AQP3-mediated activation of the AMPK/SIRT1 signaling pathway curtails gallstone formation in mice by inhibiting inflammatory injury of gallbladder mucosal epithelial cells.

BACKGROUND: Inflammatory injury of gallbladder mucosal epithelial cells affects the development of cholelithiasis, and aquaporin 3 (AQP3) is an important regulator of inflammatory response. This study reports a mechanistic insight into AQP3 regulating gallstone formation in cholelithiasis based on high-throughput sequencing. METHODS: A mouse model of cholelithiasis was induced using a high-fat diet, and the gallbladder tissues were harvested for high-throughput sequencing to obtain differentially expressed genes. Primary mouse gallbladder mucosal epithelial cells were isolated and induced with Lipopolysaccharides (LPS) to mimic an in vitro inflammatory injury environment. Cell biological phenotypes were detected by TdT-mediated dUTP Nick-End Labeling (TUNEL) assay, flow cytometry, Cell Counting Kit-8 (CCK-8) assay, and Trypan blue staining. In addition, enzyme linked immunosorbent assay (ELISA) determined the production of inflammatory factors in mouse gallbladder mucosa. RESULTS: Whole-transcriptome sequencing data analysis identified 489 up-regulated and 1007 down-regulated mRNAs. Bioinformatics analysis revealed that AQP3 was significantly down-regulated in mice with cholelithiasis. AQP3 might also confer an important role in LPS-induced gallbladder mucosal injury. Overexpression of AQP3 activated the AMPK (adenosine monophosphate-activated protein kinase) / SIRT1 (sirtuin-1) signaling pathway to reduce LPS-induced inflammatory injury of the gallbladder mucosa epithelium, thereby ameliorating gallbladder damage and repressing gallstone formation in mice. CONCLUSION: Data from our study highlight the inhibitory role of AQP3 in gallbladder damage and gallstone formation in mice by reducing inflammatory injury of gallbladder mucosal epithelial cells, which is achieved through activation of the AMPK/SIRT1 signaling pathway.

Characterization of cancer-associated fibroblasts (CAFs) and development of a CAF-based risk model for triple-negative breast cancer.

Triple-negative breast Cancer (TNBC) is a highly malignant cancer with unclear pathogenesis. Within the tumor microenvironment (TME), cancer-associated fibroblasts (CAFs) vitally influence tumor onset and progression. Thus, this research aimed to identify distinct subgroups of CAF using single-cell and TNBC-related information from the GEO and TCGA databases, respectively. The primary aim was to establish a novel predictive model based on the CAF features and their clinical relevance. Moreover, the CAFs were analyzed for their immune characteristics, response to immunotherapy, and sensitivity to different drugs. The developed predictive model demonstrated significant effectiveness in determining the prognosis of patients with TNBC, TME, and the immune landscape of the tumor. Of note, the expression of GPR34 was significantly higher in TNBC tissues compared to that in other breast cancer (non-TNBC) tissues, indicating that GPR34 plays a crucial role in the onset and progression of TNBC. In summary, this research has yielded a novel predictive model for TNBC that holds promise for the accurate prediction of prognosis and response to immunotherapy in patients with TNBC.

PDCL3 is a prognostic biomarker associated with immune infiltration in hepatocellular carcinoma.

BACKGROUND: Phosducin-like 3 (PDCL3) is a member of the photoreceptor family, characterized by a thioredoxin-like structural domain and evolutionary conservation. It plays roles in angiogenesis and apoptosis. Despite its significance, research on the biological role of PDCL3 in liver hepatocellular carcinoma (LIHC) remains limited. This study aims to explore the prognostic value and potential mechanisms of PDCL3 in cancer, particularly in LIHC, through bioinformatics analysis. METHODS: RNA-seq data and corresponding clinical information for pan-cancer and LIHC were extracted from the TCGA database to analyze PDCL3 expression and survival prognosis. Differential expression of PDCL3 was analyzed using the HPA database. GO and KEGG enrichment analysis were performed for PDCL3-associated genes. The relationship between PDCL3 expression and various immune cell types was examined using the TIMER website. Clinical samples were collected, and immunohistochemistry and immunofluorescence experiments were conducted to validate the differential expression of PDCL3 in LIHC and normal tissues. In vitro assays, including CCK-8, wound healing, Transwell, and colony formation experiments, were employed to determine the biological functions of PDCL3 in LIHC cells. RESULTS: Analysis from TIMER, GEPIA, UALCAN, and HPA databases revealed differential expression of PDCL3 in various tumors. Prognostic analysis from GEPIA and TCGA databases indicated that high PDCL3 expression was associated with poorer clinical staging and prognosis in LIHC. Enrichment analysis of PDCL3-associated genes revealed its involvement in various immune responses. TCGA and TIMER databases showed that high PDCL3 expression in LIHC decreased tumor immune activity by reducing macrophage infiltration. PDCL3 exhibited positive correlations with multiple immune checkpoint genes. Immunohistochemistry (IHC) and immunofluorescence (IF) experiments confirmed elevated PDCL3 expression in LIHC tissues compared to adjacent normal tissues. In vitro experiments demonstrated that PDCL3 promoted LIHC cell proliferation, migration, invasion, and colony-forming ability. CONCLUSION: PDCL3 is highly expressed in various cancer types. Our study suggests that elevated PDCL3 expression in hepatocellular carcinoma is associated with poorer prognosis and may serve as a potential diagnostic biomarker for LIHC. PDCL3 may regulate the biological functions of LIHC by modulating immune infiltration. However, the precise regulatory mechanisms of PDCL3 in cancer warrant further investigation.

Single-cell transcriptome sequencing reveals spatial distribution of IL34+ cancer-associated fibroblasts in hepatocellular carcinoma tumor microenvironment.

We utilized scRNA-seq, a well-established technology, to uncover the gene expression characteristics of IL34+ CAFs within HCC. We analyzed the related mechanisms through in vitro and in vivo assays. To begin, we acquired scRNA-seq datasets about HCC, which enabled us to identify distinct cell subpopulations within HCC tissues. We conducted a differential analysis to pinpoint DEGs associated with normal fibroblasts (NFs) and CAFs. Subsequently, we isolated NFs and CAFs, followed by the sorting of IL34+ CAFs. These IL34+ CAFs were then co-cultured with T cells and HCC cells to investigate their potential role in Tregs infiltration, CD8+ T cell toxicity, and the biological processes of HCC cells. We validated our findings in vivo using a well-established mouse model. Our analysis of HCC tissues revealed the presence of seven primary cell subpopulations, with the most significant disparities observed within fibroblast subpopulations. Notably, high IL34 expression was linked to increased expression of receptor proteins and enhanced proliferative activity within CAFs, with specific expression in CAFs. Furthermore, we identified a substantial positive correlation between IL34 expression and the abundance of Tregs. Both in vitro and in vivo experiments demonstrated that IL34+ CAFs promoted Tregs infiltration while suppressing CD8+ T cell toxicity. Consequently, this promoted the growth and metastasis of HCC. In summary, our study affirms that IL34+ CAFs play a pivotal role in augmenting the proliferative activity of CAFs, facilitating Tregs infiltration, and inhibiting CD8+ T cell toxicity, ultimately fostering the growth and metastasis of HCC.

Sphingosine 1-phosphate receptor 2 promotes the onset and progression of non-alcoholic fatty liver disease-related hepatocellular carcinoma through the PI3K/AKT/mTOR pathway.

PURPOSE: Recent studies have revealed an increase in the incidence rate of non-alcoholic fatty liver disease-related hepatocellular carcinoma (NAFLD-HCC). Furthermore, the association of Sphingosine 1-phosphate receptor 2 (S1PR2) with various types of tumours is identified, and the metabolism of conjugated bile acids (CBAs) performs an essential function in the onset and development of HCC. However, the association of CBA and S1PR2 with NAFLD-HCC is unclear. METHODS: The relationship between the expression of S1PR2 and the prognosis of patients suffering from NAFLD-HCC was investigated by bioinformatics techniques. Subsequently, the relationship between S1PR2 and the biological behaviours of HCC cell lines Huh 7 and HepG2 was explored by conducting molecular biology assays. Additionally, several in vivo animal experiments were carried out for the elucidation of the biological impacts of S1PR2 inhibitors on HCC cells. Finally, We used Glycodeoxycholic acid (GCDA) of CBA to explore the biological effects of CBA on HCC cell and its potential mechanism. RESULTS: High S1PR2 expression was linked to poor prognosis of the NAFLD-HCC patients. According to cellular assay results, S1PR2 expression could affect the proliferation, invasion, migration, and apoptosis of Huh 7 and HepG2 cells, and was closely associated with the G1/G2 phase of the cell cycle. The experiments conducted in the In vivo conditions revealed that the overexpression of S1PR2 accelerated the growth of subcutaneous tumours. In addition, JTE-013, an antagonist of S1PR2, effectively inhibited the migration and proliferation of HCC cells. Furthermore, the bioinformatics analysis highlighted a correlation between S1PR2 and the PI3K/AKT/mTOR pathway. GCDA administration further enhanced the expression levels of p-AKT, p-mTOR, VEGF, SGK1, and PKCα. Moreover, both the presence and absence of GCDA did not reveal any significant change in the levels of S1PR2, p-AKT, p-mTOR, VEGF, SGK1, and PKCα proteins under S1PR2 knockdown, indicating that CBA may regulates the PI3K/AKT/mTOR pathway by mediating S1PR2 expression. CONCLUSION: S1PR2 is a potential prognostic biomarker in NAFLD-HCC. In addition, We used GCDA in CBAs to treat HCC cell and found that the expression of S1PR2 was significantly increased, and the expression of PI3K/AKT/mTOR signalling pathway-related signal molecules was also significantly enhanced, indicating that GCDA may activate PI3K/AKT/mTOR signalling pathway by up-regulating the expression of S1PR2, and finally affect the activity of hepatocellular carcinoma cells. S1PR2 can be a candidate therapeutic target for NAFLD-HCC. Collectively, the findings of this research offer novel perspectives on the prevention and treatment of NAFLD-HCC.