Alterations of myelin‐specific proteins and sphingolipids characterize the brains of acid sphingomyelinase‐deficient mice, an animal model of Niemann–Pick disease type A

Abstract

Niemann-Pick disease (NPD) type A is a neurodegenerative disorder caused by sphingomyelin (SM) accumulation in lysosomes relying on reduced or absent acid sphingomyelinase (ASM) activity. NPD-A patients develop progressive neurodegeneration including cerebral and cerebellar atrophy, relevant Purkinje cell and myelin deficiency with death within 3 years. ASM 'knock-out' (ASMKO) mice, an animal model of NPD-A, develop a phenotype largely mimicking that of NPD-A. The mechanisms underlying myelin formation are poorly documented in ASMKO mice. In this study we determined the content of four myelin-specific proteins, myelin basic protein (MBP), 2¢,3¢-cyclic nucleotide 3¢-phosphodiesterase (CNP), myelin associated glycoprotein (MAG) and proteolipid protein (PLP), and that of myelin-enriched sphingolipids in the brains of ASMKO and wild-type mice in early stages of post-natal (pn) life. Protein and mRNA analysis revealed that in ASMKO mice beginning from 4 post-natal weeks (wk-pn), the expression levels of MAG, CNP, and MBP were below those observed in wild-type mice and the same applied to PLP at 10 wk-pn. Moreover, at 4 wk-pn the expression of SOX10, one of the transcription factors involved in oligodendrocyte development and maintenance was lower in ASMKO mice. Lipid analysis showed that SM and the gangliosides GM3 and GM2 accumulated in the brains of ASMKO mice, as opposed to galactocerebroside and galactosulfocerebroside that, in parallel with the mRNAs of UDP-galactose ceramide galactosyltransferase and galactose-3-O-sulfotransferase 1, the two transferases involved in their synthesis, decreased. Myelin lipid analysis showed a progressive sphingomyelin accumulation in ASMKO mice; noteworthy, of the two sphingomyelin species known to be resolved by TLC, only that with the lower Rf accumulated. The immunohistochemical analysis showed that the reduced expression of myelin specific proteins in ASMKO mice at 10 wk-pn was not restricted to the Purkinje layer of the cerebellar cortex but involved the cerebral cortex as well. In conclusion, reduced oligodendrocyte metabolic activity is likely to be the chief cause of myelin deficiency in ASMKO mice, thus shedding light on the molecular dysfunctions underlying neurodegeneration in NPD-A. Keywords: absent acid sphingomyelinase 'knock-out' mice, brain sphingolipids, myelin proteins, Niemann-Pick disease type A.

Secondary Alterations of Sphingolipid Metabolism in Lysosomal Storage Diseases

“…Patterns are representative of those obtained in three different experiments. (PE phosphatidylethanolamine, PC phosphatidylcholine) plateau around 2.5 months of age, when SM level in ASMKO brain was about sixfold higher than in WT brain, remaining basically unchanged up to the age of 5 months, in agreement with previously published results [111,112]. Total phospholipid content in brain was similar in ASMKO and WT mice at all ages (Table 3), indicating a significant reduction of glycerophospholipids in ASMKO brain along time.…”

“…At 15 days of post-natal age, when SM accumulation was already evident, the ratio between lower and upper SM band was identical in WT and ASMKO brain, while at 1 month of age this ratio was already 1.4 fold higher in ASMKO than in WT and at 5 months it reached 2.5-fold in ASMKO vs WT (Fig. 3a), clearly indicating that, in ASMKO brain, SM molecular species containing shorter fatty acyl chains (C18 and C16) accumulate at a higher level that SM species with longer fatty acids (C24), in agreement with preliminary observations in purified myelin from ASMKO mice [112].…”

“…Psychosine (galactosylsphingosine), one of the galactoslylsphingolipids that accumulates in the brain of Krabbe disease (human globoid cell leukodystrophy) patients due to the deficient activity of b-galactosylceramidase, accumulates in lipid rafts from brain and sciatic nerve from twitcher mice (the animal model for the infantile variant of the disease) and from human Krabbe patients, leading to an altered distribution of lipid raft proteins and to inhibition of protein kinase C [116]. Brain pathology in ASMKO mice is probably at least in part due to a plasma membrane involvement [117], possibly associated with a non-conventional lipid raft organization [111,112]. For SM, the shift toward molecular species with shorter fatty acids has been associated in some cases with myelination defects [20,21], and might be a consequence of the myelin deficiency observed in ASMKO mice [112].…”

“…As other SL, SM is present in all tissues as a mixture of molecular species differing for their fatty acid composition [113]. HPTLC analysis of alkalinetreated organic phase lipids under the experimental conditions used allowed to separate 2 bands for SM: the lower migrating band corresponds to SM molecular species containing C18 (mainly stearic acid) and C16 fatty acids, while the upper migrating band corresponds to SM molecular species with longer chain fatty acids, mainly C24 [112]. In extranervous tissues, both SM bands accumulated in ASMKO mice at a similar extent (Table 3; Fig.…”

Types A and B Niemann–Pick Disease

“…This has been particularly well documented in the brains of ASMKO mice (see below), where sphingomyelin storage in the neuronal membranes leads to abnormal synapse formation and function and other abnormalities [41][42][43]. It is unclear whether the activity of ASM at the plasma membrane occurs in acidified "micro" compartments, or proceeds at a non-acidic pH.…”