CXCL8 Secretion Research Articles

Inhibiting HnRNP L-mediated alternative splicing of EIF4G1 counteracts immune checkpoint blockade resistance in Castration-resistant prostate Cancer.

Immunotherapy with checkpoint inhibitors produced significant clinical responses in a subset of cancer patients who were resistant to prior therapies. However, Castration-resistant prostate cancer (CRPC) is seriously lack of T cell infiltration, which greatly limits the clinical application of immunotherapy, but the mechanism is unclear. In the present study, in silico analyses and experimental data show that HnRNP L was significantly negatively correlated with CD4+ and CD8+ T cells infiltration in patients; besides, we found deficiency of HnRNP L recruites CD4+ and CD8+ T cells infiltration and impairs tumorigenesis. Mechanically, HnRNP L enhanced the translation of c-Myc and then promoted CXCL8 secretion via alternative splicing of EIF4G1. In vivo, inhibition of EIF4G1 by the inhibitor, SBI-0640756, attenuated HnRNP l-induced tumor progression and immunosuppressive activity. And most of all, therapeutic synergy between HnRNP L knockdown and Anti-PD-1 could significantly suppress xenograft prostate cancer growth. In summary, this study revealled the molecular mechanism of HnRNP L regulating the immune infiltration, which provides a new theoretical basis for overcoming the limitation of immunotherapy for CRPC.

Smad4 deficiency in hepatocytes attenuates NAFLD progression via inhibition of lipogenesis and macrophage polarization

Nonalcoholic fatty liver disease (NAFLD), a major cause of chronic liver disorders, has become a serious public health issue. Although the Smad4 signaling pathway has been implicated in the progression of NAFLD, the specific role of Smad4 in hepatocytes in NAFLD pathogenesis remains unclear. Hepatocyte-specific knockout Smad4 mice (AlbSmad4−/−) were first constructed using the Cre-Loxp recombinant system to establish a high-fat diet induced NAFLD model. The role of Smad4 in the occurrence and development of NAFLD was determined by monitoring the body weight of mice, detecting triglycerides and free fatty acids in serum and liver tissue homogenates, staining the tissue sections to observe the accumulation of liver fat, and RT-qPCR detecting the expression of genes related to lipogenesis, fatty acid intake, and fatty acid β oxidation. The molecular mechanism of Smad4 in hepatocytes affecting NAFLD was therefore investigated through combining in vitro and in vivo experiments. Smad4 deficiency in hepatocytes mitigated NAFLD progression and decreased inflammatory cell infiltration. Moreover, Smad4 deficiency inhibited CXCL1 secretion by suppressing the activation of the ASK1/P38/JNK signaling pathway. Furthermore, targeting CXCL1 using CXCR2 inhibitors diminished hepatocyte lipogenesis and inhibited the polarization of M1-type macrophages. Collectively, these results suggested that Smad4 plays a vital role in exacerbating NAFLD and may be a promising candidate for anti-NAFLD therapy.

Long non-coding RNA Malat1 modulates CXCR4 expression to regulate the interaction between induced neural stem cells and microglia following closed head injury

BackgroundClosed head injury (CHI) provokes a prominent neuroinflammation that may lead to long-term health consequences. Microglia plays pivotal and complex roles in neuroinflammation-mediated neuronal insult and repair following CHI. We previously reported that induced neural stem cells (iNSCs) can block the effects of CXCL12/CXCR4 signaling on NF-κB activation in activated microglia by CXCR4 overexpression. Here we aim to uncover the mechanism of CXCR4 upregulation in iNSCs.MethodsWe performed bioinformatic analysis to detect the differentially expressed genes in iNSCs after co-cultured with LPS-activated microglia. Subsequently, we predicted the target genes and performed gain- and loss-of-functional studies, dualluciferase reporter, RNA immunoprecipitation, biotin-coupled miRNA pulldown, fluorescence in situ hybridization and cell transplantation assays to further elucidate the mechanism underlying the immunoregulatory effects of iNSCs. Student’s t-test and one-way analysis of variance (ANOVA) with Tukey’s post hoc test were used to determine statistical significance.ResultsOur results indicated that Malat1 could act as a sponge of miR-139-5p to modulate the expression of CXCR4 that exerted significant influence on the immunoregulatory effects of iNSCs on the secretion of CXCL12, TNF-α and IGF-1 by activated microglia. Furthermore, Malat1 inhibition blocked the immunoregulatory effects of iNSC grafts on microglial activation as well as neuroinflammation in the injured cortices of CHI mice. Interestingly, NF-κB activation in iNSCs augmented the immunoregulatory effects of iNSCs on microglial activation by activating the axis of Malat1/miR-139-5p/Cxcr4. Notably, we found that TNF-α secreted by activated microglia could bind to TNFR1 at the surface of iNSCs to trigger NF-κB activation in iNSCs.ConclusionsIn short, our findings reveal a novel role of Malat1 in the immunomodulatory effects of iNSCs on microglial activation, suggesting that transplanted iNSCs may self-perceive the changes of the activated state of microglia and thus make prudential regulation of the neuroinflammation following CHI.

CircPRKD3-loaded exosomes concomitantly elicit tumor growth inhibition and glioblastoma microenvironment remodeling via inhibiting STAT3 signaling.

Glioblastoma stem cells (GSCs) and their exosomes (exos) are involved in shaping the immune microenvironment, which is important for tumor invasion and recurrence. However, studies involving GSC-derived exosomal circular RNAs (GDE-circRNAs) in regulating tumor microenvironment (TME) remain unknown. Here, we comprehensively evaluated the significance of a novel immune-related GDE-circRNA in glioma microenvironment. GDE-circPRKD3 was screened out through high-throughput sequencing and verified by RT-PCR, sanger sequencing and RNase R assays. A series of in vitro and in vivo experiments were performed to investigate the function of GDE-circPRKD3. RNA-seq, RNA immunoprecipitation, multicolor flow cytometry and western blotting were used to explore the regulation of GDE-circPRKD3 on STAT3 signaling-mediated TME remodeling. We have characterized a circRNA PRKD3 in GSC exosomes, and lower circPRKD3 expression predicts a worse prognosis for glioblastoma patients. Overexpression of GDE-circPRKD3 significantly impairs the biological competence of glioma and prolongs the survival of xenograft mice. GDE-circPRKD3 binds to HNRNPC in an m6A-dependent manner, accelerates mRNA decay of IL6ST and inhibits downstream target STAT3. Notably, GDE-circPRKD3 promotes CXCL10 secretion by reprogramming tumor-associated macrophages, which in turn recruits CD8+ tumor infiltrating lymphocytes against GBM. Moreover, brain-targeted lipid nanoparticle delivery of circPRKD3 combined with immune checkpoint blockade therapy achieves significant combinatorial benefits. This study provides a novel mechanism by which GDE-circPRKD3 relies on STAT3 signaling to remodel immunosuppressive TME and offers a potential RNA immunotherapy strategy for GBM treatment.

Complement C3 of tumor-derived extracellular vesicles promotes metastasis of RCC via recruitment of immunosuppressive myeloid cells

Heterogeneous roles of complement C3 have been implicated in tumor metastasis and are highly context dependent. However, the underlying mechanisms linking C3 to tumor metastasis remain elusive in renal cell carcinoma (RCC). Here, we demonstrate that C3 of RCC cell-derived extracellular vesicles (EVs) contributes to metastasis via polarizing tumor-associated macrophages (TAMs) into the immunosuppressive phenotype and recruiting polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs). Mechanistically, EV C3 induces the secretion of CCL2 and CXCL1 by lung macrophages and subsequently enhances TAM polarization and PMN-MDSC recruitment. Notably, targeting the CCL2/CCR2 or CXCL1/CXCR2 axis with the inhibitors RS504393 or Navarixin, respectively, effectively suppresses lung metastasis induced by RCC-derived C3 in a mouse model. Clinically, RCC patients with high expression of C3 demonstrate poor prognosis. Collectively, our findings reveal that tumor-derived EV C3 induces an immunosuppressive tumor microenvironment via TAMs, and thus promoting RCC metastasis.

P0052 Enhancing gut epithelial integrity and reducing epithelial inflammation through supplementation of fecal metabolites, guided by novel ex vivo functional prioritization and testing

Abstract Background Inflammatory bowel disease (IBD) pathogenesis involves the gut epithelia, showing intensified inflammatory signals and a "leaky gut". However, there are no interventions targeting the epithelia. We aimed to prioritize and test whether fecal content from IBD patients, supplemented with metabolites directed by our functional assays, can reduce epithelial inflammation and improve integrity. Methods We established a model system utilizing whole human fecal samples from Crohn's disease (CD), Ulcerative colitis (UC), and healthy control subjects that were co-cultured with Caco-2 epithelial cells. Transepithelial electrical resistance (TEER) measured epithelial barrier integrity, and ELISA measured CXCL1 secretion. 16Sseq characterized fecal bacteria and metabolites were characterized using a semi-targeted predefined library (n=550). Results Fecal samples were processed by separating the supernatant, heat-killing the bacteria, and rejoining them. (Fig. 1A). We generated three pools per group (CD p#1-3, UC p#1-3, Controls p#1-3). All p#1 pools included samples from ten subjects, and p#2-3 each included samples from additional five independent subjects. Microbial characterization indicated that the fecal content pools captured the overall variations with separation between IBD and control samples (Fig. 1B). Co-culturing with p#1 revealed a significant increase in CXCL1 proinflammatory chemokine secretion to the media (Fig. 1C), and reduction in TEER (Fig. 1D) which was lower with pools obtained from CD and UC patients compared to the healthy pools. Independent validation with fecal pools p#2 and p#3 showed largely similar results (Fig. 1E-F). Characterizing metabolite levels in the nine pools and correlating them with disease state (IBD vs. controls) and the measured TEER outcomes highlighted several metabolites higher in controls and with improved TEER outcomes (Fig. 2A-B). Supplementing these prioritized metabolites with fecal contents pools, UC p#3 and CD p#2, which were linked with poorer epithelial outcomes, improved epithelial integrity, showing higher TEER when compared with original CD and UC pools (Fig. 2C). Similarly, supplementing UC p#3 and CD p#2 with those same metabolites mix significantly reduced CXCL1 secretion from epithelia in the model (Fig. 2D). Conclusion Fecal content from CD and UC patients was associated with more pathologic epithelial signals, which was improved in part with the addition of specific prioritized metabolites. This pipeline and model system highlights the potential future use of metabolite supplementations to improve epithelial function and outcome in IBD.

RNAi-based ALOX15B silencing augments keratinocyte inflammation in vitro via EGFR/STAT1/JAK1 signalling

Arachidonate 15-lipoxygenase type B (ALOX15B) peroxidises polyunsaturated fatty acids to their corresponding fatty acid hydroperoxides, which are subsequently reduced into hydroxy-fatty acids. A dysregulated abundance of these biological lipid mediators has been reported in the skin and blood of psoriatic compared to healthy individuals. RNAscope and immunohistochemistry revealed increased ALOX15B expression in lesional psoriasis samples. Using a cytokine cocktail containing IL-17A, interferon-gamma and tumour necrosis factor-alpha to produce a psoriasis-like phenotype, a role for ALOX15B in human epidermal keratinocyte inflammation was investigated. siRNA-mediated silencing of ALOX15B increased CCL2 expression and secretion. In addition to CCL2, secretion of CCL5 and CXCL10 were elevated in skin equivalents treated with lipoxygenase inhibitor ML351. Inhibition of the JAK1/STAT1 pathway reversed the enhanced CCL2 expression found with ALOX15B silencing. Previous studies have linked epidermal growth factor receptor (EGFR) inhibition with the upregulation of cytokines including CCL2, CCL5 and CXCL10. ALOX15B silencing reduced EGFR expression and inhibition of EGFR signalling potentiated the effect of ALOX15B silencing on increased CCL2, CCL5 and CXCL10 expression. Confirming previous findings, gene expression of cholesterol biosynthesis genes was reduced via reduced ERK phosphorylation. Reduced ERK phosphorylation was dependant on EGFR and NRF2 activation. Furthermore, plasma membrane lipids were investigated via confocal microscopy, revealing reduced cholesterol and lipid rafts. This study suggests a role for ALOX15B in keratinocyte inflammation through modulation of lipid peroxidation and the EGFR/JAK1/STAT1 signalling axis.

P0207 Ex-vivo colonoids derived from ulcerative colitis patients retain epithelial inflammatory “memory” even after several passages, which may contribute to UC chronicity

Abstract Background Intestinal epithelia is key in ulcerative colitis (UC) pathogenesis. However, epithelial models that capture the human variations for pre-clinical screening of epithelial-directed interventions are limited. We aimed to determine whether ex-vivo colonoids derived from different UC patients, maintain inflammatory signals associated with UC. Methods Rectal biopsies-derived epithelial colonoids were maintained through 4-9 passages (>1 month in culture) using an L-WRN-conditioned medium, differentiated for 48h, and were either left untreated or treated with inflammatory cocktails for an additional 24h. ELISA quantified CXCL1 secretion, mRNAseq and qPCR assessed mRNA levels Results 7 controls and 9 UC colonoids were generated from different subjects (median age was 16 years, 56% females). After 4-9 passages, untreated colonoids from UC patients secreted significantly higher levels of the CXCL1 chemokine compared to colonoids obtained from non-IBD controls (Fig. 1A). This phenomenon was also observed when colonoids were treated with INFγ and TNFα to mimic gut inflammation (Fig. 1B). mRNAseq further confirmed differences between UC and non-IBD colonoids. Unsupervised PCA analyses indicated the separation of 4/8 UC organoids from control organoids on PC1 (explaining 39% variance) and toward organoids treated ex-vivo with INFγ and TNFα (Fig. 1C). Differential expression (fold change >1.5 and FDR <0.05) identified 800 genes significantly upregulated, and 263 genes downregulated in untreated UC compared to control colonoids following passaging (Fig. 2A). Upregulated genes included DUOX2, DUOXA2, VDAC2, TJP1 (ZO1), TGM2, CXCL1, and IL8, as previously observed in mucosal biopsies from UC (PMID: 36655602) (Fig. 2B). Functional annotation of the upregulated genes highlighted pathways related to responses to wounding, bacterial signals, and cell-cell junctions (FDR<1E-10) (Fig. 2C). Conversely, downregulated genes included lipoprotein lipase (LPL), a key enzyme in lipid metabolism, the sulfate/anion transporter gene SLC26A1, and the mitochondrial coenzyme A transporter SLC25A42 (Fig. 2A-B). Functional enrichment analysis of the downregulated genes revealed associations with rRNA binding (FDR=1E-12) and mitochondrial gene expression (FDR=1E-6), all of which are relevant to UC pathogenesis (Fig. 2C). The expression of these genes and pathways were further induced with inflammatory triggering (Fig. 2D). Conclusion UC epithelial cells maintain a heightened inflammatory response ex-vivo after several passaging. This suggests an "inflammatory" epithelial transcriptional memory that may contribute to UC chronicity, justifying the use of this model for testing interventions targeted at the epithelium.

Transcriptomic signatures of cold acclimated adipocytes reveal CXCL12 as a Brown autocrine and paracrine chemokine.

Besides its thermogenic capacity, brown adipose tissue (BAT) performs important secretory functions that regulate metabolism. However, the BAT microenvironment and factors involved in BAT homeostasis and adaptation to cold remain poorly characterized. We therefore aimed to study brown adipocyte-derived secreted factors that may be involved in adipocyte function and/or may orchestrate intercellular communications. For this, mRNA levels in mature adipocytes from mouse adipose depots were assessed using RNA sequencing upon chronic cold acclimation, and bioinformatic analysis was used to identify secreted factors. Among 858 cold-sensitive transcripts in BAT adipocytes were 210 secreted factor-encoding genes, and Cxcl12 was the top brown adipocyte-enriched cytokine. Cxcl12 mRNA expression analysis by RT-qPCR and fluorescence in situ hybridization specified Cxcl12 distribution in various cell types, and indicated its enrichment in cold-acclimated brown adipocytes. We found that CXCL12 secretion from BAT was increased after chronic cold, yet its level in plasma remained unchanged, suggesting a local/paracrine function. Cxcl12 knockdown in mature brown adipocytes impaired thermogenesis, as assessed by norepinephrine (NE)-induced glycerol release and mitochondrial respiration. However, knockdown of Cxcl12 did not impact β-adrenergic signaling, suggesting that CXCL12 regulates adipocyte function downstream of the β-adrenergic pathway. Moreover, we provide evidence for CXCL12 to exert intercellular cross-talk via its capacity to promote macrophage chemotaxis and neurite outgrowth. Collectively, our results indicate that CXCL12 is a brown adipocyte-enriched, cold-induced secreted factor involved in adipocyte function and the BAT microenvironment communication network.

Platelet FcRγ inhibits tumor metastasis by preventing the colonization of circulating tumor cells.

Fc receptor γ subunit (FcRγ) activation plays a crucial role in cancer carcinogenesis. Here, we aimed to uncover the impact of FcRγ on circulating tumor cells (CTC) colonization and the underlying mechanism. FcRγ deficient (FcRγ-/-) mice were used to investigate the functional effects of FcRγ in cancer metastasis, and the results demonstrated that FcRγ deficiency significantly promotes metastasis. The tumor metastasis effect, antiplatelet, platelet or neutrophil infusion experiments were conducted with FcRγ deficient (FcRγ-/-) mice and wild type mice (WT), bearing B16F10 or LCC tumor cells. Blood routine test, flow cytometry, immunofluorescent staining and in vivo image were applied for analysis. Platelet adhesion and neutrophil chemotaxis were analyzed by flow cytometry and ELISA in vitro. Platelet adoptive model was used for mimicing early colonization stage. Our results indicated FcRγ deficiency significantly promoted tumor metastasis accompanied with increased number of platelet and neutrophil in the lung. Further investigation showed that FcRγ-/- platelet infusion, rather than FcRγ-/- neutrophils, promoted CTC colonization. While platelet inhibitor Aspirin abrogated the platelet-mediated infiltration of neutrophil in the lung. Mechanistically, platelet FcRγ deficiency facilitated the adhesion of platelets and cancer cells and increased secretion of CXCL5 and CXCL7 which triggered the platelet-induced neutrophil recruitment. In sum, our study indicates that FcRγ is a restrainer in controlling cancer metastasis through regulating the adhesion of platelets and cancer cells and recruiting more neutrophils, which provides a potential target for anti-metastatic therapies. The level of FcRγ expression in platelets could act as a novel biomarker for cancer metastasis.

Physiologically Relevant Coculture Model for Oral Microbial-Host Interactions.

Understanding microbial-host interactions in the oral cavity is essential for elucidating oral disease pathogenesis and its systemic implications. In vitro bacteria-host cell coculture models have enabled fundamental studies to characterize bacterial infection and host responses in a reductionist yet reproducible manner. However, existing in vitro coculture models fail to replicate the physiological oxygen gradients critical for studying these interactions. Here, we present an asymmetric gas coculture system that simulates the oral microenvironment by maintaining distinct normoxic and anaerobic conditions for gingival epithelial cells and anaerobic bacteria, respectively. Using Fusobacterium nucleatum, a key oral pathobiont, we demonstrate that the system preserves bacterial viability and supports the integrity of telomerase-immortalized gingival keratinocytes. Compared to conventional models, this system enhanced bacterial invasion, elevated intracellular bacterial loads, and elicited more robust host pro-inflammatory responses, including increased secretion of CXCL10, IL-6, and IL-8. Additionally, the model enabled precise evaluation of antibiotic efficacy against intracellular pathogens. These results underscore the utility of this coculture platform for studying oral microbial pathogenesis and screening therapeutics, offering a physiologically relevant approach to advance oral and systemic health research.

Intra-tumoral sphingobacterium multivorum promotes triple-negative breast cancer progression by suppressing tumor immunosurveillance

BackgroundIntratumor-resident bacteria represent an integral component of the tumor microenvironment (TME). Microbial dysbiosis, which refers to an imbalance in the bacterial composition and bacterial metabolic activities, plays an important role in regulating breast cancer development and progression. However, the impact of specific intratumor-resident bacteria on tumor progression and their underlying mechanisms remain elusive.Methods16S rDNA gene sequencing was used to analyze the cancerous and paracancerous tissues from breast cancer patients. The mouse models of bearing 4T1 cell tumors were employed to assess the influence of bacterial colonization on tumor growth. Tissue infiltration of regulatory T (Treg) cells and CD8+ T cells was evaluated through immunohistochemistry and flow cytometric analysis. Comparative metabolite profiling in mice tumors was conducted using targeted metabolomics. Differential genes of tumor cells stimulated by bacteria were analyzed by transcriptomics and validated by qPCR assay.ResultsWe found that Sphingobacterium displayed high abundance in cancerous tissues. Intra-tumoral colonization of Sphingobacterium multivorum (S. multivorum) promoted tumor progression in 4T1 tumor-bearing mice. Moreover, S. multivorum diminished the therapeutic efficacy of αPD-1 mAb, which was associated with the increase of regulatory T cell (Treg) infiltration, and decrese of the CD8+ T cell infiltration. Targeted metabolomics revealed a conspicuous reduction of propionylcarnitine in tumors colonized by S. multivorum Furthermore, the combination of metabolite propionylcarnitine and S. multivorum shown to suppress tumor growth compared that in S. multivorum alone in vivo. Mechanistically, S. multivorum promoted the secretion of chemokines CCL20 and CXCL8 from tumor cells. CCL20 secreted into the TME facilitated the recruitment of Treg cells and reduced CD8+ T cell infiltration, thus promoting tumor immune escape.ConclusionsThis study reveals S. multivorum suppresses immune surveillance within the TME, thereby promoting breast cancer progression.

CXCL12 ameliorates neutrophilia and disease severity in SARS-CoV-2 infection.

Neutrophils, particularly low-density neutrophils (LDNs), are believed to contribute to acute COVID-19 severity. Here, we showed that neutrophilia can be detected acutely and even months after SARS-CoV-2 infection in patients and mice, while neutrophil depletion reduced disease severity in mice. A key factor in neutrophilia and severe disease in infected mice was traced to the chemokine CXCL12 secreted by bone marrow cells and unexpectedly, endothelial cells. CXCL12 levels were negatively correlated with LDN numbers in longitudinal analyses of patient blood samples. CXCL12 blockade in SARS-CoV-2-infected mice increased blood/lung neutrophil numbers thereby accelerating disease progression without changing lung virus titers. The exaggerated mortality caused by CXCL12 blockade can be reversed by neutrophil depletion. In addition, blocking interactions between SARS-CoV-2 and Angiotensin-Converting Enzyme 2 (ACE2) reduced CXCL12 levels, suggesting a signal transduction from virus-mediated ACE2 ligation to increased CXCL12 secretion. Collectively, these results demonstrate a previously unappreciated role of CXCL12 in diminishing neutrophilia, including low density neutrophilia, and its deleterious effects in SARS-CoV-2 infections. The results also support the involvement of SARS-CoV-2-endothelial cell interactions in viral pathogenesis.

Human leukocyte antigen DR alpha inhibits renal cell carcinoma progression by promoting the polarization of M2 macrophages to M1 via the NF-κB pathway

Human leukocyte antigen DR alpha (HLA-DRA) is recognized for its inhibitory effect on the progression of clear cell renal cell carcinoma (ccRCC); high HLA-DRA expression levels are positively correlated with improved prognosis in patients with ccRCC. In this study, we evaluated HLA-DRA expression in ccRCCs, its effects on tumor-associated macrophage recruitment, and the influence of polarization. Clinical cohort analyses revealed that elevated HLA-DRA expression in ccRCC cells was correlated with enhanced tumor infiltration by M1-type macrophages. In addition, ccRCC prognosis was predicted by combining HLA-DRA expression level analysis and the M1/M2 macrophage ratio. In vitro studies demonstrated that ccRCC cells with increased HLA-DRA expression promoted THP-1 cell migration and induced macrophage polarization toward the M1 phenotype. The effect was further substantiated in a mouse xenograft model in which an increase in M1 macrophages was observed. In addition, co-culturing macrophages with the supernatant from cells overexpressing HLA-DRA induced the expression of proteins associated with both M1 and M2 macrophage polarization. HLA-DRA was intricately linked to the expression and secretion of chemokines, including CCL2, CCL5, MIP-1ɑ, and CXCL-10. Moreover, the NF-κB pathway activation promoted polarization to M1 macrophages. This study shows that HLA-DRA and the M1/M2 ratio are indicators of favorable prognosis in patients with ccRCC. HLA-DRA promotes M1-like polarization by regulating NF-κB, which can be used as a therapeutic target to enhance anti-tumor immunity.

MAF1 inhibits hepatocarcinogenesis by fostering an immunostimulatory tumor microenvironment

BackgroundThe biological significance of MAF1, a tumor suppressor, in carcinogenesis and immune response of hepatocellular carcinoma (HCC) remains unreported. Understanding the underlying mechanisms by which MAF1 enhances anti-tumor immunity in...

Self-delivering RNAi immunotherapeutic PH-762 silences PD-1 to generate local and abscopal antitumor efficacy.

Immunotherapeutic inhibition of PD-1 by systemically administered monoclonal antibodies is widely used in cancer treatment, but it may cause severe immune-related adverse events (irSAEs). Neoadjuvant PD-1 inhibition before surgery has shown promise in reducing recurrence by stimulating durable antitumor immunity. Local intratumoral (IT) immunotherapy is a potential strategy to minimize irSAEs, but antibodies have limited tumor penetration, making them less suitable for this approach. Therapeutic self-delivering RNAi (INTASYL) is an emerging modality well-suited for neoadjuvant immunotherapy. This study presents preclinical proof-of-concept for PH-762, an INTASYL designed to silence PD-1, currently in clinical development for advanced cutaneous malignancies (ClinicalTrials.gov#NCT06014086). PH-762 pharmacology was characterized in vitro, and in vivo antitumor efficacy was evaluated using a murine analogue (mPH-762) in syngeneic tumor models with varying PD-1 responsiveness. Bilateral Hepa1-6 models assessed abscopal effects of local treatment. Ex vivo analyses explored mechanisms of direct and abscopal efficacy. PH-762 was rapidly internalized by human T cells, silencing PD-1 mRNA and decreasing PD-1 surface protein, enhancing TCR-stimulated IFN-γ and CXCL10 secretion. In vivo, IT mPH-762 provided robust antitumor efficacy, local and lymphatic biodistribution, and was well tolerated. Ex vivo analyses revealed that IT mPH-762 depleted PD-1 protein, promoted leukocyte and T cell infiltration, and correlated with tumor control. IT mPH-762 also demonstrated efficacy against untreated distal tumors (abscopal effect) by priming systemic antitumor immunity. These data support PH-762 as a promising candidate for neoadjuvant immunotherapy in clinical studies.

Liraglutide promotes osteogenic differentiation of mesenchymal stem cells by inhibiting M1 macrophage polarization and CXCL9 release in vitro.

As a GLP-1 receptor agonist widely used in treating type 2 diabetes, liraglutide shows potential applications in bone tissue engineering. This study investigated liraglutide's direct effects on rat bone marrow mesenchymal stem cells (BMSCs) osteogenic differentiation and its regulatory mechanism through macrophage polarization. Results showed that liraglutide significantly enhanced BMSC migration and osteogenic differentiation. Additionally, liraglutide markedly inhibited M1 macrophage polarization induced by LPS and IFN-γ, reducing inflammatory factors CXCL9 and TNF-α secretion, possibly by partially reversing M1 macrophage regulatory signals (AMPK and NF-κB pathways). Compared to M1 macrophage-conditioned medium (M1-CM), conditioned medium from liraglutide-treated macrophages showed stronger promotion of BMSC osteogenic differentiation, though this effect was reversed by CXCL9 addition. The study demonstrates that liraglutide enhances BMSC osteogenic capacity both directly and by inhibiting M1 macrophage polarization and CXCL9 secretion, offering a new therapeutic option for severe bone defects with inflammatory responses.

Opposing effects of mycotoxins alternariol and deoxynivalenol on the immunomodulatory effects of oxaliplatin and triapine.

Mycotoxin occurrence in food worldwide is estimated to increase due to climate change. Moreover, studies on how these food contaminants interfere with medications and especially anticancer therapies are rare. With the rise of anticancer immunotherapies, particularly mycotoxins with immunomodulatory activity, such as alternariol (AOH) or deoxynivalenol (DON), are of great concern. Both mycotoxins interfere with the pro-inflammatory nuclear factor kappa B (NF-κB) pathway in myeloid cells. This pathway not only plays an important role in the anticancer immune response but also inflammatory side effects induced by chemotherapeutic drugs. Consequently, the aim of this study was to investigate possible beneficial or detrimental immunomodulatory interactions between these mycotoxins and anticancer drugs. To assess the combined influence of mycotoxins and anticancer therapies on immune cell stimulation, THP-1 NF-κB reporter cells were utilized as monocytes as well as differentiated and polarized macrophages. Parameters for activation (NF-κB activity and protein expression), differentiation (CD14 and CD71 surface marker expression) and polarization (interleukin 10 (IL10), interleukin 8 (CXCL8), tumor necrosis factor α (TNF), prostaglandin-endoperoxide synthase 2 expression and CXCL8 secretion) were assessed upon combinatory treatment. Both mycotoxins affected the immunostimulatory effects of the pre-selected anticancer drugs oxaliplatin and triapine, although in opposing directions. While AOH generally suppressed a drug-induced activation and increased anti-inflammatory IL10 levels, DON potentiated activation and pro-inflammatory markers, such as CXCL8 and TNF in immune cells. In conclusion, AOH and DON have the potential to alter the immunological effects of anticancer therapies, which should be considered during therapy as well as in their future risk assessment.

Triptolide exhibits dual anti-tumor effects through inhibiting autophagy and extracellular matrix activation in pancreatic cancer.

The tumor microenvironment in pancreatic cancer, characterized by abundant desmoplastic stroma, has been implicated in the failure of chemotherapy. Therefore, developing therapeutic strategies targeting tumor and stromal cells is essential. Triptolide, a natural compound derived from the plant Tripterygium wilfordii, has shown antitumor activity in various cancers, including pancreatic cancer. However, its effects on pancreatic cancer cells and the microenvironment remain unclear. This study aimed to explore the effect of triptolide on tumor cells and the tumor microenvironment in pancreatic cancer. Cell Counting Kit-8, colony formation, apoptosis, and cell cycle assays were performed to determine the effect of triptolide on tumor cells. Additionally, co-culture assays were performed to explore the effects of the compound on cancer-associated fibroblasts (CAFs) in vitro. Orthotopic xenograft and subcutaneous tumor models were used to explore the antitumor and antistromal activation effects of triptolide in vivo. RNA sequencing was performed to identify the pathways involved in these processes in pancreatic cancer cells. Triptolide inhibited the proliferation of pancreatic cancer cells and attenuated stromal activation in vitro and in vivo. Furthermore, it suppressed autophagy and induced apoptosis in pancreatic cancer cells by inhibiting the secretion of CXCL1. Extracellular matrix formation in CAFs was disrupted by suppressing the paracrine secretion of TGF-β from tumor cells. These findings indicate that triptolide plays a dual antitumor role against tumor cells and CAFs, thus providing new insights into treating pancreatic cancer in the future.

Adiponectin deficiency prevents chronic colitis-associated colonic fibrosis via inhibiting CXCL13 production.

Colonic fibrosis is a long-term complication of inflammatory bowel disease (IBD), often leading to functional impairment, intestinal obstruction, and surgery. Adiponectin (APN) is an adipokine derived from adipocytes that plays a pleiotropic role in fibrosis regulation, depending on tissue and cell type specific or disease context, but its role in colonic fibrosis remains unclear. To explore the role and involved mechanism of APN in chronic colitis-associated colonic fibrosis. Studies were performed in GEO database, colonic tissues of UC patients, dextran sulfate sodium (DSS)-induced colonic fibrosis in male wild-type (WT) and APN-/- mice, mouse L929 and human CCD-18Co fibroblasts treated with recombinant CXCL13 protein, and colonic fibrosis in WT mice infected with shRNA of CXCL13. APN was highly expressed in the colonic tissues of UC patients and positively correlated with the colonoscopy score and colonic fibrosis markers COL1A1 and COL3A1. APN deficiency significantly improved chronic colitis-induced colonic fibrosis in mice with down-regulating collagenase accumulation and expressions of TGF-β, α-SMA, COL1A1, COL3A1, and MMP-9 in colonic tissues. Transcriptomics showed that APN deficiency mainly affected cytokine-cytokine receptor interactions, especially CXCL13 signaling. Follow-up studies showed that APN deficiency significantly decreased the number of colonic F4/80+CD206+CXCL13+ macrophages by weakening Akt phosphorylation. Additional experiments confirmed that CXCL13 notably increased the expressions of α-SMA and COL1A1 in mouse in mouse and human fibroblasts by activating p-Akt, p-p38, p-ERK, and p-JNK. Moreover, inhibiting CXCL13 with shRNA significantly ameliorated colonic fibrosis in mice with DSS-induced chronic colitis. Immunohistochemistry analysis revealed high expression of CXCL13 in the colon tissues of patients with UC, showing a positive correlation with APN, COL1A1, and COL3A1. APN contributes to the progression of colonic fibrosis and can exacerbate this condition by regulating the secretion of CXCL13 in the colon, offering potential new perspectives on the pathophysiology of colonic fibrosis.