TCNCYH Phu tritdng 62 (3) - 2009
variants including a new variant distributed in 9. Verle~ P (2000). Glucose - 6 - phosphate Lam Dong province, Vietnam. Acta Med, dehydrogenase in northern Vietnam. Trop Med Okayama, 5 1 , pp. 2 1 3 - 2 1 9 . and Int Health, 3, pp. 203 - 206.
8. Nuchprayoon 1 (2002). G6PD Viangchan 10. Vulliamy T and Beutler E (2002).
(871 G > A) is the most common deficiency Hematologically important mutations: glucose - 6 - variant in the Thai population. Hum Mutat, 1 9, (1 85). phosphate dehydrogenase. Blood Cells Moi Dis, 28, (93).
Summary
MUTATIONS ON GLUCOSE - 6 - PHOSPHATE DEHYRDROGENASE (G6PD) GENE IN THREE ETHNIC GROUPS: MUONG, TAY, AND THAI IN VIETNAM
Glucose - 6 - phosphate dehydrogenase (G5PD) is one of the most common hereditary disorders in the world, affecting to more than 400 million people in the world. In most of the cases, G5PD deficiency was caused by mutation (s) on G6PD gene. Objectives: to find out mutant points on G6PD gene of some Vietnam ethnic groups. Methods: genomic DNA was extracted from 39 blood samples belongs to 3 ethnic groii|3s: Muong, Tay, and Thai. PCR (Polymerase Chain Reaction) was used to amplify the all of exons of G6PD gene. PCR products was purified and dyed with fluorescent then reading sequencing of DNA.
Results: there were 6 changes of nucleotide at position 143 T > C, 487 G > A, 871 G > A, 1360 C > T, 1376 G > T and 1388 G > A. The change at positions 871 G > A (G6PD Viangchan) and 1360 C > T (G6P1'3 Union) was quite predominant with prevalence 25.7% and 53.8%, respectively. Conclusion:
(i6PD Viangchan and C6Pf) Union were two common variants in the studies of G6PD among three ethnic groups: Muong, Tay, and Ihai in Vietnam.
Keywords: G6PD deficiency, Vietnam, G6PD Viangchan, G6PD Union
eiEU TRA TY LE THIEU HUT GLUCOSE 6 PHOSPHATE DEHYDROGENASE HONG CAU 6 MOT SO DAN TOC SONG TAI
D A K LAK V A L A M DONG, VIET NAM
Ta Thj TTnh, Le Minh Dao
Vien Sdt ret - Ky sinh triing - Cdn triing Trung ttang
Thieu glucose 6 pholphale dehydrogenase hong cau (G6PP) li mdt benh dl truyen cd Hen quan de'n nhiim si( lhe gidi linh X, mat benh ly rdi loan enzym thtfdng gap nhai d ngifdi. Benh hay gap d nhdng nhom dim Ioc sdng trong vung sdt ret lUu hanh. Viec xac djnh ty le thieu G6PD d cic nhdm din tge tren cd y nghia quan trgng lrong chinh sich thtidc sdt rel. Muc tieu: xic djnh ty le thieu G6PD hong cau d 4 nhdm dan Ioc lai Dak Lak va Lam Dong. Doi Tugng va phuang pbap ngbien cdu: dieu tra dt ngang tren 4 dan tdc E Dc, M'ndng (tai Dak Lak nam 2005) vi Co ho, Chu ru (tai Liim Ddng nam 2006), sd dung phuang phip nhanh (Akira hirano - 1998) de phii hien cic trtfdng hop thieu C6PD. Ket qua: ty le thieu C6PD nhu sau-
TCNCYH PhiJ trWdng 62 (3) - 2009
thieu G6PD Ciia nhdm E De li 1,3% (n = 477), M'ndng li 0,5% (n = 575), Cd ho li 1,0 (n = 485) vi Chu ru li 2,9%, (n = 418). Ty Id thie'u G6PD d Nam cao ban so vdi nd. Nhdm E de Nam 2,2%,, Nd li 0,7%o , nhdm M'ndng Nam 0,8%o , Nd 0,3%o; Nhdm Ca ho: Nam 1,4%o, Nd 0,7%o vi nhdm Chu ru: Nam 4,5%o, Nd 1,9%o.
Ket luan: ty le thieu C6PD ciia nhdm dan tdc E de va M/ndng tai Dak Lak tuang dng li 1,3%o vi 0,5%,; ty le thieu C6PD ciia nhdm dan tdc Ca ho va Chu ru tai Lam Dong lUang dng la 1,0%o vi 2,9%o.
Tfl Khoa: Thie'u G6PD, Viet Nam, E de, M'ndng, Cd ho, Chu ru, phflong phap Akira hirono
I. DAT VAN DE
Dak Lak va Lam Ddng la hai trong sd' nam tinh thupc Tay Nguyen, la 2 tinh cd r i t n h i i u dan tdc cung sinh sd'ng, cd tdi 37 dan tdc d Dak Lak va 40 dan tdc d Lam ddng. D l y cung la hai tinh tinh hinh sd't ret cdn dien b i i n phflc tap trong nhflng nam vfla qua. D l t i i n tdi loai trfl benh sd't ret, thudc dieu trj tiet can (diet t h i an d trong gan) cho den nay van chl cd duy n h i t la primaquin. Vdi c l e trfldng hpp benh nhan n h i i m P.vivax viec d i i u trj primaquin p h i i keo dai 10 ngay (phac dd Bp Y t l ) hoac 14 ngay (theo To chflc Y t l T h i gidi). Vdi cac trfldng hdp p h i i sfl dung primaquin dai ngay, cd n h i i u nguy cd g l y d i i huyet cau t d n i u benh n h i n cd t h i i u G6PD. Cho d i n nay c l e sd lieu d i i u tra v l t h i i u G6PD tai Tay Nguyen cd r i t it cdng bd'. N h l m muc dich tranh c l e tac dung khdng mong nudn do thud'c gay ra va cd c h i i n lifdc sfl dung thudc an toan d c l e vung sd't ret lflu hanh, viec dieu tra x l c djnh ty le t h i i u G6PD d cac nhdm dan tdc sd'ng tai Tay nguyen la vd cung can t b i l t . Chinh vi vay chung tdi da tien h i n h nghien cflu vdi muc tieu:
Xac dinh ty le tbieu G6PD tren 4 nhdm dan tge sdng tai 2 tinb Dak Lak va Lam Dong.
II. DOI TUONG VA PHUONG PHAP NGHIEN CLfu
1. Ddi tfldng nghien cflu
La ngfldi dan thudc 4 nhdm dan tdc : Ede, M'ndng tai Dak Lak; Chu ru, Co ho tai Lam Ddng, khdng phan biet gidi tinh, tudi tfl 1 trd len.
2. Phfldng phap nghien cflu
Dieu tra ngang md t l dfldc t i i n hanh tai Dak Lak nam 2005 va L l m Ddng nam 2006.
Cd mau cho d i i u tra c i t ngang, tfnh theo cdng thflc tfnh cd mau cho mpt ty le:
s-
Z = 1,96 neu x = 0,05; p: ty le thieu G6PD/
dan tge Ude dnh trong nghien ciiu theo mgt sd nghien cifu khac la 5°/o (p = 0,05), 8 = 0,02. Tinh ditac cd mau cho mdi nhdm dan tge la 450 ngttdi.
Pbuang phap chgn mau: chpn cd chu dfch c l e xa cd d i n tdc can d i i u tra. Trong xa d i i u tra chpn m l u theo phflong p h i p ngau nhien cd he thdng, ddn vj m l u I I hd gia dinh. T i t c l c i c t h i n h vien trong gia dinh d i u dfldc lfla chpn vao nghien cflu.
Trong nghien cflu nay chung tdi da d i i u tra 477 ngfldi E de, 575 ngfldi M'ndng, 418 ngfldi Chu ru va 485 ngfldi Cd ho.
Phuang phap xac dinh tbieu C6PD: I p dung phflpng phap nhanh cua Akira hirono - 1998 [3,4]: hoa c h i t gdm cd Glucose 5 phosphat (G5P), nicotinamid adenin dinucleotid phosphat (NADP), Dimethy - thiazolyl diphenyltetrazolium bromid (MTT), phenazin methosulphat (PMS), DEAE - Sephadex A - 50, dem Tris - HCI pH 6,5.
Nguyen ly cua phfldng phap (sa dd trang 16).
Cac budc tien hanh :
Cac h o i e h i t ndi tren dfldc pha thanh dang hdn djch gdm cd: DEAE - Sephadex 20ml, dem Tris HCI 20ml, G6P 30,4 mg, NADP 6,3 mg, MTT 5,0 mg va PMS 5,0 mg pha trong nfldc c i t , hoan
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TCNCYH PhtJ trWdng 62 (3) - 2009
G6PD
G6P •*• 6 - G P
NADP
San pham cd mau .^
(formazan)
NADPH
MTT PMS
Sa do 1. Nguyen ly phUang phap xac dinh tbieu but C6PD
thanh du 80 m l . Cho 0,8 ml hdn djch tren cho vao d'ng eppendorf, de trong hdp tdi, bao q u i n d 4''C.
Khi tien hanh: lay 5 ml m l u tfl dau ngdn tay hoac tif mau da chdng ddng, nhd trflc tiep vao dng eppendorf chfla 0,8 ml hdn djch tren, lie ky, de trong hop tdi d nhiet dp phdng, sau 30 phut dpc ket qua.
III. KETQUA
Ket qua: ngfldi binh thfldng (khdng t h i i u G6PD): dung djch c h u y i n tfl m l u vang chanh sang mau tfm khoai mdn.
Ngfldi t h i i u G5PD: dung djch khdng chuyen mau (mau v i n g chanh) n h | n biet r i t r d ' b l n g mat c i c trfldng hpp t h i i u G6PD.
TT
1 2 3 4
Ten dan
E de M'ndng Chu ru Cd ho
tdc
Tdng cdng
Bang 1. Ty le thieu C6PD theo nhom dan tdc
Ndi dieu tra
Huyen Lak, Dak Lak Huyen Lak, Dak Lak
Huyen Ddn Dfldng, Lam Ddng Huyen Dam Rdng, L l m Ddng
Sd mau dieu
477 575 418 485 1,955
tra
Thie'u n 6 3 12 5 26
G6PD
(%)
(1,3) (0,5) (2,9) (1,0) (1,3)
Nhan xet: ty le thieu G6PD cua cac nhdm dan tge ban xd tai Tay Nguyen tha'p (td 0,5 - 2,9%), trung binh li 1,3%o.
Bang 2. Ty le tbieu C6PD ciia cac dan tge theo gidi tinb
Nam NiJ
Dan tdc
So dieu tra
Thieu G6PD
Sd'^ dieu tra
Thieu G6PD
1 2 3 4
E de M'ndng Chu ru Co ho Cpng
185 231 155 212 785
n 4 2 7 3 15
(%)
(2,2) (0,8) (4,5) (1,4) (2,0)
291 344 262 "
273 1,170
n 2 1 5 2 10
(%)
(0,7) (0,3) (1,9) (0,7) (0,9)
> 0 , 0 5
> 0 , 0 5
> 0 , 0 5
> 0 , 0 5
TCNCYH PhtJ trWdng 62 (3) - 2009
Nhin xet: ty le tbieu G6PD ciia nam cao ban nd tren 2 lin, tuy nhien sd khac biet chtfa cd y nghia thd'ng ke (p > 0,05) khi sd diing test / 2 .
IV. BAN LUAN
Ke't q u i d i i u tra cua chung tdi cho t h i y ty le thie'u G5PD cua cac nhdm dan tdc b i n xfl tai d l y la kha t h i p , t h i p nhit-11 0,5% (dan tdc M'ndng) va cao n h i t la 2,9% (dan tdc Chu ru). C I c nghien cflu trong nfldc cung nhfl tren t h i gidi d i u cho t h i y ty le t h i i u G6PD cua cac dan tdc khac nhau r i t k h l c nhau [5, 6,]. Mdt nghien cflu cua D o l n Hanh N h i n nam 1 995 [1 ] , tien hanh d i i u tra tren 1,676 hpc sinh nam phd thdng cd sd tai 4 tlnh H d i Binh, H I Giang, Sdn La va Thanh H o i thudc cac dan tpc Mfldng, T h i i , Mdng. Thd va Kinh cho t h i y ty le t h i i u G6PD cao n h i t thupc nhdm dan tpc Mfldng 3 1 % (n = 261), Thd 22,6%
(n = 105), Thai 19,3% (n = 575) va t h i p la nhdm d i n tdc Mdng 0,3% (n = 390), Kinh 0,5%
(n = 202). Mdt nghien cflu k h l c cua Ta Thj TTnh nam 1999 [2], t i i n hanh tai Gia lai cho t h i y : ty le t h i i u G6PD d nhdm dan tdc Gia Rai 2,3%
(n = 523), Ba Na 1,7% (n = 300). Mdt sd tac g i l cho r i n g vung dia ly cd lien quan vdi t h i i u G6PD, d nhflng vung sd't ret lflu hanh thfldng t^ le t h i i u G5PD cao hdn d nhflng vung khdng cd sd't ret [5,6,7]. Nghien cflu cua chung tdi khdng thd'ng n h i t vdi quan diem nay. Cd mdt d i i u r i t dang quan tam la da so eac d i n tdc phfa B i c nhfl Mfldng, T h i i , Tay, Thd cd ty le thieu G6PD cao hdn rd ret so vdi mpt sd dan tpc phia Nam (d Tay Nguyen) nhfl Ba Na, Gia Lai, Xieng va trong nghien cflu n l y cua chung tdi la nhdm dan tdc E De, M'ndng, Cd ho va Chu ru. P h i i chang ngudn gde d i n tpc phan nao cd tfnh c h i t quyet djnh den bien the G5PD.
Nghien cflu cua Matsuoka va cdng sfl tfl 1997 - 2003 tai Lam Ddng [7], tren 1104 m l u mau l i y vao g i i y t h i m Whatman 111, b l n g phfldng phap
formazan t i c g i l da x l c djnh ty le thieu G6PD cua ngfldi Kinh tai day la 1,9% (19/981) va ngfldi Cd ho I I 4 , 4 % (5/1 14) v l chi vdi 25 trfldng hdp thieu G6PD tac g i l da x l c dinh cd 7 bien the, trong dd cd mdt b i i n the mdi dfldc dat ten I I bien the Bao Ldc. D i i u dd cho t h i y sfl da dang cua cac bien the t h i i u G6PD tai d l y . Trong nghien cflu nay chung tdi chfla cd ket qua p h l n tfch gen de xac djnh c i c bien the t h i i u G6PD.
Theo y van, cd r i t n h i i u c i c phflpng p h i p dinh tfnh khdng p h l t hien dflpc cac trfldng hpp t h i i u G6PD d nfl (heterczygote) trfl phfldng phap phan tfch gen. T h i i u G6PD I I benh di t r u y i n lien ket nhiem s i c the gidi tfnh X, khdng cd alen tren Y. Locus gen qui djnh c i u true G5PD n l m tren n h l n h dai cua nhiem s i c the gidi tfnh X. d nam do chl cd mdt nhiem s i c the X, nen khi bi ddt b i i n thifdng I I thieu h o l n toan. T h i i u G6PD d nfl cd 2 dang: t h i i u G6PD ddng hpp tfl (homozygote) v l t h i i u G6PD di hdp tfl (heterczygote). C I c trfldng hpp t h i i u ddng hop tfl la ca 2 X (1 nhan tfl b d va 1 tfl me) d i u bj dot bien, tuy nhien trfldng hdp nay hiem gap, hay gap n h i t I I c l e dang dj hpp tfl. Trong trfldng hpp nay cd 2 quan the hdng cau: mdt q u i n t h i binh thfldng va mdt q u i n t h i i u G6PD. Thdng thfldng 2 quan the n l y phan bd' theo ty le 1:1. Tuy nhien trong mdt so trfldng hdp c l e hdng c l u t h i i u c h i i m tdi 9 0 % , hdng c l u I l n h chl cd 10% va mdt sd'trfldng hdp k h l c lai ngfldc lai. Vi vay da sd c i c phfldng phap dinh tfnh phat hien r i t kem c l e trfldng hdp t h i i u G6PD d nfl.
Phfldng phap Akira hirond cung nam trong quy luat nay. Trong nghien cflu d i n h gia dp nhay, dd dac hieu cua phflpng phap Akira hirono so vdi phfldng p b l p dinh lifdng, Ta Thi TTnh va cdng sfl [3], da nhan xet phflpng phap Akira hirono cd dp n h | y 79 % va dp dac hieu 9 7 % , (neu p h l t hien c l 2 gidi). Tuy nhien, dp n h | y tang len 9 2 % n i u chi phat hien d nam va giam xudng 5 3 % khi p h l t hien d nfl . D i i u nay cd t h i p h l n nao ly g i i i , ty le thieu
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TCNCYH Phu trWdng 62 (3) - 2009
G5PD d nam bao gid cung cao hon d nfl trong cac dieu tra cua chung tdi, tuy nhien chfla cd sfl khac biet rd ret, cd the do cd mau chfla du Idn.
V . K E T L U A N
B l n g phfldng phap nhanh (Akira Hirono) da xac djnh ty le t h i i u G5PD cua nhdm dan tdc E de va M'ndng tai Dak Lak tfldng flng la 1,3% va 0,5%, cua nhdm dan tdc Cd hd va Chu ru tai Lam Ddng tfldng flng la 1,0% va 2,9%.
T A I L I E U T H A M K H A O
1. Doan Hanh Nhan, P.verle, Ta Thj TTnh va cong sfl,. D i i u tra t h i i u men G5PD hdng cau d mpt sd d i n tdc sd'ng trong vung sd't ret lflu hanh tai m i i n B i c , Viet Nam. Ky y i u cdng trinh nghien cflu khoa hpc (1991 - 1995), vien Sdt ret - Ky sinh trung - Cdn trung Trung ifdng, Tap I, tr. 203
~ 209.
2. Ta Thj TTnh, Tran Thj Uyen, Le Minh Dao, Nguyen Minh Hung va cong sfl. T h i i u G5PD d mdt sd' d i n tdc sd'ng trong vung sd't ret lflu hanh.
Ky yeu cdng trinh nghien cflu khoa hpc (1996 -
2000), Vien Sd't ret - Ky sinh trung - Cdn trung Trung flpng, Tap I, tr. 2 1 0 - 2 1 5.
3. Ta Thj TTnh, Le Minh Dao, Nguyin Minh Hiing. Phfldng p h i p nhanh phat hien thieu G6PD hdng c l u . Tap chf thdng tin Y dfldc, t h i n g 3, nam 2005, trang 25 - 29.
4. Akira Hirono, Hisaichi Fuji! and Shiro Miwa - 1998. An improved single - step screening method for G6PD. J. Trop. M e d . Hyg., Vol. 26.
No. 1, 1998, pp. 1 - 4 .
5. Ernest Beutler - 1994. G6PD deficiency.
Blood. Vol 84, No 11 (December 1), 1994: pp.
3 6 1 3 - 3 6 3 5 .
6. Ernest Beutler - 1997. G6PD clinical studies in Medical Biochemistry New York, Oxford University Press, pp. 91 - 95
7. Matsuoka H., Dang Thi Vinh Thuan et al.
Seven different G6PD variants including a new variants distributed in Lam Dong province in southern Viet Nam. Acta Med Okayama 2007, Vol 6 1 , N o 4 , pp. 213 - 219.
Summary.
G L U C O S E 6 P H O S P H A T E D E H Y D R O G E N A S E D E F I C I E N C Y A M O N G I N M I N O R I T I E S E T H N I C G R O U P S I N D A K LAK A N D L A M D O N G , V I E T N A M
G6PD deficiency, a hereditary X - chromosome - linked defect, is one of the most common enzymopathries in human. G6PD deficiency relates with the ethnic groups living in malaria endemic areas. Detecting G6PD deficiency of ethnic groups living in endemic areas is very important for anti malarial drug policy. Objectives: to determine the prevalence of G5PD deficiency among in four minorities ethnic groups in Dak Lak and Lam Dong, Viet Nam. Method: a crossessional surveys were carried out among in four minority ethnic groups E de and M'nong in Dak Lak in 2005, Co ho and Chu Ru in Lam Dong in 2006. Rapid method (Akira Hirono) for identification the G6PD deficiency subjects was applied. Results: the prevalence of G5PD deficiency of Ede, M'nong, Co ho, Chu Ru ethnic group is 1.3% (n = 477); 0.5% (n = 575); 1.0% (n = 485) and 2.9% (n = 418), respectively. The prevalence of G5PD deficiency of male is higher than female in all groups. In Ede group:
male 2.2% and female 0.7%; in M'nong group: male 0.8% and female 0.3%; in Co ho group: Male 1.4% and female 0.7% and Chu ru group: male 4.5% and female 1.9%. Conclusion: the prevalence of G6PD deficiency of E de and M'nong ethic groups in Dak Lak is 1.3% and 0.5%, respectively. The prevalence of G5PD deficiency of Co ho and Chu Ru ethnic groups in Lam Dong is 1.0% and 2.9%, respectively.
Key words: G6PD deficiency, Viet Nam, Ede, M'nong, Co ho, Chu ru, Akira hirono method
