CAJ | 학술논문

为探讨抵抗素成熟肽对猪葡萄糖和脂肪代谢的影响，将猪抵抗素成熟肽（M R ）编码序列克隆至pET-32a（＋），在大肠埃希菌中表达重组MR融合蛋白，并用Ni-NTA亲和层析纯化。经肠激酶消化切除融合蛋白，用Ni-NTA分离MR。随后将18头5月龄去势巴马香猪随机分为3个组，对照组肌肉注射生理盐水0．5 mL ，试验1组和2组分别肌肉注射20 ng/kg 和40 ng/kg 的M R ，于注射后0、1、2、3、4、5 h分别采血，检测血清葡萄糖及脂肪代谢相关指标。结果表明，重组M R融合蛋白分子质量为29 ku ，切除融合蛋白的MR分子质量为12 ku ，纯化的MR浓度约3 mg/L。给予外源性MR后，试验组猪血清葡萄糖显著高于对照组（P＜0．05）；脂蛋白酯酶活性亦有升高的趋势，但组间差异不显著（P＞0．05）；两个试验组猪血清甘油三酯含量均显著降低，试验2组显著低于试验1组和对照组（P＜0．05）；试验2组猪血清游离脂肪酸含量显著高于试验1组和对照组（P＜0．05）。由此可见，抵抗素能抑制组织细胞的葡萄糖摄入，并通过提高脂蛋白酯酶活性促进甘油三酯分解，在调节机体能量代谢与能量平衡中发挥重要作用。
위탐토저항소성숙태대저포도당화지방대사적영향，장저저항소성숙태（M R ）편마서렬극륭지pET-32a（＋），재대장애희균중표체중조MR융합단백，병용Ni-NTA친화층석순화。경장격매소화절제융합단백，용Ni-NTA분리MR。수후장18두5월령거세파마향저수궤분위3개조，대조조기육주사생리염수0．5 mL ，시험1조화2조분별기육주사20 ng/kg 화40 ng/kg 적M R ，우주사후0、1、2、3、4、5 h분별채혈，검측혈청포도당급지방대사상관지표。결과표명，중조M R융합단백분자질량위29 ku ，절제융합단백적MR분자질량위12 ku ，순화적MR농도약3 mg/L。급여외원성MR후，시험조저혈청포도당현저고우대조조（P＜0．05）；지단백지매활성역유승고적추세，단조간차이불현저（P＞0．05）；량개시험조저혈청감유삼지함량균현저강저，시험2조현저저우시험1조화대조조（P＜0．05）；시험2조저혈청유리지방산함량현저고우시험1조화대조조（P＜0．05）。유차가견，저항소능억제조직세포적포도당섭입，병통과제고지단백지매활성촉진감유삼지분해，재조절궤체능량대사여능량평형중발휘중요작용。
The present work was aimed to further investigate the effects of resistin mature peptide on glu-cose and fat metabolism in pigs .The fragment encoding for resistin mature peptide(MR) was cloned into the expression vector pET-32a(+ ) ,and expressed in Ecoli BL-21(DE3) .The recombinant MR fusion pro-tein was purified by Ni-NTA affinity chromatography .Then the fusion peptide was cleaved from the re-combinant MR fusion protein by enterokinase digestion ,and the purified MR was isolated by Ni-NTA af-finity chromatography subsequently .18 5-month-old castrated Bama pigs were randomly divided into 3 groups ,the pigs in control group received nomal saline 0 .5 mL intramuscularly ,and the animals in experi-mental groups respectively received 20 and 40 ng MR per kilogram body weight ,blood samples were col-lected at 0 ,1 ,2 ,3 ,4 ,5 h after administration ,and the biochemical parameters associated glucose and fat metabolism were detected .The results showed that the recombinant MR fusion protein with molecular weight of 29 ku ,MR cleaved fusion protein with molecular weight of 12 ku were detected by SDS-PAGE , and the concentration of purified MR was 3 mg/L .The results also revealed that the serum glucose levels of the pigs in the experimental groups were significantly increased (P<0 .05);the serum lipoprotein lipase activities were slightly elevated ,but no significant differences among control and experimental groups were observed(P>0 .05);the serum triglyceride contents of the animals in the experimental groups were signif-icantly decreased ,moreover ,the serum triglyceride level of the pigs in experimental group 2 was signifi-cantly lower than that of control and experimental group 1(P<0 .05);and the free fatty acid content of test group 2 was also significantly higher than that of control and test group 1(P<0 .05) .The results a-bove suggested that the glucose intake of porcine histocytes was inhibited and the triglycerides decomposi-tion was promoted along with increased serum lipoprotein lipase activity owing to high dose of MR admin-istration ,resistin may play an important role in the regulation of energy metabolism and homeostasis of an-imals .