Hyperhomocysteinemia and dyslipidemia in point mutation G307S of cystathionine β-synthase-deficient rabbit generated using CRISPR/Cas9

Abstract

Background Congenital hyper-homocysteinemia (HHcy) is caused by a defective cystathionine β-synthase (CBS) gene, and is frequently associated with dyslipdemia. The aim of this study was to further elucidate the effect of mutated CBS gene on circulating lipids using a rabbit model harboring a homozygous G307S point mutation in CBS. Methods CRISPR/Cas9 system was used to edit the CBS gene in rabbit embryos. The founder rabbits were sequenced, and their plasma homocysteine (Hcy) and lipid profile were analyzed. Results Six CBS-knockout (CBS-KO) founder lines with biallelic modifications were obtained. Mutation in CBS caused significant growth retardation and high mortality rates within 6 weeks after birth. In addition, the 6-week old CBS-KO rabbits showed higher plasma levels of Hcy, triglycerides (TG), total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) compared to the age-matched wild-type (WT) controls. Histological analysis of the mutants showed accumulation of micro-vesicular cytoplasmic lipid droplets in the hepatocytes. However, gastric infusion of vitamin B and betaine complex significantly decreased the plasma levels of TG, TC and LDL-C in the CBS-KO rabbits, and alleviated hepatic steatosis compared to the untreated animals. Conclusion A CBSG307S rabbit model was generated that exhibited severe dyslipidemia when fed on a normal diet, indicating that G307S mutation in the CBS gene is a causative factor for dyslipidemia.

Hydrogen sulphide in liver glucose/lipid metabolism and non‐alcoholic fatty liver disease

“… 70 Recent evidence supported these findings by showing that CBS KO rabbits had higher plasma levels of TG, total cholesterol and low‐density lipoprotein as well as hepatic microvesicular steatosis. 72 CBS KO mice also showed significant alteration of a broad range of hepatic phospholipids and phosphatidylcholines. 73 Similarly, inhibition of hepatic CSE expression in a foetal hepatocyte line (L02) upregulates the sterol regulatory element‐binding protein 1 (SREBP‐1c) pathway, c‐Jun N‐terminal kinase (JNK) phosphorylation and hepatic oxidative stress.…”

Mechanism of homocysteine-mediated endothelial injury and its consequences for atherosclerosis

“…Numerous studies have investigated the changes in plasma concentration of Hcy and related disease progression in experimental animals after CBS knockout. The researchers used CRISPR/Cas9 to knock out the CBS gene in rabbits and found that the plasma Hcy level in knockout rabbits (50.73 μmol/L) was almost twice as high as that in controls (27.93 μmol/L) ( 24 ). And severe HHcy was observed in CBS –/– (289 ± 58 μM) but not in CBS ± or control mice (<10 μM) after knocking out CBS in mice ( 25 ).…”

“…A clinical study involving 7,898 subjects showed that plasma Hcy was negatively associated with highdensity lipoprotein cholesterol (HDL-C) and apolipoprotein A1 (ApoA1) and positively associated with triglyceride (TG) (118). Moreover, a study (24) has found that the blood lipid levels of HHcy rabbits are significantly higher than that of the controls, the TG level showed almost 52-fold increase (3746.7/71.31 mg/dL). The total cholesterol (TC) (540.8 mg/dL) and low-density lipoprotein cholesterol (LDL-C) (72.02 mg/dL) in HHcy rabbits were almost 4.4-and 2.5-fold higher that in WT controls, respectively.…”

Catalpol inhibits HHcy-induced EndMT in endothelial cells by modulating ROS/NF-κB signaling

“…In recent years, studies on HHcy-induced AS lesion formation have continued and the same conclusions have been reached [5,6]. The following hypotheses have been suggested for the link between HHcy and endothelial dysfunction and AS: interference of HHcy with nitric oxide production [7,8]; deregulation of the hydrogen sul de signaling pathway [9,10]; oxidative stress, in ammation, and impaired lipoprotein metabolism [11,12]; protein N-homocysteinylation [13,14]; and cellular hypomethylation [15,16], among others [17].…”