NEP1-40 Regulates the Development of Hippocampal Neural Stem Cells in Schizophrenic Mice

Abstract

Objective

Schizophrenia is a complex neuropsychiatric disorder characterized by cognitive, affective, and behavioral abnormalities. Existing treatments have yielded limited effects on improving cognitive function. Recent studies have identified the abnormal differentiation of hippocampal neural stem cells (NSCs), neuronal loss, and dysregulated proliferation of astrocytes as significant pathological mechanisms contributing to the symptoms of schizophrenia. Impaired hippocampal neurogenesis may lead to emotional and cognitive deficits and biases in learning and memory, indicating that NSC differentiation is critical. NEP1-40, a Nogo-A receptor inhibitor, has shown promise for nerve protection and repair promotion. However, the effects of NEP1-40 on stem cell differentiation, the reduction in neuronal apoptosis, and the amelioration of schizophrenia-like behaviors have not been adequately investigated. This study examined the influence of NEP1-40 on NSC differentiation, hippocampal neuronal apoptosis, and proliferation in adolescent mice, along with its potential to enhance cognitive and behavioral outcomes in MK-801-induced schizophrenia mouse models.

Methods

In in vivo experiments, a schizophrenia mouse model was successfully established. Subsequently, behavioral tests were conducted, followed by Western blotting (WB) and immunofluorescence (IF) analyses. In in vitro settings, NSCs were cultured and transfected. Flow cytometry, along with WB and IF assays, was employed to evaluate the effects of NEP1-40.

Results

Schizophrenia-like behaviors in mice were significantly improved with the overexpression of NEP1-40. Compared with the model group, the NEP1-40 treatment group presented increased expression of a neuronal marker (Tuj1), reduced expression of an astroglial marker (GFAP), and decreased hippocampal neuronal apoptosis. NSC differentiation was assessed by quantifying the number of BrdU-positive cells coexpressing Tuj1 and GFAP in the hippocampal dentate gyrus. NEP1-40 treatment led to an increase in BrdU/Tuj1-positive cells and a reduction in BrdU/GFAP-positive cells. In cellular studies, NEP1-40 overexpression similarly increased the number of Tuj1-positive cells, reduced the number of GFAP-positive cells, decreased the degree of neuronal apoptosis, and promoted neuronal proliferation.

Conclusion

These findings demonstrated the neurogenic effects of NEP1-40 on NSCs and their potential to mitigate schizophrenia-like behaviors in vivo.

Keywords

Schizophrenia / Cognition / Hippocampus / Stem cells / NEP1-40

References

[1] van den HeuvelMP, ScholtensLH, de LangeSC, et al.. Evolutionary modifications in human brain connectivity associated with schizophrenia. Brain, 2019, 142(12): 3991-4002.

[2] HuhnM, LeuchtC, RotheP, et al.. Reducing antipsychotic drugs in stable patients with chronic schizophrenia or schizoaffective disorder: a randomized controlled pilot trial. Eur Arch Psychiatry Clin Neurosci, 2021, 271(2): 293-302.

[3] NeumeierMS, HomanS, VetterS, et al.. Examining side effect variability of antipsychotic treatment in schizophrenia spectrum disorders: a meta-analysis of variance. Schizophr Bull, 2021, 47(6): 1601-1610.

[4] BretthauerJ, CanuD, ThiemannU, et al.. Attention for emotion-how young adults with neurodevelopmental disorders look at facial expressions of affect. Front Psychiatry, 2022, 13: 842-896.

[5] RoeskeMJ, KonradiC, HeckersS, et al.. Hippocampal volume and hippocampal neuron density, number and size in schizophrenia: a systematic review and meta-analysis of postmortem studies. Mol Psychiatry, 2021, 26(7): 3524-3535.

[6] ArsalidouM, YapleZ, JurcikT, et al.. Cognitive brain signatures of youth with early onset and relatives with schizophrenia: evidence from fMRI meta-analyses. Schizophr Bull, 2020, 46(4): 857-868.

[7] ParkYH, ShinSJ, KimHS, et al.. Omega-3 fatty acid-type docosahexaenoic acid protects against Aβ-mediated mitochondrial deficits and pathomechanisms in Alzheimer's disease-related animal model. Int J Mol Sci, 2020, 21(11): 3879-3879.

[8] SchultheisC, RosenbrockH, MackSR, et al.. Quantitative electroencephalography parameters as neurophysiological biomarkers of schizophrenia-related deficits: A Phase II substudy of patients treated with iclepertin (BI 425809). Transl Psychiatry, 2022, 121329.

[9] ZhangX, WeiX, MeiY, et al.. Modulating adult neurogenesis affects synaptic plasticity and cognitive functions in mouse models of Alzheimer's disease. Stem Cell Reports, 2021, 16(12): 3005-3019.

[10] HazzaaSM, AbdelazizSAM, Abd EldaimMA, et al.. Neuroprotective potential of allium sativum against monosodium glutamate-induced excitotoxicity: impact on short-term memory, gliosis, and oxidative stress. Nutrients, 2020, 12(4): 1028-1028.

[11] ZhaoT, WuD, DuJ, et al.. Folic acid attenuates glial activation in neonatal mice and improves adult mood disorders through epigenetic regulation. Front Pharmacol, 2022, 13818423.

[12] Baya MdzombaJ, JolyS, RodriguezL, et al.. Nogo-A-targeting antibody promotes visual recovery and inhibits neuroinflammation after retinal injury. Cell Death Dis, 2020, 112101.

[13] SotoA, Nieto-DíazM, ReigadaD, et al.. miR-182-5p Regulates nogo-A expression and promotes neurite outgrowth of hippocampal neurons in vitro. Pharmaceuticals (Basel), 2022, 155529.

[14] DingJ, ZhangC, ZhangYW, et al.. N-methyl-D-aspartate receptor subunit 1 regulates neurogenesis in the hippocampal dentate gyrus of schizophrenia-like mice. Neural Regen Res, 2019, 14(12): 2112-2117.

[15] ZhangQ, XiaY, WangY, et al.. CK2 phosphorylating I(2)(PP2A)/SET mediates tau pathology and cognitive impairment. Front Mol Neurosci, 2018, 11146.

[16] MaP, ZhaS, ShenX, et al.. NFAT5 mediates hypertonic stress-induced atherosclerosis via activating NLRP3 inflammasome in endothelium. Cell Commun Signal, 2019, 171102.

[17] BiZ, DalinW, XushengL, et al.. NEP1-40-overexpressing neural stem cells enhance axon regeneration by inhibiting Nogo-A/NgR1 signaling pathway. Curr Neurovasc Res, 2021, 18(3): 271-278.

[18] TaiC, ChangCW, YuGQ, et al.. Tau reduction prevents key features of autism in mouse models. Neuron, 2020, 106(3): 421-437.e411.

[19] AppleDM, MahesulaS, FonsecaRS, et al.. Calorie restriction protects neural stem cells from age-related deficits in the subventricular zone. Aging (Albany NY), 2019, 11(1): 115-126.

[20] LiH, XiangY, ZhuZ, et al.. Rifaximin-mediated gut microbiota regulation modulates the function of microglia and protects against CUMS-induced depression-like behaviors in adolescent rat. J Neuroinflammation, 2021, 181254.

[21] BobotM, ThomasL, MoyonA, et al.. Uremic toxic blood-brain barrier disruption mediated by AhR activation leads to cognitive impairment during experimental renal dysfunction. J Am Soc Nephrol, 2020, 31(7): 1509-1521.

[22] NiL, XuY, DongS, et al.. The potential role of the HCN1 ion channel and BDNF-mTOR signaling pathways and synaptic transmission in the alleviation of PTSD. Transl Psychiatry, 2020, 101101.

[23] SydingLA, Kubik-ZahorodnaA, NicklP, et al.. Generation and characterization of a novel angelman syndrome mouse model with a full deletion of the Ube3a gene. Cells, 2022, 11182815.

[24] ZhangY, ZhuM, SunY, et al.. Environmental noise degrades hippocampus-related learning and memory. Proc Natl Acad Sci U S A, 2021, 1181. e2017841117

[25] TetsadeP, dos BeatrizS, FernandaGQB-A, et al.. Guanosine promotes proliferation in neural stem cells from hippocampus and neurogenesis in adult mice. Mol Neurobiol, 2020, 57(9): 3814-3826.

[26] RudenJB, DixitM, ZepedaJC, et al.. Robust expression of functional NMDA receptors in human induced pluripotent stem cell-derived neuronal cultures using an accelerated protocol. Front Mol Neurosci, 2021, 14. 777049

[27] ZhuZ, PanQ, ZhaoW, et al.. BCL2 enhances survival of porcine pluripotent stem cells through promoting FGFR2. Cell Prolif, 2021, 541. e12932

[28] WangJ, QiW, ShiH, et al.. MiR-4763-3p targeting RASD2as a potential biomarker and therapeutic target for schizophrenia. Aging Dis, 2022, 13(4): 1278-1292.

[29] DuY, GaoY, WuG, et al.. Exploration of the relationship between hippocampus and immune system in schizophrenia based on immune infiltration analysis. Front Immunol, 2022, 13. 878997

[30] CastañoBL, SilvaAA, Hernandez-VelascoLL, et al.. Sulfadiazine plus pyrimethamine therapy reversed multiple behavioral and neurocognitive changes in long-term chronic toxoplasmosis by reducing brain cyst load and inflammation-related alterations. Front Immunol, 2022, 13. 822567

[31] YuW, FangH, ZhangL, et al.. Reversible changes in BDNF expression in MK-801-induced hippocampal astrocytes through NMDAR/PI3K/ERK signaling. Front Cell Neurosci, 2021, 15. 672136

[32] LiangJQ, ChenX, ChengY. Paeoniflorin rescued MK-801-induced schizophrenia-like behaviors in mice via oxidative stress pathway. Front Nutr, 2022, 9. 870032

[33] OsackaJ, KissA, BacovaZ, et al.. Effect of haloperidol and olanzapine on hippocampal cells' proliferation in animal model of schizophrenia. Int J Mol Sci, 2022, 23147711.

[34] ChenWH, LinYX, LinL, et al.. Identification of potential candidate proteins for reprogramming spinal cord-derived astrocytes into neurons: a proteomic analysis. Neural Regen Res, 2021, 16(11): 2257-2263.

[35] ChenSH, ChenZY, LinYH, et al.. Extracellular vesicles of adipose-derived stem cells promote the healing of traumatized achilles tendons. Int J Mol Sci, 2021, 222212373.

[36] GangL, YaoYC, LiuYF, et al.. Co-culture of oligodendrocytes and neurons can be used to assess drugs for axon regeneration in the central nervous system. Neural Regen Res, 2015, 10(10): 1612-1616.

[37] FangY, YaoL, LiC, et al.. The blockage of the Nogo/NgR signal pathway in microglia alleviates the formation of Aβ plaques and tau phosphorylation in APP/PS1 transgenic mice. J Neuroinflammation, 2016, 13156.

[38] Ben-SimonY, KaeferK, VelickyP, et al.. A direct excitatory projection from entorhinal layer 6b neurons to the hippocampus contributes to spatial coding and memory. Nat Commun, 2022, 1314826.

[39] BartschJC, BehrJ. Noncanonical, dopamine-dependent long-term potentiation at hippocampal output synapses in a rodent model of first-episode psychosis. Front Mol Neurosci, 2020, 1355.

[40] MarešP, KozlováL, MikuleckáA, et al.. The GluN2B-selective antagonist Ro 25–6981 is effective against PTZ-induced seizures and safe for further development in infantile rats. Pharmaceutics, 2021, 1391482.

[41] YangH, SuY, SunZ, et al.. Gold nanostrip array-mediated wireless electrical stimulation for accelerating functional neuronal differentiation. Adv Sci (Weinh), 2022, 922. e2202376

[42] JiaB, HuangW, WangY, et al.. Nogo-C inhibits peripheral nerve regeneration by regulating Schwann cell apoptosis and dedifferentiation. Front Neurosci, 2020, 14. 616258

[43] ZhaiZY, FengJ. Constraint-induced movement therapy enhances angiogenesis and neurogenesis after cerebral ischemia/reperfusion. Neural Regen Res, 2019, 14(10): 1743-1754.

[44] De FeliceM, MelisM, AroniS, et al.. The PPARα agonist fenofibrate attenuates disruption of dopamine function in a maternal immune activation rat model of schizophrenia. CNS Neurosci Ther, 2019, 25(5): 549-561.

[45] RaniB, Silva-MarquesB, LeursR, et al.. Short- and long-term social recognition memory are differentially modulated by neuronal histamine. Biomolecules, 2021, 114555.

[46] AlbertK, HiscoxJ, BoydB, et al.. Estrogen enhances hippocampal gray-matter volume in young and older postmenopausal women: a prospective dose-response study. Neurobiol Aging, 2017, 56: 1-6.

[47] DiaoM, QuY, LiuH, et al.. Effect of carbamylated erythropoietin on neuronal apoptosis in fetal rats during intrauterine hypoxic-ischemic encephalopathy. Biol Res., 2019, 52128.

[48] LuoY, YangW, LiN, et al.. Anodal transcranial direct current stimulation can improve spatial learning and memory and attenuate Aβ(42) burden at the early stage of Alzheimer's disease in APP/PS1 transgenic mice. Front Aging Neurosci, 2020, 12134.

[49] XuW, XiaoP, FanS, et al.. Blockade of Nogo-A/Nogo-66 receptor 1 (NgR1) inhibits autophagic activation and prevents secondary neuronal damage in the thalamus after focal cerebral infarction in hypertensive rats. Neuroscience, 2020, 431: 103-114.

Funding

Natural Science Foundation of Ningxia Province(2022AAC03154)

RIGHTS & PERMISSIONS

The Author(s), under exclusive licence to Huazhong University of Science and Technology