Syarifudin, NurulWulandari (2012) Peran Regulasi Insulin terhadap Penerusan Meiosis-1 dalam Sitoplasma Oosit Ikan Lele (Clarias sp). Magister thesis, Universitas Brawijaya.
Abstract
Perkembangan budidaya ikan lele dimasyarakat indonesia sudah berkembang pesat. Penelitian yang melibatkan oosit ikan atau gonad betina banyak dilakukan, namun hingga saat ini informasi dasar mengenai maturasi oosit lele masih perlu dikaji lebih mendalam. Maturasi oosit lele tidak hanya dilakukan dengan pemberian hormon reproduksi, tetapi diketahui bahwa insulin juga dapat memicu maturasi. Oleh karenanya perlu dilakukan penelitian tentang respon oosit terhadap paparan insulin aspart . Tujuan dari penelitian ini adalah untuk mengetahui kemunculan protein p21 ras, Akt/PKB dan MAPK dalam sitoplasma oosit ikan lele yang dipapar dengan insulin aspart dan mengetahui jumlah oosit yang dapat mengalami penerusan meiosis-1 setelah dipapar dengan insulin aspart. Penelitian dilaksanakan mulai bulan April sampai September 2012. Metode penelitian yang digunakan adalah metoda eksperimen. Kultur oosit yang belum matang (vitellogenesis) dalam medium dengan konsentrasi insulin 0,01; 0,1; 1; 10 Unit/ml selama 24 jam dan pada jam ke 6, 8, 10, 12, 14, 16, 20, 24 dilakukan ekstraksi protein pada sitoplasma oosit ikan lele. Selanjutnya di amati oosit yang mengalami penerusan meiosis-1. Oosit yang mengalami penerusan meiosis-1 ditandai dengan germinal vesicle migration (GVM) atau germinal vesicle break down (GVBD). Identifikasi molekul protein p21 ras, Akt/PKB dan MAPK mempunyai berat molekul 21 kDa, 110 kDa dan 42 kDa dilakukan dengan metode elektroforesis. Analisa data dibantu dengan progam ANOVA dan jika berbeda nyata akan dilanjutkan dengan uji BNT. Oosit ikan lele memberikan respon yang positif terhadap insulin aspart yang ditandai dengan munculnya protein p21 ras, Akt/PKB dan MAPK, jika oosit tersebut dipapar dengan insulin aspart dan tidak muncul pada oosit yang tidak dipapar dengan insulin. Kemunculan protein ini diduga sebagai indikator aktivitas insulin terhadap proses penerusan meiosis-1 yang ditandai dengan adanya GVM dan GVBD. Prosentase tertinggi rata-rata oosit yang mengalami penerusan meiosis-1 sebesar 82 % terjadi pada perlakuan 10 Unit/ml dan terendah pada perlakuan 0,01 Unit/ml yaitu 14 %. Sedangkan oosit yang tidak dipapar dengan insulin aspart tidak ada yang mengalami penerusan meiosis-1. Semakin tinggi konsentrasi paparan insulin dalam medium dan semakin lama waktu inkubasi, maka jumlah oosit yang dapat mengalami penerusan meiosis-1 juga akan semakin tinggi. Peningkatan jumlah penerusan meiosis-1 paling signifikan adalah konsentrasi 1 unit/ml sebesar 81%. Jika konsentrasi tersebut ditingkatkan lagi hingga mencapai 10 Unit/ml ternyata tidak berdampak peningkatan jumlah penerusan meiosis-1 yang signifikan, sehingga diduga konsentrasi optimum insulin aspart yang dapat direspon oleh oosit ikan lele untuk merangsang terjadinya penerusan meiosis-1 adalah 1 Unit/ml. Diharapkan perlu adanya penelitian lebih lanjut mengenai tingkat kemampuan insulin aspart menurunkan level cAMP dalam oosit dan adanya penggunaan bahan antagonis terhadap aksi insulin sebagai perbandingan.
English Abstract
In the two past decades, methods of fish seed production, intensive and pond rearing of catfish in Indonesia were developed. Many studies have been using catfish gonads and oocytes to be investigated, but information about mechanisms of maturation and it`s supported is still elusive and need examined more detailed. Naturally fish final maturation in are induces by steroids produced by follicle cells surrounding oocyte. Several studies have been reported that insulin has also ability to trigger the meiotic arrest withdrawn in fish oocytes for initiation of maturation process. The recent study is thought to be deserved for understanding the catfish oocytes maturation by insulin aspart exposure. The aims of recent study were to determine: 1) the emergence of p21 ras protein, protein kinase B (PKB/Akt), and mitogen activated protein kinase (MAPK) in catfish oocyte exposed by aspartic insulin. 2) to determine the amount of catfish oocytes were conducting meiotic resumption after exposed by aspartic insulin. The research was carried out from April to September 2012. This research was applied randomized experimental design. Immature catfish oocytes were incubated for 24 hours in Hank`s-basic medium with different insulin concentration e.g 0.01, 0.1, 1.0, and 10 Unit/ml. Oocytes were incubated observed at 6, 8, 10, 12, 14, 16, 20, 24 hours. Observation aimed to the occurrence of meiotic resumption indicated by the presence of visual Germinal vesicle migration (GVM), germinal vesicle breakdown (GVBD) respectively. For further observation, we performed the electrophoresis SDS-PAGE to find out the catfish oocyte maturation involved molecules particularly p21 ras protein, PKB, and MAPK. Collected data was analysed by ANOVA and was distinguished by LSD test. The result shows that catfish oocytes were treated with insulin aspart for various concentrations and time incubation demonstrated that there was a positive correlation between the insulin aspart and the extended meiotic resumption indicated by GVM and or GVBD. For further evidence we was running the crude protein extracted from oocytes in SDS PAGE electrophoresis gels and we found presence of p21 ras protein, PKB, and MAPK kinase. These three molecules were absence in sample without exposed by insulin aspart. We suggest that the differences in terms of attendance of these molecules in gels extracted from both treated and control catfish oocytes are constitute a markedly has been meiotic resumption indicated by GVM and or GVBD. The higher meiotic resumption percentage was 82% treated by 10 U/ml insulin aspart, and the conversely the least one was 0.01U/ml i.e 14%. All of the oocytes without treated by insulin aspart had no undergone meiosis resumption. The higher concentration, the higher of resumption as well as time incubation in catfish oocytes. Increasing the number of resumption meiosis is the most significant concentration of 1 U/ml by 81%. If the concentration is increased further up to 10 U/ml did not affect resumption meiosis increase in the number of significant. So that the optimum concentration of insulin aspart expected to respond by catfish oocytes to stimulate resumption meiosis is 1 U/ml. Expected need for further research on the level of insulin aspart lower cAMP levels in the oocytes. The use of antagonists to the activation of insulin in comparison.
| Item Type: | Thesis (Magister) |
|---|---|
| Identification Number: | TES/597.49/SYA/p/041205647 |
| Subjects: | 500 Natural sciences and mathematics > 597 Cold-blooded vertebrates > 597.4 Miscellaneous superorders of Actinopterygii |
| Divisions: | S2/S3 > Magister Budidaya Perairan, Fakultas Perikanan dan Ilmu Kelautan |
| Depositing User: | Endro Setyobudi |
| Date Deposited: | 14 Jan 2013 09:40 |
| Last Modified: | 14 Jan 2013 09:40 |
| URI: | http://repository.ub.ac.id/id/eprint/157873 |
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