Epididymal RNase T2 contributes to astheno-teratozoospermia and intergenerational metabolic disorder through epididymosome-sperm interaction.

BACKGROUND: The epididymis is crucial for post-testicular sperm development which is termed sperm maturation. During this process, fertilizing ability is acquired through the epididymis-sperm communication via exchange of protein and small non-coding RNAs (sncRNAs). More importantly, epididymal-derived exosomes secreted by the epididymal epithelial cells transfer sncRNAs into maturing sperm. These sncRNAs could mediate intergenerational inheritance which further influences the health of their offspring. Recently, the linkage and mechanism involved in regulating sperm function and sncRNAs during epididymal sperm maturation are increasingly gaining more and more attention. METHODS: An epididymal-specific ribonuclease T2 (RNase T2) knock-in (KI) mouse model was constructed to investigate its role in developing sperm fertilizing capability. The sperm parameters of RNase T2 KI males were evaluated and the metabolic phenotypes of their offspring were characterized. Pandora sequencing technology profiled and sequenced the sperm sncRNA expression pattern to determine the effect of epididymal RNase T2 on the expression levels of sperm sncRNAs. Furthermore, the expression levels of RNase T2 in the epididymal epithelial cells in response to environmental stress were confirmed both in vitro and in vivo. RESULTS: Overexpression of RNase T2 caused severe subfertility associated with astheno-teratozoospermia in mice caput epididymis, and furthermore contributed to the acquired metabolic disorders in the offspring, including hyperglycemia, hyperlipidemia, and hyperinsulinemia. Pandora sequencing showed altered profiles of sncRNAs especially rRNA-derived small RNAs (rsRNAs) and tRNA-derived small RNAs (tsRNAs) in RNase T2 KI sperm compared to control sperm. Moreover, environmental stress upregulated RNase T2 in the caput epididymis. CONCLUSIONS: The importance was demonstrated of epididymal RNase T2 in inducing sperm maturation and intergenerational inheritance. Overexpressed RNase T2 in the caput epididymis leads to astheno-teratozoospermia and metabolic disorder in the offspring.

The circSPON2/miR-331-3p axis regulates PRMT5, an epigenetic regulator of CAMK2N1 transcription and prostate cancer progression.

BACKGROUND: Prostate cancer (PCa) is the most frequently diagnosed malignancy in men, and its mechanism remains poorly understood. Therefore, it is urgent to discover potential novel diagnostic biomarkers and therapeutic targets that can potentially facilitate the development of efficient anticancer strategies. METHODS: A series of functional in vitro and in vivo experiments were conducted to evaluate the biological behaviors of PCa cells. RNA pulldown, Western blot, luciferase reporter, immunohistochemistry and chromatin immunoprecipitation assays were applied to dissect the detailed underlying mechanisms. High-throughput sequencing was performed to screen for differentially expressed circRNAs in PCa and adjacent normal tissues. RESULTS: Upregulation of protein arginine methyltransferase 5 (PRMT5) is associated with poor progression-free survival and the activation of multiple signaling pathways in PCa. PRMT5 inhibits the transcription of CAMK2N1 by depositing the repressive histone marks H4R3me2s and H3R8me2s on the proximal promoter region of CAMK2N1, and results in malignant progression of PCa both in vitro and in vivo. Moreover, the expression of circSPON2, a candidate circRNA in PCa tissues identified by RNA-seq, was found to be associated with poor clinical outcomes in PCa patients. Further results showed that circSPON2 induced PCa cell proliferation and migration, and that the circSPON2-induced effects were counteracted by miR-331-3p. Particularly, circSPON2 acted as a competitive endogenous RNA (ceRNA) of miR-331-3p to attenuate the repressive effects of miR-331-3p on its downstream target PRMT5. CONCLUSIONS: Our findings showed that the epigenetic regulator PRMT5 aggravates PCa progression by inhibiting the transcription of CAMK2N1 and is modulated by the circSPON2/miR-331-3p axis, which may serve as a potential therapeutic target for patients with aggressive PCa.

M1 macrophage-derived exosomes impair beta cell insulin secretion via miR-212-5p by targeting SIRT2 and inhibiting Akt/GSK-3ß/ß-catenin pathway in mice.

AIMS/HYPOTHESIS: Macrophage levels are elevated in pancreatic islets, and the resulting inflammatory response is a major contributor to beta cell failure during obesity and type 2 diabetes mellitus. Previous studies by us and others have reported that exosomes released by macrophages play important roles in mediating cell-to-cell communication, and represent a class of inflammatory factors involved in the inflammatory process associated with type 2 diabetes mellitus. However, to date, no reports have demonstrated the effect of macrophage-derived exosomes on beta cells, and little is known regarding their underlying mechanisms in beta cell injury. Thus, we aimed to study the impact of macrophage-derived exosomes on islet beta cell injury in vitro and in vivo. METHODS: The phenotypic profiles of islet-resident macrophages were analysed in C57BL/6J mice fed a high-fat diet (HFD). Exosomes were collected from the medium of cultured bone marrow-derived macrophages (BMDMs) and from isolated islet-resident macrophages of HFD-fed mice (HFD-Exos). The role of exosomes secreted by inflammatory M1 phenotype BMDMs (M1-Exos) and HFD-Exos on beta cell function was assessed. An miRNA microarray and quantitative real-time PCR (qPCR) were conducted to test the level of M1-Exos-derived miR-212-5p in beta cells. Then, miR-212-5p was overexpressed or inhibited in M1-Exos or beta cells to determine its molecular and functional impact. RESULTS: M1-polarised macrophages were enriched in the islets of obese mice. M1 macrophages and islet-resident macrophages of HFD-fed mice impaired beta cell insulin secretion in an exosome-dependent manner. miR-212-5p was notably upregulated in M1-Exos and HFD-Exos. Enhancing the expression of miR-212-5p impaired beta cell insulin secretion. Blocking miR-212-5p elicited a significant improvement in M1-Exos-mediated beta cell insulin secretion during injury. Mechanistically, M1-Exos mediated an intercellular transfer of the miR-212-5p, targeting the sirtuin 2 gene and regulating the Akt/GSK-3ß/ß-catenin pathway in recipient beta cells to restrict insulin secretion. CONCLUSIONS/INTERPRETATION: A novel exosome-modulated mechanism was delineated for macrophage-beta cell crosstalk that drove beta cell dysfunction and should be explored for its therapeutic utility.

Ribonuclease Inhibitor 1 (RNH1) Regulates Sperm tsRNA Generation for Paternal Inheritance through Interacting with Angiogenin in the Caput Epididymis.

Environmental stressors can induce paternal epigenetic modifications that are a key determinant of the intergenerational inheritance of acquired phenotypes in mammals. Some of them can affect phenotypic expression through inducing changes in tRNA-derived small RNAs (tsRNAs), which modify paternal epigenetic regulation in sperm. However, it is unclear how these stressors can affect changes in the expression levels of tsRNAs and their related endonucleases in the male reproductive organs. We found that Ribonuclease inhibitor 1 (RNH1), an oxidation responder, interacts with ANG to regulate sperm tsRNA generation in the mouse caput epididymis. On the other hand, inflammation and oxidative stress induced by either lipopolysaccharide (LPS) or palmitate (PA) treatments weakened the RNH1-ANG interaction in the epididymal epithelial cells (EEC). Accordingly, ANG translocation increased from the nucleus to the cytoplasm, which led to ANG upregulation and increases in cytoplasmic tsRNA expression levels. In conclusion, as an antioxidant, RNH1 regulates tsRNA generation through targeting ANG in the mouse caput epididymis. Moreover, the tsRNA is an epigenetic factor in sperm that modulates paternal inheritance in offspring via the fertilization process.

Radiotherapy Enhancement for Human Pancreatic Carcinoma Using a Peptide-Gold Nanoparticle Hybrid.

Pancreatic ductal adenocarcinoma (PDAC) is radioresistant. Due to their strong X-ray absorption capacity, gold nanoparticles (AuNPs) have been used as radiosensitizers for cancer therapeutics. Herein, we describe a novel conjugate complex consisting of a peptide for targeting plectin-1 (PTP) specifically expressed on the PDAC cell membrane and AuNPs, termed AuNP-PTP, to be used for PDAC radiotherapy in vitro and in vivo. Previous studies revealed that compared with unmodified AuNPs, AuNP-PTP along with relevant low-energy X-ray irradiation of 6 MV at a dose of 2 Gy (RF) increased the targeting efficiency and induced apoptosis in treated PANC-1 cells and tumours. Importantly, extensive histopathological examination did not reveal evidence of acute or chronic injury in mice due to AuNPs or AuNP-PTP for up to six weeks despite the presence of X-ray exposure. The delicate AuNP-PTP hybrid provides a novel strategy to enhance radiotherapy efficiency in PDAC treatment.