Antiproliferative effects of levan polysaccharide against colorectal cancer cells mediated through oxidative stress-stimulated HOTAIR/Akt signaling pathway: In vitro

The overexpression of lncRNA HOTAIR can result in cancer progression through upregulation of the Akt signaling pathway. We aimed to evaluate the anticancer effects of levan polysaccharide from Erwinia herbicola on colorectal cancer cells (HT-29) and explore the role of the ROS-mediated HOTAIR/Akt signaling pathway. The HT-29 cancer cells were treated with levan either alone or along with N-acetyl-L-cysteine (NAC), as a potential antioxidant for 24 hrs and different assays including MTT, LDH, ROS, apoptosis, SOD activity, CAT activity, caspase-3 activity, qRT-PCR, and western blot analysis were performed. It was shown that levan treatment induces apparent reduction in cell viability, SOD and CAT activity, HOTAIR expression, the ratio of pAkt protein level and a significant increase in the ROS level, apoptosis induction, and caspase-3 activity, which were effectively suppressed by NAC co-treatment. These data indicated the effective antiproliferative effect of levan on colorectal cancer cells, in which downregulation of ROS-mediated HOTAIR/Akt plays an important role.     

Brassinin inhibits proliferation and induces cell cycle arrest and apoptosis in nasopharyngeal cancer C666-1 cells

BackgroundNasopharyngeal cancer is a tumor that occurs in the mucous epithelium of the nasopharynx. Due to its rapid growth and early metastatic nature, the successful treatment of nasopharyngeal cancer is highly challenging.ObjectiveHere, we intended to assess the in vitro anticancer property of brassinin against the nasopharyngeal cancer C666-1 cells.MethodologyThe in vitro free radical scavenging property of the brassinin was assessed by various free radical scavenging activities such as FRAP, DPPH, chemiluminescence (CL), and ORAC assays. The cytotoxic level of the brassinin (1–50 µM) against the nasopharyngeal cancer C666-1 cells and normal Vero cells were assessed by the MTT cytotoxicity assay. The levels of TBARS, GSH, and the SOD activity was assessed using kits. The level of ROS generation, MMP, and apoptosis were investigated by the respective fluorescent staining techniques. The flow cytometry analysis was done to scrutinize the cell cycle arrest. The Bax/Bcl-2 level and caspase activities were examined using respective kits.ResultsThe brassinin treatment effectively scavenged the free radicals, which are assessed by the FRAP, DPPH, chemiluminescence (CL), and ORAC assays. The proliferation of brassinin treated C666-1 cells were decreased remarkably, while the same concentration of brassinin did not disturbed the Vero cell viability. The 30 µM of brassinin effectively increased the ROS production, depleted the MMP, and stimulated the apoptosis in the C666-1 cells. The brassinin increased the TBARS and depleted the GSH and SOD in the C666-1 cells. The flow cytometry analysis revealed that the brassinin administration improved the G0/G1 ratio and decreased the proportion of cells with ‘S’ and ‘G2/M’ phase. The Bax, caspase-3 and ?9 were elevated and Bcl-2 level was decreased in the brassinin administered C666-1 cells.ConclusionOur findings discovered that the brassinin has the capacity to prevent the proliferation and stimulate the apoptotic cell death C666‐1 cells via blocking cell cycle and increasing oxidative stress and apoptotic markers. Hence, it can be a talented therapeutic agent to treat the nasopharyngeal cancer in the future.     

CXCR4 inhibition suppresses Cd-induced renal oxidative stress,apoptosis, and fibrosis by inhibiting the TGF-β1/Smad pathway

This study aimed to investigate the role and mechanism of CXC chemokine receptor 4 (CXCR4) in cadmium (Cd)-induced renal injury. CXCR4 and TGF-β1/Smad pathway protein levels were detected by western blotting. Indicators related to renal function and oxidative stress factors were assessed and reactive oxygen species (ROS) level was evaluated by staining. TUNEL was used to measure apoptosis rate. PAS and Masson's trichrome staining were used to detect the level of renal fibrosis. The expression of Bcl-2, Bax, Cleaved-caspase 3, fibronectin, and collagen I proteins were detected by western blotting, immunohistochemistry, or immunofluorescence. The expression of CXCR4 was increased in a Cd-induced chronic renal injury model in rats. Si-CXCR4 decreased levels of TGF-β1, TGF-βR1, p-Smad2/Smad2, p-Smad3/Smad3, the renal weight index, urine protein, blood urea nitrogen, blood creatinine, and levels of MDA but raised the levels of SOD and GSH-Px. In addition, si-CXCR4 inhibited apoptosis in Cd-treated rats. CXCR4 inhibition alleviated fibrosis levels in Cd-treated rats. In Cd-treated cells, TGF-β attenuated the suppressive effect of CXCR4 inhibition on the TGF-β1/Smad pathway. TGF-β intervention increased MDA and ROS, and downregulated SOD and GSH-Px. TGF-β attenuated the inhibitory effect of CXCR4 on apoptosis and fibrosis. CXCR4 inhibition decreased levels of Cd-induced renal oxidative stress, apoptosis, and fibrosis by inhibiting the TGF-β1/Smad pathway.     

Anticancer Effects of Propionic Acid Inducing Cell Death in Cervical Cancer Cells

Recent studies found that short-chain fatty acids (SCFAs), which are produced through bacterial fermentation in the gastrointestinal tract, have oncoprotective effects against cervical cancer. The most common SCFAs that are well known include acetic acid, butyric acid, and propionic acid, among which propionic acid (PA) has been reported to induce apoptosis in HeLa cells. However, the mechanism in which SCFAs suppress HeLa cell viability remain poorly understood. Our study aims to provide a more detailed look into the mechanism of PA in HeLa cells. Flow cytometry analysis revealed that PA induces reactive oxygen species (ROS), leading to the dysfunction of the mitochondrial membrane. Moreover, PA inhibits NF-κB and AKT/mTOR signaling pathways and induces LC3B protein levels, resulting in autophagy. PA also increased the sub-G1 cell population that is characteristic of cell death. Therefore, the results of this study propose that PA inhibits HeLa cell viability through a mechanism mediated by the induction of autophagy. The study also suggests a new approach for cervical cancer therapeutics.     

Development of LC–MS method for analysis of paclitaxel-inhibited growth and enhanced therapeutic response in human glioblastoma cells

Glioma stem cells are considered responsible for drug resistance and glioma relapse resulting in poor prognosis in glioblastoma multiforme. SU3 glioma cell is a highly invasive glioma stem cell line from the patients with glioblastoma multifrome. It is of great significance to study the efficacy and molecular mechanism for anticancer drug effects on SU3 glioma cells. In this work, we develop a liquid chromatography–mass spectrometry(LC–MS) method for direct analysis of the role of drugs(paclitaxel)on SU3 glioma cells at the molecular level. We use the specific fluorescence dyes to evaluate cell viability,the levels of ROS and GSH when the cells were treated with drugs. In addition, the LC–MS platform was successfully employed to detect the amount of 6-O-methylguanine, demonstrating that it is effective to induce cell apoptosis and enhance the cytotoxic response of SU3 glioma cells. The analytical linear equals are Y = 9.49 ? 105 X + 2.42 ? 104 for 6-O-methylguanine(R2= 0.9998) and Y = 4.72 ? 104 X + 2.21 ? 103(R2= 0.9996) for 7-methylguanine. Thus, the combination of cell-specific fluorescence dyes and LC–MS method enables us to reveal the molecular mechanism of paclitaxel-inhibited growth and enhanced therapeutic response in the chemotherapy for glioma multiforme.     

Xanthatin mediates G2/M cell cycle arrest,autophagy and apoptosis via ROS/XIAP signaling in human colon cancer cells

Abstract

Xanthatin is a natural plant bicyclic sesquiterpene lactone extracted from Xanthium plants (Asteraceae). In the present study, we demonstrated for the first time that Xanthatin inhibited cell proliferation and mediated G₂/M phase arrest in human colon cancer cells. Xanthatin also activated caspase and mediated apoptosis in these cells. Concomitantly, Xanthatin triggered cell autophagic response. We found down-regulation of X-linked inhibitor of apoptosis protein (XIAP) contribute to the induction of apoptosis and autophagy. Moreover, reactive oxygen species (ROS) production was triggered upon exposure to Xanthatin in colon cancer cells. ROS inhibitor N-acetylcysteine (NAC) significantly reversed Xanthatin-mediated XIAP down-regulation, G₂/M phase arrest, apoptosis and autophagosome accumulation. In summary, our findings demonstrated that Xanthatin caused G₂/M phase arrest and mediated apoptosis and autophagy through ROS/XIAP in human colon cancer cells. We provided molecular bases for developing Xanthatin as a promising antitumor candidate for colon cancer therapy. Abbreviations ROS reactive oxygen species

DMSO dimethyl sulfoxide

5-FU 5-Fluorouracil

3-MA 3-Methyladenine

DCFH-DA 2’7’-dichlorfluorescein-diacetate

NAC N-acetylcysteine

XIAP X-linked inhibitor of apoptosis protein

     

Chemical fingerprinting and the biological properties of extracts from Fomitopsis pinicola

This study investigated the in vitro antioxidant and anticancer properties of the Fomitopsis pinicola extract (EMFP). The antioxidant activity of EFMP was analysed via free radical scavenging (DPPH, ABTS and Hydroxyl radicals) assay and a protein oxidation assay. EFMP effectively scavenged free radicals and exhibited remarkable protection against protein oxidation. The proliferation of EMFP-treated HepG2 cells was remarkably decreased. EMFP effectively increased the reactive oxygen species (ROS) production, depleted the mitochondrial membrane potential (MMP) and promoted the apoptosis of HepG2 cells. In addition, EMFP increased the malondialdehyde (MDA) content and reduced the activities of SOD, CAT and GPx in HepG2 cells. Using UPLC-Triple-TOF-MS, 2 phenolic compounds and 14 triterpenes were identified. These compounds may be the primary contributors to the antioxidant and anticancer capacities of EMFP. Together, these findings highlight the possibility of exploiting EMFP for its desired pharmaceutical ingredients.     

Apoptosis induced by NaYF_4:Eu~(3+) nanoparticles in liver cells via mitochondria damage dependent pathway

As lanthanide-doped sodium yttrium flouride(NaYF_4)nanoparticles have great potential inbiomedical applications,their biosafety is important and has attracted significant attention.In the present work,three different sized NaYF_4:Eu~(3+)nanoparticles have been prepared.Liver BRL 3 A cell was used as a cell model to evaluate their biological effects.Cell viability and apoptosis assays were used to confirm the cytotoxicity induced by NaYF_4:Eu~(3+)NPs.Apart from the elevated malondialdehyde(MDA),the decrease of superoxide dismutase(SOD),glutathione peroxidase(GSH-PX)and catalase(CAT)activity indicated reactive oxygen species(ROS)generation,which were associated with oxidative damage.The decrease of mitochondrial membrane potential(MMP)value demonstrated the occurrence of mitochondria damage.Then,release of cytochrome c from mitochondria and activation of caspase-3 confirmed that NaYF_4:Eu~(3+)NPs induced apoptosis was mitochondria damage-dependent.     

Exploring the anticancer effects of tin oxide nanoparticles synthesized by pulsed laser ablation technique against breast cancer cell line through downregulation of PI3K/AKT/mTOR signaling pathway

Tin oxide nanoparticles (SnO₂ NPs) demonstrate potential anti-cancer functions. However, the anti-cancer mechanisms of SnO₂ NPs have not been explored in detail. In the present study, we synthesized SnO₂ NPs through laser ablation technique and examined their anticancer mechanisms and the probable involvement of the PI3K/AKT mediated pathways in human breast cancer cells (MCF-7) in vitro. The synthesized SnO₂ NPs were characterized by transmission electron microcopy (TEM), dynamic light scattering (DLS), and Fourier-transform infrared spectroscopy (FTIR) techniques. Afterwards, the breast cancer cells were incubated with increasing concentrations of SnO₂ NPs, and inhibition of cell proliferation was assessed by the viability assay. Furthermore, the quantification of reactive oxygen species (ROS) and apoptosis were examined by flow cytometry followed by superoxide dismutase (SOD) and catalase (CAT) activity as well as mitochondrial membrane potential assays. The expression levels of phosphoinositide 3-kinase (PI3K), protein kinase B (AKT), mechanistic target of rapamycin (mTOR), B-cell lymphoma 2 (Bcl-2), and Bax were also assessed by western blot and quantitative real time PCR (qRT-PCR). It was shown that SnO₂ NPs, 30 nm, with potential colloidal stability selectively prevented the proliferation of MCF-7 in comparison with MCF-10A cells and triggered ROS production, apoptosis, deactivation of SOD and CAT activity, and mitigation of mitochondrial membrane potential. Moreover, SnO₂ NPs stimulated mitochondrial-mediated apoptosis pathway by overexpression of Bax/Bcl-2 and downregulation of p-PI3K/p-AKT/p-mTOR signaling pathway. This data elucidates the possible mechanisms by which SnO₂ NPs may stimulate their anticancer effects.     

Effect of sound wave stress on antioxidant enzyme activities and lipid peroxidation of Dendrobium candidum

The effect of sound wave stress on important medicinal plant, Dendrobium candidum Wall. ex Lindl, was investigated, including the responses on malondialdehyde (MDA) content, the activities change of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD) and ascorbate peroxidase (APX). Results were found that the activities of SOD, CAT, POD and APX enhanced totally in different organs of D. candidum, as leaves, stems and roots, in response to the stress. Furthermore there happened similar shift of antioxidant enzymes activities, which increased in the initial stimulation and decreased afterwards. Data showed SOD, CAT, POD and APX activities ascended to max at day 9, 6, 9 and 12 in leaves, at day 9, 6, 12 and 9 in stems, and at day 12, 6, 9 and 9 in roots, respectively. As a lipid peroxidation parameter, MDA content in different organs increased in the beginning, dropped afterward, and increased again in the late. Anyway the total trend was the rise of MDA level compared to the control. It was interesting that the MDA content appeared the lowest levels almost when the antioxidant enzymes activities were up to the highest. Our results demonstrated the different organs of D. candidum might produce accumulation of active oxygen species (AOS) under initial treatment of sound wave stress. Later AOS might start to reduce due to the enhancement of antioxidant enzymes activities treated by the stress. The data revealed that the antioxidant metabolism was to be important in determining the ability of plants to survive in sound stress, and the up regulation of these enzymes activities would help to reduce the build up of AOS, which could protect plant cells from oxidative damage. Moreover, different cell compartments might activate different defensive system to reduce excessive amount of AOS. Finally the mechanism of this action was also discussed simply.     

Chitosan-sodium alginate-polyethylene glycol-crocin nanocomposite treatment inhibits esophageal cancer KYSE-150 cell growth via inducing apoptotic cell death

BackgroundEsophageal cancer is a sixth most cause of cancer-associated mortalities worldwide with increased global prevalence in each year. It has a poor prognosis with 5-year survival rate are less than 10%.ObjectiveThe present study was focused to fabricate the chitosan-sodium alginate-polyethylene glycol-crocin nanocomposites (CSP-Cr-NCs) and evaluate its in vitro anticancer potential against the esophageal cancer KYSE-150 cells.MethodologyThe fabricated CSP-Cr-NCs were characterized using different techniques such as UV–visible spectroscopy, photoluminescence assay, DLS analysis, XRD, SEM and EDX analyses. The antimicrobial study was conducted by well diffusion technique against the S. pneumoniae, K. pneumoniae, E. coli, S. aureus, and C. albicans. The viability of CSP-Cr-NCs treated KYSE-150 and Het-1A cells were assessed by the MTT assay. The ROS production, MMP level, and apoptotic cell death in the CSP-Cr-NCs administered cells were assessed by using different fluorescent staining techniques. The cell migration of CSP-Cr-NCs treated KYSE-150 cells were assessed by wound scratch assay. The levels of TBARS, GSH, and SOD activity in the CSP-Cr-NCs treated KYSE-150 cells were assessed by kits.ResultsThe outcomes from the different characterization analyses witnessed the formation of CSP-Cr-NCs with the 100–210 nm size, tetragonal and agglomerated morphological appearances. The CSP-Cr-NCs effectively repressed the microbial growth. The CSP-Cr-NCs treated KYSE-150 cells were demonstrated the decreased cell viability and IC50 was found at 2.5 µg, furthermore it did not affected the normal Het-1A cells. The formulated CSP-Cr-NCs treatment at 2.5 and 5 µg improved the ROS production, and decreased the MMP status in the KYSE-150 cells. The elevated incidences of apoptotic cells death was found in the CSP-Cr-NCs treated KYSE-150 cells. The CSP-Cr-NCs also inhibited the migration of KYSE-150 cells. The increased TBARS content and decreased GSH and SOD activity was also found in the CSP-Cr-NCs treated KYSE-150 cells.ConclusionOur findings proved that the formulated CSP-Cr-NCs treatment effectively inhibited the esophageal cancer KYSE-150 cell growth via increasing ROS production and apoptotic cell death. Therefore, it could be a promising anticancer candidate in the future for the esophageal cancer treatment.     

Continuous hypoxia attenuates paraquat-induced cytotoxicity in the human A549 lung carcinoma cell line

Paraquat (1,1'-dimethyl-4,4'-bipyridinium dichloride; PQ), an effective and widely used herbicide, was commercially introduced in 1962. It is reduced by the electron donor NADPH, and then reduced PQ transfers the electrons to molecular oxygen, resulting in the production of reactive oxygen species (ROS), which are related to cellular toxicity. However, the influence of continuous hypoxia on PQ-induced ROS production has not fully been investigated. We evaluated in vitro the protective effect of continuous hypoxia on PQ-induced cytotoxicity in the human carcinogenic alveolar basal epithelial cell line (A549 cells) by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay and live and dead assay, and by measuring lactate dehydrogenase (LDH) release. To elucidate the mechanism underlying this effect, we monitored the immunofluorescence of intracellular ROS and measured malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione peroxidase (GPx) activities. Continuous hypoxia protected the A549 cells from PQ-induced cytotoxicity. Continuous hypoxia for a period of 24 h significantly reduced intracellular ROS, decreased MDA concentration in the supernatant, and normalized SOD and GPx activities. Continuous hypoxia attenuated PQ-induced cell toxicity in A549 cells. This protective effect might be attributable to the suppression of PQ-induced ROS generation.     

The potential antioxidant ability of hydroxytyrosol on Caenorhabditis elegans against oxidative damage via the insulin signaling pathway

Hydroxytyrosol (HY) deriving from olive leaves is a phenolic component which has been proven to possess a strong antioxidant ability. However, the underling mechanism is still unclear. To evaluate the antioxidant ability of HY comprehensively, assays in vitro and in vivo (Caenorhabditis elegans (C. elegans) was used as a model organism) were conducted. The results showed HY could scavenge 2,2-Diphenyl-1-picryl-hydrazyl (DPPH) radicals with a strong total reducing power. Pretreated with HY for 48 h, the cell viability of Chinese hamster ovary (CHO) cells was enhanced under oxidative stress by reducing the level of reactive oxygen species (ROS) and malondialdehyde (MDA). A suitable concentration of HY showed no side effects on the development, fertility, and movement of C. elegans. With the treatment of HY, the survival was enhanced by 15.79% under thermal stress. The ROS and MDA contents were also reduced, which might be associated with the increasing abilities of antioxidant enzymes such as superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) and the heat shock protein HSP-16.2. Nuclear localization of DAF-16 was greatly increased after treated with HY. More outcomes demonstrated HY exhibited an excellent antioxidant capacity via the insulin signaling pathway by upregulating daf-16 and sod-3 and downregulating the genes of age-1 and daf-2.     

Apoptosis induced by NaYF<Subscript>4</Subscript>:Eu<Superscript>3+</Superscript> nanoparticles in liver cells via mitochondria damage dependent pathway

As lanthanide-doped sodium yttrium flouride (NaYF₄) nanoparticles have great potential in biomedical applications, their biosafety is important and has attracted significant attention. In the present work, three different sized NaYF₄:Eu³⁺ nanoparticles have been prepared. Liver BRL 3A cell was used as a cell model to evaluate their biological effects. Cell viability and apoptosis assays were used to confirm the cytotoxicity induced by NaYF₄:Eu³⁺ NPs. Apart from the elevated malondialdehyde (MDA), the decrease of superoxide dismutase (SOD), glutathione peroxidase (GSH-PX) and catalase (CAT) activity indicated reactive oxygen species (ROS) generation, which were associated with oxidative damage. The decrease of mitochondrial membrane potential (MMP) value demonstrated the occurrence of mitochondria damage. Then, release of cytochrome c from mitochondria and activation of caspase-3 confirmed that NaYF₄:Eu³⁺ NPs induced apoptosis was mitochondria damage-dependent.     

Effect of berberine on the viability of adipose tissue-derived mesenchymal stem cells in nutrients deficient condition

This study was designed to examine whether berberine protects rat adipose tissue-derived stem cells (ASCs) against glucose and serum deprivation (GSD)-induced cell death. ASCs were cultured for 24 h in GSD condition in the presence of berberine and then cell viability, apoptosis and generation of reactive oxygen species (ROS) were evaluated. The GSD condition significantly decreased ASCs viability and increased ROS generation and apoptosis. Incubation with 0.75–3 μM berberine partially increased cell viability and decreased ROS generation and apoptosis in GSD condition. In conclusion, berberine partially protects ASCs in nutrients deficient condition and may help ASCs to preserve their survival during cell therapy of ischemia.     

Ruthenacarborane and Quinoline: A Promising Combination for the Treatment of Brain Tumors

Gliomas and glioblastomas are very aggressive forms of brain tumors, prone to the development of a multitude of resistance mechanisms to therapeutic treatments, including cytoprotective autophagy. In this work, we investigated the role and mechanism of action of the combination of a ruthenacarborane derivative with 8-hydroxyquinoline (8-HQ), linked via an ester bond (complex 2), in rat astrocytoma C6 and human glioma U251 cells, in comparison with the two compounds alone, i.e., the free carboxylic acid (complex 1) and 8-HQ, and their non-covalent combination ([1 + 8-HQ], in 1:1 molar ratio). We found that only complex 2 was able to significantly affect cellular viability in glioma U251 cells (IC₅₀ 11.4 μM) via inhibition of the autophagic machinery, most likely acting at the early stages of the autophagic cascade. Contrary to 8-HQ alone, complex 2 was also able to impair cellular viability under conditions of glucose deprivation. We thus suggest different mechanisms of action of ruthenacarborane complex 2 than purely organic quinoline-based drugs, making complex 2 a very attractive candidate for evading the known resistances of brain tumors to chloroquine-based therapies.     

Neuroprotective activity of lavender oil on transient focal cerebral ischemia in mice

The air-dried aerial parts of Lavandula angustifolia Mill, a traditional Uygur herbal drug, is used as resuscitation-inducing therapy to treat neurodisfunctions, such as stroke. This study was designed to assess the neuroprotective effects of lavender oil against ischemia/reperfusion (IR) injury in mice. Focal cerebral ischemia was induced by the intraluminal occlusion method with a nylon string. The neurodysfuntion was evaluated by neurological deficit and the infarct area was showed by 2,3,5-triphenyltetrazolium chloride (TTC) staining. The histopathological changes were observed by hematoxylin and eosin staining. The levels of mitochondria-generated reactive oxygen species (ROS), malondialdehyde (MDA) and carbonyl, the ratio of reduced glutathione (GSH)/glutathione disulfide (GSSG), the activities of superoxide dismutase (SOD), catalase (CAT) and glutathion peroxidase (GSH-Px) in brain tissue were measured to estimate the oxidative stress state. Neurological deficit, infarct size, histopathology changes and oxidative stress markers were evaluated after 22 h of reperfusion. In comparison with the model group, treatment with lavender oil significantly decreased neurological deficit scores, infarct size, the levels of MDA, carbonyl and ROS, and attenuated neuronal damage, upregulated SOD, CAT, GSH-Px activities and GSH/GSSG ratio. These results suggested that the neuroprotective effects of lavender oil against cerebral ischemia/reperfusion injury may be attributed to its antioxidant effects.     

Wnt/GSK3β/β-catenin signaling pathway regulates calycosin-mediated anticancer effects in glioblastoma multiforme cells

Calycosin, an O-methylated isoflavone, has been widely reported to induce anticancer activity in different cancer cells in vitro. Nonetheless, the associated mechanism of calycosin in glioblastoma multiforme cells (U87) still remains unknown. To explore the anticancer effects, the apoptotic mechanism of calycosin via Wnt/GSK3β/β-catenin signaling was explored in U87 cells. Different assays including: cytotoxicity, free radical determination, SOD and CAT activity, GSH content, qPCR, mitochondrial membrane potential, caspase activity, and western blotting assays were performed. It was shown that calycosin mitigated cell viability in U87 cells, whereas it showed no apparent effect on BV2 microglial cells. Calycosin triggered apoptosis via upregulating the mitochondria-associated caspase pathway in U87 cells. Calycosin induced the reduction of the mitochondrial membrane potential, overexpression of Bax, downexpression of Bcl-2, and activation of caspase-9 and caspase-3. Calycosin-stimulated apoptosis was associated with the upregulation of free radical scavenging through the modulation of antioxidant enzymes, such as SOD and CAT as well as the level of GSH. The apoptotic activity of calycosin was mediated by suppression of pGSK-3βser9, β-catenin, and c-Myc at protein level. The present study suggested that calycosin triggers U87 cell death through an antioxidant effect mediated by Wnt/GSK3β/β-catenin signaling pathway.     

Plasma extracellular vesicle delivery of miR-210-3p by targeting ATG7 to promote sepsis-induced acute lung injury by regulating autophagy and activating inflammation

Extracellular vesicles (EVs) can be used for intercellular communication by facilitating the transfer of miRNAs from one cell to a recipient cell. MicroRNA (miR)-210-3p is released into the blood during sepsis, inducing cytokine production and promoting leukocyte migration. Thus, the current study aimed to elucidate the role of plasma EVs in delivering miR-210-3p in sepsis-induced acute lung injury (ALI). Plasma EVs were isolated from septic patients, after which the expression of various inflammatory factors was measured using enzyme-linked immunosorbent assay. Cell viability and apoptosis were measured via cell counting kit-8 and flow cytometry. Transendothelial resistance and fluorescein isothiocyanate fluorescence were used to measure endothelial cell permeability. Matrigel was used to examine the tubulogenesis of endothelial cells. The targeting relationship between miR-210-3p and ATG7 was assessed by dual-luciferase reporter assays. The expression of ATG7 and autophagy-related genes was determined to examine autophagic activation. A sepsis mouse model was established by cecal ligation and puncture (CLP)-induced surgery. The level of miR-210-3p was highly enriched in septic EVs. MiR-210-3p enhanced THP-1 macrophage inflammation, BEAS-2B cell apoptosis, and HLMVEC permeability while inhibiting angiogenesis and cellular activity. MiR-210-3p overexpression reduced ATG7 and LC3II/LC3I expression and increased P62 expression. Improvements in vascular density and autophagosome formation, increased ATG7 expression, and changes in the ratio of LC3II/LC3I were detected, as well as reduced P62 expression, in adenovirus-anti-miR-210-3p treated mice after CLP injury. Taken together, the key findings of the current study demonstrate that plasma EVs carrying miR-210-3p target ATG7 to regulate autophagy and inflammatory activation in a sepsis-induced ALI model.Subject terms: Infection, Immunological disorders     

Cephalotaxine Inhibits the Survival of Leukemia Cells by Activating Mitochondrial Apoptosis Pathway and Inhibiting Autophagy Flow

Cephalotaxine (CET) is a natural alkaloid with potent antileukemia effects. However, its underlying molecular mechanism has not been well understood. In this study, we verified that CET significantly inhibited the viability of various leukemia cells, including HL-60, NB4, Jurkat, K562, Raji and MOLT-4. RNA-sequencing and bioinformatics analysis revealed that CET causes mitochondrial function change. Mechanism research indicated that CET activated the mitochondrial apoptosis pathway by reducing the mitochondrial membrane potential, downregulating anti-apoptotic Bcl-2 protein and upregulating pro-apoptotic Bak protein. In addition, the autophagy signaling pathway was highly enriched by RNA-seq analysis. Then, we found that CET blocked the fluorescence colocation of MitoTracker Green and LysoTracker Red and upregulated the level of LC3-II and p62, which indicated that autophagy flow was impaired. Further results demonstrated that CET could impair lysosomal acidification and block autophagy flow. Finally, inhibiting autophagy flow could aggravate apoptosis of HL-60 cells induced by CET. In summary, this study demonstrated that CET exerted antileukemia effects through activation of the mitochondria-dependent pathway and by impairing autophagy flow. Our research provides new insights into the molecular mechanisms of CET in the treatment of leukemia.