Phosphorus commonly reduces lipid deposition in the vertebrates. However, the underlying mechanisms involved in the process remain unclear. Yellow catfish were given three experimental diets with dietary phosphate levels of 3.22, 6.47 and 7.99 g Pi kg− 1, respectively, for 8 weeks. The contents of triglyceride, non-esterified free fatty acids, adenosine triphosphate, nicotinamide adenine dinucleotide, nicotinamide adenine dinucleotide, enzymatic activities, mRNA and protein expression were determined in the intestinal tissues. Hematoxylin and eosin, Oil Red O staining, and transmission electron microscope were performed for intestinal tissues. Primary intestinal epithelial cells were isolated from yellow catfish intestine. Western blot analysis, Immunoprecipitation assays, Immunofluorescence staining, and RNA extraction and quantitative real-time PCR were decided. Luciferase reporter assays and electrophoretic mobility shift assay were used to evaluate the function of Sirt3, PPARα and Lcad promoters. High dietary phosphate intake activated intestinal phosphate absorption and excretion, and reduced lipid deposition through increasing lipolysis in the intestine. Moreover, phosphate incubation increased the mRNA and protein expression of krüppel like factor 4 (klf4), silent mating-type information regulation 2 homolog 3 (sirt3), peroxisome proliferator activated receptor alpha (pparα) and long chain acyl-CoA dehydrogenase (lcad) in the intestinal epithelial cells (IECs), and klf4 knockdown attenuated the phosphate-induced increase of protein levels of Sirt3, Pparα and Lcad. Further investigation found that Klf4 overexpression increased the activity of sirt3 and pparα promoters, which in turn reduced the acetylation and protein level of Lcad. Dietary Pi excess induced lipid degradation by the activation of the Klf4-Sirt3/Pparα-Lcad pathway in the intestine and primary IECs.

中文翻译：

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Klf4-Sirt3/Pparα-Lcad 通路有助于高磷酸盐诱导的脂质降解

磷通常会减少脊椎动物的脂质沉积。然而，该过程中涉及的潜在机制仍不清楚。给黄鲶鱼喂食磷酸盐水平分别为 3.22、6.47 和 7.99 g Pi kg− 1 的三种实验日粮，持续 8 周。测定肠道组织中甘油三酯、非酯化游离脂肪酸、三磷酸腺苷、烟酰胺腺嘌呤二核苷酸、烟酰胺腺嘌呤二核苷酸、酶活性、mRNA和蛋白表达量的含量。对肠组织进行苏木精和伊红、油红O染色和透射电镜。从黄鲶肠中分离出原代肠上皮细胞。蛋白质印迹分析、免疫沉淀分析、免疫荧光染色、并决定了RNA提取和定量实时PCR。荧光素酶报告分析和电泳迁移率变动分析用于评估 Sirt3、PPARα 和 Lcad 启动子的功能。高膳食磷酸盐摄入量激活肠道磷酸盐吸收和排泄，并通过增加肠道脂肪分解来减少脂质沉积。此外，磷酸盐孵育增加了 krüppel 样因子 4 (klf4)、沉默交配型信息调节 2 同系物 3 (sirt3)、过氧化物酶体增殖物激活受体 α (pparα) 和长链酰基辅酶 A 脱氢酶 (lcad) 的 mRNA 和蛋白质表达在肠上皮细胞 (IEC) 中，klf4 敲低减弱了磷酸盐诱导的 Sirt3、Pparα 和 Lcad 蛋白质水平的增加。进一步研究发现，Klf4 过表达增加了 sirt3 和 pparα 启动子的活性，进而降低了 Lcad 的乙酰化和蛋白水平。膳食 Pi 过量通过激活肠道和初级 IEC 中的 Klf4-Sirt3/Pparα-Lcad 通路诱导脂质降解。