Application of dipeptide pro-asp in promoting secretion of igf-1 from animal hepatocytes
A technology of IGF-1, pro-asp, applied in the field of animal growth and metabolism, to achieve the effect of good promotion and application prospects
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Embodiment 1
[0028] Example 1 Dipeptide Pro-Asp (rather than amino acid monomer Pro+Asp) promotes the secretion of IGF-1 from animal liver cells
[0029] 1. Experimental method
[0030] (1) HepG2 cells were cultured in H-DMEM medium containing 10% fetal bovine serum, and primary piglet hepatocytes were cultured in Hepato ZYME-SFM serum-free medium.
[0031] (2) The above-mentioned HepG2 cells and piglet primary liver cells were treated with Pro-Asp (50, 200, 500 μM) and Pro+Asp (500 μM) for 24 hours, respectively.
[0032] (3) Cells were collected, RNA and protein were extracted, and the expressions of IGF-1 mRNA and protein were detected by qPCR and western blot respectively.
[0033] The cell supernatant was collected, and the IGF-1 secretion in the cell supernatant was detected by RIA or ELSIA.
[0034] 2. The results are attached Figure 1~6 shown. The results showed that 500 μM Pro-Asp could significantly improve the HepG2 cells ( figure 1 ) and primary piglet hepatocytes ( fig...
Embodiment 2
[0035] Example 2 The effect of tail vein injection of dipeptide Pro-Asp on expression of IGF-1 and signaling pathway in mouse liver cells
[0036] 1. Experimental method
[0037] (1) Animal feeding conditions: 1 animal / box, single breeding; feeding temperature and humidity: 20-26 ℃, 40-70%; adopt 10h: 14 h day and night intermittent lighting; the conditions of the breeding room are always kept stable to ensure the test results reliability. Free access to food and water, feed and drinking water were provided by the Guangdong Provincial Medical Experimental Animal Center; Quarantine: The purchased SPF grade C57BL BKS-DB mice were pre-fed for 7 days. Animals were checked once a day during this period.
[0038] (2) Animal grouping and treatment: 4-week-old mice were randomly divided into normal control group, T1 group and T2 group, with 6 mice in each group, 3 groups in total. The mice in the normal control group were injected with normal saline into the tail vein, the mice in ...
Embodiment 3
[0042] Example 3 Pro-Asp has the effect of promoting IGF-1 secretion similar to GH
[0043] 1. Culture HepG2 cells in H-DMEM medium containing 10% fetal bovine serum, treat cells with Pro-Asp (500 μM) or / and GH (20 nM) for 24 hours, collect cells, extract cell proteins, and use western blot method The expression of IGF-1 protein and related signaling pathway proteins were carried out respectively.
[0044] 2. The results are attached Figure 8 shown. Figure 8 It shows that adding GH and Pro-Asp alone can significantly promote the phosphorylation level of JAK2 and STAT5 and the expression level of IGF-1 protein; Phosphorylation level, but can not significantly increase the expression level of IGF-1 protein, suggesting that Pro-Asp has a similar effect of GH.
[0045] In summary, the experiments showed that the dipeptide Pro-Asp (rather than the monomeric amino acid Pro+Asp) can promote the secretion of IGF-1 from animal liver cells, and in terms of promoting the secretion o...
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