Akurasi IP-10 dan hsa-miR-15a-5p, hsa-miR-16-5p, hsa-miR-21-5p sebagai Regulator IP-10 pada Diagnosis Tuberkulosis Anak

STEFANI CANDRA FIRMANTI, Prof. dr. Sofia Mubarika, M.Med.Sc., Ph.D.; Prof. dr. Tri Wibawa, Ph.D, Sp.MK(K).; dr. Rina Triasih, M.Med(Paed), Sp.A(K), Ph.D

2019 | Disertasi | DOKTOR ILMU KEDOKTERAN DAN KESEHATAN

Latar Belakang: IP-10 merupakan protein yang telah banyak dilaporkan sebagai biomarker potensial untuk diagnosis TB. Akan tetapi nilai diagnosis IP-10 pada TB anak masih bervariasi. MiRNA berperan penting sebagai regulator ekspresi protein dan juga dapat menjadi marker potensial diagnosis TB. Regulasi IP-10 oleh miRNA plasma pada mekanisme infeksi dan diagnosis TB anak menjadi menarik untuk diteliti. Tujuan: menilai akurasi IP-10, hsa-miR-15a-5p, hsa-miR-16-5p, dan hsa-miR21-5p sebagai regulator IP-10 pada diagnosis TB anak, serta mengetahui regulasi miRNA- miRNA tersebut pada ekspresi IP-10. Metode: rekruitmen dilakukan pada anak usia < 15 tahun baik yang bergejala TB maupun tidak melalui dua pendekatan: aktif melalui investigasi kontak dan pasif dari pasien rawat jalan maupun rawat inap yang dirawat di dua rumah sakit di Yogyakarta. Pada seluruh anak dilakukan anamnesis, pemeriksaan fisik, pemeriksaan Rontgen dada AP dan lateral, pengambilan darah untuk pemeriksaan IGRA, IP-10. Pada anak bergejala dilakukan pemeriksaan sputum BTA dan tes molekular cepat TB. Profiling miRNA plasma dilakukan pada pooled plasma 7 anak sakit TB, 7 anak TB laten, dan 7 anak sehat terpapar TB. Validasi miRNA plasma terpilih dilakukan pada individual plasma dari 10 anak sakit TB, 10 anak TB laten, dan 10 anak sehat terpapar TB. Hasil: Dari total 79 anak yang direkrut, 12 anak sakit TB, 16 anak TB laten, 40 anak sehat terpapar TB, dan 11 anak bukan TB tidak terpapar. IP-10 memiliki sensitivitas 92,3%, spesifisitas 95,4%, PPV 92,3%, dan NPV 95,4% untuk diagnosis infeksi TB anak. IP-10 dapat membedakan sakit TB terkonfirmasi mikrobiologis dari TB laten dengan nilai sensitivitas 100%, spesifisitas 93,7%, PPV 66,7%, dan NPV 100%. IP-10 tidak berbeda bermakna pada kelompok sakit TB dan TB laten. Pada diagnosis infeksi TB, Hsa-miR-15a-5p dengan normalisasi hsa-miR-21-5p memiliki nilai sensitivitas 75%, spesifisitas 80%, PPV 88,2%, dan NPV 61,5% sedangkan dengan normalisasi hsa-miR- 16-5p memiliki nilai sensitivitas 85%, spesifisitas 80%, PPV 89,5%, dan NPV 72,7%. Hsa-miR-16-5p dan hsa-miR-21-5p masing-masing memiliki nilai sensitivitas, spesifitas, PPV dan NPV sebesar 70% untuk diagnosis sakit TB. Hsa-miR-15a-5p memiliki korelasi negatif yang bermakna dengan ekspresi IP-10 (r=-0,4, p<0,05). Kesimpulan: IP-10 dapat digunakan untuk diagnosis infeksi TB dan diagnosis sakit TB terkonfirmasi mikrobiologis, namun tidak dapat membedakan sakit TB dan TB laten. Hsa-miR-15a-5p dapat digunakan untuk diagnosis infeksi TB. Hsa-miR-16-5p dan hsa-miR-21-5p dapat digunakan untuk diagnosis sakit TB. Hsa-miR-15a-5p merupakan regulator IP-10 pada TB anak.

Background: IP-10 is a protein that has been widely reported as a potential biomarker for TB diagnosis. However, the diagnosis value of IP-10 in pediatric TB still varies. MiRNA plays an important role as a regulator of protein expression and can also be a potential marker of TB diagnosis. The regulation of IP-10 by plasma miRNA on the mechanism of infection and diagnosis of pediatric TB is interesting to study. Objective: to assess the accuracy of IP-10, hsa-miR-15a-5p, hsa-miR-16- 5p, and hsa-miR21-5p as regulators of IP-10 in the diagnosis of TB in children, and to determine the regulation of these miRNAs in expression of IP-10. Methods: We recruited eligible symptomatic and asymptomatic children (aged < 15 years) actively (by contact investigation) and passively (from inpatient and outpatient clinics in two hospitals, in Yogyakarta, Indonesia). We conducted clinical examination and chest X-ray in all eligible children. Sputum smear and Xpert MTB/RIF assay were performed in children with TB symptoms. All participants underwent blood sampling for QuantiFERON-TBA Gold In-tube (QFT-IT) assay, IP- 10 test. Profiling plasma miRNA were performed in pooled plasma from group of TB disease, latent TB infection, and TB exposed only, 7 children per group. Validation of selected plasma miRNA were performed in individual plasma from 10 children with TB disease, l0 children with latent TB infection, and l0 children with TB exposed only. Results: A total of 79 children were recruited to this study. Twelve children were with TB disease, 16 with latent TB infection (LTBI), 40 were TB exposed only and 11 were non-TB. Children with evidence of TB infection (either with TB disease or LTBI) had higher levels of antigen-stimulated IP-10 compared to non-infected children (TB exposed only and Non-TB) (p=0.000). A cut-off 408.74 pg/mL for antigen-stimulated IP-10 showed high diagnostic accuracy for diagnosis of TB infection (AUC=0.97, 95%CI: 0.92-1.00, sensitivity 92.3%, and specificity 91.9%). However, the stimulated levels of IP-10 between children with TB disease and LTBI were not significantly different (p=0.268). For the diagnosis of TB infection, hsa-miR-15a-5p with normalization of hsa-miR-21-5p has a sensitivity 75%, specificity 80%, PPV 88.2%, and NPV 61.5% whereas with normalization of hsa-miR- 16-5p has a sensitivity 85%, specificity 80%, PPV 89.5%, and NPV 72.7%. Hsa-miR-16-5p and hsa-miR-21-5p have a sensitivity, specificity, PPV and NPV of 70%, respectively, for the diagnosis of TB disease. Hsa-miR-15a-5p has a significant negative correlation with IP-10 expression (r = -0.4, p <0.05). Conclusion: IP-10 can be used for the diagnosis of TB infection and the diagnosis of microbiological confirmed TB disease, but cannot distinguish TB disease and latent TB infection. Hsa-miR-15a-5p can be used for diagnosis of TB infection. Hsa-miR-16-5p and hsa-miR-21-5p can be used for the diagnosis of TB disease. Hsa- miR-15a-5p is an IP-10 regulator in pediatric TB.

Kata Kunci : TB anak, IP-10, hsa-miR-15a-5p, hsa-miR-16-5p, hsa-miR-21-5p, sakit TB, TB laten