The skin of atopic dermatitis patients contains a novel enzyme, glucosylceramide sphingomyelin deacylase, which cleaves the N-acyl linkage of sphingomyelin and glucosylceramide

Abstract

We have demonstrated previously that there is an abnormal expression of sphingomyelin (SM) deacylase in the epidermis of patients with atopic dermatitis (ADe). In the present study, we have prepared N-[palmitic acid-1-(14)C]SM and N-[palmitic acid-1-(14)C]glucosylceramide (GCer) to use as substrates and have quantified SM deacylase activity by detecting the release of [(14)C]palmitic acid in extracts of the stratum corneum or the epidermis of ADe patients. In studies using [palmitic acid-1-(14)C]SM as a substrate, a pH dependency of catalytic activity with a peak at pH 5.0 was found. Preparative SDS/PAGE using an extract of ADe epidermis revealed that the molecular mass of SM deacylase is 40000 Da, which is consistent with its apparent molecular mass of 42000 Da estimated by gel-filtration analysis of stratum corneum extracts. Analytical isoelectric focusing (IEF) chromatography demonstrated that the pI values of SM deacylase, beta-glucocerebrosidase (GlcCDase), sphingomyelinase (SMase) and acid ceramidase were 4.2, 7.4, 7.0 and 5.7, respectively. In enzymic analysis using pI-4.2 SM deacylase partially purified by IEF, which had no detectable contamination with acid ceramidase, GlcCDase or SMase, radio-TLC analysis revealed that radiolabelled sphingosylphosphocholine or [1-(14)C]palmitic acid was enzymically liberated from [choline-methyl-(14)C]SM or N-[palmitoyl-1-(14)C]GCer, respectively, used as substrates. Further the pI-4.2 protein purified from extracts of the stratum corneum of ADe patients was able to hydrolyse N-[palmitoyl-1-(14)C]SM and GCer, but not N-[palmitoyl-1-(14)C]ceramide. These results indicate that a hitherto undiscovered epidermal enzyme, termed here glucosylceramide sphingomyelin deacylase, is expressed in the skin of ADe patients, which plays an important role in ceramide deficiency (including acylceramides) in the stratum corneum.

Sphingosylphosphorylcholine is upregulated in the stratum corneum of patients with atopic dermatitis

“…A similar accumulation of SPC has been observed in Niemann-Pick disease, which is associated with defects of SMase and results in the lipidosis for SM (17), although no data suggest that this SPC accumulation is linked to the expression of an SM deacylase-like enzyme. Although SPC accumulation has been speculated to result from a defect of SMase [because SPC is a substrate for SMase (17)], it would be intriguing to determine whether there is a similar up-regulated expression of SM deacylase in Niemann-Pick disease as that could also provide a mechanism for the production of SPC in that disease as it does for AD (6,7,16). A similar accumulation of substrates and reaction products by corresponding N-deacylase enzymes has been found in Gaucher disease, in which there is an accumulation of glucosylceramide and GS due to a defect of GlCdase activity (18).…”

Assessment of Epidermal Barrier Function by Photoacoustic Spectrometry in Relation to Its Importance in the Pathogenesis of Atopic Dermatitis

“…Dysfunction of the stratum corneum seems to be associated with the increased permeability and penetration of environmental irritants and allergens in clinically normal AD skin (Cooper, 1994;Deleuran et al, 1998;Fitzharris and Riley, 1999;Gloor et al, 1981;Ogawa and Yoshiike, 1993;Rothe and Grant-Kels, 1996;Tan et al, 1996;Tanaka et al, 1994;Tupker et al, 1990), which predisposes it to inflammation (Broberg et al, 1992;Lammintausta et al, 1993). The impaired barrier function of the stratum corneum in AD has been found to result from the decreased production of ceramides (Imokawa et al, 1991(Imokawa et al, , 1994, which are generated from sphingomyelin by sphingomyelinase Higuchi et al, 2000;Murata et al, 1996;Wood et al, 1996). However, the activity of sphingomyelin deacylase, which hydrolyzes sphingomyelin at the acyl site to yield sphingosylphosphorylcholine, is abnormally high in clinically normal and lesional skin in AD , resulting in the ceramide deficiency (Imokawa et al, 1991).…”

Sphingosylphosphorylcholine acts in an anti-inflammatory manner in renal mesangial cells by reducing interleukin-1β-induced prostaglandin E2 formation

“…Furthermore, patients with atopic dermatitis have increased SPC and reduced ceramide levels in the skin, which correlate with an enhanced activity of a sphingomyelin (SM) deacylase in the skin of these patients (60)(61)(62)(63). This suggests that SPC is formed by a phospholipase A 2 (PLA 2 )-like enzymatic activity.…”

“…Whether this is changed under inflammatory or fibrotic disease conditions has not been addressed. SPC formation is theoretically possible: i) from SM by the action of a PLA 2 -like enzyme such as the SM deacylase (60,61); ii) from sphingosine by phosphocholine transfer; or iii) from S1P by choline transfer. The generation of SPC from SM by PLA 2 activity is an intriguing hypothesis and is further stressed by the finding that cells isolated from Niemann-Pick patients, which possess an acid sphingomyelinase deficiency and consequently accumulate SM in the lysosomes, also show an accumulation of SPC (64).…”