Injury In PC12 Cells Research Articles

Expression of Concern regarding Lei Fan, Huifeng Zhang, Xiaobin Li, Guang Yang, Jingtao Ru, Tao Liu. Emodin protects hyperglycemia-induced injury in PC-12 cells by up-regulation of miR-9. Molecular and Cellular Endocrinology. 2018; 474. DOI: 10.1016/j.mce.2018.03.009

Expression of Concern regarding Lei Fan, Huifeng Zhang, Xiaobin Li, Guang Yang, Jingtao Ru, Tao Liu. Emodin protects hyperglycemia-induced injury in PC-12 cells by up-regulation of miR-9. Molecular and Cellular Endocrinology. 2018; 474. DOI: 10.1016/j.mce.2018.03.009

Overexpression of miR-338-5p in exosomes derived from mesenchymal stromal cells provides neuroprotective effects by the Cnr1/Rap1/Akt pathway after spinal cord injury in rats

Growing evidence has shown that microRNAs (miRNAs) play crucial roles in the physiopathology of spinal cord injury (SCI). Recent studies have confirmed that miR-338-5p regulates myelination, suggesting a potential role in the treatment of SCI. However, the molecular mechanism of miR-338-5p on SCI is still unknown. Recently, exosomes have emerged as an ideal vector to deliver therapeutic molecules such as miRNAs. Here, we explored the effects of miR-338-5p-overexpressing exosomes derived from bone marrow-derived mesenchymal stromal cells (BMSCs) on SCI. In vivo, a model of contusion SCI in rats was established, and we observed that overexpression of miR-338-5p in exosomes profoundly increased the expression levels of neurofilament protein-M and growth-associated protein-43 and decreased those of myelin-associated glycoprotein and glial fibrillary acidic protein, which provided neuroprotective effects after acute SCI. In an in vitro study, we found that overexpression of miR-338-5p in exosomes repressed cell apoptosis following H2O2-induced oxidative stress injury in PC12 cells. Additionally, we confirmed that cannabinoid receptor 1 (Cnr1) was the target gene of miR-338-5p by dual-luciferase reporter assays and that Rap1 was the downstream gene by the KEGG pathway analysis. We found that miR-338-5p increased cAMP accumulation as a consequence of downregulated expression of the target gene Cnr1, and then, Rap1 was activated by cAMP. Eventually, the activation of the PI3K/Akt pathway attenuated cell apoptosis and promoted neuronal survival by cAMP-mediated Rap1 activation. In brief, these findings showed that exosomes overexpressing miR-338-5p were a promising treatment strategy for SCI.

Polycyclic polyprenylated acylphloroglucinols with immunosuppressive activity from Hypericum perforatum and absolute configurations assignment of previously reported analogues

Hyperformitins A–I (1–9), nine undescribed polycyclic polyprenylated acylphloroglucinols (PPAPs) with double-bond migration, along with four new isomers hyperformitins J–M (10–13), were isolated from Hypericum perforatum. Their structures and absolute configurations were determined by spectroscopic analyses including HRESIMS, IR, UV, NMR, and ECD, as well as optical rotation (OR) calculations. The absolute configurations of previously reported analogues, garsubellins D and C as well as garcinielliptones L and M, were assigned for the first time by NMR spectra and specific rotations analyses assisting with OR calculations. Selected compounds were tested for their immunosuppressive activities against lipopolysaccharide (LPS)-induced B lymphocyte proliferation. Compounds 1, 3, 4, 5, 7, and 11 showed inhibition activities against the proliferation of B lymphocyte with IC50 values ranging from 4.1 to 9.7 μM. Furthermore, the neuroprotective activities of the isolates against corticosterone (CORT)-induced injury in PC12 cells were also tested, and compounds 1, 12, and 13 exhibited neuroprotective effects with cell viabilities of 68.0%, 71.3%, and 68.4%, respectively under the concentration of 10 μM.

Production of bioactive recombinant human fibroblast growth factor 12 using a new transient expression vector in E. coli and its neuroprotective effects.

In recent years, an increasing number of studies have shown that fibroblast growth factor 12 (FGF12) plays important roles in regulating neural development and function. Importantly, changes of FGF12 expression are thought to be related to the pathophysiology of many neurological diseases. However, little research has been performed to explore the protective effect of FGF12 on nerve damage. This study aims to explore its neuroprotective effects using our recombinant humanized FGF12 (rhFGF12). The hFGF12 gene was cloned and ligated into an expression vector to construct a recombinant plasmid pET-3a-hFGF12. Single colonies were screened to obtain high expression engineering strains, and fermentation and purification protocols for rhFGF12 were designed and optimized. The biological activities and related mechanisms of rhFGF12 were investigated by MTT assay using NIH3T3 and PC12 cell lines. The in vitro neurotoxicity model of H2O2-induced oxidative injury in PC12 cells was established to explore the protective effects of rhFGF12. The results indicate that the beneficial effects of rhFGF12 were most likely achieved by promoting cell proliferation and reducing apoptosis. Moreover, a transgenic zebrafish (islet) with strong GFP fluorescence in the motor neurons of the hindbrain was used to establish a central injury model caused by mycophenolate mofetil (MMF). The results suggested that rhFGF12 could ameliorate central injury induced by MMF in zebrafish. In conclusion, we have established an efficient method to express and purify active rhFGF12 using an Escherichia coli expression system. Besides, rhFGF12 plays a protective effect of on nerve damage, and it provides a promising therapeutic approach for nerve injury. KEY POINTS: • Effective expression and purification of bioactive rhFGF12 protein in E. coli. • ERK/MAPK pathway is involved in rhFGF12-stimulated proliferation on PC12 cells. • The rhFGF12 has the neuroprotective effects by inhibiting apoptosis.

Neuroprotective effects of Tiaogeng decoction against H2O2-induced oxidative injury and apoptosis in PC12 cells via Nrf2 and JNK signaling pathways

Ethnopharmacological relevanceTiaogeng decoction (TGD), a mixture of 10 traditional Chinese herbs, has been used clinically for over 30 years in treating menopause-related symptoms such as cognitive changes, mood disorders, vasomotor symptoms, and sleep disorders. These central nervous system symptoms are closely associated with declined ovarian function, which dramatically increases the risk of neurodegenerative disease. Previous studies revealed that TGD may have anti-oxidative and anti-apoptotic properties, potentially preventing neurodegenerative conditions; however, the underlying pharmacological mechanism remains unclear. Aim of the studyThis study aimed to examine whether TGD could activate the Nrf2 and C-Jun N-terminal kinase (JNK) signaling pathways to effectively reduce oxidative injury and apoptosis in PC12 cells and elucidate the mechanism by which this medicine may prevent neurodegenerative disease. Materials and methodsPC12 cells were exposed to different concentrations of TGD (125, 250, 500 μg/mL) and H2O2 (150 μM). 17β-estradiol (0.05 μg/mL) was used as the positive control. A cell counting kit-8 (CCK-8) and a lactate dehydrogenase (LDH) assay were used to detect cell viability and cytotoxicity, while Hoechst and flow cytometry were performed to evaluate apoptosis levels. Mitochondrial function was assessed by measuring mitochondrial membrane potential (MMP), and superoxide dismutase (SOD), and reactive oxygen species (ROS) levels were used to measure oxidative stress (OS). Western blot analysis was used to identify the levels of Nrf2, phospho-JNK (p-JNK), phospho-mitogen-activated protein kinase kinase 7 (p-MKK7), Kelch-like ECH-associated protein 1 (Keap1), heme oxygenase-1 (HO-1), Caspase3 (Casp3), Caspase9 (Casp9), Bax, and Bcl-2 proteins. Moreover, JNK agonist anisomycin and Nrf2 inhibitor ML385 were used to validate pathways. ResultsTGD pretreatment significantly alleviated H2O2-induced cytotoxicity, apoptosis, MMP, and OS levels. H2O2 stimulated the activation of Nrf2 and JNK signaling pathways, but TGD increased the extent of Nrf2 antioxidant activation, decreased the activation of JNK, and eventually reversed the H2O2-induced protein expression of p-MKK7, Keap1, HO-1, Cleaved Caspase3 (CL-Casp3), Cleaved Caspase9 (CL-Casp9), Bax, and Bcl-2. ConclusionsThis study's findings suggest that TGD may attenuate oxidative injury and apoptosis via the Nrf2 and JNK signaling pathways, making it a potential therapeutic candidate for neurodegenerative diseases.

Protective effects of the knockdown of lncRNA AK139328 against oxygen glucose deprivation/reoxygenation-induced injury in PC12 cells.

Cerebral ischemic stroke is a major cause of adult morbidity and mortality worldwide. Several long non-coding RNAs (lncRNAs) have been reported to participate in cerebral ischemia/reperfusion injury (IRI). However, to the best of our knowledge, the role of lncRNA AK139328 in cerebral ischemic stroke remains poorly understood. The present study aimed to determine the expression and function of lncRNA AK139328 in the progression of IRI. PC12 cells were injured by oxygen glucose deprivation/reoxygenation (OGD/R) to establish an in vitro ischemic stroke model. An MTT assay was performed to determine cell viability. Reverse transcription-quantitative PCR was used to analyze the expression levels of AK139328 and Netrin-1 in blood samples from patients who had suffered a cerebral ischemic stroke and healthy individuals or OGD/R PC12 cells. ELISAs were used to determine the levels of inflammatory cytokines. In addition, oxidative stress levels and the levels of cell apoptosis were evaluated by reactive oxygen species (ROS) kits, flow cytometry and western blotting. Immunofluorescence staining was used for the detection of cell neurite outgrowth. The results of the present study revealed that AK139328 expression levels were upregulated in patients who had suffered a cerebral ischemic stroke and in PC12 cells following stimulation with OGD/R. The knockdown of AK139328 alleviated OGD/R-induced decreases in cell viability, downregulation in Netrin-1 expression and increases in inflammatory cytokines levels, including TNF-α, IL-1β and IL-6. Moreover, AK139328 silencing suppressed oxidative stress and cell apoptosis in OGD/R-treated PC12 cells. Furthermore, the expression levels of microtubule associated protein 2 and growth associated protein 43 in OGD/R-injured PC12 cells were upregulated following the knockdown of AK139328 expression. In conclusion, these findings suggested that the knockdown of AK139328 expression may protect PC12 cells against OGD/R injury by regulating inflammatory responses, oxidative stress and cell apoptosis. The data suggested a potential therapeutic target for the diagnosis and treatment of cerebral ischemic stroke.

Phthalimide-(N-alkylbenzylamine) cysteamide hybrids as multifunctional agents against Alzheimer's disease: Design, synthesis, and biological evaluation.

The complex pathogenesis of Alzheimer's disease (AD) calls for multi-target approach for disease treatment. Herein, based on the MTDLs strategy, a series of phthalimide-(N-alkylbenzylamine) cysteamide hybrids were designed, synthesized, and investigated in vitro for the purpose. Most of the target compounds were found to be potential multi-target agents. In vitro results showed that compound 9e was the representative compound in this series, endowed with high EeAChE and HuAChE inhibitory potency (IC50 =1.55µm and 2.23µm, respectively), good inhibitory activity against self-induced Aβ1-42 aggregation (36.08% at 25µm), and moderate antioxidant capacity (ORAC-FL value was 0.68 Trolox equivalents). Molecular docking studies rationalized the binding mode of 9e in both PAS and CAS of AChE. Moreover, 9e displayed excellent ability to against H2 O2 -induced PC12 cell injury and penetrate BBB. Overall, these results highlighted that compound 9e was an effective and promising multi-target agent for further anti-AD drug development.

Sesquiterpenoids and 2-(2-Phenylethyl)chromone Derivatives from the Resinous Heartwood of Aquilaria sinensis

One novel spirolactone, aquilarisinolide (1), three new sesquiterpenoids, (2R,4S,5R,7R)-2-hydroxyeremophila-9,11-dien-8-one (2), (1R,4S,5S,7R,11R)-13-hydroxyepidaphnauran-9-en-8-one (3), and (4R,5S,7R,8S,10S,13R)-8,13-dihydroxyrotunda-1,11-dien-3-one (4), together with 13 known compounds (5–17) were isolated from the resinous heartwood of Aquilaria sinensis (Thymelaeaceae). The structures of the new compounds were elucidated based on the analysis of NMR and MS data and theoretical calculations their ECD spectra. The isolated compounds were evaluated for their protective activities against PC12 cell injury induced by corticosterone (CORT) and 1-methyl-4-phenylpyridine ion (MPP+), as well as inhibitory activities against BACE1. Compound 4, 5,6-dihydroxy-2-(2-phenylethyl)chromone (5), daphnauranol B (7), 6-methoxy-2-[2-(3-methyoxyphenyl)ethyl]chromone (10), isoagarotetrol (14), and 1-hydroxy-1,5-diphenylpentan-3-one (16) showed significant protective effects on CORT-induced injury in PC12 cells at a concentration of 20 μM (P < 0.001). Isoagarotetrol (14) showed a significant protective effect on MPP+-induced injury in PC12 cells at a concentration of 20 μM (P < 0.001), while compound 4 showed a moderate activity (P < 0.01). The BACE1-inhibitory activities of all tested compounds were very weak with less than 30% inhibition at a concentration of 20 μM.Graphic

A method for the expression of fibroblast growth factor 14 and assessment of its neuroprotective effect in an Alzheimer's disease model.

BackgroundFibroblast growth factor (FGF) 14 is a member of the FGF family that is mainly expressed in the central nervous system. FGF14 has a close association with the occurrence of neurodegenerative conditions; however, its significance in Alzheimer’s disease (AD) has yet to be evaluated. Therefore, we sought to obtain a large amount of exogenous FGF14 protein and explore its effect in a cellular model of AD.MethodsFGF14 protein was expressed in an Escherichia coli system using gene recombination technology. Purified protein was obtained through washing and renaturation of inclusion bodies combined with nickel column affinity chromatography. The AD model was established via Aβ25-35-induced injury in PC12 cells. Changes in the levels of lactate dehydrogenase and malondialdehyde were detected, and the neuroprotective effect of recombinant human FGF14 (rhFGF14) was evaluated through double-fluorescence staining and flow cytometry apoptosis detection. For further exploration of rhFGF14-mediated regulation of mitogen-activated protein kinase (MAPK) signaling, western blot was employed.ResultsWe successfully induced large amounts of insoluble rhFGF14. Following solubilization and refolding of the rhFGF14 from inclusion bodies, high purity rhFGF14 was purified by Nickel affinity column chromatography. The results showed that rhFGF14 alleviated Aβ25-3-induced PC12 cell injury by inhibiting the phosphorylation of p38, extracellular signal-regulated kinase 1/2, and c-Jun N-terminal kinase, thus suppressing the MAPK signaling pathway.ConclusionsFGF14 performed a neuroprotective role in our in vitro AD model via its inhibition of MAPK signaling, highlighting its potential as a therapeutic drug for neurodegenerative conditions.

Knockdown of XIST Attenuates Cerebral Ischemia/Reperfusion Injury Through Regulation of miR-362/ROCK2 Axis.

Long non-coding RNAs (lncRNAs) are considered as critical regulators in the pathogenesis of cerebral ischemia. In this present study, we aimed to investigate the impact and underlying mechanism of lncRNA X-inactive specific transcript (XIST) in cerebral ischemia/reperfusion (I/R) injury. An oxygen-glucose deprivation/reperfusion (OGD/R) model in PC12 cells was applied to mimic cerebral I/R injury in vitro and middle cerebral artery occlusion/reperfusion (MCAO/R) model was performed in mice to mimic cerebral I/R injury in vivo. Real-time PCR, fluorescence in situ hybridization (FISH) assay, and western blotting assay were carried out to detect the expression levels of XIST, miR-362, and Rho-related coiled-coil containing protein kinase 2 (ROCK2). The functional experiments were measured by CCK-8 assay, immumofluorescence assay, ELISA assay, TUNEL, and TTC staining. Results displayed that XIST was elevated in PC12 cells with OGD/R, as well as in the ischemic penumbra of mice with MCAO/R. In vitro, knockdown of XIST facilitated cell survival, inhibited apoptosis, and alleviated inflammation injury in OGDR PC12 cells. In vivo, inhibition of XIST remarkably reduced the neurological impairments, promoted neuron proliferation, and suppressed apoptosis in MCAO mice. Mechanistically, XIST acted as a competing endogenous RNA of miR-362 to regulate the downstream gene ROCK2. In conclusion, depletion of XIST attenuated I/R-induced neurological impairment and inflammatory response via the miR-362/ROCK2 axis. These findings offer a potential novel strategy for ischemic stroke therapy.

Separation and purification of magnoflorine, spinosin, and 6‴-feruloyspinosin from Ziziphi Spinosae Semen by high-speed counter-current chromatography.

In the study, high-speed counter-current chromatography was used for separation and purification of magnoflorine, spinosin, and 6‴-feruloyspinosin from Ziziphi Spinosae Semen. With n-butanol-ethyl acetate-water (2:3:5, v/v) as the optimum solvent system, about 75mg of magnoflorine, 110mg of spinosin, and 40mg of 6‴-feruloyspinosin were isolated from 0.5g of crude extract of Z. Spinosae Semen, with the purity of 95.7, 97.2, and 96.4%, respectively. The chemical structures were identified by MS and NMR spectroscopy. In addition, the antidepressant activity of the isolated components was evaluated by PC12 cells injury model and chronic unpredictable mild stress depression mouse model. The results showed that high-speed counter-current chromatography could be used to realize the one-time rapid preparation and separation of magnoflorine, spinosin, and 6‴-feruloyspinosin from Z. Spinosae Semen and compatibility of these isolated components has certain antidepressant activity.

Protective effect of 1, 25(OH)_2D_3 on Aβ_(1-42)-induced pyrolysis in PC12 cells

To investigate the protective effect of 1, 25(OH)_2D_3 on Aβ_(1-42)-induced pyrolysis in PC12 cells. The Alzheimer& apos; s disease model in PC12 cells was established with 20 μmol/L Aβ_(1-42). The experiment was divided into control group, model group(20 μmol/L Aβ_(1-42)) and 1, 25(OH)_2D_3 groups(1, 10, 100 nmol/L 1, 25(OH)_2D_3+20 μmol/L Aβ_(1-42)). Cell activity was detected by CCK-8, cell membrane permeability was detected by AO/EB staining, lactic dehydrogenase(LDH)and interleukin-1β(IL-1β)were detected by colorimetry and ELISA, NOD-like receptor family protein 1(NLRP1), cysteinyl aspartate specific proteinase-1(caspase-1)and gasdermin D(GSDMD)protein expression were detected by Western Blot. Compared with the control group, the cell activitywas significantly decreased(P& lt; 0. 01), cell membrane permeability, the level of LDH and IL-1β, and the expression of NLRP1, caspase-1 and GSDMD were significantly increased(P& lt; 0. 01). Compared with the model group, the cell activity was significantly increased(P& lt; 0. 01), cell membrane damage was decreased in PC12 cells exposed to 1, 25(OH)_2D_3. The level of LDH and IL-1β were significantly decreased(P& lt; 0. 01) in PC12 cells exposed to 10 and 100 nmol/L 1, 25(OH)_2D_3. The expression of NLRP1 and GSDMD in 1 nmol/L 1, 25(OH)_2D_3 group was decreased(P& lt; 0. 05), and the decrease was more significant in 10 and 100 nmol/L 1, 25(OH)_2D_3 groups(P& lt; 0. 01). The expression of caspase-1 was significantly decreased in 10 and 100 nmol/L 1, 25(OH)_2D_3 groups(P& lt; 0. 05, P& lt; 0. 01). 1, 25(OH)_2D_3 exerts a significant protective effect against Aβ_(1-42)-induced PC12 cells injury through inhibition of neuronal pyrolysis.

Paeoniflorin protects PC12 cells from oxygen-glucose deprivation/reoxygenation-induced injury via activating JAK2/STAT3 signaling.

Ischemic stroke is the most common type of stroke, and it has become a major health issue as it is characterized by high mortality and morbidity rates. Paeoniflorin (PF) is a natural compound and the main active ingredient of Radix Paeoniae. The aim of the present study was to investigate the role of PF in oxygen-glucose deprivation/reoxygenation (OGD/R)-induced injury of PC12 cells and its association with the Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) pathway. An in vitro model of OGD/R injury was established in PC12 cells. Subsequently, Cell Counting Kit-8 assay and ELISA were used to evaluate cell viability and the secretion of inflammatory factors, respectively, in PC12 cells subjected to OGD/R and treated with PF. The levels of oxidative stress indicators and inflammatory factors were measured using corresponding commercial kits. In addition, the apoptosis rate of PC12 cells subjected to OGD/R and treated with PF was determined by flow cytometry, and the expression of apoptosis-related proteins was analyzed by western blotting. Additionally, the expression levels of JAK2/STAT3 pathway-related proteins were also evaluated. The cell viability, levels of oxidative stress, inflammation and apoptosis were also measured in OGD/R-induced PC12 cell injury models following co-treatment of cells with PF and FLLL32, a specific inhibitor of JAK2/STAT3 signaling. Cell viability was reduced, while oxidative stress and inflammation were increased after OGD/R-induced injury. However, the treatment of cells with PF significantly enhanced cell viability, and alleviated oxidative stress, inflammation and apoptosis of OGD/R-treated PC12 cells. Furthermore, PF activated the JAK2/STAT3 signaling pathway. Following FLLL32 intervention, the effects of PF on oxidative stress, inflammation and apoptosis of OGD/R-treated PC12 cells were reversed. In conclusion, the findings of the present study suggested that PF may protect PC12 cells from OGD/R-induced injury via activating the JAK2/STAT3 signaling pathway, thus providing novel insight into the mechanism through which PF may alleviate ischemic stroke and indicating a potential strategy for ischemic stroke treatment.

Knockdown of lncRNA TTTY15 alleviates ischemia/reperfusion-induced inflammation and apoptosis of PC12 cells by targeting miR-766-5p.

The pathogenesis of ischemic stroke is extremely complex and has a significant impact on the quality of life of the patients. Accumulating studies have reported that long non-coding RNAs (lncRNAs) may be associated with the progression of ischemic stroke. However, the role and underlying mechanism of action of the lncRNA testis-specific transcript Y-linked 15 (TTTY15) in ischemic stroke remains unknown. The present study analyzed the expression levels of TTTY15 in PC12 cells injured by oxygen-glucose deprivation/reperfusion (OGD/R). The effects of the knockdown of TTTY15 expression on the levels of the inflammatory cytokines TNF-α, IL-1β, IL-18 and IL-10, cell apoptosis and the expression levels of the apoptosis-associated proteins Bcl-2, Bax, cleaved caspase-3, caspase-3, cleaved caspase-9 and caspase-9, were subsequently analyzed in OGD/R-treated PC12 cells using ELISA, flow cytometry and western blotting, respectively. In addition, the downstream target gene of TTTY15 was verified using a dual luciferase reporter assay. The effects of TTTY15 on the inflammation and apoptosis of PC12 cells treated with OGD/R were determined by targeting miR-766-5p. The results of the present study revealed that TTTY15 expression was upregulated in OGD/R-treated PC12 cells. The knockdown of TTTY15 significantly decreased the concentrations of the proinflammatory factors TNF-α, IL-1β and IL-18, while it increased the concentration of the anti-inflammatory cytokine IL-10 in OGD/R-treated PC12 cells. Apoptosis was also suppressed following gene silencing of TTTY15. Subsequently, miR-766-5p was identified as a target gene of TTTY15 using a dual luciferase reporter assay and the expression levels of TTTY15 and miR-766-5p were found to be negatively correlated. The overexpression of miR-766-5p alleviated the stimulatory effect of TTTY15 overexpression on the inflammation and apoptosis of PC12 cells treated with OGD/R. Therefore, the present study revealed that TTTY15 knockdown improved the OGD/R-induced injury of PC12 cells by upregulating miR-766-5p expression.

LncRNA-MIAT promotes neural cell autophagy and apoptosis in ischemic stroke by up-regulating REDD1

BackgroundIschemic stroke (IS) accounts for 80% of stroke incidence, which has an impact on the life quality of patients. Long non-coding RNA (LncRNA), a class of non-coding transcripts greater than 200 nucleotidesin length, has been extensively studied in cerebrovascular diseases. Myocardial infarction associated transcript (MIAT) is highly expressed in nervous system. Therefore this study aims to explore the role of LncRNA MIAT in IS and to clarify its underlying mechanism, providing therapeutic value for the treatment of IS. MethodsThe neurological function of rats was evaluated by neurological deficit score. Triphenyltetrazolium chloride (TTC) staining was used to detect infarct area in brain tissues. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to examine the expression of MIAT. Western blotting was used to detect the expressions of REDD1, p-mTOR, autophagy-related proteins LC3 and p62, and apoptotic-related proteins Bax, cleaved-caspase3, Bcl-2. Flow cytometry was applied to examine neuronal cell apoptosis. RNA pull-down and RIP assay was used to verify the binding of MIAT and REDD1. The level of REDD1 ubiquitination was detected by ubiquitination and Co-immunoprecipitation (Co-IP) assay. ResultsThe expressions of MIAT and REDD1 were increased in IS rats and oxygen-glucose deprivation/reoxygenation (OGD/R)-induced PC12 cell injury. After interference with si-MIAT, the results of flow cytometry showed that the rate of apoptosis was reduced. Western blotting results showed that the expression of LC3II/LC3I, Bax, and cleaved-caspase3 was decreased, while the expression of p-mTOR, p62, and Bcl-2 was increased. RNA pull-down and RIP assay found the binding relationship between MIAT and REDD1, and interference with si-MIAT down-regulated the expression of REDD1. The level of REDD1 ubiquitination was increased and the expression of REDD1 was decreased after interference with si-MIAT in PC12 cells. Co-IP results showed that interference with si-MIAT enhanced the binding ability of CUL4A-DDB1 and REDD1. ConclusionAltogether, MIAT promotes autophagy and apoptosis of neural cells and aggravates IS by up-regulating the expression of REDD1.

Knockdown of lncRNA SNHG14 alleviates LPS-induced inflammation and apoptosis of PC12 cells by regulating miR-181b-5p.

Spinal cord injury (SCI) is a traumatic central nervous system disorder that leads to permanent functional loss, and unavailable treatment of this disease results in poor quality of life. However, the specific role of long non-coding RNA small nucleolar RNA host gene 14 (lncRNA SNHG14) in SCI has not been fully studied. The aim of the current study was to investigate the role of SNHG14 and its regulatory mechanism in lipopolysaccharide (LPS)-induced PC-12 cells. LPS was used to stimulate PC-12 cells to simulate inflammatory injury following SCI in vitro. Cell viability and apoptosis were respectively assessed by Cell Counting Kit-8 assay and TUNEL assay. Western blotting was performed to detect the expressions of apoptosis-related proteins. The mRNA levels of SNHG14 and microRNA (miR)-181b-5p were detected by reverse transcription-quantitative PCR. The target of SNGH14 was predicted by bioinformatics analysis and subsequently validated by a luciferase reporter assay. ELISA was then used to detect the levels of inflammatory factors. The results indicated that LPS induced inflammation, decreased cell viability and increased the apoptosis of PC-12 cells. Interference of SNHG14 alleviated this type of injury of PC-12 cells. Bioinformatics prediction and luciferase reporter assay demonstrated that miR-181b-5p could directly bind to SNHG14. Moreover, mechanistic investigations revealed that the miR-181b-5p inhibitor could reverse the inhibitory effects of SNHG14 silencing on cell viability, inflammation and apoptosis of PC-12 cells. To conclude, the present results showed that SNHG14 knockdown alleviated PC-12 cell inflammation and apoptosis induced by LPS via regulating miR-181b-5p, which might provide a novel insight into the treatment of SCI.

P53-mediated ferroptosis is required for 1-methyl-4-phenylpyridinium-induced senescence of PC12 cells

Parkinson's disease (PD) is characterized by the loss of dopaminergic neurons in the substantia nigra and striatum. Aging is the most important risk factor of PD. Ferroptosis is an iron-dependent form of cell death associated with PD. However, it is not clear whether ferroptosis accelerates PD by promoting cellular senescence. This study investigated the mechanism of 1-methyl-4-phenylpyridinium (MPP+) -induced PC12 cells injury. We found that MPP+ induced cell senescence with increased β-galactosidase activity and the expression of p53, p21 and p16 activation in cells. In addition, MPP+ treatment showed smaller mitochondria and increased membrane density, downregulation of ferritin heavy chain 1 expression and upregulation of acyl-CoA synthetase long chain family member 4 expression, and enhanced levels of oxidative stress, which were important characteristics of ferroptosis. Ferrostatin-1 (Fer-1), a ferroptosis inhibitor, was tested to eliminate MPP+-induced cell senescence. Fer-1 downregulated the expression of p53 and upregulated the expression of solute carrier family 7 member 11 (SLC7A11) and glutathione peroxidase-4 (GPX4) in MPP+-induced ferroptosis. Inhibition of p53 eliminated cell senescence by upregulation the expression of of SLC7A11 and GPX4. Thus, these results suggest that MPP+ induces senescence in PC12 cells via the p53/ SLC7A11/ GPX4 signaling pathway in the ferroptosis regulation mechanism.

Editorial Expression of Concern: Long non-coding RNA Mirt2 relieves lipopolysaccharide-induced injury in PC12 cells by suppressing miR-429.

An Editorial Expression of Concern to this paper has been published: https://doi.org/10.1007/s13105-019-00691-7

A New Brominated Norsesquiterpene Glycoside From the Rhizomes of Acorus tatarinowii Schott

A new brominated norsesquiterpene glycoside, acoruside (1), has been isolated from the rhizomes of Acorus tatarinowii Schott, together with 8 known compounds (2-9). Their structures were elucidated mainly based on 1-dimensional (1D) and 2D nuclear magnetic resonance spectra. The absolute configuration of compound 1 was determined by comparing its experimental and calculated electronic circular dichroism spectra. The in vitro tests indicated that at 10 µM, compounds 2, 3, and 4 aggravated serum deprivation injuries of PC12 cells, compound 2 aggravated rotenone-induced injuries of PC12 cells, and compounds 3 and 4 aggravated the oxygen-glucose deprivation-induced injuries of PC12 cells.

Pretreatment of Indobufen and Aspirin and their Combinations with Clopidogrel or Ticagrelor Alleviates Inflammasome Mediated Pyroptosis Via Inhibiting NF-κB/NLRP3 Pathway in Ischemic Stroke.

Increasing studies showed that several anti-platelet drugs turned out to be a promising strategy for inflammatory response. In this study, we investigated the protective efficiency of pretreatment of indobufen or aspirin combined with clopidogrel or ticagrelor (IACT) on cerebral ischemic injury via NF-κB/NLRP3 pathway. Ischemia/reperfusion (I/R) injury was simulated in vivo by middle cerebral artery occlusion/reperfusion (MCAO/R) model, and rats were pretreated with indobufen and aspirin and their combinations with clopidogrel or ticagrelor respectively. The platelet aggregation, cerebral infarct size, water content, neurological impairment and LDH release were measured. The relative expression of inflammasome mediated pyroptosis was determined by ELISA, RT-PCR, Tunel, Immunofluorescence and Western blotting as appropriate. In vitro, I/R injury was simulated in PC12 cells using oxygen glucose deprivation/reperfusion (OGD/R) and Lipopolysaccharide (LPS) to induce pyroptosis. The effect of combinations were significantly greater than MCAO/R group on decreasing the platelet aggregation, infarct size, brain edema, LDH release and neurologic impairment. LPS aggravated I/R-induced PC12 cell injury, which was significantly suppressed by pretreatment of IACT and Bay11-7082. Mechanistically, IACT alleviated transcriptionally encoded IL-1β, IL-18 and NLRP3 via inhibiting nuclear transportation of NF-κB. Importantly, at protein level, NLRP3, Caspase-1, IL-18, IL-1β and GSDMD were significantly decreased in combination groups both in vivo and vitro. IACT reduce inflammasome mediated pyroptosis in MCAO/R rats and OGD/R PC12 cells through inhibiting NF-κB/NLRP3 signaling pathway, which suggests that drug combination is a protective strategy with clinical potential against I/R-induced injury. Graphical abstract.