Esophageal carcinoma is one of the most common malignancies with high cancer-related morbidity and mortality worldwide. MicroRNAs (miRNAs) are a class of small non-coding RNAs that regulate a wide variety of cellular processes, and also play an important role in the development and progression of cancers. In a previous microarray study, we demonstrated that miR-130b was upregulated in esophageal squamous cell carcinoma (ESCC) tissues. However, the biologic functions and the molecular mechanism of miR-130b in ESCC remain to be elucidated. qRT-PCR assays were used to quantify miR-130b expression levels in ESCC samples. Novel targets of miR-130b were identified via a bioinformatics search and confirmed using a dual-luciferase reporter system. Western blotting and qRT-PCR assays were used to quantify the expression of the target gene PTEN (phosphatase and tensin homolog) and the downstream effector, Akt. ESCC cells over- or underexpressing miR-130b were analyzed for in vitro biologic functions. High levels of miR-130b were identified in 20 ESCC samples following comparison with adjacent non-neoplastic tissues. We confirmed that miR-130b interacted with the 3′-untranslated region of PTEN, and that an increase in the expression level of miR-130b negatively affected the protein level of PTEN. However, the dysregulation of miR-130b had no obvious impact on PTEN mRNA. As Akt is a downstream effector of PTEN, we explored if miR-130b affected Akt expression, and found that miR-130b indirectly regulated the level of phosphorylated Akt, while total Akt protein remained unchanged. Overexpression of miR-130b increased the proliferation of ESCC cells and enhanced their ability to migrate and invade. In contrast, the proliferation, migration, and invasion of ESCC cells were weakened when miR-130b expression was suppressed, which was reversed by PTEN-targeted siRNA. The results indicate that miR-130b plays an oncogenic role in ESCC cells by repressing PTEN expression and Akt phosphorylation, which would be helpful in developing miRNA-based treatments for ESCC.

Aberrant expression of miR-224 is associated with tumor development and progression. This study investigated the role of miR-224 in esophageal squamous cell carcinoma (ESCC) ex vivo and in vitro. A total of 103 esophageal intraepithelial neoplasia, ESCC tissue specimens, and their matched distant normal tissues were collected to test miR-224 expression using qRT-PCR analysis. Western blot was used to quantify the level of PH domain leucine-rich repeat protein phosphatase 1 (PHLPP1) and PHLPP2 in ESCC tissues. Cell viability, apoptosis, invasion, and colony formation assays were used to assess the altered phenotypes of esophageal cancer cell lines after miR-224 expression or inhibition. A luciferase reporter assay was used to confirm miR-224 binding to PHLPP1 and PHLPP2 mRNA. miR-224 was significantly overexpressed in esophageal intraepithelial neoplasia and ESCC tissues, while the expression of PHLPP1 and PHLPP2 proteins, the target genes of miR-224, was downregulated in ESCC tissues. miR-224 expression was associated with advanced clinical TNM stage, pathologic grade, and the level of PHLPP1 and PHLPP2 proteins in ESCC tissues. Ectopic overexpression of miR-224 promoted proliferation, migration, and invasion, but suppressed apoptosis of ESCC cells. miR-224 was able to bind to the 3′ untranslated region (3′-UTR) of PHLPP1 and PHLPP2 mRNA to suppress their expression. The current study demonstrated that miR-224 acts as an oncogenic miRNA in ESCC, possibly by targeting PHLPP1 and PHLPP2.

Accurate prediction of lymph node metastasis (LNM) in T1 esophageal squamous cell cancer is critical for guiding treatment decisions after endoscopic submucosal dissection (ESD). We developed a deep learning-based artificial intelligence model using whole slide images (WSIs) to predict LNM and reduce overtreatment. The model was trained, validated, and internally tested on 160 surgically resected cases (72 LNM+, 88 LNM–) from 374 patients without prior ESD, achieving an AUC of 0.949 (95% CI: 0.912–0.986) on internal test. Further validation was performed on an external ESD cohort comprising clinically high-risk cases with invasion depths from MM to SM2. The model attained an accuracy of 90.1%, sensitivity of 81.8%, specificity of 91.4%, and an F1-score of 69.2%. It correctly classified 90.1% of samples, with a negative predictive value (NPV) of 96.9%. The high NPV and specificity underscore the model’s utility in minimizing overtreatment while preserving diagnostic accuracy in high-risk T1 esophageal cancer.

Tumour-associated microbiota directly colonise lesion sites and interact with the local immune microenvironment, yet their compositional shifts during the adenoma-to-cancer transition remain poorly characterised, particularly in archival tissue. Using five-region 16S ribosomal RNA (5R 16S rRNA) sequencing with a four-category contamination control framework, we compared tumour-associated microbiomes in formalin-fixed paraffin-embedded (FFPE) tissues from 25 colonic tubular adenoma and 25 colon cancer patients in a case–control design. Alpha diversity was significantly reduced in the colon cancer group across three indices (Shannon, Simpson, and Pielou’s evenness), reflecting shifts in community evenness rather than overall species loss. Beta diversity analysis, however, revealed no significant differences in global community composition between groups (ANOSIM: R = 0.026, p = 0.106; PERMANOVA: R ² = 0.024, p = 0.137), indicating that observed compositional differences are taxon-specific rather than community-wide. Across four independent statistical methods (ANCOM-BC2, LEfSe, Mann–Whitney U test, and Fisher’s exact test), Bacteroides caccae and Prevotella intermedia were consistently enriched in colon cancer tissue, suggesting compositional shifts in the tumour-associated microbiome that may be associated with colorectal carcinogenesis. Within the colon cancer group, Alistipes finegoldii abundance showed a negative correlation with tumour diameter across all colon cancer samples (Spearman r = −0.520, p = 0.008; n = 25, including 20 samples at the detection limit as tied ranks; only 5 samples had detectable abundance); given the very limited number of detectable samples, this finding should be regarded as strictly exploratory and requires prospective validation. These findings demonstrate that tumour-associated microbiome profiling in archival FFPE tissue is feasible under a rigorous contamination control framework, and provide preliminary evidence for taxon-specific compositional differences during colonic adenoma-to-carcinoma progression. The identified candidate taxa warrant prospective validation in larger cohorts and mechanistic investigation to clarify their roles in colon carcinogenesis.