β-防御素（sBD-2）在绵羊胎儿肺、肠、生殖道内的表达变化与分布研究
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摘要
作为在动物体先天性免疫中起着重要作用的内源性抗菌肽之一的β-防御素是防御素家族的重要成员之一,它在先天性免疫及获得性免疫中均起着重要的作用,而且随着研究的深入越来越多的关于β-防御素的其他作用正在被发现。但是关于绵羊胎儿发育过程中β-防御素-2(sBD-2) mRNA表达与分布的研究报道很少,对此作者进行了研究,其结果如下:
1.克隆了绵羊胎儿十二指肠组织β-防御素(sBD-2)基因,经BLAST搜索比对发现绵羊胎儿十二指肠组织的sBD-2 cDNA与绵羊、山羊、牛、猪和骆驼的β-防御素之间有较高的同源性(大于70%),与人、猴和鼠的β-防御素除个别的有50%左右的同源性外,其余大部分的同源性在20%以下。
2.采用原位杂交技术研究了sBD-2mRNA在绵羊5月龄胎儿肺、十二指肠、子宫组织中的分布与定位规律。结果表明:在肺泡上皮细胞胞质和子宫内膜上皮细胞胞质内均有杂交信号,而子宫内膜固有层无杂交信号;在肠的粘膜上皮细胞胞质内有强的杂交信号,而其固有层则无杂交信号。证明这些组织中都有sBD-2 mRNA的分布,且存在于上皮细胞中。
3.应用实时荧光定量PCR技术研究了绵羊胎儿从2月龄到5月龄过程中sBD-2 mRNA在肺、肠、生殖道的表达变化规律。结果表明绵羊胎儿从2月龄到5月龄,sBD-2基因在肺、肠和生殖道中的表达均呈现逐渐增加的趋势,说明sBD-2的表达与胎儿的发育阶段有密切的关系。且sBD-2 mRNA在肠中的表达较高,在肺及生殖道中的表达相对较低,说明sBD-2基因的表达存在组织差异性。
β-defense is one of the endogenous antimicrobial peptides. As an important member of the defensins family,β-defense played an important part in both of innate and acquired immunity, and the other effects have being found with more and more in-depth study on theβ-defense. But the expression and distribution about sheepβ-defensin-2 (sBD-2) mRNA on the development of the sheep’s fetal were reported rarely, so we studied on it, the results go as bellow:
1.By cloning sBD-2 gene of duodenum of sheep’s fetus and comparing the sequences, we found there was a higher degree of homology (above70%) between the sBD-2 cDNA of duodenal of sheep’s fetal with the sheep, goats, cattle, pigs and camels’β-defense, and about 50% homology with the human, monkey, mouse’β-defense, and bellow 20% homology with the other species’β-defense.
2.Studied on the distribution and location of the sBD-2 in lung, duodenum and uterus of sheep’s fetal of 5-month-old by in situ hybridization. The results showed that there were hybridization signal in the cytoplasm of pulmonary alveoli’s epithelial cells and endometrial epithelial cells, but no hybridization signal in the proper layer. And there was a strong hybridization signal in the cytoplasm of intestinal epithelial cell, but no hybridization signal in the proper layer. It was pointed out that sBD-2 mRNA distributed in above tissues, and existed in the epithelial cells.
3.Studied on the expression changes of sBD-2 mRNA in the lung, intestine, genital duct of sheep’s fetus from the 2-month-old to 5-month-old by real-time PCR. The results showed that the expression of sBD-2 mRNA in lung, intestine and genital duct of sheep’s fetus from the 2-month-old to 5-month-old increased gradually. It indicated that the expression of sBD-2 was closely related to the fetal stage of development. The expression of sBD-2 mRNA was the highest in intestine, while it was relatively lower in lung and genital duct. This result indicated that the expression of sBD-2 gene existed different distribution in distinct tissues.
1.克隆了绵羊胎儿十二指肠组织β-防御素(sBD-2)基因,经BLAST搜索比对发现绵羊胎儿十二指肠组织的sBD-2 cDNA与绵羊、山羊、牛、猪和骆驼的β-防御素之间有较高的同源性(大于70%),与人、猴和鼠的β-防御素除个别的有50%左右的同源性外,其余大部分的同源性在20%以下。
2.采用原位杂交技术研究了sBD-2mRNA在绵羊5月龄胎儿肺、十二指肠、子宫组织中的分布与定位规律。结果表明:在肺泡上皮细胞胞质和子宫内膜上皮细胞胞质内均有杂交信号,而子宫内膜固有层无杂交信号;在肠的粘膜上皮细胞胞质内有强的杂交信号,而其固有层则无杂交信号。证明这些组织中都有sBD-2 mRNA的分布,且存在于上皮细胞中。
3.应用实时荧光定量PCR技术研究了绵羊胎儿从2月龄到5月龄过程中sBD-2 mRNA在肺、肠、生殖道的表达变化规律。结果表明绵羊胎儿从2月龄到5月龄,sBD-2基因在肺、肠和生殖道中的表达均呈现逐渐增加的趋势,说明sBD-2的表达与胎儿的发育阶段有密切的关系。且sBD-2 mRNA在肠中的表达较高,在肺及生殖道中的表达相对较低,说明sBD-2基因的表达存在组织差异性。
β-defense is one of the endogenous antimicrobial peptides. As an important member of the defensins family,β-defense played an important part in both of innate and acquired immunity, and the other effects have being found with more and more in-depth study on theβ-defense. But the expression and distribution about sheepβ-defensin-2 (sBD-2) mRNA on the development of the sheep’s fetal were reported rarely, so we studied on it, the results go as bellow:
1.By cloning sBD-2 gene of duodenum of sheep’s fetus and comparing the sequences, we found there was a higher degree of homology (above70%) between the sBD-2 cDNA of duodenal of sheep’s fetal with the sheep, goats, cattle, pigs and camels’β-defense, and about 50% homology with the human, monkey, mouse’β-defense, and bellow 20% homology with the other species’β-defense.
2.Studied on the distribution and location of the sBD-2 in lung, duodenum and uterus of sheep’s fetal of 5-month-old by in situ hybridization. The results showed that there were hybridization signal in the cytoplasm of pulmonary alveoli’s epithelial cells and endometrial epithelial cells, but no hybridization signal in the proper layer. And there was a strong hybridization signal in the cytoplasm of intestinal epithelial cell, but no hybridization signal in the proper layer. It was pointed out that sBD-2 mRNA distributed in above tissues, and existed in the epithelial cells.
3.Studied on the expression changes of sBD-2 mRNA in the lung, intestine, genital duct of sheep’s fetus from the 2-month-old to 5-month-old by real-time PCR. The results showed that the expression of sBD-2 mRNA in lung, intestine and genital duct of sheep’s fetus from the 2-month-old to 5-month-old increased gradually. It indicated that the expression of sBD-2 was closely related to the fetal stage of development. The expression of sBD-2 mRNA was the highest in intestine, while it was relatively lower in lung and genital duct. This result indicated that the expression of sBD-2 gene existed different distribution in distinct tissues.
引文
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