1,25(OH)2D3及VDR敲除对山羊附睾头上皮细胞β防御素家族表达的影响

doi:10.11843/j.issn.0366-6964.2023.05.020

Abstract

Abstract: The aim of this study was to explore whether 1,25(OH)₂D₃ regulates beta-defensin gene expression in the caprine epididymal caput epithelial cells by the VDR signaling pathway. Three 6-month-old Taihang black goats were selected, and the epididymal caput tissues were collected. The caprine epididymal caput epithelial cells were isolated by differential adherence time, and the purity of epithelial cells was identified by cellular immunofluorescence; The epididymal caput epithelial cells were treated with 100 nmol·L^-1 1,25(OH)₂D₃; And the pCas9/gRNA1 plasmid vector with the highest knockout efficiency was selected for cell transfection. And negative control group and blank control group were set up, with three replicate in each group. After the cells were treated with 1,25(OH)₂D₃ or VDR gene knockout, the expressions of VDR and 17 β-defensin genes were detected by qRT-PCR, and the expression of VDR protein and 3 β-defensin proteins were detected by Western blot. The results showed that 1,25(OH)₂D₃ could significantly increase mRNA and protein expressions of VDR, gBD124, gBD126 and gBD104a(P<0.01), and markedly increased the expression of gBD104, gBD109tr1, gBD109tr2, gBD113, gBD116, gBD120, gBD121 and gBD123 genes(P<0.01), and significantly increased the expression of gBD106, gBD127, gBD129 and gBD134 genes (P<0.05), but had no significant effect on gBD110 like and gBD128 expression (P<0.05); After the three gene knockout vectors were transfected into cells, the expression of VDR protein in pCas9-VDR-V1 group was significantly decreased (P<0.01). VDR gene knockout significantly increased gBD124 mRNA and protein expression (P<0.01), and significantly reduced the mRNA and protein expression of gBD126 and gBD104a (P<0.05), the mRNA level of gBD109tr1, gBD109tr2, gBD116, gBD123, gBD127, gBD128 and gBD134 genes in VDR gene knockout group were significantly lower than those in the other groups (P<0.01), the mRNA level of gBD104, gBD106, gBD120 and gBD129 genes in VDR gene knockout group were significantly lower than those in the other groups (P<0.05), but had no significant effect on gBD121, gBD110 like and gBD113 (P<0.05). In conclusion, 1,25(OH)₂D₃ could up-regulate the expression of VDR and some beta-defensins, VDR gene knockout increased the expression of some beta-defensins. The results showed that 1,25(OH)₂D₃ could increase the expression of part beta-defensins in caprine epididymal caput epithelial cells by up-regulating the expression of VDR and activating the VD/VDR signaling pathway.

Key words: beta-defensin, 1,25(OH)₂D₃, vitamin D receptor, gene knockout, epididymal caput epithelial cells

CLC Number:

S827.3

WANG Li, GUO Yaru, ZHANG Junmei, LEI Mingkai, WANG Zhenguo, ZHANG Chunxiang, REN Youshe. Effect of 1,25(OH)₂D₃ and VDR Knockout on the Expression of β-defensins Family in the Caprine Epididymal Caput Epithelial Cells[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(5): 1990-2000.

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Effect of 1,25(OH)₂D₃ and VDR Knockout on the Expression of β-defensins Family in the Caprine Epididymal Caput Epithelial Cells

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Effect of 1,25(OH)2D3 and VDR Knockout on the Expression of β-defensins Family in the Caprine Epididymal Caput Epithelial Cells

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Effect of 1,25(OH)₂D₃ and VDR Knockout on the Expression of β-defensins Family in the Caprine Epididymal Caput Epithelial Cells