Alterations in endo-lysosomal function induce similar hepatic lipid profiles in rodent models of drug-induced phospholipidosis and Sandhoff disease

Abstract

Drug-induced phospholipidosis (DIPL) is characterized by an increase in the phospholipid content of the cell and the accumulation of drugs and lipids inside the lysosomes of affected tissues, including in the liver. Although of uncertain pathological significance for patients, the condition remains a major impediment for the clinical development of new drugs. Human Sandhoff disease (SD) is caused by inherited defects of the β subunit of lysosomal β-hexosaminidases (Hex) A and B, leading to a large array of symptoms, including neurodegeneration and ultimately death by the age of 4 in its most common form. The substrates of Hex A and B, gangliosides GM2 and GA2, accumulate inside the lysosomes of the CNS and in peripheral organs. Given that both DIPL and SD are associated with lysosomes and lipid metabolism in general, we measured the hepatic lipid profiles in rodent models of these two conditions using untargeted LC/MS to examine potential commonalities. Both model systems shared a number of perturbed lipid pathways, notably those involving metabolism of cholesteryl esters, lysophosphatidylcholines, bis(monoacylglycero)phosphates, and ceramides. We report here profound alterations in lipid metabolism in the SD liver. In addition, DIPL induced a wide range of lipid changes not previously observed in the liver, highlighting similarities with those detected in the model of SD and raising concerns that these lipid changes may be associated with underlying pathology associated with lysosomal storage disorders.

Keywords: ceramides; lipidomics; lysophospholipid; lysosome; mass spectrometry; storage diseases; toxicology.

Fig. 1.

LPC concentrations in liver tissue. Metabolites were extracted from homogenized tissue, and LPCs were detected by using positive-ion mode LC/MS in control and Hexb^−/− mice (A) and in control and chloroquine-treated rats (B). Results are mean ± SEM. Significance level is quoted for Student’s t test: * P ≤ 0.05; ** P ≤ 0.01.

Fig. 2.

BMP concentrations in liver tissue. Metabolites were extracted from homogenized tissue, and BMPs were detected by using negative-ion mode LC/MS in control and Hexb^−/− mice (A) and in control and chloroquine-treated rats (B). Results are mean ± SEM. Significance level is quoted for Student’s t test: * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001.

Fig. 3.

CE concentrations in liver tissue. Mass spectrum of CE 18:2 represented by (M+NH₄)⁺ (m/z 666.6) (A). The typical MS source fragment representative of CEs is also present (m/z 369.4). Metabolites were extracted from homogenized tissue, and CEs were detected by using positive-ion mode LC/MS in control and Hexb^−/− mice (B) and in control and chloroquine-treated rats (C). Results are mean ± SEM. Significance level is quoted for Student’s t test: * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001.

Fig. 4.

Cers in liver tissue. Metabolites were extracted from homogenized tissue, and Cers were detected by using positive-ion mode LC/MS in control and Hexb^−/− mice (A) and in control and chloroquine-treated rats (B). Ratio of long-chain to VLC Cers in liver tissue from control and Hexb^−/− mice (C) and from control and chloroquine-treated rats (D). Fold change in CerS mRNA expression in the liver of rats treated with chloroquine compared with controls (set to 1 for each transcript) as measured by real-time qPCR (E). Results are mean ± SEM. Significance level is quoted for Student’s t test: * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001.