Promoted Cell Viability Research Articles

MicroRNA miR-125b can suppress ovarian granulosa cell functions: Interrelationships with FSH.

This study aimed to evaluate the involvement of miR-125b and its interrelationship with follicle-stimulating hormone (FSH)in the control of basic ovarian granulosa cell functions. The effect of miR-125b mimics on basic functions of porcine ovarian granulosa cells cultured with and without FSH, and the effect of FSH on the expression of endogenous miR-125b was examined. Expression levels of miR-125b, viability, proliferation (accumulation of PCNA and cyclin B1), apoptosis (accumulation of bax and caspase 3), the accumulation of FSH receptors (FSHR), steroid hormones, insulin-like growth factor I (IGF-I), oxytocin, and prostaglandin E2 release were analysed by reverse transcription-quantitative polymerase chain reaction, Trypan blue exclusion test, quantitative immunocytochemistry, and ELISA. Transfection of cells with miR-125b mimics inhibited cell viability, proliferation, apoptosis, the occurrence of FSHR, progesterone, testosterone, estradiol, and oxytocin release but stimulated prostaglandin E2 output. FSH promoted cell viability, proliferation, steroid hormones, IGF-I, oxytocin, and prostaglandin E2 output and reduced the expression of miR-125b and apoptosis. Furthermore, miR-125b mimics supported the effect of FSH on the release of estradiol, IGF-I, and prostaglandin E2, and inverted FSH influence on cell viability, proliferation, apoptosis, progesterone, and testosterone output. FSH supported both inhibitory and stimulatory action of miR-125b on ovarian cell functions. Present observations indicate that: miR-125b can be involved in the control of basic ovarian functions and that miR-125b and FSH are antagonists in their actions on ovarian cell functions. The ability of FSH to reduce miR-125b expression and the ability of miR-125b mimics to decrease the occurrence of FSHR and to modify FSH effects indicate the existence of the self-inhibiting FSH-miR-125b axis and that miR-125b can mediate the actions of FSH on ovarian cells.

UHPLC-MS Chemical Fingerprinting and Antioxidant, Enzyme Inhibition, Anti-Inflammatory In Silico and Cytoprotective Activities of Cladonia chlorophaea and C. gracilis (Cladoniaceae) from Antarctica.

The lichen species Cladonia chlorophaea and C. gracilis (Cladoniaceae) are widely distributed in the island archipelago of maritime Antarctica and represent a natural resource of scientific interest. In this work, the metabolomic characterization of the ethanolic extracts of these species and the determination of the antioxidant activity, enzymatic inhibition and anti-inflammatory potential of selected compounds on the 5-lipoxygenase enzyme by molecular docking and cytoprotective activity in the SH-SY5Y cell line were carried out. Nineteen compounds were identified by liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry (UHPLC-ESI-QTOF-MS) in each of the species. The contents of phenolic compounds, antioxidant activity, the inhibition of cholinesterases (acetylcholinesterase and butyrylcholinesterase) and digestive enzymes (α-glucosidase and pancreatic lipase) were variable among species, with better results in C. chlorophaea. Molecular docking evidenced significant binding affinities of some compounds for the 5-lipoxygenase enzyme, together with outstanding pharmacokinetic properties. Both extracts were shown to promote cell viability and a reduction in reactive oxygen species production in an H2O2-induced oxidative stress model. This study contributes to the chemical knowledge of the Cladonia species and demonstrates the biological potential for the prevention and promising treatment of central nervous system pathologies, inflammatory disorders and metabolic alterations.

Bioactive Oligopeptides from Ginseng (Panax ginseng Meyer) Suppress Oxidative Stress-Induced Senescence in Fibroblasts via NAD+/SIRT1/PGC-1α Signaling Pathway.

The physicochemical properties and multiple bioactive effects of ginseng oligopeptides (GOPs), plant-derived small molecule bioactive peptides, suggest a positive influence on health span and longevity. Given this, cellular senescence is the initiating factor and key mechanism of aging in the organism, and thus the current study sought to explore the effects of GOPs on H2O2-induced cellular senescence and its potential mechanisms. Senescence was induced in mouse embryonic fibroblasts NIH/3T3 by 4 h of exposure to 200 µM H2O2 and confirmed using CCK-8 assay and Western blot analyses of p16INK4A and p21Waf1/Cip1 after 24 h of growth medium administration with or without GOPs supplementation (25, 50, and 100 µg/mL). We found that GOPs delayed oxidative stress-induced NIH/3T3 senescence by inhibiting the G1 phase arrest, increasing DNA synthesis in the S phase, decreasing the relative protein expression of p16INK4A and p21Waf1/Cip1, promoting cell viability, protecting DNA, and enhancing telomerase (TE) activity. Further investigation revealed that the increase in antioxidative capacity and anti-inflammation capacity might form the basis for the retarding of the senescence effects of GOPs. Furthermore, GOPs supplementation significantly improved mitochondrial function and mitochondrial biogenesis via the NAD+/SIRT1/PGC-1𝛼 pathway. These findings indicate that GOPs may have a positive effect on health span and lifespan extension via combating cellular senescence, oxidative stress, and inflammation, as well as modulating longevity regulating pathway NAD+/SIRT1/PGC-1𝛼.

SGLT2 inhibitor empagliflozin promotes revascularization in diabetic mouse hindlimb ischemia by inhibiting ferroptosis.

Gliflozins are known as SGLT2 inhibitors, which are used to treat diabetic patients by inhibiting glucose reabsorption in kidney proximal tubules. Recent studies show that gliflozins may exert other effects independent of SGLT2 pathways. In this study we investigated their effects on skeletal muscle cell viability and paracrine function, which were crucial for promoting revascularization in diabetic hindlimb ischemia (HLI). We showed that treatment with empagliflozin (0.1-40 μM) dose-dependently increased high glucose (25 mM)-impaired viability of skeletal muscle C2C12 cells. Canagliflozin, dapagliflozin, ertugliflozin, ipragliflozin and tofogliflozin exerted similar protective effects on skeletal muscle cells cultured under the hyperglycemic condition. Transcriptomic analysis revealed an enrichment of pathways related to ferroptosis in empagliflozin-treated C2C12 cells. We further demonstrated that empagliflozin and other gliflozins (10 μM) restored GPX4 expression in high glucose-treated C2C12 cells, thereby suppressing ferroptosis and promoting cell viability. Empagliflozin (10 μM) also markedly enhanced the proliferation and migration of blood vessel-forming cells by promoting paracrine function of skeletal muscle C2C12 cells. In diabetic HLI mice, injection of empagliflozin into the gastrocnemius muscle of the left hindlimb (10 mg/kg, every 3 days for 21 days) significantly enhanced revascularization and blood perfusion recovery. Collectively, these results reveal a novel effect of empagliflozin, a clinical hypoglycemic gliflozin drug, in inhibiting ferroptosis and enhancing skeletal muscle cell survival and paracrine function under hyperglycemic condition via restoring the expression of GPX4. This study highlights the potential of intramuscular injection of empagliflozin for treating diabetic HLI.

Knockdown of DAPK1 inhibits IL-1β-induced inflammation and cartilage degradation in human chondrocytes by modulating the PEDF-mediated NF-kB and NLRP3 inflammasome pathway.

Osteoarthritis (OA) is a common joint disease that is characterized by inflammation and cartilage degradation. Death-associated protein kinase 1 (DAPK1) is a multi-domain serine/threonine kinase and has been reported to be involved in the progression of OA. However, its role and mechanism in OA remain unclear. Here, we found the expression of DAPK1 in OA cartilage tissues was higher than that in normal cartilage tissues. The expression of DAPK1 in chondrocytes was up-regulated by IL-1β. Knockdown of DAPK1 promoted cell viability and anti-apoptotic protein expression, while it inhibited the apoptosis rate and pro-apoptotic protein expressions in IL-1β-induced chondrocytes. In addition, DAPK1 inhibition reduced the levels of inflammatory cytokines and expressions of matrix metalloproteinases (MMPs), and increased the expressions of collagen II and aggrecan. The data of mechanistic investigation indicated that the expression of pigment epithelium-derived factor (PEDF) was positively regulated by DAPK1. Overexpression of PEDF attenuated the effects of DAPK1 knockdown on IL-1β-induced cell viability, apoptosis, inflammation, and cartilage degradation. Furthermore, PEDF overexpression restored the activity of the NF-κB pathway and NLRP3 inflammasome after DAPK1 knockdown. Collectively, down-regulation of DAPK1 inhibited IL-1β-induced inflammation and cartilage degradation via the PEDF-mediated NF-κB and NLRP3 inflammasome pathways.

Knockdown of lncRNA XIST Ameliorates IL-1β-Induced Apoptosis of HUVECs and Change of Tissue Factor Level via miR-103a-3p/HMGB1 Axis in Deep Venous Thrombosis by Regulating the ROS/NF-κB Signaling Pathway.

The effect of lncRNA X inactive-specific transcript (XIST) inducing cardiovascular diseases on deep vein thrombosis (DVT) and its mechanism has not been reported. In this study, we uncovered the mystery that lncRNA XIST causes DVT with HUVEC dysfunction. The expression levels of lncRNA XIST and miR-103a-3p were detected by qRT-PCR, and HMGB1 expression was determined by qRT-PCR and western blot. The correlations among the expression levels of lncRNA XIST, miR-103a-3p, and HMGB1 were determined by Spearman's rank-order correlation test. XIST siRNA (si-XIST) was transfected into HUVECs to knock down the intrinsic expression of lncRNA XIST. The influences of si-XIST on interleukin-1 beta- (IL-1β-) treated HUVEC viability and apoptosis and the level of tissue factor (TF) were detected by MTT, flow cytometry, and ELISA kit, respectively. The relationships between lncRNA XIST, miR-103a-3p, and HMGB1 were predicted by the Encyclopedia of RNA Interactomes (ENCORI) database and verified by dual luciferase reporter assay. The effects of lncRNA XIST and miR-103a-3p on HMGB1 expression were detected by qRT-PCR, western blot, and immunofluorescence analysis. The levels of ROS/NF-κB pathway-related proteins were detected to study the regulatory mechanism of lncRNA XIST/miR-103a-3p/HMGB1 on IL-1β-treated HUVECs apoptosis and change of TF level. The upregulated expression levels of lncRNA XIST and HMGB1 and downregulated level of miR-103a-3p were found in the plasma of DVT patients and IL-1β-treated HUVECs. Si-XIST promoted cell viability and inhibited HUVEC apoptosis and ameliorated the change of TF level triggered by IL-1β. lncRNA XIST sponged miR-103a-3p and miR-103a-3p targeted HMGB1. Si-XIST inhibited the ROS/NF-κB pathway to suppress HUVEC apoptosis and ameliorate the change of TF level induced by IL-1β via the miR-103a-3p/HMGB1 axis. lncRNA XIST sponged miR-103a-3p improving HMGB1 expression to exacerbate DVT by activating the ROS/NF-κB signaling pathway. Our findings indicated that lncRNA XIST can be used as a potential therapeutic target in DVT.

Bioinformatics analysis of miRNAs in the neuroblastoma 11q-deleted region reveals a role of miR-548l in both 11q-deleted and MYCN amplified tumour cells

Neuroblastoma is a childhood tumour that is responsible for approximately 15% of all childhood cancer deaths. Neuroblastoma tumours with amplification of the oncogene MYCN are aggressive, however, another aggressive subgroup without MYCN amplification also exists; rather, they have a deleted region at chromosome arm 11q. Twenty-six miRNAs are located within the breakpoint region of chromosome 11q and have been checked for a possible involvement in development of neuroblastoma due to the genomic alteration. Target genes of these miRNAs are involved in pathways associated with cancer, including proliferation, apoptosis and DNA repair. We could show that miR-548l found within the 11q region is downregulated in neuroblastoma cell lines with 11q deletion or MYCN amplification. In addition, we showed that the restoration of miR-548l level in a neuroblastoma cell line led to a decreased proliferation of these cells as well as a decrease in the percentage of cells in the S phase. We also found that miR-548l overexpression suppressed cell viability and promoted apoptosis, while miR-548l knockdown promoted cell viability and inhibited apoptosis in neuroblastoma cells. Our results indicate that 11q-deleted neuroblastoma and MYCN amplified neuroblastoma coalesce by downregulating miR-548l.

Influence of Molecular Weight and Grafting Density of PEG on the Surface Properties of Polyurethanes and Their Effect on the Viability and Morphology of Fibroblasts and Osteoblasts.

Grafting polyethylene glycol (PEG) onto a polymer's surface is widely used to improve biocompatibility by reducing protein and cell adhesion. Although PEG is considered to be bioinert, its incorporation onto biomaterials has shown to improve cell viability depending on the amount and molecular weight (MW) used. This phenomenon was studied here by grafting PEG of three MW onto polyurethane (PU) substrata at three molar concentrations to assess their effect on PU surface properties and on the viability of osteoblasts and fibroblasts. PEG formed a covering on the substrata which increased the hydrophilicity and surface energy of PUs. Among the results, it was observed that osteoblast viability increased for all MW and grafting densities of PEG employed compared with unmodified PU. However, fibroblast viability only increased at certain combinations of MW and grafting densities of PEG, suggesting an optimal level of these parameters. PEG grafting also promoted a more spread cell morphology than that exhibited by unmodified PU; nevertheless, cells became apoptotic-like as PEG MW and grafting density were increased. These effects on cells could be due to PEG affecting culture medium pH, which became more alkaline at higher MW and concentrations of PEG. Results support the hypothesis that surface energy of PU substrates can be tuned by controlling the MW and grafting density of PEG, but these parameters should be optimized to promote cell viability without inducing apoptotic-like behavior.

The Therapeutic Effects of Seven Lycopodium Compounds on Cell Models of Alzheimer's Disease.

As an acetylcholinesterase inhibitor (AChEI), Huperzine-A (Hup-A) is marketed for treatment of mild to moderate Alzheimer's disease (AD) for decades in China. However, Hup-A causes some side effects. To search for new analogs or derivatives of Hup-A, we produced five Lycopodium alkaloids and two analogues by chemical synthesis: Lyconadins A-E, H-R-NOB, and 2JY-OBZ4. To systematically evaluate the therapeutic effects of the seven compounds on AD cell models. We assessed the effects of the seven compounds on cell viability via CCK-8 kit and used HEK293-hTau cells and N2a-hAPP cells as AD cell models to evaluate their potential therapeutic effects. We examined their effects on cholinesterase activity by employing the mice primary neuron. All compounds did not affect cell viability; in addition, Lyconadin A and 2JY-OBZ4 particularly increased cell viability. Lyconadin D and Lyconadin E restored tau phosphorylation at Thr231, and H-R-NOB and 2JY-OBZ4 restored tau phosphorylation at Thr231 and Ser396 in GSK-3β-transfected HEK293-hTau cells. 2JY-OBZ4 decreased the level of PP2Ac-pY307 and increased the level of PP2Ac-mL309, supporting that 2JY-OBZ4 may activate PP2A. Lyconadin B, Lyconadin D, Lyconadin E, H-R-NOB, and 2JY-OBZ4 increased sAβPPα level in N2a-hAPP cells. 2JY-OBZ4 decreased the levels of BACE1 and sAβPPβ, thereby reduced Aβ production. Seven compounds exhibited weaker AChE activity inhibition efficiency than Hup-A. Among them, 2JY-OBZ4 showed the strongest AChE inhibition activity with an inhibition rate of 17% at 10μM. Among the seven Lycopodium compounds, 2JY-OBZ4 showed the most expected effects on promoting cell viability, downregulating tau hyperphosphorylation, and Aβ production and inhibiting AChE in AD.

KLF2 reduces dexamethasone-induced injury to growth plate chondrocytes by inhibiting the Runx2-mediated PI3K/AKT and ERK signalling pathways

Dexamethasone (Dex) is a type of glucocorticoid drug. Long term use can induce growth plate chondrocytes (GPCs) apoptosis, impair differentiation, and inhibit cell proliferation and bone growth. It has been reported that Krüppel-like factor 2 (KLF2) inhibits osteoblast damage induced by Dex, but the role in Dex-induced GPCs remains unclear. Dex was used to construct a model of growth plate injury in vitro. CCK-8 and TUNEL kits were used to determine cell viability and apoptosis. A model of growth plate injury was established by intraperitoneal injection of Dex. Immunohistochemistry was used to investigate the expression of KLF2 in rats. The results showed that KLF2 expression of rat tibial GPCs was down-regulated after Dex stimulation. Overexpression of KLF2 promoted cell viability and cell cycle, while inhibited apoptosis of growth plate Dex-induced chondrocytes. Moreover, KLF2 inhibited Runx2-mediated PI3K/AKT and ERK signalling pathways. And PI3K/AKT and ERK signalling pathways, which were involved in the regulation of KLF2 on GPCs. Further studies showed that KLF2 alleviated growth plate injury in vivo. In conclusion, our study found that KLF2 promoted proliferation and inhibited apoptosis of Dex-induced GPCs by targeting the Runx2-mediated PI3K/AKT and ERK signalling pathways.

AhR regulates VEGF expression by promoting STAT1 transcriptional activity, thereby affecting endothelial angiogenesis in acute limb ischemia

BackgroundAngiogenesis has great potential in the treatment of acute limb ischemia (ALI). Here, we aimed to investigate the effect and mechanism of Aryl hydrocarbon receptor (AhR) on angiogenesis in ALI. MethodsThe ALI mouse model was constructed by femoral artery ligation, and the cell ischemia injury was induced by Hypoxia/serum deprivation. The laser doppler perfusion imaging was executed to detect the limb blood flow velocity. The tube formation assay was performed to evaluate angiogenesis. The cell viability was measured by 3-(45)-dimethylthiahiazo(-z-y1)-35-di-phenytetrazoliumromide. The cell migration was detected by wound healing assay. Hematoxylin-eosin, immunohistochemistry, immunofluorescence, dual-luciferase reporter gene assay, and Chromatin immunoprecipitation assay were conducted. ResultsIn ALI models, AhR expression was increased and translocated from cytoplasm to nucleus. Besides, necrosis and inflammatory infiltration were also increased in gastrocnemius tissues of model mice. In addition, AhR loss (LV-sh-AhR) promoted cell viability, angiogenesis, and migration, and also elevated the levels of vascular endothelial growth factor (VEGF), Tie2, and Ang2 in HUVEC models with Hypoxia/serum deprivation injury. Meanwhile, the interaction between AhR and signal transducer and activator of transcription 1 (STAT1), as well as STAT1 and VEGF, has also been confirmed. Co-transfection of LV-sh-AhR and LV-STAT1 suppressed cell viability, angiogenesis, and migration of injured HUVECs. Furthermore, injection of AAV2/9-shAhR in vivo also promoted angiogenesis, which was consistent with the in vitro experimental results. ConclusionsIn ALI models, activated AhR was translocated to the nucleus and down-regulated VEGF expression by promoting the transcriptional activity of STAT1, thereby inhibiting endothelial angiogenesis.

Structural characterization and immunomodulatory activity of an acidic heteropolysaccharide isolated from the fermented burdock residue mediated by Rhizopus nigricans

• An acidic polysaccharide (RBAPS) was gained from fermented burdock residue mediated by R. nigricans. • The structural characteristics of RBAPS were clarified. • RBAPS promoted cell viability and phagocytosis of macrophages. • RBAPS induced the macrophages polarization and pro-inflammatory cytokines secretion. An acidic heteropolysaccharide (RBAPS) with a relative molecular weight of 140.9 kDa was extracted from Rhizopus nigricans mediated burdock residue fermentation. Structural analysis indicated that RBAPS was mainly composed of Rha, Ara, Gal, GalA, GlcA, Glc and Xyl in a molar ratio of 14.5: 14.3: 30.1: 31.0: 5.5: 0.8: 3.8. The dominant backbone of RBAPS consisted of α-Rha p , α-GalA p , and α-Gal p with the branched chains of β-Gal p and α-Ara f . In macrophage RAW264.7 cells, RBAPS promoted proliferation, enhanced phagocytosis, hindered apoptosis, and promoted cytokines (NO, IL-6, TNF-α, and IL-1β) secretion. RBAPS also promoted M1 macrophage transformation and induced the secretion of NO and IL-6 in differentiated macrophages. The findings suggested that RBAPS has the potential immunomodulatory and anti-tumor effects, giving it application prospect in the health and pharmaceutical fields.

LncGBP9 knockdown alleviates myocardial inflammation and apoptosis in mice with acute viral myocarditis via suppressing NF-κB signaling pathway.

Myocardial inflammation and apoptosis are key processes in coxsackievirus B3 (CVB3)-induced acute viral myocarditis (AVMC). Accumulating evidence reveals the essential roles of long noncoding RNAs (lncRNAs) in the pathogenesis of AVMC. Here, we aimed to evaluate the biological functions of a novel lncRNA guanylate-binding protein 9 (lncGBP9) in AVMC progression and further explore its underlying mechanisms. Initially, mouse models of AVMC were constructed by CVB3 infection. The expression and localization of lncGBP9 in heart tissues were analyzed using RT-qPCR and FISH. Adeno-associated virus serotype 9 (AAV9)-mediated lncGBP9 knockdown was then employed to clarify its roles in survival, cardiac function, and myocardial inflammation and apoptosis. Moreover, the mRNA and protein levels of pro-inflammatory cytokines (TNF-α, IL-6, and IL-1β) were detected by RT-qPCR and ELISA, and the regulation of lncGBP9 knockdown on the NF-κB signaling pathway was investigated by Western blotting. Using an in vitro model of HL-1 cardiomyocytes exposed to CVB3 infection, the effects of lncGBP9 knockdown on cell viability, inflammation, and apoptosis were further evaluated in vitro. Increased lncGBP9 expression was detected in the heart tissues of AVMC mice and CVB3-infected HL-1 cells, and was mainly located in the cytoplasm. Knockdown of lncGBP9 remarkably alleviated the severity of AVMC in CVB3-infected mice, as verified by improved cardiac function, and reduced myocardial inflammation and apoptosis. Additionally, lncGBP9 knockdown suppressed the NF-κB signaling pathway and consequently reduced productions of pro-inflammatory cytokines in vivo. In vitro functional assays further confirmed that lncGBP9 knockdown promoted cell viability, inhibited cell apoptosis, and reduced pro-inflammatory cytokines release in CVB3-infected HL-1 cells through suppressing NF-κB activation. Collectively, lncGBP9 knockdown exerts anti-inflammatory and anti-apoptotic effects in CVB3-induced AVMC, which may be mediated in part via NF-κB signaling pathway. These findings highlight lncGBP9 as an attractive target for therapeutic interventions.

Trilayered Hydrogel Scaffold for Vocal Fold Tissue Engineering.

The lamina propria within the vocal fold (VF) is a complex multilayered tissue that increases in stiffness from the superficial to deep layer, where this characteristic is crucial for VF sound production. Tissue-engineered scaffolds designed for VF repair must mimic the biophysical nature of the native vocal fold and promote cell viability, cell spreading, and vibration with air flow. In this study, we present a unique trilayered, partially degradable hydrogel scaffold that mimics the multilayered structure of the VF lamina propria. Using thiol-norbornene photochemistry, trilayered hydrogel scaffolds were fabricated via layer-by-layer stacking with increasing polymer concentration from the top to middle to deep layer. Mechanical analysis confirmed that hydrogel modulus increased with increasing polymer concentration. Partially degradable hydrogels promoted high cell viability and cell spreading in three dimensions as assessed via live/dead and cytoskeleton staining, respectively. Importantly, partially degradable hydrogels maintained some degree of the three dimensional polymer network following protease exposure, while still enabling encapsulated cells to remodel their local environment via protease secretion. Finally, the trilayered hydrogel scaffold successfully vibrated and produced sound in proof-of-concept air flow studies. This work represents a critical first step toward the design of a multilayered, hydrogel scaffold for vocal fold tissue engineering.

Lncrna FGD5-AS1 Aggravates Myocardial Ischemia-Reperfusion Injury by Sponging Mir-129-5p

LncRNA FGD5-AS1 regulates the pathogenesis of many human diseases. We aimed to elucidate the function of lncRNA FGD5-AS1 and the regulatory mechanism of lncRNA FGD5-AS1/miR-129-5p in myocardial ischemia-reperfusion (I/R) injury. Myocardial I/R injury mice model and H/R treated H9c2 cells were established. RT-qPCR and Western blot analysis were used to detect the mRNA and protein expression. Cell viability was detected by MTT assay. Dual luciferase reporter assay was applied to confirm the relationship between lncRNA FGD5-AS1 and miR-129-5p. LncRNA FGD5-AS1 was upregulated in myocardial I/R injury mice models and H/R treated H9c2 cells. Functionally, knockdown of lncRNA FGD5-AS1 promoted cell viability and inhibited apoptosis in H/R treated H9c2 cells. In addition, lncRNA FGD5-AS1 directly targets miR-129-5p. Upregulation of lncRNA FGD5-AS1 weakened the protective effect of miR-129-5p on myocardial I/R injury. LncRNA FGD5-AS1 aggravates myocardial I/R injury by downregulating miR-129-5p.

Comprehensive analysis of lncRNA-mediated ceRNA networkfor hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a high-burden cancer. The molecular mechanism of HCC has not been fully elucidated. Notably, current research has revealed a significant function for long non-coding RNAs (lncRNAs) in the prognosis of patients with HCC. Here, this study aims to construct a regulated lncRNA-mediated ceRNA network and find biological targets for the treatment of HCC. Based on the RNA expression patterns from the TCGA, we did an analysis to determine which genes were expressed differently between liver tumor tissues and noncancerous tissues. Then, using bioinformatic tools, we built a lncRNA-miRNA-mRNA ceRNA network and used GO and KEGG functional analyses on the DEmRNAs connected to ceRNA networks. The main lncRNAs in the subnetwork were chosen, and we next looked at the relationships between these lncRNAs and the clinical characteristics of patients with HCC. The prognosis-related genes and immune cells were identified using Kaplan-Meier and Cox proportional hazard analyses, and CIBERSORT was utilized to separate the 22 immune cell types. CCK8 assay was performed to measure cell viability in HCC cells after lncRNA HOTTIP modulation. Differentially expressed mRNA and lncRNAs in HCC and paracancerous tissues were identified. There are 245 lncRNAs, 126 miRNAs, and 1980 mRNAs that are expressed differently in liver tumour tissues than in noncancerous cells. Function analysis showed that mRNAs in ceRNA network were significantly enriched in G1/S transition of mototiv cell cycle, positive regulation of cell cycle process, hepatocellular carcinoma, and cancer related pathways. CD8 T cells and T follicular helper cells had a favourable link with a 0.65 correlation coefficient. Additionally, there was a strong correlation between Eosinophils, activated NK cells, and B memory cells. Strikingly, depletion of lncRNA HOTTIP inhibited viability of HCC cells. In addition, miR-205 upregulation suppressed viability of HCC cells, while miR-205 downregulation repressed viability of HCC cells. Notably, miR-205 depletion rescued HOTTIP depletion-mediated suppression of cell viability in HCC. A ceRNA network was created by examining the lncRNA, miRNA, and mRNA expression profiles of liver tumours from the TCGA database. LncRNA HOTTIP promoted cell viability via inhibition of miR-205 in HCC cells.

Alternative splicing of apoptosis genes promotes human T cell survival.

Alternative splicing occurs in the vast majority of human genes, giving rise to distinct mRNA and protein isoforms. We, and others, have previously identified hundreds of genes that change their isoform expression upon T cell activation via alternative splicing; however, how these changes link activation input with functional output remains largely unknown. Here, we investigate how costimulation of T cells through the CD28 receptor impacts alternative splicing in T cells activated through the T cell receptor (TCR, CD3) and find that while CD28 signaling alone has minimal impact on splicing, it enhances the extent of change for up to 20% of TCR-induced alternative splicing events. Interestingly, a set of CD28-enhanced splicing events occur within genes encoding key components of the apoptotic signaling pathway; namely caspase-9, Bax, and Bim. Using both CRISPR-edited cells and antisense oligos to force expression of specific isoforms, we show for all three of these genes that the isoform induced by CD3/CD28 costimulation promotes resistance to apoptosis, and that changes in all three genes together function combinatorially to further promote cell viability. Finally, we show that the JNK signaling pathway, induced downstream of CD3/CD28 costimulation, is required for each of these splicing events, further highlighting their co-regulation. Together, these findings demonstrate that alternative splicing is a key mechanism by which costimulation of CD28 promotes viability of activated T cells.

Evaluation of Human Osteoblasts on NIPS Micro-Patterned PCL Carriers Containing Nanohydroxyapatite and Reduced Graphene Oxide Using PSµM.

The content and surface topology of tissue engineering scaffolds are two important parameters in regulating the cell behavior. In this study, a phase separation micromolding (PSµM) method was implemented to develop micro-groove-imprinted poly(ε-caprolactone) (PCL)–nano hydroxyapatite (nHAp)–reduced graphene oxide (rGO) ternary blend constructs. Physical and chemical characterizations of cell-devoid constructs were performed by FTIR, XRD, TGA, DSC, porosity, swelling, wettability analysis, tensile and compression mechanical tests. The in vitro biological performance of human osteoblasts cultured on micro-patterned blend constructs was evaluated by MTT and alamarBlue viability assays. The findings revealed that nHAp and rGO significantly promote cell viability and proliferation, while the micro-pattern determines the direction of cell migration. Alkaline phosphatase and Ca2+ analyses were carried out to determine the osteogenic properties of cell-laden constructs. This study describes a simple method to generate topologically modified ternary blend PCL/nHAp/rGO constructs using the PSµM method, which contributes to cell proliferation and migration, which is particularly important in regenerative medicine.

PBX1 attenuates H2O2-induced oxidant stress in human trabecular meshwork cells via promoting NANOG-mediated PI3K/AKT signaling pathway.

Oxidative stress-induced excessive extracellular matrix (ECM) deposition in trabecular meshwork (TM) tissue is considered the major pathological procedure of glaucoma. This study aimed to explore the role and regulatory mechanism of pre-B-cell leukemia transcription factor 1 (PBX1) in H2O2-induced human trabecular meshwork cells (HTMCs). Expressions of PBX1, NANOG, ECM, and pathway-related factors were detected by qRT-PCR and western blot. Cell viability and apoptosis of HTMCs were measured using CCK-8 and flow cytometry assays. Reactive oxygen species (ROS), superoxide dismutase (SOD), and L-glutathione (GSH) levels were detected to evaluate oxidative stress. Through luciferase reporter assay, the association between PBX1 and NANOG was verified. Results presented that PBX1 was significantly upregulated in H2O2-induced HTMCs. Functionally, PBX1 and NANOG promoted cell viability, inhibited cell apoptosis and ECM deposition, suppressed ROS accumulation, and enhanced the productions of SOD and GSH in H2O2-stimulated HTMCs, while PBX1 inhibition showed the opposite effects. In addition, PBX1 promoted the transcription of NANOG by upregulating the promoter activity of NANOG which activated the PI3K-AKT signaling pathway. What's more, the inhibitions of PI3K-AKT signaling pathway or NANOG reversed the protective effect of PBX1 on H2O2-stimulated HTMCs. In summary, our study firstly revealed that PBX1 attenuated the oxidative damage in HTMCs via regulating NANOG-mediated PI3K/AKT signaling, suggesting that PBX1 might be a potential treatment target for glaucoma patients.

Overexpression of miR-383-3p protects cardiomyocytes against hypoxia/reoxygenation injury via regulating PTEN/PI3K/AKT signal pathway.

MicroRNAs are widely reported as biomarkers and therapeutic targets in cardiovascular diseases. This study is aimed to expound on the regulatory responsibility of miR-383-3p in H/R-induced injury of H9c2 cells. In this study, H9c2 cells were administrated with H/R. MiR-383-3p expression was measured using qRT-PCR. ELISA was used to determine lactate dehydrogenase (LDH), superoxide dismutase (SOD), and malondialdehyde (MDA) levels. Reactive oxygen species (ROS) were detected with 2,7-Dichlorodihydrofluorescein diacetate probe. 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di- phenytetrazoliumromide, flow cytometry, and TUNEL experiments were conducted to measure cell viability and apoptosis. Cleaved caspase-3, caspase-3, Bax, Bcl-2, PTEN, PI3K, p-PI3K, Akt, p-AKT expression levels were examined by Western blot. Cleaved caspase-3 expression was also measured by immunofluorescence staining. Dual-luciferase reporter gene assay was applied to validate the binding sites in miR-383-3p and the 3'UTR of PTEN. We reported that, miR-383-3p expression in H9c2 cells treated with H/R was remarkably decreased. MiR-383-3p overexpression ameliorated oxidative stress and apoptosis and promoted cell viability in H9c2 cells treated with H/R, while miR-383-3p inhibitor showed the reverse effects. PTEN was identified as a target gene of miR-383-3p. Additionally, enhancement of PTEN expression abolished the influences of miR-383-3p on H9c2 cells. MiR-383-3p mimics could significantly decrease PTEN expression in H9c2 cells while increasing p-PI3K expression and p-AKT expression, while the miR-383-3p inhibitors showed the opposed effects. In conclusion, miR-383-3p protected H9c2 cells from H/R-induced injury via regulating PTEN/PI3K/AKT signal pathway.