Exosomal MiR-381 from M2-polarized macrophages attenuates urethral fibroblasts activation through YAP/GLS1-regulated glutaminolysis.

OBJECTIVE AND DESIGN: Post-traumatic urethral stricture is a clinical challenge for both patients and clinicians. Targeting glutamine metabolism to suppress excessive activation of urethral fibroblasts (UFBs) is assumed to be a potent and attractive strategy for preventing urethral scarring and stricture. MATERIAL OR SUBJECTS: In cellular experiments, we explored whether glutaminolysis meets the bioenergetic and biosynthetic demands of quiescent UFBs converted into myofibroblasts. At the same time, we examined the specific effects of M2-polarized macrophages on glutaminolysis and activation of UFBs, as well as the mechanism of intercellular signaling. In addition, findings were further verified in vivo in New Zealand rabbits. RESULTS: It revealed that glutamine deprivation or knockdown of glutaminase 1 (GLS1) significantly inhibited UFB activation, proliferation, biosynthesis, and energy metabolism; however, these effects were rescued by cell-permeable dimethyl α-ketoglutarate. Moreover, we found that exosomal miR-381 derived from M2-polarized macrophages could be ingested by UFBs and inhibited GLS1-dependent glutaminolysis, thereby preventing excessive activation of UFBs. Mechanistically, miR-381 directly targets the 3'UTR of Yes-associated protein (YAP) mRNA to reduce its stability at the transcriptional level, ultimately downregulating expression of YAP, and GLS1. In vivo experiments revealed that treatment with either verteporfin or exosomes derived from M2-polarized macrophages significantly reduced urethral stricture in New Zealand rabbits after urethral trauma. CONCLUSION: Collectively, this study demonstrates that exosomal miR-381 from M2-polarized macrophages reduces myofibroblast formation of UFBs and urethral scarring and stricture by inhibiting YAP/GLS1-dependent glutaminolysis.

Integrative Analysis Developing and Validating Potential Candidate Biomarkers for Cancer Stemness Features of Pan-Renal Cell Carcinoma.

In our study, 49 key genes significantly associated with renal cell carcinoma (RCC) stemness were obtained. Next, we developed a molecular prognostic signature associated with stemness features of pan-RCC. The difference in overall survival (OS) between the high- and low-risk groups was statistically significant (p < .05). The area under the receiver operating characteristic curve for 1-year OS, 5-year OS, and 10-year OS was 0.759, 0.712, and 0.918, respectively. The results of validation in The Cancer Genome Atlas cohort and International Cancer Genome Consortium cohort revealed the predictive capability of this signature. Furthermore, we selected three genes and further validation showed that these three hub genes were potential hub biomarkers for pan-RCC stemness features.

ARPC1A correlates with poor prognosis in prostate cancer and is up-regulated by glutamine metabolism to promote tumor cell migration, invasion and cytoskeletal changes.

OBJECTIVE: This study aimed to identify potential biomarkers for prostate cancer (PCa) progression and metastasis, and to discern their biological functions. METHODS: Bioinformatics methods were used to screen for hub genes. The expression level of key hub genes in PCa was determined and their prognostic significance was examined. A series of functional assays were performed to investigate the function of the highest-ranking hub gene. RESULTS: Actin related protein 2/3 complex subunit 1A (ARPC1A) was identified as the hub gene. ARPC1A was highly expressed in PCa tissues and cell lines, and was an independent prognostic factor for predicting biochemical recurrence after radical prostatectomy and overall survival of PCa patients. Knockdown of ARPC1A inhibited PCa cell migration, invasion and cytoskeleton formation, but had no impact on cell proliferation and cell cycle progression. In vivo, ARPC1A overexpression promoted lung metastasis of PCa, but had no efffect on tumor growth. Additionally, glutamine metabolism was identified as an upstream regulator of ARPC1A, and promoted migration, invasion and cytoskeletal changes of PCa cell through ARPC1A. CONCLUSION: These findings suggested that ARPC1A, which correlates with poor prognosis in PCa, functions downstream of glutamine metabolism to regulate cytoskeletal changes, cellular migration and cellular invasion in this disease.

Development and Validation of Novel Biomarkers Related to M2 Macrophages Infiltration by Weighted Gene Co-Expression Network Analysis in Prostate Cancer.

M2-tumor-associated macrophages (TAMs) work as a promoter in the processes of bone metastases, chemotherapy resistance, and castration resistance in prostate cancer (PCa), but how M2-TAMs affect PCa has not been fully understood. In this study, we analyzed the proportion of tumor-infiltrating immune cells using the CIBERSORT algorithm, based on samples from the Cancer Genome Atlas database. Then we performed weighted gene co-expression network analysis to examine the modules concerning infiltrated M2-TAMs. Gene Ontology analysis and pathway enrichment analysis were performed for functional annotation and a protein-protein interaction network was constructed. The International Cancer Genomics Consortium cohort was used as a validation cohort. The red module showed the most correlation with M2-TAMs in PCa. Biological processes and pathways were mainly associated with the immune-related processes, as revealed by functional annotation. Four hub genes were screened: ACSL1, DLGAP5, KIF23 and NCAPG. Further validation showed that the four hub genes had a higher expression level in tumor tissues than that in normal tissues, and they were good prognosis biomarkers for PCa. In conclusion, these findings contribute to understanding the underlying molecular mechanisms of how M2-TAMs affect PCa, and looking for the potential biomarkers and therapeutic targets for PCa patients.

Mesenchymal Stem Cell-Derived Exosomes Induced by IL-1ß Attenuate Urethral Stricture Through Let-7c/PAK1/NF-κB-Regulated Macrophage M2 Polarization.

BACKGROUND: Urethral stricture is a clinical challenge for both patients and clinicians. Post-traumatic urethral stricture is associated with formation of scar tissue caused by excessive inflammation. The aim of this study is exploring potential therapeutic strategies for this condition. METHODS: In vivo experiments on New Zealand rabbits and in vitro experiments on THP-1 monocytes and urethral fibroblasts were performed to investigate the effects on post-traumatic urethral stricture of exosomes isolated from IL-1ß-treated mesenchymal stem cells (Exo-MSCsIL-1ß) and the role of macrophage M2 polarization in this process. Additionally, related signaling and mechanism behind were explored. RESULTS: In a New Zealand rabbit model of post-traumatic urethral stricture, injection of Exo-MSCsIL-1ß significantly reduced urethral stricture and collagen fiber accumulation compared with Exo-MSCs. Addition of Exo-MSCsIL-1ß to THP-1 monocytes in vitro induced M2 macrophage polarization, which, in turn, inhibited activation of urethral fibroblasts and synthesis of collagen. Mechanistically, Exo-MSCsIL-1ß were found to contain high levels of the microRNA let-7c, and luciferase reporter assays showed that let-7c interacted with the 3'UTR of PAK1 mRNA. Transfection of THP-1 cells with a let-7c mimic downregulated PAK1 expression and inhibited activation of the NF-κB signaling pathway. CONCLUSION: These results support a role for let-7c-containing Exo-MSCsIL-1ß in reducing urethral stricture via inhibition of PAK1-NF-κB signaling, M2 macrophage polarization, and differentiation of urethral myofibroblasts.

The prognostic value of homeobox A9 (HOXA9) methylation in solid tumors: a systematic review and meta-analysis.

BACKGROUND: The prognosis of homeobox A9 (HOXA9) methylation have been assessed in a variety of cancers; nevertheless, the results remain undetermined due to discrete outcome and the limitations of small sample size. Therefore, we conducted a meta-analysis to explore the effect of HOXA9 methylation on the prognostic outcomes of patients with solid tumors. METHODS: Qualified studies were verified by searching PubMed, Excerpta Medica Database and Web of Science until September, 2020. Clinicopathological factors and hazard ratio (HR) of 95% confidence interval (95% CI) were selected. Subgroup analysis including carcinoma category, analysis method and sample size were adopted. RESULTS: In the meta-analysis 1,031 patients with solid carcinoma from 7 eligible investigations were involved. Among human cancer we discovered that the high HOXA9 methylation level was negative correlative with overall survival (OS) (HR =2.36; 95% CI: 1.70-3.26). In the subgroup analysis, we found HOXA9 methylation over-expression had statistical significance with poorer OS in lung cancer patients (HR =3.08, 95% CI: 1.70-5.55, P=0.002) and non-lung cancer (HR =2.10, 95% CI: 1.42-3.10, P=0.0002). Similar result was found in sample size. Greater than or equal to 100 (HR =2.31, 95% CI: 1.54-3.45, P<0.0001) and less than 100 (HR =2.45, 95% CI: 1.42-4.23, P=0.001). DISCUSSION: HOXA9 methylation has a significantly estimable biomarker of predicting poor prognosis and a potential target for therapy in solid malignant carcinoma from our meta-analysis.

Identification of novel genes in testicular cancer microenvironment based on ESTIMATE algorithm-derived immune scores.

Testicular cancer is the most common solid malignancy among young men. We downloaded data of testicular cancer patients from The Cancer Genome Atlas database to find novel genes in the testicular cancer microenviroment based on ESTIMATE algorithm-derived immune scores. A total of 156 cases of testicular cancer were included in this study and 165 cases of normal testicular tissues were used. We divided the testicular cancer patients into high- and low-score groups based on their immune scores. We identified 1,226 differentially expressed genes (fold change > 2, false discovery rate < 0.05), including 688 downregulated genes and 538 upregulated genes, between these two groups. The top Gene Ontology terms were involved in the immune response-regulating cell surface receptor signaling pathway, immune response-activating cell surface receptor signaling pathway, external side of the plasma membrane, and receptor ligand activity. By performing the Kyoto Encyclopedia of Genes and Genomes analysis, we demonstrated that cAMP signaling pathway was highly enriched among these differentially expressed genes. High expression of LINC01564, LINC02208, ODAM, RNA5SP111, and RNU6-196P were found to be associated with poor overall survival. The expression of genes was further validated by the Human Protein Atlas and only ALB and IFNG were demonstrated to be differentially expressed between testis tissue and testicular cancer tissue.

Development and validation of a molecular prognostic index of bladder cancer based on immunogenomic landscape analysis.

BACKGROUND: Bladder cancer (BCa) is one of the important tumors that have been proven to be treatable with immunotherapy. This study aims to identify and validate a molecular prognostic index of BCa based on immunogenomic landscape analysis. METHODS: The cancer genome atlas (TCGA) database and immunology database and analysis portal (ImmPort) database were used to identified differentially expressed immune-related genes (IRGs). Prognostic IRGs were screened and protein-protein interaction (PPI) network was constructed. Multivariate Cox analysis was performed to develop a molecular prognostic index of BCa. Internal and external validation were then performed in TCGA cohort and GEO cohort, respectively. Besides, we also explore the relationship between this index and clinical characteristics, immune cell infiltration and tumor microenvironment. RESULTS: A total of 61 prognostic IRGs were identified and a molecular prognostic index was developed. The top four hub genes included MMP9, IGF1, CXCL12 and PGF. The difference in overall survival between high-risk group and low-risk group was statistically significant. The area under curve of the receiver operating characteristic (ROC) curve was 0.757, suggesting the potential for this index. Besides, Internal validation using TCGA cohort and external validation using GEO cohort indicated that this index was of great performance in predicting outcome. T cells CD8, T cells CD4 memory activated, T cells follicular helper, macrophages M0, macrophages M2 and neutrophils were significantly associated with prognosis of BCa patients. Female, high grade, stage III&IV, N1-3 and T3-4 were associated significantly with higher risk score compared with male, low grade, stage I&II, N0 and T1-2, respectively. High risk score had a positive association with higher stromal score and ESTIMATE score while high risk score had a negative association with tumor purity. CONCLUSIONS: This study identified several prognostic immune-related genes of clinical value. Besides, we developed and validated a molecular index based on immunogenomic landscape analysis, which performed well in predicting prognosis of BCa. Further researches are needed to verify the effectiveness of this index and these vital genes.

Development and validation of hub genes for lymph node metastasis in patients with prostate cancer.

Lymph node metastasis is one of the most important independent risk factors that can negatively affect the prognosis of prostate cancer (PCa); however, the exact mechanisms have not been well studied. This study aims to better understand the underlying mechanism of lymph node metastasis in PCa by bioinformatics analysis. We analysed a total of 367 PCa cases from the cancer genome atlas database and performed weighted gene co-expression network analysis to explore some modules related to lymph node metastasis. Gene Ontology analysis and pathway enrichment analysis were conducted for functional annotation, and a protein-protein interaction network was built. Samples from the International Cancer Genomics Consortium database were used as a validation set. The turquoise module showed the most relevance with lymph node metastasis. Functional annotation showed that biological processes and pathways were mainly related to activation of the processes of cell cycle and mitosis. Four hub genes were selected: CKAP2L, CDCA8, ERCC6L and ARPC1A. Further validation showed that the four hub genes well-distinguished tumour and normal tissues, and they were good biomarkers for lymph node metastasis of PCa. In conclusion, the identified hub genes facilitate our knowledge of the underlying molecular mechanism for lymph node metastasis of PCa.