College of Ocean and Earth Sciences, Xiamen University, Xiamen, 361102, China; Key Laboratory of Marine Biogenetic Resources, Third Institute of Oceanography, State Oceanic Administration, Xiamen, 361005, China; Key Laboratory of Marine Biotechnology of Fujian Province, Institute of Oceanology, Fujian Agriculture and Forestry University, Fuzhou, 350002, China.
Key Laboratory of Marine Biotechnology of Fujian Province, Institute of Oceanology, Fujian Agriculture and Forestry University, Fuzhou, 350002, China.
Fish Shellfish Immunol. 2019 Jul;90:20-29. doi: 10.1016/j.fsi.2019.04.048. Epub 2019 Apr 19.
Granulocyte colony-stimulating factor (GCSF) is a pleiotropic cytokine that plays a key role in regulation of hematopoiesis, innate and adaptive immune responses in mammals. However, bioactivity of GCSF in teleost fish remains largely unknown. In this study, a GCSFb homologue from large yellow croaker (Larimichthys crocea) (LcGCSFb) was cloned by RACE-PCR techniques. The open reading frame (ORF) of LcGCSFb is 603 bp long and encoded a protein precursor of 200 amino acids (aa), with a 19-aa signal peptide and a 181-aa mature peptide. In healthy fish, the LcGCSFb was constitutively expressed in all examined tissues, with the highest levels in mucous tissues, such as gills, intestine, and stomach. Its transcripts in head kidney, spleen, gills, intestine and stomach were significantly induced by Vibrio alginolyticus challenge. LcGCSFb transcripts were also detected in primary head kidney leukocytes (PKL), primary head kidney macrophages (PKM), primary head kidney granulocytes (PKG) and head kidney cell line (LYCK), and markedly upregulated by inactivated V. alginolyticus. These data suggested that LcGCSFb may play a role in immune response against bacterial infection. In vivo administration of recombinant LcGCSFb protein (rLcGCSFb) significantly upregulated the expression levels of the inflammatory cytokines (IL-6 and TNFα), and transcription factor C/EBPβ, which is required for proliferation of neutrophils. Furthermore, rLcGCSFb showed an ability to strengthen the phagocytosis of PKL in vitro. Taken together, LcGCSFb may be involved in antibacterial immunity via promoting the inflammatory response and the phagocytic activity of leukocytes. To our knowledge, this is the first report on immunoregulatory roles of GCSF in teleost.
粒细胞集落刺激因子 (GCSF) 是一种多功能细胞因子，在哺乳动物的造血、先天和适应性免疫反应中发挥关键作用。然而，鱼类的 GCSF 生物活性在很大程度上仍然未知。在这项研究中，通过 RACE-PCR 技术克隆了大黄鱼 (Larimichthys crocea) 的 GCSFb 同源物 (LcGCSFb)。LcGCSFb 的开放阅读框 (ORF) 长 603bp，编码一个 200 个氨基酸 (aa) 的前体蛋白，带有 19 个 aa 的信号肽和 181 个 aa 的成熟肽。在健康鱼类中，LcGCSFb 在所有检测到的组织中均呈组成型表达，在粘液组织（如鳃、肠和胃）中的表达水平最高。在头肾、脾脏、鳃、肠和胃中，其转录物在受到弧菌刺激后显著上调。LcGCSFb 的转录物也在原头肾白细胞 (PKL)、原头肾巨噬细胞 (PKM)、原头肾粒细胞 (PKG) 和头肾细胞系 (LYCK) 中检测到，并被失活的弧菌显著上调。这些数据表明 LcGCSFb 可能在针对细菌感染的免疫反应中发挥作用。体内给予重组 LcGCSFb 蛋白 (rLcGCSFb) 可显著上调炎症细胞因子 (IL-6 和 TNFα) 和转录因子 C/EBPβ 的表达水平，后者是中性粒细胞增殖所必需的。此外，rLcGCSFb 在体外显示出增强 PKL 吞噬作用的能力。总之，LcGCSFb 可能通过促进白细胞的炎症反应和吞噬活性参与抗菌免疫。据我们所知，这是鱼类 GCSF 在免疫调节中的作用的首次报道。
