Agaricus Blazei Murill Ameliorates Myocardial Ischemia-Reperfusion Injury · PDF file 2011....

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Transcript of Agaricus Blazei Murill Ameliorates Myocardial Ischemia-Reperfusion Injury · PDF file 2011....

  • |叫.一ticle 叫[1J!I"li!ti1.. 11lI川:“:235-41

    Basic Scicnιe

    Agaricus Blazei Murill Ameliorates Myocardial Ischemia-Reperfusion Injury

    Chicl1 -j-lu(I /-I11011g. U Hon- Ni TS lI Î , .I Ren-Hu ll' P.的嗨• I }'i- /-Isua l1 Hsie !J, 3 Wοei-.Jer CI1IIong.J../ Cα-Chillg HSlf , 可 I'Velll叫 HlI{lI1g~.4. 1I1ld I'Ven-.Jone Clrel1 l.~ 均

    8ackgrolllld: AgariclIs bla~ei MlIri ll (ABM) is a widcly acceptcd heallh food and is known 10 prescnt anli-ll1 morigenic

    activities . ABM has also been rcported 10 11l0dll late oxidati ve strcss aner inJl a111111a tion and infec tion. Reactive

    oxygen fl閃閃dica l s havc been shown to bc criti ca ll y in vo lved in i schc l1l ia-rcpcr仇, sion injllry cascadcs. We ail1lcd to

    stlldy Ihe antioxidant activ圳的 ofABM aga inst tiss lIc injllry causcd by 111yO巳ard i a l ischcmia- repcrfllsion proccsses

    MethOlls: A rat card io l1lyocy tc H9c2 ccl l line was sct up and lIsed for thc stlldy. Cc ll viabi li tics afler di仟erent

    dosages 0 1' ABM ex tracl trcat l11cnt and hydrogcn peroxidc injuries wc rc c he氾kcd . A card iac ische l1l ia-rcpcrfllsion

    injllry modcl was set lIP to elllcidate the cardioprotcctive roles 0 1" ABM cxtrac t. Diffcrcnt doses of 22.5 I11 g/kg of

    body wcighl and 45 I1lg/k且 of body wc ight AB M cxlraCI wcrc p悶-Ircatcd al 24 and 4日 hO llrs bctòre indllcing the ischc l11 ia-rcpcrl"lIsion injllry

    Re.", lts: ABM cx tract trCatll1enl increased thc slI rviva l o f ca rdiomyocytes witholll cytotoxicily. Pret rcatment with ABM ex tract redllced hydrogen pcrox ide- indllced cc ll da mage and increased ccll slI rviva l (0 日 6 :t 0. 1 5 in ABM 6.0

    mg/ l1l l vs. 0.45 :t 0.03 in control, p = 0.0 19). ABM-pretreatcd rats lhat lInderwent myoca rdial ischcmia-rcpcrfllsion had greatl y rcdllced infarcl areas comparcd to those in thc control grollp (14.3 :t 3.3% in contro l grollp . 4.6 士 1.9% in

    22.5 111 g/kg grollp, 4.9 士 2. 1 % in 45 mg/kg grollp. p < 0.05) COll cIIlS;O Il: ABM incrcascs anlioxidant ac tivi旬, which grea tl y amc li ora tcs myoca rdial injllries callsed by

    l1lyocardi al ischc l11 ia- repcrfllsion injllrics. Us ing ABM as a hea lth food l11ay providc ca rdioprotect ivc cffect aga inst

    ischemia-rcperfll sion i 叮 uncs

    Key WO l"ds : AgariclIs h1a~ei Murill • An ti ox idant • Ischcmi a• cpcrlùsion . Myocardia l infa rcti on

    INTRODUCTION known fo r it s med ic ina l propertics in diabetcs ‘ osteopo-

    ros悶、 and ca nce r. 1.1 The ABM mushroom is anti c las- Agaric lIs bla工的 M lI rill (ABM) , an edi bl e Basid i o-ωogeni c in m刊I ce W I川1叫山h t仙u l11 0 r xe釘enog♂ra ft心s

    l11ycota r,心1I叫11川ngll s f,ωOll叫l叮II1 d pril川maril抄Y 11川n SOllt仙h America , i的s well ca別te的5 t山h a削t ABM i陌s t仙u叫lmor i cida l. AB M also like ly plays a

    ro le in preventing tumor initiation by reducing geno-

    Rece ived: JanLHlly 7. 20 10 Accepled: .I unc 2X. 20 10 I Depart mcnt 01' Emc l 且c l1 cy Mcdicinc; 2Dcp i:l rt Il1 Ci11 of Int crn al Mcdicinc司 National Taiwan Un ivers ity Hospita l‘ Taipei: 3Bas ic Med ica l Sc iel時的、 Medica l Laboratory Sc iencc and ßiotechnology: -I Biochcmistl)'. Collegc ofMed icÎ nc可 Nationnl Chcng-Kul1且 Univcrs i ty. Tainan: 雪Simpso l1 B i o l cchno\o且y Il1c.. Taoyu él ll ‘ 4、 Dcpartmcn t 01 Intemal Mcdic ine可 Shin Chu Gcneral Hospilal‘ Shin Chu. Taiwan Addrcss correspondcnce and reprint reqllc心的 1 0: Dr. Wcn- .I ol1e Chcl1 , Depanment of Emcr且cncy Medicinc. Nat Îona l Tai wan University Hospilal , No. 7‘ Zhollgshall S. Rd ‘ Taipci City 100. Ta iwall. Tel : XX6- 2-2356-2831 ‘ Fax: 886-2-2322-31 50: E-mai l: wjchcI1 1 955(ç~lllu.cdll. tw The two cO lTcsponding au t l叮叮 contribll tcd eq llally

    235

    tox ic itics ca used by DNA-damaging agents5 ABM has

    a lso becn shown to cnhance innatc as we ll as adapt ivc 6.7 i l11 mune rcsponses in l11 ice.".' It st imulates the prod uc-

    tion of inter le ll kin and cyto kin的 , which inc rcase host

    immuniti cs against bacteria l and viral infect ions. S. 'O It

    also induccs inter leukin-12- and intcrfero n-y-mcdiated

    natura l kill er cell ac tivation ." These find ings inva ri ab ly

    suggest that A BM is a promising “hea lth food" thal pro- tects hos ts from infcctions and cancers. O nc recent

    study' l reported that ABM prevented di ethylnit ros3m ine,

    AC(fI Cflrdiol Sill 2010:2 f1:l35- 41

  • Chien-Hlla Hllang et al

    a strong hepatic cytotoxin and genotoxin,丘。 111 c3 11 sing fied Eagle l11 edill m (DMEM ) sllpp lemented with 10% fe-

    liver ce ll injury; thi s indicated thal ABM helps maintain tal bov ine serum (FBS), 2 111M L-g lll ta l11 ine, 1 x non-

    tissue in tegrily. The deh ydrated ABM fruct ifica tion body essent ia l amino ac id , and 1 x antibiotic/anti l11 ycotic

    is ri ch in proteins and carboh ydrates. The po lysaccharide mixtllre (Gibco BRL, Grand Island , NY). 執le grcw lhe cOl11ponent,但-glllcans , i 日 key to the l11 edicina l effect s of ce ll s at 37 oC in a 5% CO2 的mosphere. ABM. 1 The ß-glucans exlracted from ABM 1110dll laled T he dried ABM powdcr was offered by Si l11 pson

    im l11 une responses by stimu lati ng the re lease of cylo- Biolechnologies, lnc . (Taoyuan, Taiwan). We mixed 2.5 kines , which comprise ABM's il11l11 une regu latory eι grams of ABM powder with 100 mL of phosphate bllι fects U fered sa li ne (PBS) [pH 7.4] and shook it at room temper-

    Two parallel but somew hal contradictory immune alure for 1 h. The ABM so luti on was filt ered us in g

    reg lllato ry effec ts of ABM have been reported . O ne Whatman No. 1 filter pape r. The filtra惚 , which com-

    stlld/ reported that, in macrophages , ABM stimu lated prised the water-soluble cO l11ponents of ABM、 was ster- nitri c oxide production and increased infla l11 mati on, i1 ized by pass ing il through a 0. 22-~lm-sy rin ge fi lter. It which creates cell ular oxidative stress; a second stlldy,14 was then slored al -20 oC unti l ll se.

    in li ver and lung ce ll s, reported that ABM redllced the

    prod llction of reacti ve oxygen species (ROS) and main- MTT assay

    tained DNA and ce ll ul ar integrity. The mo lecular mecha- A 3-阱, 5-d imethylthiazo l-2-y 1 ] 孔, 5-diphenyl l e tn卜

    ni sms to reconcile these two di stinc t pheno l11ena have zo liu l11 bro l11 ide (MTT) ce ll pro liferation assay was l1sed

    not yct been identified . However, ABM l11ay affect ox i- to detect ce ll viability by eva lllating the l11itochondrial

    da ti ve responses in different ti ss ll e-types in di stinct redllctase ac ti vity after treatments with ABM extract and

    ways. hyd rogen pe rox ide. The protoco ls bas ica ll y fo ll owed

    In lhe presenl s rudy, we wanted to kno w whelher those previously described 22 Brie fl y, lhe ce ll s that were ABM is an anti oxidant in the card iac sys tem and g ro wn to 5 x 104/we ll in a 24-we ll clllture pl ale we re

    whether it protec ts cardiomyocytes [rom permanent trealed with vario l1 s doses of ABM exlrac l for 24 h al

    damage callsed by aCllte myoca rdial infarction, which 37 oC and then treated with 100μM hydrogen peroxide

    know whether ABM protecls cardiomyocytes [rom the Intracellular reactive oxygen species (ROS)

    ox idative stress callsed by [巴perflls i on therapy. measurement

    lntracellu la r ROS level was sludi ed by measuring

    the changes in flllorescence resulting from intrace ll ulal

    MATERIALS AND METHODS probe ox idation. The probe 2' ,7'-d ich loroflllorescin dia-

    cetate (DCFH -DA、 5.0 mM 、 S i gma , St. LOll is , MO ‘

    Cell lines and the ABM mushroom U.S.A.) entered the ce ll and th e acetate g ro l1 p on

    We used rat cardiomyocy le H9c2 cell s for thi s study. DC FH-DA was c leaved by cell l1 lar es te ra ses. S l1 bsc-

    The cell s were 111aintained in reglllar Dulbecco 's mod i- quent oxidation by ROS , particu larly hydroxyl radica l 、

    ACfα C(I川Iiο1 Sil/ 2010:26:235-41 236

  • Antioxidant Activity in AgariclIs B/azei Muri ll

    y ic lded thc fluo rescent prodllc t DCF. AB M cx trac t- complele ly liga ting the margi na l coronary a rtery fo r 30

    lrcated and contro ll ed 1-1 9c2 ce ll s ( 1 x 10") wc rc loaded min . After 30 min 0 1' isc hem ia, the S UI 且ica l li e was rc- w ith 10 ml DCF I-I -DA at 4 ho urs a fte r treatmenl w ith leased and the ischcmic myocardium was reperfused fo r

    1-1 20 2, and ana lyzed by f ACSca n ( Bec to n Di ck in son 2.5 h . Myoca rd ial ischemia was con fÏ rmed visua ll y by H e i de l be l 且, Gerl1lany) . reg iona l cyanos is of the myocard ia l Sllrface.

    Th e infa rct s ize was de termined as prev io us ly

    Cell damage assays by flow cytometry analysis dcscr ibed 24 us ing 1% tripheny l-tetrazo lium ch lo ride

    Damagcd ce ll s were detected by pro pid ill l1l iod ide (S igma-Aldrich, U.S.) in phosphate bll ffer [pH 7.4]. The

    (叭 , 50 I1lg/ l1l l) afte r trcatment with 1-1 20 2 fo r 24 h as de- lell ventri cul ar area at risk and the area 0 1' in fa rcted ti s- sc ribed prcvio us ly. 23 Labe ling was pc rlo r l1led with the s lIe were measured by an independent, blindcd obsc rver

    sta ining ana lyzed on a FACScan (Becton Dickinson He i- using CO ll1pllter plani l1letry.

    dc l berg、 Germany). When nuclear membrane d isrllpted,

    which is a s ign of advanced ce ll dama且ed , the 1'1 we nt Statistical analysis

    into thc n