过表达HMOX1基因对无量山乌骨鸡原代前脂肪细胞增殖的影响

doi:10.16431/j.cnki.1671-7236.2026.01.032

Abstract

Abstract:

Objective This experiment aimed to construct an overexpression vector of heme oxygenase 1 （HMOX1） gene in Wuliangshan Black-boned chickens and transfect it into chicken primary preadipocytes， and investigate the effect of HMOX1 gene overexpression on the proliferation of chicken primary preadipocyte. Method Collect the subcutaneous fat tissue from 1-day-old chicks of Wuliangshan Black-boned chicken， extract total RNA and reverse transcribe it into cDNA， and amplify the CDS region of HMOX1 gene. The overexpression vector pcDNA3.1-HMOX1-mRFP was constructed using the plasmid pcDNA3.1-mRFP， followed by restriction enzyme digestion and sequencing for identification. The pcDNA3.1-HMOX1 overexpression vector was transfected into chicken primary preadipocytes by using transfection reagents， and the overexpression efficiency was detected. Cell viability was measured using CCK-8 method， and the cell cycle was analyzed by flow cytometry. Real-time quantitative PCR and Western blotting were used to detect the mRNA and protein expression levels of proliferation-related genes （P21， CDK1 and PCNA）. Result Restriction enzyme digestion and sequencing indicated that the pcDNA3.1-HMOX1-mRFP overexpression vector was successfully constructed. Real-time quantitative PCR and Western blotting test results showed that， compared with control group， the mRNA and protein expression of HMOX1 gene in cells transfected with pcDNA3.1-HMOX1-mRFP were extremely significantly upregulated （P<0.01）， demonstrated that HMOX1 was successfully overexpressed in chicken primary preadipocytes. CCK-8 assay results showed that compared with control group， the cell viability of HMOX1 overexpression group significantly increased at 12 hours after transfection （P<0.05）， and decreased extremely significantly at 36 and 48 hours after transfection （P<0.01）. The results of flow cytometry test showed that，compared with control group， the number of cells in the S phase in HMOX1 overexpression group increased extremely significantly at 12 hours （P<0.01）， while the number of cells in the G1 phase decreased extremely significantly （P<0.01）. The number of cells in the S phase at 48 hours was significantly reduced （P<0.05）. Detection results of proliferation-related genes showed that there was no significant difference in the mRNA and protein expression of P21 and CDK1 genes in the cells between the two groups （P>0.05）. The mRNA and protein expression of PCNA gene in overexpression group were significantly lower than those in control group （P<0.05）. Conclusion The HMOX1 gene overexpression vector was successfully constructed. After transfection of the original preadipocytes of Wuliangshan Black-boned chicken， cell proliferation was inhibited. The experimental results could provide theoretical guidance for further study of the effect of HMXO1 on the differentiation of primary preadipocytes of Wuliangshan Black-boned chicken.

Key words: Wuliangshan Black-boned chicken; HMOX1 gene; preadipocyte; proliferation

CLC Number:

S831.2

YANG Pingyuan, OU Xiaoman, OU Zhengmiao, CHEN Fenfen. Effect of HMOX1 Gene Overexpression on the Proliferation of Primary Preadipocyte in Wuliangshan Black-boned Chicken[J]. China Animal Husbandry & Veterinary Medicine, 2026, 53(1): 359-367.

Figures/Tables 6

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References 27

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基因 Genes	引物序列 Primer sequences （5′→3′）	产物长度 Product length/bp	GenBank登录号 GenBank accession No.
HMOX1	F：GAGCCAGGAGAACGGTCAT R：CCTGCTTGTCCTCTCACTCT	115	NC_052532.
β-actin	F：GAGAAATTGTGCGTGACATCA R：CCTGAACCTCTCATTGCCA	152	NC_006101.5
CDK1	F：TGCCATTCAAGACGAGTTCTG R：AGGAGTGGAATAA	198	NM_205314.1
P21	F：GAAGAGTTGTCCACGATAAGC R：TTCCAGTCCTCCTCA	247	AF513031.1
PCNA	F：ATGCGGATACGTTGGCTCTA R：CAATGTGGCTGAG	180	NM_204170.2

Effect of HMOX1 Gene Overexpression on the Proliferation of Primary Preadipocyte in Wuliangshan Black-boned Chicken

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