Purification of antimicrobial factors from human cervical mucus.
Purification of antimicrobial factors from human cervical mucus.
复制标题
DOI：
10.1093/humrep/dem128
复制
发表时间：
2007-07
影响因子：
6.1
通讯作者：
Li Ming;Xiaoling Pan;Li Yan;W. Lili;Wu Qi;Xiyong Yu;W. Boyao;H. Ning
中科院分区：
文献类型：
--
作者：
Li Ming;Xiaoling Pan;Li Yan;W. Lili;Wu Qi;Xiyong Yu;W. Boyao;H. Ning
本研究的目的是从健康女性宫颈粘液中分离杀菌蛋白。方法收集宫颈粘液，用1%醋酸溶解。通过凝胶覆盖试验检测酸溶性提取物对大肠杆菌ATCC 43827的抗菌活性。提取物具有明显的抑菌活性，条带清晰可见。用反相高效液相色谱法对抗菌带进行纯化，用放射扩散法测定其抗菌活性。结果分离纯化得到两种抗菌蛋白，并通过Tricine SDS-聚丙烯酰胺凝胶电泳、N-末端测序和基质辅助激光解吸电离飞行时间质谱对其进行了鉴定。经鉴定，这两种蛋白分别为高迁移率族核小体结合结构域2（HMG N2）和分泌型白细胞肽酶抑制剂（SLPI）。SLPI是宫颈粘液中已知的抗菌肽，而宫颈粘液中的HMG N2先前尚未报道。RT-PCR检测宫颈上皮细胞和宫颈癌细胞中HMG N2 mRNA的表达。狭缝杂交显示大量的HMG N2蛋白在宫颈粘液。结论HMG N2和SLPI在健康女性宫颈粘液中的表达可能与其对生殖系统潜在病原体的免疫监视和防御有关。
BACKGROUND The aim of this study was to separate bactericidal proteins from healthy female cervical mucus. METHODS Cervical mucus was collected and dissolved in 1% acetic acid. The antimicrobial activity of acid-soluble extracts was detected by gel overlay assay against Escherichia coli ATCC 43827. The extracts showed considerable amount of antibacterial activity with a clearly visible band. The bactericidal band was purified by reverse-phase high performance liquid chromatography and the antibacterial activity of the eluate was examined using radial diffusion assay. RESULTS Two antimicrobial proteins were purified and were further characterized by Tricine sodium dodecyl sulphate-polyacrylamide gel electrophoresis, N-terminal sequencing and matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The proteins were identified as high-mobility group nucleosomal-binding domain 2 (HMG N2) and secretory leukocyte peptidase inhibitor (SLPI). SLPI is an antimicrobial peptide already known in the cervical mucus while HMG N2 in the cervical mucus had not been previously reported. The expression of HMG N2 mRNA was detected in Hela cells and cervical epithelial cells by RT-PCR. Slit hybridization showed abundant amounts of the HMG N2 protein in the cervical mucus. CONCLUSIONS These results suggest that the expression of HMG N2 and SLPI in the healthy female cervical mucus may be relevant to their immune surveillance and defense against potential pathogens in human reproductive system.