USF1-mediated ALKBH5 stabilizes FLII mRNA in an m6A-YTHDF2-dependent manner to repress glycolytic activity in prostate adenocarcinoma.

Upstream-stimulating factor 1 (USF1) is a ubiquitously expressed transcription factor implicated in multiple cellular processes, including metabolism and proliferation. This study focused on the function of USF1 in glycolysis and the malignant development of prostate adenocarcinoma (PRAD). Bioinformatics predictions suggested that USF1 is poorly expressed in PRAD. The clinical PRAD samples revealed a low level of USF1, which was correlated with an unfavorable prognosis. Artificial upregulation of USF1 significantly repressed glycolytic activity in PRAD cells and reduced cell growth and metastasis in vitro and in vivo. Potential downstream genes of USF1 were probed by integrated bioinformatics analyses. The chromatin immunoprecipitation and luciferase assays indicated that USF1 bound to the α-ketoglutarate-dependent dioxygenase alkB homolog 5 (ALKBH5) promoter for transcription activation. Flightless I (FLII) was identified as the gene showing the highest degree of correlation with ALKBH5. As an m6A demethylase, ALKBH5 enhanced FLII mRNA stability by inducing m6A demethylation in an m6A-YTH N6-methyladenosine RNA-binding protein F2 (YTHDF2)-dependent manner. Either silencing of ALKBH5 or FLII blocked the role of USF1 in PARD cells and restored glycolysis, cell proliferation, and invasion. This study demonstrates that USF1 activates ALKBH5 to stabilize FLII mRNA in an m6A-YTHDF2-dependent manner, thereby repressing glycolysis processes and the progression of PRAD.

Tenuigenin activates the IRS1/Akt/mTOR signaling by blocking PTPN1 to inhibit autophagy and improve locomotor recovery in spinal cord injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Tenuigenin (TEN) is a main pharmacologically active component of Polygala tenuifolia Willd. (Polygalaceae), which has shown neuroprotective functions in Alzheimer's disease. Moreover, TEN also demonstrated an anti-oxidative impact in an in vitro model of Parkinson's disease, reducing damage and loss of dopaminergic neurons. AIM: This work focuses on the impact of TEN on locomotor recovery following spinal cord injury (SCI) and underpinning molecules involved. METHODS: A rat model of SCI was generated, and the rats were treated with TEN, oe-PTPN1 (PTP non-receptor type 1), a protein kinase B (Akt)/mammalian target of rapamycin (mTOR) antagonist LY294002, or an autophagy inhibitor 3-methyladenine (3-MA). Subsequently, locomotor function was detected. Pathological changes and neuronal activity in the spinal cord tissues were analyzed by hematoxylin and eosin staining, Nissl staining, and TUNEL assays. Protein expression of Beclin-1 and microtubule associated protein 1 light chain 3 beta (LC3B)-II/LC3B-I, PTPN1, IRS1, mTOR, and phosphorylated Akt (p-Akt) was analyzed by western blot assays. The LC3B expression was further examined by immunofluorescence staining. RESULTS: Treatment with TEN restored the locomotor function of SCI rats, reduced the cavity area and cell apoptosis, upregulated growth-associated protein 43 and neurofilament 200, and decreased the Beclin-1 and LC3B-II/LC3B-I levels in the spinal cord. TEN suppressed PTPN1 protein level, while PTPN1 suppressed IRS1 protein to reduce the p-Akt and mTOR levels. Either PTPN1 overexpression or LY294002 treatment blocked the promoting effect of TEN on SCI recovery. However, treatment with 3-MA suppressed autophagy, which consequently rescued the locomotor function and reduced neuron loss induced by PTPN1. CONCLUSION: This study demonstrates that TEN suppresses autophagy to promote function recovery in SCI rats by blocking PTPN1 and rescuing the IRS1/Akt/mTOR signaling.

Long non-coding RNA CRNDE regulates the growth and migration of prostate cancer cells by targeting microRNA-146a-5p.

The function of lncRNA CRNDE and its role in prostate cancer (PC) remains unclear. The aim of this study was to determine the expression level of lncRNA CRNDE in PC tissues and to elucidate its role in PC. The expression levels of lncRNA CRNDE were measured by quantitative reverse transcription polymerase chain reaction. The role of lncRNA CRNDE in PC cells was studied using loss-of-function assays in vitro. Cell proliferation, migration, invasion, and apoptosis were assessed via Cell Counting Kit-8, colony formation, flow cytometry, wound healing, and transwell chamber assays. A luciferase reporter assay was used to characterize the interaction between lncRNA CRNDE and miR-146a-5p. In PC tissues, the expression level of lncRNA CRNDE was upregulated. Moreover, knockdown of lncRNA CRNDE suppressed PC cell proliferation and migration and induced apoptosis in vitro. miR-146a-5p was verified as a direct target of lncRNA CRNDE. Moreover, the inhibition of miR-146a-5p partially counteracted the effects of lncRNA CRNDE on PC cell proliferation, migration, and invasion. In conclusion, lncRNA CRNDE may serve as a cancer promoter in PC by targeting miR-146a-5p. Therefore, lncRNA CRNDE could be a promising target for the clinical treatment of PC.

Bone marrow mesenchymal stem cells-derived exosomal microRNA-19b-1-5p reduces proliferation and raises apoptosis of bladder cancer cells via targeting ABL2.

BACKGROUND: Exosomes are involved in intercellular communication via specialized molecular cargo, such as microRNAs (miRNAs). However, the mechanisms underlying exosomal miR-19b-1-5p in bladder cancer remain largely unknown, thus, we aim to investigate the effect of exosomal miR-19b-1-5p on bladder cancer with the involvement of non-receptor protein tyrosine kinase Arg (ABL2). METHODS: miR-19b-1-5p and ABL2 expression were tested in bladder cancer. miR-19b-1-5p inhibition/elevation assays were conducted to determine its role in bladder cancer. Exosomes were extracted from bone marrow mesenchymal stem cells (BMSCs). Exosomes and T24 cells were co-cultured to verify their function in biological characteristics of bladder cancer cells. RESULTS: miR-19b-1-5p was down-regulated while ABL2 was upregulated in bladder cancer. Exosomal miR-19b-1-5p suppressed malignant behaviors of bladder cancer cells, and also inhibited tumor growth in vivo. Up-regulated ABL2 mitigated the effects of miR-19b-1-5p up-regulation on bladder cancer cells. CONCLUSION: BMSCs-derived exosomal miR-19b-1-5p suppresses bladder cancer growth via decreasing ABL2.

Trelagliptin Mitigates Macrophage Infiltration by Preventing the Breakdown of the Blood-Brain Barrier in the Brain of Middle Cerebral Artery Occlusion Mice.

Stroke is a significant cardiovascular disease that influences the health of human beings all over the world, especially the elderly population. It is reported that the blood-brain barrier (BBB) can be easily destroyed by stroke, which is one of the main factors responsible for macrophage infiltration and central nervous inflammation. Here, we report the protective effects of Trelagliptin against BBB injury and macrophage infiltration. Our results indicate that the infraction volume, the neurological score, and macrophage infiltration staining with CD68 were increased in middle cerebral artery occlusion (MCAO) mice but significantly reversed by treatment with Trelagliptin. Additionally, Trelagliptin reduced the permeability of the BBB by increasing the expression of the tight junction zonula occludens protein-1 (ZO-1) in the cerebral cortex. In an in vitro hypoxia model of endothelial cells, the increased migration of macrophages, enlarged permeability of endothelial monolayer, downregulation of ZO-1, and elevated expression level of CXCL1 by hypoxic conditions were all reversed by treatment with Trelagliptin in a dose-dependent manner. Our results demonstrate that Trelagliptin might mitigate macrophage infiltration by preventing the breakdown of the blood-brain barrier in the brains of MCAO mice.

Circular RNA TTC3 regulates cerebral ischemia-reperfusion injury and neural stem cells by miR-372-3p/TLR4 axis in cerebral infarction.

BACKGROUND: Stroke serves as a prevalent cerebrovascular disorder with severe cerebral ischemia/reperfusion (CIR) injury, in which neural stem cells (NSCs) play critical roles in the recovery of cerebral function. Circular RNAs (circRNAs) have been widely found to participate in stroke and NSC modulation. However, the role of circRNA TTC3 (circTTC3) in the regulation of CIR injury and NSCs remains elusive. Here, we aimed to explore the impact of circTTC3 on CIR injury and NSCs. METHODS: The middle cerebral artery occlusion/repression (MCAO/R) model was established in C57BL/6J mice. The primary astrocytes were isolated from the cerebellum from C57BL/6J mice. The primary NSCs were obtained from rat embryos. The effect of circTTC3 on CIR injury and NSCs was analyzed by TTC staining, qPCR, Western blot, LDH colorimetric kits, MTT assays, Annexin V-FITC Apoptosis Detection Kit, luciferase reporter gene assays, and others in the system. RESULTS: Significantly, the expression of circTTC3 was elevated in the MCAO/R mice and oxygen and glucose deprivation (OGD)-treated astrocytes. The depletion of circTTC3 attenuated cerebral infarction, neurological score, and brain water content. The OGD treatment induced apoptosis and the levels of lactate dehydrogenase (LDH) in the astrocytes, in which circTTC3 depletion reduced this phenotype in the system. Moreover, the depletion of circTTC3 promoted the proliferation and upregulated the nestin and ß-tubulin III expression in NSCs. Mechanically, circTTC3 was able to sponge miR-372-3p, and miR-372-3p can target Toll-like receptor 4 (TLR4) in NSCs. The miR-372-3p inhibitor or TLR4 overexpression could reverse circTTC3 depletion-mediated astrocyte OGD injury and NSC regulation. CONCLUSION: Thus, we conclude that circTTC3 regulates CIR injury and NSCs by the miR-372-3p/TLR4 axis in cerebral infarction. Our finding presents new insight into the mechanism by which circTTC3 modulates CIR injury and NSC dysfunction. CircTTC3, miR-372-3p, and TLR4 may serve as potential targets for the treatment of CIR injury during stroke.

Melatonin Plays a Protective Role by Regulating miR-26a-5p-NRSF and JAK2-STAT3 Pathway to Improve Autophagy, Inflammation and Oxidative Stress of Cerebral Ischemia-Reperfusion Injury.

BACKGROUND: Melatonin (MT) has potential protective effect on cerebral ischemia-reperfusion injury (CIRI), but its underlying regulatory mechanism has not been identified. PURPOSE: This study aimed to explore the role of miR-26a-5p-neuron-restrictive silencing factor (NRSF/REST), Janus kinase-2 (JAK2)-signal transducer and activator of transcription-3 (STAT3) pathway in the protection mechanism of MT against CIRI in vivo and in vitro. METHODS: Sprague Dawley rats were induced with ischemia-reperfusion (IR) in vivo model; PC12 cells were induced with oxygen-glucose deprivation/reperfusion (OGD/R) in vitro model; and MT intervention was conducted before the model was established. The effect of MT on autophagy factors (LC3II/LC3I, P62), inflammatory factors (TNF-α, IL-6, IL-10) and oxidative stress indexes (MDA, GSHPx, SOD) was explored, and then the above three indexes were determined by real-time quantitative PCR, ELISA, and detection kit corresponding to oxidative stress indexes. The neuroprotective effect of MT pretreatment on brain IR injury was evaluated by neurological deficit scores and TUNEL method. The levels of miR-26a-5p and NRSF were detected by real-time quantitative PCR and Western blot, and the interaction between them was evaluated by dual luciferase report. The role of JAK2-STAT3 pathway in MT protection mechanism was verified by pathway blocker (AG490) and Western blot. RESULTS: MT pretreatment can significantly reduce neurological deficit score and neuronal apoptosis, inhibit CIRI autophagy, inflammation and oxidative stress in vivo and in vitro, reduce LC3II/LC3I, TNF-α, IL-6, MDA and increase P62, IL-10, GSHPx, SOD. Further analysis identifies that downregulating miR-26a-5p or upregulating NRSF can eliminate the protective effect of MT, and NRSF is the direct target of miR-26a-5p. The protective effect of MT can also be eliminated under AG490 intervention. CONCLUSION: MT plays a protective role by regulating miR-26a-5p-NRSF and JAK2-STAT3 pathway to improve CIRI autophagy, inflammation and oxidative stress.

MiR-631/ZAP70: A novel axis in the migration and invasion of prostate cancer cells.

MicroRNAs (miRNAs) are short, endogenous non-coding RNA molecules involved in cancer initiation and progression. Using transwell migration and invasion assays, we found that miR-631 inhibited the migration and invasion of prostate cancer (PCa) cells. Bioinformatic algorithms indicated the 3'-untranslated region (3'-UTR) of zeta-associated protein 70 (ZAP70) has a putative binding site for miR-631. We found that miR-631 can bind to the 3'-UTR of ZAP70 and decrease its expression. Further studies confirmed that ZAP70 facilitates PCa cell migration and invasion. Interestingly, using gain- and loss-of function experiments, we found that ZAP70 is a major target of miR-631 and largely mediates its activity. In addition, we further discovered that miR-631 was downregulated and ZAP70 was overexpressed in PCa cell lines and PCa tissues. A concordant inverse correlation between miR-631 and ZAP70 was also found in PCa tissues. In all, our study demonstrates that miR-631 decreases PCa cell migration and invasion by dampening ZAP70 expression.