Search

Objective: Long noncoding RNAs (lncRNAs) and microRNAs (miRNAs) are widely expressed in the brain and are associated with the development of neurological and neurodegenerative diseases. However, their roles and molecular mechanisms in major depressive disorder (MDD) remain largely unknown. This study aimed to identify lncRNAs and miRNAs involved in the development of MDD and elucidate their molecular mechanisms., Methods: Transcriptome and bioinformatic analyses were performed to identify miRNAs and lncRNAs related to MDD. C57 mice were subjected to chronic unpredictable mild stress (CUMS) to establish a depression model. Lentiviruses containing either lncRNA NPTN-IT1-201 or miR-142-5p were microinjected into the hippocampal region of these mice. Behavioral tests including the sucrose preference test (SPT), tail suspension test (TST), and forced swim test (FST) were conducted to evaluate depressive-like behaviors., Results: The results revealed that overexpression of lncRNA NPTN-IT1-201 or inhibition of miR-142-5p significantly ameliorated depressive-like behaviors in CUMS-treated mice. Dual-luciferase reporter assays confirmed interactions between miR-142-5p with both brain-derived neurotrophic factor (BDNF) and NPTN-IT1-201. ELISA analysis revealed significant alterations in relevant biomarkers in the blood samples of MDD patients compared to healthy controls. Histological analyses, including HE and Nissl staining, showed marked structural changes in brain tissues following CUMS treatment, which were partially reversed by lncRNA NPTN-IT1-201 overexpression or miR-142-5p inhibition. Immunofluorescence imaging demonstrated significant differences in the levels of BAX, Bcl2, p65, Iba1 among different treatment groups. TUNEL assays confirmed reduced apoptosis in brain tissues following these interventions. Western blotting showed the significant differences in BDNF, BAX, and Bcl2 protein levels among different treatment groups., Conclusion: NPTN-IT1-201 regulates inflammation and apoptosis in MDD by targeting BDNF via miR-142-5p, making it a potential therapeutic target for MDD., (© 2024. Huazhong University of Science and Technology.)

Follicular fluid meiosis-activating sterol (FF-MAS) is a small molecule compound found in FF, named for its ability to induce oocyte resumption of meiosis. Granulosa cells (GCs) within the follicle are typically located in a hypoxic environment under physiologic conditions due to limited vascular distribution. Previous research suggests that hypoxia-induced cell cycle arrest and apoptosis in GCs may be crucial triggering factors in porcine follicular atresia. However, the impact of FF-MAS on GCs within follicles has not been explored so far. In this study, we uncovered a novel role of FF-MAS in facilitating GC survival under hypoxic conditions by inhibiting STAT4 expression. We found that STAT4 expression was upregulated in porcine GCs exposed to 1% O2. Both gain and loss of function assays confirmed that STAT4 was required for cell apoptosis under hypoxia conditions, and that the GC apoptosis caused by hypoxia was markedly attenuated following FF-MAS treatment through inhibition of STAT4 expression. Correlation analysis in vivo revealed that GC apoptosis was associated with increased STAT4 expression, while the FF-MAS content in follicular fluid was negatively correlated with STAT4 mRNA levels and cell apoptosis. These findings elucidate a novel role of FF-MAS-mediated protection of GCs by inhibiting STAT4 expression under hypoxia, which might contribute to the mechanistic understanding of follicular development., (© The Author(s) 2024. Published by Oxford University Press on behalf of the American Society of Animal Science. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Many studies have indicated that histone deacetylase inhibitors (HDACis) have a significant antitumor effect in cancer. Here we report a compound named LT-548-133-1 that not only acts as an HDAC inhibitor but also interferes with microtubule assembly to inhibit MCF-7 cell proliferation and induce apoptosis. Consistent with Chidamide, LT-548-133-1 inhibited HDAC activity and increased histone H3 acetylation. But the difference is that it significantly induced cell cycle G2/M arrest while Chidamide caused G0/G1 arrest in MCF-7 cells. By Western blotting, we found the accumulation of CyclinB1 and phosphorylated histone H3 in LT-548-133-1 treated cells. Immunofluorescence based microtubule-repolymerization experiments and immunofluorescence staining of cell microtubules and nuclei showed that LT-548-133-1inhibited microtubule-repolymerization and induced mitotic abnormalities. The decreased expression of Bcl-2 and the increased expression of Bax, p53, p21, and cleaved-Caspase3 indicated the occurrence of apoptosis. Flow cytometry results also showed an increase in the proportion of apoptotic cells after administration of LT-548-133-1 or Chidamide. Therefore, we demonstrated that LT-548-133-1 could act as an HDAC inhibitor while inhibiting microtubule-repolymerization, causing mitosis to be arrested in G2/M. These two effects ultimately lead to proliferation inhibition and apoptosis of MCF-7 cells., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by proliferation and insufficient apoptosis of fibroblast-like synoviocytes (FLSs).The biology and functions of interleukin (IL)-34 are only beginning to be uncovered. We previously demonstrated IL-34 could upregulate the expression of IL-17 in RA patients. In this study, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry of Annexin V and PI staining were performed to assess cell proliferation and apoptosis progression in RA-FLSs after stimulated with increasing concentrations of IL-34, respectively. Inflammatory cytokines and angiogenic factors were measured using quantitative real-time PCR, Western blotting and ELISA. We explored the association between IL-34 and RA-FLS proliferation and apoptosis in the context of RA. Stimulating RA-FLSs with different concentrations of IL-34 significantly promoted the proliferation and inhibited the apoptosis of RA-FLSs in a concentration-dependent manner. Neutralization of IL-17 with the IL-17 inhibitor plumbagin (PB) reduced the effects of IL-34. Proinflammatory cytokine (IL-17A IL-6 and tumor necrosis factor-α, TNF-α) and angiogenic factor (vascular endothelial growth factor, VEGF and hypoxia-inducible factor-1α, HIF-1α) expression was markedly upregulated in RA-FLSs stimulated by IL-34. PB-mediated inhibition of IL-17A also decreased the expression of IL-6, TNF-α, HIF-1α and VEGF in RA-FLSs. Taken together, these findings suggest that targeting IL-34 production in RA-FLSs may be a therapeutic strategy for RA., (© 2021. The Author(s).)

MicroRNAs (miRNAs) are involved in many pathological and biological processes, such as ischemia/reperfusion (I/R) injury by modulating gene expression. Increasing evidence indicates that miR-378a-3p might provide a potential cardioprotective effect against ischemic heart disease. Cell apoptosis is a crucial mechanism in I/R injury. As such, this study evaluated the protective effects and underlying mechanisms of action of miR-378a-3p on H9C2 cardiomyocyte apoptosis following I/R injury. We found that I/R-induced H9C2 cardiomyocytes exhibited a decrease in miR-378a-3p expression, while treatment with a miR-378a-3p mimic suppressed cell apoptosis, JNK1/2 activation, cleavage of PARP and caspase-3, and Bax/Bcl-2 ratio but increased DUSP1 expression, which subsequently inhibited JNK1/2 phosphorylation. TRIM55 was shown to be a target of miR-378a-3p and its downregulation inhibited the miR-378a-3p inhibitor-induced increase in cell apoptosis and JNK1/2 activation. TRIM55 inhibited DUSP1 protein expression through ubiquitination of DUSP1. Moreover, DUSP1 overexpression inhibited the TRIM55 overexpression-induced increase in cell apoptosis and JNK1/2 activation. The protective effect of miR-378a-3p was subsequently confirmed in a rat myocardial I/R model, as evidenced by a decrease in cardiomyocyte apoptosis of cardiomyocytes, TRIM55 expression, and JNK1/2 activation. Taken together, these results suggest that miR-378a-3p may protect against I/R-induced cardiomyocyte apoptosis via TRIM55/DUSP1/JNK signaling.

Introduction: Rare-earth nanoparticles in the environment and human body pose a potential threat to human health. Although toxic effects of rare-earth nanoparticles have been extensively studied, the effects on the early development are not well understood. In this study, we attempted to explain the toxic effects of neodymium oxide (Nd 2 O 3 ) nanoparticles on early development., Methods: We added the Nd 2 O 3 nanoparticles at different concentrations and recorded the mortality and malformation rate per 24 hrs under a microscope. The live embryos treated with Nd 2 O 3 nanoparticles were imaged as movies and Z step lapses with a confocal microscope, and heart rates were counted for 30 s to measure the cardiac function. The live Tg ( Flk1 :EGFP) transgenic embryos exposed to Nd 2 O 3 nanoparticles were observed under confocal microscope to measure the cerebrovascular development. Subsequently, we extracted the total protein for Western blot at 5 days post-fertilisation (dpf). Embryos were collected to undergo TUNEL staining for apoptosis detection., Results: Nd 2 O 3 nanoparticles disturbed embryo development at high concentrations (>200 μg/mL). The mortality and malformation rate gradually increased in a dose-dependent manner by morphological observation, while the Nd 2 O 3 median lethal concentration (LD50) was 203.4 μg/mL at 120 hrs post-fertilisation (hpf). Furthermore, the Nd 2 O 3 -treated embryos showed severe arrhythmia and reduced heart rate. We also observed the markedly cerebrovascular disappearance at middle concentration (100 and 200 μg/mL). The downregulated autophagy flux in brain blood vessels and increased apoptosis level in neurons might affect vessels sprouting and contribute to the vanished cerebrovascular., Conclusion: The results suggested that the embryos exposed to Nd 2 O 3 activated the apoptosis pathway and induced toxicity and abnormal cardiac/cerebrovascular development., Competing Interests: The authors report no conflicts of interest in this work., (© 2020 Chen et al.)

Objective: Gastrodin, a glucoside of gastrodigenin, inhibits cerebral oxidant stress and apoptosis in multiple central nervous system injury, but its effect in intracerebral hemorrhage (ICH) remains unclear. This study investigated the effect of gastrodin on neuronal apoptosis and neurological deficits in rat ICH model., Methods: In vitro experiments were performed using hematoma lysate-induced cell damage model in primary cortical neurons. Rat ICH model was produced by a caudatum injection of collagenase. Gastrodin was intraperitoneal injected after 2 hours following ICH. Cell viability, brain water content, neurological score, western blot, and immunofluorescence experiments were performed., Results: Gastrodin significantly decreased hematoma lysate-induced reduction of cell viability and cell apoptosis in primary cortical neurons. Gastrodin significantly improved brain edema and neurological deficits post-ICH. Moreover, gastrodin administration significantly reduced levels of ROS, 8-OHDG, 3-Nitrotyrosine and MDA, while increased GSH-Px and SOD activity, and stimulated the upregulation of Keap1, Nrf2, and HO-1 signaling at 72 hours post-ICH. Furthermore, gastrodin significantly increased Bcl-2 expression, while reduced level of Bax, active caspase-3 and active caspase-9, also reduced the number of active caspase-3 or TUNEL positive neurons at 72 hours post-ICH., Conclusion: These results suggest that gastrodin is neuroprotective after ICH and the mechanism may be associated with the inhibition of oxidative stress and neuronal apoptosis., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Excessive plasma triglyceride (TG) and cholesterol levels promote the progression of several prevalent cardiovascular risk factors, including atherosclerosis, which is a leading death cause. Perilipin 5 (Plin5), an important perilipin protein, is abundant in tissues with very active lipid catabolism and is involved in the regulation of oxidative stress. Although inflammation and oxidative stress play a critical role in atherosclerosis development, the underlying mechanisms are complex and not completely understood. In the present study, we demonstrated the role of Plin5 in high-fat-diet-induced atherosclerosis in apolipoprotein E null (ApoE -/- ) mice. Our results suggested that Plin5 expressions increased in the artery tissues of ApoE -/- mice. ApoE/Plin5 double knockout (ApoE -/- Plin5 -/- ) exacerbated severer atherogenesis, accompanied with significantly disturbed plasma metabolic profiles, such as elevated TG, total cholesterol, and low-density lipoprotein cholesterol levels and reduced high-density lipoprotein cholesterol contents. ApoE -/- Plin5 -/- exhibited a higher number of inflammatory monocytes and neutrophils, as well as overexpression of cytokines and chemokines linked with an inflammatory response. Consistently, the IκBα/nuclear factor kappa B pathway was strongly activated in ApoE -/- Plin5 -/- . Notably, apoptosis was dramatically induced by ApoE -/- Plin5 -/- , as evidenced by increased cleavage of Caspase-3 and Poly (ADP-ribose) polymerase-2. In addition, ApoE -/- Plin5 -/- contributed to oxidative stress generation in the aortic tissues, which was linked with the activation of phosphatidylinositol 3-kinase/protein kinase B and mitogen-activated protein kinases pathways. In vitro, oxidized low-density lipoprotein (ox-LDL) increased Plin5 expression in RAW264.7 cells. Its knockdown enhanced inflammation, apoptosis, oxidative stress, and lipid accumulation, while promotion of Plin5 markedly reduced all the effects induced by ox-LDL in cells. These studies strongly supported that Plin5 could be a new regulator against atherosclerosis, providing new insights on therapeutic solutions., (© 2019 Wiley Periodicals, Inc.)

Aims: Sepsis is a syndrome of inflammatory response induced by infection. Cellular adhesion molecules may involve in sepsis-induced myocardial dysfunction (SIMD) which is a major predictor of morbidity and mortality of sepsis. Here we studied the role of FAM46C in AC16 cells and c57 mice with lipopolysaccharides (LPS) treatment., Main Methods: Real-time PCR and western blot were used to detect the expression level of relative genes and protein. Cell proliferation and apoptosis were evaluated., Key Findings: Interestingly, negative correlation between Toll-like receptor 4 (TLR4) and FAM46C in sepsis was observed. The overexpression of FAM46C reduced the apoptosis induced by LPS in AC16 cells. Inhibition of apoptosis contributed by FAM46C was mediated by adhesion molecule via blocking p38 and ERK/MAPK signaling pathway. Moreover, overexpression of Fam46c and inhibition of TLR4 by TAK-242 could attenuate apoptosis induced by LPS in vivo., Significance: FAM46C played an important role in SIMD via inhibiting LPS-induced myocardial dysfunction by downregulating cellular adhesion molecules and inhibiting apoptosis. It was the first time to explore the role of FAM46C in SIMD in this study., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Pathological stimuli, such as bacterial activity, dental bleaching, and nonpolymerized resin monomers, can cause death of dental pulp cells (DPCs) through oxidative stress- (OS-) induced mitochondrial dysfunction. However, the crucial molecular mechanisms that mediate such a phenomenon remain largely unknown. OS is characterized by the overproduction of reactive oxygen species (ROS), e.g., H 2 O 2 , O 2 - , and · OH. Mitochondria are a major source of ROS and the principal attack target of ROS. Cyclophilin D (CypD), as the only crucial protein for mitochondrial permeability transition pore (mPTP) induction, facilitates the opening of mPTP and causes mitochondrial dysfunction, leading to cell death. In the present study, we hypothesized that CypD-mediated mitochondrial molecular pathways were closely involved in the process of OS-induced death of human DPCs (HDPCs). We tested the phenotypic and molecular changes of HDPCs in a well-established OS model-H 2 O 2 treatment. We showed that H 2 O 2 dramatically reduced the viability and increased the death of HDPCs in a time- and dose-dependent manner by performing MTT, flow cytometry, and TUNEL assays and quantifying the expression changes of Bax and Bcl-2 proteins. H 2 O 2 also induced mitochondrial dysfunction, as reflected by the increased mitochondrial ROS, reduced ATP production, and activation of mPTP (decreased mitochondrial membrane potential and enhanced intracellular Ca 2+ level). An antioxidant (N-acetyl-L-cysteine) effectively preserved mitochondrial function and significantly attenuated H 2 O 2 -induced cytotoxicity and death. Moreover, H 2 O 2 treatment markedly upregulated the CypD protein level in HDPCs. Notably, genetic or pharmacological blockade of CypD significantly attenuated H 2 O 2 -induced mitochondrial dysfunction and cell death. These findings provided novel insights into the role of a CypD-dependent mitochondrial pathway in the H 2 O 2 -induced death in HDPCs, indicating that CypD may be a potential therapeutic target to prevent OS-mediated injury in dental pulp.

Accumulating evidence indicates that post-injury inflammation characterized by activated microglia contributes much to the neuropathology of ischemic injury. Several studies have demonstrated that microglia exhibit two entirely different functional activation states, referred to as classically activated (M1) and alternatively activated (M2) phenotype. Promoting microglial phenotype to switch from M1 dominant to M2 dominant might be a promising approach for handling ischemic injury. However, the comprehensive mechanism that underlines microglia polarization in ischemic brain remains unclear. Neuronal erythropoietin-producing human hepatocellular carcinoma cell receptor 4 (EphA4), the richest Eph receptor in the central nervous system (CNS), upregulate after ischemia and may have the potential to regulate microglia activation. We hypothesized that modulating EphA4/ephrin signaling could affect ischemic injury through controlling microglia polarization. We therefore knocked down neuronal EphA4 with short hairpin RNA (shRNA) and determined the role of EphA4/ephrin signaling in oxygen-glucose deprivation and reperfusion (OGD/R)-induced injury. We found that EphA4 shRNA treatment attenuated OGD/R-induced apoptosis and microglia proliferation. Neuronal EphA4 knockdown also promoted microglial M2 polarization, which reduced pro-inflammatory mediators and released anti-inflammatory cytokines as well as neurotrophic factors. We further revealed that EphA4 shRNA treatment functioned through RhoA/Rho-associated kinase 2 (ROCK2) signaling, a key mediator of microglia alternative activation. Together, these data suggested that blockage of EphA4/ephrin signaling between neuron and microglia decreased OGD/R-induced injury by promoting alternative activation of microglia via RhoA/ROCK2 signaling.

Background: We demonstrated growing evidence supports a protective role of chlorogenic acid of rat hepatocytes elicited by two compounds, i.e. thapsigargin and palmitic acid. Nevertheless, little is known about the mechanisms of palmitic acid induced endoplasmic reticulum (ER) stress and cell death., Methods: The proliferation of primary rat hepatocytes was detected by MTT assay. The expression of GRP78, CHOP and GRP94 was detected by Western blot analyses. Caspase-3 activity was detected by a Caspase-3 substrate kit. Cell apoptosis was detected by Hoechst 33342 staining., Results: We demonstrated that incubation of hepatocytes for 16 h with palmitic acid elevated cell death. Moreover, Western blot analyses demonstrated increased levels of the endoplasmic reticulum stress markers - glucose regulated protein 78 (GRP78), C/EBP homologous protein (CHOP), and glucose regulated protein 94 (GRP94). Chlorogenic acid could inhibit ER stress induced cell death and levels of indicators of ER stress caused by palmitic acid. The effect of thapsigargin, which evokes ER stress were reversed by chlorogenic acid., Conclusions: Altogether, our data indicate that in primary rat hepatocytes, chlorogenic acid prevents ER stress-mediated apoptosis of palmitic acid.

The identification and development of novel therapeutic strategies for acute lung injury is urgently required. It has been previously demonstrated that microRNA (miR)‑16 suppresses the level of transforming growth factor (TGF)‑β in acute lung injury (ALI). Therefore, the present study investigated the role of miR‑16 in the phenotype, cell proliferation and apoptosis, and the involvement of TGF‑β/Smad family member 2 (Smad2) and JAK/signal transducer and activator of transcription (STAT)3 signaling, of primary human alveolar type II epithelial cells (AECII). Following transfection with miR‑16 mimics, AECII cells were exposed to hyperoxia for 24 h. Subsequently, immunofluorescence staining of surfactant protein‑A (SP‑A) was performed, and cell proliferation and apoptosis were investigated by Cell Counting Kit‑8 assays and annexin V‑fluorescein isothiocyanate/propidium iodide staining, respectively. Furthermore, the expression levels of miR‑16, TGF‑β, Smad2, phosphorylated‑Smad2, JAK and STAT3 were detected by western blotting and/or reverse transcription‑quantitative polymerase chain reaction. The results demonstrated that miR‑16 levels and SP‑A fluorescence were markedly inhibited by hyperoxia. Furthermore, transfection of AECII cells with miR‑16 mimics increased SP‑A fluorescence in hyperoxia‑treated AECII cells, significantly reversed hyperoxia‑induced reductions in cell proliferation and inhibited hyperoxia‑induced apoptosis. Finally, miR‑16 mimics modulated the mRNA and protein expression of components of the TGF‑β/Smad2 and JAK/STAT3 pathways in AECII cells following hyperoxia. In conclusion, the results of the present study indicate that overexpression of miR‑16 may exert a protective effect in AECII cells against cell apoptosis and ALI, which may be associated with TGF‑β/Smad2 and JAK/STAT3 signaling pathways. This may also represent a promising target for novel therapeutic strategies for acute lung injury.

Background/aims: Gastric cancer is considered as a common malignancy with a poor prognosis as well as unsatisfactory treatment. Neutrophil gelatinase-associated lipocalin (NGAL) has been reported to affect multiple aspects of human tumor, including gastric cancer. This study aims to explore the effects of NGAL gene silencing on the proliferation as well as apoptosis of human gastric cancer MGC-803 cells., Methods: This study included 87 patients with gastric cancer. MGC-803 cells were collected and mainly treated with siRNA against NGAL and recombinant NGAL plasmid. The expression of NGAL mRNA and the expressions of NGAL protein and apoptosis-related proteins were determined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis, respectively. Cell cycle and apoptosis were tested by flow cytometry, and cell proliferation was detected by water soluble tetrazolium-1 (WST-1) assay. The effect of NGAL gene silencing on tumorigenicity of MGC-803 cells in vivo was detected through establishment of xenograft in nude mice., Results: NGAL was highly expressed in gastric cancer tissues. The protein and mRNA expressions of NGAL gene in MGC-803 cells treated with NGAL-siRNA were obviously reduced, and the amount of cells in G0/G1 phase was increased. Moreover, MGC-803 cells treated with NGAL-siRNA exhibited inhibited proliferation, enhanced apoptosis, decreased expressions of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) as well as B-cell lymphoma-2 (Bcl-2) and increased expressions of cysteine-aspartic acid specific protease-9 (caspase-9) and Bcl2-associated X (Bax), as well as repressed tumorigenicity in vivo., Conclusion: NGAL gene silencing inhibits proliferation and promotes apoptosis of MGC-803 cells, which can provide a novel theory for treatment of gastric cancer., (© 2018 The Author(s). Published by S. Karger AG, Basel.)

Nearly all clinical trials that have attempted to develop effective strategies against ischemic stroke have failed, excluding those for thrombolysis, and most of these trials focused only on preventing neuronal loss. However, astrocytes have gradually become a target for neuroprotection in stroke. In previous studies, we showed that the newly identified molecular N-myc downstream-regulated gene 2 (Ndrg2) is specifically expressed in astrocytes in the brain and involved in some neurodegenerative diseases. However, the role of NDRG2 in ischemic stroke remained unclear. In this study, we investigated the role of NDRG2 in middle cerebral artery occlusion (MCAO)-induced focal cerebral ischemia and in oxygen-glucose deprivation (OGD)-induced cellular apoptosis in the M1800 astrocyte cell line. NDRG2 mRNA and protein expression began to increase at 6 and 2 h after reperfusion and peaked at 24 h in the ischemic penumbra and in M1800 cells, as detected by RT-PCR and Western blotting. Double immunofluorescence staining showed that the number of apoptotic cells increased as the NDRG2-positive signal increased and that the NDRG2 signal was sometimes co-localized with TUNEL-positive cells and translocated from the cytoplasm to the nucleus in both the ischemic penumbra and the M1800 cells. Using a lentivirus, we successfully constructed two stable astrocytic cell lines in which NDRG2 expression was significantly up- or down-regulated. NDRG2 silencing had a proliferative effect and reduced the percentage of apoptotic cells, reactive oxygen species (ROS) production, and cleaved Caspase-3 protein expression following OGD, whereas NDRG2 over-expression had the opposite effects. In conclusion, NDRG2 is involved in astrocyte apoptosis following ischemic-hypoxic injury, and inhibiting NDRG2 expression significantly reduces ROS production and astrocyte apoptosis. These findings provide insight into the role of NDRG2 in ischemic-hypoxic injury and provide potential targets for future clinical therapies for stroke.

Quercetin is a potent cancer therapeutic agent and dietary antioxidant present in fruit and vegetables. Quercetin prevents tumor proliferation by inducing cell cycle arrest and is a well known cancer therapeutic agent and autophagy mediator. Recent studies showed that drug delivery by nanoparticles have enhanced efficacy with reduced side effects. In this regard, gold-quercetin into poly(DL-lactide-co-glycolide) nanoparticles was examined. In this study, we explored the role and possible underlying mechanisms of quercetin nanoparticle in regulation of antitumor activity in liver cancer cells. Treatment with quercetin nanoparticle effectively inhibited the liver cancer cell proliferation, cell migration and colony formation, thus suppressing liver cancer progression. Quercetin nanoparticle also upregulated apoptosis markedly. Further study suggested that quercetin nanoparticle accelerated the cleavage of caspase-9, caspase-3, and induced the up-releasing of cytochrome c (Cyto-c), contributing to apoptosis in liver cancer cells. Quercetin nanoparticles also promoted telomerase reverse transcriptase (hTERT) inhibition through reducing AP-2β expression and decreasing its binding to hTERT promoter. In addition, quercetin nanoparticle had an inhibitory role in cyclooxygenase 2 (COX-2) via suppressing the NF-κB nuclear translocation and its binding to COX-2 promoter. Quercetin nanoparticle also inactivated Akt and ERK1/2 signaling pathway. Taken together, our results suggested that quercetin nanoparticle had an antitumor effect by inactivating caspase/Cyto-c pathway, suppressing AP-2β/hTERT, inhibiting NF-κB/COX-2 and impeding Akt/ERK1/2 signaling pathways. Our results provided new mechanistic basis for further investigation of quercetin nanoparticles to find potential therapeutic strategies and possible targets for liver cancer inhibition.

Nischarin, a novel integrin binding protein, has been demonstrated its negative effects on cell migration and invasion. However, the biological role of Nischarin in breast cancer has not been fully elucidated yet. Our study aimed to analyze the association between Nischarin expression and clinical features of breast cancer patients, and further investigate the role of Nischarin in breast cancer cells apoptosis, migration and invasion. Results showed that Nischarin expression was significantly lower in breast cancer tissues (37.8%, 23/67) than in normal tissues (61.8%, 21/34; P < 0.05), and the expression of Nischarin significantly negatively correlated with estrogen receptor status. Similarly, Nischarin expression was highest in normal breast cell line HBL-100 while triple-negative breast cancer cell line MDA-MB-231 had the lowest expression of Nischarin. Further experiments demonstrated that overexpression of Nischarin may induce apoptosis, and inhibit cell migration and invasion. The present data confirmed that Nishcharin might be a novel tumor suppressor and plays an important role in breast cancer cell apoptosis and metastasis, which can be used as a potential therapeutic target for breast cancer treatment., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Galangin is an active pharmacological ingredient from propolis and Alpinia officinarum Hance, and has been reported to have anti-cancer and antioxidative properties. Berberine, a major component of Berberis vulgaris extract, exhibits potent anti-cancer activities through distinct molecular mechanisms. However, the anticancer effect of galangin in combination with berberine is still unknown. In the present study, we demonstrated that the combination of galangin with berberine synergistically resulted in cell growth inhibition, apoptosis and cell cycle arrest at G2/M phase with the increased intracellular reactive oxygen species (ROS) levels in oesophageal carcinoma cells. Pretreatment with ROS scavenger promoted the apoptosis dramatically induced by co-treatment with galangin and berberine. Treatment with galangin and berberine alone caused the decreased expressions of Wnt3a and β-catenin. Interestingly, combination of galangin with berberine could further suppress Wnt3a and β-catenin expression and induce apoptosis in cancer cells. Additionally, in nude mice with xenograft tumors, the combinational treatment of galangin and berberine significantly inhibited the tumor growth without obvious toxicity. Overall, galangin in combination with berberine presented outstanding synergistic anticancer role in vitro and in vivo, indicating that the beneficial combination of galangin and berberine might provide a promising treatment for patients with oesophageal carcinoma., (Copyright © 2016. Published by Elsevier Masson SAS.)

Growing evidence suggests that microRNA plays an essential role in the development and metastasis of many tumors, including gastric cancer. Aberrant miR‑370 expression has been indicated in tumor growth, but the mechanism of miR‑370 inhibits both the proliferation and metastatic ability for gastric cancer remains unclear. Accumulating evidence reported that PTEN signaling pathway plays an important role in the cellular processes, such as apoptosis, cell growth and proliferation. The goal of this study was to identify whether miR‑370 could inhibit the growth, migration, invasion, proliferation and metastasis of gastric cancer through targeting PTEN. Real-time PCR (RT-PCR) was used to quantify miR-370 expression in vitro experiments. The biological functions of miR‑370 were determined via cell proliferation. Our study indicated that miR‑370 targeted PTEN leading to activation of apoptosis signaling and the cell proliferation of cervical cancer cells, ameliorating gastric cancer growth and progression. In addition, the combination of miR‑370 and PTEN inactivated AKT, MDM2 and mTOR while stimulated caspase-3, p53 and GSK3β expression, promoting apoptosis and suppressing proliferation of gastric cancer cells. Therefore, our study revealed the mechanistic links between miR‑370 and PTEN in the pathogenesis of gastric cancer through modulation of cell apoptosis and proliferation. Additionally, targeting miR‑370 could serve as a novel strategy for future gastric cancer therapy clinically.

Although Beclin 1 has been demonstrated to exert an important role in cell autophagy during carcinogenesis, its biological function in lung cancer has yet to be fully elucidated. A previous study by our laboratory identified that knockdown of Beclin 1 promoted cell growth and inhibited apoptosis in the A549 lung cancer cell line. In the present study, a Beclin 1 lentiviral expression vector was constructed, and an A549 cell line was established with a steady expression of Beclin 1. Furthermore, the effect of Beclin 1 overexpression on cell invasion and apoptosis, changes in the activities of the apoptosis‑associated caspases‑3 and ‑9, and the overexpression of esophageal cancer‑related gene 4 (ECRG4) were examined. The results demonstrated that the overexpression of Beclin 1 in A549 cells reduced cell invasion by Matrigel invasion assay and promoted apoptosis by flow cytometric analysis (P<0.01) compared with Lenex‑packaged lentiviral particles and non‑transfected control groups. Furthermore, the overexpression of Beclin 1 in A549 cells increased the activities of caspases‑3 and ‑9 and the expression of ECRG4 (P<0.01) compared with Lenex‑packaged lentiviral particles and non‑transfected control groups. In conclusion, the overexpression of Beclin 1 promoted apoptosis and decreased invasion by upregulating the expression of ECRG4 in A549 lung adenocarcinoma cells. Therefore, the selection of Beclin l as a target for gene therapy represents a more effective method for the treatment of lung cancer.

Galangin, a flavonoid extracted from the root of the Alpinia officinarum Hence, has been shown to have anticancer properties against several types of cancer cells. However, the influence of galangin on human renal cancer cells remains to be elucidated. In the present study, proliferation of 786‑0 and Caki‑1 cells was suppressed following exposure to various doses of galangin. Cell invasion and wound healing assays were used to observe the effect of galangin on invasion and migration. The results demonstrated that Galangin inhibited cell invasion by suppressing the epithelial mesenchymal transition (EMT), with an increase in the expression of E‑cadherin and decreased expression levels of N‑cadherin and vimentin. The apoptosis induced by galangin was analyzed by flow cytometry. The results revealed that galangin induced apoptosis in a dose‑dependent manner. The accumulation of reactive oxygen species (ROS) is an important contributing factor for the apoptosis of various types of cancer cell. The dichlorofluorescein-diacetate method was used to determine the level of ROS. Galangin induced the accumulation of intracellular ROS and malondialdehyde, and decreased the activities of total antioxidant and superoxide dismutase in renal cell carcinoma cells. Galangin exerted an antiproliferative effect and inhibited renal cell carcinoma invasion by suppressing the EMT. This treatment also induced apoptosis, accompanied by the production of ROS. Therefore, the present data suggested that galangin may have beneficial effects by preventing renal cell carcinoma growth, inhibiting cell invasion via the EMT and inducing cell apoptosis.

Background/aims: Global cerebral ischemia/reperfusion (GCIR) may incur neurocognitive impairment. Tea polyphenols (TP) have strong anti-oxidant capacity. This study planned to investigate the protective effect of TP against the neurocognitive impairment caused by GCIR and its mechanism., Methods: One-stage anterior approach for cerebral four-vessel occlusion (4VO) was used to construct the GCIR model. Sprague Dawley rats were randomly classified into Sham group, GCIR group, and TP group (n = 50 per group). Besides receiving the same 4VO, the rats in TP group were treated with TP (6.4%) injection from the tail vein 30 minutes before cerebral ischemia. Morris water-maze test was used to evaluate the changes in space recognition and memory and open field activity test to assess the activity and motor function of rats. The cell apoptotic study in hippocampal CA1 region at specified time points (12, 24, 48, and 72 hours after surgery) was carried out by the flow cytometry, histology (hematoxylin and eosin staining), and immunohistochemical (terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining) examinations. One-way analysis of variance and least significant difference t-test were used and statistical significance considered at P < 0.05., Results: Compared with the GCIR group, the TP group was significantly attenuated in the impairment of space recognition and memory caused by GCIR and so was the neuronal apoptosis in the hippocampal CA1 region (P < 0.05)., Conclusion: TP may attenuate the impairment of space recognition and memory caused by GCIR via anti-apoptosis.

Caffeic acid phenyl ester (CAPE) is a potent anti-inflammatory agent and it can eliminate the free radicals. This study aimed to investigate the radioprotective effects of CAPE on X-ray irradiation induced intestinal injury in rats. Rats were intragastrically administered with 10 μmol/kg/d CAPE for 7 consecutive days before exposing them to a single dose of X-ray irradiation (9Gy) to abdomen. Rats were sacrificed 72 h after exposure to radiation. We found that pretreatment with CAPE effectively attenuated intestinal pathology changes, apoptosis, oxidative stress, bacterial translocation, the content of nitric oxide and myeloperoxidase as well as the concentration of plasma tumor necrosis factor-α. Pretreatment with CAPE also reversed the activation of p38MAPK and the increased expression of intercellular cell adhesion molecule-1 induced by radiation in intestinal mucosa. Taken together, these results suggest that pretreatment with CAPE could be a promising candidate for treating radiation-induced intestinal injury., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Objective: Apogossypolone (ApoG2), a new derivative of gossypol, is a potent cell-growth inhibitor. ApoG2 has been demonstrated to have superior anti-tumor activity than gossypol in Bcl-2 transgenic mice. The purpose of this study was to investigate the inhibitory effect of ApoG2 on breast cancer cell line MCF-7 in vitro and in vivo, and to investigate its anti-tumor mechanism., Methods: MCF-7 cell line in culture was treated with ApoG2. The inhibitory effects of ApoG2 on cell growth were measured by MTT and colony-formation assay. The cell apoptotic rate and cell cycle were analyzed by use of flow cytometry (FCM). The ultrastructural changes were observed by transmission electron microscopy. Autophagy was detected by acridine orange staining. Expression of Bcl-2, Bax, and Beclin 1 proteins was measured by western blot analysis., Results: The inhibitory effect of ApoG2 on MCF-7 cell proliferation was dose and time-dependent. The maximum effect was observed when cells were incubated for 72 h with 40 μM ApoG2. ApoG2 at 5 μM also inhibited colony formation. FCM assay indicated that ApoG2 induced cell apoptosis and caused cell arrest in the S phase and G2/M phase. Transmission electron microscopic examination and acridine orange staining showed that ApoG2 induced intracellular autolysosome formation. Furthermore, ApoG2 reduced Bcl-2 expression, and enhanced expression of Bax and Beclin 1. Xenografting of MCF-7 cells in mice can also be inhibited by ApoG2., Conclusion: ApoG2, a novel anti-apoptotic Bcl-2 agent, inhibits proliferation of breast cancer cell line MCF-7 by inducing cell apoptosis and autophagy.

Suberoylanilide hydroxamic acid (SAHA), a potent pan-histone deacetylase (HDAC) inhibitor, has been clinically approved for the treatment of cutaneous T-cell lymphoma (CTCL). SAHA has also been shown to exert a variety of anticancer activities in many other types of tumors, however, few studies have been reported in large-cell lung carcinoma (LCC). Our study aimed to investigate the potential antitumor effects of SAHA on LCC cells. Here, we report that SAHA was able to inhibit the proliferation of the LCC cell line NCI-H460 in a dose- and time-dependent manner, induced cell apoptosis and G2/M cell cycle arrest, decreased AKT and ERK phosphorylation, inhibited the expression of pro-angiogenic factors (VEGF, HIF-1α) in vitro, and suppressed tumor progression in an NCI-H460 cell nude mouse xenograft model in vivo. These results indicate that SAHA can exert its strong antitumor effects in LCC patient.

Invasion of human intestinal epithelial cells (HCT-8) by Cryptosporidium parvum resulted in a rapid induction of host cell spermidine/spermine N(1)-acetyltransferase 1 (hSSAT-1) mRNA, causing a 4-fold increase in SSAT-1 enzyme activity after 24 h of infection. In contrast, host cell SSAT-2, spermine oxidase, and acetylpolyamine oxidase (hAPAO) remained unchanged during this period. Intracellular polyamine levels of C. parvum-infected human epithelial cells were determined, and it was found that spermidine remained unchanged and putrescine increased by 2.5-fold after 15 h and then decreased after 24 h, whereas spermine decreased by 3.9-fold after 15 h. Concomitant with these changes, N(1)-acetylspermine and N(1)-acetylspermidine both increased by 115- and 24-fold, respectively. Increased SSAT-1 has previously been shown to be involved in the endoplasmic reticulum (ER) stress response leading to apoptosis. Several stress response proteins were increased in HCT-8 cells infected with C. parvum, including calreticulin, a major calcium-binding chaperone in the ER; GRP78/BiP, a prosurvival ER chaperone; and Nrf2, a transcription factor that binds to antioxidant response elements, thus activating them. However, poly(ADP-ribose) polymerase, a protein involved in DNA repair and programmed cell death, was decreased. Cumulatively, these results suggest that the invasion of HCT-8 cells by C. parvum results in an ER stress response by the host cell that culminates in overexpression of host cell SSAT-1 and elevated N(1)-acetylpolyamines, which can be used by a parasite that lacks ornithine decarboxylase.

The expression of BECLIN1 is significantly reduced in non‑small cell lung cancer (NSCLC) compared with non‑cancerous tissue. However, the role of BECLIN1 in lung cancer is unclear. Using the RNA interference (RNAi) technique the present study investigated the effect of the knockdown of BECLIN1 on the cell growth and proliferation of the A549 human lung cancer cell line. The target site for the RNAi technique was designed and the lentivirus vector for the small interfering (si)RNA expression was constructed according to the encoding sequence of the mRNA for BECLIN1. The A549 cells were transfected with the siRNA virus against BECLIN1. BECLIN1 expression was detected by reverse transcription (RT)‑PCR and western blot analysis. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method was applied to detect cell growth. Flow cytometry was used to determine cell apoptosis. The activity of caspase‑3 and caspase‑9 was also detected in the A549 cells with BECLIN1 knockdown. The results showed that siRNA virus transfection significantly decreased the mRNA and protein expression of BECLIN1 in the transfected A549 cells. The knockdown of BECLIN1 promoted cell growth and decreased apoptosis. Caspase‑3 and ‑9 activity in the A549 cells with BECLIN1 knockdown was significantly reduced. In conclusion, the siRNA expression vector effectively inhibited the expression of BECLIN1 in the A549 human lung cancer cell line and promote the growth and proliferation of the A549 cells.

Objective: Resveratrol shows chemopreventive activity in a variety of human cancers by targeting mitochondria and triggering apoptosis. The purpose of this study was to investigate the antitumor action of resveratrol in bladder cancer and its underlying mechanism., Methods: Using two different bladder cell lines, BTT739 and T24, the cytotoxicity of resveratrol were determined by MTT assay. The apoptosis induced by resveratrol was assayed by transferase dUTP nick end labeling staining. To show whether the mitochondrial dysfunction involved in the effects of resveratrol, mitochondrial function was detected by mitochondrial membrane potential, reactive oxygen species production and adenosine 5'-triphosphate content. In addition, the markers of apoptosis in the intrinsic mitochondrial-dependent pathway were analyzed by the release of cytochrome c and the activities of caspase-9 and caspase-3., Results: Resveratrol effectively decreased cell viability and induced apoptosis in a concentration- and time-dependent manner. In addition, resveratrol significantly disrupted the mitochondrial membrane potential in both intact cells and isolated mitochondria. Resveratrol also increased reactive oxygen species production and reduced adenosine 5'-triphosphate concentrations. Western blot analysis showed that resveratrol provoked the release of cytochrome c from mitochondria to the cytosol. Furthermore, resveratrol significantly promoted the activation of caspase-9 and caspase-3., Conclusions: These findings suggest that resveratrol efficiently triggers apoptosis in bladder cancer cells through the intrinsic mitochondrial-dependent pathway, which is associated with mitochondrial dysfunction. Resveratrol might have great pharmacological promise in the treatment of bladder cancer., (© 2012 The Japanese Urological Association.)

Many pro-apoptotic factors, such as nuclear factor-kappa B (NF-κB) and Fas, play crucial roles in the process of Leydig cell apoptosis, ultimately leading to male sterility, such as in Sertoli cell only syndrome (SCO) and hypospermatogenesis. However, the molecular mechanism of such apoptosis is unclear. Recent reports on N-myc downstream-regulated gene 2 (ndrg2) have suggested that it is involved in cellular differentiation, development, and apoptosis. The unique expression of NDRG2 in SCO and hypospermatogenic testis suggests its pivotal role in those diseases. In this study, we analyzed NDRG2 expression profiles in the testes of normal spermatogenesis patients, hypospermatogenesis patients, and SCO patients, as well as in vivo and in vitro models, which were Sprague-Dawley rats and the Leydig cell line TM3 treated with the Leydig cell-specific toxicant ethane-dimethanesulfonate (EDS). Our data confirm that NDRG2 is normally exclusively located in the cytoplasm of Leydig cells and is up-regulated and translocates into the nucleus under apoptotic stimulations in human and murine testis. Meanwhile, transcription factor NF-κB was activated by EDS administration, bound to the ndrg2 promoter, and further increased in expression, effects that were abolished by NF-κB inhibitor Pyrrolidine dithiocarbamate (PDTC). Furthermore, siRNA knock-down of ndrg2 led to increased proliferative or decreased apoptotic TM3 cells, while over-expression of ndrg2 had the reverse effect. This study reveals that ndrg2 is a novel gene that participates in Leydig cell apoptosis, with essential functions in testicular cells, and suggests its possible role in apoptotic Leydig cells and male fertility., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Background: Spinal cord injury (SCI), including immediate mechanical injury and secondary injury, is associated with the inflammatory response, apoptosis and oxidative stress in response to traumatic injury. Tanshinone IIA (TIIA) is one of the major extracts obtained from Salvia miltiorrhiza BUNGE, which has anti-inflammatory and anti-apoptotic effects on many diseases. However, little is known about the effects of TIIA treatment on SCI. Therefore, the aim of the present study is to evaluate the pharmacological action of TIIA on secondary damage and the underlying mechanisms of experimental SCI in rats., Methodology/principal Findings: SCI was generated using a weight drop device on the dorsal spinal cord via a two-level T9-T11 laminectomy. SCI in rats resulted in severe trauma, characterized by locomotor disturbance, edema, neutrophil infiltration, the production of astrocytes and inflammatory mediators, apoptosis and oxidative stress. TIIA treatment (20 mg/kg, i.p.) after SCI induced significant effects: (1) improved motor function (Basso, Beattie and Bresnahan scores), (2) reduced the degree of tissue injury (histological score), neutrophil infiltration (myeloperoxidase activity) and the expression of astrocytes, (3) inhibited the activation of SCI-related pathways, such as NF-κB and MAPK signaling pathways, (4) decreased the production of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6) and iNOS, (5) reduced apoptosis (TUNEL staining, and Bcl-2 and caspase-3 expression) and (6) reversed the redox state imbalance., Conclusions/significance: The results clearly show that TIIA has a prominent protective effect against SCI through inhibiting the inflammatory response and apoptosis in the spinal cord tissue after SCI.

Objective: To investigate the effect of Bcl-2 overexpression on Fas and TNFR1-mediated apoptosis and its possible mechanism in rat hippocampus following global ischemia/reperfusion (IR)., Methods: Ninety healthy male SD rats were randomly divided into sham operated group, IR group and Bcl-2 overexpression group (BT group). Rat model of global IR was established by the 4-V0 method. The expressions of Bcl-2, Fas and TNFR1 and the cell apoptosis in the CA1 and CA3 regions were examined by HE staining, immunohistochemistry and TUNEL method., Results: In IR group, the neurons in the CA1 region showed an obvious reduction in number with disordered arrangement and interstitial edema 48 h after global IR. Such changes were not obvious in BT group. Immunohistochemistry showed that Fas expression in the CA1 region reached the peak level at 6 h in IR group with a greater expression intensity than that in BT group (P<0.05). TNFR1 was expressed at a higher level in IR group than in BT group (P<0.05), reaching the peak level at 24 h. In the sham group, the expression of Fas and TNFR1 was not detected the in CA1 and CA3 regions. Global IR caused increased cell apoptosis in the CA1 and CA3 regions, starting at 6 h and reached peak at 24 to 48 h. The cell apoptosis was less obvious in BT group (P<0.05)., Conclusion: Fas and TNFR1 are expressed in the CA1 and CA3 regions after global IR in rats, suggesting the involvement of death receptor in cerebral IR injury. Bcl-2 overexpression decreases the expression of Fas and TNFR1 and cell apoptosis after global IR, thus offering protective effect against cerebral IR injury.

Objective: To determine the effect of exogenous GM3 on proliferation, apoptosis and VEGF expression in human lung adenocarcinoma cell line A549 cells., Methods: A549 cells were treated with GM3 at different concentrations for 48 hours. MTT assay was used to detect the cell proliferation and flow cytometry was applied to analyze cell apoptosis. RT-PCR was used to detect the expression level of VEGF mRNA and confocal laser scanning microscopy was applied to observe the localization and fluorescence intensity of VEGF., Results: Comparing with the control, being treated with higher than 10 µmol/L GM3 significantly inhibited A549 cell proliferation (P < 0.05), and the suppressive effect could be enhanced following increasing doses. The IC(50) was 412 µmol/L. Comparing with the control, being treated with higher than 40 µmol/L GM3 significantly promoted the apoptotic rate of A549 cells (P < 0.05). Comparing with the control, being treated with higher than 40 µmol/L GM3 significantly decreased the VEGF mRNA level of A549 cells (P < 0.05), and the fluorescence intensity of VEGF distinctly weakened., Conclusions: Exogenous ganglioside GM3 can inhibit the proliferation, promote apoptosis, and down-regulate the VEGF expression level in A549 cells. This may be considered as two mechanisms of GM3 for its anti-tumor effect by modulating cell apoptosis and angiogenesis.

Objectives: The objective of the study was to evaluate the therapeutic effects of carbon quantum dots (CQDs) in immunomodulation on non‐obese diabetic (NOD) mice, as the model for Sjögren's syndrome (SS). Methods: Carbon quantum dots were generated from Setaria viridis via a hydrothermal process. Their toxic effects were tested by cell viability and blood chemistry analysis, meanwhile therapeutic effects were investigated in NOD mice in the aspects of saliva flow, histology, and immune cell distribution. Results: Carbon quantum dots, with rich surface chemistry and unique optical properties, showed non‐cytotoxicity in vitro or no damage in vivo. Intravenously applied CQDs alleviated inflammation in the submandibular glands in NOD mice after 6‐week treatments. The inflammatory area index and focus score were significantly decreased in CQD‐treated mice. Besides, the levels of anti‐SSA and anti‐SSB were decreased in the presence of CQDs. The stimulated saliva flow rates and weight of submandibular glands were significantly increased in CQD‐treated mice by reducing the apoptosis of cells. The CD3+ and CD4+ T cells distributed around the ducts of submandibular glands were significantly decreased, while the percentage of Foxp3+ cells was higher in CQD‐treated mice than that in the control group. Conclusions: Our findings suggest that CQDs may ameliorate the dysregulated immune processes in NOD mice. [ABSTRACT FROM AUTHOR]

The developmental fate of ovarian follicles is primarily determined by the survival status (proliferation or apoptosis) of granulosa cells (GCs). Owing to the avascular environment within follicles, GCs are believed to live in a hypoxic niche. Follicle‐stimulating hormone (FSH) has been reported to improve GCs survival by governing hypoxia‐inducible factor‐1α (HIF‐1α)‐dependent hypoxia response, but the underlying mechanisms remain poorly understood. Growth arrest‐specific gene 6 (GAS6) is a secreted ligand of tyrosine kinase receptors, and has been documented to facilitate tumor growth. Here, we showed that the level of GAS6 was markedly increased in mouse ovarian GCs after the injection of FSH. Specifically, FSH‐induced GAS6 expression was accompanied by HIF‐1α accumulation under conditions of hypoxia both in vivo and in vitro, whereas inhibition of HIF‐1α with small interfering RNAs/antagonist repressed both expression and secretion of GAS6. As such, Luciferase reporter assay and chromatin immunoprecipitation assay showed that HIF‐1α directly bound to a hypoxia response element site within the Gas6 promoter and contributed to the regulation of GAS6 expression in response to FSH. Notably, blockage of GAS6 and/or its receptor Axl abrogated the pro‐survival effects of FSH under hypoxia. Moreover, phosphorylation of Axl by GAS6 is required for FSH‐mediated Akt activation and the resultant pro‐survival phenotypes. Finally, the in vitro findings were verified in vivo, which showed that FSH‐induced proliferative and antiapoptotic effects in ovarian GCs were diminished after blocking GAS6/Axl using HIF‐1α antagonist. These findings highlight a novel function of FSH in preserving GCs viability against hypoxic stress by activating the HIF‐1a‐GAS6‐Axl‐Akt pathway. [ABSTRACT FROM AUTHOR]

Simple Summary: Tumor-associated macrophages (TAMs) and cancer-associated fibroblasts (CAFs) in the tumor microenvironment are critical factors in the curative effects of cancer therapies. The aim of our study was to explore the interactions between TAMs and CAFs in the context of ionizing radiation. We confirmed that M2 macrophages correlate with poor prognosis and induce radioresistance in cervical cancer. M2 polarization was increased after high-dose IR in both mouse models and patients with cervical cancer. M2 macrophage contents were increased in CAF-positive regions in patients with cervical cancer who relapsed after receiving radical radiotherapy. In addition, high-dose irradiated CAFs promote macrophage M2 polarization in cervical cancer through the secretion of chemokine (C-C motif) ligand 2 (CCL2). Our data provide a new insight into the relation between CAFs and TAMs under IR, which is of significance for further exploration of the mechanism of radioresistance in cervical cancer. Radiotherapy, including brachytherapy, is a major therapeutic regimen for cervical cancer. Radioresistance is a decisive factor in radiation treatment failure. Tumor-associated macrophages (TAMs) and cancer-associated fibroblasts (CAFs) in the tumor microenvironment are critical factors in the curative effects of cancer therapies. However, the interactions between TAMs and CAFs in the context of ionizing radiation are not fully understood. This study was undertaken to investigate whether M2 macrophages induce radioresistance in cervical cancer and to explore the TAMs' phenotypic transformation after IR and its underlying mechanisms. The radioresistance of cervical cancer cells was enhanced after being co-cultured with M2 macrophages. TAMs tended to undergo M2 polarization after high-dose irradiation, which was strongly associated with CAFs in both mouse models and patients with cervical cancer. Additionally, cytokine and chemokine analysis was performed to find that high-dose irradiated CAFs promoted macrophage polarization towards the M2 phenotype through chemokine (C-C motif) ligand 2. Collectively, our results highlight the crucial role that high-dose irradiated CAFs play in the regulation of M2 phenotype polarization, which ultimately induces radioresistance in cervical cancer. [ABSTRACT FROM AUTHOR]