Tesis

Pengembangan teknik ko-kultur adiposed derived mesenchymal stem cells dan kardiomiosit pada amnion bilayer = Development of a co-culture technique for adiposed derived mesenchymal stem cells and cardiomyocytes in the amnion bilayer.

Latar belakang: Tata laksana penyakit jantung koroner (PJK) saat ini seperti medikamentosa, intervensi koroner dan operasi pintas jantung koroner, hanya meningkatkan kualitas hidup pasien, tetapi tidak menghentikan dan memperbaiki kerusakan otot jantung yang terjadi sehingga pasien berujung gagal jantung. Terapi sel punca saat ini dikembangkan untuk memperbaiki kerusakan otot jantung yang terjadi. Terapi sel punca menggunakan mesenchymal stem cells (MSCs) telah muncul sebagai sumber sel punca yang potensial, dikarenakan memiliki sifat multipoten, anti inflamasi dan dapat berdiferensiasi menjadi kardiomiosit. Untuk mendukung repopulasi sel punca yang diberikan secara langsung pada pasien PJK, diperlukan teknik rekayasa jaringan yang menggunakan pendekatan sistem ko-kultur pada perancah/scaffold yang dikenal dengan istilah ”scaffold-based cell therapy”. Penelitian ini bertujuan untuk mengetahui rasio densitas ko-kultur antara kardiomiosit dan adiposed-derived mesenchymal stem cells (ADMSCs), serta usia donor ADMSCs yang optimal pada perancah untuk menghasilkan differensiasi sel kardiomiosit baru Metode: Penelitian ini merupakan studi eksperimental in vitro yang dilakukan di Pelayanan Jantung Terpadu Rumah Sakit Umum Pusat Nasional dr. Cipto Mangunkusumo (PJT RSUPNCM) dan laboratorium Institute of Medical Education and Research in Indonesia, Universitas Indonesia (IMERI-UI). Jumlah pendonor sel kardiomiosit sebanyak 4 subjek, pendonor ADMSc usia tua (60-70 tahun) sebanyak 3 subjek dan pendonor ADMSc usia muda (30-40 tahun) sebanyak 3 subjek. Kardiomiosit dan ADMSc yang telah diisolasi dilakukan seeding pada perancah, kemudian sel kardiomiosit hasil ko-kultur dilakukan uji viabilitas serta pemeriksaan aktivitas kardiomiogenesis. Hasil: Hasil ko-kultur pada rasio densitas sel kardiomiosit dengan ADMSCs adalah 1:5 dan 1:6, tidak berbeda bermakna baik pada sel yang viabel maupun sel mati pada hari ke2 (64 banding 95 sel) dan pada hari ke-8 (173 banding 72 sel) pascako-kultur (p > 0,05). Pada uji viabilitas dengan ATPlite pada hari ke-9 terpadat perbedaan bermakna antara kelompok ADMSCs-60 dan kelompok ADMSCs-30 (502.600 banding 937.947, p = 0,004). Namun, penilaian aktivitas kardiomiogenesis, kelompok ADMSCs-30 memiliki ekspresi gen PPARƳ yang lebih tinggi pada hari ke-5 dan ke-9, serta α-aktinin, MHC dan cTnT hari ke-5 juga ditemukan lebih tinggi secara signifikan (p < 0,005). Simpulan: Tidak ada perbedaan yang bermakna antara densitas ko-kultur sel kardiomiosit dengan ADMSCs 1:5 dan 1:6, serta hasil ko-kultur antara sel kardiomiosit dengan ADMSCs kelompok usia muda memiliki hasil yang jauh lebih baik dari segi morfologi, viabilitas sel dan kapasitas kardiomiogenesis.
Kata Kunci : Kardiomiosit; ADMSCs, scaffold-based cell therapy


Background: The current management of coronary heart diseases (CHD), such as medication, coronary intervention, and coronary heart bypass surgery, merely improves patients' quality of life but does not stop and repair defects in cardiomyocytes leading to heart failure. Stem cell therapy is currently developed to repair damaged cardiomyocytes. Stem cell therapy using mesenchymal stem cells (MSCs) has emerged as a potential resource of stem cells due to its multipotent, anti-inflammatory properties and may differentiate into cardiomyocytes as well. To support the repopulation of stem cells administered directly to CHD patients, a tissue engineering technique that uses a coculture system approach on a scaffold is needed, known as "scaffold-based cell therapy". This study aims to determine the co-culture density ratio between cardiomyocytes and adipose-derived mesenchymal stem cells (ADMSCs), as well as the optimal age of donor ADMSCs on the scaffold to produce new cardiomyocyte cell differentiation. Methods: This research was an in vitro experimental study conducted at the Integrated Cardiac Service of the dr. Cipto Mangunkusumo General Hospital and the Institute of Medical Education and Research laboratory in Indonesia, University of Indonesia (IMERI-UI). The number of cardiomyocyte cell donors was four subjects, three subjects were elder ADMSc donors (60-70 years), and 3 were young ADMSc donors (30-40 years). The isolated cardiomyocytes and ADMSc were seeded on the scaffold, and then the co-cultured cardiomyocytes were assessed for viability and examined for cardiomyogenesis activity. Results: Co-culture results with density ratio of cardiomyocytes to ADMSCs were 1:5 and 1:6, not significantly different in both live and dead cells on day 2 (64 vs 95 cells) and day 8 (173 vs 72 cells). postco-culture (p >0.05). Viability test with ATPlite on day 9, there was a significant difference between the ADMSCs-60 group and the ADMSCs30 group (502,600 vs 937,947, p = 0.004). In addition, assessing cardiomyogenic activity, the ADMSCs-30 group had higher PPARƳ gene expression at days 5 and 9, and αactinin, MHC and cTnT at day 5 were also found to be significantly higher (p < 0.005). Conclusion: No significant difference between the co-culture density of cardiomyocytes with ADMSCs 1:5 and 1:6, and the co-culture between cardiomyocytes and ADMSCs in the young age group had significantly better results morphologically, cell viability and cardiomyogenesis capacity.
Key Words : Cardiomyocytes, ADMSCs, scaffold-based cell therapy