Biology Direct最新文献

Background & aims: Gallbladder cancer (GBC) is characterised by a desmoplastic microenvironment rich in collagen fibres and extracellular matrix, contributing to increased matrix stiffness. SEMAs are overexpressed in various cancers but their expression and role in the crosstalk between activated gallbladder fibroblasts (GFs) and GBC cells within a stiff microenvironment remain unclear. Herein, we aimed to elucidate the expression patterns and tumour-promoting effects of SEMAs in activated GFs under the matrix stiffness microenvironment.

Methods: Masson staining assessed collagen deposition in GBC stroma. GFs were isolated from gallbladder tissues and cultured on silicon or polyacrylamide hydrogels. SEMA expression in GFs under different stiffness conditions was determined via RNA-seq and RT-qPCR. Transwell assays, tumorsphere-formation and Western blot assays were used to investigate the effects of GFs-derived SEMA7A on GBC epithelial-mesenchymal transition (EMT) and cancer stem-like traits. Subcutaneous tumour models were established by co-injecting GFs and GBC cells to assess SEMA7A's role in vivo. Co-immunoprecipitation, WB, Elisa and mutation assays were employed to elucidate the mechanism of SEMA7A involvement in GFs-GBC cell crosstalk.

Results: This study revealed a high-stiffness microenvironment in GBC, inducing GFs activation and SEMA7A paracrine through YAP/TAZ signalling. GFs-secreted SEMA7A under matrix stiffness microenvironment promoted GBC EMT and cancer stem-like traits. Mechanistically, GFs-secreted SEMA7A bound to its receptor integrin β1 instead of plexinC1 on GBC cells to induce the phosphorylation of p300 at S1834 and maintain the malignant phenotypes of GBC cells. Moreover, the SEMA7A/integrin β1 axis-induced p300 phosphorylation at S1834 was mediated by Akt activation (p-Akt at S473) in GBC cells.

Conclusions: Our findings demonstrate a complex stiffness/SEMA7A/integrin β1/AKT/p300 cascade mediating reciprocal interactions between GFs and GBC cells, offering a potential therapeutic target for patients with GBC.

Background: Metabolic alterations are closely associated with the exhaustion and immune deficiency of CD8⁺ tumor-infiltrating lymphocytes (TILs), while little is known about diffuse large B-cell lymphoma (DLBCL). This study aimed to elucidate the significance of the metabolic alterations in exhausted CD8⁺TILs and its underlying regulatory mechanism in DLBCL.

Methods: The metabolic alterations in exhausted CD8⁺TILs in DLBCL were evaluated through single-cell RNA sequencing (scRNA-seq). The crucial metabolic pathway and its significance in the biological function of exhausted CD8⁺TILs were investigated by scRNA-seq and RNA sequencing. The marker gene in crucial metabolic pathway, and its correlations between exhaustion status, the tumor microenvironment (TME) composition, clinicopathological characteristics, prognosis, and immune checkpoint blockade (ICB) therapy efficacy were evaluated by scRNA-seq, RNA sequencing, immunohistochemistry, and RT-qPCR. Furthermore, the underlying regulatory mechanism involved in the metabolic alteration related to CD8⁺TILs exhaustion was explored through scRNA-seq, RNA sequencing, and somatic mutation analysis.

Results: Our study illustrated the metabolic heterogeneity in CD8⁺TILs, and demonstrated that oxidative phosphorylation (OXPHOS) was the crucial pathway in CD8⁺TILs exhaustion. The high OXPHOS activity indicated the immune deficiency in exhausted CD8⁺TILs, and UQCRFS1 was identified as a marker gene. High UQCRFS1 indicated the immunosuppressive TME, severe clinicopathological characteristics, including activated B-cell-like subtype, high IPI and PS score, advanced stage, dismal prognosis, and resistance to ICB therapy. Furthermore, MYC-related signaling and P2RY8 mutation in DLBCL may regulate the UQCRFS1 expression in exhausted CD8⁺TILs.

Conclusions: Our study highlights the importance of OXPHOS activity in CD8⁺TILs exhaustion and suggests its possible regulatory mechanism, which is feasible in clinical evaluation and beneficial for novel immunotherapeutic approaches in DLBCL.

Background: Globally, HCC is still one of the most common cancers. N6-methyladenosine (m⁶A) modifications and long-stranded noncoding RNAs (lncRNAs) play key roles in regulating HCC progression. The role of the lncRNA RNF144A-AS1, a newly identified lncRNA, in HCC is unclear.

Methods: In HCC, RNF144A-AS1 expression level and its effect on prognosis were investigated by bioinformatics. CCK-8, EdU, scratch assay, and Transwell assays were used to detect the impact of RNF144A-AS1 on hepatocellular carcinoma malignancy. Assays using MeRIP-qPCR, RIP, and Actinomycin D were used to study the effects of m⁶A methylation on hepatocellular carcinoma malignant phenotypes. Revealing the potential mechanism of action of RNF144A-AS1 by luciferase reporter gene assay, PCR, and Western blot assays, and Nude mice subcutaneous load cell and lung metastasis models were used to verify the effect of RNF144A-AS1 on the malignant phenotype of tumors in vivo.

Results: The lncRNA RNF144A-AS1 was significantly upregulated in HCC, and it was significantly associated with poor prognosis. Functionally, HCC cells with RNF144A-AS1 knockdown were inhibited in terms of proliferation, migration, and invasion. Further studies in vivo confirmed that RNF144A-AS1 knockdown inhibited tumor cell growth and metastasis. Mechanistically, METTL3 increased the m⁶A modification and stability of RNF144A-AS1 in an IGF2BP1-associated manner. In addition, RNF144A-AS1 was inhibited by sponge-like miR-1301-3p to inhibit RNF38 degradation, thereby promoting the HCC malignant phenotype.

Conclusion: The RNF144A-AS1 gene is affected by METTL3/IGF2BP1 methylation and encourages liver cancer proliferation and metastasis by increasing expression of RNF38 through sponge-like miR-1301-3p. RNF144-AS1 promises to be a therapeutic target for HCC.

Background: The mucin-degrading bacterium Akkermansia muciniphila has attracted enormous interest for its beneficial effects on human health. However, growing evidence suggests that the Akkermansia genus is populated by several species that differ in phenotypic characteristics and association with human traits.

Results: We present the most comprehensive phylotaxonomic analysis of Akkermansia genomes in terms of sample size and host representation. By applying approaches based on average nucleotide identities and on the biological species concept, we show that the Akkermansia genus comprises at least 31 species, 13 of which can be detected in humans. The largest species diversity is contributed by non-human and non-mouse animals, and limited evidence of species-specificity is evident, with several Akkermansia species detected in phylogenetically distant animals. Analysis of accessory gene content among species also failed to reveal species-specific or diet-specific associations, but rather reflected genome size. Thus, A. muciniphila and A. ignis have, on average, small genomes and retain a part of genes that characterize either A. massiliensis or A. sp004167605/A. biwaensis. Finally, investigation of the population structure of A. muciniphila, the species that has been more intensely investigated due to its effects on human health, clearly distinguished two phylogroups corresponding to AmIa and AmIb. However, analysis of laboratory mouse-derived genomes revealed that additional populations, specific to these animals, exist. Such populations show limited evidence of admixture, suggesting bottleneck or competition effects.

Conclusions: Our data support the concept that the genetic diversity of Akkermansia should be taken into account in experimental settings. They also call for sequencing efforts to characterize the wider genetic diversity of Akkermansia bacteria.

Chloride intracellular channels (CLICs) are a family of six evolutionarily conserved proteins with diverse functions. Previously, we identified CLIC2, as the fifth-ranked master regulator associated with diffuse-type gastric cancer (dGC) showing increased expression in tumors. Here we used bulk, as well as single cell sequencing datasets of dGC, to demonstrate for the first time a direct association of CLIC2 with the microsatellite stable GC and, furthermore, the expression of CLIC2 in macrophages (MCs), and endothelial cells (ECs) populating gastric tissue. We generated CLIC2 knock-out THP-1 monocytic cells (THP-1CLIC2_KO) determining that while CLIC2 deletion had no observable effect on monocytes, THP-1CLIC2_KO macrophages exhibited significant morphological changes, including increased membrane protrusions, and upregulated expression of CD11b, CD11c, CD80, and CD86 markers. Furthermore, cytokine secretion profiling of THP-1CLIC2_KO differentiated cells revealed elevated secretion of CCL8, alongside reduced secretion of IL-1β, IL-6, and osteoprotegerin (OPG). Additionally, we observed increased phosphorylation of Shp1 phosphatase with the concomitant absence of Stat3 phosphorylation, which impaired downstream signaling, in line with the evidence that Clic2 interacts with both Shp1 and Stat3. Based on these findings, we suggest that CLIC2 plays a pivotal role in regulating monocyte-to-macrophage differentiation by modulating the Stat3 signaling pathway, thus enhancing gastric cancer progression by establishing a tumor-permissive microenvironment.

Background: Ferroptosis and disulfidptosis are newly discovered forms of regulated cell death that play critical roles in cancer progression, metabolism, and immune evasion. However, their interplay and combined influence on colorectal cancer (CRC) progression remain insufficiently understood.

Methods: We developed a ferroptosis-disulfidptosis-related gene (FDRG) score using machine-learning algorithms to analyze gene modifications associated with these pathways in CRC, utilizing data from the TCGA and GEO databases. The model was externally validated, and associations with clinical outcomes, immune infiltration, mutational landscapes, immunotherapy responses, and drug sensitivity were explored. Key genes were further investigated through bioinformatics and in vitro experiments.

Results: We constructed an 8-gene risk model with strong prognostic value, stratifying CRC patients into high- and low-risk groups with significant differences in clinical characteristics, immune cell infiltration, and therapeutic responses. Among these genes, CHMP6 was identified as a previously uncharacterized tumor suppressor in CRC. Beyond its inhibitory effect on tumor cell proliferation, migration, and invasion, CHMP6 was found to play a critical role in modulating anti-tumor immunity. Our findings established CHMP6 as a dual-function tumor suppressor that not only restrains tumor progression but also enhances immune-mediated tumor control.

Conclusions: The FDRG score is a robust tool for predicting CRC prognosis, tumor microenvironment dynamics, and response to immunotherapy. CHMP6 emerged as a promising tumor suppressor and potential therapeutic target, offering new insights into CRC treatment strategies.

Background: Despite significant advancements in therapeutic approaches for lung cancer, the prognosis of lung squamous cell carcinoma (LUSC) remains suboptimal, underscoring the critical need to identify novel molecular targets and develop targeted therapeutic strategies. Through bioinformatic analysis, SH3RF2 was identified as a gene significantly upregulated in LUSC patients, and its high expression was strongly associated with lower survival rates. However, no significant differences in expression or survival correlation were observed in lung adenocarcinoma. Notably, SH3RF2, an E3 ubiquitin ligase characterized by three SH3 domains, has not been systematically investigated in LUSC pathogenesis.

Results: Mechanistic investigations found that SH3RF2 promoted tumor cell proliferation, upregulated M2 markers (Arg-1, CD163, IL-10), increased CD206 + subpopulation of M0 THP-1 cells and enhanced migration and invasion of M0 THP-1 cells. SH3RF2 promoted the nuclear translocation of β-catenin. Furthermore, ICG-001, the inhibitors of β-catenin pathway, alleviated the above effects of SH3RF2. In vivo tumorigenesis experiments found that SH3RF2 promoted tumor growth and increased the proportion of M2 cells. Proteomic analysis revealed that SH3RF2 interacted with LZTS2 and regulated the ubiquitination of LZTS2 with RING domain. Overexpression of LZTS2 attenuated SH3RF2-induced nuclear translocation of β-catenin, suppressed cellular migration and invasion, and inhibited M2 polarization promoted by SH3RF2 overexpression. The combination of SH3RF2 knockdown and radiotherapy inhibited the growth of tumor compared with SH3RF2 knockdown or radiotherapy alone.

Conclusions: This study demonstrates the functionality of SH3RF2 in both potentiating tumor progression and inducing M2 macrophage polarization through coordinated regulation of LZTS2 degradation and β-catenin nuclear translocation. These findings establish a novel mechanistic framework and propose SH3RF2-associated signaling axes as promising therapeutic targets for LUSC.

Background: Iron overload and ferroptosis are associated with intestinal ischemia and reperfusion (II/R)-induced acute lung injury (ALI). However, the mechanisms underlying the regulation of iron homeostasis remain unclear. Nrf2 regulates cellular iron homeostasis; nonetheless, its impact on ALI pathology and its underlying mechanism of action requires further investigation. Ubiquitin ligase E3B (UBE3B) plays a critical role in the proteasome pathway, which is essential for protein turnover and ubiquitin-mediated signaling. A recent study found that UBE3B plays a role in oxidative stress; nevertheless, it remains unknown whether its role is related to Nrf2. Furthermore, the exact role of UBE3B in ALI and its underlying mechanism remain largely uncharacterized.

Methods and results: In the present study, immunohistochemical analysis of UBE3B expression in type II alveolar epithelial cells (AECII) was conducted, and its expression was found to be decreased in II/R-ALI. Western blot analysis indicated that UBE3B hypoactivation may aggravate oxidative stress, thereby promoting ALI. Moreover, UBE3B was involved in iron metabolism dysfunction and ferroptosis. UBE3B deficiency enhanced the process of nuclear receptor coactivator 4 (NCOA4)-mediated ferritinophagy and increased ferrous ion content, whereas overexpression of UBE3B reversed the harmful effects of Nrf2 knockdown on AECII, which may promote AECII ferroptosis.

Conclusions: This study highlights the role of the Nrf2/UBE3B/NCOA4 axis in AECII ferroptosis and II/R-ALI pathogenesis, suggesting that Nrf2 activation may be a promising strategy for ALI treatment.

Doxorubicin (DOX) is a potent chemotherapeutic widely used against various cancers, but its clinical application is limited by DOX-induced cardiotoxicity (DIC). This study explored the cardioprotective potential of extracellular vesicle-enriched secretome derived from adipose stem cells (EVS_ASC) in mitigating DOX-induced apoptosis in cardiomyocytes. Adipose-derived stem cells were cultured, and their conditioned medium and extraceullular vesicles were isolated and characterized according to the Minimal Information for Studies of Extracellular Vesicles 2023 guidelines. HL-1 cardiomyocytes were pretreated with EVS_ASC before exposure to 1 µM DOX. Cell viability was assessed via the cell counting kit-8 assay, while apoptosis markers and survival mediators were evaluated through Western blotting. RNA sequencing identified differentially expressed genes, including clusterin (Clu), which was further quantified using an enzyme-linked immunosorbent assay. The functional role of clusterin was validated through siRNA-mediated knockdown. EVS_ASC significantly improved cell viability in DOX-exposed cardiomyocytes and reduced the cleaved caspase-3 to procaspase-3 ratio. Clusterin expression was highest in EVS_ASC-treated cells, and its knockdown markedly increased caspase-3 cleavage, confirming its pivotal role in cardioprotection. Moreover, EVS_ASC enhanced the phosphorylation of AKT, Bcl2-associated agonist of cell death, and glycogen synthase kinase-3β, implicating PI3K/AKT pathway activation in clusterin upregulation and anti-apoptotic effects. These findings demonstrate that EVS_ASC mitigates DOX-induced apoptosis in cardiomyocytes through clusterin upregulation and PI3K/AKT pathway activation. Clusterin is identified as a potential biomarker for evaluating EVS_ASC efficacy. While EVS_ASC shows promise as a cardioprotective strategy against DIC, further studies are needed to optimize its therapeutic safety by addressing potential oncogenic risks.