conducted using a dextran sulfate sodium (DSS)-induced colitis model in mice. RESULTS: CD103 + CD11b + DC exhibited DCreg-like functionality and was identified as DCreg for subsequent investigation. Analysis of Foxp3 + Treg percentages within a co-culture system of CD4 + CD25 - T-cells and DCregs, with or without FLT3L, demonstrated the involvement of the FLT3/FLT3L axis in driving the differentiation of precursor T-cells into Foxp3 + Tregs induced by DCregs. Cell migration and co-culture assays revealed that the FLT3/FLT3L axis enhanced DCreg migration toward Tregs via the Rho pathway. Additionally, it was observed that DCregs could promote Treg proliferation through the Notch pathway, as inhibition of Notch signaling by DAPT (N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester) suppressed Treg expansion within the co-culture system of DCregs and CD4 + T-cells or CD4 + CD25 + T-cells. Furthermore, the FLT3/FLT3L axis influenced JAG1 expression in DCregs, indirectly modulating Treg expansion. In vivo experiments further established that FLT3L promoted DCreg expansion and restored Treg balance in DSS-induced colitis models, thereby ameliorating colitis symptoms in mice. CONCLUSION The FLT3/FLT3L axis is integral to the maintenance of DCreg function in Treg expansion.
Animals ; T-Lymphocytes, Regulatory/immunology* ; Dendritic Cells/immunology* ; Mice ; Mice, Inbred BALB C ; Membrane Proteins/metabolism* ; Receptors, Notch/metabolism* ; Lymphoid Tissue/metabolism* ; Signal Transduction/physiology* ; Coculture Techniques ; Flow Cytometry

The cyclic guanosine monophosphate-adenosine monophosphate synthase-stimulator of interferon genes (cGAS-STING) pathway is a cornerstone of host innate immunity, playing a central role in detecting cytosolic double-stranded DNA of both endogenous and exogenous origins. Upon activation, cGAS synthesizes the second messenger 2'3'-cyclic GMP-AMP (cGAMP), which binds and activates STING to trigger downstream immune responses, including the production of type I interferons and proinflammatory cytokines. Emerging studies highlight the cGAS-STING pathway as a promising therapeutic target for preventing and treating diverse pathologies, with particularly transformative potential in anticancer therapies. In this review, we dissect the key findings, functions, and associated components of the cGAS-STING pathway. In addition, we emphasize the factors that upregulate or downregulate the pathway, as well as the role of the cGAS-STING pathway in health and disease. By integrating mechanistic insights with clinical perspectives, this review aims to bridge fundamental discoveries with therapeutic applications of cGAS-STING biology.
Humans ; Nucleotidyltransferases/metabolism* ; Membrane Proteins/metabolism* ; Animals ; Immunity, Innate/physiology* ; Signal Transduction/physiology* ; Neoplasms/metabolism*

The etiology of nervous system diseases is complicated, posing significant harm to patients and often resulting in poor prognoses. In recent years, the role of dopaminergic system in nervous system diseases has attracted much attention, and its complex regulatory mechanism and therapeutic potential have been gradually revealed. This paper reviews the role of dopaminergic neurons, the neurotransmitter dopamine, dopamine receptors and dopamine transporters in neurological diseases (including Alzheimer's disease, Parkinson's disease and schizophrenia), with a view to further elucidating the disease mechanism and providing new insights and strategies for the treatment of neurological diseases.
Humans ; Dopamine/metabolism* ; Nervous System Diseases/physiopathology* ; Parkinson Disease/physiopathology* ; Receptors, Dopamine/metabolism* ; Dopaminergic Neurons/physiology* ; Dopamine Plasma Membrane Transport Proteins/metabolism* ; Alzheimer Disease/physiopathology* ; Schizophrenia/physiopathology* ; Animals

Objective To explore the effect of the knockdown of transmembrane 9 superfamily protein member 2 (TM9SF2) on the replication of vesicular stomatitis virus (VSV), and investigate its role in the mechanism of antiviral innate immunity. Methods Small interfering RNA (siRNA) was used to knock down the TM9SF2 gene in human non-small cell lung cancer A549 cells. The CCK-8 method was used to assess cell proliferation. A VSV-green fluorescent protein (VSV-GFP) infected cell model was established. The plaque assay was used to measure the viral titer in the supernatant. RT-qPCR and Western blotting were employed to quantify the mRNA and protein levels of VSV genome replication in A549 cells following VSV infection, as well as the expression of interferon β (IFN-β) mRNA and interferon regulatory factor 3 (IRF3) protein phosphorylation following polyinosinic-polycytidylic acid (poly(I:C)) stimulation. Results Compared to the negative control, the knockdown of TM9SF2 exhibited a significant effect, with no observed impact on A549 cell proliferation. The VSV-GFP infected A549 cell model was successfully established. After viral stimulation, fluorescence intensity was reduced following TM9SF2 knockdown, and the mRNA and protein levels of VSV were significantly downregulated. The viral titer of VSV was decreased. After poly(I:C) stimulation, TM9SF2 knockdown significantly upregulated the mRNA level of IFN-β and the phosphorylation level of IRF3 protein. Conclusion The knockdown of TM9SF2 inhibits the replication of vesicular stomatitis virus, and positively regulates the type I interferon signaling pathway, thus enhancing the host's antiviral innate immune response.
Humans ; Virus Replication/genetics* ; Signal Transduction ; Membrane Proteins/metabolism* ; A549 Cells ; Vesiculovirus/physiology* ; Interferon-beta/metabolism* ; Interferon Regulatory Factor-3/genetics* ; Interferon Type I/metabolism* ; Vesicular Stomatitis/immunology* ; Gene Knockdown Techniques ; Vesicular stomatitis Indiana virus/physiology* ; RNA, Small Interfering/genetics*

Virus-induced senescence (VIS) is a significant biological phenomenon, which is associated with declining immune function, accelerating aging process and causing aging-related diseases. A variety of common viruses, including RNA viruses (such as SARS-CoV-2), DNA viruses (such as herpesviruses and hepatitis B virus), and prions can cause VIS in host cells. The primary mechanisms include abnormal activation of the cGAS-STING signaling pathway, DNA damage response, and potential correlations with the integrated stress response due to intracellular phase separation. Viral infection and cellular senescence influence each other: cellular senescence serves as a defense to restrict viral replication and transmission, while some viruses exploit cellular senescence to enhance their infectivity and replication. Understanding the mechanisms of VIS is conducive to the development of therapeutic strategies for viral infections and promotion of healthy aging. However, there is lack of research on therapeutic targets and drug development in this field so far. Although senolytics may be effective for anti-senescent cells therapy, their efficacy for VIS needs evidence from further clinical trials. This article reviews the research progress on the connection between viral infection and cellular senescence, to provide insights for the prevention and treatment of aging related diseases.
Humans ; Cellular Senescence/physiology* ; Virus Diseases/physiopathology* ; Signal Transduction ; Nucleotidyltransferases/metabolism* ; DNA Damage ; Virus Replication ; COVID-19 ; Membrane Proteins/metabolism* ; SARS-CoV-2

OBJECTIVES: The neurotoxicity of carbon monoxide (CO) to the central nervous system is a key pathogenesis of delayed encephalopathy after acute carbon monoxide poisoning (DEACMP). Our previous study found that retinoic acid (RA) can suppress the neurotoxic effects of CO. This study further explores, in vivo and in vitro, the molecular mechanisms by which RA alleviates CO-induced central nervous system damage. METHODS: A cytotoxic model was established using the mouse hippocampal neuronal cell line HT22 and primary oligodendrocytes exposed to CO, and a DEACMP animal model was established in adult Kunming mice. Cell viability and apoptosis of hippocampal neurons and oligodendrocytes were assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and Annexin V/propidium iodide (PI) double staining. The transcriptional and protein expression of each gene was detected using real-time fluorescence quantitative PCR (RT-qPCR) and Western blotting. Long noncoding RNA (lncRNA) SNHG15 and LINGO-1 were knocked down or overexpressed to observe changes in neurons and oligodendrocytes. In DEACMP mice, SNHG15 or LINGO-1 were knocked down to assess changes in central nervous tissue and downstream protein expression. RESULTS: RA at 10 and 20 μmol/L significantly reversed CO-induced apoptosis of hippocampal neurons and oligodendrocytes, downregulation of SNHG15 and LINGO-1, and upregulation of brain-derived neurotrophic factor (BDNF) and tyrosine kinase receptor B (TrkB) (all P<0.05). Overexpression of SNHG15 or LINGO-1 weakened the protective effect of RA against CO-induced cytotoxicity (all P<0.05). Knockdown of SNHG15 or LINGO-1 alleviated CO-induced apoptosis of hippocampal neurons and oligodendrocytes and upregulated BDNF and TrkB expression levels (all P<0.05). Experiments in DEACMP model mice showed that knockdown of SNHG15 or LINGO-1 mitigated central nervous system injury in DEACMP (all P<0.05). CONCLUSIONS RA alleviates CO-induced apoptosis of hippocampal neurons and oligodendrocytes, thereby reducing central nervous system injury and exerting neuroprotective effects. LncRNA SNHG15 and LINGO-1 are key molecules mediating RA-induced inhibition of neuronal apoptosis and are associated with the BDNF/TrkB pathway. These findings provide a theoretical framework for optimizing the clinical treatment of DEACMP and lay an experimental foundation for elucidating its molecular mechanisms.
Animals ; RNA, Long Noncoding/physiology* ; Brain-Derived Neurotrophic Factor/genetics* ; Carbon Monoxide Poisoning/complications* ; Mice ; Tretinoin/pharmacology* ; Nerve Tissue Proteins/metabolism* ; Membrane Proteins/metabolism* ; Apoptosis/drug effects* ; Hippocampus/cytology* ; Receptor, trkB/metabolism* ; Neurons/drug effects* ; Male ; Brain Diseases/etiology* ; Oligodendroglia/drug effects* ; Signal Transduction ; Cell Line

Alzheimer's disease (AD) is the most prevalent neurodegenerative disorder worldwide, causing dementia and affecting millions of individuals. One prominent characteristic in the brains of AD patients is glucose hypometabolism. In the context of galactose metabolism, intracellular glucose levels are heightened. Galactose mutarotase (GALM) plays a crucial role in maintaining normal galactose metabolism by catalyzing the conversion of β-D-galactose into α-D-galactose (α-D-G). The latter is then converted into glucose-6-phosphate, improving glucose metabolism levels. However, the involvement of GALM in AD progression is still unclear. In the present study, we found that the expression of GALM was significantly increased in AD patients and model mice. Genetic knockdown of GALM using adeno-associated virus did not change the expression of amyloid precursor protein (APP) and APP-cleaving enzymes including a disintegrin and metalloprotease 10 (ADAM10), β-site APP-cleaving enzyme 1 (BACE1), and presenilin-1 (PS1). Interestingly, genetic overexpression of GALM reduced APP and Aβ deposition by increasing the maturation of ADAM10, although it did not alter the expression of BACE1 and PS1. Further electrophysiological and behavioral experiments showed that GALM overexpression significantly ameliorated the deficits in hippocampal CA1 long-term potentiation (LTP) and spatial learning and memory in AD model mice. Importantly, direct α-D-G (20 mg/kg, i.p.) also inhibited Aβ deposition by increasing the maturation of ADAM10, thereby improving hippocampal CA1 LTP and spatial learning and memory in AD model mice. Taken together, our results indicate that GALM shifts APP processing towards α-cleavage, preventing Aβ generation by increasing the level of mature ADAM10. These findings indicate that GALM may be a potential therapeutic target for AD, and α-D-G has the potential to be used as a dietary supplement for the prevention and treatment of AD.
Animals ; ADAM10 Protein/metabolism* ; Alzheimer Disease/pathology* ; Amyloid Precursor Protein Secretases/metabolism* ; Disease Models, Animal ; Humans ; Mice ; Amyloid beta-Peptides/metabolism* ; Male ; Mice, Transgenic ; Membrane Proteins/metabolism* ; Cognitive Dysfunction/pathology* ; Mice, Inbred C57BL ; Amyloid beta-Protein Precursor/metabolism* ; Female ; Hippocampus/metabolism* ; Long-Term Potentiation/physiology*

This study aimed to investigate the effect of treadmill exercise on neuropathic pain and to determine whether mitophagy of the anterior cingulate cortex (ACC) contributes to exercise-mediated amelioration of neuropathic pain. Chronic constriction injury of the sciatic nerve (CCI) was used to establish a neuropathic pain model in Sprague-Dawley (SD) rats. Von-Frey filaments were used to assess the mechanical paw withdrawal threshold (PWT), and a thermal radiation meter was used to assess the thermal paw withdrawal latency (PWL) in rats. qPCR was used to evaluate the mRNA levels of Pink1, Parkin, Fundc1, and Bnip3. Western blot was used to evaluate the protein levels of PINK1 and PARKIN. To determine the impact of the mitophagy inducer carbonyl cyanide m-chlorophenylhydrazone (CCCP) on pain behaviors in CCI rats, 24 SD rats were randomly divided into CCI drug control group (CCI+Veh group), CCI+CCCP low-dose group (CCI+CCCP0.25), CCI+CCCP medium-dose group (CCI+CCCP2.5), and CCI+CCCP high-dose group (CCI+CCCP5). Pain behaviors were assessed on 0, 1, 3, 5, and 7 days after modeling. To explore whether exercise regulates pain through mitophagy, 24 SD rats were divided into sham, CCI, and CCI+Exercise (CCI+Exe) groups. The rats in the CCI+Exe group underwent 4-week low-moderate treadmill training one week after modeling. The mechanical pain and thermal pain behaviors of the rats in each group were assessed on 0, 7, 14, 21, and 35 days after modeling. Western blot was used to detect the levels of the mitophagy-related proteins PINK1, PARKIN, LC3 II/LC3 I, and P62 in ACC tissues. Transmission electron microscopy was used to observe the ultrastructure of mitochondrial morphology in the ACC. The results showed that: (1) Compared with the sham group, the pain thresholds of the ipsilateral side of the CCI group decreased significantly (P < 0.001). Meanwhile, the mRNA and protein levels of Pink1 were significantly higher, and those of Parkin were lower in the CCI group (P < 0.05). (2) Compared with the CCI+Veh group, each CCCP-dose group showed higher mechanical and thermal pain thresholds, and the levels of PINK1 and LC3 II/LC3 I were elevated significantly (P < 0.05, P < 0.01). (3) The pain thresholds of the CCI+Exe group increased significantly compared with those of the CCI group after treadmill intervention (P < 0.001, P < 0.01). Compared with the CCI group, the protein levels of PINK1 and P62 were decreased (P < 0.001, P < 0.01), and the protein levels of PARKIN and LC3 II/LC3 I were increased in the CCI+Exe group (P < 0.01, P < 0.05). Rod-shaped mitochondria were observed in the ACC of CCI+Exe group, and there were little mitochondrial fragmentation, swelling, or vacuoles. The results suggest that the mitochondrial PINK1/PARKIN autophagy pathway is blocked in the ACC of neuropathic pain model rats. Treadmill exercise could restore mitochondrial homeostasis and relieve neuropathic pain via the PINK1/PARKIN pathway.
Rats ; Animals ; Mitophagy/physiology* ; Rats, Sprague-Dawley ; Carbonyl Cyanide m-Chlorophenyl Hydrazone/pharmacology* ; Gyrus Cinguli ; Neuralgia ; Ubiquitin-Protein Ligases/metabolism* ; Protein Kinases ; Membrane Proteins/metabolism* ; Mitochondrial Proteins/metabolism*

Human salivary histatin 1 (Hst1) exhibits a series of cell-activating properties, such as promoting cell spreading, migration, and metabolic activity. We recently have shown that fluorescently labeled Hst1 (F-Hst1) targets and activates mitochondria, presenting an important molecular mechanism. However, its regulating signaling pathways remain to be elucidated. We investigated the influence of specific inhibitors of G protein-coupled receptors (GPCR), endocytosis pathways, extracellular signal-regulated kinases 1/2 (ERK1/2) signaling, p38 signaling, mitochondrial respiration and Na+/K+-ATPase activity on the uptake, mitochondria-targeting and -activating properties of F-Hst1. We performed a siRNA knockdown (KD) to assess the effect of Sigma-2 receptor (S2R) /Transmembrane Protein 97 (TMEM97)-a recently identified target protein of Hst1. We also adopted live cell imaging to monitor the whole intracellular trafficking process of F-Hst1. Our results showed that the inhibition of cellular respiration hindered the internalization of F-Hst1. The inhibitors of GPCR, ERK1/2, phagocytosis, and clathrin-mediated endocytosis (CME) as well as siRNA KD of S2R/TMEM97 significantly reduced the uptake, which was accompanied by the nullification of the promoting effect of F-Hst1 on cell metabolic activity. Only the inhibitor of CME and KD of S2R/TMEM97 significantly compromised the mitochondria-targeting of Hst1. We further showed the intracellular trafficking and targeting process of F-Hst1, in which early endosome plays an important role. Overall, phagocytosis, CME, GPCR, ERK signaling, and S2R/TMEM97 are involved in the internalization of Hst1, while only CME and S2R/TMEM97 are critical for its subcellular targeting. The inhibition of either internalization or mitochondria-targeting of Hst1 could significantly compromise its mitochondria-activating property.
Endocytosis/physiology* ; Histatins/pharmacology* ; Humans ; Membrane Proteins ; Mitochondria/metabolism* ; RNA, Small Interfering/pharmacology* ; Receptors, G-Protein-Coupled/metabolism* ; Receptors, sigma

The aim of the present study was to investigate the effects of exercises with different durations and intensities on mitochondrial autophagy and FUNDC1 in rat skeletal muscles. Sixty male Sprague-Dawley rats were randomly divided into 2- and 4-week control groups (Con), moderate-intensity exercise groups (M-ex groups, treadmill exercise, 16 m/min, 1 h/d, 6 d/week), and high-intensity exercise groups (Hi-ex groups, treadmill exercise, 35 m/min, 20 min/d, 6 d/week). The bilateral soleus muscles were separated after the intervention, and paraffin sections were prepared for transmission electron microscopy. ELISA method was used to detect the content of citrate synthase (CS). The co-localizations of microtubule-associated protein 1 light chain 3 (LC3)/cytochrome c oxidase IV (COX-IV), FUNDC1/COX-IV and LC3/FUNDC1 were observed by immunofluorescent staining in frozen sections. The skeletal muscle mitochondria were extracted, and the expression of autophagy-related proteins, including AMPKα, p-AMPKα, Unc-51 like kinase 1 (ULK1), FUNDC1, LC3 and p62, were detected by Western blot. The results showed that exercise increased mitochondrial function, i.e. peroxisome proliferator-activated receptor γ co-activator-1α (PGC-1α), COX-I protein expression levels and CS content. There was no difference of mitochondrial function parameters between 2-week M-ex and 2-week Hi-ex groups, while mitochondrial function of 4-weeks Hi-ex group was significantly lower than that of 4-week M-ex group. Under the same exercise intensity, mitochondrial autophagy activation in skeletal muscle of 4-week exercise was higher than that in 2-week exercise group; Under the same duration of exercise, mitochondrial autophagy activation of Hi-ex group was higher than that in M-ex group. Both 2- and 4-week exercise intervention increased LC3/COX-IV, COX-IV/FUNDC1, and FUNDC1/LC3 co-localizations. Exercise increased LC3-II/LC3-I ratio, down-regulated p62 protein expression level, up-regulated FUNDC1, ULK1 protein expression levels and AMPKα phosphorylation, and the changes of these proteins in 4-week Hi-ex group were significantly greater than those in 4-week M-ex group. These results suggest exercise induces mitochondrial autophagy in skeletal muscles, and the activity of autophagy is related to the duration and intensity of exercise. The induction mechanism of exercise may involve the mediation of FUNDC1 expression through AMPK-ULK1 pathway.
Animals ; Autophagy ; Exercise Therapy ; Humans ; Male ; Membrane Proteins/physiology* ; Mitochondria ; Mitochondrial Proteins/physiology* ; Muscle, Skeletal/metabolism* ; Rats ; Rats, Sprague-Dawley