Fan Mingzhen, Miao Yuyang, Yan Yutong, Zhu Kunyuan, Zhao Xiaoe, Pan Menghao, Ma Baohua, Wei Qiang
Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Northwest A&F University, Xianyang, China.
College of Veterinary Medicine, Northwest A&F University, Xianyang, China.
Front Vet Sci. 2022 Apr 12;9:822165. doi: 10.3389/fvets.2022.822165. eCollection 2022.
During infection, the infected tissue secretes a variety of endogenous peptides to resist further invasion of pathogens. Among these endogenous peptides, the natriuretic peptides and the antimicrobial peptides attracted the most attention. C-type natriuretic peptide (CNP) and its receptor natriuretic peptide receptor B (NPR-B) were members of the natriuretic peptide system. The antimicrobial peptide S100A7 plays an important role to resist infection of bacteria in mastitis. It is reported that the expression of S100A7 is regulated by an activator protein-1 (AP-1)-responsive promoter. As a subunit of AP-1, c-Jun is a downstream target of CNP/NPR-B signaling pathway. Therefore, it is a hypothesis that the CNP/NPR-B signaling pathway induces the expression and secretion of S100A7 in mammary glands to take part in local mammary gland innate immunity. To verify this hypothesis, goat mammary gland and isolated mammary epithelial cells (MECs) were used to explore the expression of CNP/NPR-B and their physiological roles in goat mammary gland. The results showed that goat mammary gland expressed NPR-B, but not CNP. The expression and secretion of S100A7 in goat MECs were obviously induced by CNP/NPR-B signaling pathway. After treatment with CNP, the cyclic guanosine monophosphate (cGMP) level in goat MECs was significantly upregulated. Along with the upregulation of cGMP level, the phosphorylation levels of c-Jun N-terminal kinase (JNK) and its target c-Jun were also increased gradually. KT5823 is a specific inhibitor for protein kinase G (PKG). KT5823 remarkably inhibited the phosphorylation of JNK and c-Jun induced by CNP. Correspondingly, KT5823 evidently inhibited the expression and secretion of S100A7 induced by CNP. On the other hand, the expression of NPR-B and S100A7 was upregulated in the mastitis goat mammary gland. But, there was no significant difference in expression of CNP between healthy and mastitis goat mammary gland tissues. The goat mastitis model was established using goat MECs treated by lipopolysaccharide (LPS). LPS treatment also could increase the expression of NPR-B and S100A7. In conclusion, goat mammary gland expressed NPR-B, indicating mammary gland was the target organ for natriuretic peptide system. Moreover, CNP, through NPR-B/JNK/c-Jun signaling pathway to regulate the expression and secretion of S100A7 in MECs, played an important role in mammary gland innate immunity.
在感染过程中，受感染组织会分泌多种内源性肽以抵抗病原体的进一步入侵。在这些内源性肽中，利钠肽和抗菌肽最受关注。C型利钠肽（CNP）及其受体利钠肽受体B（NPR-B）是利钠肽系统的成员。抗菌肽S100A7在抵抗乳腺炎细菌感染中起重要作用。据报道，S100A7的表达受激活蛋白-1（AP-1）反应性启动子调控。作为AP-1的一个亚基，c-Jun是CNP/NPR-B信号通路的下游靶点。因此，有一个假说认为，CNP/NPR-B信号通路诱导乳腺中S100A7的表达和分泌，以参与局部乳腺先天免疫。为验证这一假说，使用山羊乳腺和分离的乳腺上皮细胞（MECs）来探究CNP/NPR-B在山羊乳腺中的表达及其生理作用。结果表明，山羊乳腺表达NPR-B，但不表达CNP。CNP/NPR-B信号通路明显诱导山羊MECs中S100A7的表达和分泌。用CNP处理后，山羊MECs中的环磷酸鸟苷（cGMP）水平显著上调。随着cGMP水平的上调，c-Jun氨基末端激酶（JNK）及其靶点c-Jun的磷酸化水平也逐渐升高。KT5823是蛋白激酶G（PKG）的特异性抑制剂。KT5823显著抑制CNP诱导的JNK和c-Jun的磷酸化。相应地，KT5823明显抑制CNP诱导的S100A7的表达和分泌。另一方面，乳腺炎山羊乳腺中NPR-B和S100A7的表达上调。但是，健康山羊和乳腺炎山羊乳腺组织中CNP的表达没有显著差异。使用经脂多糖（LPS）处理的山羊MECs建立山羊乳腺炎模型。LPS处理也可增加NPR-B和S100A7的表达。总之，山羊乳腺表达NPR-B，表明乳腺是利钠肽系统的靶器官。此外，CNP通过NPR-B/JNK/c-Jun信号通路调节MECs中S100A7的表达和分泌，在乳腺先天免疫中起重要作用。
