Hubungan Mutasi Gena Hemoglobin Beta (HBB) dan PemodifikasGenetik XmnI, BCL11A, dan HBS1L-MYB Terhadap Fenotip Thalassaemia Beta

LANTIP RUJITO, Prof.dr. Abdul Salam M Sofro, PhD.; Dr.dr.Sri Mulatsih, SpA (K)

2015 | Disertasi | S3 Kedokteran Umum

Thalassaemia Beta memiliki heterogenitas genetik dan klinis yang berbeda dalam berbagai populasi. Tujuan penelitian adalah untuk menentukan jenis mutasi pada gena Beta, gena pengubah genetik lokus XmnI, BCL11A, HBS1L-MYB, dan aspek fenotipe pasien Thalassaemia di Banyumas, Jawa Tengah, Indonesia. Penelitian menggunakan desain penelitian cross sectional study. Subyek adalah semua pasien dengan diagnosis Thalassaemia dalam database Yayasan Thalassaemia Indonesia (YTI) Banyumas. Jenis mutasi pada gena globin Beta ditentukan dengan teknik Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP), Amplification-Refractory Mutation System (ARMS), dan direct sequencing. Alel-alel pada lokus XmnI, SNPs rs11886868 , rs766432, rs9399137 dikarakterisasi menggunakan teknik PCR-RFLP dan ARMS sesuai dengan primer dan enzim restriksi yang sesuai. Status klinis pasien Thalassaemia diukur dengan skor derajat klinis Sriciphai. Data dianalisis secara deskriptif selanjutnya dilakukan uji multivariat sesuai dengan karakteristik data. Hasil pengujian dianggap bermakna bila p<0,05 pada interval kepercayaan 95%. Penelitian menemukan bahwa frekuensi alel mutasi gena Beta sebagai berikut : IVS-1-5 (G>C) (43,5 %), Cd26 (HbE) (28,2%), IVS-1-1 (G>A) (5%), Cd15 (TGG>TAG) (3,8%), IVS-1-1(G>T)(3,1%), Cd35 (-C) (2,4%), serta di bawah 1 % adalah Cd41/42 (-TTCT), Cd8/9 +G, Cd19 (AAC>AGC), Cd123/124/125(- ACCCCACC), IVS-1-2 (T>C), Cd17 (AAG>TAG), Cd40 (-G), CAP +1 (A>C). Data penelitian memperlihatkan bahwa alel XmnI memiliki frekuensi alel minor 14 %. Alel C pada rs11886868 pada populasi subyek di Banyumas adalah alel mayor dengan frekuensi 78 %. Distribusi alel rs766432 dan rs9399137 memiliki distribusi yang hampir serupa pada berbagai populasi dunia (18-19 %). XmnI, rs766432 dan rs11886868 pada kelompok HbE/Thalassaemia Beta berhubungan secara signifikan baik pada kadar HbF maupun derajat klinis pasien (p<0,05). Sedangkan alel s9399137 HBS1L-MYB tidak menunjukkan hubungan yang serupa (p>0,05). Pada kelompok homozigot Thalassaemia Beta-0/Beta-0 maupun Thalassaemia Beta-0/Beta-+ (severe) polimorfisme lokus XmnI, rs766432, rs11886868, dan rs9399137 tidak berhubungan secara signifikan dengan derajat klinis pasien (p>0,05). Kesimpulan penelitian adalah sebagai berikut; mutasi gena Beta paling banyak adalah alel IVS-1-5 (G>C) dan Cd26 (HbE) dengan persentase 43,5 % dan 28,2 %. Polimorfisme lokus XmnI, rs766432, rs11886868 (BCL11A) pada pasien HbE/Thalassaemia Beta berhubungan secara signifikan dengan derajat klinis, sedangkan rs9399137 (HBS1L-MYB) tidak berhubungan. Polimorfisme lokus XmnI, rs766432, rs11886868, dan rs9399137 pada pasien homozigot Thalassaemia Beta-0/Beta-0 maupun Thalassaemia Beta-0/Beta-+ (severe) tidak berhubungan secara signifikan dengan derajat klinis.

Beta Thalassemia has different genetics and clinical heterogeneity in various populations. The research objective was to determine the types of mutations in Beta gene, a genetic modifier gene of XmnI locus, BCL11A, HBS1L-MYB, and phenotype aspects of Thalassemia patients in Banyumas, Central Java, Indonesia. The study used cross sectional study design. The subjects were all patients with a diagnosis of thalassemia in the database of the Indonesia Thalassaemia Foundation (YTI) Banyumas. The type of mutation in the Beta globin genes was determined by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP), Amplification-Refractory Mutation System (ARMS), and direct sequencing. Alleles at the XmnI locus, SNPs rs11886868, rs766432, rs9399137 were characterized using PCR-RFLP and ARMS technique in accordance with appropriate primers and restriction enzymes. Clinical status of Thalassaemia patients were measured using Sriciphai clinical score. Data were analyzed descriptively then performed multivariate test in accordance with the characteristics of the data. The test results are considered significant if p <0.05 on 95% confidence intervals. The study found that the mutation allele frequency of Beta genes were as follows: IVS-1-5 (G>C) (43.5%), Cd26 (HbE) (28.2%), IVS-1-1 (G>A) (5 %), Cd15 (TGG>TAG) (3.8%), IVS-1-1 (G>T) (3.1%), Cd35 (-C) (2.4%), and below 1% were CD41/42 (-TTCT), CD8/9 + G, CD19 (AAC>AGC), Cd123/124/125 (- ACCCCACC), IVS-1-2 (T>C), Cd17 (AAG>TAG), CD40 (-G), CAP +1 (A>C). Data showed that the XmnI allele have a minor allele frequency of 14%. C allele at rs11886868 in the subject of Banyumas population is a major allele with a frequency of 78%. Rs766432 and rs9399137 allele distribution has a similar distribution to the various populations of the world (18-19%). XmnI, rs766432 and rs11886868 in the HbE/Beta Thalassaemia group associated significantly with HbF levels or clinical score patients (p<0.05), while s9399137 HBS1L-MYB alleles did not show a similar relationship (p>0.05). In homozygous Beta-0/Beta-0 Thalassaemia and Beta-0/Beta-+ (severe) Thalassaemia XmnI polymorphisms, rs766432, rs11886868, and rs9399137 did not significantly associate with the degree of clinical scores (p>0.05). The conclusion of the study was as follows; most mutations in Beta genes are alleles of IVS-1-5 (G> C) and Cd26 (HbE) with a percentage of 43.5% and 28.2%. XmnI polymorphism loci, rs766432, rs11886868 (BCL11A) in patients HbE/Beta Thalassaemia significantly associated with clinical degrees, while rs9399137 (HBS1L-MYB) was not related. XmnI polymorphism loci, rs766432, rs11886868, and rs9399137 in patients with homozygous Thalassaemia Beta-0/Beta-0 and Thalassaemia Beta-0/Beta-+(severe) was not significantly related to the clinical degree.

Kata Kunci : Thalassaemia Beta, XmnI, BCL11A, HBS1L-MYB, clinical appearance