Disertasi

Potensi platelet rich plasma untuk pemulihan fungsi ovarium pada tikus model insufisiensi ovarium prematur = The potential of platelet rich plasma for ovarian function revivement at prematur ovarian insufficiency’s rat model.

Latar Belakang: Insufisiensi ovarium prematur (IOP) dapat menyebabkan penurunan kualitas hidup penderitanya akibat berbagai dampak kesehatan jangka panjang sejak usia di bawah 40 tahun. Platelet-rich plasma (PRP) menjadi salah satu pilihan terapi yang mulai dikembangkan, namun belum diketahui pasti efektivitasnya dan keterlibatan oogonial stem cell (OSC). Penelitian ini bertujuan untuk menganalisis potensi PRP dalam pemulihan fungsi ovarium pada dua tingkat keparahan yang berbeda serta dalam mengaktivasi OSC untuk menghasilkan folikel baru pada tikus model IOP yang diinduksi kemoterapi. Metode: Tikus Sprague Dawley betina (usia 10-12 minggu) diinduksi menjadi model IOP ringan dan berat dengan perbedaan lama paparan kemoterapi lalu diberikan PRP intraovarium. Obat kemoterapi penginduksi IOP dipilih menggunakan analisis in silico. Terbentuknya tikus model IOP dikonfirmasi dengan adanya gangguan siklus estrus, peningkatan FSH, penurunan estradiol dan AMH, serta gangguan pada gambaran mikroskopis folikel ovarium. Perbaikan fungsi ovarium dinilai dari siklus estrus, kadar hormon FSH, estradiol dan AMH, gambaran folikel ovarium pada pemeriksaan mikroskopis, serta ekspresi gen FOXO3 sebagai penanda folikulogenesis. Ekspresi gen dan protein DDX4 dipilih sebagai penanda aktivasi OSC. Analisis differentially-expressed genes (DEG) terhadap hasil RNA sequencing jaringan ovarium dilakukan untuk mengeksplorasi proses biologis dan jalur pensinyalan yang terkait dengan pemulihan fungsi ovarium. Hasil: Cisplatin diprediksi memiliki potensi besar menyebabkan IOP. Tikus model IOP ringan dan berat dapat dibuat dengan injeksi intraperitoneal cisplatin 3 mg/kgBB per minggu selama 3 dan 5 minggu. Tikus model IOP ringan menunjukkan perbaikan fungsi ovarium dan peningkatkan aktivasi OSC. Hal ini belum terlihat pada tikus model IOP berat. Hasil analisis DEG mengkonfirmasi perbaikan di tingkat transkriptomik untuk berbagai proses biologis terkait fungsi ovarium pada tikus model IOP berat yang diberikan PRP, terutama melalui jalur pensinyalan PI3K/Akt dengan gen targen PI3K dan Akt. Kesimpulan: Potensi PRP dalam memperbaiki fungsi ovarium lebih nyata pada tikus model IOP ringan. Pemberian PRP juga meningkatkan aktivasi OSC.
Kata kunci: Insufisiensi ovarium prematur; platelet-rich plasma; tikus model, fungsi ovarium; cisplatin


Background: Premature ovarian insufficiency (POI) may decrease the quality of life of women under 40-years old due to various long-term health impacts. Currenttly, platelet-rich plasma (PRP) has been proposed as one of therapeutic options, however its effectiveness and involvement of oogonial stem cells (OSC) are not known clearly. This study aimed to analize the potential of PRP in the revivement the ovarian function at two different severity levels of POI rat model and its effect on OSC activation to produce new follicles. Methods: Female Sprague Dawley rats (aged 10-12 weeks) were induced into mild and severe POI models with different length of chemotherapy exposure, and treated with an intraovarian injection of PRP. Chemotherapy agent to induced POI was selected using in silico analysis. The formation ofPOI rat model was confirmed by the estrus cycle disorder, increased FSH, decreased estradiol and AMH, and pathology in the microscopic features of ovarian follicles. Improvement in ovarian function was assessed through the estrus cycle, levels of FSH, estradiol and AMH, ovarian follicle images on microscopic examination, and expression of FOXO3 gene as a marker of foliculogenesis. Expression of gene and DDX4 protein were chosen as markers of OSC activation. To explore the biological processes and signaling pathways associated with the restoration of ovarian function, a differentially-expressed genes (DEG) analysis of ovarian tissue RNA sequencing data was conducted. Results: Cisplatin was predicted to have a great potential to cause IOP. Mild- and severe-IOP rat models can be established with intraperitoneal injection of 3 mg/kgBW cisplatin per week for 3 and 5 weeks. Mild-IOP rat models showed improvement of ovarian function and OSC activation. These enhancement was not found in severe-IOP rat models. DEG analysis emphasized improvements at the transcriptomic level for various biological processes related to ovarian function in severe-POI rat model treated with PRP, primarily PI3K/Akt signaling pathway with PI3K and Akt as target genes. Conclusion: The potential for PRP to revive ovarian function was more pronounced in mild-POI rat model. The provision of PRP also increased OSC activation.
Keywords: premature ovarian insufficiency; platelet-rich plasma; rat model; ovarian function; cisplatin