CAJ | 학술논문

本试验旨在研究葡萄糖转运载体（ GLUT）4、CLUT2在罗非鱼不同组织中的表达及其对注射葡萄糖的响应。利用实时荧光PCR的方法从罗非鱼肌肉中克隆得到GLUT4的cDNA片段，其GenBank登录号为JN900493，大小为603 bp，编码200个氨基酸。通过实时荧光半定量PCR检测，比较GLUT4和GLUT2在肌肉、心脏和肝脏中的表达差异，结果显示GLUT4在肌肉和心脏中的表达量较高，而在肝脏中GLUT4的表达量则较低；GLUT2在肝脏中的表达量最高，而在肌肉和心脏中则表现出极低的表达量。选取体重约为80 g的罗非鱼150尾，随机分2个组，每组3个重复，每个重复25尾。试验组腹腔注射葡萄糖（每100 g体重30 mg），对照组以相同剂量腹腔注射0．7％的无菌生理盐水。在注射前（0 h）和注射后的1、3、6和12 h分别进行采样测定。结果显示：1）试验组血浆葡萄糖含量在注射葡萄糖后1 h时达到最高，并显著高于对照组（ P＜0．05），而后开始下降，3 h后恢复到正常水平；试验组血浆胰岛素含量在葡萄糖注射后3 h时达到最高，并显著高于对照组（P＜0．05），而后开始下降，12 h后恢复到正常水平。2）试验组肌肉中GLUT4 mRNA的相对表达量在注射葡萄糖后3 h开始升高，在12 h时达到最高，此时显著高于对照组（ P＜0．05）；试验组心脏中GLUT4 mRNA的相对表达量在注射葡萄糖后1 h即较对照组显著升高（ P＜0．05），而后随着时间的推移逐渐降低，在6 h时恢复到正常水平；注射葡萄糖后，肝脏中GLUT2 mRNA的相对表达量没有显著变化（ P＞0．05）。结果表明，注射葡萄糖后瞬时升高了罗非鱼的血浆葡萄糖含量，相对于血浆葡萄糖含量的升高，血浆胰岛素含量的升高相对延迟，而肌肉中GLUT4 mRNA相对表达量的提高又延迟于胰岛素含量的升高，从而加重了罗非鱼对葡萄糖的代谢负担。本試驗旨在研究葡萄糖轉運載體（ GLUT）4、CLUT2在囉非魚不同組織中的錶達及其對註射葡萄糖的響應。利用實時熒光PCR的方法從囉非魚肌肉中剋隆得到GLUT4的cDNA片段，其GenBank登錄號為JN900493，大小為603 bp，編碼200箇氨基痠。通過實時熒光半定量PCR檢測，比較GLUT4和GLUT2在肌肉、心髒和肝髒中的錶達差異，結果顯示GLUT4在肌肉和心髒中的錶達量較高，而在肝髒中GLUT4的錶達量則較低；GLUT2在肝髒中的錶達量最高，而在肌肉和心髒中則錶現齣極低的錶達量。選取體重約為80 g的囉非魚150尾，隨機分2箇組，每組3箇重複，每箇重複25尾。試驗組腹腔註射葡萄糖（每100 g體重30 mg），對照組以相同劑量腹腔註射0．7％的無菌生理鹽水。在註射前（0 h）和註射後的1、3、6和12 h分彆進行採樣測定。結果顯示：1）試驗組血漿葡萄糖含量在註射葡萄糖後1 h時達到最高，併顯著高于對照組（ P＜0．05），而後開始下降，3 h後恢複到正常水平；試驗組血漿胰島素含量在葡萄糖註射後3 h時達到最高，併顯著高于對照組（P＜0．05），而後開始下降，12 h後恢複到正常水平。2）試驗組肌肉中GLUT4 mRNA的相對錶達量在註射葡萄糖後3 h開始升高，在12 h時達到最高，此時顯著高于對照組（ P＜0．05）；試驗組心髒中GLUT4 mRNA的相對錶達量在註射葡萄糖後1 h即較對照組顯著升高（ P＜0．05），而後隨著時間的推移逐漸降低，在6 h時恢複到正常水平；註射葡萄糖後，肝髒中GLUT2 mRNA的相對錶達量沒有顯著變化（ P＞0．05）。結果錶明，註射葡萄糖後瞬時升高瞭囉非魚的血漿葡萄糖含量，相對于血漿葡萄糖含量的升高，血漿胰島素含量的升高相對延遲，而肌肉中GLUT4 mRNA相對錶達量的提高又延遲于胰島素含量的升高，從而加重瞭囉非魚對葡萄糖的代謝負擔。
본시험지재연구포도당전운재체（ GLUT）4、CLUT2재라비어불동조직중적표체급기대주사포도당적향응。이용실시형광PCR적방법종라비어기육중극륭득도GLUT4적cDNA편단，기GenBank등록호위JN900493，대소위603 bp，편마200개안기산。통과실시형광반정량PCR검측，비교GLUT4화GLUT2재기육、심장화간장중적표체차이，결과현시GLUT4재기육화심장중적표체량교고，이재간장중GLUT4적표체량칙교저；GLUT2재간장중적표체량최고，이재기육화심장중칙표현출겁저적표체량。선취체중약위80 g적라비어150미，수궤분2개조，매조3개중복，매개중복25미。시험조복강주사포도당（매100 g체중30 mg），대조조이상동제량복강주사0．7％적무균생리염수。재주사전（0 h）화주사후적1、3、6화12 h분별진행채양측정。결과현시：1）시험조혈장포도당함량재주사포도당후1 h시체도최고，병현저고우대조조（ P＜0．05），이후개시하강，3 h후회복도정상수평；시험조혈장이도소함량재포도당주사후3 h시체도최고，병현저고우대조조（P＜0．05），이후개시하강，12 h후회복도정상수평。2）시험조기육중GLUT4 mRNA적상대표체량재주사포도당후3 h개시승고，재12 h시체도최고，차시현저고우대조조（ P＜0．05）；시험조심장중GLUT4 mRNA적상대표체량재주사포도당후1 h즉교대조조현저승고（ P＜0．05），이후수착시간적추이축점강저，재6 h시회복도정상수평；주사포도당후，간장중GLUT2 mRNA적상대표체량몰유현저변화（ P＞0．05）。결과표명，주사포도당후순시승고료라비어적혈장포도당함량，상대우혈장포도당함량적승고，혈장이도소함량적승고상대연지，이기육중GLUT4 mRNA상대표체량적제고우연지우이도소함량적승고，종이가중료라비어대포도당적대사부담。
This experiment was conducted to determine the expression of glucose transporter ( GLUT) 4 and GLUT2 in different tissues of tilapia ( Oreochromis niloticus) and its response to glucose injection. A partial cDNA of GLUT4 was cloned from muscle of tilapia by real-time PCR method. The partial cDNA of GLUT4 ( GenBank accession No. was JN900493) was 603 bp, and encoded 200 amino acids. The expression differ-ences of GLUT4 and GLUT2 in muscle, heart and liver were compared by semi-quantitative real-time PCR de-tection. The results showed that the expression level of GLUT4 in muscle and heart was higher, while that in liver was lower; the expression level of GLUT2 in liver was higher, while that in muscle and heart was very low. A total of 150 tilapia with the body weight about 80 g were randomly allotted to 2 groups with 3 replicates per group and 25 fish per replicate. In experimental group, glucose with the dose of 30 mg per 100 g body weight was injected intraperitoneally into the fish, while the fish in control group were injected with 0.7% ster-ile saline water as the same dose and method. Samples were collected and measured before infection ( 0 h) and at 1, 3, 6 and 12 h after infection, respectively. The results showed as follows:1) the maximum plasma glu-cose content of experimental group was observed at 1 h after injection which was significantly higher than that of control group ( P<0.05) , and then began to decreaseed and returned to normal level at 3 h after injection;the maximum plasma insulin content of experimental group was observed at 3 h after injection which was signif-icantly higher than that of control group ( P<0.05) , and then began to decrease and returned to normal level at 12 h after injection. 2) The relative expression level of GLUT4 mRNA in muscle of experimental group began to increase at 3 h after injection, and reached the maximum at 12 h after injection which was significantly high-er than that of control group ( P<0.05);the relative expression level of GLUT4 mRNA in heart at 1 h after in-jection of experimental group was significantly higher than that of control group ( P<0.05) , and then decreased gradually with the passage of time and return to normal level at 6 h after injection;the relative expression level of GLUT4 mRNA in liver had no significant change after glucose injection ( P>0.05) . The results suggest that injecting glucose can instantaneously increase the plasma glucose content of tilapia, but the increase of plasma insulin content is relatively delayed compare with plasma glucose content, and the relative expression level of GLUT4 mRNA in muscle show a delayed increase relative to plasma insulin content, then the burden of glucose metabolism in tilapia is increased.