Abstract 3653: Akt3 Offers Stronger Protection than Akt1 Against Brain Injury by Differentially Promoting Mtor Protein Levels In Both In Vitro and In Vivo Stroke Models

Background and Objective: Akt is a serine-threonine kinase that plays critical role in promoting cell survival. Akt consists of three isoforms (Akt1, 2, 3), with Akt3 predominantly expressed in the brain. Although Akt pathway has been shown to mediate neuronal survival in cerebral ischemic injury, it is unclear how these Akt isoforms contribute to neuronal protection, and whether exogenous Akt can protect the brain against ischemic injury or not. In this study, we over-expressed Akt isoforms and its downstream signaling proteins such as FKHR and PRAS40 to investigate the role of the Akt pathway along with its potential relationship with the mTOR pathway in stroke. Methods: Sprauge Dawley rats (250∼280g) were used for all studies. A lentiviral vector consists of a CMV promoter driving IRES-eGFP was used to clone an active Akt 1 and 3 (cAKt 1 and 3), dominant-negative Akt (AktDN), active FKHR (AAA FKHR), and PRAS40. Lentivirus expressing these genes were added to primary mixed cortical cultures for two days prior to oxygen glucose deprivation (OGD) (MOI=1:5). Neuronal survival was measured by LDH release. Lentivirus were stereotaxically injected into the cortex, and rats were subjected to focal ischemia induced by distal MCA occlusion combined with bilateral CCA occlusion. Western blotting and immunofluorescent confocal microscopy were used to detect the expression of Akt isoforms and other proteins in both the Akt and mTOR pathways. Results: Western blotting analysis showed that both endogenous Akt1 and 3 proteins degraded as early as 1 h after stroke, while Akt2 protein remained unchanged until 24 h after stroke. In vitro studies showed that over-expression of both constitutively active cAkt1 and cAkt3 decreased LDH release after OGD, while AktDN worsened neuronal death ( P <0.05). In vivo over-expression of cAkt1, cAkt3 and PRAS40 reduced infarct size after stroke ( P <0.01). Gene transfer of cAkt1 and 3 also promoted protein levels of pAkt (phosphorylated Akt), pPRAS40, pFKHR, pPTEN, pmTOR, but not pGSK3β. Both in vitro and in vivo studies showed that over-expression of cAkt3 resulted in a stronger protection than cAkt1 ( P <0.05). Interestingly, cAkt3 gene transfer preserved both endogenous protein levels of Akt1 and 3, whereas cAkt1 gene transfer only preserved endogenous Akt1. Furthermore, cAkt3 promoted higher pmTOR levels than cAkt1. Treatment of rapamycin, an mTOR inhibitor, blocked the protective effects of both cAkt1 and cAkt3 both in vitro and in vivo. Conclusion: Lentiviral-mediated overexpression of cAkt3 confers stronger protection than that of cAkt1, by maintaining both endogenous Akt1 and Akt3, as well as promoting higher mTOR activities after stroke.

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Mn Inhibits GSH Synthesis via Downregulation of Neuronal EAAC1 and Astrocytic xCT to Cause Oxidative Damage in the Striatum of Mice

Oxidative Medicine and Cellular Longevity ◽

10.1155/2018/4235695 ◽

2018 ◽

Vol 2018 ◽

pp. 1-15 ◽

Cited By ~ 7

Author(s):

Xinxin Yang ◽

Haibo Yang ◽

Fengdi Wu ◽

Zhipeng Qi ◽

Jiashuo Li ◽

...

Keyword(s):

Oxidative Stress ◽

Oxidative Damage ◽

Dependent Manner ◽

Protein Levels ◽

Key Factor ◽

Protein Sulfhydryl ◽

Mrna And Protein Expression ◽

The Brain

Excessive manganese (Mn) can accumulate in the striatum of the brain following overexposure. Oxidative stress is a well-recognized mechanism in Mn-induced neurotoxicity. It has been proven that glutathione (GSH) depletion is a key factor in oxidative damage during Mn exposure. However, no study has focused on the dysfunction of GSH synthesis-induced oxidative stress in the brain during Mn exposure. The objective of the present study was to explore the mechanism of Mn disruption of GSH synthesis via EAAC1 and xCT in vitro and in vivo. Primary neurons and astrocytes were cultured and treated with different doses of Mn to observe the state of cells and levels of GSH and reactive oxygen species (ROS) and measure mRNA and protein expression of EAAC1 and xCT. Mice were randomly divided into seven groups, which received saline, 12.5, 25, and 50 mg/kg MnCl2, 500 mg/kg AAH (EAAC1 inhibitor) + 50 mg/kg MnCl2, 75 mg/kg SSZ (xCT inhibitor) + 50 mg/kg MnCl2, and 100 mg/kg NAC (GSH rescuer) + 50 mg/kg MnCl2 once daily for two weeks. Then, levels of EAAC1, xCT, ROS, GSH, malondialdehyde (MDA), protein sulfhydryl, carbonyl, 8-hydroxy-2-deoxyguanosine (8-OHdG), and morphological and ultrastructural features in the striatum of mice were measured. Mn reduced protein levels, mRNA expression, and immunofluorescence intensity of EAAC1 and xCT. Mn also decreased the level of GSH, sulfhydryl, and increased ROS, MDA, 8-OHdG, and carbonyl in a dose-dependent manner. Injury-related pathological and ultrastructure changes in the striatum of mice were significantly present. In conclusion, excessive exposure to Mn disrupts GSH synthesis through inhibition of EAAC1 and xCT to trigger oxidative damage in the striatum.

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Modulator of apoptosis-1 is a potential therapeutic target in acute ischemic injury

Journal of Cerebral Blood Flow & Metabolism ◽

10.1177/0271678x18794839 ◽

2018 ◽

Vol 39 (12) ◽

pp. 2406-2418 ◽

Cited By ~ 3

Author(s):

Su Jing Chan ◽

Hui Zhao ◽

Kazuhide Hayakawa ◽

Chou Chai ◽

Chong Teik Tan ◽

...

Keyword(s):

Cortical Neurons ◽

Infarct Volume ◽

Neuronal Loss ◽

Ischemic Injury ◽

Glucose Deprivation ◽

Artery Occlusion ◽

In Vivo Models ◽

The Brain

Modulator of apoptosis 1 (MOAP-1) is a Bax-associating protein highly enriched in the brain. In this study, we examined the role of MOAP-1 in promoting ischemic injuries following a stroke by investigating the consequences of MOAP-1 overexpression or deficiency in in vitro and in vivo models of ischemic stroke. MOAP-1 overexpressing SH-SY5Y cells showed significantly lower cell viability following oxygen and glucose deprivation (OGD) treatment when compared to control cells. Consistently, MOAP-1−/− primary cortical neurons were observed to be more resistant against OGD treatment than the MOAP-1+/+ primary neurons. In the mouse transient middle cerebral artery occlusion (tMCAO) model, ischemia triggered MOAP-1/Bax association, suggested activation of the MOAP-1-dependent apoptotic cascade. MOAP-1−/− mice were found to exhibit reduced neuronal loss and smaller infarct volume 24 h after tMCAO when compared to MOAP-1+/+ mice. Correspondingly, MOAP-1−/− mice also showed better integrity of neurological functions as demonstrated by their performance in the rotarod test. Therefore, both in vitro and in vivo data presented strongly support the conclusion that MOAP-1 is an important apoptotic modulator in ischemic injury. These results may suggest that a reduction of MOAP-1 function in the brain could be a potential therapeutic approach in the treatment of acute stroke.

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Abstract WMP38: Inhibition of Rip1 Kinase Improves Brain Functional Recovery After Ischemic Stroke via Reducing Astrocytic Scar Formation

Stroke ◽

10.1161/str.48.suppl_1.wmp38 ◽

2017 ◽

Vol 48 (suppl_1) ◽

Author(s):

Huiling Zhang ◽

Zhong-Sheng Li ◽

Yong Ni ◽

Xian-Yong Zhou ◽

Shi-Gang Qiao

Keyword(s):

Ischemic Stroke ◽

Western Blotting ◽

Functional Recovery ◽

Recovery Phase ◽

Glial Scar ◽

Scar Formation ◽

Western Blotting Analysis ◽

Protein Levels

During the recovery phase of ischemic stroke, one of the major barriers for the spontaneous neuronal axon regeneration is the formation of astrogliosis and glial scar, and targeting astrogliosis becomes a therapeutic strategy for ischemic stroke. However, the mechanism regulating the process of scar components after ischemia still remains poorly understood. The aim of this study was to observe the role of RIP1 kinase (RIP1K), the key regulator of necroptosis (programmed necrosis) in the brain functional recovery after ischemic stroke and in the ischemic stroke-induced astrogliosis and glial scar formation in both in vitro and in vivo glial scar models. The glial scar formation model in vitro or in vivo was established by using primary cultured astrocyte subjected to 6 hours of oxygen-glucose deprivation (OGD) following 12 hours or 24 hours reperfusion, or by 90 min of transient middle cerebral artery occlusion (tMCAO) and reperfusion in rats. Western blotting analysis and immunohistochemical assay showed that knockdown of RIP1K by lentivirally-delivered shRNAs against RIP1K (shRNA RIP1K) could decrease several protein levels of glial scar markers such as glial fibillary acidic protein (GFAP), neurocan and phosphacan both in in vitro and in vivo glial scar models. Furthermore, western blotting analysis showed that knockdown of RIP1K reduced the protein levels of VEGF-D receptor 3 in in vitro glial scar models. In addition, knockdown of RIP1K also notably reduced the shrinking volume and ameliorated the behavioral symptoms in the recovery phase of rats after tMCAO. And immunocytochemistry assay demonstrated that RIP1K knockdown promoted the neuronal axonal generation in a neuron and astrocyte co-culture system. Our data indicates that RIP1K might play an important role in the formation of glial scar after ischemic stroke via promoting the function of VEGF-D receptor 3 in astrocytes.

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Eriodictyol ameliorated cognitive dysfunction in APP/PS1 mice through inhibited ferroptosis via vitamin D receptor mediated Nrf2 activation

10.21203/rs.3.rs-1046295/v1 ◽

2021 ◽

Author(s):

Lin Li ◽

Wenjun Li ◽

Xiangru Zheng ◽

Qinglong Liu ◽

Qian Du ◽

...

Keyword(s):

Vitamin D ◽

Signaling Pathway ◽

Western Blotting ◽

Vitamin D Receptor ◽

Amyloid Β ◽

Qpcr Assay ◽

Aβ Aggregation ◽

The Brain

Abstract Background Alzheimer's disease (AD) is the most common type of neurodegenerative disease in contemporary era, and it is still clinically incurable. Eriodictyol, a natural flavonoid compound mainly exists in citrus fruits and some Chinese herbal medicine, has been reported with its effect of anti-inflammatory, antioxidant, anti-cancer and neuroprotective effects. However, there are few studies on the anti-AD effect and molecular mechanism of eriodictyol. Methods APP/PS1 mice were treated with eriodictyol and the cognitive function of mice was assessed by behavioral tests. The level of amyloid-β (Aβ) aggregation and hyper-phosphorylation of Tau in the brain of mice were detected by histological analysis and Western blotting. Meanwhile, HT-22 cells which induced by amyloid-β peptide (1-42) (Aβ1−42) oligomer were treated with eriodictyol after which cell viability was determined and the production of p-Tau was tested by Western blotting. Then, the characteristics of ferroptosis, including iron aggregation, lipid peroxidation and the expression of glutathione peroxidase type 4(GPX4), were determined both in vivo and in vitro by Fe straining, Western blotting and qPCR assay. Additionally, the expression level of Vitamin D receptor (VDR) and the activity of nuclear factor erythroid 2-related factor 2/heme oxygenase-1 (Nrf2/HO-1) signaling pathway were tested by Western blotting and qPCR assay. After that, the HT-22 cells with VDR knockout were used to explore the potential mechanisms and the relationship between VDR and Nrf2 was further assessed by coimmunoprecipitation assay and bioinformatics analysis. Results Eriodictyol obviously ameliorated cognitive deficits in APP/PS1 mice, suppressed Aβ aggregation and the phosphorylated level Tau in the brain of APP/PS1 mice. Meanwhile, eriodictyol could inhibit Tau hyper-phosphorylation and neurotoxicity in HT-22 cells induced by Aβ1−42 oligomer. Furthermore, both in vivo and in vitro, eriodictyol showed the anti-ferroptosis effect and its mechanism may connected with the activation of Nrf2/HO-1 signaling pathway. Additionally, the further experiment explains that the activation of Nrf2/HO-1 signaling pathway with eriodictyol treatment mediated by VDR. Conclusions Eriodictyol alleviated memory impairment and AD-like pathological changes via activating Nrf2/HO-1 signaling pathway mediated by VDR, which provide a new possibility for the treatment of AD.

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Long Non-Coding RNA FOXD1-AS1 Promotes the Progression and Glycolysis of Nasopharyngeal Carcinoma by Sustaining FOXD1 Expression

10.21203/rs.3.rs-52587/v1 ◽

2020 ◽

Author(s):

Zhanwang Wang ◽

Dong He ◽

Yuxing Zhu ◽

Xueying Hu ◽

Yi Jin ◽

...

Keyword(s):

Nasopharyngeal Carcinoma ◽

Molecular Mechanisms ◽

Bioinformatics Analysis ◽

Regulatory Role ◽

Loss Of Function ◽

Over Expression ◽

Protein Levels ◽

Non Coding Rna

Abstract Background: Increasing evidence have emphasized the importance of long non-coding RNAs (lncRNAs) in various human cancers progression. Forkhead box D1 antisense RNA1 (FOXD1-AS1) is a novel lncRNA and plays vital regulatory role in diverse biological processes of cancers. However, the biological function, molecular mechanism and clinical significance of FOXD1-AS1 in nasopharyngeal carcinoma is still unknown.Methods: Comprehensive bioinformatics analysis and qRT-PCR was conducted to detect the expression level of FOXD1-AS1. Loss-of-function and gain-of-function experiments were performed to verify the functions of FOXD1-AS1 in proliferation, migration, invasion, apoptosis and glycolysis of nasopharyngeal carcinoma in vitro and in vivo. Further bioinformatics analysis and experiments were carried out to explore the underlying molecular mechanisms of FOXD1-AS1. Results: FOXD1-AS1 was significantly overexpressed in nasopharyngeal carcinoma and associated with poor survival in patients. Knockdown of FOXD1-AS1 significantly inhibited cell proliferation, migration, invasion and glycolysis, and promotes apoptosis in nasopharyngeal carcinoma, whereas over-expression of FOXD1-AS1 has the opposite effect. Mechanistically, we found that FOXD1-AS1 could upregulate the expression of FOXD1 through stabilizing the FOXD1 expression at mRNA and protein levels, and FOXD1 increased the glycolysis level by transcriptionally upregulating the expression of LDHA, PKM and ENO1, thus playing an oncogenic role in nasopharyngeal carcinoma progression. Conclusion: FOXD1-AS1 plays a critical regulatory role in nasopharyngeal carcinoma. The identified FOXD1-AS1/FOXD1 axis may serve as a potential prognostic biomarker and therapeutic target for patients with nasopharyngeal carcinoma.

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NFKBIZ3′ UTR Mutations Confer Selective Growth Advantage and Activate Genes with Therapeutic Implications in Diffuse Large B-Cell Lymphoma

Blood ◽

10.1182/blood-2019-127421 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 296-296

Author(s):

Sarah E Arthur ◽

Nicole Thomas ◽

Christopher Rushton ◽

Miguel Alcaide ◽

Shannon Healy ◽

...

Keyword(s):

B Cell ◽

Research Funding ◽

B Cell Lymphoma ◽

Selective Growth ◽

Over Expression ◽

Protein Levels ◽

Selective Growth Advantage ◽

Mutant Lines

Introduction: The activated B-cell-like (ABC) subtype of diffuse large B-cell lymphoma (DLBCL) is characterized by activation of NF-κB signaling and increased mortality. Recurrent mutations affecting genes such as MYD88, CD79A/B and TNFAIP3 have been shown to be involved in activation of the NF-κB pathway in ABC DLBCL; however there still remain cases with no known genetic basis for this pathway activation suggesting that our understanding of the drivers of ABC DLBCL remains incomplete. Previously, NFKBIZ was shown to be amplified in ~10% of ABC DLBCLs and to contribute to activation of NF-κB signaling. We have recently described a novel pattern of mutations affecting the 3′ UTR of NFKBIZ resulting in an overall mutation rate of 34% for this gene (UTR mutations or amplifications) in ABC DLBCL. These NFKBIZ UTR mutations are mutually exclusive with MYD88 mutations, thus suggesting they may also lead to activation of NF-κB signaling. NFKBIZ encodes the IκB-ζ protein, which interacts with NF-κB transcription factors and is thought to regulate canonical NF-κB signaling. We hypothesized that NFKBIZ UTR mutations affect the normally rapid degradation of mRNA by disrupting secondary structures recognized by RNA-binding proteins such as ribonucleases. The resulting elevated mRNA levels would in turn lead to accumulation of IκB-ζ protein as a novel mechanism to promote cell growth and survival in ABC DLBCL. Methods: NFKBIZ 3′ UTR mutations were introduced into a DLBCL cell line (WSU-DLCL2) using the CRISPR-Cas9 system. NFKBIZ mRNA and protein levels were evaluated using custom designed droplet digital PCR assays and western blot analyses. Cells were stimulated with LPS to induce NFKBIZ expression and mRNA and protein levels were measured in wild-type (WT) and CRISPR-mutant lines to compare rates of mRNA decay and protein expression. A competitive growth assay with WT and CRISPR-mutant lines was performed to assess whether UTR mutations provide a growth advantage in culture (in vitro) and in mouse xenografts (in vivo). The composition of the pool of WT and mutant lines was determined by comparing WT and mutant DNA sequence proportions. RNA-sequencing was then performed on WT and a subset of CRISPR-mutant cell lines to identify genes up-regulated by IκB-ζ in mutant lines. Findings from these models were compared to the effects of NFKBIZ over-expression in patient tissues. Results: Introduction of NFKBIZ mutations into a DLBCL cell line confirmed that UTR deletions lead to increased mRNA and protein levels. LPS stimulation showed prolonged mRNA elevation in mutant lines, consistent with our model wherein these mutations disrupt post-transcriptional regulatory mechanisms. NFKBIZ UTR deletions gave DLBCL cells a selective growth advantage over WT both in vitro (cell culture) and in vivo (xenograft mouse model). RNA-sequencing of mutant and WT lines revealed possible transcriptional targets of IκB-ζ including some NF-κB targets and genes commonly over-expressed in ABC DLBCL (BATF, MAML2, and TNFRSF13B). HCK, a gene known to be activated by MYD88 was also upregulated in NFKBIZ mutant lines. This is consistent with our hypothesis that mutations in MYD88 and the NFKBIZ UTR are mutually exclusive because they activate similar pathways. HCK is also a target of ibrutinib, suggesting the potential utility of ibrutinib in these patients. Novel targets of IκB-ζ were also discovered through this analysis including CD274, the gene encoding PD-L1. This could be a novel mechanism for DLBCL tumours to express PD-L1 and therefore suggest that these tumours may be susceptible to anti-PD1/PD-L1 immunotherapies. Conclusions: This work highlights the role of NFKBIZ and 3′ UTR mutations in driving ABC DLBCL. We demonstrate that these mutations can lead to over-expression of NFKBIZ and provide a selective growth advantage to cells both in vitro and in vivo. In addition, we described multiple targets of IκB-ζ that may have implications in treatment susceptibility and/or resistance in ABC DLBCL. These findings contribute to a better understanding of the genetic basis of DLBCL, which is necessary to guide personalized therapeutic strategies. Disclosures Scott: Roche/Genentech: Research Funding; NanoString: Patents & Royalties: Named inventor on a patent licensed to NanoSting [Institution], Research Funding; Janssen: Consultancy, Research Funding; Celgene: Consultancy. Steidl:Bristol-Myers Squibb: Research Funding; Nanostring: Patents & Royalties: Filed patent on behalf of BC Cancer; Juno Therapeutics: Consultancy; Tioma: Research Funding; Bayer: Consultancy; Roche: Consultancy; Seattle Genetics: Consultancy.

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Nonclassical Mechanisms of Progesterone Action in the Brain: I. Protein Kinase C Activation in the Hypothalamus of Female Rats

Endocrinology ◽

10.1210/en.2008-0712 ◽

2008 ◽

Vol 149 (11) ◽

pp. 5509-5517 ◽

Cited By ~ 26

Author(s):

Bhuvana Balasubramanian ◽

Wendy Portillo ◽

Andrea Reyna ◽

Jian Zhong Chen ◽

Anthony N. Moore ◽

...

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Intracellular Signaling ◽

Estradiol Benzoate ◽

Female Rats ◽

Protein Levels ◽

Kinase C ◽

The Brain

The modulation of gene regulation by progesterone (P) and its classical intracellular regulation by progestin receptors in the brain, resulting in alterations in physiology and behavior has been well studied. The mechanisms mediating the short latency effects of P are less well understood. Recent studies have revealed rapid nonclassical signaling action of P involving the activation of intracellular signaling pathways. We explored the involvement of protein kinase C (PKC) in P-induced rapid signaling in the ventromedial nucleus of the hypothalamus (VMN) and preoptic area (POA) of the rat brain. Both the Ca2+-independent (basal) PKC activity representing the activation of PKC by the in vivo treatments and the Ca+2-dependent (total) PKC activity assayed in the presence of exogenous cofactors in vitro were determined. A comparison of the two activities demonstrated the strength and temporal status of PKC regulation by steroid hormones in vivo. P treatment resulted in a rapid increase in basal PKC activity in the VMN but not the POA. Estradiol benzoate priming augmented P-initiated increase in PKC basal activity in both the VMN and POA. These increases were inhibited by intracerebroventricular administration of a PKC inhibitor administered 30 min prior to P. The total PKC activity remained unchanged demonstrating maximal PKC activation within 30 min in the VMN. In contrast, P regulation in the POA significantly attenuated total PKC activity ± estradiol benzoate priming. These rapid changes in P-initiated PKC activity were not due to changes in PKC protein levels or phosphorylation status.

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TRIB3-Regulated Akt Signal Pathway Affects Trophoblast Invasion in the Development of Preeclampsia

American Journal of Perinatology ◽

10.1055/s-0041-1735872 ◽

2021 ◽

Author(s):

Xin Sui ◽

Lei Zhang ◽

Xu-Feng Zhang ◽

Ya Zhang

Keyword(s):

Western Blotting ◽

Serum Levels ◽

Urinary Protein ◽

Trophoblast Invasion ◽

Akt Pathway ◽

Invasion And Migration ◽

Placental Efficiency ◽

And Migration

Objective The aim of the study is to explore the mechanism of tribbles pseudokinase 3 (TRIB3)-regulated Akt pathway in the development of preeclampsia (PE). Methods TRIB3 expression in the placenta of PE patient was determined by quantitative reverse transcriptase polymerase chain reaction and western blotting. Then HTR-8/SVneo or JEG-3 cells were transfected and divided into Mock, Control siRNA, TRIB3 siRNA-1, and TRIB3 siRNA-2 groups. Cell proliferation, invasion, and migration were determined by MTT assay, Transwell assay, and wound healing test, while the expression of TRIB3 and Akt pathway was measured by western blotting. PE rats were treated with TRIB3 siRNA, and blood pressure, 24-hour urinary protein, as well as serum levels of sFlt-1 and vascular endothelial growth factor (VEGF) were measured. Results The placenta of PE patients presented with increased TRIB3 expression. In comparison with Mock group, the proliferation, invasion, and migration of HTR-8/SVneo and JEG-3 cells in TRIB3 siRNA-1 group and TRIB3 siRNA-2 group increased, with decreased TRIB3 expression but enhanced expression of p-Akt/Akt, MMP-2, and MMP-9. Rats in PE group showed increases in mean arterial pressure, SBP, 24-hour urinary protein, and serum sFlt-1 levels, but decreases in serum VEGF levels, fetal weight, and placental efficiency. Moreover, TRIB3 expression was upregulated, while p-Akt/Akt was downregulated in the placenta of rats in PE group. However, indicators above were significantly improved in rats treated with TRIB3 siRNA. Conclusion TRIB3 was upregulated in the PE placenta, while silencing TRIB3 activated the Akt signaling pathway to promote the invasion and migration of trophoblast both in vitro and in vivo and ameliorated the development of PE symptoms in the PE rat model. Key Points

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Novel Ubiquitin Specific Protease-13 Inhibitors Alleviate Neurodegenerative Pathology

Metabolites ◽

10.3390/metabo11090622 ◽

2021 ◽

Vol 11 (9) ◽

pp. 622

Author(s):

Xiaoguang Liu ◽

Kaluvu Balaraman ◽

Ciarán C. Lynch ◽

Michaeline Hebron ◽

Christian Wolf ◽

...

Keyword(s):

Neurodegenerative Diseases ◽

Small Molecule ◽

Alpha Synuclein ◽

Protein Levels ◽

Neurodegenerative Pathology ◽

Specific Protease ◽

Ubiquitin Specific Protease ◽

The Brain

Ubiquitin Specific Protease-13 (USP13) promotes protein de-ubiquitination and is poorly understood in neurodegeneration. USP13 is upregulated in Alzheimer’s disease (AD) and Parkinson’s disease (PD), and USP13 knockdown via shRNA reduces neurotoxic proteins and increases proteasome activity in models of neurodegeneration. We synthesized novel analogues of spautin-1 which is a non-specific USP13 inhibitor but unable to penetrate the brain. Our synthesized small molecule compounds are able to enter the brain, more potently inhibit USP13, and significantly reduce alpha-synuclein levels in vivo and in vitro. USP13 inhibition in transgenic mutant alpha-synuclein (A53T) mice increased the ubiquitination of alpha-synuclein and reduced its protein levels. The data suggest that novel USP13 inhibitors improve neurodegenerative pathology via antagonism of de-ubiquitination, thus alleviating neurotoxic protein burden in neurodegenerative diseases.

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Cpne3 Regulates the Cell Proliferation and Apoptosis in Human Gbm via the Activation of Pi3k/akt Signaling Pathway

10.21203/rs.3.rs-241485/v1 ◽

2021 ◽

Author(s):

Dainan Zhang ◽

Xiaoyin Wang ◽

Xi Wang ◽

Zemin Wang ◽

Shunchang Ma ◽

...

Keyword(s):

Cell Proliferation ◽

Cell Apoptosis ◽

Bioinformatic Analysis ◽

Akt Pathway ◽

Clinical Samples ◽

Protein Markers ◽

Mice Model ◽

Over Expression

Abstract Background: Even with decades of intensive study, the signaling regulative network of the progression of GBM remains unclear, a deeper understanding of the molecular crosstalk with pathways in GBM is needed to identify new potential targets for treatment. Methods: To investigated the expression of CPNE3 in GBM, we applied bioinformatic analysis and clinical samples validation. Then the functional validation of carried out in commercially available glioma cell lines and nude mice model. Also, the GSEA analysis was used to identify the relevant pathways. The role of activated pathway was further validated by pharmacology method.Results: We found that CPNE3 was significantly up-regulated in GBM when compared with adjacent normal tissues, and the overexpression of CPNE3 promoted cell proliferation and inhibiting cell apoptosis in vitro and in vivo. Also, the principal protein markers of PI3K/AKT pathway were found to be phosphorylated by CPNE3 over-expression, and pathway inhibitor, LY294002, alleviated the cell proliferation enhancement induced by CPNE3 over-expression. Conclusion: Our results showed that the expression of CPNE3 promotes cell proliferation by inhibiting cell apoptosis via activating PI3K/AKT pathway, thereby enhancing the progression of GBM, which suggesting that CPNE3 may play as a tumorigenesis gene and its crosstalk with PI3K/AKT pathway may become a promising potential therapeutic target for human GBMs.

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