Skip to main content

Advertisement

SpringerLink
Log in

Tetrahydrocurcumin Ameliorates Homocysteinylated Cytochrome-c Mediated Autophagy in Hyperhomocysteinemia Mice after Cerebral Ischemia

  • Published:
Journal of Molecular Neuroscience Aims and scope Submit manuscript

Abstract

High levels of homocysteine (Hcy) known as hyperhomocysteinemia (HHcy), contribute to autophagy and ischemia/reperfusion injury (I/R). Previous studies have shown that I/R injury and HHcy cause increased cerebrovascular permeability; however, the associated mechanism remains obscure. Interestingly, during HHcy, cytochome-c becomes homocysteinylated (Hcy-cyto-c). Cytochrome-c (cyto-c) transports electrons and facilitates bioenergetics in the system. However, its role in autophagy during ischemia/reperfusion injury is unclear. Tetrahydrocurcumin (THC) is a major herbal antioxidant and anti-inflammatory agent. Therefore, the objective of this study was to determine whether THC ameliorates autophagy during ischemia/reperfusion injury by reducing homocysteinylation of cyto-c in hyperhomocysteinemia pathological condition. To test this hypothesis, we employed 8–10-week-old male cystathionine-beta-synthase heterozygote knockout (CBS+/−) mice (genetically hyperhomocystemic mice). Experimental group was: CBS+/−, CBS+/− + THC (25 mg/kg in 0.1% DMSO dose); CBS (+/−)/I/R, and CBS (+/−)/I/R + THC (25 mg/kg in 0.1% DMSO dose). Ischemia was performed for 30 min and reperfusion for 72 h. THC was injected intra-peritoneally (I.P.) once daily for a period of 3 days after 30 min of ischemia. The infarct area was measured using 2,3,5-triphenyltetrazolium chloride staining. Permeability was determined by brain edema and Evans Blue extravasation. The brain tissues were analyzed for oxidative stress, matrix metalloproteinase-9 (MMP-9), damage-regulated autophagy modulator (DRAM), and microtubule-associated protein 1 light chain 3 (LC3) by Western blot. The mRNA levels of S-adenosyl-l-homocysteine hydrolases (SAHH) and methylenetetrahydrofolate reductase (MTHFR) genes were measured by quantitative real-time polymerase chain reaction. Co-immunoprecipitation was used to determine the homocysteinylation of cyto-c. We found that brain edema and Evans Blue leakage were reduced in I/R + THC-treated groups as compared to sham-operated groups along with reduced brain infarct size. THC also decreased oxidative damage and ameliorated the homocysteinylation of cyto-c in-part by MMP-9 activation which leads to autophagy in I/R groups as compared to sham-operated groups. This study suggests a potential therapeutic role of dietary THC in cerebral ischemia.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  • Abe K, Yuki S, Kogure K (1988) Strong attenuation of ischemic and postischemic brain edema in rats by a novel free radical scavenger. Stroke 19:480–485

    Article  PubMed  CAS  Google Scholar 

  • Adhami F, Liao G, Morozov YM, Schloemer A, Schmithorst VJ, Lorenz JN, Dunn RS, Vorhees CV, Wills-Karp M, Degen JL, Davis RJ, Mizushima N, Rakic P, Dardzinski BJ, Holland SK, Sharp FR, Kuan CY (2006) Cerebral ischemia-hypoxia induces intravascular coagulation and autophagy. Am J Pathol 169:566–583

    Article  PubMed  CAS  Google Scholar 

  • Adhami F, Schloemer A, Kuan CY (2007) The roles of autophagy in cerebral ischemia. Autophagy 3:42–44

    PubMed  CAS  Google Scholar 

  • Asahi M, Asahi K, Jung JC, del Zoppo GJ, Fini ME, Lo EH (2000) Role for matrix metalloproteinase 9 after focal cerebral ischemia: effects of gene knockout and enzyme inhibition with BB-94. J Cereb Blood Flow Metab 20:1681–1689

    Article  PubMed  CAS  Google Scholar 

  • Asahi M, Wang X, Mori T, Sumii T, Jung JC, Moskowitz MA, Fini ME, Lo EH (2001) Effects of matrix metalloproteinase-9 gene knock-out on the proteolysis of blood–brain barrier and white matter components after cerebral ischemia. J Neurosci 21:7724–7732

    PubMed  CAS  Google Scholar 

  • Beard RS Jr, Bearden SE (2011) Vascular complications of cystathionine beta-synthase deficiency: future directions for homocysteine-to-hydrogen sulfide research. Am J Physiol Heart Circ Physiol 300:H13–H26

    Article  PubMed  CAS  Google Scholar 

  • Bederson JB, Pitts LH, Germano SM, Nishimura MC, Davis RL, Bartkowski HM (1986) Evaluation of 2,3,5-triphenyltetrazolium chloride as a stain for detection and quantification of experimental cerebral infarction in rats. Stroke 17:1304–1308

    Article  PubMed  CAS  Google Scholar 

  • Belayev L, Busto R, Zhao W, Ginsberg MD (1996) Quantitative evaluation of blood–brain barrier permeability following middle cerebral artery occlusion in rats. Brain Res 739:88–96

    Article  PubMed  CAS  Google Scholar 

  • Chen H, Yoshioka H, Kim GS, Jung JE, Okami N, Sakata H, Maier CM, Narasimhan P, Goeders CE, Chan PH (2011) Oxidative stress in ischemic brain damage: mechanisms of cell death and potential molecular targets for neuroprotection. Antioxid Redox Signal 14:1505–1517

    Article  PubMed  CAS  Google Scholar 

  • Damdimopoulos AE, Miranda-Vizuete A, Pelto-Huikko M, Gustafsson JA, Spyrou G (2002) Human mitochondrial thioredoxin. Involvement in mitochondrial membrane potential and cell death. J Biol Chem 277:33249–33257

    Article  PubMed  CAS  Google Scholar 

  • Dennis PB, Mercer CA (2009) The GST-BHMT assay and related assays for autophagy. Methods Enzymol 452:97–118

    Article  PubMed  CAS  Google Scholar 

  • Finkelstein JD (1998) The metabolism of homocysteine: pathways and regulation. Eur J Pediatr 157(Suppl 2):S40–S44

    Article  PubMed  CAS  Google Scholar 

  • Fujimura M, Gasche Y, Morita-Fujimura Y, Massengale J, Kawase M, Chan PH (1999) Early appearance of activated matrix metalloproteinase-9 and blood–brain barrier disruption in mice after focal cerebral ischemia and reperfusion. Brain Res 842:92–100

    Article  PubMed  CAS  Google Scholar 

  • Fukui S, Fazzina G, Amorini AM, Dunbar JG, Marmarou A (2003) Differential effects of atrial natriuretic peptide on the brain water and sodium after experimental cortical contusion in the rat. J Cereb Blood Flow Metab 23:1212–1218

    Article  PubMed  CAS  Google Scholar 

  • Gasche Y, Copin JC, Sugawara T, Fujimura M, Chan PH (2001) Matrix metalloproteinase inhibition prevents oxidative stress-associated blood–brain barrier disruption after transient focal cerebral ischemia. J Cereb Blood Flow Metab 21:1393–1400

    Article  PubMed  CAS  Google Scholar 

  • Gasche Y, Fujimura M, Morita-Fujimura Y, Copin JC, Kawase M, Massengale J, Chan PH (1999) Early appearance of activated matrix metalloproteinase-9 after focal cerebral ischemia in mice: a possible role in blood–brain barrier dysfunction. J Cereb Blood Flow Metab 19:1020–1028

    Article  PubMed  CAS  Google Scholar 

  • Jakubowski H (2006) Pathophysiological consequences of homocysteine excess. J Nutr 136:1741S–1749S

    PubMed  CAS  Google Scholar 

  • Jakubowski H (2007) The molecular basis of homocysteine thiolactone-mediated vascular disease. Clin Chem Lab Med 45:1704–1716

    Article  PubMed  CAS  Google Scholar 

  • Jakubowski H (2008) The pathophysiological hypothesis of homocysteine thiolactone-mediated vascular disease. J Physiol Pharmacol 59(Suppl 9):155–167

    PubMed  Google Scholar 

  • Jiang J, Wang W, Sun YJ, Hu M, Li F, Zhu DY (2007) Neuroprotective effect of curcumin on focal cerebral ischemic rats by preventing blood–brain barrier damage. Eur J Pharmacol 561:54–62

    Article  PubMed  CAS  Google Scholar 

  • Kleinschnitz C, Schwab N, Kraft P, Hagedorn I, Dreykluft A, Schwarz T, Austinat M, Nieswandt B, Wiendl H, Stoll G (2010) Early detrimental T-cell effects in experimental cerebral ischemia are neither related to adaptive immunity nor thrombus formation. Blood 115:3835–3842

    Article  PubMed  CAS  Google Scholar 

  • Kraus JP, Janosik M, Kozich V, Mandell R, Shih V, Sperandeo MP, Sebastio G, de Franchis R, Andria G, Kluijtmans LA, Blom H, Boers GH, Gordon RB, Kamoun P, Tsai MY, Kruger WD, Koch HG, Ohura T, Gaustadnes M (1999) Cystathionine beta-synthase mutations in homocystinuria. Hum Mutat 13:362–375

    Article  PubMed  CAS  Google Scholar 

  • Kumar M, Tyagi N, Moshal KS, Sen U, Kundu S, Mishra PK, Givvimani S, Tyagi SC (2008) Homocysteine decreases blood flow to the brain due to vascular resistance in carotid artery. Neurochem Int 53:214–219

    Article  PubMed  CAS  Google Scholar 

  • Lee JE, Yoon YJ, Moseley ME, Yenari MA (2005) Reduction in levels of matrix metalloproteinases and increased expression of tissue inhibitor of metalloproteinase-2 in response to mild hypothermia therapy in experimental stroke. J Neurosurg 103:289–297

    Article  PubMed  CAS  Google Scholar 

  • Lee SR, Tsuji K, Lee SR, Lo EH (2004) Role of matrix metalloproteinases in delayed neuronal damage after transient global cerebral ischemia. J Neurosci 24:671–678

    Article  PubMed  CAS  Google Scholar 

  • Lominadze D, Roberts AM, Tyagi N, Moshal KS, Tyagi SC (2006) Homocysteine causes cerebrovascular leakage in mice. Am J Physiol Heart Circ Physiol 290:H1206–H1213

    Article  PubMed  CAS  Google Scholar 

  • Longa EZ, Weinstein PR, Carlson S, Cummins R (1989) Reversible middle cerebral artery occlusion without craniectomy in rats. Stroke 20:84–91

    Article  PubMed  CAS  Google Scholar 

  • Manolescu BN, Oprea E, Farcasanu IC, Berteanu M, Cercasov C (2010) Homocysteine and vitamin therapy in stroke prevention and treatment: a review. Acta Biochim Pol 57:467–477

    PubMed  CAS  Google Scholar 

  • Mishra PK, Givvimani S, Metreveli N, Tyagi SC (2010) Attenuation of beta2-adrenergic receptors and homocysteine metabolic enzymes cause diabetic cardiomyopathy. Biochem Biophys Res Commun 401:175–181

    Article  PubMed  CAS  Google Scholar 

  • Mun-Bryce S, Rosenberg GA (1998) Matrix metalloproteinases in cerebrovascular disease. J Cereb Blood Flow Metab 18:1163–1172

    Article  PubMed  CAS  Google Scholar 

  • Nappo F, De RN, Marfella R, De LD, Ingrosso D, Perna AF, Farzati B, Giugliano D (1999) Impairment of endothelial functions by acute hyperhomocysteinemia and reversal by antioxidant vitamins. JAMA 281:2113–2118

    Article  PubMed  CAS  Google Scholar 

  • Nishiyama T, Mae T, Kishida H, Tsukagawa M, Mimaki Y, Kuroda M, Sashida Y, Takahashi K, Kawada T, Nakagawa K, Kitahara M (2005) Curcuminoids and sesquiterpenoids in turmeric (Curcuma longa L.) suppress an increase in blood glucose level in type 2 diabetic KK-Ay mice. J Agric Food Chem 53:959–963

    Article  PubMed  CAS  Google Scholar 

  • Obeid R, Herrmann W (2006) Mechanisms of homocysteine neurotoxicity in neurodegenerative diseases with special reference to dementia. FEBS Lett 580:2994–3005

    Article  PubMed  CAS  Google Scholar 

  • Okada K, Wangpoengtrakul C, Tanaka T, Toyokuni S, Uchida K, Osawa T (2001) Curcumin and especially tetrahydrocurcumin ameliorate oxidative stress-induced renal injury in mice. J Nutr 131:2090–2095

    PubMed  CAS  Google Scholar 

  • Pan MH, Huang TM, Lin JK (1999) Biotransformation of curcumin through reduction and glucuronidation in mice. Drug Metab Dispos 27:486–494

    PubMed  CAS  Google Scholar 

  • Pari L, Murugan P (2004) Protective role of tetrahydrocurcumin against erythromycin estolate-induced hepatotoxicity. Pharmacol Res 49:481–486

    Article  PubMed  CAS  Google Scholar 

  • Perla-Kajan J, Marczak L, Kajan L, Skowronek P, Twardowski T, Jakubowski H (2007) Modification by homocysteine thiolactone affects redox status of cytochrome c. Biochemistry 46:6225–6231

    Article  PubMed  CAS  Google Scholar 

  • Perla-Kajan J, Stanger O, Luczak M, Ziolkowska A, Malendowicz LK, Twardowski T, Lhotak S, Austin RC, Jakubowski H (2008) Immunohistochemical detection of N-homocysteinylated proteins in humans and mice. Biomed Pharmacother 62:473–479

    Article  PubMed  CAS  Google Scholar 

  • Qipshidze N, Tyagi N, Sen U, Givvimani S, Metreveli N, Lominadze D, Tyagi SC (2010) Folic acid mitigated cardiac dysfunction by normalizing the levels of tissue inhibitor of metalloproteinase and homocysteine-metabolizing enzymes postmyocardial infarction in mice. Am J Physiol Heart Circ Physiol 299:H1484–H1493

    Article  PubMed  CAS  Google Scholar 

  • Rami A, Langhagen A, Steiger S (2008) Focal cerebral ischemia induces upregulation of Beclin 1 and autophagy-like cell death. Neurobiol Dis 29:132–141

    Article  PubMed  CAS  Google Scholar 

  • Reggiori F, Klionsky DJ (2002) Autophagy in the eukaryotic cell. Eukaryot Cell 1:11–21

    Article  PubMed  CAS  Google Scholar 

  • Romanic AM, White RF, Arleth AJ, Ohlstein EH, Barone FC (1998) Matrix metalloproteinase expression increases after cerebral focal ischemia in rats: inhibition of matrix metalloproteinase-9 reduces infarct size. Stroke 29:1020–1030

    Article  PubMed  CAS  Google Scholar 

  • Rosenberg GA, Estrada EY, Dencoff JE (1998) Matrix metalloproteinases and TIMPs are associated with blood–brain barrier opening after reperfusion in rat brain. Stroke 29:2189–2195

    Article  PubMed  CAS  Google Scholar 

  • Selhub J (1999) Homocysteine metabolism. Annu Rev Nutr 19:217–246

    Article  PubMed  CAS  Google Scholar 

  • Sen U, Herrmann M, Herrmann W, Tyagi SC (2007) Synergism between AT1 receptor and hyperhomocysteinemia during vascular remodeling. Clin Chem Lab Med 45:1771–1776

    Article  PubMed  CAS  Google Scholar 

  • Strickland S, Gualandris A, Rogove AD, Tsirka SE (1996) Extracellular proteases in neuronal function and degeneration. Cold Spring Harb Symp Quant Biol 61:739–745

    Article  PubMed  CAS  Google Scholar 

  • Tejima E, Zhao BQ, Tsuji K, Rosell A, van Leyen K (2007) Astrocytic induction of matrix metalloproteinase-9 and edema in brain hemorrhage. J Cereb Blood Flow Metab 27:460–468

    Article  PubMed  CAS  Google Scholar 

  • Thiyagarajan M, Sharma SS (2004) Neuroprotective effect of curcumin in middle cerebral artery occlusion induced focal cerebral ischemia in rats. Life Sci 74:969–985

    Article  PubMed  CAS  Google Scholar 

  • Tyagi N, Givvimani S, Qipshidze N, Kundu S, Kapoor S, Vacek JC, Tyagi SC (2010) Hydrogen sulfide mitigates matrix metalloproteinase-9 activity and neurovascular permeability in hyperhomocysteinemic mice. Neurochem Int 56:301–307

    Article  PubMed  CAS  Google Scholar 

  • Tyagi N, Moshal KS, Ovechkin AV, Rodriguez W, Steed M, Henderson B, Roberts AM, Joshua IG, Tyagi SC (2005a) Mitochondrial mechanism of oxidative stress and systemic hypertension in hyperhomocysteinemia. J Cell Biochem 96:665–671

    Article  PubMed  CAS  Google Scholar 

  • Tyagi N, Ovechkin AV, Lominadze D, Moshal KS, Tyagi SC (2006) Mitochondrial mechanism of microvascular endothelial cells apoptosis in hyperhomocysteinemia. J Cell Biochem 98:1150–1162

    Article  PubMed  CAS  Google Scholar 

  • Tyagi N, Sedoris KC, Steed M, Ovechkin AV, Moshal KS, Tyagi SC (2005b) Mechanisms of homocysteine-induced oxidative stress. Am J Physiol Heart Circ Physiol 289:H2649–H2656

    Article  PubMed  CAS  Google Scholar 

  • Utepbergenov DI, Mertsch K, Sporbert A, Tenz K, Paul M, Haseloff RF, Blasig IE (1998) Nitric oxide protects blood–brain barrier in vitro from hypoxia/reoxygenation-mediated injury. FEBS Lett 424:197–201

    Article  PubMed  CAS  Google Scholar 

  • Uyama O, Okamura N, Yanase M, Narita M, Kawabata K, Sugita M (1988) Quantitative evaluation of vascular permeability in the gerbil brain after transient ischemia using Evans blue fluorescence. J Cereb Blood Flow Metab 8:282–284

    Article  PubMed  CAS  Google Scholar 

  • Veltkamp R, Siebing DA, Sun L, Heiland S, Bieber K, Marti HH, Nagel S, Schwab S, Schwaninger M (2005) Hyperbaric oxygen reduces blood–brain barrier damage and edema after transient focal cerebral ischemia. Stroke 36:1679–1683

    Article  PubMed  Google Scholar 

  • Ventruti A, Cuervo AM (2007) Autophagy and neurodegeneration. Curr Neurol Neurosci Rep 7:443–451

    Article  PubMed  CAS  Google Scholar 

  • Wang J, Tsirka SE (2005) Neuroprotection by inhibition of matrix metalloproteinases in a mouse model of intracerebral haemorrhage. Brain 128:1622–1633

    Article  PubMed  Google Scholar 

  • Wang X, Jung J, Asahi M, Chwang W, Russo L, Moskowitz MA, Dixon CE, Fini ME, Lo EH (2000) Effects of matrix metalloproteinase-9 gene knock-out on morphological and motor outcomes after traumatic brain injury. J Neurosci 20:7037–7042

    PubMed  CAS  Google Scholar 

  • Watanabe M, Osada J, Aratani Y, Kluckman K, Reddick R, Malinow MR, Maeda N (1995) Mice deficient in cystathionine beta-synthase: animal models for mild and severe homocyst(e)inemia. Proc Natl Acad Sci U S A 92:1585–1589

    Article  PubMed  CAS  Google Scholar 

  • Xue M, Hollenberg MD, Yong VW (2006) Combination of thrombin and matrix metalloproteinase-9 exacerbates neurotoxicity in cell culture and intracerebral hemorrhage in mice. J Neurosci 26:10281–10291

    Article  PubMed  CAS  Google Scholar 

  • Yodkeeree S, Chaiwangyen W, Garbisa S, Limtrakul P (2009) Curcumin, demethoxycurcumin and bisdemethoxycurcumin differentially inhibit cancer cell invasion through the down-regulation of MMPs and uPA. J Nutr Biochem 20:87–95

    Article  PubMed  CAS  Google Scholar 

  • Yodkeeree S, Garbisa S, Limtrakul P (2008) Tetrahydrocurcumin inhibits HT1080 cell migration and invasion via downregulation of MMPs and uPA. Acta Pharmacol Sin 29:853–860

    Article  PubMed  CAS  Google Scholar 

  • Zhao BQ, Tejima E, Lo EH (2007) Neurovascular proteases in brain injury, hemorrhage and remodeling after stroke. Stroke 38:748–752

    Article  PubMed  CAS  Google Scholar 

  • Zhao J, Zhu W, Liu T, Yang J, Li G (2010) Electrochemical probing into cytochrome c modification with homocysteine-thiolactone. Anal Bioanal Chem 397:695–701

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

This work was supported by NIH grants: HL- 71010, NS-51568 to SCT and HL-107640 to NT.

Rationale

Cystathionine-beta synthase deficient homozygous mice suffer from severe growth retardation and a majority of them are dead by 5 weeks of age. Histological examination showed that the hepatocytes of homozygotes were enlarged, multinucleated, and filled with microvesicular lipid droplets. Plasma homocysteine levels of the homozygotes were approximately 40 times normal. Heterozygous mutants have an approximately 50% reduction in cystathionine beta-synthase mRNA and enzyme activity and have twice normal plasma homocysteine levels. Thus, the heterozygous mutants are promising for studying the in vivo role of elevated levels of homocysteine.

Author information

Authors and Affiliations

  1. Department of Physiology and Biophysics, School of Medicine, Health Sciences Center, A-1115, University of Louisville, Louisville, KY, 40202, USA

    Neetu Tyagi, Natia Qipshidze, Charu Munjal, Jonathan C. Vacek, Naira Metreveli, Srikanth Givvimani & Suresh C. Tyagi

Authors
  1. Neetu Tyagi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Natia Qipshidze
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Charu Munjal
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jonathan C. Vacek
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Naira Metreveli
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Srikanth Givvimani
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Suresh C. Tyagi
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Neetu Tyagi.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Tyagi, N., Qipshidze, N., Munjal, C. et al. Tetrahydrocurcumin Ameliorates Homocysteinylated Cytochrome-c Mediated Autophagy in Hyperhomocysteinemia Mice after Cerebral Ischemia. J Mol Neurosci 47, 128–138 (2012). https://doi.org/10.1007/s12031-011-9695-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12031-011-9695-z

Keywords

Search

Navigation