本研究应用双吲哚基马来酰亚胺I(BSM),蛋白激酶C(PKC)酶活性的药理学竞争性抑制剂,以1.25pmol虾-1持续60分钟,以研究PKC在凡纳滨对虾血细胞信号转导通路中的潜在参与。鉴定并鉴定了南美白对虾中的一种新型PKC(LvnPKC),并确定其参与介导神经内分泌-免疫调节网络。接受BSM的南美白对虾乳杆菌的血细胞表现出显著降低的PKC活性和LvnPKC基因和蛋白质表达水平。此外,总血细胞计数,透明细胞,半颗粒细胞显着增加,颗粒细胞显着减少,同时,显著增加的酚氧化酶活性,呼吸爆发,超氧化物歧化酶(SOD)活性,吞噬活性,观察到中性粒细胞胞外诱捕网;然而,吞噬活性显著下降。在分子模型中,脂多糖和β-1,3-葡聚糖结合蛋白的基因表达,过氧化物酶素,胞浆锰SOD,线粒体锰SOD,接受BSM的凡纳滨对虾血细胞中的铜/锌SOD明显降低,但酚氧化酶原I显著增加。在儿茶酚胺生物合成中,酪氨酸,多巴胺,接受BSM的凡纳滨对虾的血细胞中去甲肾上腺素显著减少,和1-二羟基苯丙氨酸增加。此外,酪氨酸羟化酶(TH)活性显著增加,而TH和二羟苯丙氨酸脱羧酶基因表达显著下降。这些发现表明BSM抑制血细胞中的PKC活性,其中LvnPKC基因和蛋白质表达也被抑制。此外,血细胞的免疫能力,包括它们的酚氧化酶原和抗氧化系统,吞噬活性,和儿茶酚胺生物合成,被打乱了,证实LvnPKC在介导血细胞神经内分泌-免疫调节网络中的作用。
This
study applied bisindolylmaleimide I (BSM), a pharmacological competitive inhibitor of protein kinase C (PKC) enzymatic activity, at 1.25 pmol shrimp-1 for 60 min to investigate the potential involvement of PKC in signal transduction pathways in the hemocytes of Litopenaeus vannamei. A novel PKC in L. vannamei (LvnPKC) was identified and characterized and was determined to be involved in mediating the neuroendocrine-immune regulatory network. The hemocytes of L. vannamei that receive BSM exhibit significantly decreased PKC activity and LvnPKC gene and protein expression levels. Furthermore, the total hemocyte count, hyaline cells, and semigranular cells increased significantly along with significant decreases in granular cells, and meanwhile, the significantly increased phenoloxidase activity, respiratory bursts, superoxide dismutase (SOD) activity, phagocytic activity, and neutrophil extracellular trap were observed; however, phagocytic activity decreased significantly. In a molecular model, the gene expressions of lipopolysaccharide- and β-1,3-glucan-binding protein, peroxinectin, cytosolic manganese SOD, mitochondrial manganese SOD, and copper/zinc SOD in the hemocytes of L. vannamei that had received BSM decreased significantly, but prophenoloxidase I increased significantly. In catecholamine biosynthesis, tyrosine, dopamine, and norepinephrine decreased significantly in the hemocytes of L. vannamei that had received BSM, and l-dihydroxyphenylalanine increased. Moreover, tyrosine hydroxylase (TH) activity increased significantly, whereas TH and dihydroxyphenylalanine decarboxylase gene expression decreased significantly. These findings suggest that BSM inhibits PKC activity in hemocytes in which LvnPKC gene and protein expression are also inhibited. Additionally, the hemocytes\' immunocompetence, including their prophenoloxidase and antioxidant systems, phagocytic activity, and catecholamine biosynthesis, was disrupted, confirming the roles of LvnPKC in mediating the neuroendocrine-immune regulatory network in hemocytes.