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Exosomal microRNA-23a-3p contributes to the progression of cholangiocarcinoma by interaction with Dynamin3
No full textCholangiocarcinoma (abbreviated as CCA) accounts for about 3% of digestive tract tumors, which is a rare disease with relatively low incidence. Herein, we firstly discovered overexpression of microRNA-23a-3p (abbreviated as miR-23a-3p) in CCA tissues, as well as cell lines via bioinformatics prediction. Next, by conducting miR-23a-3p knockdown system in HUCCT1 cells and miR-23a-3p overexpression system in RBE cells, we investigated the biological effects of miR-23a-3p. Based on our findings, inhibition of miR-23a-3p was able to prevent cancer cell proliferation via colony formation, CCK-8, as well as EdU assays. Moreover, invasion as well as migration abilities of cells was examined by transwell assay and wound healing test. Animal study further verified that knockdown miR-23a-3p slowed down tumor growth and lung metastasis. In addition, we identified cholangiocarcinoma cells transferred miR-23a-3p through exosomes by a series of assays. Functional experiments have confirmed that exosomal miR-23a-3p could benefit for cancer cell growth and metastasis, serving as a cancer promoting gene. Furthermore, we found Dynamin3 (abbreviated as DNM3) turned out to be a target of miR-23a-3p, while DNM3 was down-regulated in cholangiocarcinoma. Knockdown DNM3 accelerated cancer cell development. Collectively, our findings firstly pointed out that exosomal miR-23a-3p was conducive to the progression of cholangiocarcinoma by interaction with DNM3, which provided potential evidence for cancer treatment.</p
Additional file 1 of Epigenetic regulation of ferroptosis via ETS1/miR-23a-3p/ACSL4 axis mediates sorafenib resistance in human hepatocellular carcinoma
No full textAdditional file 1: Supplementary Figure 1. (A) Sample clustering and (B) the analysis of the scale-free index for various soft-thresholding powers (β). (C) Scatterplot show the correlations between gene module membership in the blue module and gene significance for sorafenib response. (D) Overall survival analysis of the top-10 enhanced miRNAs in the blue module. Data was retrieved from Kaplan-Meier Plotter of liver cancer with default setting. (E) MiR-23a-3p expression between HCC and NAT. Unpair t-test, P > 0.05. (F) MiR-23a-3p expression among different grades of HCC. One-way ANOVA, P > 0.05. (G) Univariable analysis of the association between survival and clinicopathologic factors. Supplementary Figure 2. (A) Body weight of mice was recorded every 3 days. The arrow indicates the start of sorafenib administration. WT: vehicle group; R1–5: sorafenib-treated group. (B) Body weight of re-injected mice (n = 5). WT: mice with parental cells; R1/3/5: mice with in vivo-generated sorafenib resistant cells. Supplementary Figure 3. (A) The IC50 value of sorafenib in MHCC97L and PLC/PRF/5 by MTT assay. (B) The expression of p-ERK was downregulated after different doses of sorafenib treatment, indicating the effective response to sorafenib (C) The inhibition of ETS1 siRNAs on ETS1 mRNA and protein expression. Three biological replicates were conducted independently in all experiments above. One-way ANOVA, *P < 0.05, **P < 0.01, ***P < 0.005, ****P < 0.0001. Supplementary Figure 4. (A) The knockout of miR-23a-3p in 23a-KO cells was determined by qRT-PCR. (B) The flow scheme illustrates orthotopic HCC mouse model establishment. (C) The body weight of mouse model. (D) The accumulation of cleaved caspase 3 and PARP was detected by immunoblotting. Total caspase 3 and PARP were determined as reference. (E) The expression of miR-23a-3p upon transfection of miR-23a-3p mimics and Anti-miR-23a. Ten nanometer of miR-23a-3p and 30 nM of Anti-miR-23a were used in the transfection experiments. (F) The downregulated phosphorylated-ERK indicated that miR-23a-3p expression did not influence sorafenib efficiency. Three biological replicates were conducted independently in all experiments above. Unpair t-test a, ****P < 0.0001, or One-way ANOVA c and e, *P < 0.05, **P < 0.01, ***P < 0.005, ****P < 0.0001. Supplementary Figure 5. (A) Predicted miR-23a-3p binding sites in the 3’UTR of CPOX mRNA according to the computational algorithms of RNA hybrid. (B) The inhibitory effect on ACSL4 mRNA and protein expression by siRNA interference. (C) The ACSL4 expression on cotreatment of Anti-miR-23a and sorafenib. p-Akt was induced by cellular ROS and showed a consistent pattern with ACSL4. (D) Cellular ROS was determined by DCFDA staining. (E) Tissues in data analysis were numbered from 1 to 88, two of HCC tissues were excluded due to the severe damage. (F) Correlation between miR-23a-3p and ACSL4. Three biological replicates were conducted independently in all experiments above. One-way ANOVA, *P < 0.05, **P < 0.01, ***P < 0.005, ****P < 0.0001. Table S1. Patient information. Table S2. Sequence of primer sets
Supplementary Material for: Exosome miR-23a-3p From Osteoblast Alleviates Spinal Cord Ischemia/Reperfusion Injury by Down-regulating KLF3-activated CCNL2 Transcription
No full textBackground: Spinal cord ischemia/reperfusion injury (SCIRI) is usually caused by spinal surgery or aortic aneurysm surgery and can eventually lead to paralysis or paraplegia and neurological dysfunction. Exosomes are considered as one of the most promising therapeutic strategies for SCIRI as they can pass the blood-spinal barrier. Previous studies have proved that exosomes secreted by osteocytes have a certain slowing effect on SCIRI.
Aim: We aimed to explore the effect of osteoblast secreted exosomes on SCIRI.
Methods: Firstly, neurons and osteoblasts were co-cultured under different conditions. GEO database was utilized to detect the expression of miR-23a-3p in osteoblast exosomes. SCIRI cells were treated with exosomes, and the detection was taken to prove whether miR-23a-3p could slow the progression of SCIRI. Downstream gene and the potential regulatory mechanism were explored through database and functional experiments.
Results: MiR-23a-3p was highly expressed in exosomes and it slowed down the process of SCIRI. Downstream mRNA KLF3 could bind to miR-23a-3p and was highly expressed in IRI. Moreover, CCNL2 was regulated by KLF3 and was highly expressed in IRI. Rescue experiments verified that miR-23a-3p suppressed the transcription of CCNL2 by targeting KLF3.
Conclusion: Exosome miR-23a-3p from osteoblast alleviates SCIRI by down-regulating KLF3-activated CCNL2 transcription
MiR-23a-3p Mediates Sorafenib Resistance in Human Hepatocellular Carcinoma: Study on The Regulatory Mechanisms and Potential Natural Suppressor from Chinese Herbal Medicine
No full text
Hepatocellular carcinoma (HCC) is the most histotye of liver cancer, which has brought great concern to global health due to the high incidence and mortality. Due to the poor diagnosis of HCC, patients are often diagnosed at late stages and have to adopt systemic therapies for the rest of their lives. Until now, although various agents have been developed since sorafenib first came out, the efficiency of these agents did not get much better improvement. Drug resistance is the main hindrance for HCC patients to benefit from these drugs. Current studies have demonstrated the mechanisms of sorafenib resistance in HCC are involved in drug transporter systems, regulated cell death (RCD), cancer stem-like features, and so on. They can be subject to epigenetic modifications, especially microRNAs (miRNAs).
In this study, the identification of deregulated miRNA, miR-23a-3p, was through weighted correlation network analysis (WGCNA) on the miRNA expression profile in a sorafenib-treated patients’ cohort and the correlation analysis on the clinical relevance. The role of miR-23a-3p in facilitating sorafenib resistance in HCC was confirmed by establishing an in vivo generated sorafenib resistant (IV-SR) HCC cell line and conducting loss-of-function (LOF) experiments in orthotopic HCC mouse models. In the mechanism studies, proteomic analysis was performed to figure out the key upstream regulator and the essential downstream signalling. It was found that the ETS Proto-Oncogene 1 (ETS1) was the transcription factor (TF) that directly activated miR-23a-3p transcription upon sorafenib treatment, and ferroptosis was the core downstream signalling suppressed by miR-23a-3p overexpression. Specifically, Acyl-CoA synthetase long chain family member 4 (ACSL4) was proved to be the target of miR-23a-3p, which is responsible for catalysing the polyunsaturated fatty acid-containing phospholipid synthesis, an important prerequisite for ferroptosis activation.
Given the clinical significance and the regulatory mechanisms of miR-23a-3p in sorafenib-treated HCC, miR-23a-3p was suggested as a target to enhance sorafenib efficacy in anti-HCC activity. Then a screening approach to look for a miR-23a-3p-targeted Chinese compound was conducted and identified that fangchinoline and HL23 could prevent sorafenib-induced miR-23a-3p transcription. HL23 showed better cytotoxicity on HCC than fangchinoline and showed a synergistic effect with sorafenib. The combination treatment of HL23 and sorafenib in the orthotopic HCC mouse model indicated that HL23 could remarkably enhance sorafenib efficacy.
In conclusion, this study demonstrated that miR-23a-3p contributed to sorafenib resistance by targeting ACSL4-mediated ferroptosis in HCC and could be a target by HL23 to enhance the anti-HCC activity of sorafenib.</p
Inhibition of lncRNA NEAT1 sensitizes medulloblastoma cells to cisplatin through modulating the miR-23a-3p-glutaminase (GLS) axis
No full textMedulloblastoma (MB) is a commonly occurring brain malignancy in adolescence. Currently, the combination of chemotherapy with subsequent irradiation is a regular therapeutic strategy. However, high dosage of chemotherapy is associated with drug resistance and side effects. The long non-coding RNA nuclear paraspeckle assembly transcript 1 (NEAT1), which is frequently overexpressed in diverse human tumors, is correlated with worse survival rate in cancer patients. Currently, the precise roles of NEAT1 in MB and chemoresistance remain unclear. Our study aimed to investigate the biological functions of NEAT1 in cisplatin-resistant medulloblastoma. We report that NEAT1 was significantly upregulated in medulloblastoma patient specimens. Silencing NEAT1 significantly suppressed MB cell proliferation and sensitized MB cells to cisplatin. In cisplatin-resistant MB cell line, DAOY Cis R, NEAT1 expression, and glutamine metabolism were remarkably upregulated in cisplatin-resistant cells. Under low glutamine supply, cisplatin-resistant cells displayed increased cisplatin sensitivity. Bioinformatical analysis and luciferase assay uncovered that NEAT1 functions as a ceRNA of miR-23a-3p to downregulate its expressions in MB cells. Moreover, miR-23a-3p was apparently downregulated in MB patient tissues and cisplatin resistant MB cells. We identified GLS (glutaminase), a glutamine metabolism enzyme, was directly targeted by miR-23a-3p in MB cells. Rescue experiments demonstrated restoration of miR-23a-3p in NEAT1-overexpressing DAOY cisplatin resistant cells successfully overcame the NEAT1-promoted cisplatin resistance by targeting GLS. In general, our results revealed new molecular mechanisms for the lncRNA-NEAT1-mediated cisplatin sensitivity of MB.</p
Banking Affiliate Regulation Under Section 23A of the Federal Reserve Act
Get PDFBefore committing a bank\u27s financial resources to an affiliate, bankers must be aware of the scope of the term affiliate under Section 23A of the Federal Reserve Act. Section 23A places restrictions on the financial dealings between banks and their affiliate companies. The author analyzes Section 23A and the relevant regulatory and compliance issues that have recently surfaced. The author concludes that in the event of Glass-Steagall repeal, interaffiliate regulation of the financial dealings between banks and securities affiliates, as accomplished by Section 23A, would be a viable method of permitting the merger of investment and commercial banking
Validation of differential expression of miR-151-3p and miR-23a-3p in the dentate gyrus middle molecular layer 5 h after tetanisation.
No full textUpregulation of miR-151-3p and miR-23a-3p was confirmed by single-plex RT-qPCR (dual criteria: one-tailed Student's t-test p < 0.05; fold change ± 0.15). Expression values: individual and mean fold changes. * p < 0.05; n = 7. Dotted lines indicate cut-off for fold change criterion (± 15%). All data were normalised to miR-301b.</p
ROC curves of the BrS prediction model using miR-23a-3p, miR-423a-3p, and miR-223–3P.
No full textThe miR-23a-3p, miR-423a-3p, and miR-223–3P remained as independent predictors for BrS patients. The ROC curve using these three miRNAs showed good discrimination of the BrS patients from the controls with an area under the curve (AUC) of 0.871 and a sensitivity and specificity of 84.3% and 82.4%, respectively (Fig 3, left). Internal validation was performed by the leave-one-out cross-validation technique. The AUC based on cross-validation was 0.834 with a sensitivity and specificity of 83.5% and 81.1%, respectively.</p
IRF1 is directly repressed by miR-23a.
No full text<p>(A) As predicted by the TargrtScan and PicTar database, the IRF1 3′UTR contained a miR-23a binding site. The mutated IRF1 3′UTR containing several mutated nucleotides within the miR-23a binding site is shown. (B) The direct interaction of miR-23a and IRF1 3′UTR was confirmed by using a fluorescent reporter assay. MGC803 cells were transfected with the EGFP-IRF1 3′UTR reporter gene together with pcDNA3, pri-miR-23a, ASO-NC or ASO-23a. The cells were lysed 72 h after transfection, and the EGFP intensity was measured by spectrophotometry (n = 3, * p<0.05). (C) MGC803 cells were transfected with the EGFP vector, the IRF1 3′UTR reporter or the mutant EGFP-3′UTR reporter in addition to pcDNA3, pcDNA3/pri-miR-23a, ASO-NC or ASO-23a. The fluorescence intensity was detected through the method described previously (n = 3, * p<0.05). (D) MGC803 and BGC823 cells were transfected with pcDNA3, pcDNA3/pri-miR-23a, ASO-NC or ASO-23a. RNA was extracted from the transfected cells, and the expression of IRF1 mRNA was measured by real-time PCR (n = 3, * p<0.05). (E) We determined the protein expression level of IRF1 in MGC803 and BGC823 cells by western blot. The numerals above the western blot image show the ratios of the densitometry of IRF1 and GAPDH when compared with the control group.</p
Calcineurin-NFAT signaling regulates atrogin-1 and MuRF1 via microRNA-23a (miR-23a) during muscle atrophy
No full textMuscle atrophy is prevalent in chronic kidney disease (CKD) patients. MicroRNAs play a critical role in biological processes including muscle atrophy. MicroRNA-23a (miR-23a) negatively regulates the expression of two atrophy-related ubiquitin ligases, atrogin-1 and MuRF1; it is reduced in muscle during atrophy. Although miR-23a expression was recently shown to be positively regulated by NFATc3, the underlying mechanism of miR-23a suppression during atrophy remains unknown. We previously reported that the activity of calcineurin (Cn), the calcium-activated phosphatase that regulates NFATc proteins, is decreased when insulin signaling is decreased. Since CKD causes muscle atrophy, and glucocorticoids are required for the response, we investigated how dexamethasone (DEX) affects Cn activity, NFATc3 signaling, and miR-23a expression. C2C12 or L6 myotubes were treated with 100 uM DEX to induce atrophy. Within 1 h, Cn activity was reduced and less NFATc3 was located in the nucleus. Further, miR-23a was also decreased within 30 minutes. After 48 h, expression of the NFATC3 target gene, MCIP1.4, and miR-23a were decreased. Expression of atrogin-1 and MuRF1 were also increased 48 h after DEX. Collectively, these findings indicate the Cn-NFAT signaling pathway may play an important role in the regulation of atrogin-1 and MuRF1 by suppressing miR23a during CKD and glucocorticoid-related muscle atrophy. Support: NIH DK007656; AHA GRNT766002
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