PCMT1 regulates the migration, invasion, and apoptosis of prostate cancer through modulating the PI3K/AKT/GSK-3ß pathway.

Protein L-isoaspartate (D-aspartate) O-methyltransferase (PCMT1) is a repair enzyme that catalyzes the conversion of isomerized aspartic acid (iso-Asp) residues into their normal structure, thereby restoring the configuration and function of proteins. Studies have shown that PCMT1 is overexpressed in several tumors and affects patients' prognosis. However, there are few reports on the role of PCMT1 in prostate cancer (PCa). In the present research, with the assistance of The Cancer Genome Atlas Program (TCGA) database, we found that PCMT1 was overexpressed in PCa tissues. The results of quantitative reverse transcription-polymerase chain reaction (qRT-PCR), western blot and immunohistochemistry staining also showed that PCMT1 expression was significantly increased in PCa tissues and cell lines. In PCa clinical samples, PCMT1 expression was closely related to Gleason score, clinical stage, lymph node metastasis and bone metastasis. The experiments of overexpression and knockdown of PCMT1 in vitro or in vivo showed that PCMT1 can significantly promote the proliferation, migration and invasion of PCa cells, inhibit cell apoptosis, and promote the growth of PCa. We furthermore confirmed that PCMT1 regulated the migration, invasion and apoptosis of PCa cells by modulating the phosphatidylinositol 3-kinase/AKT kinase/glycogen-synthase kinase-3ß (PI3K/AKT/GSK-3ß) signaling pathway. Collectively, PCMT1 plays a cancer-facilitative role in PCa by promoting the proliferation, migration and invasion of PCa cells, and inhibiting apoptosis. Therefore, PCMT1 is considered to represent a novel target for treating PCa.

Enhancer of zeste homolog 2 modulates oxidative stress-mediated pyroptosis in vitro and in a mouse kidney ischemia-reperfusion injury model.

Enhancer of zeste homolog 2 (EZH2), a well-known methyltransferase, mediates histone H3 lysine 27 trimethylation (H3K27me3) and plays a crucial role in several kidney disease models. However, its role in renal ischemia/reperfusion (I/R) injury still remains unclear. In this study, we found that EZH2 was positively related to renal I/R injury and inhibition of EZH2 with DZNeP alleviated I/R injury and blocked the activation of oxidative stress and pyroptosis in vivo. Similarly, inhibition of EZH2 with either DZNeP or si-RNA also exerted an inhibitory effect on hypoxia/reoxygenation (H/R)-induced oxidative stress and pyroptosis in vitro. Moreover, further study revealed that ablation of reactive oxygen species (ROS) with N-acetyl-cysteine (NAC) suppressed pyroptosis in human renal proximal tubular epithelial cell line cells exposed to H/R stimulation. Furthermore, Nox4, which was positively related to the generation of ROS, was upregulated during H/R process, while it could be reversed by EZH2 inhibition. Consistently, Nox4-mediated ROS generation was attenuated upon inhibition of EZH2 with DZNeP or si-RNA. Additionally, the transcriptional activity of Nox4 was enhanced by the activation of ALK5/Smad2/3 signaling pathway, which was abolished by ALK5 knockdown in vitro. Finally, EZH2 inhibition blocked H/R and I/R-activated ALK5/Smad2/3 pathway and also resulted in an obvious decrease in the transcriptional activity and protein expression levels of Nox4. In conclusion, our results proved that EZH2 inhibition alleviated renal pyroptosis by blocking Nox4-dependent ROS generation through ALK5/Smad2/3 signaling pathway, indicating that EZH2 could be a potential therapeutic target for renal I/R injury.

Inhibition of PRMT5 Attenuates Oxidative Stress-Induced Pyroptosis via Activation of the Nrf2/HO-1 Signal Pathway in a Mouse Model of Renal Ischemia-Reperfusion Injury.

BACKGROUND: Extensive evidence has demonstrated that oxidative stress, pyroptosis, and proinflammatory programmed cell death are related to renal ischemia/reperfusion (I/R) injury. However, the underlying mechanism remains to be illustrated. Protein arginine methylation transferase 5 (PRMT5), which mediates arginine methylation involved in the regulation of epigenetics, exhibits a variety of biological functions and essential roles in diseases. The present study investigated the role of PRMT5 in oxidative stress and pyroptosis induced by I/R injury in a mouse model and in a hypoxia/reoxygenation (H/R) model of HK-2 cells. METHODS: C57 mice were used as an animal model. All mice underwent right nephrectomy, and the left renal pedicles were either clamped or not. Renal I/R injury was induced by ligating the left renal pedicle for 30 min followed by reperfusion for 24 h. HK-2 cells were exposed to normal conditions or stimulation through H/R. EPZ015666(EPZ)-a selective potent chemical inhibitor-and small interfering RNA (siRNA) were administered to suppress the function and expression of PRMT5. The levels of urea nitrogen and creatinine in the serum and renal tissue injury were assessed. Immunohistochemistry, western blotting, and reverse transcription-polymerase chain reaction were used to evaluate pyroptosis-related proteins including nod-like receptor protein-3, ASC, caspase-1, caspase-11, GSDMD-N, and interleukin-1ß. Cell apoptosis and cell viability were detected through flow cytometry, and the levels of reactive oxygen species (ROS) and hydrogen peroxide (H2O2) were measured. Ki-67 was used to assess the proliferation of renal tubular epithelium. In addition, the activity of malondialdehyde and superoxide dismutase was determined. RESULTS: I/R or H/R induced an increase in the expression of PRMT5. Inhibition of PRMT5 by EPZ alleviated oxidative stress and I/R- or H/R-induced pyroptosis. In renal tissue, the application of EPZ promoted the proliferation of tubular epithelium. In addition, H/R-induced pyroptosis in HK-2 cells was dependent on oxidative stress in vitro. Administration of either EPZ or siRNA led to decreased expression of pyroptosis-related proteins. Inhibition of PRMT5 also attenuated the I/R- or H/R-induced oxidative stress in vivo and in HK-2 cells, respectively. It also resulted in a distinct decrease in the levels of malondialdehyde and H2O2, and an apparent increase in superoxide dismutase activity in mouse renal tissue. Moreover, it led to a significant decrease in the levels of ROS and H2O2 in HK-2 cells. When activated, NF-E2-related factor/heme oxygenase-1 (Nrf2/HO-1)-a key regulator of various cytoprotective proteins that withstand oxidative damage-can decrease the generation of ROS. Nrf2/HO-1 was downregulated during I/R in tissues and H/R in HK-2 cells, and this effect was reversed by the PRMT5 inhibitor. Furthermore, the expressions of Nrf2 and HO-1 proteins were markedly upregulated by EPZ or siRNA against PRMT5. CONCLUSION: PRMT5 is involved in ischemia- and hypoxia-induced oxidative stress and pyroptosis in vitro and in vivo. Inhibition of PRMT5 may ameliorate renal I/R injury by suppressing oxidative stress and pyroptosis via the activation of the Nrf2/HO-1 pathway, as well as promoting the proliferation of tubular epithelium. Therefore, PRMT5 may be a promising therapeutic target.

Inhibition of Brd4 alleviates renal ischemia/reperfusion injury-induced apoptosis and endoplasmic reticulum stress by blocking FoxO4-mediated oxidative stress.

Ischemia/reperfusion injury (I/R) is one of the leading causes of acute kidney injury (AKI) that typically occurs in renal surgeries. However, renal I/R still currently lacks effective therapeutic targets. In this study, we proved that inhibition of Brd4 with its selective inhibitor, JQ1, could exert a protective role in renal I/R injury in mice. Inhibiting Brd4 with either JQ1 or genetic knockdown resulted in reduction of endoplasmic reticulum stress (ERS)-associated protein and proapoptotic protein expression both in I/R-induced injury and hypoxia/reoxygenation (H/R) stimulation in HK-2 cells. H/R-induced apoptosis and ERS depended on oxidative stress in vitro. Moreover, FoxO4, which is involved in the generation of hydrogen peroxide, was up-regulated during H/R stimulation-mediated apoptosis and ERS, and this upregulation could be abolished by Brd4 inhibition. Consistently, FoxO4-mediated ROS generation was attenuated upon inhibition of Brd4 with JQ1 or siRNA against Brd4. Further, the transcriptional activity of FoxO4 was suppressed by PI3K and AKT phosphorylation, which are upstream signals of FoxO4 expression, and were enhanced by Brd4 both in vivo and in vitro. In conclusion, our results proved that Brd4 inhibition blocked renal apoptotic and ERS protein expression by preventing FoxO4-dependent ROS generation through the PI3K/AKT pathway, indicating that Brd4 could be a potential therapeutic target for renal I/R injury.

Aged kidneys are refractory to autophagy activation in a rat model of renal ischemia-reperfusion injury.

BACKGROUND: Ischemia-reperfusion (I/R) injury is the most common cause of acute kidney injury (AKI). Numerous therapeutic approaches for I/R injury have been studied, including autophagy, particularly in animal models of renal I/R injury derived from young or adult animals. However, the precise role of autophagy in renal ischemia-reperfusion in the aged animal model remains unclear. The purpose of this study was to demonstrate whether autophagy has similar effects on renal I/R injury in young and aged rats. MATERIALS AND METHODS: All rats were divided into two age groups (3 months and 24 months) with each group being further divided into four subgroups (sham, I/R, I/R+Rap (rapamycin, an activator of autophagy), I/R+3-MA (3-methyladenine, an inhibitor of autophagy)). The I/R+Rap and I/R+3-MA groups were intraperitoneally injected with rapamycin and 3-MA prior to ischemia. We then measured serum levels of urea nitrogen, creatinine and assessed damage in the renal tissue. Immunohistochemistry was used to assess LC3-II and caspase-3, and Western blotting was used to evaluate the autophagy-related proteins LC3-II, Beclin-1 and P62. Apoptosis and autophagosomes were evaluated by TUNEL and transmission electron microscopy, respectively. RESULTS: Autophagy was activated in both young and aged rats by I/R and enhanced by rapamycin, although the level of autophagy was lower in the aged groups. In young rats, the activation of autophagy markedly improved renal function, reduced apoptosis in the renal tubular epithelial cells and the injury score in the renal tissue, thereby exerting protective effects on renal I/R injury. However, this level of protection was not present in aged rats. CONCLUSION: Our data indicated that the activation of autophagy was ineffective in aged rat kidneys. These discoveries may have major implications in that severe apoptosis in aged kidneys might be refractory to antiapoptotic effect induced by the activation of autophagy.

miR-181a-5p is downregulated and inhibits proliferation and the cell cycle in prostate cancer.

Prostate cancer (PCa) is one of the most common cancers in men worldwide. However, the detailed molecular mechanisms underlying PCa tumorigenesis and progression remain largely unclear. MicroRNAs are key regulators of gene post-transcriptional expression in human cancer. In this study, we used public datasets, including GSE21036, GSE14857 and GSE45604 to analyze the expression of miR-181a in PCa. We also explored the potential role of miR-181a by using bioinformatics analysis and gain of function assay. miR-181a was down-regulated in PCa. Bioinformatics analysis revealed miR-181a was significantly involved in regulating cell metabolic process and gene expression. Of note, gain of function assay results showed overexpression of miR-181a could significantly inhibit cell proliferation by inducing G1 cell cycle arrest. Our results suggest miR-181a-5p may be adiagnostic and therapeutic biomarker for prostate cancer.