PurposeLong non-coding RNAs (lncRNAs) play important roles in the malignant behavior of cancer. HOTAIR, a well-studied lncRNA, contributes to breast cancer development, and overexpression of HOTAIR predicts a poor prognosis. However, the regulatory role of HOTAIR in the cancer stem-like cell (CSC) subpopulation remains largely unknown. Our goal was to determine the regulatory functions of HOTAIR in the processes of self-renewal capacity, tumor formation and proliferation of CSCs derived from breast cancer.MethodsWe first enriched and incubated the CSC population derived from breast cancer cell line MCF7 (CSC-MCF7) or MDA-MB-231 (MB231, CSC-MB231). Self-renewal capacity and tumor formation were assessed in vitro and in vivo to determine the stemness of CSCs. We assessed the impact on ectopically upregulated or downregulated expression of HOTAIR in CSCs by soft agar, self-renewal capacity and CCK-8 assays. The functional domain of HOTAIR was determined by truncation. RT-qPCR and semiquantitative Western blotting were performed to detect the expression levels of genes of interest. Chromatin IP (ChIP) was employed to detect the transcriptional regulatory activity of p53 on its target gene.ResultsAfter the identification of CSC properties, RT-qPCR analysis revealed that HOTAIR, but not other cancer-associated lncRNAs, is highly upregulated in both CSC-MCF7 and CSC-MB231 populations compared with MCF7 and MB231 populations. By modulating the level of HOTAIR expression, we showed that HOTAIR tightly regulates the proliferation, colony formation, migration and self-renewal capacity of CSCs. Moreover, full-length HOTAIR transcriptionally inhibits miR-34a specifically, leading to upregulation of Sox2, which is targeted by miR-34a. Ectopic introduction of miR-34a mimics reverses the effects of HOTAIR on the physiological processes of CSCs, indicating that HOTAIR affects these processes, including self-renewal capacity; these effects are dependent on the regulation of Sox2 via miR-34a. Interestingly, tight transcriptional regulation of p53 by HOTAIR was found; accordingly, p21 is indirectly regulated by HOTAIR, resulting in cell cycle entry.ConclusionThese results suggest that HOTAIR is a key regulator of proliferation, colony formation, invasion and self-renewal capacity in breast CSCs, which occurs in part through regulation of Sox2 and p53.
The tumour‑suppressor protein p53 is a key regulator of multiple cellular processes and exerts its tumour‑suppressor function by inducing apoptotic cell death. However, emerging evidence indicates that p53 is also involved in inducing ferroptosis, which is a unique iron‑dependent form of non‑apoptotic cell death triggered by the RAS‑selective lethal small molecule erastin. Previous studies have shown that erastin exposure induces increased ROS accumulation and oxidative stress. In the present study, we incubated A549 cells with erastin and detected ROS accumulation. Semi‑quantitative western blotting was performed to analyse the effect of the induced ROS on p53 activity. To determine how ROS activate p53, NAC, an ROS scavenger, and KU‑55933, an ATM kinase inhibitor, were employed to co‑incubate with erastin, followed by western blot analysis. Either p53 or SLC7A11 siRNA was introduced into A549 cells to silence the target‑gene expression, followed by ROS detection to illustrate the regulatory role of ROS‑activated p53 on its target gene SLC7A11. Annexin V‑FITC/PI staining was performed to detect the induction of apoptotic cell death by erastin exposure. To further assess the effects of erastin treatment on cellular proliferation, EdU staining and cell cycle flow cytometric analysis were performed. Erastin exposure upregulated and activated p53 and thus, transcriptionally activated its downstream target genes, including p21 and Bax, in lung cancer A549 cells dependent on erastin‑induced ROS. Subsequently, activated p53 by erastin treatment suppressed SLC7A11 and induced ROS accumulation, indicating the potential feedback loop between p53 and erastin‑induced ROS. By employing the caspase inhibitor Z‑VAD‑FMK, it was revealed that erastin‑induced p53 contributed to both ferroptotic and apoptotic cell death and inhibited cell proliferation via arresting the cell cycle at G1 phase. Collectively, these results indicated that p53 may contribute to the cytotoxic and cytostatic effects associated with establishing a feedback loop with ROS induced by erastin.
BACKGROUND HSK3486 (ciprofol) is a 2,6-disubstituted phenol derivative that acts like propofol as an agonist at the gamma-aminobutyric acid-A (GABA A ) receptor. OBJECTIVE To investigate the efficacy and safety of HSK3486 for general anaesthesia induction and maintenance. DESIGN A single-blinded, randomised, parallel-group, phase 3 trial. SETTING Involving 10 study centres, from November 24, 2020 to January 25, 2021. PATIENTS A total of 129 patients undergoing nonemergency, noncardiothoracic, and nonneurosurgical elective surgery. INTERVENTION Patients were randomly assigned at a 2:1 ratio into HSK3486 or propofol groups, to receive HSK3486 (0.4 mg kg −1 ) or propofol (2.0 mg kg −1 ) for induction before a maintenance infusion at initial rates of 0.8 and 5.0 mg kg −1 h −1 , and were adjusted to maintain a bispectral index (BIS) of 40–60 until the end of surgery. MAIN OUTCOME MEASURES Noninferiority between the drugs was evaluated as the lower limit of the 95% confidence interval (CI) for the between-group difference in the success rate of anesthetic maintenance (primary outcome) >−8%. Secondary outcomes included successful anaesthetic induction, full alertness and spontaneous breathing recovery, time until leaving the postanaesthesia care unit and changes in BIS. Safety profiles were also measured. RESULTS Of 129 enrolled patients, 128 completed the trial, with 86 in the HSK3486 group and 42 in the propofol group. The success rate for the maintenance of general anaesthesia was 100% for both groups, and noninferiority of HSK3486 was confirmed (95% CI −4.28% to 8.38%). No significant differences were found between the two groups of patients with regard to secondary outcomes (all P > 0.05). There appeared to be a comparable incidence of treatment for emergency adverse events (TEAEs) (80.2% vs. 81.0%, P = 1.000) and drug-related TEAEs (57.0% vs. 64.3%, P = 0.451) in the HSK3486 and propofol groups. CONCLUSION HSK3486 had a noninferior efficacy profile compared to propofol, exhibiting excellent tolerance. TRIAL REGISTRATION Clinicaltrials.gov, identifier: NCT04511728.
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