Aerobic Training Inhibits Hippocampal Inflammation by Activating the Hippocampus TLR4/miR-223/NLRP3 Signaling Pathway Axis in Mice with CUMS-induced Depression

QU Honglin1,2 XIE Jun2 CHEN Jiaqin LIU Ruilian2 TANG Changfa CHEN Yilin CHEN Wei LI Di PENG Qi CHEN Rui1

(1.Hunan Normal University , Changsha, China 420012)
(2.Yichun University, Yichun, China 336000)
【Knowledge Link】pyknosis; Nissl body

【Abstract】 Objective: The purpose of this study was to explore the effects of aerobic training on the hippocampal inflammation by activating the hippocampal TLR4/miR-223/NLRP3 signal pathway in mice with chronic unpredictable mild stress (CUMS)-induced depression. Methods: Sixty 8-week-old male C57BL/6 mice were divided into control group (CG), CUMS model group (MG), CUMS model plus exercise group (ME), TLR4 inhibitor group (TG), and TLR4 inhibitor plus exercise group (TE) (n = 12). Except CG, the CUMS was molded for mice of the other four groups with chronic stress for 28 days. The mice in both ME group and TE group were trained using a treadmill at a moderate intensity for 8 weeks. The mice in TG group and TE group were received intraperitoneal injection of TAK-242, an inhibitor of TLR4, for 4 weeks. The neurobehavioral scores were tested after the 8-week training for all the mice. The samples were collected the day after their last training. The levels of blood IL-1β and IL-10 were measured using ELISA kit; the morphology of Nissl body in hippocampus was detected by Nissl staining; the positive expression areas of TLR4, IL-1β, IL-10, and NF-κB proteins were detected by immunohistochemistry; the mRNA expression levels of NLRP3, TLR4, IL-1β, IL-10, NF-κB, and miR-223 in hippocampus were examined by RT-PCR; the protein expression levels of TLR4, IL-1β, and IL-10 in hippocampus were tested by Western blot. Results: The mice with CUMS showed significantly decreased neurobehavioral function, significantly increased inflammatory cytokines such as IL-1β in blood, and the appearance of Nissl body pyknosis. The 8-week aerobic training significantly improved the hippocampus function in the mice with CUMS, reduced the level of IL-1β in blood, Nissl body pyknosis, and the desperate behaviors, and increased the mouse desire to survive. High-throughput sequencing and correlation analysis of miRNA and mRNA showed that there was a targeting regulatory relationship between miR-223 and NLRP3, which was influenced by exercise intervention. The expressions of inflammatory cytokines such as NLRP3, TLR4, IL-1β, and NF-κB were down-regulated while that of the anti-inflammatory cytokine IL-10 was up-regulated in ME mice. The expressions of NF-κB and NLRP3 were down-regulated and that of miR-223 was up-regulated, and the anti-inflammatory effect was increased in TG mice. The consistent anti-inflammatory effect was also found in TE mice. Conclusion: Aerobic training significantly reduced the expression of TLR4 in the hippocampus of mice with CUMS-induced depression while activated the TLR4/miR-223/NLRP3 pathway axis, thus improving the hippocampus function to inhibit inflammation and enhance the repair of hippocampal inflammatory injury.

【Keywords】 depression; chronic unpredictable mild stress; microRNA-223; aerobic training; hippocampus; TLR4/miR-223/NLRP3;


【Funds】 Science and Technology Project of Health and Family Planning Commission of Jiangxi Province (20171862) Science and Technology Key Project of Hunan Provincial Department of Education (1293)

Download this article

(Translated by PXJ1)


    Chen, W., Chen, J. Mao, H. et al. Chinese Journal of Arteriosclerosis (中国动脉硬化杂志), 25(8): 783–790 (2017).

    Dong, X. master’s thesis, Yunnan Normal University (2017).

    Du, J. China Sport Science and Technology (中国体育科技), 53(6): 101–107 (2017).

    Gao, P., Luo, X. Yu, Y. et al. Chinese Journal of Sports Medicine (中国运动医学杂志), 35(11): 1094–1097 (2015).

    Ge, H., Wei, H. Zhang, Y. et al. Journal of Beijing Sport University (北京体育大学学报), 45(5): 39–45 (2017).

    Geng, Y., Lin, Q. Ma, H. et al. Journal of Beijing Sport University (北京体育大学学报), 41(1): 70–74 (2018).

    Guo, Y., Luo, C. Chen, J. et al. China Sport Science and Technology (中国体育科技), 53(4): 119–124, 137 (2017).

    Hong, D., Qi, Y. Zhang, Y. et al. China Health Industry (中国卫生产业), 8(24): 3–4 (2011).

    Li, H. master’s thesis, Yangzhou University (2017).

    Li, X. master’s thesis, Taiyuan University of Technology (2016).

    Lin, Q. Geng, Y. & Tian, Z. China Sport Science (体育科学), 37(7): 44–49 (2017).

    Lin, X., Lu, L. & Wang, X. Journal of Shanghai University of Sport (上海体育学院学报), 41(4): 49–56 (2017).

    Liu, M., Feng, L. & Wang, X. Journal of Shanghai University of Sport (上海体育学院学报), 39(3): 87–89, 94 (2015).

    Liu, W., Guo, W. Xu, Q. et al. Acta Pharmaceutica Sinica (药学学报), 51(10): 1505–1512 (2016).

    Qu, H. & Chen, H. Chinese Journal of Rehabilitation Medicine (中国康复医学杂志) 23(8): 768–770 (2008).

    Qu, H., Xie, J. Chen, J. et al. Journal of Tianjin University of Sport (天津体育学院学报), 33(2): 148–155 (2018).

    Tang, M., Wu, Q. Zhang, H. et al. Chinese Journal of Gerontology (中国老年学杂志), 37(21): 5462–5464 (2017).

    Wang, H., Liu, S. Xia, Z. et al. Chinese Journal of Rehabilitation Medicine (中国康复医学杂志), 32(12): 1351–1355 (2017).

    Wei, Y., Yang, J. Yin, Z. et al. Journal of Trauma and Emergency (Electronic Version) (创伤与急诊电子杂志), 4(3): 166–184 (2016).

    Wen, R., Chang, H. & Bai, F. China Medical Engineering (中国医学工程), 16(1): 91–93 (2018).

    Wu, W., Yue, J, Wang, G. et al. Chinese Journal of Rehabilitation Medicine (中国康复医学杂志), 31(11): 1219–1224 (2016).

    Wu, X., Liu, L. Yin, Y. et al. Journal of Fujian Normal University(Natural Science Edition) (福建师范大学学报 (自然科学版)), 32(5): 106–112 (2016).

    Wu, Y., Sun, J. & Chi, T. Chinese Heart Journal (心脏杂志), 30(2): 141–145 (2018).

    Xia, X. doctoral thesis, Tianjin Medical University (2016).

    Zhang, T. & Xiong, X. Medical Recapitulate (医学综述), 21(2): 199–201 (2015).

    Zhang, X. doctoral thesis, East China Normal University (2016).

    Zhang, Y. doctoral thesis, The Second Military Medical University (2016).

    Zhen, F., Wang, L. & An, C. Chinese Journal of Behavioral Medicine and Brain Science (中华行为医学与脑科学杂志), 26(8): 760–764 (2017).

    AFSHARI M, YANG A, BEGA D, 2017. Motivators and barriers to exercise in Parkinson’s disease. J Parkinson’s Disease, 7(4): 703–711.

    AREND WP, PALMER G, GABAY C. 2008. IL-1, IL-18, and IL-33 families of cytokines. Immunological Reviews, 223: 20–38.

    BAUERNFEIND F, RIEGER A, SCHILDBERG FA, et al., 2012. NLRP3 inflammasome activity is negatively controlled by miR-223. J Immunol, 189(8): 4175–4181.

    BEDFORD TG, TIPTON CM, WILLSON NC, et al., 1979. Maximum oxygen consumption of rats and its changes with various experimental procedures. J Appl Physiol. 47(6): 1278–1283.

    CAMKURT MA, ACAR Ş, COŞKUN S, et al., 2015. Comparison of plasma MicroRNA levels in drug naive, first episode depressed patients and healthy controls. J Psychiatric Res, 69: 67–71.

    CAMKURT MA, GÜNEŞ S, COŞKUN S, et al., 2017. Peripheral Signatures of Psychiatric Disorders: MicroRNAs. Clin Psychopharmacology Neurosci, 15(4): 313–319.

    CHARLES L, CAPURON L, MILLERAH, 2006. Cytokines sing the blues: Inflammation and the pathogenesis of depression. Trends Immunol, 27(1): 24–31.

    CHENG Y, PARDO M, ARMINI RS, et al., 2016. Stress-induced neuroinflammation is mediated by GSK3-dependent TLR4 signaling that promotes susceptibility to depression-like behavior. Brain Behav Immunity, 53: 207–222.

    DUNN AJ, SWIERGIEL AH, DE BEAUREPAIRE R, 2005. Cytokines as mediators of depression: What can we learn from animal studies? Neurosci Biobehavioral Rev, 29(4): 891–909.

    HAAPAKOSKI R, EBMEIER KP, ALENIUS H, et al., 2016. Innate and adaptive immunity in the development of depression: An update on current knowledge and technological advances. Prog Neuropsychopharmacology Biolo Psychiatry, 66(3): 63–72.

    HANEKLAUS M, GERLIC M, KUROWSKA-STOLARSKA M, et al., 2012. Cutting edge: MiR-223 and EBV miR-BART15 regulate the NLRP3 inflammasome and IL-1β production. J Immunol, 189(8): 3795–3799.

    HE WT, WAN H, HU L, et al., 2015. Gasdermin D is an executor of pyroptosis and required for interleukin-1 β secretion. Cell Res, 25(12): 1285–1298.

    KAYAGAKI N, WONG MT, STOWE IB, et al., 2013. Noncanonical inflammasome activation by intracellular LPS independent of TLR4. Sci, 341(6151): 1246–1249.

    LATZ E, XIAO TS, STUTZ A, 2013. Activation and regulation of the inflammasomes. Nature Rev Immunol, 13(6): 397–411.

    LI T, MORGAN MJ, CHOKSI S, et al., 2010. MicroRNAs modulate the noncanonical transcription factor NF-kappaB pathway by regulating expression of the kinase IKKalpha during macrophage differentiation. Nat Immunol, 11(9): 799–805.

    LIN KM, HU W, TROUTMAN TD, et al., 2014. IRAK-1 bypasses priming and directly links TLRs to rapid NLRP3 inflammasome activation. Proc Nat Acad Sci USA, 111(8): 775–780.

    LIU W, SHENG H, XU Y, et al., 2013. Swimming exercise ameliorates depression-like behavior in chronically stressed rats: Relevant to proinflammatory cytokines and IDO activation. Brain Res, 242(2): 110–116.

    MCNEELA EA, BURKE A, NEILL DR, et al., 2010. Pneumolysin activates the NLRP3 Inflammasome and promotes proinflammatory cytokines independently of TLR4. PloS Pathogens, 6(11): e1001191.

    MAN SM, KANNEGANTI TD, 2015. Converging roles of caspases in inflammasome activation, cell death and innate immunity. Nat Rev Immunol, 16(1): 7.

    NEUDECKER V, HANEKLAUS M, JENSEN O, et al., 2017. Myeloidderived miR-223 regulates intestinal inflammation via repression of the NLRP3 inflammasome. J Exp Med, 214(6): 1737–1752.

    PLAYFORD ED, 2011. Exercise and Parkinson’s disease. J Neurol Neurosury Pshychiatry. 82(11): 1185.

    RADOMAIZIK S, ZALDIVAR F, OLIVER S, et al., 2010. Evidence for microRNA involvement in exercise-associated neutrophil gene expression changes. J Appl Physiol, 109(1): 252–261.

    SOLHAUG HI, ROMULD EB, ROMILD U, et al., 2012. Increased prevalence of depression in cohorts of the elderly: an 11-year followup in the generla population—the HUNT study. Int Psychogeriatrics, 24(1): 151–158.

    SUBRAMANIAN I, 2017. Complementary and alternative medicine and exercise in nonmotor symptoms of Parkinson’s disease. Int Rev Neurobiology, 134: 1163–1188.

    SUN W, JULIE LI YS, HUANG HD, et al., 2010. microRNA: A master regulator of cellular processes for bioengineering systems. Annu Rev Biomed Engin, 12(1): 1–27.

    WANG J, BAI XJ, SONG Q, et al., 2015. miR-223 inhibits lipid deposition and inflammation by suppressing Toll-like receptor 4 signaling in macrophages. Int J Mol Sci, 16: 24965–24982.

    WANG XH, HUANG W, YANG Y, et al., 2014. Loss of miR-223 duplex (5p and 3p) aggravates myocardial depression and mortality in polymicrobial sepsis. Biochim et Biophys Acta, 1842(5): 701–711.

    XU YJ, SHENG H, BAO QY, et al., 2016. NLRP3 inflammasome activation mediates estrogen deficiency-induced depression- and anxiety-like behavior and hippocampal inflammation in mice. Brain Behav Immunity, 56: 175–186.

    YAN H, LI L, LIU F, et al., 2011. Screening of the anti-inflammatory peptides against MD-2 and its depression on TLR4 activation stimulated by LPS. Inflammation Res, 60: S150.

    YANG Z, ZHONG L, XIAN R, et al., 2015. MicroRNA-223 regulates inflammation and brain injury via feedback to NLRP3 inflammasome after intracerebral hemorrhage. Mol Immunol, 65(2): 267–276.

This Article


CN: 11-1295/G8

Vol 39, No. 02, Pages 39-50

February 2019


Article Outline



  • 1 Materials and methods
  • 2 Results
  • 3 Discussion
  • 4 Conclusions
  • References