Functional genomics reveal that the serine synthesis pathway is essential in breast cancer

Exosomes are small (∼30–140 nm) lipid bilayer-enclosed particles of endosomal origin. They are a subset of extracellular vesicles (EVs) that are secreted by most cell types. There has been growing interest in exosome research in the last decade due to their emerging role as intercellular messengers and their potential in disease diagnosis. Indeed, exosomes contain proteins, lipids, and RNAs that are specific to their cell origin and could deliver cargo to both nearby and distant cells. As a result, investigation of exosome cargo contents could offer opportunities for disease detection and treatment. Moreover, exosomes have been explored as natural drug delivery vehicles since they can travel safely in extracellular fluids and deliver cargo to destined cells with high specificity and efficiency. Despite significant efforts made in this relatively new field of research, progress has been held back by challenges such as inefficient separation methods, difficulties in characterization, and lack of specific biomarkers. In this review, we summarize the current knowledge in exosome biogenesis, their roles in disease progression, and therapeutic applications and opportunities in bioengineering. Furthermore, we highlight the established and emerging technological developments in exosome isolation and characterization. We aim to consider critical challenges in exosome research and provide directions for future studies.

Challenges and opportunities in exosome research—Perspectives from biology, engineering, and cancer therapy

In recent years, circular RNAs (circRNAs), a new star of non-coding RNA, have been emerged as vital regulators and gained much attention for involvement of initiation and progression of diverse kinds of human diseases, especially cancer. However, regulatory role, clinical significance and underlying mechanisms of circRNAs in triple-negative breast cancer (TNBC) still remain largely unknown. Here, the expression profile of circRNAs in 4 pairs of TNBC tissues and adjacent non-tumor tissues was analyzed by RNA-sequencing. Quantitative real-time PCR and in situ hybridization were used to determine the level and prognostic values of circAGFG1 in two TNBC cohorts. Then, functional experiments in vitro and in vivo were performed to investigate the effects of circAGFG1 on tumor growth and metastasis in TNBC. Mechanistically, fluorescent in situ hybridization, dual luciferase reporter assay, RNA pull-down and RNA immunoprecipitation experiments were performed to confirm the interaction between circAGFG1 and miR-195-5p in TNBC. We found that circAGFG1 was evidently up-regulated in TNBC, and its level was correlated with clinical stage, pathological grade and poor prognosis of patients with TNBC. The results indicated that circAGFG1 could promote TNBC cell proliferation, mobility and invasion as well as tumorigenesis and metastasis in vivo. Mechanistic analysis showed that circAGFG1 may act as a ceRNA (competing endogenous RNA) of miR-195-5p to relieve the repressive effect of miR-195-5p on its target cyclin E1 (CCNE1). Our findings suggest that circAGFG1 promotes TNBC progression through circAGFG1/miR-195-5p/CCNE1 axis and it may serve as a new diagnostic marker or target for treatment of TNBC patients.

/pdf/the-circrna-circagfg1-acts-as-a-sponge-of-mir-195-5p-to-3z0yw5l81v.pdf

The circRNA circAGFG1 acts as a sponge of miR-195-5p to promote triple-negative breast cancer progression through regulating CCNE1 expression

Colorectal cancer is an important public health problem. Several screening methods have been shown to be effective in reducing colorectal cancer mortality. The objective of this review was to assess the cost-effectiveness of the different colorectal cancer screening methods and to determine the preferred method from a cost-effectiveness point of view. Five databases (MEDLINE, EMBASE, the Cost-Effectiveness Analysis Registry, the British National Health Service Economic Evaluation Database, and the lists of technology assessments of the Centers for Medicare and Medicaid Services) were searched for cost-effectiveness analyses published in English between January 1993 and December 2009. Fifty-five publications relating to 32 unique cost-effectiveness models were identified. All studies found that colorectal cancer screening was cost-effective or even cost-saving compared with no screening. However, the studies disagreed as to which screening method was most effective or had the best incremental cost-effectiveness ratio for a given willingness to pay per life-year gained. There was agreement among studies that the newly developed screening tests of stool DNA testing, computed tomographic colonography, and capsule endoscopy were not yet cost-effective compared with the established screening options.

cost effectiveness of colorectal cancer screenting

Objective High-fat diet (HFD)-induced metabolic disorders can lead to impaired sperm production. We aim to investigate if HFD-induced gut microbiota dysbiosis can functionally influence spermatogenesis and sperm motility. Design Faecal microbes derived from the HFD-fed or normal diet (ND)-fed male mice were transplanted to the mice maintained on ND. The gut microbes, sperm count and motility were analysed. Human faecal/semen/blood samples were collected to assess microbiota, sperm quality and endotoxin. Results Transplantation of the HFD gut microbes into the ND-maintained (HFD-FMT) mice resulted in a significant decrease in spermatogenesis and sperm motility, whereas similar transplantation with the microbes from the ND-fed mice failed to do so. Analysis of the microbiota showed a profound increase in genus Bacteroides and Prevotella, both of which likely contributed to the metabolic endotoxaemia in the HFD-FMT mice. Interestingly, the gut microbes from clinical subjects revealed a strong negative correlation between the abundance of Bacteroides-Prevotella and sperm motility, and a positive correlation between blood endotoxin and Bacteroides abundance. Transplantation with HFD microbes also led to intestinal infiltration of T cells and macrophages as well as a significant increase of pro-inflammatory cytokines in the epididymis, suggesting that epididymal inflammation have likely contributed to the impairment of sperm motility. RNA-sequencing revealed significant reduction in the expression of those genes involved in gamete meiosis and testicular mitochondrial functions in the HFD-FMT mice. Conclusion We revealed an intimate linkage between HFD-induced microbiota dysbiosis and defect in spermatogenesis with elevated endotoxin, dysregulation of testicular gene expression and localised epididymal inflammation as the potential causes. Trial registration number NCT03634644.

https://gut.bmj.com/content/gutjnl/early/2020/01/02/gutjnl-2019-319127.full.pdf

Impairment of spermatogenesis and sperm motility by the high-fat diet-induced dysbiosis of gut microbes

Summary
Exosomes are membranous nanovesicles of endocytic origin that carry and transfer regulatory bioactive molecules and mediate intercellular communication between cells and tissues. Although seminal exosomes have been identified in human seminal plasma, their exact composition and possible physiologic function remain unknown. The objective of this study was to perform a comprehensive proteomics analysis of exosomes derived from human seminal plasma. Seminal exosomes were isolated and purified from 12 healthy donors using a 30% sucrose cushion-based exosome-isolation protocol, followed by characterization by western blot, transmission electron microscopy, and nanoparticle tracking analysis before performing extensive liquid chromatography tandem mass spectrometry proteomics analysis. The identified proteins were analyzed by bioinformatics analysis, and seminal exosomes-associated proteins were selectively validated by western blot. A total of 1474 proteins were identified in all seminal exosomes samples, with Gene Ontology analysis demonstrating that these identified seminal exosomes-associated proteins were mostly linked to ‘exosomes,’ ‘cytoplasm,’ and ‘cytosol.’ Bioinformatics analysis indicated that these proteins were mainly involved in biologic processes, including metabolism, energy pathways, protein metabolism, cell growth and maintenance, and transport. Of these identified proteins, PHGDH, LGALS3BP, SEMG1, ACTB, GAPDH, and the exosomal-marker protein ALIX were validated by western blot. This study provided a more comprehensive description of the seminal exosomes proteome and could also be a resource for further screening of biomarkers and comparative proteomics studies, including those associated with male infertility and prostate cancer.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/andr.12412

Comprehensive proteomics analysis of exosomes derived from human seminal plasma

Cysteine-rich secretory protein 2 (CRISP2) is an important sperm protein and plays roles in spermatogenesis, modulation of flagellar motility, acrosome reaction, and gamete fusion. Clinical evidence shows a reduced CRISP2 expression in spermatozoa from asthenozoospermic patients, but the molecular mechanism underlying its reduction remains unknown. Herein, we carried out a study focusing on the CRISP2 reduction and its roles in asthenozoospermia. Initially, through analyzing CRISP2 expression and methylation on CRISP2 promoter activity in sperm, we observed a decreased expression of CRISP2 protein rather than its mRNA in the ejaculated spermatozoa from asthenozoospermic patients and no methylation in the CRISP2 promoter, suggesting CRISP2 expression may be regulated in the sperm at the posttranscriptional level. Subsequently, we found that microRNA 27b (miR-27b), predicted as a candidate regulator of CRISP2 using bioinformatics, was highly expressed in the ejaculated spermatozoa from asthenozoosp...

The Expression of Cysteine-Rich Secretory Protein 2 (CRISP2) and Its Specific Regulator miR-27b in the Spermatozoa of Patients with Asthenozoospermia

BACKGROUND Semenogelin 1 (SEMG1) is an important secretory protein in spermatozoa involved in the formation of a gel matrix encasing ejaculated spermatozoa. Previous studies show that the SEMG1 gene is highly expressed in spermatozoa from patients with asthenozoospermia (AZS); however, the underlying molecular mechanisms are not yet clear. OBJECTIVES To study the molecular mechanism of high expression of SEMG1 gene and its potential roles in AZS. MATERIALS AND METHODS Western blot and real-time PCR were used to detect the expression levels of SEMG1 protein and mRNA in the ejaculated spermatozoa from normozoospermic males and AZS patients. Bioinformatics analysis was used to predict miRNAs targeting for SEMG1 3'-untranslated region detection of the expression levels of all the candidate miRNAs in ejaculatory spermatozoa in AZS patients or normozoospermic volunteers. Luciferase reporter assays were performed to confirm it can directly bind to SEMG1. Correlation of miR-525-3p and SEMG1 mRNA expression with clinical sperm parameters were also analyzed. Finally, we conducted a follow-up study of reproductive history about all the subjects. RESULTS SEMG1 mRNA and protein level were significantly higher in AZS patients compared to that in normozoospermic volunteers (p < 0.001). Subsequently, microRNA-525-3p (miR-525-3p) which was predicted as a candidate regulator of SEMG1 was found lower expressed in ejaculatory spermatozoa in AZS patients (p = 0.0074). Luciferase experiment revealed that microRNA-525-3p could directly target SEMG1 3'-untranslated region and suppress its expression. Importantly, our retrospective follow-up study showed that both low miR-525-3p expression and high SEMG1 expression level was significantly associated with low progressive sperm motility, abnormal sperm morphology, and infertility. DISCUSSION AND CONCLUSION The elevated expression of SEMG1 and reduced expression of miR-525-3p are associated with AZS and male infertility. Our study provides a potential therapeutic target for the treatment of male infertility or for male contraception.

Expressions of miR‐525‐3p and its target gene SEMG1 in the spermatozoa of patients with asthenozoospermia

Uncontrolled proliferation is thought to be the most fundamental characteristic of cancer. Detailed knowledge of cancer cell proliferation mechanisms would not only benefit understanding of cancer progression, but may also provide new clues for developing novel therapeutic strategies. In vitro function of MNX1 (Motor neuron and pancreas homeobox 1) in bladder cancer cell was evaluated using MTT assay, colony formation assay, and bromodeoxyuridine incorporation assay. Real-time PCR and western blotting were performed to detect MNX1 and CCNE1/2 expressions. In vivo tumor growth was conducted in BALB/c-nu mice. We reported that MNX1 is responsible for sustaining bladder cancer cell proliferation. Abnormal MNX1 upregulation in bladder cancer cell lines and 167 human tissue specimens; high MNX1 expression levels correlated significantly with shorter 5-year overall and relapse-free survival in the bladder cancer patients. Furthermore, MNX1 overexpression accelerated bladder cancer cell proliferation and tumorigenicity both in vitro and in vivo, whereas MNX1 downregulation arrested it. In addition, MNX1 transcriptionally upregulated CCNE1 and CCNE2 by directly bounding to their promoters, which promoted G1–S transition in the bladder cancer cells. These findings reveal an oncogenic role and novel regulatory mechanism of MNX1 in bladder cancer progression and suggest that MNX1 is a potential prognostic biomarker and therapeutic target.

/pdf/motor-neuron-and-pancreas-homeobox-1-hlxb9-promotes-28z6blvwdk.pdf

Motor neuron and pancreas homeobox 1/HLXB9 promotes sustained proliferation in bladder cancer by upregulating CCNE1/2

Arginine methyltransferase 5 (PRMT5) is involved in a variety of cancers. We used bioinformatics analysis to investigate PRMT5 overexpression in bladder urothelial cancer (BUC) and its clinical significance. We also conducted molecular biology experiments to investigate the effect of PRMT5 on the phenotype of BUC cells in vitro and in vivo. PRMT5 was found to be upregulated in BUC tissue in the Oncomine and The Cancer Genome Atlas databases. We validated the results from these databases in a cohort of BUC samples. Kaplan-Meier and Cox multivariate analyses demonstrated that PRMT5 upregulation is an independent prognostic risk factor for BUC. The in vitro and in vivo phenotypic experiments found that downregulated expression of PRMT5 in BUC cells inhibits BUC cell proliferation and aggression. In addition, gene set enrichment analysis demonstrated that PRMT5 knockdown leads to cell cycle G1/S arrest, deactivation of Akt, and mTOR phosphorylation in BUC cells. These results suggest that PRMT5 could be used as a potential molecular marker for BUC in the future.

/pdf/high-prmt5-expression-is-associated-with-poor-overall-3jjrg7jkuf.pdf

Mingkun Chen

Papers

Comprehensive proteomics analysis of exosomes derived from human seminal plasma

The Expression of Cysteine-Rich Secretory Protein 2 (CRISP2) and Its Specific Regulator miR-27b in the Spermatozoa of Patients with Asthenozoospermia

Expressions of miR‐525‐3p and its target gene SEMG1 in the spermatozoa of patients with asthenozoospermia

Motor neuron and pancreas homeobox 1/HLXB9 promotes sustained proliferation in bladder cancer by upregulating CCNE1/2

High PRMT5 expression is associated with poor overall survival and tumor progression in bladder cancer.