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Studies of egg production between Leghorn layers and Taiwan country chickens using ovarian follicular cell functionality
|關鍵字:||Taiwan country chicken|
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|摘要:||雞隻產蛋效率受到卵巢濾泡細胞之生長發育影響，而台灣土雞雖然其肉質具有良好風味且廣受國人喜愛，但產蛋效率普遍較來航蛋雞（Leghorn）差，此將影響台灣土雞隻市場之推廣。因此，本研究之目的在於探討台灣土雞卵巢濾泡生成過程中其細胞凋亡敏感性（susceptibility）、激性腺素受體表現以及其類固醇生成能力之改變，並進一步分析比較於高低產蛋雞隻之差異性表現與影響機制。此外，本研究亦嘗試於實驗室建立雞隻卵巢濾泡顆粒細胞體外培養系統，期供後續更深入探討土雞卵巢濾泡細胞之功能。試驗一，觀察雞隻卵巢各階段成熟度不同之濾泡顆粒細胞與鞘膜細胞中P450scc、P450arom、FSH-R、LH-R、Bcl-2、Cas-3 mRNA之表現，結果顯示於Leghorn line、L2 line、B line中，隨濾泡逐漸成熟而顆粒細胞層內P450scc、LH-R mRNA表現量增加，於鞘膜細胞層中，LH-R mRNA隨濾泡成熟而增加至F3，F2~F1則逐漸下降；P450arom mRNA大多表現於鞘膜細胞中，但於Leghorn line和B line F5、F6顆粒細胞層中仍可偵測到P450arom mRNA之表現；FSH-R mRNA之表現，於Leghorn line的顆粒細胞層及鞘膜細胞層及L2 line的顆粒細胞層中，隨濾泡成熟而有下降的趨勢；Bcl-2 mRNA於L2 line濾泡顆粒細胞及鞘膜細胞中之表現，並未隨濾泡發育階段而有顯著性差異，而於B line濾泡顆粒細胞及鞘膜細胞中，Bcl-2 mRNA皆隨濾泡逐漸成熟而減少；Cas-3 mRNA於L2 line、B line之表現未隨著發育過程而有顯著性差異。試驗二，體外培養卵巢濾泡顆粒細胞，並觀察P450scc、P450arom、FSH-R、LH-R、Bcl-2、Cas-3 mRNA之表現，結果顯示於Leghorn line、L2 line細胞中，經FSH、LH、8Br-cAMP處理之濾泡顆粒細胞內P450scc mRNA之表現量隨濾泡顆粒細胞發育而有上升趨勢，但於B line中，經由FSH、8Br-cAMP處理後顆粒細胞內P450scc mRNA表現量下降，可能導致P4無法生成，進一步影響排卵；經FSH、TGF-α處理後，顆粒細胞內FSH-R mRNA之表現量隨濾泡發育而下降；LH-R mRNA之表現量於各別添加FSH、LH、8Br-cAMP、TGF-β1之培養下，隨濾泡顆粒細胞發育而漸增；Bcl-2 mRNA之表現量於各別添加FSH、LH、8Br-cAMP之培養下有上升趨勢。試驗三，將雞隻濾泡生成過程中，濾泡細胞激性腺素受體之表現、細胞凋亡基因之表現與類固醇生成能力之變化，和具代表性之高低產蛋雞隻做相關性之比較研究，結果顯示B line濾泡進入排序後，顆粒細胞P450scc, FSH-R mRNA之表現量較Leghorn、L2 line有顯著地減少；Cas-3 mRNA之表現量較 L2 line高；而B line F1、F2中，LH-R mRNA之表現量較L2 line低；Bcl-2 mRNA 之表現量於B line隨濾泡逐漸成熟而減少，於L2 line則無顯著差異。綜合上述結果，推測B line產蛋性能偏低之原因可能為其濾泡細胞對激性腺素刺激反應較不敏感，導致其較差之類固醇內泌素生成能力，進而影響其產蛋效率。但詳細機制仍需更進一步探討。|
The development and differentiation of follicular cells are associated with ovarian function and in hens exect effects on egg production. Although the rich flavor of Taiwan country chicken meat is preferred in the market in Taiwan, the poor performance in egg production, compared to Leghorn layers, greatly hinders the production of Taiwan country chicken. Therefore, the aims of this study were conducted in relation to egg production to ovarian function using susceptibility of cell apoptosis, expressions of gonadotrophin receptors and acquirement of steroidogenetic ability on the follicular cells during folliculogenesis in the hen. The comparisons of the differential expressions between Taiwan chicken lines with high and low egg production were also evaluated. Additionally, an in vitro follicular cell culture system was established. In Experiment 1, The granulose cells (GC) and theca cells (TC) were isolated from Leghorn, L2 and B lines at the different developmental stages, and the expressions of P450scc, P450arom, FSH-R, LH-R, Bcl-2 and Caspase-3 were analyzed by semi quantitative RT-PCR. The results showed that the expressions of LH-R and P450scc in B and L2 lines increased with the maturation of follicles in GC, whereas FSH-R decreased. The expression of Bcl-2 decreased with the maturation of follicles in B line's GC. No significant differences were found in the expressions of Caspase-3 during folliculogenesis. In Experiment 2, an in vitro follicular cell culture system was established. The effects of various growth factors, including FSH, LH, 8Br-cAMP, TGF-β1 and TGF-α on the expression of genes in the granulosa cells were analyzed. Results showed that the expression of P450scc, LH-R and Bcl-2 in Leghorn and L2 lines increased with the maturation of granulose cells in the presence of FSH/LH/8Br-cAMP/TGF-β1/TGF-α after 24 h culture, whereas FSH-R decreased. In Experiment 3, the expressions of P450scc, P450arom, FSH-R, LH-R, Bcl-2 and Caspase-3 in follicular cells among Leghorn, L2 and B lines were compared. It is shown that P450scc, FSH-R and LH-R in B line were lower compared to those in Leghorn and L2 lines, whereas Cas-3 is higher. Bcl-2 decreased with the maturation of follicles in B line's GC, but no significant differences were found in L2 line. In conclusion, the low egg production in B line is very likely due to the poor responsiveness to gonadotrophin and thereby leading to low production of steroid hormones. Further investigations are required in order to dig the depth of the mechanism between ovarian function and egg production among different lines of chickens.
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