Chronic Disease Program, Ottawa Hospital Research Institute, Ottawa, Ontario, Canada.
Department of Chemistry and Biomolecular Sciences, Faculty of Science, University of Ottawa, Ottawa, Ontario, Canada.
Hum Reprod. 2018 Dec 1;33(12):2175-2183. doi: 10.1093/humrep/dey315.
Do the truncated LL-37 peptides, GI-20 and GF-17, have spermicidal activity and microbicidal effects on the sexually transmitted infection (STI) pathogen Neisseria gonorrhoeae with equivalent potency to LL-37?
GI-20 and GF-17 exhibited spermicidal effects on both mouse and human sperm as well as microbicidal action on N. gonorrhoeae with the same efficacy as LL-37.
The antimicrobial peptide LL-37 exerts microbicidal activity against various STI pathogens as well as spermicidal effects on both mouse and human sperm.
STUDY DESIGN, SIZE, DURATION: Spermicidal activities of GI-20 and GF-17 were evaluated in vitro in mouse and human sperm and in vivo in mice. Finally, in vitro antimicrobial effects of LL-37, GI-20 and GF-17 on an STI pathogen, N. gonorrhoeae were determined. All experiments were repeated three times or more. In particular, sperm samples from different males were used on each experimental day.
PARTICIPANTS/MATERIALS, SETTING, METHODS: The plasma membrane integrity of peptide-treated sperm was assessed by cellular exclusion of Sytox Green, a membrane impermeable fluorescent DNA dye. Successful mouse in vitro fertilization was revealed by the presence of two pronuclei in oocytes following co-incubation with capacitated untreated/peptide-pretreated sperm. Sperm plus each peptide were transcervically injected into female mice and the success of in vivo fertilization was scored by the formation of 2-4 cell embryos 42 h afterward. Reproductive tract tissues of peptide pre-exposed females were then assessed histologically for any damage. Minimal inhibitory/bactericidal concentrations of LL-37, GI-20 and GF-17 on N. gonorrhoeae were determined by a standard method.
Like LL-37, treatment of sperm with GI-20 and GF-17 resulted in dose-dependent increases in sperm plasma membrane permeabilization, reaching the maximum at 18 and 3.6 μM for human and mouse sperm, respectively (P < 0.0001, as compared with untreated sperm). Mouse sperm treated with 3.6 μM GI-20 or GF-17 did not fertilize oocytes either in vitro or in vivo. Moreover, reproductive tract tissues of female mice pre-exposed to 3.6 μM GI-20 or GF-17 remained intact with no lesions, erosions or ulcerations. At 1.8-7.2 μM, LL-37, GI-20 and GF-17 exerted bactericidal effects on N. gonorrhoeae.
N/A.
LIMITATIONS, REASONS FOR CAUTION: Direct demonstration of the inhibitory effects of GI-20 and GF-17 on human in vitro and in vivo fertilization cannot be performed due to ethical issues.
Like LL-37, GI-20 and GF-17 acted as spermicides and microbicides against N. gonorrhoeae, without adverse effects on female reproductive tissues. With lower synthesis costs, GI-20 and GF-17 are attractive peptides for further development into vaginal spermicides/microbicides.
STUDY FUNDING/COMPETING INTEREST(S): This work was supported by Canadian Institutes of Health Research (MOP119438 and CCI82413 to N.T.) and NIH (R01 AI105147 to G.W.). There are no competing interests to declare.
截短的 LL-37 肽 GI-20 和 GF-17 是否具有杀精作用和对性传播感染（STI）病原体淋病奈瑟菌的杀菌作用，其效力与 LL-37 相当？
GI-20 和 GF-17 对人和小鼠精子均具有杀精作用，对淋病奈瑟菌也具有杀菌作用，其效果与 LL-37 相同。
抗菌肽 LL-37 对各种 STI 病原体具有杀菌作用，对人和小鼠精子具有杀精作用。
研究设计、大小和持续时间：在人和小鼠精子中评估 GI-20 和 GF-17 的杀精活性，并在小鼠体内进行评估。最后，测定 LL-37、GI-20 和 GF-17 对 STI 病原体淋病奈瑟菌的体外抗菌作用。所有实验均重复三次或更多次。特别是，在每个实验日使用不同男性的精子样本。
参与者/材料、设置、方法：通过 Sytox Green 细胞外排除评估肽处理精子的质膜完整性，Sytox Green 是一种不可渗透的膜荧光 DNA 染料。用未经处理/肽预处理的精子与获能卵母细胞共孵育后，出现两个原核，表明小鼠体外受精成功。将精子与每种肽一起经宫颈注入雌性小鼠体内，42 小时后形成 2-4 细胞胚胎，则判定体内受精成功。然后评估肽预暴露的雌性生殖道组织是否有任何损伤。通过标准方法确定 LL-37、GI-20 和 GF-17 对淋病奈瑟菌的最小抑菌/杀菌浓度。
与 LL-37 一样，用 GI-20 和 GF-17 处理精子会导致精子质膜通透性呈剂量依赖性增加，对人和小鼠精子的最大增加分别达到 18 和 3.6 μM（P < 0.0001，与未处理的精子相比）。在体外或体内，用 3.6 μM GI-20 或 GF-17 处理的小鼠精子均未使卵母细胞受精。此外，用 3.6 μM GI-20 或 GF-17 预暴露的雌性小鼠生殖道组织保持完整，没有损伤、侵蚀或溃疡。在 1.8-7.2 μM 时，LL-37、GI-20 和 GF-17 对淋病奈瑟菌具有杀菌作用。
无。
局限性、谨慎的原因：由于伦理问题，无法直接证明 GI-20 和 GF-17 对人类体内和体外受精的抑制作用。
与 LL-37 一样，GI-20 和 GF-17 对淋病奈瑟菌具有杀精和杀菌作用，对女性生殖组织没有不良影响。GI-20 和 GF-17 的合成成本较低，是进一步开发阴道杀精剂/杀菌剂的有吸引力的肽。
研究资金/竞争利益：这项工作得到了加拿大卫生研究院（MOP119438 和 CCI82413 给 N.T.）和 NIH（R01 AI105147 给 G.W.）的支持。没有竞争利益需要申报。
