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PCR primers are designed to obtain the gene sequence of 20-311 amino acids of the rabbit APOE protein according to the total sequence of the rabbit APOE gene. The 10xHis-B2M-tag gene and the Myc tag gene are added to the N-terminal and C-terminal of the gene of interest, respectively, obtaining the target gene. The target gene is amplified by PCR. The amplified target genes are cloned into expression vectors and then transformed into E.coli cells. The transformed E.coli cells are cultured in a suitable medium containing corresponding antibiotics, and IPTG is added to induce protein expression. The expressed cells are harvested by centrifugation and then lysed to release the APOE protein. Affinity chromatography is used to purify the collected supernatant to obtain pure rabbit APOE protein. The purity of recombinant rabbit APOE is greater than 90% as measured by SDS-PAGE.
Apolipoprotein E (APOE) is a critical protein involved in lipid metabolism and cardiovascular health, particularly in the context of atherosclerosis. APOE is a glycoprotein that plays a significant role in the transport and clearance of lipoproteins, including chylomicrons and very low-density lipoproteins (VLDL) from circulation. Its primary function is to facilitate the binding of lipoproteins to hepatic receptors, thereby regulating cholesterol homeostasis and lipid metabolism [1]. In rabbits, the APOE gene exhibits a high degree of homology with the human APOE ε3 isoform, the most prevalent in humans, sharing approximately 80% sequence similarity [2].
Research has demonstrated that the absence of APOE in knockout rabbit models leads to significant alterations in lipid profiles and a predisposition to atherosclerosis. Studies involving APOE knockout rabbits have shown that these animals develop atherosclerotic lesions similar to those observed in humans, making them valuable models for studying cardiovascular diseases [3][4][5].
APOE is also implicated in neurodegenerative diseases. APOE in the brain is associated with clearing amyloid-beta, a peptide that accumulates in Alzheimer's disease. The structural similarities between rabbit and human APOE suggest that rabbit models can be effectively utilized to study the pathophysiology of Alzheimer's disease and related cognitive deficits [2]. Furthermore, alterations in APOE expression have been linked to conditions such as diabetes, where insulin deficiency can downregulate APOE mRNA expression, further complicating lipid metabolism [6].
References:
[1] F. Simon, A. Larena‐Avellaneda, & S. Wipper, Experimental atherosclerosis research on large and small animal models in vascular surgery, Journal of Vascular Research, vol. 59, no. 4, p. 221-228, 2022. https://doi.org/10.1159/000524795
[2] C. Weiss, N. Bertolino, D. Procissi, Q. Smith, K. Viola, S. Bartley, et al., Diet‐induced alzheimer's‐like syndrome in the rabbit, Alzheimer S & Dementia Translational Research & Clinical Interventions, vol. 8, no. 1, 2022. https://doi.org/10.1002/trc2.12241
[3] J. McNally, A. Havenon, S. Kim, C. Wang, S. Wang, M. Zabriskie, et al., Rabbit models of intracranial atherosclerotic disease for pathological validation of vessel wall mri, The Neuroradiology Journal, vol. 34, no. 3, p. 193-199, 2020. https://doi.org/10.1177/1971400920980153
[4] M. Zabriskie, C. Wang, S. Wang, & M. Alexander, Apolipoprotein e knockout rabbit model of intracranial atherosclerotic disease, Animal Models and Experimental Medicine, vol. 3, no. 2, p. 208-213, 2020. https://doi.org/10.1002/ame2.12125
[5] S. Wang, T. Yu, C. Hung, C. Yang, M. Lin, C. Su, et al., Inhibition of semicarbazide-sensitive amine oxidase reduces atherosclerosis in cholesterol-fed new zealand white rabbits, Scientific Reports, vol. 8, no. 1, 2018. https://doi.org/10.1038/s41598-018-27551-6
[6] C. Phillips, C. Madigan, D. Owens, P. Collins, & G. Tomkin, Defective chylomicron synthesis as a cause of delayed particle clearance in diabetes?, Journal of Diabetes Research, vol. 3, no. 3, p. 171-178, 2002. https://doi.org/10.1080/15604280214277
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KEGG: ocu:100009337
UniGene: Ocu.6224