第10卷，第5期

Kirsten rat sarcoma (KRAS) mutant non-small cell lung cancer (NSCLC) hasdistinctco-mutationalprofiles. Approximately 20% of KRAS mutant NSCLC cases underwent loss-of-function mutation of Kelch-like ECH-associated protein 1 (KEAP1) gene, namely KRAS-KEAP1 comutation (KK) type, which exhibited a poorer immune checkpoint inhibitor (ICI) response compared with individual KRAS mutation (K) type tumors. Recent studies suggest that K-type tumors are referred to as "hot tumors" while KK-type tumors are as "cold tumors". Therefore, clarifying the molecular mechanism of the poor ICI-responsive phenotype of KK-type tumors is the key to improving the clinical efficacy and breaking through the therapy bottleneck. Co-mutations could regulate the immune contexture to reshape the tumor immune microenvironment (TIME), which has been identified as a major influence factor of ICI response. KK-type tumors were confirmed to contain few tumor-infiltrating lymphocytes, especially CD8⁺ T cells, leading to a suppressive TIME, which is also defined as a "cold tumor". Here, we demonstrated that NRF2, the target of KEAP1, exerted its role as a negative regulator of CD8⁺ T cells recruitment by decreasing CCL5 and CXCL10 chemokines in KK-type NSCLC. Mechanistically, NRF2 promoted the transcription and expression of BRCA1 to repair DNA damage, resulting in STING pathway inactivation. We propose the combination of NRF2 inhibitor or STING agonist with ICI may be a promising therapeutic approach for patients with KK-type NSCLC.

1135

Currently, the primary treatment for hepatocellular carcinoma (HCC) is a comprehensive treatment based on surgery. In the case of advanced HCC that may not be removed surgically, additional issues, such as drug resistance and drug inefficacy with long-term use of chemotherapeutic drugs, highlight the pressing need for new treatment strategies. Autophagy plays an essential role in cellular physiology, which was reported to modulate components of the immune system. Autophagy-related immune genes (ARIGs) are linked to both autophagy and immunity. Here we identified histone deacetylase 1 (HDAC1), an ARIG strongly expressed in HCC, as a therapeutic target. Then valproic acid (VPA), a specific inhibitor of HDAC1, was used to treat liver cancer in vitro and in vivo. The results demonstrated that VPA could significantly induce autophagy and apoptosis of Hepa1-6 cells and inhibit tumor growth in vivo. This effect could be related to the regulation of autophagy and tumor immune microenvironment by VPA.

1251

1472

Poly (ADP-ribose) polymerase inhibitors (PARPi) are used to treat ovarian cancer and triple-negative breast cancer (TNBC) with defective homologous recombination repair pathways. However, de novo and acquired PARPi resistance limits clinical benefits. The MRE11-RAD50-NBS1 (MRN) complex mediates the sensing, processing, and signaling of DNA double-strand breaks (DSBs) and plays important roles in the efficacy of PARPi and radiation treatment, and yet the mechanisms for the regulation and degradation of the MRN complex are not well understood. ZRANB1, also known as Trabid, is a breast cancer-promoting deubiquitinase that preferentially cleaves K29-, K33-, and K63-linked ubiquitin chains, but its role in therapy resistance remains unknown.

1236

Glycosylation is a ubiquitous molecular modification in the process of cellular metabolism regulation, which plays a vital role in maintaining and regulating cellular functions. Deficiency in glycosylation enzymes may jeopardize the glycosylation process and eventually lead to the metabolic disease, manifesting with either a neurologic or multisystem phenotype, which is also known as congenital disorder of glycosylation (CDG). It could be classified into type I CDG and type II CDG, and the latter was referred to as the abnormal formation of nascent glycoprotein. Phosphomannomutase 2 CDG (PMM2-CDG), the most prevalent CDG with an incidence rate of 1 in 20, 000 individuals, is mainly due to the metabolic disorder of phosphomannomutase 2 (PMM2). PMM2 participates in the conversion of mannose-6-phosphatetomannose-1-phosphate. The cysteine residues, as one of the least abundant amino acids, have unique attributes to maintain the stability of protein structure, especially for catalytic activity and protein folding. Therefore, cysteine mutations have been identified in various diseases including PMM2-CDG. Suggested by the crystal structure of PMM2 (PDB ID: 7O5Z), four cysteine mutations in the hPMM2, C9Y, C103F, C192G, and C241S, have been found in PMM2-CDG. After bioinformatic analysis by PROVEAN, the four cysteine mutations were predicted with "Deleterious", ranking with the best two were C103F and C192G (C9Y: -5.493, C103F: -6.339, C192G: -11.176, C241S: -2.986). However, the underlying pathological mechanism is still unclear. Among them, C103F and C192G have only been reported in clinical study without a detailed explanation, so they were selected for our research. Two mutants were located near the linker region 1 and 2; C103F was located on the No. 4 a helix and C192G was located on the No. 7 a helix. The purpose of this study was to explore the structure and function roles of PMM2 clinical mutation and investigate the role of cysteine in PMM2 which might provide new insights into preventive and therapeutic strategies for PMM2-CDG.

1814

Akt aberrant activation accelerated tumor development and metastasis, and our previous study identified silent information regulator 6 (SIRT6) as a novel and critical tumor suppressor in colorectal cancer (CRC) downstream of Akt. However, the function of SIRT6 in cancer progression has not been fully elucidated. In the present study, we found that except for inducing apoptosis, SIRT6 could serve as a crucial regulator to initiate autophagy by directlyinteractingwithandactivatingULK1. More importantly, we also reported autophagy regulation in a SIRT6-dependentindirectmanner, namely, SIRT6 competitively bound to PUMA, which led to the release of ULK1. Of note, autophagy inhibition enhanced the proapoptotic effect of Midostaurin both in vitro and in vivo. Summarily, SIRT6 induced apoptosis accompanied by protective autophagy, and SIRT6 promoted autophagy by directly interacting with ULK1 and competitive binding to PUMA. This new insight identified the dual function of SIRT6 on apoptosis and autophagy, which has attractive clinical applications in cancer therapy.

1384

N⁶-methyladenosine (m⁶A) is the most abundant modification in eukaryotic messenger RNA. This modification is dynamic, reversible, and mediated by proteins characterized as methyltransferases and demethylases. Recent investigations have found that aberrant expression of methyltransferases and demethylases results in m⁶A dysregulation and, in consequence, affects the biological functions in which this modification is involved. Indeed, m⁶A dysregulation affects the development and maintenance of various diseases, including cancer. For this reason, we explored the potential role that m⁶A modification has in invasive breast cancer of no special type (IBC-NST) and its molecular subtypes luminal A, luminal B HER2-, luminal B HER2+, HER2+, and triple-negative breast cancer (TNBC) whose degree of global m⁶A methylation has not yet been studied.

1234

Acralmelanoma (AM) is a rare subtypeof cutaneousmelanoma linked to poor prognosis, largely due to a lacking of effective targeted therapeutic strategies. Whole-genome sequencing (WGS) data revealed that AM showed a different mutation landscape from cutaneous melanoma. BRCA1 and BRCA2 mutations appear in about 3%-16% of AMs. Pharmacologic inhibition of the DNA repair enzyme PARP has been approved by the FDA as monotherapy in patients with deleterious germline BRCA1/2 mutated advanced ovarian cancer, and the usage has been expanded to metastatic breast cancer, pancreatic cancer, and prostate cancer with homologous recombination repair (HRR) gene defects. However, whether AM with BRCA mutations is also sensitive to PARP inhibition is unknown. We identified a stageⅣAM patient with a germline BRCA1 frameshift mutation (BRAC1 G1384Nfs*7) who was resistant to anti-PD1 therapy. Both patient-derived xenograft and cells (PDX/PDC) models from the same AM patient were established. PARP inhibitor olaparib significantly decreased cell proliferation and slowed tumor growth by increasing DNA double-strand breakage in AM cancer cells. Administration of olaparib to the patient achieved stable disease for 3 months. This study provides preclinical and clinical evidence that PARP inhibitors can slow tumor growth in BRCA1-mutant advanced acral melanoma.

1236

Hereditary tyrosinemia type 1 (HT1) is a life-threatening disease caused by the patient’s inability to break down tyrosine due to loss-of-function mutations in the fumarylacetoacetate hydrolase (FAH) enzyme. Currently, the only available life-saving treatment is nitisinone (NTBC). However, nitisinone therapy comes with debilitating side effects and requires a strict drug regime combined with a tyrosine- and phenylalanine-restricted diet. Consequently, therapy adherence is often experienced as an additional burden. In this study, transcriptional profiling was conducted parallel to high-resolution metabolomics on, respectively, liver tissue and serum samples of Fah-deficient mice. The experimental workflow of our study is shown in. Canonical pathway analyses showed manifest activation of the NRF2-stress response, associated with hepatocellular damage and mainly driven by a markedly increased oxidative burden. Other altered canonical pathways were primarily a downstream consequence of the latter. Furthermore, we observed an overall aminoacidemia and changes related to aminoacyl-tRNA biosynthesis, which postulates that short-term nitisinone discontinuation can already put the patient at risk for hepatocellular carcinoma development. Finally, we did not only identify changes in numerous pathways, directly and indirectly, related to the metabolism of tyrosine and glutathione but also the presence of “side-chain” metabolites, such as y-glutamyl- and N-acetyl-coupled amino acids, indicating that non-directly related pathways like phase II biotransformation reactions are also affected.

1282

The maternally expressed gene 3 (MEG3) acts as an antitumor component in different cancer cells. However, MEG3 variant has been shown to confer cancer susceptibilityand certain polymorphisms within MEG3 are implicated in cancer risk (rs7158663, rs4081134, and rs11160608). Moreover, MEG3-TTCC and MEG3-CCCA promote the differentiation of SCs by activating the JAK2/STAT3 signaling pathway in different degrees. In particular, the alleles of MEG3 (AA, GA + AA) and MEG3 (rs7158663) were associated with an increased risk for human liver cancer. Moreover, Sirt2 is a specific NAD-dependent histone deacetylase for H4eK16 and has a strong preference for histone H4K16Ac in their deacetylation activity. Nevertheless, the functions and mechanism of MEG3 variant on hepatocarcinogenesis have not yet been well-demonstrated.

1255

Glioblastoma multiforme (GBM) is the most common primary malignant tumor of the central nervous system. Grade IV GBM is lethal, and has a high rate of relapse despite aggressive therapy. The prognosis is poor because of therapeutic resistance and tumor relapse. Very low rates of clinical responses and frequent treatment failures are common. Previous studies have focused on identifying genetic alterations in GBM that have diagnostic and prognostic values and may help to define the subclasses of the GBM patients, including mutations in isocitrate dehydrogenase 1 (IDH1), amplification of epidermal growth factor receptors (EGFR), and mutations and polymorphisms of telomerase reverse transcriptase (TERT) promoter. These markers are useful for identifying the pathogenesis of malignant glioma formation, but they are all associated with tumorigenesis and do not currently influence the treatment of most GBM patients.

1254

1468

p53 is an important tumor suppressor gene. The p53 pathway is activated in response to cellular stress stimulation. However, in more than 50% of breast cancers, p53 is mutated or inactivated, which permits cancer growth. Although a large number of co-regulators of p53 have been identified, the integrated molecular network of p53 and the key co-factors that could transactivate p53 remain unclear. Myeloid Zinc Finger 1 (MZF1), a member of the SCAN-ZFP (SCAN-Zinc finger protein) family, has been involved in the occurrence and development of various types of malignant tumors, including breast cancer. However, the exact mechanism of action remains unclear. Our study aims to investigate the cross-regulatory loop between MZF1 and p53 in breast cancer cells.

1222

1258

Wilms tumor (WT) is the most common childhood kidney cancer. Although WT is largely curable, current treatments fail in up to 15 percent of patients. Moreover, survivors suffer from the complications and late effects of the aggressive treatments. Thus, there is a critical need to improve our understanding of tumorigenesis to develop novel therapies to reduce the treatment burden while maintaining excellent survival rates. WT is believed to arise from the immature kidney cells, nephron progenitor cells (NPCs), which have failed to differentiate properly. Previous studies revealed that Wilms cells share a transcriptional and epigenetic landscape with normal renal stem cells. Although several studies have shown the positive associations between WT in children and embryonic exposure to adverse environments, the underlying mechanisms remain unknown. Altered epigenetics is central to oncogenesis in manypediatriccancers. The critical contribution of epigenetic dysregulation to pediatric tumors provides a compelling rationale for the therapeutic potential of epigeneticdrugs. Histonedeacetylases (HDACs) and Enhancer of Zeste Homolog 2 (EZH2, a histone H3K27 methyltransferase), have been demonstrated to play a critical role in self-renewal and differentiation of mouse NPCs. In addition, altered expression and mutations of HDACs and EZH2 have been linked to many human cancers, including WT. Thus, they are among the most promising therapeutic targets for cancer treatment. We reasoned that WT would result from the unrestrained proliferation of progenitor cells due to overactive HDAC1/2 (HDAC1 and HDAC2), and EZH2. We tested this hypothesis by analyzing a clinical specimen received from the left kidney tumor of an 11-year-old male patient diagnosed with WT and four other human WT specimens. We have the approval from Tulane Human Research Protection Office & Institutional Review Boards (Study number: 2019-623) to study WT specimens. The tumor (9.5 cm -8 cm -7.5 cm) is classified as favorable for the histology of indeterminate cell tumors. As shown in Figure 1A, the tumor exhibits a biphasic pattern with significant blastemal component admixed with stromal component. No significant epithelial component is present. The blastema represents the undifferentiated and malignant component, consisting of small round blue cells with overlapping nuclei and brisk mitotic activity. The blastemal component shows somewhat a basaloid growth pattern. The stromal component is also prominent and includes hypercellular undifferentiated mesenchymal cells.

1256

Phospholipase D (PLD) and its product phosphatidic acid (PA) function as pleiotropic factors in the regulation of cancer progression, which includes the promotion of cell growth, survival, cell migration, and angiogenesis. Chemical inhibition of the enzymatic activity of PLD1 and PLD2, or genetic down-regulation of their expression both induce tumor suppression, while the compounds for PLD expression inhibition are barely reported. Here, we astonishingly found that the anti-hyperglycemic agents biguanides decreased PLD1 protein expression in cancer cells and further investigated its underlying regulatory mechanism.

1937

Mantle cell lymphoma (MCL) is an aggressive subtype of non-Hodgkin lymphoma (NHL) characterized by the overexpression of cyclin D1 and deregulated cell cycle. Ganetespib (STA-9090), a second-generation HSP90 inhibitor, dramatically disrupted oncogenic cellular processes resulting in the inhibition of client protein-derived tumors in preclinical studies. The synthetic lethal strategy using poly (ADP-ribose) polymerase (PARP) inhibitors (PARPis) has been reported as a powerful therapeutic intervention in MCL and their effectiveness can be increased by a deficiency in DNA damage repair (DDR), especially homologous recombination deficiency. Since we found earlier that transient ganetespib treatment can induce defects in DDR, we hypothesized that ganetespib treatment can possibly enhance the sensitivity of MCL cells to PARPis and HSP90 and PARP is can be demonstrated as promising combination therapies for MCL.

1203

The corneal endothelial monolayer is responsible for maintaining corneal transparency through its barrier and pump functions. Endothelial cells do not proliferate in vivo, and aging-related reductions in cell density make the endothelium fragile. However, the molecular mechanism associated with the aging corneal endothelium remains elusive. In this study, we used ultraviolet A (UVA)-light-induced senescence to mimic the degenerative endothelial changes during aging. MALAT1, the most abundant long non-coding RNA (lncRNA) gene in the cornea, was markedly down-regulated after UVA irradiation. The inhibition or activation of Malat1 expression genetically led to the aggravation or remission of the cellular aging phenotype after UVA irradiation. Furthermore, an in vitro model established by the human corneal endothelial cell line (HCEC) recapitulated the morphological and molecular changes during aging, encouraging the investigation of the underlying mechanisms. We observed changes in the mitochondrial bioenergetic profiles of HCECs accompanied by extensive cell aging and reduced reproductive capacity after MALAT1 was silenced by gene-targeting ASOs. Finally, Malat1 knockout (KO) aggravated mouse corneal endothelial senescence and dysfunction, which reinforced our conclusions. Our findings indicate that MALAT1 may contribute to the delay of aging processes involving endothelial cells and provide a new therapeutic target for treating aging-related corneal endothelium disorders.

1246

Pancreatic cancer (PaCa) is one of the most aggressive and lethal malignancies with rapid progression and poor prognosis with the 5-year survival rate remaining less than 5% because approximately 80% of PaCa patients are diagnosed at an advanced stage and lose the chance of curative resection. Accumulated evidence has revealed that noncoding RNAs (ncRNAs) are involved in the pathogenesis of PaCa and serve as potential biomarkers for PaCa. Exosomal lncRNAs also play a vital role in tumorigenesis and some exosomal lncRNAs have been reported to accelerate tumor progression. In this study, we first employed RNA-seq of PaCa plasma samples to identify significantly differentially expressed lncRNAs in PaCa patients and revealed that LINC01268 was up-regulated in PaCa tumor tissues and plasma. LINC01268 is an emerging lncRNA associated with glioma malignancy grade. Thus. we further analyzed the malignant biological behaviors of this lncRNA in PaCa and found that LINC01268 promoted cell proliferation, migration, and invasion as an oncogene, which promoted epithelial-mesenchymal transition (EMT) in PaCa via the miR-217-KIF2A-PI3K/AKT axis.

1253

Glioblastoma (GBM) is the most common and lethal malignancy in the central nervous system. One of the major difficulties in treatment is that the initial clinical diagnosis of GBM is already WHO grade IV, without recognizable lower-grade precursor lesions. Copy number variations (CNVs) were found to appear in malignant cells several years before the initial diagnosis of GBM. Less differentiation and more aggressive phenotypes were observed in GBM cells with a higher degree of CNVs. Additionally, CNVs provide more accurate stratification of clinical outcomes than does the WHO grade system. Therefore, we reasoned that differentially expressed genes (DEGs) among GBM cells with different CNV statuses would be significant for the aggressiveness of GBM. Here we leveraged the single-cell RNA-sequencing (scRNA-seq) to construct the CNV profile of GBM at single-cell resolution, divided GBM cells into different clusters according to their CNV statuses, and investigated the molecular functions of DEGs among GBM clusters. Through a series of experiments, we identified anaphase-promoting complex subunit 11 (ANAPC11) as a switch controlling the neuronal differentiation of GBM cells, providing a novel alternative for the development of differentiation-inducing therapy to overcome GBM.

1199

Single-cellRNAsequencing (scRNA-seq) andmachine learning technologies, developed rapidly in recent years, offer an unparalleled opportunity to study the mechanism of tumorigenesis. Changes in metabolism-related genes (MRGs) result in hepatocellular carcinoma (HCC) progression, and MRGs have the potential to be used as a clinical prognostic indicator. In this study, a machine learning model was built using LASSO regression to find prognostic genes and create a new MRG risk signature of HCC patients. The patients were divided into the MRG high-and low-risk groups based on their risk scores. Significant overall survival advantages were observed in the MRG low-risk group. In addition, risk score and cancer status were used to establish a nomogram with high accuracy. The calibration curves and decision curves further demonstrated the satisfactory agreement between the predicted and observed values in the probability of overall survival. Subsequently, we discovered the different characteristics of the MRG highand low-risk group patients based on tumor mutation burden, gene set enrichment, and immune infiltration. Our study identifies a novel MRG signature for risk-stratification and accurate identification of HCC patients with poor subtypes, which could precisely predict the prognosis of HCC and may guide personalized treatment for HCC patients.

1209

Triple-negative breast cancer (TNBC) is highly malignant and refractory to immunotherapy through impeding the immune cell infiltration and inflammation in the tumor microenvironment (TME). DNA-dependent protein kinase catalytic subunit (DNA-PKcs) is a member of the phosphatidylinositol 3-kinase-related kinase family, which is required for the nonhomologous end joining repair. The effect of DNA-PKcs on the generation of cytosolic DNA and inflammation response in tumor immuno-environment is not defined. We found a specific DNA-PKcs inhibitor, NU7441, induced cytosolic DNA, stimulator of interferon genes (STING), and retinoic acidinducible gene I (RIG-I) signals in vitro. In Balb/c immunecompetent mice bearing 4T1 TNBC cells, NU7441 impaired the tumor growth and metastasis, and increased the CD45⁺ leukocytes, CD4⁺ T cells, CD8⁺ T cells, and CD1a⁺ antigenpresenting cells, as well as MHC-I and interferon alpha receptor (IFNAR) inTME. However, inBalb/c athymic nudemice without IFNAR and CD8⁺ T cells in TME, NU7741 did not influence tumor growth. These results show that inhibition of DNA-PKcs triggers cytosolic DNA sensing and induces an inflamed TME to promote anti-tumoral immunity, which provides a strategy to alter the inflammation and lymphocyte infiltration in TME to increase the efficacy of immunotherapy in TNBC and other cancers with an immune-suppressive TME.

1193

The m⁶A modification involves almost all aspects of RNA biology, including the alternative splicing of mRNA precursors, mRNA transport and stability, and miRNA processing and regulation of target genes. Alternative splicing controlling the information storage and RNA translation involves the regulation of various biological processes. Here, we integrated the genomic information of 161 esophageal cancer (EC) samples to comprehensively evaluate the m⁶A modification patterns and correlated the m⁶A modification pattern with the prognosis of EC patients, where two distinct m⁶A modification patterns were proposed. The combined effects of high m⁶Ascore and low TMB correlated with a better prognosis of EC patients. In addition, we found an inherent correlation between m⁶A modification and the occurrence of alternative splicing events in EC patients. Altogether, we established a scoring system to quantify the m⁶A modification pattern with RNA alternative splicing events in individual EC patients.

1205

Hengel et al recently reported that bi-allelic loss-of-function mutations in UDP-Glucose 6-Dehydrogenase (UGDH) caused a severe epileptic encephalopathy syndrome-Jamuar syndrome (OMIM#618792). The functional studies partially recapitulated the clinical phenotypes in the patient-derived cerebral organoid. A reduced number of proliferating neuronal progenitors in cerebral organoids was shown, which is a critical mechanism in congenital microcephaly (CM) whose patients were born with an occipitofrontal circumference (OCF) more than 2 standard deviations below average for age and sex. However, none of the reported patients in the article presented the phenotype as CM.

1187

Studies on carcinogenesis continue to provide new information about different disease-related processes. Among others, much research has focused on the involvement of cancer stem cells (CSCs) in tumor initiation and progression. Studying the similarities and differences between CSCs and physiological stem cells (SCs) allows for a better understanding of cancer biology. Recently, it was shown that stem cell identity is partially governed by the Krϋppel-associated box domain zinc finger proteins (KRAB-ZFPs), the biggest family of transcription regulators. Several KRAB-ZFP factors exert a known effect in tumor cells, acting as tumor suppressor genes (TSGs) or oncogenes, yet their role in CSCs is still poorly characterized. Here, we review recent studies regarding the influence of KRAB-ZFPs and their cofactor protein TRIM28 on CSCs phenotype, stemness features, migration and invasion potential, metastasis, and expression of parental markers.

1286

Higd1a is a conserved gene in evolution which is widely expressed in many tissues in mammals. Accumulating evidence has revealed multiple functions of Higd1a, as a mitochondrial inner membrane protein, in the regulation of metabolic homeostasis. It plays an important role in anti-apoptosis and promotes cellular survival in several cell types under hypoxic condition. And the survival of porcine Sertoli cells facilitated by Higd1a helps to support reproduction. In some cases, Higd1a can serve as a sign of metabolic stress. Over the past several years, a considerable amount of studies about how tumor fate is determined and how cancerous proliferation is regulated by Higd1a have been performed. In this review, we summarize the physiological functions of Higd1a in metabolic homeostasis and its pathophysiological roles in distinct diseases including cancer, nonalcoholic fatty liver disease (NAFLD), type II diabetes and mitochondrial diseases. The prospect of Higd1a with potential to preserve mammal health is also discussed. This review might pave the way for Higd1a-based research and application in clinical practice.

1266

Severe insulin resistance has been linked to some of the most globally prevalent disorders, such as diabetes mellitus, nonalcoholic fatty liver disease, polycystic ovarian syndrome, and hypertension. Hereditary severe insulin resistance syndrome (H-SIRS) is a rare disorder classified into four principal categories: primary insulin receptor defects, lipodystrophies, complex syndromes, and obesity-related H-SIRS. Genes such as INSR, AKT2, TBC1D4, AGPAT2, BSCL2, CAV1, PTRF, LMNA, PPARG, PLIN1, CIDEC, LIPE, PCYT1A, MC4R, LEP, POMC, SH2B1, RECQL2, RECQL3, ALMS1, PCNT, ZMPSTE24, PIK3R1, and POLD1 have been linked to H-SIRS. Its clinical features include insulin resistance, hyperglycemia, hyperandrogenism, severe dyslipidemia, fatty liver, abnormal topography of adipose tissue, and low serum leptin and adiponectin levels. Diagnosis of H-SIRS is based on the presence of typical clinical features associated with the various H-SIRS forms and the identification of mutations in HSIRS-linked genes by genetic testing. Diet therapy, insulin sensitization, exogenous insulin therapy, and leptin replacement therapy have widely been adopted to manage H-SIRS. The rarity of H-SIRS, its highly variable clinical presentation, refusal to be tested for genetic mutations by patients' family members who are not severely sick, unavailability of genetic testing, and testing expenses contribute to the delayed or underdiagnoses of H-SIRS. Early diagnosis facilitates early management of the condition, which results in improved glycemic control and delayed onset of diabetes and other complications related to severe insulin resistance. The use of updated genetic sequencing technologies is recommended, and long-term studies are required for genotype-phenotype differentiation and formulation of diagnostic and treatment protocols.

2017

Hepatocellular carcinoma (HCC) has a very high incidence and fatality rate, and in most cases, it is already at an advanced stage when diagnosed. Therefore, early prevention and detection of HCC are two of the most effective strategies. However, the methods recommended in the practice guidelines for the detection of HCC cannot guarantee high sensitivity and specificity except for the liver biopsy, which is known as the "gold standard". In this review, we divided the detection of HCC into pre-treatment diagnosis and post-treatment monitoring, and found that in addition to the traditional imaging detection and liver biopsy, alpha fetoprotein (AFP), lens culinaris-agglutinin-reactive fraction of AFP (AFP-L3), protein induced by vitamin K absence or antagonist-II (PIVKA-II) and other biomarkers are excellent biomarkers for HCC, especially when they are combined together. Most notably, the emerging liquid biopsy shows great promise in detecting HCC. In addition, lactic dehydrogenase (LDH), suppressor of cytokine signaling (SOCS) and other relevant biomarkers may become promising biomarkers for HCC post-treatment monitoring. Through the detailed introduction of the diagnostic technology of HCC, we can have a detailed understanding of its development process and then obtain some enlightenment from the diagnosis, to improve the diagnostic rate of HCC and reduce its mortality.

1290

Hydrogen sulfide (H₂S) is one of the three known gas signal transducers, and since its potential physiological role was reported, the literature on H₂S has been increasing. H₂S is involved in processes such as vasodilation, neurotransmission, angiogenesis, inflammation, and the prevention of ischemia-reperfusion injury, and its mechanism remains to be further studied. At present, the role of post-translational processing of proteins has been considered as a possible mechanism for the involvement of H₂S in a variety of physiological processes. Current studies have shown that H₂S is involved in S-sulfhydration, phosphorylation, and S-nitrosylation of proteins, etc. This paper focuses on the effects of protein modification involving H₂S on physiological and pathological processes, looking forward to providing guidance for subsequent research.

1198

Cellular metabolism-induced epigenetic regulation is essential for the maintenance of cellular homeostasis. Nicotinamide N-methyltransferase (NNMT) is emerging as a key point of intersection between cellular metabolism and epigenetic regulation and has a central role in various physiological and pathological processes. NNMT catalyzes the methylation of nicotinamide (NAM) using the universal methyl donor S-adenosyl methionine (SAM) to yield S-adenosyl-L-homocysteine (SAH) and N1-methylnicotinamide (MNAM), directly linking methylation balance with nicotinamide adenosine dinucleotide (NAD⁺) contents. NNMT acts on either the SAM-methylation balance or both NAD⁺ metabolism, depending on the tissue involved or pathological settings where metabolic demand is increased. Under physiological conditions, the liver act as an essential metabolic organ with abundant NNMT expression, while NNMT hepatic function is not mediated by its methyltransferase activity due to other major methyltransferases such as glycine N-methyltransferase (GNMT) in the liver. However, hepatic NNMT, as well as its metabolite is improperly regulated and linked to the worse pathological states in liver diseases, including alcoholic liver disease, non-alcoholic fatty liver disease (NAFLD), liver cirrhosis, and hepatocellular carcinoma (HCC), suggesting a potential role in the process of liver diseases. In this review, we summarize how NNMT regulates cell methylation balance and NAD metabolism, and extensively outline the current knowledge concerning the functions of NNMT in hepatic metabolism including glucose, lipid and energy, with a specific focus on the contribution of NNMT to the pathophysiology of liver-related diseases. NNMT is involved in the development and progression of liver diseases. Understanding the complex NNMT regulatory network and its effects on pathogenesis could provide new therapeutic strategies in the context of liver diseases.

1289

Exosomes carry and transmit signaling molecules used for intercellular communication. The generation and secretion of exosomes is a multistep interlocking process that allows simultaneous control of multiple regulatory sites. Protein molecules, mainly RAB GTPases, cytoskeletal proteins and soluble N-ethylmaleimide-sensitive fusion attachment protein receptor (SNARE), are specifically regulated in response to pathological conditions such as altered cellular microenvironment, stimulation by pathogenic factors, or gene mutation. This interferes with the smooth functioning of endocytosis, translocation, degradation, docking and fusion processes, leading to changes in the secretion of exosomes. Large numbers of secreted exosomes are disseminated by the flow of body fluids and absorbed by the recipient cells. By transmitting characteristic functional proteins and genetic information produced under disease conditions, exosomes can change the physiological state of the recipient cells and their microenvironment. The microenvironment, in turn, affects the occurrence and development of disease. Therefore, this review will discuss the mechanism by which exosome secretion is regulated in cells following the formation of mature secretory multivesicular bodies (MVBs). The overall aim is to find ways to eliminate disease-derived exosomes at their source, thereby providing an important new basis for the clinical treatment of disease.

1212

Neurexin-3 is primarily localized in the presynaptic membrane and forms complexes with various ligands located in the postsynaptic membrane. Neurexin-3 has important roles in synapse development and synapse functions. Neurexin-3 mediates excitatory presynaptic differentiation by interacting with leucine-rich-repeat transmembrane neuronal proteins. Meanwhile, neurexin-3 modulates the expression of presynaptic a-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors and g-aminobutyric acid A receptors by interacting with neuroligins at excitatory and inhibitory synapses. Numerous studies have documented the potential contribution of neurexin-3 to neurodegenerative and neuropsychiatric disorders, such as Alzheimer's disease, addiction behaviors, and other diseases, which raises hopes that understanding the mechanisms of neurexin-3 may hold the key to developing new strategies for related illnesses. This review comprehensively covers the literature to provide current knowledge of the structure, function, and clinical role of neurexin-3.

1191

Circular RNAs (circRNAs) are a novel class of noncoding RNAs that widely exist in eukaryotes. As a new focus in the field of molecular regulation, circRNAs have attracted much attention in recent years. Previous studies have confirmed that circRNAs are associated with many physiological and pathological processes. CircRNAs also participate in the regulation of stem cells. Stem cells have the properties of self-renewal and differentiation, which make stem cell therapy popular. CircRNAs may serve as new targets in stem cell therapy due to their regulation in stem cells. However, the underlying relationships between circRNAs and stem cells are still being explored. In this review, we briefly summarize the effects of circRNAs on stem cells, in the context of biological activities, aging and apoptosis, and aberrant changes. Moreover, we also examine the biological roles of stem cell-derived exosomal circRNAs. We believe our review will provide insights into the effects of circRNAs on stem cells.

1382

Gene therapy holds great promise for curing cancer by editing the deleterious genes of tumor cells, but the lack of vector systems for efficient delivery of genetic material into specific tumor sites in vivo has limited its full therapeutic potential in cancer gene therapy. Over the past two decades, increasing studies have shown that lentiviral vectors (LVs) modified with different glycoproteins from a donating virus, a process referred to as pseudotyping, have altered tropism and display cell-type specificity in transduction, leading to selective tumor cell killing. This feature of LVs together with their ability to enable high efficient gene delivery in dividing and non-dividing mammalian cells in vivo make them to be attractive tools in future cancer gene therapy. This review is intended to summarize the status quo of some typical pseudotypings of LVs and their applications in basic anti-cancer studies across many malignancies. The opportunities of translating pseudotyped LVs into clinic use in cancer therapy have also been discussed.

1223

In recent years, cardiovascular health problems are becoming more and more serious. At the same time, mechanical stimulation closely relates to cardiovascular health. In this context, Piezo1, which is very sensitive to mechanical stimulation, has attracted our attention. Here, we review the critical significance of Piezo1 in mechanical stimulation of endothelial cells, NO production, lipid metabolism, DNA damage protection, the development of new blood vessels and maturation, narrowing of blood vessels, blood pressure regulation, vascular permeability, insulin sensitivity, and maintenance of red blood cell function. Besides, Piezo1 may participate in the occurrence and development of atherosclerosis, diabetes, hypertension, and other cardiovascular diseases. It is worth noting that Piezo1 has dual effects on maintaining cardiovascular health. On the one hand, the function of Piezo1 is necessary to maintain cardiovascular health; on the other hand, under some extreme mechanical stimulation, the overexpression of Piezo1 may bring adverse factors such as inflammation. Therefore, this review discusses the Janus-faced role of Piezo1 in maintaining cardiovascular health and puts forward new ideas to provide references for gene therapy or nanoagents targeting Piezo1.

1254

N⁶-methyladenosine (m⁶A) RNA modification is widely perceived as the most abundant and common modification in transcripts. This modification is dynamically regulated by specific m⁶A "writers", "erasers" and "readers" and is reportedly involved in the occurrence and development of many diseases. Since m⁶A RNA modification was discovered in the 1970s, with the progress of relevant research technologies, an increasing number of functions of m⁶A have been reported, and a preliminary understanding of m⁶A has been obtained. In this review, we summarize the mechanisms through which m⁶A RNA modification is regulated from the perspectives of expression, posttranslational modification and protein interaction. In addition, we also summarize how external and internal environmental factors affect m⁶A RNA modification and its functions in tumors. The mechanisms through which m⁶A methylases, m⁶A demethylases and m⁶A-binding proteins are regulated are complicated and have not been fully elucidated. Therefore, we hope to promote further research in this field by summarizing these mechanisms and look forward to the future application of m⁶A in tumors.

1259

Malignant tumor is still a major problem worldwide. During tumorigenesis or tumor development, tumor suppressor p53-binding protein 2 (TP53BP2), also known as apoptosis stimulating protein 2 of p53 (ASPP2), plays a critical role in p53 dependent and independent manner. Expression of TP53BP2 is highly correlated with the prognosis and survival rate of malignant tumor patients. TP53BP2 can interact with p53, NF-κB p65, Bcl-2, HCV core protein, PP1, YAP, CagA, RAS, PAR3, and other proteins to regulate cell function. Moreover, TP53BP2 can also regulate the proliferation, apoptosis, autophagy, migration, EMT and drug resistance of tumor cells through downstream signaling pathways, such as NF-κB, RAS/MAPK, mevalonate, TGF-b1, PI3K/AKT, aPKC-i/GLI1 and autophagy pathways. As a potential therapeutic target, TP53BP2 has been attracted more attention. We review the role of TP53BP2 in tumorigenesis or tumor development and the signal pathway involved in TP53BP2, which may provide more deep insight and strategies for tumor treatment.

1261

Circular RNAs (circRNAs) are a special class of single-stranded RNA molecules with covalently closed loops widely expressed in eukaryotic organisms. CircRNAs have long been considered to play important roles in various physiological and pathological processes as non-coding RNAs. However, circRNAs have recently garnered considerable attention due to their ability to be translated into peptides/proteins via internal ribosome entry site-or N6-methyladenosine-mediated pathways or rolling translation mechanisms. Furthermore, dysregulation of translatable circRNAs and their encoded proteins has been associated with developing and progressing diseases such as cancer. This review aims to summarize the driving mechanisms of circRNA translation and the available strategies in circRNA translation research. The main focus is on the emerging biological functions of translatable circRNAs, their regulatory mechanisms, and potential clinical applications in human diseases to provide new perspectives on disease diagnosis, prognosis, and targeted therapy.

1273

Alternative splicing (AS) produces the different mRNA splicing bodies, which are then translated into multiple protein isoforms and participate in various biological functions. With a deeper understanding of alternative splicing through the study of transcriptomes using high-throughput sequencing-based methods, the correlation between aberrant AS and diseases triggered a great concern, especially abnormal AS and cancer. Medulloblastoma (MB) is an intracranial tumor in children. Sonic hedgehog MB (SHH-MB) accounted for approximately 30% of MB, which is associated with the activation of SHH signaling. Growing evidence shows that aberrant AS is closely related to the tumorigenesis of MB. Here, we briefly introduced the AS and its mechanism. Next, we described canonical/noncanonical hedgehog signaling and its correlation with MB. The main description focused on AS of various regulators in canonical hedgehog signaling in MB. In addition, we also described AS of various regulators in noncanonical hedgehog signaling. Meanwhile, activated hedgehog signaling also induces AS in MB. Then, we pointed out that aberrant AS of hedgehog signaling is associated with different MB subgroups. Finally, we summarized the therapeutic applications of targeted AS in cancer treatment. In summary, further understanding of AS in SHH-MB could develop therapeutic targets for splicing factors which may be a novel therapeutic strategy.

1177

Lactate is an end product of glycolysis. Owing to the lactate shuttle concept introduced in the early 1980s, increasing researchers indicate lactate as a critical energy source for mitochondrial respiration and as a precursor of gluconeogenesis. Lactate also acts as a multifunctional signaling molecule through receptors expressed in various cells, resulting in diverse biological consequences including decreased lipolysis, immune regulation, and anti-inflammation wound healing, and enhanced exercise performance in association with the gut microbiome. Furthermore, increasing evidence reveals that lactate contributes to epigenetic gene regulation by lactylating lysine residues of histones, which accounts for its key role in immune modulation and maintenance of homeostasis. Here, we summarize the function and mechanism of lactate and lactylation in tumor metabolism and microenvironment.

1339

NK2 genes (NKX2 gene cluster in humans) encode for homeodomain-containing transcription factors that are conserved along the phylogeny. According to the most detailed classifications, vertebrate NKX2 genes are classified into two distinct families, NK2.1 and NK2.2. The former is constituted by NKX2-1 and NKX2-4 genes, which are homologous to the Drosophila scro gene; the latter includes NKX2-2 and NKX2-8 genes, which are homologous to the Drosophila vnd gene. Conservation of these genes is not only related to molecular structure and expression, but also to biological functions. In Drosophila and vertebrates, NK2 genes share roles in the development of ventral regions of the central nervous system. In vertebrates, NKX2 genes have a relevant role in the development of several other organs such as the thyroid, lung, and pancreas. Loss-of-function mutations in NKX2-1 and NKX2-2 are the monogenic cause of the brain-lung-thyroid syndrome and neonatal diabetes, respectively. Alterations in NKX2-4 and NKX2-8 genes may play a role in multifactorial diseases, autism spectrum disorder, and neural tube defects, respectively. NKX2-1, NKX2-2, and NKX2-8 are expressed in various cancer types as either oncogenes or tumor suppressor genes. Several data indicate that evaluation of their expression in tumors has diagnostic and/or prognostic value.

1167

In Pseudomonas aeruginosa (P. aeruginosa), transcription factors (TFs) are important mediators in the genetic regulation of adaptability and pathogenicity to respond to multiple environmental stresses and host defences.The P. aeruginosa genome harbours 371 putative TFs; of these, about 70 have been shown to regulate virulence-associated phenotypes by binding to the promoters of their target genes. Over the past three decades, several techniques have been applied to identify TF binding sites on the P. aeruginosa genome, and an atlas of TF binding patterns has been mapped. The virulence-associated regulons of TFs show complex crosstalk in P. aeruginosa's regulatory network. In this review, we summarise the recent literature on TF regulatory networks involved in the quorum-sensing system, biofilm formation, pyocyanin synthesis, motility, the type III secretion system, the type VI secretion system, and oxidative stress responses.We discuss future perspectives that could provide insights and targets for preventing clinical infections caused by P. aeruginosa based on the global regulatory network of transcriptional regulators.

1224

Small nucleolar RNAs (snoRNAs) play critical roles in various biological processes. The aberrant expression or depletion of snoRNAs is related to various diseases. In previous research, most of the snoRNAs were categorized as C/D box snoRNAs and H/ACA box snoRNAs, whose typical functions were thought of as regulation of 2′-O-ribose methylation and pseudouridylation of ribosome RNAs, respectively. However, in the past two decades, studies have revealed an increasing number of snoRNAs without specific targets or determined cell functions. These findings indicated that some potential roles of snoRNAs are still unknown. Numerous studies have indicated the correlation of snoRNAs with human diseases. SnoRNAs play various roles in abundant biological processes, and they have great potential in controlling human diseases. This new and rising field could benefit from investigations of the disease pathogenesis, biomarker identification, and the determination of novel therapeutic targets. This review summarized the reports on snoRNAs and the regulation of different diseases in recent years.

1256

Accumulating evidence supports the association of somatic mutations with tumor occurrence and development. We aimed to identify somatic mutations with important implications in hepatocellular carcinoma (HCC) and explore their possible mechanisms. The gene mutation profiles of HCC patients were assessed, and the tumor mutation burden was calculated. Gene mutations closely associated with tumor mutation burden and patient overall survival were identified. In vivo and in vitro experiments were performed to verify the effects of putative genes on proliferation, invasion, drug resistance, and other malignant biological behaviors of tumor cells. Fourteen genes with a high mutation frequency were identified. The mutation status of 12 of these genes was closely related to the mutation burden. Among these 12 genes, LRP1B mutation was closely associated with patient prognosis. Nine genes were associated with immune cell infiltration. The results of in vivo and in vitro experiments showed that the knockdown of LRP1B promotes tumor cell proliferation and migration and enhances the resistance of tumor cells to liposomal doxorubicin. LRP1B could directly bind to NCSTN and affect its protein expression level, thereby regulating the PI3K/AKT pathway. Our mutational analysis revealed complex and orchestrated liposomal alterations linked to doxorubicin resistance that may also render cancers less susceptible to immunotherapy and also provides new treatment alternatives.

1412

Osteoarthritis (OA) has been considered non-reversible as articular cartilage wears down with limited repair capacity. Enhanced chondrocyte hypertrophy and increased type X collagen gene (COL10A1) expression have been associated with OA. Therefore, regulators controlling collagen X expression and chondrocyte hypertrophy may play a role in OA intervention. Here, we investigated how Distal-less homeobox 5 (DLX5), the distal-less homeobox family member, controls murine Col10a1 gene expression and chondrocyte hypertrophy in chondrogenic cell models and its role in a murine OA model. Through qRT-PCR and Western blot analyses, we detected significantly increased levels of COL10A1 and DLX5 in hypertrophic MCT and ATDC5 cells compared to their proliferative stage. Forced expression of Dlx5 further increases, while knockdown of Dlx5 decreases COL10A1 expression in hypertrophic MCT cells. We have performed dual-luciferase reporter and ChIP assays and demonstrated that DLX5 promotes reporter activity through direct interaction with Col10a1 cis-enhancer. We established a murine OA model and detected markedly increased COL10A1 and DLX5 in the articular cartilage and subchondral bone of the OA mice compared with the controls. Notably, forced overexpression of DLX5 in hypertrophic MCT cells up-regulates RUNX2, and adjacent DLX5 and RUNX2 binding sites have previously been found within the Col10a1 cis-enhancer. Together, our data suggest that DLX5 may cooperate with RUNX2 to control cell-specific Col10a1 expression and chondrocyte hypertrophy and is involved in OA pathogenesis.

1278

This study aims to identify the inflammatory factor-related genes which help to predict the prognosis of patients with colorectal cancer. GSEA (Gene Set Enrichment Analysis) was used to acquire inflammation-related genes and the corresponding expression information was collected from TCGA database to determine the DEGs (differentially-expressed genes) in CRC patients. We conducted enrichment analysis and PPI (proteineprotein interaction) of these DEGs. Besides, key genes that are both differentially-expressed and prognosis-related were screened out, which were used to establish the prognostic model. We obtained 79 DEGs and 19 prognostic genes, 10 prognostic-related differential genes were eventually screened. These genes were used to construct the prognostic model. We also identified that the immune infiltration score of macrophages between different risk groups was significantly different and similar distinction was witnessed in immune function score of APC (antigen-presenting cell) costimulation and type I IFN (interferon) response.

1348

Osteosarcoma is a common malignant tumor occurring in children and young adults. Chondroitin sulfate (CS) participates in cell adhesion, cell division, and the formation of neural networks in the body, the biosynthesis of which requires the participation of glycosyltransferases. CHPF, a glycosyltransferase, plays a role in the extension of CS. Recently, CHPF's biological roles and functional importance in human diseases including malignant tumors have been widely discussed. However, whether CHPF is involved in osteosarcoma development and growth has not been revealed. The present work aimed to investigate the expression levels, functional significance and molecular mechanism of CHPF in osteosarcoma progression. Our results revealed that CHPF is strongly expressed in osteosarcoma tissues and cells. Furthermore, CHPF serves as a tumor promoter in the development and progression of osteosarcoma through enhancing cell proliferation and migration while suppressing apoptosis. Exploration of the mechanism by which CHPF promotes osteosarcoma indicated that CHPF promotes osteosarcoma through counteracting SKP2's ubiquitination and activating the Akt signaling pathway. For the first time, we clarified the roles of CHPF in osteosarcoma, and our results suggested that CHPF might be a novel therapeutic target in the treatment strategies for osteosarcoma.

1323

Aptamers, short single DNA or RNA oligonucleotides, have shown immense application potential as molecular probes for the early diagnosis and therapy of cancer. However, conventional cell-SELEX technologies for aptamer discovery are time-consuming and laborious. Here we discovered a new aptamer BC-3 by using an improved rapid X-Aptamer selection process for human bladder carcinoma, for which there is no specific molecular probe yet. We show that BC-3 exhibited excellent affinity in bladder cancer cells but not normal cells. We demonstrate that BC-3 displayed high selectivity for tumor cells over their normal counterparts in vitro, in mice, and in patient tumor tissue specimens. Further endocytosis pathway analysis revealed that BC-3 internalized into bladder cancer cells via clathrin-mediated endocytosis. Importantly, we identified ribosomal protein S7 (RPS7) as the binding target of BC-3 via an integrated methodology (mass spectrometry, colocalization assay, and immunoblotting). Together, we report that a novel aptamer BC-3 is discovered for bladder cancer and its properties in the disease are unearthed. Our findings will facilitate the discovery of novel diagnostic and therapeutic strategies for bladder cancer.

1349

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have attracted attention in the field of regenerative medicine due to their potential ability to repair damaged hearts. However, the immature phenotype of these cells limits their clinical application. Cardiomyocyte maturation is accompanied by changes in mitochondrial quality. PTEN-induced putative kinase 1 (PINK1) has been linked to mitochondrial quality control. However, whether the changes in mitochondrial quality in hiPSC-CMs are associated with PINK1, and the impact of PINK1 on hiPSC-CMs development are not clear. In this study, we found that knockdown of PINK1 in hiPSC-CMs resulted in mitochondrial fragmentation and impaired mitochondrial functions such as mitophagy and mitochondrial biogenesis. PINK1 deletion also inhibited the maturation of hiPSC-CMs, reverting them to a naive structural and functional state. We found that restoring the mitochondrial structure did not completely rescue the mitochondrial dysfunction caused by PINK1 deletion, while activation of PINK1 kinase activity using kinetin promoted mitochondrial fusion, increased the mitochondrial membrane potential and ATP production, and maintained the development and maturation of hiPSC-CMs. In conclusion, PINK1 regulates the mitochondrial structure and function of hiPSC-CMs, and is essential for the maturation of hiPSC-CMs.

1223

Despite significant improvements in five-year survival rates due to early diagnosis and combination therapy, triple-negative breast cancer (TNBC) treatment remains a major challenge. Finding new and effective targets for diagnosis and drug therapy is urgent for TNBC patients. Jagged-1 (JAG1), one of the canonical ligands of the Notch signaling pathway, is involved in vascular budding and is a poor prognostic factor of TNBC. In this study, combined with quantitative real-time PCR, database analysis, animal experiments, and other means, JAG1 was confirmed to be related to the poor prognosis of TNBC patients. JAG1 was highly expressed in MDA-MB-231 Bone (231B) cells, with stronger invasion and metastasis ability than MDA-MB-231 (231) cells. Treatment of human vascular endothelial cells (HUVEC) with TNBC conditioned medium showed that TNBC JAG1 promoted the angiogenesis of HUVEC. Next, we detected the exosomes extracted from TNBC conditioned medium and found that JAG1 promoted the exosome secretion from 231 cells via ALIX-RAB11A/RAB35. In addition, we also found that the exosomes from JAG1 overexpressed TNBC cells contained more long non-coding RNA (lncRNA) MALAT1, and MALAT1 promoted angiogenesis of HUVEC by targeting miR-140-5p. Finally, the angiogenesis-promoting effect of JAG1 in TNBC was further investigated by matrix gel assay. In conclusion, we reveal that JAG1 has a pro-invasion effect on TNBC and is involved in microenvironment angiogenesis by promoting exosome secretion and the MALAT1-miR-140-5p-JAG1/VEGFA pathway.

1321

Uveitis, a vision-threatening inflammatory disease worldwide, is closely related to resident microglia. Retinal microglia are the main immune effector cells with strong plasticity, but their role in uveitis remains unclear. N6-methyladenosine (m⁶A) modification has been proven to be involved in the immune response. Therefore, we in this work aimed to identify the potentially crucial m⁶A regulators of microglia in uveitis. Through the single-cell sequencing (scRNA-seq) analysis and experimental verification, we found a significant decrease in the expression of fat mass and obesity-associated protein (FTO) in retinal microglia of uveitis mice and human microglia clone 3 (HMC3) cells with inflammation. Additionally, FTO knockdown was found to aggravate the secretion of inflammatory factors and the mobility/chemotaxis of microglia. Mechanistically, the RNA-seq data and rescue experiments showed that glypican 4 (GPC4) was the target of FTO, which regulated microglial inflammation mediated by the TLR4/NF-κB pathway. Moreover, RNA stability assays indicated that GPC4 upregulation was mainly regulated by the downregulation of the m⁶A "reader" YTH domain family protein 3 (YTHDF3). Finally, the FTO inhibitor FB23-2 further exacerbated experimental autoimmune uveitis (EAU) inflammation by promoting the GPC4/TLR4/NF-κB signaling axis, and this could be attenuated by the TLR4 inhibitor TAK-242. Collectively, a decreased FTO could facilitate microglial inflammation in EAU, suggesting that the restoration or activation of FTO function may be a potential therapeutic strategy for uveitis.

1519