Spinal cord injury (SCI) is one of the leading causes of neurological disability and death. So far, there is no satisfactory treatment for SCI, because of its complex and ill-defined pathophysiology. Recently, autophagy has been implicated as protective in acute SCI rat models. Here, we investigated the therapeutic value of a dietary intervention, namely, intermittent fasting (IF), on neuronal survival after acute SCI in rats, and its underlying mechanism related to autophagy regulation. We found remarkable improvement in both behavioral performance and neuronal survival at the injured segment of the spinal cord of animals previously subjected to IF. Western blotting revealed a marked decrease in apoptosis-related markers such as cleaved caspase 3 levels and the bax/bcl-2 ratio in the IF group, which suggested an inhibition of the intrinsic apoptosis pathway. In addition, the expression of the autophagy markers LC3-II and beclin 1 was also increased in the IF group compared with ad libitum fed animals. In parallel, IF decreased the levels of the substrate protein of autophagy, p62, indicative of an upregulation of the autophagic processes. Treatment with 3-methyladenine (3-MA), a selective inhibitor of autophagy, reversed the downregulated apoptosis-related markers by IF. Finally, IF could activate the adenosine monophosphate (AMP)-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) pathway and enhance lysosome function by upregulating transcription factor (TF)EB expression. Altogether, the present findings suggest that IF exerts a neuroprotective effect after acute SCI via the upregulation of autophagy, and further points to dietary interventions as a promising combinatorial treatment for SCI.
ChenH., JiH., ZhangM., LiuZ., LaoL., DengC., ChenJ., and ZhongG. (2017). An agonist of the protective factor SIRT1 improves functional recovery and promotes neuronal survival by attenuating inflammation after spinal cord injury. J. Neurosci. 37, 2916–2930.
3.
RabchevskyA.G., PatelS.P., and SpringerJ.E. (2011). Pharmacological interventions for spinal cord injury: where do we stand?. How might we step forward? Pharmacol. Ther. 132, 15–29.
4.
ArediaF., Guaman OrtizL.M., GiansantiV., and ScovassiA.I. (2012). Autophagy and cancer. Cells, 1, 520–534.
5.
BanerjeeR., BealM.F., and ThomasB. (2010). Autophagy in neurodegenerative disorders: pathogenic roles and therapeutic implications. Trends Neurosci. 33, 541–549.
6.
KannoH., OzawaH., SekiguchiA., and ItoiE. (2009). The role of autophagy in spinal cord injury. Autophagy, 5, 390–392.
7.
WangZ.Y., LinJ.H., MuharramA., and LiuW.G. (2014). Beclin-1-mediated autophagy protects spinal cord neurons against mechanical injury-induced apoptosis. Apoptosis, 19, 933–945.
8.
TangP., HouH., ZhangL., LanX., MaoZ., LiuD., HeC., DuH., and ZhangL. (2014). Autophagy reduces neuronal damage and promotes locomotor recovery via inhibition of apoptosis after spinal cord injury in rats. Mol. Neurobiol. 49, 276–287.
9.
GodarR.J., MaX., LiuH., MurphyJ.T., WeinheimerC.J., KovacsA., CrosbyS.D., SaftigP., and DiwanA. (2015). Repetitive stimulation of autophagy-lysosome machinery by intermittent fasting preconditions the myocardium to ischemia-reperfusion injury. Autophagy, 11, 1537–1560.
10.
QinJ., ZhouJ., DaiX., ZhouH., PanX., WangX., ZhangF., RaoJ., and LuL. (2016). Short-term starvation attenuates liver ischemia-reperfusion injury (IRI) by Sirt1-autophagy signaling in mice. Am. J. Transl. Res. 8, 3364–3375.
11.
RichN.J., Van LandinghamJ.W., FigueiroaS., SethR., CorniolaR.S., and LevensonC.W. (2010). Chronic caloric restriction reduces tissue damage and improves spatial memory in a rat model of traumatic brain injury. J. Neurosci. Res. 88, 2933–2939.
12.
PlunetW.T., LamC.K., LeeJ.H., LiuJ., and TetzlaffW. (2010). Prophylactic dietary restriction may promote functional recovery and increase lifespan after spinal cord injury. Ann. N. Y. Acad. Sci. 1198, Suppl. 1, E1–11.
13.
JeongM.A., PlunetW., StreijgerF., LeeJ.H., PlemelJ.R., ParkS., LamC.K., LiuJ., and TetzlaffW. (2011). Intermittent fasting improves functional recovery after rat thoracic contusion spinal cord injury. J. Neurotrauma, 28, 479–492.
YangL., LicastroD., CavaE., VeroneseN., SpeltaF., RizzaW., BertozziB., VillarealD.T., HotamisligilG.S., HolloszyJ.O., and FontanaL. (2016). Long-term calorie restriction enhances cellular quality-control processes in human skeletal muscle. Cell Rep. 14, 422–428.
16.
YuanW., and ZhuY. (2019). 141. Autophagy induction contributes to the neuroprotective effect of intermittent fasting in experimental spinal cord injury. Spine J. 19, S68.
17.
DaiS., WangB., LiW., WangL., SongX., GuoC., LiY., LiuF., ZhuF., WangQ., WangX., ShiY., WangJ., ZhaoW., and ZhangL. (2016). Systemic application of 3-methyladenine markedly inhibited atherosclerotic lesion in ApoE(-/-) mice by modulating autophagy, foam cell formation and immune-negative molecules. Cell Death Dis. 7, e2498.
18.
YaoT., YingX., ZhaoY., YuanA., HeQ., TongH., DingS., LiuJ., PengX., GaoE., PuJ., and HeB. (2015). Vitamin D receptor activation protects against myocardial reperfusion injury through inhibition of apoptosis and modulation of autophagy. Antioxid. Redox Signal. 22, 633–650.
19.
BassoD.M., BeattieM.S., and BresnahanJ.C. (1995). A sensitive and reliable locomotor rating scale for open field testing in rats. J. Neurotrauma, 12, 1–21.
ZhouK.L., ZhouY.F., WuK., TianN.F., WuY.S., WangY.L., ChenD.H., ZhouB., WangX.Y., XuH.Z., and ZhangX.L. (2015). Stimulation of autophagy promotes functional recovery in diabetic rats with spinal cord injury. Sci. Rep. 5, 17130.
22.
SahaniM.H., ItakuraE., and MizushimaN. (2014). Expression of the autophagy substrate SQSTM1/p62 is restored during prolonged starvation depending on transcriptional upregulation and autophagy-derived amino acids. Autophagy, 10, 431–441.
23.
HeX., YuanW., LiZ., HouY., LiuF., and FengJ. (2017). 6-Hydroxydopamine induces autophagic flux dysfunction by impairing transcription factor EB activation and lysosomal function in dopaminergic neurons and SH-SY5Y cells. Toxicol. Lett. 283, 58–68.
24.
SettembreC., Di MaltaC., PolitoV.A., Garcia ArencibiaM., VetriniF., ErdinS., ErdinS.U., HuynhT., MedinaD., ColellaP., SardielloM., RubinszteinD.C., and BallabioA. (2011). TFEB links autophagy to lysosomal biogenesis. Science, 332, 1429–1433.
25.
BaylissJ.A., LemusM.B., StarkR., SantosV.V., ThompsonA., ReesD.J., GalicS., ElsworthJ.D., KempB.E., DaviesJ.S., and AndrewsZ.B. (2016). Ghrelin-AMPK signaling mediates the neuroprotective effects of calorie restriction in Parkinson's disease. J. Neurosci. 36, 3049–3063.
26.
YuJ., ParkhitkoA.A., and HenskeE.P. (2010). Mammalian target of rapamycin signaling and autophagy: roles in lymphangioleiomyomatosis therapy. Proc. Am. Thorac. Soc. 7, 48–53.
27.
UnoM., HonjohS., MatsudaM., HoshikawaH., KishimotoS., YamamotoT., EbisuyaM., YamamotoT., MatsumotoK., and NishidaE. (2013). A fasting-responsive signaling pathway that extends life span in C. elegans. Cell Rep. 3, 79–91.
28.
LongoV.D., and MattsonM.P. (2014). Fasting: molecular mechanisms and clinical applications. Cell Metab. 19, 181–192.
29.
LevineM.E., SuarezJ.A., BrandhorstS., BalasubramanianP., ChengC.W., MadiaF., FontanaL., MirisolaM.G., Guevara-AguirreJ., WanJ., PassarinoG., KennedyB.K., WeiM., CohenP., CrimminsE.M., and LongoV.D. (2014). Low protein intake is associated with a major reduction in IGF-1, cancer, and overall mortality in the 65 and younger but not older population. Cell Metab. 19, 407–417.
30.
FannD.Y., NgG.Y., PohL., and ArumugamT.V. (2017). Positive effects of intermittent fasting in ischemic stroke. Exp. Gerontol. 89, 93–102.
31.
DavisL.M., PaulyJ.R., ReadnowerR.D., RhoJ.M., and SullivanP.G. (2008). Fasting is neuroprotective following traumatic brain injury. J. Neurosci. Res. 86, 1812–1822.
32.
MalinowskiB., ZalewskaK., WesierskaA., SokolowskaM.M., SochaM., LicznerG., Pawlak-OsinskaK., and WicinskiM. (2019). Intermittent fasting in cardiovascular disorders–an overview. Nutrients, 11, 673.
33.
PlunetW.T., StreijgerF., LamC.K., LeeJ.H., LiuJ., and TetzlaffW. (2008). Dietary restriction started after spinal cord injury improves functional recovery. Exp. Neurol. 213, 28–35.
34.
StreijgerF., PlunetW.T., PlemelJ.R., LamC.K., LiuJ., and TetzlaffW. (2011). Intermittent fasting in mice does not improve hindlimb motor performance after spinal cord injury. J. Neurotrauma, 28, 1051–1061.
35.
ChengE.H., KirschD.G., ClemR.J., RaviR., KastanM.B., BediA., UenoK., and HardwickJ.M. (1997). Conversion of Bcl-2 to a Bax-like death effector by caspases. Science, 278, 1966–1968.
36.
Loncarevic-VasiljkovicN., PesicV., TodorovicS., PopicJ., SmiljanicK., MilanovicD., RuzdijicS., and KanazirS. (2012). Caloric restriction suppresses microglial activation and prevents neuroapoptosis following cortical injury in rats. PLoS One, 7, e37215.
37.
ShenY., KapfhamerD., MinnellaA.M., KimJ.E., WonS.J., ChenY., HuangY., LowL.H., MassaS.M., and SwansonR.A. (2017). Bioenergetic state regulates innate inflammatory responses through the transcriptional co-repressor CtBP. Nat. Commun. 8, 624.
38.
SullivanP.G., RippyN.A., DorenbosK., ConcepcionR.C., AgarwalA.K., and RhoJ.M. (2004). The ketogenic diet increases mitochondrial uncoupling protein levels and activity. Ann. Neurol. 55, 576–580.
39.
LevineB., and KroemerG. (2008). Autophagy in the pathogenesis of disease. Cell, 132, 27–42.
40.
BhatP., KrielJ., Shubha PriyaB., Basappa, ShivananjuN.S., and LoosB. (2017). Modulating autophagy in cancer therapy: Advancements and challenges for cancer cell death sensitization. Biochem. Pharmacol. 147, 170–182.
41.
DelbridgeL.M., MellorK.M., TaylorD.J., and GottliebR.A. (2015). Myocardial autophagic energy stress responses–macroautophagy, mitophagy, and glycophagy. Am. J. Physiol. Heart Circ. Physiol. 308, H1194–1204.
42.
LumkwanaD., du ToitA., KinnearC., and LoosB. (2017). Autophagic flux control in neurodegeneration: progress and precision targeting–Where do we stand?. Prog. Neurobiol. 153, 64–85.
43.
SekiguchiA., KannoH., OzawaH., YamayaS., and ItoiE. (2012). Rapamycin promotes autophagy and reduces neural tissue damage and locomotor impairment after spinal cord injury in mice. J. Neurotrauma, 29, 946–956.
44.
BaiL., MeiX., WangY., YuanY., BiY., LiG., WangH., YanP., and LvG. (2017). The role of netrin–1 in improving functional recovery through autophagy stimulation following spinal cord injury in rats. Front. Cell Neurosci. 11, 350.
45.
ZhaoH., ChenS., GaoK., ZhouZ., WangC., ShenZ., GuoY., LiZ., WanZ., LiuC., and MeiX. (2017). Resveratrol protects against spinal cord injury by activating autophagy and inhibiting apoptosis mediated by the SIRT1/AMPK signaling pathway. Neuroscience, 348, 241–251.
46.
LiH.T., ZhaoX.Z., ZhangX.R., LiG., JiaZ.Q., SunP., WangJ.Q., FanZ.K., and LvG. (2016). Exendin-4 enhances motor function recovery via promotion of autophagy and inhibition of neuronal apoptosis after spinal cord injury in rats. Mol. Neurobiol. 53, 4073–4082.
47.
WangP., LinC., WuS., HuangK., WangY., BaoX., ZhangF., HuangZ., and TengH. (2017). Inhibition of autophagy is involved in the protective effects of ginsenoside Rb1 on spinal cord injury. Cell Mol. Neurobiol. 38, 679–690.
48.
BisicchiaE., LatiniL., CavallucciV., SassoV., NicolinV., MolinariM., D'AmelioM., and ViscomiM.T. (2017). Autophagy inhibition favors survival of rubrospinal neurons after spinal cord hemisection. Mol. Neurobiol. 54, 4896–4907.
49.
WuF., WeiX., WuY., KongX., HuA., TongS., LiuY., GongF., XieL., ZhangJ., XiaoJ., and ZhangH. (2018). Chloroquine promotes the recovery of acute spinal cord injury by inhibiting autophagy-associated inflammation and endoplasmic reticulum stress. J. Neurotrauma, 35, 1329–1344.
50.
MoscatJ., and Diaz–MecoM.T. (2009). p62 at the crossroads of autophagy, apoptosis, and cancer. Cell, 137, 1001–1004.
51.
SardielloM. (2016). Transcription factor EB: from master coordinator of lysosomal pathways to candidate therapeutic target in degenerative storage diseases. Ann. N. Y. Acad. Sci. 1371, 3–14.
52.
SardielloM., PalmieriM., di RonzaA., MedinaD.L., ValenzaM., GennarinoV.A., Di MaltaC., DonaudyF., EmbrioneV., PolishchukR.S., BanfiS., ParentiG., CattaneoE., and BallabioA. (2009). A gene network regulating lysosomal biogenesis and function. Science, 325, 473–477.
53.
HeX., YuanW., LiZ., and FengJ. (2017). An autophagic mechanism is involved in the 6-hydroxydopamine-induced neurotoxicity in vivo. Toxicol. Lett. 280, 29–40.
