味觉受体T1R1和T1R3在从江香猪附睾不同区段的差异表达

Differential expression of taste receptors T1R1 and T1R3 in different regions of epididymis in Congjiang Xiang pigs

  • 摘要: 【目的】明确从江香猪附睾中不同区段味觉受体T1R1和T1R3的定位及表达情况,为揭示猪附睾不同区段形成特殊微环境保障精子成熟的作用机制提供参考依据。【方法】采集初情期(30d)和性成熟期(180d)从江香猪附睾组织样品,分别采用实时荧光定量PCR、免疫组织化学、蛋白免疫印迹(Western blotting)等检测从江香猪附睾不同区段味觉受体T1R1和T1R3的定位及表达情况。【结果】味觉受体T1R1/T1R3的编码基因TAS1R1/TAS1R3均表现为附睾Ⅳ区的相对表达量显著高于其他区段(P<0.05,下同),而Ⅰ区的相对表达量显著低于其他区段,具体排序为Ⅳ区>Ⅴ区>Ⅱ区> Ⅲ区>Ⅰ区。免疫组织化学定位分析结果显示,在从江香猪附睾不同区段均检测到T1R1/T1R3不同程度的免疫阳性信号。其中,T1R1在附睾Ⅰ区主要定位于肌样细胞层与基细胞核;在附睾Ⅱ区主要定位于附睾上皮细胞、肌样细胞和精子细胞;在附睾 Ⅲ 区强烈定位于附睾管腔和微绒毛根部;附睾 Ⅳ区为5个区段中表达最强烈的部位,定位于附睾管腔精子、管腔内缘和狭窄细胞膜上;在附睾Ⅴ区主要定位于附睾间质组织和狭窄细胞膜上。T1R3定位于附睾Ⅰ区血管内皮细胞、脱落生精细胞和附睾上皮细胞(尤其是基细胞);在附睾II区的精子、晕细胞和狭窄细胞上有较强表达;在附睾 Ⅲ区和 Ⅳ区主要定位于空泡化的附睾上皮细胞;在附睾Ⅴ区定位于附睾管不规则处和空泡化附睾上皮细胞,其表达强度仅次于 Ⅲ区。Western blotting检测结果显示,从江香猪附睾Ⅳ区的T1R1/T1R3表达水平最高,其次是Ⅱ区,二者显著高于其他区段。【结论】初情期从江香猪附睾T1R1和T1R3的表达具有区段特异性和细胞特异性,以Ⅳ区的表达水平最高,且主要定位于附睾上皮细胞顶端、狭窄细胞和微绒毛根部,提示T1R1/T1R3在建立促进猪附睾精子成熟和储存的特殊腔内微环境中发挥重要作用。

     

    Abstract: 【Objective】To clarify the location and expression of taste receptors T1R1 and T1R3 in different regions of the epididymis of Congjiang Xiang pigs, and to provide reference for revealing the mechanism of the formation of unique microenvironment in different segment of the epididymis to ensure sperm maturation. 【Method】The epididymis tissue samples at the puberty(30 d) and sexual maturity(180 d) were collected, and real-time fluorescence quantitative PCR, immunohistochemistry, and Western blotting were used to detect the location and expression of taste receptors T1R1 and T1R3 in different regions of epididymis of Congjiang Xiang pigs. 【Result】The expression level of TAS1R1/TAS1R3 coding gene of taste receptor T1R1/T1R3 was significantly higher in epididymis IV region than in other regions(P<0.05, the same below), while the relative expression level of epididymis I region was significantly lower than that of other regions, and the rank was IV>V>II>III>I region. Immunohistochemical localization analysis showed that T1R1/T1R3 immunopositive signals at different extents were detected in different parts of the epididymis of Congjiang Xiang pigs. Among them, T1R1 was mainly located in myoid cell layer and basal nucleus in epididymis region I. In epididymis II region, it mainly located in epididymal epithelial cells, myoid cells and sperm cells. In epididymis III region, it was strongly localized in epididymis lumen and microvilli root. Epididymis IV region was the most strongly expressed part among the five regions, which was located in the epididymis lumen sperm, lumen inner margin and narrow cell membrane. The V region of the epididymis was mainly located on the epididymis interstitial tissue and narrow cell membrane of the epididymis. T1R3was localized in vascular endothelial cells, exfoliated spermatogenic cells and epididymal epithelial cells(especially basal cells) in epididymal region I. It was strongly expressed on sperm, halo cells and narrow cells in epididymis II region. In areas III and IV of the epididymis, it was mainly located in the vacuolated epididymal epithelial cells. The expression intensity of epididymal epithelial cells located in irregularity and vacuolated epididymal duct in epididymal region V was second only to that in region III. Western blotting test results showed that the expression level of T1R1/T1R3 in epididymis region IV of Congjiang Xiang pigs was the highest, followed by that in epididymis region II, which was significantly higher than that other regions. 【Conclusion】The expression of T1R1 and T1R3 from the epididymis of Congjiang Xiang pigs during at puberty is region-specific and cell-specific, with the highest expression level in region IV, which is mainly located in the apex of epididymal epithelial cells, narrow cells and microvilli roots, suggesting that T1R1/T1R3 plays an important role in establishing a unique intracellular microenvironment that promotes maturation and storage of porine epididymal sperm.

     

/

返回文章
返回