小鼠精子各項參數和Hedgehog在精卵交互作用中的功能

本實驗室利用死亡後，環境中溫度和時間對精子的影響的模式，觀察精子的狀態。之後再根據先前文獻指出，Hedgehog 蛋白對於精子的發育和運動有ㄧ定的影響力，對於卵子則有促進濾泡發育的功用，於是觀察在體外受精的環境裡加入 Hedgehog 蛋白或其抑制劑 (Cyclopamine、5E1)，對於授精的影響。設計在小鼠死亡後，將小鼠屍體存放在 4℃ 或 20℃ 環境下，每隔 12 hr 觀察ㄧ次精子狀態。結果顯示：精子若無法在個體死亡後立即取出，保存遺體條件的溫度較高的情形下，會使精子大量死亡 （對照組: 96%，20℃ 12-36 hr:＜82%，p<0.05）、嚴重傷害精子活動力（獲能後活力分級“3”：對照組:27%，20℃ 12-36 hr:＜2.5%，p<0.05）和精子的外型大量趨向不正常（不正常外型比例：對照組：<1%，20℃ 12-36 hr:>50%，p<0.05）；在較低溫的環境下，雖然存活率可以維持一定程度，但是時間ㄧ長，精子的活動力仍會下降，而且對精子的外型仍然有著嚴重的傷害（不正常外型比例：對照組：<1%，4℃ 12-36 hr:>31%，p<0.05）。體外受精試驗設計四組：對照組、模擬 Hedgehog 功能的 rHh 組和阻斷Hedgehog 功用的 5E1 和 cyclpamine 兩組。體外受精的果顯示： rHH 可以增加受孕機率達 100%，然而在 5E1 和 cyclopamine 中，除了會降低受孕機率到 68% 和50% 外（p<0.05）還會延緩受精卵的發育進度，在 5E1 組中受精卵有發育型態不正常的情形出現。因此若希望保有良好的精子品質，最好的方式是盡快取出體外作適當的保存；而體外受精的結果，提供了將來醫學上提高受精機率的一個方針。

In this study, a postmortem model was used to observe the sperm parameters under various environmental conditions (4℃ and 20℃ in 12 hr, 24hr, 36hr). Furthermore, since Hedgehog had been reported to affect sperm development and motility, the effects of Hedgehog and Hedgehog inhibitors (cyclopamine and 5E1) in sperm-egg interactions were also investigated. The results showed that sperm survival rates (control: 96%, Exp:<82%), motility (control: 27%, Exp:<2.5%), and morphology (abnormal morphology : control: <1%, Exp:>50%) were highly affected under higher temperature (p<0.05). Under lower temperature, sperm motility and morphology were also affected (abnormal morphology: control: <1%, Exp:>31%), although survival rate seemed unaffected. Sperm-egg interaction data showed that fertilization rates were improved with Hedgehog addition (100%), in contrast to the lower rates with cyclopamine (50%) and Hedgehog antibody (5E1) (68%) addition. In addition, embryo morphology was abnormal under the effect of 5E1. In conclusion, postmortem sperm parameters were highly affected by environmental factors and Hedgehog protein supplementation may be a good method to improve the fertilization rates under in vitro conditions.