Tgp chi Cdng ngheSinh hgc 11(2): 203-210, 2013
X A C D I N H D A H I N H N U C L E O T I D E D O N T R O N G V U N G P R O M O T E R D O A N G E N M A T R I X M E T A L L O P R O T E I N A S E - 9 Of B E N H N H A N H Q I C H l f N G M A C H V A N H C A P Do Hihi Chi', Le Thi Bich Thao', Nguyen Tiin Dung', Pham Dure Dan', Bui Thj Huyin', Nguyen Thi Minh Phirong', Do Doan Lgi^ Nguyin Bich Nhi', Phan Van Clii'
Vien Cong nghe sinh hgc, Vien Hdn ldm Khoa hoc vd Cdng nghe Viet Nam Vi4n Tim mach, Benh vien Bgch Mai
TOM T A T
Theo thong ke ciia Hiep hoi tim mach the giai (World Heart Federation), hang nam cac benh ly ve tim mach da cirop di mang song ciia khoang 17,5 trieu nguoi tren toan the gioi. Hoi chimg mach vanh cap hien dang la van de sue khoe rat dugc quan tam tren ihe gioi do benh thucmg de doa tinh mang con nguoi mot each dot nggt, ly Ie tir vong cao, di chirng nang ne cho moi ca nhan, gia dinh va loan xa hpi, tham chi dang co xu huong tre hoa do tuoi nguai mac can benh nguy hiem nay. Gan day, cac nghien ciiu da tim ra moi lien quan di truyen giua kieu hinh trong cac gen chi thi voi chimg vOa xo dpng mach. Nhom gen matrix melalloproleinase (MMPs) la mpt trong nhirng he thong gen chi thi tiem nang: trong do, MMP-9 dupc xem la yeu to co vai tro quan trpng trong qua trinh suy giam chat nen ngoai bao (matrix extracellular) va su tai tao cac mang xo viia voi co chat chinh la collagen lyp IV va elastin. Cho den nay, day la gen dugc nghien cuu nhieu nhat trong nhom gen MMPs lien quan den hpi chirng mach vanh cap. Trong nghien ciru nay, chimg toi xac dinh da hinh viing promoter doan gen MMP-9 (tiJ vi tri 1809 toi 1374) cua 41 ca the ngudi Viet Nam bao gom 9 mau nguai binh thucmg khoe manh, 14 mau benh nhan dau that nguc khong on dinh va 18 mau benh nhan nhoi mau co tim cap. Ket qua cho thay co mot da hinh nucleotide dan a vi tri -1562C>T trong vimg promoter ciia doan gen MMP-9 a 3 benh nhan dau that nguc khong on dinh va 7 benh nhan nhoi mau co tim. Day la nhirng dan lieu dau tien ve irinh tu doan gen MMP-9 cua mot so ca the nguoi Viet Nam mac hpi chiing mach vanh cap.
Tirkhoa: Da hinb, hoi chirng mach vanh cdp. n lallopn e-9. nucleotide, pnimoier
MO DAU
Hpi chirng mach vanh cap (Acute Coronary Syndrome, ACS) van dang la nguyen nhan tir vong hang dau tren toan the gioi: iheo nhirng bao cao moi day tai Hoa Ky, hang nam co hon 90 nghin nguoi tir vong vi benh nay. Hgi chung mach vanh cung la dang benh pho bien nhat trong cac benh lien quan den tim mach a Chau Au. Tai Phap hien co hon 3 tneu nguoi mac benh va moi nam co hon 50 nghin ca tii vong. Tai Viet Nam, Iheo thong ke ciia Hoi Tim mach hpc Viet Nam (6/2010) cii 3 nguoi Viet Nam irucmg thanh thi co I nguai co nguy co mac b?nh tim mach, chii yeu la hgi chirng mach vanh cap (HCMVC)-mQl benh man tinh vai nhiing dot tien triln cSp tinh ma khi do, co the co nhimg bieu hien lam sang, di?n hpc va sinh hoa khac nhau (con dau that nguc khong on dinh, nhoi mau co tim cap (NMCT) vai song Q xuyen thanh, NMCT c5p khong CO s6ng Q hay NMCT duoi ngi tam mac,,.). Nhirng
bieu hien nay hien nay dupc gpi duai mol ten chung la HCMVC vi chung deu co chung mol co so sinh ly benh hgc la su nut hoac vo ciia cac mang vira xa tren thanh dpng mach vanh, gay hep hoac tac long mach dan den mau khong luu thong dugc trong co the Viia xo dgng mach hinh thanh la do nhirng ton thuong 6 cac te bao ngi mac, lam cho cac te bao do mat chiic nang bao ve thanh mach. Ton thuong co the do anh huong cua dong mau co ap luc cao lien tuc tac dpng den nhu trong benh tang huyet ap, roi loan lipid mau hay cac yeu to mien dich Khi bi ton ihuong, cac le bao ngi mac tai cho mai kha nang liet ra prostacyclin. Tieu cau lap tire lach ra khoi dong mau de tap trung vao cho do va kel dinh lai, phong thich ra nhieu chat trong do co yeu to tang trucmg.
yeu to nay kich thich su di chuyen cac te bao co Iron 6 lop trung mac ra Icrp noi mac va phal Irien manh tai do Cac bach cau don nhan ttr dong mau ciing den ngay cho ton thuang va dupc chuyen dang thanh dai thuc bao. Cac dai thuc bao nay "an" cac LDL-C va 203
tra thanh cac "te bao bgt" tich day ma. Den khi qua tai, cac te bao nay bi v& va do cholesterol ra ngoai lam cho lop duoi npi mac day len tao nen cac vach lipid hay cac mang. xo viia dac trung ciia benh (Nguyen Quang Tuan,, 2011). Nguyen nhan ciia HCMVC chua dugc xac dinh ro rang, khi noi den nguyen nhan cua benh nguoi ta dimg den khai niem yeu to nguy ca ciia benh dpng mach vanh, do la: dp luoi, giai tinh, hiit thuoc la, tang huyet ap, tang ham lugng cholesterol trong mau, benh tieu dudng, beo phi va loi song it van dpng, Gan day, cac nghien ciiu da tim ra moi lien quan di truyen giiia kieu hinh trong cac gen chi thi vai chimg viia xo dong mach, Nhom gen malnx melalloproleinase (MMPs) la mpt trong nhiing he thong gen chi thi tiem nang (Wang el al., 201 I). MMPs la enzyme thuoc ho endopeptidases lien ket vdi kem (Zn) trong trung tam hoat dong co boat tinh phan giai protein de ngan ngira su suy giam ciia mpt so prolein d mo lien ket nhu collagen, proteoglycan va elastin (Opdenakker el al., 2001), MMPs tham gia vao trong qua trinh suy giam ciia chat nen ngoai bao d qua trinh sinh ly binh thudng ctia co the nhu su phat Irien phoi thai, sinh san, qua trinh biet hoa cua te bao, su hinh thanh mach va tai tao md,.. tang hoat dgng va bieu hien ciia MMPs cung da dugc xac dmh trong cac qua trinh benh ly nhu chirng viem khdp, cac benh dudng ho hap, tdn thuong co tim, chiing phinh mach, cac khdi u ac tinh (Creemers et al, 2001), cac benh tu mien nhu benh lupus ban dd he thdng, viem khdp dang ihap va benh da xo cimg, Hau het cac MMPs tiel ra cac protein khdng boat dpng dugc hoat hoa khi phan cat bdi cac proteinase ngoai bao. Nghien ciru d loai khi nau (rhesus macaque) cho thay cac enzyme nay gop phan tao ra lL-8 do huy dgng ciia cac lien te bao mau tao ra lir nly suong, cac nghien ciru tren chugt cung cho thay vai tro hen quan ciia MMP-9 vdi qua trinh tai tao mo. Henney va ddng tac gia nam 2000 da bao cao sy thay doi kieu gen se anh huong den su bieu hien cua MMPs va co the gdp phan vao sir xuat hien ciia benh tim mach Gen MMP-9 nSm tren nhiem sac the 20, vimg trinh lu ma hda gdm 2387 nucleotide MMP-9 cdn duoc ggi la collagenase type IV (CLG4) hay gelatinase B (GELB) la y^u td quan trong tac dgng den qua trinh suy giam collagen va sy tai tao cac mang xa vira Cac nghien ciiu dich te hgc Cling cho thay rang mirc dp bieu hien ciia MMP-9 tang len trpng tuan hoan mau d benh nhan hgi chirng mach vanh MMP-9 dugc quan tam nhu la muc lieu dieu tri hudng dich tiem nang Dya tren co sd nhirng nghien ciiu nay chimg toi da lien hanh xac dinh da hinh viing promoter doan gen MMP-9 ciia mgt s6 benh nhan mac hgi chiing mach vanh cap Day la
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nhung din lieu &i\x lien vh trinh tu doan gen MMP-9 ciia mpt s6 ca t h i ngudi Viet Nam co hgi chung mach vanh cap.
VAT LIEU V A PHU'ONG PHAP NGHIEN CUtJ V a t Ueu
Mau mau ngoai vi ngudi binh thudng dugc chgn lua theo cac tieu chi cua de tai cd hd so ro rang va dugc khang dinh la ngudi khde manh tai Vien Tim mach, Benh vien Bach Mai thong qua cac xet nghiem: (i) Cac chi lieu hda sinh huyet thanh (Glucose huyet, Cholesterol loan phan. Triglyceride, HDL-cholesterol, LDL-chplesterol, SGOT, SGPT, Creatinine mau, Troponin, CK-MB, BNP) d ngudng binh thudng; (li) Cac chi lieu huyet ap, dien tim, sieu am tim. chup X quang tim phoi d miic binh thudng.
Mau mau ngoai vi benh nhan HCMVC gdm benh nhan dau that nguc khdng dn djnh (DT) va nhoi mau ca tim cap (NM) dupc khang dinh qua tham kham lam sang, cac xet nghiem hda sinh huyet thanh va chan doan hpi chirng mach vanh cap ngoai cac chi lieu xet nghiem nhu ngudi binh thudng cdn them xet nghiem chup mach vanh Mau mau ngoai vi {60 mau) dugc lay lit mau nguyen va dugc bao quan d • 86''C chp qua trinh phan tich.
Phirong p h a p
Tdch chiit DNA long so mdu
DNA dugc chiet xuat tir cac bach cau ciia mau ngoai vi bang bd kit Gene JET^"^ Genomic DNA Purification (Fermentas, Dire) theo quy trinh ldm tat nhu sau: Cho 400fil dung djch trich ly va 20pl dung dich proteinase K vao 200pl mau loan phan, trpn deu bang may vortex lai khi dung dich ddng nh^t. U cac miu d 56''C trong 10 phut. Them 200^1 ethanol (96%) . Chuyen dung dich sang cdt tinh sach GenJET''"'^ genomic and, ly lam 8000 vdng/phiit.
Chuyen cot Igc vao dng eppendorf mdi. Them 500pl Wash Buffer 1 (da bd sung ethanol). Ly tam 1000 vdng/phiit. Loai bd djch luu thdng qua cdt. Them vao 500jil Wash Buffer II (da bd sung ethanol). Ly tam 13000 vdng/3phut. Dat cot loc vao ong eppendprf mdi. Them 200pl Elution Buffer vao tam mang ciia cot, U d nhiet dp phdng trong 2 phut va ly tam 8000 vdng/2phiit
Sau khi tinh sach, DNA duac kiem tra ndng do tren may quang phd Sp-3000 nano Optima d budc sdng 260/280 nm. Dien di DNA tdng s6 tren gel agarose 0,8% vdi cac bang cd kich thudc tren 21 kb
Tgp chi Cdng ngh4 Sinh hgc 11(2): 203-210, 2013 Tiep do, DNA dugc pha loang dkn ndng dd 40 ng/pl, chia nhd va bao quan d -20°C.
Thi& ki moi de nhdn dogn gen MMP-9 Doan gen can khuech dai cd kich thudc 435bp nam tir vi tri 1809 tdi 1374 tren gen MMP-9, Dya vao trinh ty gen chuan MMP-9 tren NCBI vdi sd dang ky NM_004994, chiing toi thiet ke cap mdi:
Mdi xudl (F): 5'-GCCTGGC AC AT AGTAGGCCC-3'
tir san pham PCR da dupc tinh sach tren he thdng may ABI PRISM® 3100 Avanl Genetic Analyzer theo nguyen ly cua Sanger vdi bp kit BigDye®
Terminator v3.1 Cycle Sequencing. Trinh tu nay sau do dupc xii ly va so sanh vdi cac trinh ly dang ky Iren ngan hang dti lieu gen qudc te NCBI bang cac ph§n mSm BioEdit, Clustal X...
KET QUA
Mdi ngugc (R): 5'-CTTCCTAGCCAGCCGGCATC-3' Ket qua nhan ban PCR doan gen MMP-9 Khuech ddi gen bdngphdn ung PCR
Khuech dai DNA bang phan iing PCR, san pham DNA tinh sach dupc khuech dai bang PCR (Polymerase Chain Reaction) sir dung may gia nhiet Gene Amp"^ PCR System 9700 ciia hang Applied Biosystems, Cac pimer sir dung cho phan img PCR dugc mo la d bang tren. Phan iing PCR cd chu trinh nhiet nhu sau: bien tinh DNA d 94''C trong 5 phiit, tiep theo la khuech dai DNA vdi 35 chu ky (gdm bi^n tinh d 94''C/45 giay , bat cap d 62''C/l phiit va keo dai chudi d 72''C/1 phut 30 giay, cuoi ciing ii d 72"C trong 7 phut).
San pham PCR ti6p tuc dupc lam sach bang Qiagen Kil (QIA quik® Gel extraction Kit, Dire) tao dieu kien thuan lgi cho viec xac dmh trinh lu gen.
Xdc dinh trinh tugen
Trinh tu cac nucleotide duoc xac dinh tryc tiep
Mau DNA genome lir mau ngoai vi cua 41 ca the ngudi viet nam bao gdm 9 ngudi binh thudng khde manh, 14 benh nhan dau that ngyc khong dn dinh va 18 benh nhan nhoi mau ca tim cap dupc diing lam khudn cho phan img PCR vdi cac cap moi duac thiet ke dac hieu va chu trinh nhiet da dupc tdi uu hda nhu da md ta trong phan vat lieu va phuang phap nghien ciru, Doan gen MMP-9 duoc nhan vdi cap mdi (F-R), san pham PCR duoc tinh sach va kiem tra bang dien di gel tren agarose 1,5%, nhudm vdi elhidium bromide va dgc dudi anh sang ciia Ua tir ngoai, nhu tren hinh 1 minh hpa ket qua PCR.
Ket qua giai trinh ttr doan gen MMP-9 Trong 41 mau nghien ciru da dugc xac dinh trinh tu doan gen MMP-9 chiing tdi phat hien thay cd 10 mau benh co da hinh nucleotide dan trong viing promoter doan gen MMP-9.
10 11 i ;
Hinh 1. Oi^n di i56 sSn pham PCR duoc tinh sach ciia do^n gen MMP-9 tren gel agarose 1,5% cua 12 mhu AND genome ti>
mSu ngoai vi K^t q u ^ cho IhSy, san ph^m PCR thu d u g c co ktch thuac phan li> khoang 435bp, kich thudc nay phij hgp vfri kich t h u * c Iinh loan tren ly thuyet. Bang san pham PCR sang, dam va ro net dii dieu kien cho vi#c giai trinh t u gen
D d Hiiu Chi et al.
MMP9 BTOl BT02 DT09 DTll DT22 NH06 NHI 5 NM17 NM36 NM50 NM64 NM66
MMP9 BTOl BT02 DT09 DTll DT22 NM06 NM15 NM17 HM36 NM50 NM64 NM66
MMP9 BTOl BT02 DT09 DTll DT22 NM06 NM15 NMI 7 NM36 NM50 NM64 NH66
1460 1470 1480 1490 1500 1510
.1 ....|....| ....|....| ....|....| ....|....| ....i....| ....|.--
TG GGCAGATCAC TTGAGTCAGA AGTTCGAAAC CAGCCTGGTC AACGTAGTGA AACCCCAT
1520 1530 1540 1550 1560 1570
.1 ....|....| ....|....| ....|....| ....|....| ....|....| ....|...
CT CTACTAAAAA TACAAAAAAT TTAGCCAGGC GTGGTGGCGC ACGCCTATAA TACCAGCT
1580 1590 1600 1610 1620 1630
.1 .,..|....| ....|....| ....|....l ....|....| ....|....| ....|...
AC TCGGGAGGCT GAGGCAGGAG A_ATTGCTTGA ACCCGGGAGG CAGATGTTGC AGTGAGCC
Hinh 2. So sanh trinh tu nucleotide ciia doan gen MMP-9 tip 12 ca the ngudi Viet Nam (BTOl, BT02, DT09, DT 11. DT22, NM06, NM15, NM17, NM36, NM50, NM64, NM66) vdi trinh lu gen MMP-9 chu^n trgn ngan hang NCBI. Ket qua phan tich bang phan mem bioEdit cho thay trinh tu doan gen MMP-9 nhan duoc tiF cac mau ADN mau ngoai vl cua cac nhdm b|nh va nhom doi chirng deu co kich thudc va do tuong d6ng cao vdi trinh tu chuan NT_011362 10 tren NCBI. Co mgt diem sai khac la da hinh nucleotide don tai vi td 1562C>T da duoc tim thay d 3 miu dau thit ngirc khong on djnh va 7 miu nhbi mau CO tim cap so voi miu dot chirng
Tgp chi Cdng nghe Sinh hgc 11(2) 203-210, 2013
THAO LUAN
Sau con nhoi mau c a tim cap, nhieu cytokines va enzymes phan giai protein da dugc giai phdng.
MMP-9 dupc tiet ra la zymogen 92kDa tir bach c5u trung tinh, dai Ihyc bao, mpt s6 loai xk bao:
monocytes, macrophages, neutrophils, keratinocytes, fibroblasts, osteoclasts va endothelial. MMP-9 cd cau tnic gom 3 viing: viing gidng nhu fibronectin cd chiic nang trong qua Irinh gin ciia gelatin, viing dac trung tuong tu cua collagen loai V giau serine/proline'tlireonine dong vai trd nhu vi tri gan 0-linked oLgosaccharides da lien ket va vimg lugng ty hemopexin co tan ciing la duoi carboxyi, MMP-9 co lien quan d&n phan iing viem, tai tao md, lam lanh vet thucmg, phat trien khoi u va di can. MMP-9 ddng vai trd quan Irgng trong sy pha vo lien ket peptide trong phan tir prolein (endopeptidases) lam suy giam protein chat nen ngoai bao va trong viec di chuyen ciia bach cau, cung nhu trong tai hap thu le bao hiiy xuong, MMP-9 phan ra thanh collagen dang IV va dang V trong ba phan tu doan large C-terminal va mot phan tu doan shorter N-terminal. MMP-9 cdn cd the lam giam fibronectin nhung khdng lam giam laminin ho5c Pz-peptide, MMP-9 tai to hgp trong E.coli khdng bi glycosyl hda, chuoi polypeptide co chira 338 doan aa (113-450) tuong iing vdi viing xiic tac cua protein co tong trpng iugng phan tti 42,03kDa va hpp nhat vdi 4.5kDa amino-terminal hexahistidine tag.
Danh gia lam sang mdt benh nhan nghi ngd mac HCMVC thudng bi han che khi tneu chirng khdng dien hinh hoac tham lang gay khd khan cho viec chan doan ciing nhu bd sdt, lam cham tre qua trinh cap ciru cho benh nhan. Ngoai ra, dien tim liic ban dau thudng ciing kJiong giup ich cho chan doan trong phan ldn benh nhan do do nhay luong ddi thap (35- 50%) (Lee et al., 1985, Selker el al, 1997). Tiap can hi^n nay tap trung vao cac dau an tim (cadiac markers) nhu la nen lang trong chan doan va nghien Cliu benh sinh cua HCMVC. Cac dau an tim la xet nghiem thudng dugc sir dung nhat hien nay nhu chi thi phan lir ciia tdn thuang vach lim: creatine kinase (CK) va creatine kinase-MB (CK-MB), cardiac troponin T (cTnT) and 1 (cTnl), B-lype natriuretic peptide (BNP), C-reactive prolein (CRP), Chi thi phan tir ciia mang xo viia khong dn dinh: placental growth factor (PIGF). myeloperoxidase (MPO), pregnancy-associated plasma protein (PAPP-A), metalloproleinase-9. Chi thi phan tii ciia chimg thieu mau cyc bp va ho^i tii md: fatty acid binding
proteins (FABPs), free fatty acids unbound to albumin (FFAu), ischemia modifi ed albumin (IMA) (Loria et ai, 2008), Cac bang chiing hien nay tir nhung nghien ciiu co ban den lam sang deu cho thay MMP-9 ddng mdt vai Ird quan trong trong viia xa dpng mach va nhoi mau co tim cip (Derosa et ai, 2007; Inokubo et ai. 2001). Da hinh gen MMP-9 lien quan vdi lang boat ddng ciia enzyme MMP-9 trong huyet Urong va gdp phan vao qua trinh phat trien ciia benh dgng mach vanh d giai doan dau (Saedi et ai, 2012). Ket qua nghien cihi ciia chiing tdi cung cho thay cd mgl da hinh nucleotide dan d vi tri 1562C/T trong viing promoter cua doan gen MMP-9 d berdi nhan dau that nguc khdng dn dinh va nhoi mau co tim cap, dang luu y la khong tim thay da hinh nay d nhirng ngudi khoe manh. Da hinh d benh nhan bi nhdi mau co tim (7/18) cao hon d benh nhan dau that ngyc khdng on dinh (3/14), Da hinh nucleotide dan (Single nucleotide polymorphism, SNP) trong promoter gen MMP-9, cu the la qua trinh chuyen ddi ciia C (cylosine) thanh T (thymidine) d vi tri -1562 dugc biet den la yeu td nguy co nhoi mau CO tim cip ( K o h ^ / ( i / , 2 0 0 8 , Wang ef a/,, 2011), Tin sd ciia da hinh dan nucleotide MMP-9 trong nhdi mau CO tim cap tinh la cao hon so vdi nhdm ddi chimg (Koh et ai, 2008, Zhang et cd., 1999;
Pollanen et ai, 2001) Mgt sd cdng bd ket luan ddi lap nhau ve vai trd cua da hinh MMP-9 vdi mirc do nghiem trgng ciia benh dgng mach vanh (Pollanen el al., 2001; Abilleira et al., 2006; Dalepiane el ai.
2007). Nhirng diem luang dong va khac biet trong cac ket qua ciia nhirng nghien ciru trudc day cd the vi da hinh xay ra trong khu vuc promoter se anh hudng den phien ma gen va bieu hien tang nong do ciia MMP-9. Han nCra trong su kich boat MMP-9 tit pro-MMP, mol sd luang tac vin can thiet nhu TIMP (tissue inhibitor of melalloproleinase), MT-MMP (membrane-type matrix metalloproteinase), plasmin, va CO che phan hoi ciia chinh MMP-9 (Rutschow el ai, 2006).
Su quan tam ciia cdng dong cac nha khoa hgc den chirc nang gen MMP-9 trong nhiing nSm qua da tang len nhanh chdng, Nghien ciru ciia Kim nam 2005; Wang va ddng tac gia nam 2011 deu cho thay da hinh gen MMP-9 cd vai tro quan trgng trong qua trinh co tim bi nhdi mau va muc do bieu hien tang cao ciia MMP-9 irong huyel thanh o ngudi Han Qudc va d tdc ngudi Uygur (Trung Qudc). Kai va ddng lac gia nam 1998 da do ham luang MMP-9 trong benh nhan ACS va benh nhan dau thai ngyc dn dinh so sanh vdi ngudi khde manh, Benh nhan dau that nguc dn dinh c6 ham
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lugng MMP-9 luong tu nhu ngudi khde manh, tat ca cac benh nhan dau that nguc on dinh cd ndng do cua MMP-9 cao liic nhap vien, giam d i n sau trong ihdi gian nam vien. Vai trd tiem nang ciia MMP-9 nhu la mpt chi thj sinh hgc ciia su phan loai nguy CO benh nhan ACS da duoc kiem tra bdi Blankenberg nam 2003, hg da nghien ciiu 795 benh nhan dau that ngyc on dmh va 332 dau that ngyc khong pn djnh, ket qua cho thay MMP-9 cd lien quan den tir vong d benh nhan lim mach. Gia t n chuan doan ciia MMP-9 chi xep sau cac chi sd CRP, fibrinogen, IL-6, IL-18. That vay, su giai phdng MMP-9 cd y nghTa bii dap cho cac md ciia lim sau khi bi ton thuang, mac dii cac dir lieu trudc day cho thay d ddng vat chuyen gen de bi vd tam that dugc bao ve qua thieu hut cua gen MMP-9 (Heymas et ai, 1999), Nghien cim mdi nhit nam 2012 ciia Selianto va ddng tac gia cung cap 2 phat hien: thit nhal, kel qua do miic do bieu hien MMP-9 trong huyel thanh d thdi gian dau tang len dang kh cua nhung benh nhan nhdi mau c o tim cap tinh vdi ST chenh ien (STEMI), udc tinh cao gap 4 lin so vdl khi binh thudng; thii hai, da hinh MMP-9 (- 1562C>T) lien quan vdi miic bieu hien cao ciia MMP-9 trong huySt thanh benh nhan STEMI, Nhin chung, van con mdt sd ban che nghien ciiu lam sang ve MMPs d cac benh mach vanh cap tinh, Vi du, ndng dp trong huyet tuang hoac hoat ddng cua MMP-9 trong cac tdn thucmg xa viia dpng mach van cdn chua rd rang. Hon niia, d giai doan thrombin ciing cd the kich boat d^ bxku hien MMP- 9. Nhu vay, mirc do cao ciia MMP-9 Irong huyet thanh cd the chi don ihuan la mdt he qua, chii khdng phai la nguyen nhan gay ra cac cue huyet khdi ben trong Idng mach mau.
Chimg tdi da nhan thanh cdng doan DNA cd kich thudc 435bp chiia doan gen MMP-9 tir cap mdi (F-R) ciia 41 mau nghien ciru va xac dinh dupc trinh lu doan gen MMP-9 bang phuang phap dgc true ti^p lir san pham PCR ciia cac mau. Da phan tich va so sanh trinh tu doan gen MMP-9 cua 41 ca thd ngudi Viet Nam vdi trinh ty chuin tir NCBI, Tim t h i y diem da hinh nucleotide don vi tri 1562C>T d 10 mau trong tdng sd 32 mau benh. Cac di^m da hinh nay deu lim thiy d benh nhan mic hpi chiing mach vanh cap ma khdng thiy d ngudi binh thudng. K.61 qua tren la nghien ciiu diu tien da danh gia mdi lien Ne tiem nang giiia da hinh MMP-9 d nhirng benh
nhan cd hgi chirng mach vanh cap voi ngudi binh thudng.
Lcri cam cm: Cong trinh ndy dugc hodn thdnh vai su hd trer ve kinh phi cua de tdi dgc lap thugc Phdng Thi nghiem Trgng diem Quoc gia, Bg Khoa hgc vd Cdng nghe DT- PTNTD. 2011-G/03 (201J- 2014): "Tim kiem cdc chi thi sinh hoc lien quan hgi chimg mach vdnh bang cdc ky thugt proteomics".
TAI LIEU THAM KHAO
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D E T E C T I O N O F A S I N G L E N U C L E O T I D E P O L Y M O R P H I S M IN T H E M M P - 9 P R O M O T E R R E G I O N F R O M P A T I E N T S W I T H A C U T E C O R O N A R Y S Y N D R O M E Do Huu C h i ' , Le Thi Bich T h a o ' . Nguyen Tien Dung'. Pham Duc Dan', Bui Thi Huyen', Nguyen Thi Minh Phuong', Do Doan Lol^ Nguyen Bich Nhi', Phan Van Chi'*
' Inslilule of Biotechnology. Vietnam Academy of Science and Technology 'Vietnam Heart Institute, Bach Mai Hospital
SUMMARY
According to the Worid Heart Federation, Cardiovascular diseases are the world's largest killers, claiming 17.5 million lives a year. Acute coronary syndrome (ACS) is a health issue of great concern woridwide due lo ils sudden attack, high mortality rate and serious sequelae, wilh the number of patients increasing rapidly in both developing and developed countries. Recent studies have discovered genetic association belween polymorphic variants in candidate genes and atherosclerosis The mairix
• Aulhor for correspondence • Tel ^S~l-4-379l3657: L-mail chi@ibia
Dd Hiru Chi et ai metalloproteinase (MMP) family is a potential candidate gene systems for ACS, of which MMP-9 is the most attractive target research due lo ils vital role in extracellular malnx degradation and atherosclerotic plaque reproduction. In this study, sequencing of MMP-9 gene from forty-one inviduals (9 normal samples, 14 unstable angina and 18 acute myocardial infarction patients were Vietnamese carried out. By comparing MMP9 gene sequence between ACS group and the control without ACS, the -1562C>T polymorphisms were detected in MMP9 genes of 3 unstable angina patients and 7 acute myocardial infarction patients Our results show the first data of MMP-9 gene sequence from Vietnamese ACS patients.
Keywords: Polymorphism, acute coronary sydrome, matrix metalloproteinase-9, nucleotide, promoter
