Abnormal lactate metabolism and lysine lactylation (Kla) have emerged as critical factors driving tumor immune evasion through dual mechanisms of tumor metabolic reprogramming and epigenetic regulation. Tumor cells produce lactate through multiple metabolic pathways, which is then exported via monocarboxylate transporters to accumulate in the tumor microenvironment (TME). Within the TME, lactate acts as a key substrate driving the Kla of both histone and non-histone proteins. Lactate and Kla synergistically drive tumor immune evasion by acting on both tumor cells and immune cells, ultimately driving tumor initiation, proliferation, metastasis, and therapeutic resistance. Currently, therapeutic strategies targeting lactate metabolism have emerged as a research hotspot for breaking tumor immune evasion, which encompass three directions: inhibiting lactate production, blocking lactate transport, and reversing abnormal Kla. Some candidate drugs have entered clinical trials and demonstrated preliminary efficacy, yet challenges remain, including insufficient specificity, off-target toxicity, and resistance mediated by tumor metabolic plasticity. Therefore, this review systematically summarizes the mechanisms of lactate production and transport within the TME, the regulation of Kla, and their role in mediating tumor immune evasion. Anti-tumor therapeutic strategies and recent research advances targeting lactate metabolism are also clarified, which provide a theoretical basis for elucidating the metabolic mechanisms of tumor immune evasion and developing novel metabolic-immune combination therapies.

Background:Increasing evidence indicates that tumor cellular senescence can impair antitumor immunity and promote skin cutaneous melanoma (SKCM) progression. However, effective methods for assessing tumor cellular senescent status and tumor immune microenvironment (TIME) status remain lacking. This study intends to establish a novel Senescence-TIME Risk Score (STIRS) based on senescence and TIME related genes to predict prognosis and immunotherapy responsiveness in SKCM patients, thereby providing new strategies for current clinical personalized treatment. Methods:We identified distinct senescent microenvironment patterns using t-distributed stochastic neighbor embedding (t-SNE) based on a set of senescence marker genes and predicted the TIME in SKCM using the estimation of stromal and immune cells in malignant tumor tissues using expression data (ESTIMATE) algorithm. Based on this, we divided the SKCM cohort into three groups: low-senescence & high-immunity, high-senescence & low-immunity, and mixed. We analyzed differentially expressed genes (DEGs) between the first two groups. Gene ontology (GO) enrichment analysis, kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis, gene set enrichment analysis (GSEA), and the construction of protein-protein interaction (PPI) network were used to investigate the functional relevance of DEGs. We screened DEGs using least absolute shrinkage and selection operation (LASSO) regression and random forest (RF) algorithm to construct the STIRS. Patients were grouped by the median of STIRS, and differences in expression of immune cells and immune checkpoints between groups were examined. The predictive capability of STIRS for immunotherapy was validated. Finally, we knocked down the core risk gene in the B16 cell line to validate its function. Results:We identified 994 DEGs predominantly enriched in TIME- and senescence-related pathways. The constructed STIRS comprises six signature genes. Patients in the high-STIRS group exhibited significantly poorer survival than those in the low-STIRS group, and STIRS negatively correlated with immune response and immunotherapy responsiveness. A nomogram integrating STIRS and clinical indicators demonstrated satisfactory predictive performance for SKCM patient prognosis. These findings validate the STIRS model as a reliable independent prognostic indicator. Additionally, knockdown of the core risk gene keratin 17 (KRT17) inhibited the invasion and proliferation of B16 cells, demonstrating the role of KRT17 in the progression of SKCM. Conclusion:This study proposed a novel STIRS model and selected the core risk gene KRT17 for functional validation, which had potential as a prognostic tool and a guide for creating personalized therapies for SKCM patients.

RNA binding proteins could regulate transcription through modulating RNA polymerase II involved transcriptional condensates with the aid of nascent RNA. However, whether this mechanism also contributes to tumorigenesis is still unknown. Here, we show the RBP AKAP95 is required for MLL rearranged leukemogenesis. Phase separation and RNA binding properties of AKAP95 modulate Pol II recruitment into condensates at genome-wide target sites. AKAP95 interact with MLL1 translocated fragment and their partial co-condensation leads to stronger AKAP95 binding towards MLL-AF9 target genes. Loss of AKAP95 significantly downregulates expression of MLL-AF9 targets and impairs MLL-AF9 driven leukemogenesis. AKAP95 is not essential for normal and regenerative hematopoiesis, rendering it an ideal target for leukemia therapeutics. As an endeavor for modifying condensates, we design a peptide JD-PI95 bridging AKAP95 to protein quality surveillance component HSP70. The peptide successfully impairs AKAP95 phase separation, attenuates gene transcription and inhibits MLLr leukemia cells proliferation. Our work demonstrates an important role of RBP condensates in tumorigenesis through regulation of transcription and a new method to perturb specific condensates for tumor inhibition. The role of RNA-binding protein (RBP) phase separation in transcriptional regulation and its contribution to tumorigenesis remains to be characterised. Here, the authors show that RBP AKAP95 physically co-condenses with the mixed lineage leukemia 1 (MLL1) translocated fragment commonly found in MLL rearranged (MLLr) leukemia to promote leukemogenesis.

BACKGROUND:The comparative prognostic performance of lymph node-related prognostic parameters-positive lymph nodes, log odds of positive lymph nodes, and lymph node ratio-in duodenal adenocarcinoma remains undefined. This study aims to compare their predictive performance in both Western and Eastern cohorts and develops a prognostic nomogram based on the optimal indicator. METHODS:Data were obtained from the Surveillance, Epidemiology, and End Results (N = 1,693) database and Wuhan Union Hospital (N = 295). Using 7 evaluation indicators, this study assessed the prognostic performance of 3 lymph node-related prognostic parameters in both their continuous and categorical form. A nomogram was constructed based on the optimal lymph node-related prognostic parameter and independent prognostic factors. External validation was carried out using the Wuhan Union Hospital cohort. RESULTS:Among continuous lymph node-related prognostic parameters, standardized log odds of positive lymph nodes demonstrated superior predictive accuracy in both Surveillance, Epidemiology, and End Results cohort (c-index: 0.639; 5-year area under the curve: 0.685) and Wuhan Union Hospital cohort (c-index: 0.679; 5-year area under the curve: 0.697). Similarly, its categorical counterpart, log odds of positive lymph nodes classification, also exhibited robust prognostic discrimination (Surveillance, Epidemiology, and End Results c-index: 0.619, 5-year area under the curve: 0.666; Wuhan Union Hospital c-index: 0.637, 5-year area under the curve: 0.640). The nomogram, incorporating log odds of positive lymph nodes classification, age, T-stage, retrieved lymph nodes, positive lymph nodes, and chemotherapy, demonstrated an area under the curve of 0.735, 0.708, and 0.698 for 1-, 3-, and 5-year predictions, respectively, in the Surveillance, Epidemiology, and End Results cohort, and 0.734, 0.715, and 0.691, respectively, in the Wuhan Union Hospital cohort. Calibration curves and decision curve analysis confirmed the model's accuracy and clinical utility. The online tool facilitates clinical use. CONCLUSION:Log odds of positive lymph nodes classification is identified as the most powerful lymph node-related prognostic parameter for prognostic prediction in duodenal adenocarcinoma. The proposed nomogram, incorporating log odds of positive lymph nodes classification, provides an accurate and clinically useful tool for individualized survival estimation.

BACKGROUND:Aphids are one of the most damaging piercing-sucking pests in rice production, whose population outbreaks cause crop losses amounting to hundreds of millions of dollars annually worldwide. To address this challenge, it is highly imperative to develop aphid-resistant rice varieties through techniques such as genetic modification. RESULTS:In this study, we successfully targeted the (E)-β-farnesene synthase (EβFS) to the chloroplast using the promoter of the rice Rubisco small subunit gene (rbcS) and its chloroplast transit peptide (ctp), which significantly enhanced the production of (E)-β-farnesene (EβF) in transgenic rice. At the seedling stage, the EβFS protein content was 17.6-fold higher in transgenic rice compared with that in the rice previously generated using the cauliflower mosaic virus 35S promoter (35S::EβFS). At the tillering stage, the EβF release rate reached 12.7 μg per plant per day in the transgenic rice. Olfactory behavioral assays demonstrated that the transgenic rice line generated in this study had significantly enhanced ability to repel bird cherry-oat aphids (Rhopalosiphum padi) ('Push' effect) and attract its natural enemy multicolored Asian lady beetles (Harmonia axyridis) ('Pull' effect) compared with the 35S::EβFS lines. In addition, the rbcS promoter can achieve specific expression of the EβFS gene in photosynthetic organs, avoiding unnecessary consumption of energy and resources associated with constitutive expression. CONCLUSION:This Push-Pull strategy could protect rice plants from aphid infestation by controlling the aphid populations, providing an ecologically friendly and sustainable approach for integrated pest management in rice fields. © 2025 Society of Chemical Industry.

Cancer cells are characterized by their altered energy metabolism. A hallmark of cancer metabolism is aerobic glycolysis, also called the Warburg effect. Hexokinase 2 (HK2), a crucial glycolytic enzyme converting glucose to glucose-6-phosphate, has been identified as a central player in the Warburg effect. Deletion of HK2 decreases cancer cell proliferation in animal models without explicit side effects, suggesting that targeting HK2 is a promising strategy for cancer therapy. In this study, we discovered a correlation between HK2 and the tumor immune response in triple-negative breast cancer. Inhibition of HK2 led to a reduction in G-CSF expression in 4T1 cells and a decrease in the development of myeloid-derived suppressor cells which, in turn, enhanced T cell immunity and prolonged the survival of 4T1 tumor-bearing mice. Furthermore, the HK2 inhibitor 3-BrPA improved the therapeutic efficacy of anti-PD-L1 therapy in 4T1 tumor-bearing mouse models. This study highlights the potential of glycolysis-targeting interventions as a novel treatment strategy, which can be combined with immunotherapy for the treatment of triple-negative breast cancer.

This study aims to evaluate bowel and urogenital function in patients with locally advanced low rectal cancer treated with immunotherapy combined with neoadjuvant therapy followed by surgery, compared to surgery alone. We conducted a retrospective analysis of 162 patients with locally advanced low rectal cancer treated at Union Hospital, affiliated with Tongji Medical College, Huazhong University of Science and Technology, from May 2021 to May 2024. Seventy-three patients underwent short-course radiotherapy combined with chemotherapy and immunotherapy followed by surgery (SCRT-CIT group), while 89 patients received surgery alone (non-SCRT-CIT group). Standardized questionnaires assessed bowel and urogenital function. Logistic regression analysis was used to identify independent predictors of functional outcomes. The incidence of major low anterior resection syndrome (LARS) was significantly higher in the SCRT-CIT group (59.0

Oxidative stress has been implicated as a pivotal factor in the pathogenesis of numerous malignancies. However, the association between oxidative stress and the prognosis of patients with colorectal cancer liver metastasis (CRCLM) is not well elucidated. We conducted a retrospective analysis of 424 patients with CRCLM who underwent primary resection at the Union Hospital, Tongji Medical College, Wuhan, between July 2013 and September 2018. Patients were randomly divided into a training set (n = 300) and a test set (n = 124) in a 7:3 ratio.To develop the CRCLM-integrated Oxidative Stress Score (CLIOSS) for CRCLM, we utilized individual oxidative stress markers. The overall survival (OS) and disease-free survival (DFS) were estimated using the Kaplan-Meier method, and the log-rank test was employed to assess prognostic factors.To validate the predictive performance of CLIOSS, we constructed Receiver Operating Characteristic (ROC) curves and calculated the Area Under the Curve (AUC) values. Additionally, Decision Curve Analysis (DCA) and calibration plots were used to evaluate the clinical utility and predictive consistency of the model. The CLIOSS prognostic model was constructed based on the following variables and their corresponding β coefficients: 0.042 × total bilirubin (TBIL, µmol/L) + 0.032 × blood urea nitrogen (BUN, mmol/L) − 0.001 × uric acid (UA, µmol/L). Higher CLIOSS were associated with poorer OS (2.934;95% CI 2.167–3.974;P < 0.001) and DFS(2.707; 95% CI 2.000–3.664; P < 0.001. The 3-year OS AUC values were 0.803 in the training set and 0.851 in the testset, while the 3-year DFS AUC values were 0.892 and 0.898, respectively. Decision curve analysis demonstrated that both predictive models have significant clinical utility in practice. CLIOSS is a comprehensive prognostic index derived from oxidative stress markers, designed to predict long-term survival in patients with CRCLM. Our study shows that higher CLIOSS are significantly associated with poorer outcomes, making it an important tool for assessing patient risk.

The transition from acute kidney injury (AKI) to chronic kidney disease (CKD) remains a critical clinical challenge with limited therapeutic options. To our knowledge, this was the first study to investigate the renoprotective effects of silibinin, a flavonolignan from Silybum marianum, in mitigating AKI-to-CKD progression via modulation of MAPK and PI3K/AKT signaling pathways-addressing a critical unmet need, as no targeted therapies currently exist to block this transition. For in vivo evaluation, a mice renal ischemia-reperfusion injury (IRI) model was established, where 45 min of ischemia was followed by reperfusion for 1 day (AKI) or 14 days (CKD) across experimental groups. Human renal proximal tubular (HK-2) cells were utilized for in vitro modeling, including hypoxia/reoxygenation (H/R) model and TGF-β1-induced fibrosis. Bioinformatics analysis was employed for target prediction of silibinin. Results showed that silibinin significantly decreased serum creatinine, blood urea nitrogen, oxidative stress, inflammation, and apoptosis induced by IRI. The HE, Masson and Sirius Red staining showed that silibinin decreased the kidney damage and collagen deposition dramatically. Mechanistically, silibinin enhanced PI3K/AKT phosphorylation, suppressed phosphorylation of MAPK components (p38, ERK1/2, JNK), elevated antioxidant enzyme activity, reduced ROS/malondialdehyde levels, inhibited pro-inflammatory cytokine release, downregulated Bax/cleaved caspase-3 expression, and upregulated the anti-apoptotic factor Bcl-2. These effects were recapitulated in vitro, where silibinin mitigated H/R and TGF-β1-driven cellular injury by restoring redox balance and modulating dual signaling axes. Collectively, our findings demonstrated silibinin as a multi-target therapeutic candidate that impedes AKI-CKD transition via coordinated regulation of PI3K/AKT and MAPK pathways. These findings position silibinin as a promising agent for clinical intervention, potentially improving long-term renal outcomes in AKI patients at risk of progressing to CKD.

Lymphovascular invasion (LVI) is associated with poor prognosis in a variety of malignancies; however, its prognostic value has not been fully defined in patients with colorectal cancer with liver metastases (CRCLM). The aim of this study was to investigate the impact of LVI on long-term postoperative recurrence and survival in patients with CRCLM. Clinicopathologic data were retrospectively collected from patients who underwent primary resection for CRCLM at Wuhan Union Hospital from 2013 to 2018. To reduce potential confounders and selection bias, we used propensity score matching (PSM) to compare the clinicopathologic characteristics and long-term prognostic outcomes of patients in the LVI (+) and LVI (-) groups. Cox unifactorial and multifactorial analyses were used to screen relevant factors affecting patient prognosis, and Kaplan-Meier curves were plotted to compare differences in patient overall survival (OS) and disease-free survival (DFS). The predictive power of independent factors on patients’ long-term prognosis was assessed using receiver operating characteristic ROC) curves and area under the curve (AUC). After PSM, 230 patients were enrolled in the study (n = 115 per group). Multifactorial analysis revealed that LVI was an independent prognostic factor for OS and DFS (hazard ratio [HR], 1.424; 95

Terpene synthases (TPSs) are key enzymes in terpenoids synthesis of plants and play crucial roles in regulating plant defence against pests and diseases. Here, we report the functional characterization of OsTPS19 and OsTPS20, which were upregulated by the attack of brown planthopper (BPH). BPH female adults performed concentration-dependent behavioural responses to (S)-limonene showing preference behaviour at low concentrations and avoidance behaviour at high concentrations. Overexpression lines of OsTPS19 and OsTPS20, which emitted higher amounts of the monoterpene (S)-limonene, decreased the hatching rate of BPH eggs, reduced the lesion length of sheath blight caused by Rhizoctonia solani and bacterial blight caused by Xanthomonas oryzae. While knockout lines of OsTPS19 and OsTPS20, which emitted lower amounts of (S)-limonene, were more susceptible to these pathogens. Overexpression of OsTPS19 and OsTPS20 in rice plants had adverse effects on the incidence of BPH, rice blast, and sheath blight in the field and had no significant impacts on rice yield traits. OsTPS19 and OsTPS20 were found to be involved in fine-tuning the emission of (S)-limonene in rice plants and play an important role in defence against both BPH and rice pathogens.

BACKGROUND:Immunotherapy for gastric cancer remains a challenge due to its limited efficacy. Metabolic reprogramming toward glycolysis has emerged as a promising avenue for enhancing the sensitivity of tumors to immunotherapy. Pyruvate dehydrogenase kinases (PDKs) play pivotal roles in regulating glycolysis. The importance of PDKs in the context of gastric cancer immunotherapy and their potential as therapeutic targets have not been fully explored. METHODS:PDK and PD-L1 expression was analyzed using data from the GSE66229 and The Cancer Genome Atlas (TCGA) cohorts. Additionally, the Immune Checkpoint Blockade Therapy Atlas (ICBatlas) database was utilized to assess PDK expression in an immune checkpoint blockade (ICB) therapy group. Subsequently, the upregulation of PD-L1 and the enhancement of anticancer effects achieved by targeting PDK were validated through in vivo and in vitro assays. The impact of PDK on histone acetylation was investigated using ChIP‒qPCR to detect changes in histone acetylation levels. RESULTS:Our analysis revealed a notable negative correlation between PD-L1 and PDK expression. Downregulation of PDK led to a significant increase in PD-L1 expression. PDK inhibition increased histone acetylation levels by promoting acetyl-CoA generation. The augmentation of acetyl-CoA production and concurrent inhibition of histone deacetylation were found to upregulate PD-L1 expression in gastric cancer cells. Additionally, we observed a significant increase in the anticancer effect of PD-L1 antibodies following treatment with a PDK inhibitor. CONCLUSIONS:Downregulation of PDK in gastric cancer cells leads to an increase in PD-L1 expression levels, thus potentially improving the efficacy of PD-L1 immune checkpoint blockade therapy.

Alteration in lipid metabolism is recognized as a hallmark feature of colorectal cancer (CRC). Protein S-palmitoylation plays a critical role in many different cellular processes including protein-lipid interaction. Zinc Finger DHHC-Type Containing 1 (ZDHHC1, also known as ZNF377) belongs to the palmitoyl-transferase ZDHHC family, and is a potential tumor suppressor. However, our knowledge of the functional roles of ZDHHC1 in CRC is limited. We discovered that ZDHHC1 expression was downregulated in CRC tissues and that low levels of ZDHHC1 were associated with unfavorable prognosis. Functional studies showed that ZDHHC1 inhibited CRC cell proliferation and invasion in vitro and in vivo. We also found that lipase G (LIPG) is negatively regulated by ZDHHC1 and plays a key role in CRC cell growth through lipid storage. Additionally, we demonstrated that ZDHHC1 functions as a IGF2BP1-palmitoylating enzyme that induces S-palmitoylation at IGF2BP1-C337, which results in downregulated LIPG expression via m6A modification. Mechanistic investigations revealed that the ZDHHC1/IGF2BP1/LIPG signaling axis is associated with inhibition of CRC cell growth. Our study uncovers the potential role of ZDHHC1 in CRC, including inhibition of CRC growth by reducing the stability of LIPG mRNA in an m6A dependent-manner by palmitoylation of IGF2BP1.

Glioblastoma (GBM) is one of the most malignant tumors of the central nervous system. The pattern of immune checkpoint expression in GBM remains largely unknown. We performed snRNA-Seq and spatial transcriptomic (ST) analyses on untreated GBM samples. 8 major cell types were found in both tumor and adjacent normal tissues, with variations in infiltration grade. Neoplastic cells_6 was identified in malignant cells with high expression of invasion and proliferator-related genes, and analyzed its interactions with microglia, MDM cells and T cells. Significant alterations in ligand-receptor interactions were observed, particularly between Neoplastic cells_6 and microglia, and found prominent expression of VISTA/VSIG3, suggesting a potential mechanism for evading immune system attacks. High expression of TIM-3, VISTA, PSGL-1 and VSIG-3 with similar expression patterns in GBM, may have potential as therapeutic targets. The prognostic value of VISTA expression was cross-validated in 180 glioma patients, and it was observed that patients with high VISTA expression had a poorer prognosis. In addition, multimodal cross analysis integrated SnRNA-seq and ST, revealing complex intracellular communication and mapping the GBM tumor microenvironment. This study reveals novel molecular characteristics of GBM, co-expression of immune checkpoints, and potential therapeutic targets, contributing to improving the understanding and treatment of GBM.

To the Editor: Salivary adenoid cystic carcinoma (SACC) is characterized by the high frequency of perineural invasion (PNI) behavior during tumor progression. However, the impact of intercellular communication between SACC cells and surrounding non-malignant cells on PNI remains unclear. As the main constituent cells of the peripheral nervous system, Schwann cells have been reported to be closely associated with the process of PNI. Cancer cell-derived exosomes often serve as important signaling mediators between cancer cells and the local microenvironment. Therefore, this study aimed to elucidate the reciprocal interplay between cancer cells and Schwann cells via exosomes during cancer PNI and to identify potential therapeutic target for PNI. This research was approved by the Institutional Ethical Committee of West China Hospital of Stomatology (No. WCHSIRB-D-2022-020). The animal experiments were approved by the Institutional Animal Care and Use Committee of the West China Hospital, Sichuan University (No. 20211282A). To explore the role of the interaction of tumor exosomes and Schwann cells in the occurrence of PNI, the exosomes from the conditioned medium (CM) of SACC cells including SACC-83 and SACC-LM were isolated and identified as described before.[1] To enable observation of the delivery of exosomes, the SACC cells and Schwann cells (RSC96) were infected with CD63-red fluorescence protein (RFP) lentivirus and green fluorescence protein (GFP) lentivirus, respectively. Then, incubated exosomes and transfected Schwann cells, and used the confocal microscopy to observe whether the exosomes derived from SACC cells could be delivered to the Schwann cells. Transwell migration and invasion assays, CCK8 viability and flow cytometry apoptosis assays were conducted to determine the effect of Schwann cells educated by exosomes on SACC cells. To investigate how tumor-derived exosomes mediate Schwann cells to regulate SACC cells behaviors, microarray analysis of micro RNAs (miRNAs) in exosomes from SACC cells and HSG cells was performed, and species homology analysis and expression screening were also performed.[2] Then, the mimics of selected miRNAs were separately transfected into Schwann cells to evaluate the role of the miRNA on SACC. Besides, wound healing, cell invasion, cell viability, and apoptosis assays were performed to investigate the role of exosomal miRNAs, following transfection of Schwann cells with either the inhibitor or mimics of miRNA. To elucidate the role of exosomal miRNA in SACC PNI in vivo, a sciatic nerve graft tumor model was used. The sciatic nerve was treated with exosomal miRNA agomir, and the expression of miRNA in the sciatic nerve was detected by quantitative polymerase chain reaction (qPCR) assay. Then, SACC-LM cells were transplanted around the nerve to establish the PNI model. The function of the sciatic nerve was evaluated by recording the distance (mm) from the first toe to the fifth toe of the hind limb paw and calculating the sciatic nerve score.[3] In addition, an invasive dorsal root ganglion (DRG) model was constructed. DRGs from rats were cultured in the Matrigel for two days and stimulated with miRNA agomir. Then, SACC-LM cells were seeded far away from the DRG and the cancer cells invaded the Matrigel were calculated for quantitative analysis. Transcriptome sequencing and gene set enrichment analysis (GSEA) were performed to unravel the mechanism by which exosomal miRNA reprograms Schwann cells to promote SACC invasion. Then, qPCR and Western blot assays were conducted to detect the expression of related genes. In addition, an inhibitor of related gene was used to prove the function of identified gene. Reagents, consumables, and instruments used are shown in Supplementary Table 1, https://links.lww.com/CM9/C114. GraphPad Prism 8.0 software (https://www.graphpad.com/) was used for data statistical analysis. Differences were considered significant if P value was less than 0.05. Data were presented as mean ± standard deviation. The results of cup-shaped structure, size of the isolated particles, and the characteristic of proteins such as CD9, CD63, ALIX, and TSG101 indicated exosomes of SACC cells and HSG cells were successfully extracted [Supplementary Figure 1, https://links.lww.com/CM9/C114]. The virus transfection results showed that SACC cells and Schwann cells were infected with RFP and GFP, respectively [Supplementary Figure 2A, https://links.lww.com/CM9/C114]. Then, the exosomes of SACC cells were also labeled with RFP. Confocal images showed that red exosomes could be delivered into the Schwann cells by co-culture [Supplementary Figure 2B, https://links.lww.com/CM9/C114]. As for the outcomes of function experiments, Schwann cells educated by SACC exosomes could promote SACC cell migration and invasion [Supplementary Figure 2C, https://links.lww.com/CM9/C114], but not in SACC cell proliferation or apoptosis [Supplementary Figure 3, https://links.lww.com/CM9/C114]. Microarray analysis of exosomes of SACC cells and HSG cells showed that a total of 153 miRNAs were upregulated in SACC-83 and SACC-LM cell-derived exosomes [Supplementary Figure 4A, B, https://links.lww.com/CM9/C114]. By species homology analysis and expression screening, six miRNAs were identified [Supplementary Figure 4C, D, https://links.lww.com/CM9/C114]. Migration assays indicated that among six miRNAs, only exosomal miR-130b-3p obviously promoted SACC migration by regulating Schwann cells [Supplementary Figure 5, https://links.lww.com/CM9/C114]. When Schwann cells were transfected with mimic of miR-130b-3p, the results of SACC wound healing, cell invasion, cell viability, and apoptosis assays were consistent with those obtained for the exosomes [Figure 1A, B and Supplementary Figure 6A, B, https://links.lww.com/CM9/C114]. Then, Schwann cells were stably transfected with miR-130b-3p inhibitor, as expected, the effects of Schwann cell CM on SACC migration and invasion were abolished by miR-130b-3p inhibitor [Figure 1C, D and Supplementary Figure 6C, https://links.lww.com/CM9/C114], which indicates that SACC cell-derived miR-130b-3p medicates the ability of Schwann cells to regulate cell migration and invasion of SACC cells.Figure 1: The function verification of exosomal miR-130b-3p of SACC cells. (A, B) Wound healing (A) and invasion assay (B) of SACC cells treated by the CM of Schwann cells transfected miR-130b-3p mimic or normal control. (C, D) Wound healing (C) and invasion assay (D) of SACC cells treated by the CM of Schwann cells transfected miR-130b-3p inhibitor or normal control. Migrated cells and distance were counted and representative images were shown. Scale bar of wound healing, 200 μm. Scale bar of invasion assay, 100 μm. * P <0.05; † P <0.01, ‡ P <0.001. SACC: Salivary adenoid cystic carcinoma; CM: Conditioned medium; NC: Negative control.For the experiments in vivo, the expression of miR-130b-3p in the sciatic nerve treated with miR-130b-3p agomir showed that the expression of miR-130b-3p was upregulated [Supplementary Figure 7A, https://links.lww.com/CM9/C114]. Then, SACC-LM cells were transplanted around the nerve to establish the PNI model [Supplementary Figure 7B, C, https://links.lww.com/CM9/C114]. Based on the volume of the tumor, no statistically significant difference was found between the negative control group and the experimental group (miR-130b-3p agomir pre-treatment) [Supplementary Figure 7D, https://links.lww.com/CM9/C114]. However, the mice pre-treated with the miR-130b-3p agomir had significantly lower sciatic nerve index and scores than did the control group [Supplementary Figure 7E, https://links.lww.com/CM9/C114]. Besides, the DRG treated by miR-130b-3p agomir significantly increased the number of migrated SACC cells [Supplementary Figure 7F, https://links.lww.com/CM9/C114]. These results demonstrated that the pre-treatment of nerves with the miR-130b-3p agomir contributed to the progression of SACC. The GSEA results showed that compared with the CM of control cells (CM-mi-nc), the CM of Schwann cells transfected with miR-130b-3p mimics (CM-mi-130b) significantly affected cholesterol homeostasis and the expression of related genes in SACC-LM cells [Supplementary Figure 7G, https://links.lww.com/CM9/C114]. The results of qPCR showed that Proprotein convertase subtilisin/kexin type 9 (PCSK9) and the low-density lipoprotein receptor (LDLR) were significantly upregulated [Supplementary Figure 8A, https://links.lww.com/CM9/C114]. Therefore, we transfected the Schwann cells with miR-130b-3p inhibitors to further prove the changed expression of genes. The results of qPCR showed that PCSK9 and LDLR were significantly downregulated as expected [Supplementary Figure 8B, https://links.lww.com/CM9/C114]. Then, Western blot assays were performed, which showed high expression level of PCSK9 in SACC cells treated with the CM-mi-130b [Supplementary Figure 8C, https://links.lww.com/CM9/C114]. To prove that whether PCSK9 is the key functional gene that leads to the increased invasion ability of SACC cells, a PCSK9 inhibitor was used to inhibit its expression. The following three groups were compared: SACC + CM-mi-nc vs. SACC + CM-mi-130b vs. SACC + CM-mi-130b + PCSK9 inhibitor. The results of the Western blot and migration assays showed that the enhanced migration ability induced by CM-mi-130b was associated with the expression of PCSK9 [Supplementary Figure 8D, E, https://links.lww.com/CM9/C114]. This study indicated that SACC cell-derived exosomal miR-130b-3p could educate and reprogram Schwann cells to affect the PNI of SACC by regulating PCSK9. However, there are some limitations to this study. For example, SACC cell-derived exosomes medicated Schwann cells to form a pre-metastatic niche, but the changes in Schwann cells were not further explored. Similarly, the association between PCSK9 and cholesterol homeostasis in SACC cells was just predicted by sequencing analysis, but a more detailed mechanism needs to be fully elucidated in future studies. In summary, the results of this study revealed the role of miR-130b-3p derived from SACC exosomes in the progression of PNI from the perspective of the perineural microenvironment and Schwann cells. Mechanistically, the increased invasion of SACC cells was associated with the expression of PCSK9. The findings of our study will contribute to a better understanding of the crosstalk between cancer cells and Schwann cells, especially by providing novel insights into how Schwann cells facilitate the PNI of SACC. Acknowledgments We thank Mier Li for the statistics support. We thank Shanghai OE Biotech (China) for the sequencing analysis. Funding This work is supported by the National Natural Science Foundation of China (Nos. 82141130 and 82372735) and the Joint Project of Medical Science and Technology of Henan Province (No. LHGJ20230308). Conflicts of interest None.

Myocardial ischemia/reperfusion (I/R) decreases cardiac function and efficiency. Accumulating evidence suggests that long noncoding RNAs (lncRNAs) have been linked to the cellular processes of myocardial I/R injury. The present investigation elucidated the function of lncRNA colon cancer -associated transcript 2 (CCAT2) in myocardial I/R injury and the related mechanisms. AC16 cardiomyocytes were exposed to hypoxia (16 hours)/reoxygenation (6 hours) (H/R) to mimic myocardial I/R models in vitro . CCAT2 and microRNA (miR)-539-3p expressions in AC16 cardiomyocytes were measured using real-time quantitative polymerase chain reaction. B -cell -specific Moloney murine leukemia virus insertion region 1 (BMI1) protein levels in AC16 cardiomyocytes were determined by western blotting. Cell viability, lactate dehydrogenase (LDH) leakage, reactive oxygen species (ROS) levels, mitochondrial membrane potential, and apoptosis were detected using Counting Kit -8, LDH Assay Kit, dihydroethidium assay, 5,5',6,6'tetrachloro1,1',3,3'-tetramethylbenzimidazolylcarbocyanine iodide staining, flow cytometry, and western blotting, respectively. The interactions between the molecules were confirmed using the dual-luciferase gene reporter. The wingless/integrated/beta-catenin (Wnt/(3-catenin) pathway under the H/R condition was detected by western blotting. CCAT2 and BMI1 mRNA expressions were reduced in H/R-exposed AC16 cardiomyocytes. CCAT2 overexpression exerted protective effects against H/R-induced cardiomyocyte injury, as demonstrated by increased cell viability and mitochondrial membrane potential and decreased LDH leakage, ROS levels, and apoptosis. In addition, CCAT2 positively regulated BMI1 expression by binding to miR-539-3p. CCAT2 knockdown or miR539-3p overexpression restrained the protective effects of BMI1 against H/R-induced cardiomyocyte injury. In addition, miR-539-3p overexpression reversed the protective effects of CCAT2. Furthermore, CCAT2 activated the Wnt/(3-catenin pathway under the H/R condition via the miR-539-3p/BMI1 axis. Overall, this investigation showed the protective effects of the CCAT2/miR-539-3p/BMI1/Wnt/(3-catenin regulatory axis against cardiomyocyte injury induced by H/R.

Figure S1: Effects of the PRC2 complex on the IFNï§-mediated Th1-type chemokine production in colon cancer cells

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