BACKGROUND: The underlying mechanisms of gastric cancer (GC) remain unknown. Therefore, in this study, we employed a comprehensive approach, combining computational and experimental methods, to identify potential key genes and unveil the underlying pathogenesis and prognosis of GC.
METHODS: Gene expression profiles from GEO databases (GSE118916, GSE79973, and GSE29272) were analyzed to identify DEGs between GC and normal tissues. A PPI network was constructed using STRING and Cytoscape, followed by hub gene identification with CytoHubba. Investigations included expression and promoter methylation analysis, survival modeling, mutational and miRNA analysis, gene enrichment, drug prediction, and in vitro assays for cellular behaviors.
RESULTS: A total of 83 DEGs were identified in the three datasets, comprising 41 up-regulated genes and 42 down-regulated genes. Utilizing the degree and MCC methods, we identified four hub genes that were hypomethylated and up-regulated: COL1A1, COL1A2, COL3A1, and FN1. Subsequent validation of their expression and promoter methylation on clinical GC samples through targeted bisulfite sequencing and RT-qPCR analysis further confirmed the hypomethylation and overexpression of these genes in local GC patients. Furthermore, it was observed that these hub genes regulate tumor proliferation and metastasis in in vivo and exhibited mutations in GC patients.
CONCLUSIONS: We found four potential diagnostic and prognostic biomarkers, including COL1A1, COL1A2, COL3A1, and FN1 that may be involved in the occurrence and progression of GC.

Esophageal squamous cell carcinoma (ESCC) is a common malignancy of the gastrointestinal tract with a single therapeutic option and a lack of effective clinical therapeutic biomarkers. Extracellular matrix (ECM) remodeling plays a pro-carcinogenic role in a variety of malignancies, but its role in esophageal squamous carcinoma remains to be elucidated. In this study, we examined the expression levels of ECM remodeling markers in 71 pairs of esophageal squamous carcinoma tissues and normal tissues adjacent to the carcinoma using immunohistochemical staining, and analyzed their relationship with clinicopathological features and prognosis. The results suggested that extracellular matrix remodeling markers (integrin αV, fibronectin, MMP9) were abnormally highly expressed in esophageal squamous carcinoma tissues. There was a statistically significant difference between the positive expression of ECM remodeling and the TNM stage of esophageal squamous carcinoma, and there was no statistically significant correlation with age, gender and carcinoembryonic antigen expression, differentiation degree, T stage, and lymph node metastasis. Overall survival rate and overall survival time were significantly lower in patients with positive ECM remodeling expression, which was an independent risk factor for poor prognosisof esophageal squamous carcinoma.

Renal fibrosis is the most common pathway in progressive kidney diseases. The unilateral ureteral obstruction (UUO) model is used to induce progressive renal fibrosis. We evaluated the effects of irisin on renal interstitial fibrosis in UUO mice. The GSE121190, GSE36496, GSE42303, and GSE96101 datasets were downloaded from the Gene Expression Omnibus (GEO) database. In total, 656 differentially expressed genes (DEGs) were identified in normal and UUO mouse renal samples. Periostin and matrix metalloproteinase-2 (MMP-2) were selected to evaluate the effect of irisin on renal fibrosis in UUO mice. In UUO mice, irisin ameliorated renal function, decreased the expression of periostin and MMP-2, and attenuated epithelial-mesenchymal transition and extracellular matrix deposition in renal tissues. In HK-2 cells, irisin treatment markedly attenuated TGF-β1-induced expression of periostin and MMP-2. Irisin treatment also inhibited TGF-β1-induced epithelial-mesenchymal transition, extracellular matrix formation, and inflammatory responses. These protective effects of irisin were abolished by the overexpression of periostin and MMP-2. In summary, irisin treatment can improve UUO-induced renal interstitial fibrosis through the TGF-β1/periostin/MMP-2 signaling pathway, suggesting that irisin may be used for the treatment of renal interstitial fibrosis.

OBJECTIVE: To investigate the effect of tofacitinib, a pan-Janus kinase (JAK) inhibitor, on transforming growth factor-beta 1 (TGF-β1)-induced fibroblast to myofibroblast transition (FMT) and to explore its mechanism. To provide a theoretical basis for the clinical treatment of connective tissue disease-related interstitial lung disease (CTD-ILD).
METHODS: (1) Human fetal lung fibroblast 1 (HFL-1) were cultured in vitro, and 6 groups were established: DMSO blank control group, TGF-β1 induction group, and TGF-β1 with different concentrations of tofacitinib (0.5, 1.0, 2.0, 5.0 μmol/L) drug intervention experimental groups. CCK-8 was used to measure the cell viability, and wound-healing assay was performed to measure cell migration ability. After 48 h of combined treatment, quantitative real-time PCR (RT-PCR) and Western blotting were used to detect the gene and protein expression levels of α-smooth muscle actin (α-SMA), fibronectin (FN), and collagen type Ⅰ (COL1). (2) RT-PCR and enzyme-linked immunosorbnent assay (ELISA) were used to detect the interleukin-6 (IL-6) gene and protein expression changes, respectively. (3) DMSO carrier controls, 1.0 μmol/L and 5.0 μmol/L tofacitinib were added to the cell culture media of different groups for pre-incubation for 30 min, and then TGF-β1 was added to treat for 1 h, 6 h and 24 h. The phosphorylation levels of Smad2/3 and signal transducer and activator of transcription 3 (STAT3) protein were detected by Western blotting.
RESULTS: (1) Tofacitinib inhibited the viability and migration ability of HFL-1 cells after TGF-β1 induction. (2) The expression of α-SMA, COL1A1 and FN1 genes of HFL-1 in the TGF-β1-induced groups was significantly up-regulated compared with the blank control group (P < 0.05). Compared with the TGF-β1 induction group, α-SMA expression in the 5.0 μmol/L tofacitinib intervention group was significantly inhi-bited (P < 0.05). Compared with the TGF-β1-induced group, FN1 gene was significantly inhibited in each intervention group at a concentration of 0.5-5.0 μmol/L (P < 0.05). Compared with the TGF-β1-induced group, the COL1A1 gene expression in each intervention group did not change significantly. (3) Western blotting results showed that the protein levels of α-SMA and FN1 in the TGF-β1-induced group were significantly higher than those in the control group (P < 0.05), and there was no significant difference in the expression of COL1A1. Compared with the TGF-β1-induced group, the α-SMA protein level in the intervention groups with different concentrations decreased. And the differences between the TGF-β1-induced group and 2.0 μmol/L or 5.0 μmol/L intervention groups were statistically significant (P < 0.05). Compared with the TGF-β1-induced group, the FN1 protein levels in the intervention groups with different concentrations showed a downward trend, but the difference was not statistically significant. There was no difference in COL1A1 protein expression between the intervention groups compared with the TGF-β1-induced group. (4) After TGF-β1 acted on HFL-1 cells for 48 h, the gene expression of the IL-6 was up-regulated and IL-6 in culture supernatant was increased, the intervention with tofacitinib partly inhibited the TGF-β1-induced IL-6 gene expression and IL-6 in culture supernatant. TGF-β1 induced the increase of Smad2/3 protein phosphorylation in HFL-1 cells for 1 h and 6 h, STAT3 protein phosphorylation increased at 1 h, 6 h and 24 h, the pre-intervention with tofacitinib inhibited the TGF-β1-induced Smad2/3 phosphorylation at 6 h and inhibited TGF-β1-induced STAT3 phosphorylation at 1 h, 6 h and 24 h.
CONCLUSIONS: Tofacitinib can inhibit the transformation of HFL-1 cells into myofibroblasts induced by TGF-β1, and the mechanism may be through inhibiting the classic Smad2/3 pathway as well as the phosphorylation of STAT3 induced by TGF-β1, thereby protecting the disease progression of pulmonary fibrosis.

RBM15 functions as an oncogene in multi-type cancers. However, the reports on the roles of RBM15 in cervical cancer are limited. The purpose of this study was to investigate the potentials of RBM15 in cervical cancer. RT-qPCR was conducted to determine mRNA levels. Western was carried out to detect protein expression. CCK-8, colony formation and EdU assays were conducted to determine cell proliferation. Scratch and transwell assays were conducted to determine cell migration and invasion. MeRIP assay was conducted to determine N6-methyl adenosine (m6A) levels. Luciferase assay was conducted to verify the m6A sites of EZH2 and binding sites between EZH2 and promoter of FN1. ChIP assay was conducted to verify the interaction between EZH2 and FN1. The results showed that RBM15 was upregulated in cervical cancer patients and cells. Moreover, high levels of RBM15 predicted poor clinical outcomes. RBM15 knockdown inhibited the proliferation and epithelial-mesenchymal transition (EMT) of cervical cancer cells. RBM15 promoted the m6A modification of EZH2 as well as its protein translation. Additionally, EZH2 bound to the promoter of fibronectin 1 (FN1) and EZH2-FN1 axis is the cascade downstream of RBM15. Overexpressed EZH2 antagonized the effects of RBM15 knockdown and promoted the aggressiveness of cervical cancer cells. In summary, RBM15/EZH2/FN1 signaling cascade induces the proliferation and EMT of cervical cancer. Therefore, RBM15/EZH2/FN1 signaling may be a promising strategy for cervical cancer.

Triple-negative breast cancer (TNBC) is more aggressive and has a higher metastasis rate compared with other subtypes of breast cancer. Due to the lack of drug-targetable receptors, chemotherapy is now the only available systemic treatment for TNBC. However, some patients might still develop drug resistance and have poor prognosis. Therefore, novel molecular biomarkers and new treatment targets are urgently needed for patients with TNBC. To provide molecular insights into TNBC progression, we investigated the function and the underlying mechanism of Defective in cullin neddylation 1 domain containing 5 (DCUN1D5) in the regulation of TNBC. By TCGA dataset and surgical specimens with immunohistochemical (IHC) staining method, DCUN1D5 was identified to be significantly upregulated in TNBC tumor tissues and negatively associated with prognosis. A series of in vitro and in vivo experiments were performed to confirm the oncogenic role of DCUN1D5 in TNBC. Overexpression of FN1 or PI3K/AKT activator IGF-1 could restore the proliferative and invasive ability induced by DCUN1D5 knockdown and DCUN1D5 could act as a novel transcriptional target of transcription factor Yin Yang 1 (YY1). In conclusion, YY1-enhanced DCUN1D5 expression could promote TNBC progression by FN1/PI3K/AKT pathway and DCUN1D5 might be a potential prognostic biomarker and therapeutic target for TNBC treatment.

UNASSIGNED: Atherosclerosis is significantly influenced by endothelial cell activation and dysfunction. Studies have demonstrated the substantial presence of fibronectin (Fn) within atherosclerotic plaques, promoting endothelial inflammation and activation. However, cellular Fn (cFn) secreted by various cell types, including endothelial cells and smooth muscle cells, and plasma Fn (pFn) produced by hepatocytes. They are distinct forms of Fn that differ in both structure and function. The specific contribution of different types of Fn in promoting endothelial cell activation and dysfunction remain uncertain. Therefore, this study aimed to investigate the respective roles of pFn and endothelial cell-derived Fn (FnEC) in promoting endothelial cell activation and dysfunction.
UNASSIGNED: Initially, endothelial cell injury was induced by exposing the cells to oxidized low-density lipoprotein (ox-LDL) and subsequently we generated a mutant strain of aortic endothelial cells with Fn knockdown (FnEC-KD). The impact of the FnEC-KD arel the addition of pFn on the expression levels of inflammatory factors, vasoconstrictors, and diastolic factors were compared.
UNASSIGNED: The results showed that the FnEC-KD significantly inhibited ox-LDL-induced intercellular adhesion molecule 1 (ICAM-1, p < 0.05), vascular cell adhesion molecule (VCAM-1, p < 0.05), and endothelin (p < 0.05) expression, and nuclear factor kappa-B (NFκB, p < 0.05) activation. These results implied that FnEC-KD inhibited both endothelial cell activation and dysfunction. Surprisingly, the addition of pFn significantly inhibited the ox-LDL-induced ICAM-1 (p < 0.05), VCAM-1 (p < 0.05), and endothelin (p < 0.05) expression and NFκB (p < 0.05) activation. Implying that pFn inhibits endothelial cell activation and dysfunction. Additionally, the study revealed that ox-LDL stimulation enhanced the production of excessive nitric oxide, leading to severe endothelial cell damage.
UNASSIGNED: Aortic FnEC promotes endothelial cell activation and endothelial dysfunction, whereas pFn inhibits ox-LDL-induced endothelial cell activation and endothelial dysfunction.

BACKGROUND: Gastric cancer (GC) ranks fifth in global cancer incidence and third in mortality rate among all cancer types. Circular RNAs (circRNAs) have been extensively demonstrated to regulate multiple malignant biological behaviors in GC. Emerging evidence suggests that several circRNAs derived from FNDC3B play pivotal roles in cancer. However, the role of circFNDC3B in GC remains elusive.
METHODS: We initially screened circFNDC3B with translation potential via bioinformatics algorithm prediction. Subsequently, Sanger sequencing, qRT-PCR, RNase R, RNA-FISH and nuclear-cytoplasmic fractionation assays were explored to assess the identification and localization of circ0003692, a circRNA derived from FNDC3B. qRT-PCR and ISH were performed to quantify expression of circ0003692 in human GC tissues and adjacent normal tissues. The protein-encoding ability of circ0003692 was investigated through dual-luciferase reporter assay and LC/MS. The biological behavior of circ0003692 in GC was confirmed via in vivo and in vitro experiments. Additionally, Co-IP and rescue experiments were performed to elucidate the interaction between the encoded protein and c-Myc.
RESULTS: We found that circ0003692 was significantly downregulated in GC tissues. Circ0003692 had the potential to encode a novel protein FNDC3B-267aa, which was downregulated in GC cells. We verified that FNDC3B-267aa, rather than circ0003692, inhibited GC migration in vitro and in vivo. Mechanistically, FNDC3B-267aa directly interacted with c-Myc and promoted proteasomal degradation of c-Myc, resulting in the downregulation of c-Myc-Snail/Slug axis.
CONCLUSIONS: Our study revealed that the novel protein FNDC3B-267aa encoded by circ0003692 suppressed GC metastasis through binding to c-Myc and enhancing proteasome-mediated degradation of c-Myc. The study offers the potential applications of circ0003692 or FNDC3B-267aa as therapeutic targets for GC.

Mitochondrial dysfunction and oxidative stress are important mechanisms for secondary injury after traumatic brain injury (TBI), which result in progressive pathophysiological exacerbation. Although the Fibronectin type III domain-containing 5 (FNDC5) was reported to repress oxidative stress by retaining mitochondrial biogenesis and dynamics, its possible role in the secondary injury after TBI remain obscure. In present study, we observed that the level of serum irisin (the cleavage product of FNDC5) significantly correlated with the neurological outcomes of TBI patients. Knockout of FNDC5 increased the lesion volume and exacerbated apoptosis and neurological deficits after TBI in mice, while FNDC5 overexpression yielded a neuroprotective effect. Moreover, FNDC5 deficiency disrupted mitochondrial dynamics and function. Activation of Sirtuin 3 (SIRT3) alleviated FNDC5 deficiency-induced disruption of mitochondrial dynamics and bioenergetics. In neuron-specific SIRT3 knockout mice, FNDC5 failed to attenuate TBI-induced mitochondrial damage and brain injuries. Mechanically, FNDC5 deficiency led to reduced SIRT3 expression via enhanced ubiquitin degradation of transcription factor Nuclear factor erythroid 2-related factor 2 (NRF2), which contributed to the hyperacetylation and inactivation of key regulatory proteins of mitochondrial dynamics and function, including OPA1 and SOD2. Finally, engineered RVG29-conjugated nanoparticles were generated to selectively and efficiently deliver irisin to the brain of mice, which yielded a satisfactory curative effect against TBI. In conclusion, FNDC5/irisin exerts a protective role against acute brain injury by promoting SIRT3-dependent mitochondrial quality control and thus represents a potential target for neuroprotection after TBI.

Leucine-rich α2-glycoprotein-1 (LRG1) is overexpressed in various cancers, including non-small cell lung cancer (NSCLC), but its role in NSCLC cell metastasis is not well understood. In this study, NSCLC cell exosomes were analyzed using different techniques, and the impact of exosomal LRG1 on NSCLC cell behavior was investigated through various assays both in vitro and in vivo. The study revealed that LRG1, found abundantly in NSCLC cells and exosomes, enhanced cell proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT). Exosomal LRG1 was shown to promote NSCLC cell metastasis in animal models. Additionally, the interaction between LRG1 and fibronectin 1 (FN1) in the cytoplasm was identified. It was observed that FN1 could counteract the effects of LRG1 knockdown on cell regulation induced by exosomes derived from NSCLC cells. Overall, the findings suggest that targeting exosomal LRG1 or FN1 may hold therapeutic potential for treating NSCLC.