Analysis of differential neonatal lethality in cystathionine β‐synthase deficient mouse models using metabolic profiling

Abstract

Cystathionine beta‐synthase (CBS) is a key enzyme of the trans‐sulfuration pathway that converts homocysteine to cystathionine. Loss of CBS activity due to mutation results in CBS deficiency, an inborn error of metabolism characterized by extreme elevation of plasma total homocysteine (tHcy). C57BL6 mice containing either a homozygous null mutation in the cystathionine β‐synthase (Cbs−/−) gene or an inactive human CBS protein (Tg‐G307S Cbs−/−) are born in mendelian numbers, but the vast majority die between 18 and 21 days of age due to liver failure. However, adult Cbs null mice that express a hypomorphic allele of human CBS as a transgene (Tg‐I278T Cbs−/−) show almost no neonatal lethality despite having serum tHcy levels similar to mice with no CBS activity. Here, we characterize liver and serum metabolites in neonatal Cbs+/−, Tg‐G307S Cbs−/−, and Tg‐I278T Cbs−/− mice at 6, 10, and 17 days of age to understand this difference. In serum, we observe similar elevations in tHcy in both Tg‐G307S Cbs−/− and Tg‐I278T Cbs−/− compared to control animals, but methionine is much more severely elevated in Tg‐G307S Cbs−/− mice. Large scale metabolomic analysis of liver tissue confirms that both methionine and methionine‐sulfoxide are significantly more elevated in Tg‐G307S Cbs−/− animals, along with significant differences in several other metabolites including hexoses, amino acids, other amines, lipids, and carboxylic acids. Our data are consistent with a model that the neonatal lethality observed in CBS‐null mice is driven by excess methionine resulting in increased stress on a variety of related pathways including the urea cycle, TCA cycle, gluconeogenesis, and phosphatidylcholine biosynthesis.

Homozygous whole body Cbs knockout in adult mice features minimal pathology during ageing despite severe homocysteinemia

“…Both the Tg-I278T and the Cre/LoxP Cbs −/− mouse models were induced at comparable ages (30-40 days and 63 days, respectively) and did not show biochemical or histological evidence of severe damage in the liver or kidney. 26,34 Though histological SAF scoring of Cbs −/− livers suggested ballooning and inflammation, this was not confirmed by biochemical analysis. Moreover, these findings challenge the HHCy 'threshold' theory, which presupposes a buffering mechanism which fails to protect if HHCy exceeds 169 µM.…”

“…Second, our model also presented impaired weight gain soon after induction. Both the Tg‐I278T and the Cre/LoxP Cbs −/− mouse models were induced at comparable ages (30–40 days and 63 days, respectively) and did not show biochemical or histological evidence of severe damage in the liver or kidney 26,34 . Though histological SAF scoring of Cbs −/− livers suggested ballooning and inflammation, this was not confirmed by biochemical analysis.…”

Biosynthesis, Quantification and Genetic Diseases of the Smallest Signaling Thiol Metabolite: Hydrogen Sulfide

“…Results from a recent study indicate that lethality in CBS knockout mice is brought about by abnormal buildup of methionine in the liver. Accumulation of Met in the liver was accompanied by downstream disturbances in central pathways of energy metabolism [199]. Strikingly, neither a methionine restricted diet nor cystathionine injections could rescue lethality in CBS-null mice [199].…”

“…Accumulation of Met in the liver was accompanied by downstream disturbances in central pathways of energy metabolism [199]. Strikingly, neither a methionine restricted diet nor cystathionine injections could rescue lethality in CBS-null mice [199]. A CBS heterozygous knockout (CBS +/− ) mouse model was developed by Jensen et al and the formation of H 2 S in CBS +/− and wild type animals was measured [200].…”